hcn2 neuronal cells  (ATCC)


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    Structured Review

    ATCC hcn2 neuronal cells
    GRP78 plays a key role in regulating the expression of APP and Tau after exposure of <t>neuronal</t> <t>cells</t> and microglia to AR12 and neratinib. <t>HCN2</t> and BV2 cells were transfected with a scrambled siRNA or with siRNA molecules to knock down the expression of GRP78, HSP70 or HSP90 and in parallel co-transfected to express Tau or APP. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of APP, Tau and ERK2. (n = 3 +/-SD) * p
    Hcn2 Neuronal Cells, supplied by ATCC, used in various techniques. Bioz Stars score: 94/100, based on 27 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Images

    1) Product Images from "AR12 increases BAG3 expression which is essential for Tau and APP degradation via LC3-associated phagocytosis and macroautophagy"

    Article Title: AR12 increases BAG3 expression which is essential for Tau and APP degradation via LC3-associated phagocytosis and macroautophagy

    Journal: Aging (Albany NY)

    doi: 10.18632/aging.204337

    GRP78 plays a key role in regulating the expression of APP and Tau after exposure of neuronal cells and microglia to AR12 and neratinib. HCN2 and BV2 cells were transfected with a scrambled siRNA or with siRNA molecules to knock down the expression of GRP78, HSP70 or HSP90 and in parallel co-transfected to express Tau or APP. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of APP, Tau and ERK2. (n = 3 +/-SD) * p
    Figure Legend Snippet: GRP78 plays a key role in regulating the expression of APP and Tau after exposure of neuronal cells and microglia to AR12 and neratinib. HCN2 and BV2 cells were transfected with a scrambled siRNA or with siRNA molecules to knock down the expression of GRP78, HSP70 or HSP90 and in parallel co-transfected to express Tau or APP. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of APP, Tau and ERK2. (n = 3 +/-SD) * p

    Techniques Used: Expressing, Transfection, Immunostaining

    Knock down of Beclin1 or ATG5 prevents autophagosome formation and autophagic flux in HCN2 cells. HCN2 neuronal cells were transfected with a scrambled siRNA or with an siRNA to knock down the expression of ATG5 or Beclin1 and were co-transfected with a plasmid to express LC3-GFP-RFP. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 4h and 8h. The mean number of intense GFP+RFP+ and RFP+ punctae per cell was determined (n = 3 +/-SD) # p
    Figure Legend Snippet: Knock down of Beclin1 or ATG5 prevents autophagosome formation and autophagic flux in HCN2 cells. HCN2 neuronal cells were transfected with a scrambled siRNA or with an siRNA to knock down the expression of ATG5 or Beclin1 and were co-transfected with a plasmid to express LC3-GFP-RFP. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 4h and 8h. The mean number of intense GFP+RFP+ and RFP+ punctae per cell was determined (n = 3 +/-SD) # p

    Techniques Used: Transfection, Expressing, Plasmid Preparation

    AR12 and neratinib reduce chaperone expression in neuronal cells. HCN2 neuronal cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of GRP78 (total and cell surface), HSP70, HSP90, eIF2α and ERK2, and the phosphorylation of eIF2α S51. (n = 3 +/-SD) * p
    Figure Legend Snippet: AR12 and neratinib reduce chaperone expression in neuronal cells. HCN2 neuronal cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of GRP78 (total and cell surface), HSP70, HSP90, eIF2α and ERK2, and the phosphorylation of eIF2α S51. (n = 3 +/-SD) * p

    Techniques Used: Expressing, Immunostaining

    Enhanced BAG3 expression requires autophagy and ER stress signaling. HCN2 cells were transfected with a scrambled siRNA control or with siRNA molecules to knock down the expression of Beclin1, ATG5, ULK1, eIF2α or PERK. After 24h, cells were treated for 6h with vehicle control or with AR12 (2 μM). Cells were fixed in place and immunostaining performed to detect the expression of BAG3 and ERK2 (n = 3 +/-SD) # p
    Figure Legend Snippet: Enhanced BAG3 expression requires autophagy and ER stress signaling. HCN2 cells were transfected with a scrambled siRNA control or with siRNA molecules to knock down the expression of Beclin1, ATG5, ULK1, eIF2α or PERK. After 24h, cells were treated for 6h with vehicle control or with AR12 (2 μM). Cells were fixed in place and immunostaining performed to detect the expression of BAG3 and ERK2 (n = 3 +/-SD) # p

    Techniques Used: Expressing, Transfection, Immunostaining

    BAG3 is essential for drug-induced degradation of GRP78. ( A – C ) HCN2 cells were transfected with a scrambled siRNA or with an siRNA to knock down BAG3 expression. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of GRP78 (total and cell surface), HDAC6, HSP90, p62, LAMP2, PERK, P-PERK T980, eIF2α and P-eIF2α S51 and ERK2. (n = 3 +/-SD) * p
    Figure Legend Snippet: BAG3 is essential for drug-induced degradation of GRP78. ( A – C ) HCN2 cells were transfected with a scrambled siRNA or with an siRNA to knock down BAG3 expression. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of GRP78 (total and cell surface), HDAC6, HSP90, p62, LAMP2, PERK, P-PERK T980, eIF2α and P-eIF2α S51 and ERK2. (n = 3 +/-SD) * p

    Techniques Used: Transfection, Expressing, Immunostaining

    ER stress signaling plays a key role in facilitating autophagy and protein degradation in neuronal cells. Upper Graphs. HCN2 cells and BV2 cells were transfected with a scrambled siRNA or with an siRNA to knock down the expression of eIF2α and were co-transfected with a plasmid to express LC3-GFP-RFP. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 4h and 8h. The mean number of intense staining GFP+RFP+ and RFP+ punctae per cell was determined (n = 3 +/-SD) # p
    Figure Legend Snippet: ER stress signaling plays a key role in facilitating autophagy and protein degradation in neuronal cells. Upper Graphs. HCN2 cells and BV2 cells were transfected with a scrambled siRNA or with an siRNA to knock down the expression of eIF2α and were co-transfected with a plasmid to express LC3-GFP-RFP. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 4h and 8h. The mean number of intense staining GFP+RFP+ and RFP+ punctae per cell was determined (n = 3 +/-SD) # p

    Techniques Used: Transfection, Expressing, Plasmid Preparation, Staining

    Over-expression of GRP78 suppresses the drug-induced expression of BAG3. ( A ) HCN2 cells were transfected with a scrambled siRNA or with siRNA molecules to knock down the expression of GRP78, HSP70 or HSP90. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of BAG3 and ERK2. (n = 3 +/-SD) # p
    Figure Legend Snippet: Over-expression of GRP78 suppresses the drug-induced expression of BAG3. ( A ) HCN2 cells were transfected with a scrambled siRNA or with siRNA molecules to knock down the expression of GRP78, HSP70 or HSP90. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of BAG3 and ERK2. (n = 3 +/-SD) # p

    Techniques Used: Over Expression, Expressing, Transfection, Immunostaining

    ER stress signaling plays a key role in facilitating autophagy and HDAC6 protein degradation. ( A ) HCN2 cells were transfected with a scrambled siRNA or with an siRNA molecule to knock down the expression of eIF2α. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of HDAC6, LAMP2, p62 and ERK2. (n = 3 +/-SD) * p
    Figure Legend Snippet: ER stress signaling plays a key role in facilitating autophagy and HDAC6 protein degradation. ( A ) HCN2 cells were transfected with a scrambled siRNA or with an siRNA molecule to knock down the expression of eIF2α. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of HDAC6, LAMP2, p62 and ERK2. (n = 3 +/-SD) * p

    Techniques Used: Transfection, Expressing, Immunostaining

    Knock down of Rubicon suppresses drug-induced autophagosome formation but does not appear to alter autophagic flux. HCN2 neuronal cells and BV2 microglial cells were transfected with a scrambled siRNA or with an siRNA to knock down the expression of Rubicon and were co-transfected with a plasmid to express LC3-GFP-RFP. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 4h and 8h. The mean number of intense GFP+RFP+ and RFP+ punctae per cell was determined (n = 3 +/-SD) # p
    Figure Legend Snippet: Knock down of Rubicon suppresses drug-induced autophagosome formation but does not appear to alter autophagic flux. HCN2 neuronal cells and BV2 microglial cells were transfected with a scrambled siRNA or with an siRNA to knock down the expression of Rubicon and were co-transfected with a plasmid to express LC3-GFP-RFP. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 4h and 8h. The mean number of intense GFP+RFP+ and RFP+ punctae per cell was determined (n = 3 +/-SD) # p

    Techniques Used: Transfection, Expressing, Plasmid Preparation

    AR12 and the drug combination reduce the expression of mutant forms of TAU and APP. HCN2 and BV2 cells were transfected with plasmids to express wild type TAU or TAU 301L. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of TAU, TAU 301L and ERK2. (n = 3 +/-SD) * p
    Figure Legend Snippet: AR12 and the drug combination reduce the expression of mutant forms of TAU and APP. HCN2 and BV2 cells were transfected with plasmids to express wild type TAU or TAU 301L. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of TAU, TAU 301L and ERK2. (n = 3 +/-SD) * p

    Techniques Used: Expressing, Mutagenesis, Transfection, Immunostaining

    Degradation of APP and Tau by AR12 and neratinib requires LAP and macroautophagy in HCN2 neuronal cells and in BV2 microglia. ( A , B ) HCN2 cells were transfected with a scrambled siRNA or with siRNA molecules to knock down expression of Rubicon, Beclin1 or ATG5. In parallel, cells were transfected with an empty vector plasmid in ( B ), or transfected with plasmids to express APP or Tau in ( A ). After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of Tau, APP and ERK2. (n = 3 +/-SD) Endogenous expression of APP was 5% of the value for APP expressed from a plasmid. Endogenous expression of Tau was 6% of the value for Tau expressed from a plasmid. * p
    Figure Legend Snippet: Degradation of APP and Tau by AR12 and neratinib requires LAP and macroautophagy in HCN2 neuronal cells and in BV2 microglia. ( A , B ) HCN2 cells were transfected with a scrambled siRNA or with siRNA molecules to knock down expression of Rubicon, Beclin1 or ATG5. In parallel, cells were transfected with an empty vector plasmid in ( B ), or transfected with plasmids to express APP or Tau in ( A ). After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of Tau, APP and ERK2. (n = 3 +/-SD) Endogenous expression of APP was 5% of the value for APP expressed from a plasmid. Endogenous expression of Tau was 6% of the value for Tau expressed from a plasmid. * p

    Techniques Used: Transfection, Expressing, Plasmid Preparation, Immunostaining

    Expression of Tau or APP does not significantly alter the regulation of protein phosphorylation or protein expression caused by AR12 and neratinib. HCN2 cells were transfected with an empty vector plasmid or with plasmids to express Tau or APP. Twenty-four h afterwards, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the phosphorylation and expression of the indicated proteins (n = 3 +/-SD). * p
    Figure Legend Snippet: Expression of Tau or APP does not significantly alter the regulation of protein phosphorylation or protein expression caused by AR12 and neratinib. HCN2 cells were transfected with an empty vector plasmid or with plasmids to express Tau or APP. Twenty-four h afterwards, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the phosphorylation and expression of the indicated proteins (n = 3 +/-SD). * p

    Techniques Used: Expressing, Transfection, Plasmid Preparation, Immunostaining

    Control data showing protein expression knock down and protein over-expression. Left: BV2 and HCN2 cells as indicated were transfected with siRNA molecules to knock down the expression of the indicated proteins or transfected with plasmids to over-express the indicated proteins. The percentage remaining after knock-down or the percentage over-expression above basal levels is indicated. (n = 3 +/-SD) (total ERK2 is included as an invariant total protein loading control). Right: Images of HCN2 cells transfected to express TAU or APP.
    Figure Legend Snippet: Control data showing protein expression knock down and protein over-expression. Left: BV2 and HCN2 cells as indicated were transfected with siRNA molecules to knock down the expression of the indicated proteins or transfected with plasmids to over-express the indicated proteins. The percentage remaining after knock-down or the percentage over-expression above basal levels is indicated. (n = 3 +/-SD) (total ERK2 is included as an invariant total protein loading control). Right: Images of HCN2 cells transfected to express TAU or APP.

    Techniques Used: Expressing, Over Expression, Transfection

    Degradation of chaperones and eIF2α S51 phosphorylation requires LAP and macroautophagy. ( A ) HCN2 and ( B ) BV2 cells were transfected with a scrambled siRNA or with siRNA molecules to knock down the expression of Rubicon, Beclin1 or ATG5. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of GRP78 (total and cell surface / plasma membrane), HSP70, HSP90, eIF2α and ERK2, and the phosphorylation of eIF2α S51. (n = 3 +/-SD) * p
    Figure Legend Snippet: Degradation of chaperones and eIF2α S51 phosphorylation requires LAP and macroautophagy. ( A ) HCN2 and ( B ) BV2 cells were transfected with a scrambled siRNA or with siRNA molecules to knock down the expression of Rubicon, Beclin1 or ATG5. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of GRP78 (total and cell surface / plasma membrane), HSP70, HSP90, eIF2α and ERK2, and the phosphorylation of eIF2α S51. (n = 3 +/-SD) * p

    Techniques Used: Transfection, Expressing, Immunostaining

    AR12 and neratinib cause degradation of p70 S6K in HCN2 neuronal cells. ( A ) HCN2 cells were transfected with a scrambled siRNA control or with an siRNA to knock down expression of AMPKa. After 24h, cells were treated with vehicle control or with [AR12 (2 μM) + neratinib (50 nM)] for 6h. Cells were fixed in place and immunostaining performed to determine the phosphorylation and expression of the indicated proteins (n = 3 +/-SD). * p
    Figure Legend Snippet: AR12 and neratinib cause degradation of p70 S6K in HCN2 neuronal cells. ( A ) HCN2 cells were transfected with a scrambled siRNA control or with an siRNA to knock down expression of AMPKa. After 24h, cells were treated with vehicle control or with [AR12 (2 μM) + neratinib (50 nM)] for 6h. Cells were fixed in place and immunostaining performed to determine the phosphorylation and expression of the indicated proteins (n = 3 +/-SD). * p

    Techniques Used: Transfection, Expressing, Immunostaining

    2) Product Images from "MicroRNAs serve as prediction and treatment-response biomarkers of attention-deficit/hyperactivity disorder and promote the differentiation of neuronal cells by repressing the apoptosis pathway"

    Article Title: MicroRNAs serve as prediction and treatment-response biomarkers of attention-deficit/hyperactivity disorder and promote the differentiation of neuronal cells by repressing the apoptosis pathway

    Journal: Translational Psychiatry

    doi: 10.1038/s41398-022-01832-1

    Illustration and comparisons of the differentiation patterns of HCN-2 cells transfected with miRNA mimic. a HCN-2 cells were transfected with scrambled control mimic (control), hsa-miR-30e-5p mimic (miR-30e), hsa-miR-140-3p mimic (miR-140), or hsa-miR-126-5p mimic (miR-126) for 24 h. Then, they were seeded on new plates for monitoring and recording growth with a camera. Each transfection treatment had three independent wells, and three pictures were randomly taken from each well, resulting in 3*3 = 9 pictures. ( b – f ) We determined the differentiation patterns of HCN-2 cells in terms of different indices, including ( b ) vessel area, ( c ) vessel length, ( d ) junction number, and ( e ) mean lacunarity. The t -test p values were calculated by comparing the values of the control set on a specific day. f For a systematic comparison, the values were normalized to the average values of the control set at day 9. *, **, *** and **** denote p value
    Figure Legend Snippet: Illustration and comparisons of the differentiation patterns of HCN-2 cells transfected with miRNA mimic. a HCN-2 cells were transfected with scrambled control mimic (control), hsa-miR-30e-5p mimic (miR-30e), hsa-miR-140-3p mimic (miR-140), or hsa-miR-126-5p mimic (miR-126) for 24 h. Then, they were seeded on new plates for monitoring and recording growth with a camera. Each transfection treatment had three independent wells, and three pictures were randomly taken from each well, resulting in 3*3 = 9 pictures. ( b – f ) We determined the differentiation patterns of HCN-2 cells in terms of different indices, including ( b ) vessel area, ( c ) vessel length, ( d ) junction number, and ( e ) mean lacunarity. The t -test p values were calculated by comparing the values of the control set on a specific day. f For a systematic comparison, the values were normalized to the average values of the control set at day 9. *, **, *** and **** denote p value

    Techniques Used: Transfection

    3) Product Images from "MicroRNAs serve as prediction and treatment-response biomarkers of attention-deficit/hyperactivity disorder and promote the differentiation of neuronal cells by repressing the apoptosis pathway"

    Article Title: MicroRNAs serve as prediction and treatment-response biomarkers of attention-deficit/hyperactivity disorder and promote the differentiation of neuronal cells by repressing the apoptosis pathway

    Journal: Translational Psychiatry

    doi: 10.1038/s41398-022-01832-1

    Illustration and comparisons of the differentiation patterns of HCN-2 cells transfected with miRNA mimic. a HCN-2 cells were transfected with scrambled control mimic (control), hsa-miR-30e-5p mimic (miR-30e), hsa-miR-140-3p mimic (miR-140), or hsa-miR-126-5p mimic (miR-126) for 24 h. Then, they were seeded on new plates for monitoring and recording growth with a camera. Each transfection treatment had three independent wells, and three pictures were randomly taken from each well, resulting in 3*3 = 9 pictures. ( b – f ) We determined the differentiation patterns of HCN-2 cells in terms of different indices, including ( b ) vessel area, ( c ) vessel length, ( d ) junction number, and ( e ) mean lacunarity. The t -test p values were calculated by comparing the values of the control set on a specific day. f For a systematic comparison, the values were normalized to the average values of the control set at day 9. *, **, *** and **** denote p value
    Figure Legend Snippet: Illustration and comparisons of the differentiation patterns of HCN-2 cells transfected with miRNA mimic. a HCN-2 cells were transfected with scrambled control mimic (control), hsa-miR-30e-5p mimic (miR-30e), hsa-miR-140-3p mimic (miR-140), or hsa-miR-126-5p mimic (miR-126) for 24 h. Then, they were seeded on new plates for monitoring and recording growth with a camera. Each transfection treatment had three independent wells, and three pictures were randomly taken from each well, resulting in 3*3 = 9 pictures. ( b – f ) We determined the differentiation patterns of HCN-2 cells in terms of different indices, including ( b ) vessel area, ( c ) vessel length, ( d ) junction number, and ( e ) mean lacunarity. The t -test p values were calculated by comparing the values of the control set on a specific day. f For a systematic comparison, the values were normalized to the average values of the control set at day 9. *, **, *** and **** denote p value

    Techniques Used: Transfection

    4) Product Images from "Development of Lipidoid Nanoparticles for siRNA Delivery to Neural Cells"

    Article Title: Development of Lipidoid Nanoparticles for siRNA Delivery to Neural Cells

    Journal: The AAPS Journal

    doi: 10.1208/s12248-021-00653-2

    Cellular uptake of Cy5 siRNA determined using fluorescent microscopy. HCN-2 cells (human cortical neuron cell line) were incubated for 2 h ( a ), 4 h ( b ), and 24 h ( c ) with the indicated samples containing 50 nM Cy5 siRNA. Panel ( d ) represents the zoomed-in image depicting HCN-2 cells incubated with LNPs with PEG-DMG. Scale bar = 50 µm. Cells were imaged using an Olympus IX 73 epifluorescent inverted microscope to detect Cy5 signals at excitation and emission wavelengths of 651 nm and 670 nm, respectively. Images are representative of n = 3 independent wells
    Figure Legend Snippet: Cellular uptake of Cy5 siRNA determined using fluorescent microscopy. HCN-2 cells (human cortical neuron cell line) were incubated for 2 h ( a ), 4 h ( b ), and 24 h ( c ) with the indicated samples containing 50 nM Cy5 siRNA. Panel ( d ) represents the zoomed-in image depicting HCN-2 cells incubated with LNPs with PEG-DMG. Scale bar = 50 µm. Cells were imaged using an Olympus IX 73 epifluorescent inverted microscope to detect Cy5 signals at excitation and emission wavelengths of 651 nm and 670 nm, respectively. Images are representative of n = 3 independent wells

    Techniques Used: Microscopy, Incubation, Inverted Microscopy

    5) Product Images from "Transcriptomic Analysis of HCN-2 Cells Suggests Connection among Oxidative Stress, Senescence, and Neuron Death after SARS-CoV-2 Infection"

    Article Title: Transcriptomic Analysis of HCN-2 Cells Suggests Connection among Oxidative Stress, Senescence, and Neuron Death after SARS-CoV-2 Infection

    Journal: Cells

    doi: 10.3390/cells10092189

    ( a ) Evidence of a significant decrease in SOD1 expression in HCN-2 culture medium after SARS-CoV-2 infection, suggesting impairment of ROS reduction. ( b ) Densitometric analysis of SOD1, * p
    Figure Legend Snippet: ( a ) Evidence of a significant decrease in SOD1 expression in HCN-2 culture medium after SARS-CoV-2 infection, suggesting impairment of ROS reduction. ( b ) Densitometric analysis of SOD1, * p

    Techniques Used: Expressing, Infection

    ( a ) Evidence of significant increase in caspase 3 expression in HCN-2 culture medium after SARS-CoV-2 infection, suggesting that apoptosis is taking place in the infected cells; ( b ) densitometric analysis of caspase 3, * p
    Figure Legend Snippet: ( a ) Evidence of significant increase in caspase 3 expression in HCN-2 culture medium after SARS-CoV-2 infection, suggesting that apoptosis is taking place in the infected cells; ( b ) densitometric analysis of caspase 3, * p

    Techniques Used: Expressing, Infection

    SARS-CoV-2-triggered pathway depiction of DEGs found in RNA-seq analyses after infection of the HCN-2. The red genes are upregulated and, thus, more expressed after the infection. The green downregulated genes are more expressed in the control. The yellow frames show increased activities.
    Figure Legend Snippet: SARS-CoV-2-triggered pathway depiction of DEGs found in RNA-seq analyses after infection of the HCN-2. The red genes are upregulated and, thus, more expressed after the infection. The green downregulated genes are more expressed in the control. The yellow frames show increased activities.

    Techniques Used: RNA Sequencing Assay, Infection

    Enriched biological processes terms involved in oxidative stress and death by differentially expressed genes (DEGs) found in HCN-2 cells after exposure to SARS-CoV-2. For each Gene Ontology term represented, the length of the bar shows the number of DEGs observed in that specific category. The color of the bar points to the ratio between the number of DEGs found in our analysis and the number of genes included in the term from white (no DEGs of the term) to green (all DEGs of the term). The terms are alphabetically sorted.
    Figure Legend Snippet: Enriched biological processes terms involved in oxidative stress and death by differentially expressed genes (DEGs) found in HCN-2 cells after exposure to SARS-CoV-2. For each Gene Ontology term represented, the length of the bar shows the number of DEGs observed in that specific category. The color of the bar points to the ratio between the number of DEGs found in our analysis and the number of genes included in the term from white (no DEGs of the term) to green (all DEGs of the term). The terms are alphabetically sorted.

    Techniques Used:

    Enriched biological processes terms involved in cell cycle by differentially expressed genes (DEGs) found in HCN-2 cells after exposure to SARS-CoV-2. For each Gene Ontology term represented, the length of the bar shows the number of DEGs observed in that specific category. The color of the bar points to the ratio between the number of DEGs found in our analysis and the number of genes included in the term from white (no DEGs of the term) to green (all DEGs of the term). The terms are alphabetically sorted.
    Figure Legend Snippet: Enriched biological processes terms involved in cell cycle by differentially expressed genes (DEGs) found in HCN-2 cells after exposure to SARS-CoV-2. For each Gene Ontology term represented, the length of the bar shows the number of DEGs observed in that specific category. The color of the bar points to the ratio between the number of DEGs found in our analysis and the number of genes included in the term from white (no DEGs of the term) to green (all DEGs of the term). The terms are alphabetically sorted.

    Techniques Used:

    ( a ) Evidence of significant increase in COX2 expression in HCN-2 culture medium after SARS-CoV-2 infection, suggesting inflammation in the infected cells; ( b ) densitometric analysis of COX2, * p
    Figure Legend Snippet: ( a ) Evidence of significant increase in COX2 expression in HCN-2 culture medium after SARS-CoV-2 infection, suggesting inflammation in the infected cells; ( b ) densitometric analysis of COX2, * p

    Techniques Used: Expressing, Infection

    6) Product Images from "Development of lipidoid nanoparticles for siRNA delivery to neural cells"

    Article Title: Development of lipidoid nanoparticles for siRNA delivery to neural cells

    Journal: bioRxiv

    doi: 10.1101/2021.07.28.454207

    Cellular uptake of Cy5 siRNA determined using fluorescent microscopy. HCN-2 cells (human cortical neuron cell line) were incubated for 2 hours ( a ) 4 hours ( b ) and 24 hours ( c ) with the indicated samples containing 50 nM Cy5 siRNA. Scale bar = 50 μm. Cells were imaged using an Olympus IX 73 epifluorescent inverted microscope to detect Cy5 signals at excitation and emission wavelengths of 651 nm and 670 nm, respectively. Images are representative of n=3 independent wells.
    Figure Legend Snippet: Cellular uptake of Cy5 siRNA determined using fluorescent microscopy. HCN-2 cells (human cortical neuron cell line) were incubated for 2 hours ( a ) 4 hours ( b ) and 24 hours ( c ) with the indicated samples containing 50 nM Cy5 siRNA. Scale bar = 50 μm. Cells were imaged using an Olympus IX 73 epifluorescent inverted microscope to detect Cy5 signals at excitation and emission wavelengths of 651 nm and 670 nm, respectively. Images are representative of n=3 independent wells.

    Techniques Used: Microscopy, Incubation, Inverted Microscopy

    7) Product Images from "SARS-CoV-2 Infected Pediatric Cerebral Cortical Neurons: Transcriptomic Analysis and Potential Role of Toll-like Receptors in Pathogenesis"

    Article Title: SARS-CoV-2 Infected Pediatric Cerebral Cortical Neurons: Transcriptomic Analysis and Potential Role of Toll-like Receptors in Pathogenesis

    Journal: International Journal of Molecular Sciences

    doi: 10.3390/ijms22158059

    Western blot for ACE2 and TLR4. HCN-2 cells expressed TLR4 but not ACE2. A549 and A549-hACE2 expressed ACE2 and TLR4.
    Figure Legend Snippet: Western blot for ACE2 and TLR4. HCN-2 cells expressed TLR4 but not ACE2. A549 and A549-hACE2 expressed ACE2 and TLR4.

    Techniques Used: Western Blot

    Virus replication in HCN-2 cortical neurons. N1 and N2 copy number increased in a time dependent manner. * p
    Figure Legend Snippet: Virus replication in HCN-2 cortical neurons. N1 and N2 copy number increased in a time dependent manner. * p

    Techniques Used:

    8) Product Images from "Photoactivatable metabolic warheads enable precise and safe ablation of target cells in vivo"

    Article Title: Photoactivatable metabolic warheads enable precise and safe ablation of target cells in vivo

    Journal: Nature Communications

    doi: 10.1038/s41467-021-22578-2

    Compound 15 selectively ablates human glioblastoma cancer cells under clinical PDT conditions and in the presence of non-cancerous human brain cells. a Procedure for coculturing human non-cancerous brain HCN-2 cells (stained with CellMask Green) and human glioblastoma U87-nlsCrimson cells. Cocultures (15,000 HCN-2 cells and 10,000 U87 cells per well) were incubated with compound 15 (100 μM) and illuminated with ThorLabs M530L3 LED (37 J cm −2 ). The fluorescence emission from E2Crimson is reduced in U87-nlsCrimson glioblastoma cells upon cell death. ( b , c ) Representative brightfield and fluorescent confocal microscopy images (from three independent experiments) of cocultured HCN-2 (green, λ em : 520 nm) and U87-nlsCrimson (magenta, λ em : 645 nm) before ( b ) and after treatment ( c ). Yellow arrowheads in ( c ) highlight dead U87h cells devoid of Crimson fluorescence. Scale bars: 10 μm. d Phototoxicity analysis under different concentrations of compound 15 and 5-ALA in U87 glioblastoma cells (20,000 cells/well) using the same irradiation settings (10 mW, 37 J cm −2 ). Data presented as mean values ± SEM ( n = 3 independent experiments). e HCN-2 cell viability (10,000 cells/well) after incubation with concentrations under the phototoxicity threshold [i.e., compound 15 (100 μM, red) and 5-ALA (200 μM, blue)] and light exposure (37 J cm −2 ). Data presented as mean values ± SEM ( n = 4 independent experiments). Source data are available.
    Figure Legend Snippet: Compound 15 selectively ablates human glioblastoma cancer cells under clinical PDT conditions and in the presence of non-cancerous human brain cells. a Procedure for coculturing human non-cancerous brain HCN-2 cells (stained with CellMask Green) and human glioblastoma U87-nlsCrimson cells. Cocultures (15,000 HCN-2 cells and 10,000 U87 cells per well) were incubated with compound 15 (100 μM) and illuminated with ThorLabs M530L3 LED (37 J cm −2 ). The fluorescence emission from E2Crimson is reduced in U87-nlsCrimson glioblastoma cells upon cell death. ( b , c ) Representative brightfield and fluorescent confocal microscopy images (from three independent experiments) of cocultured HCN-2 (green, λ em : 520 nm) and U87-nlsCrimson (magenta, λ em : 645 nm) before ( b ) and after treatment ( c ). Yellow arrowheads in ( c ) highlight dead U87h cells devoid of Crimson fluorescence. Scale bars: 10 μm. d Phototoxicity analysis under different concentrations of compound 15 and 5-ALA in U87 glioblastoma cells (20,000 cells/well) using the same irradiation settings (10 mW, 37 J cm −2 ). Data presented as mean values ± SEM ( n = 3 independent experiments). e HCN-2 cell viability (10,000 cells/well) after incubation with concentrations under the phototoxicity threshold [i.e., compound 15 (100 μM, red) and 5-ALA (200 μM, blue)] and light exposure (37 J cm −2 ). Data presented as mean values ± SEM ( n = 4 independent experiments). Source data are available.

    Techniques Used: Staining, Incubation, Fluorescence, Confocal Microscopy, Irradiation

    9) Product Images from "Tumor-responsive, multifunctional CAR-NK cells cooperate with impaired autophagy to infiltrate and target glioblastoma"

    Article Title: Tumor-responsive, multifunctional CAR-NK cells cooperate with impaired autophagy to infiltrate and target glioblastoma

    Journal: bioRxiv

    doi: 10.1101/2020.10.07.330043

    In vitro effector activity of multifunctional genetically-engineered NK cells against patient-derived GBM. ( A ) In vitro cytotoxicity of NK-92 and CD73.mCAR-NK92 cells against GBM43 cells at indicated E/T ratios over 4 h. ( B ) Degranulation of NK-92 and CD73.mCAR-NK92 cells [% CD107 and (MFI) CD107] after 4 h coculture with GBM43 cells (E/T ratio, 5:1). NK cells were analyzed by flow cytometry for surface CD107a expression as a marker of degranulation. ( C ) IFN-γ production by NK-92 and CD73.mCAR-NK92 cells (% IFN-γ) after 4 h coculture with GBM43 cells (E/T ratio, 5:1). ( D ) In vitro cytotoxicity of CD73.mCAR-NK92 and CD73.mCAR-NK92 (following aCD73 scFv cleavage) cells against GBM43 cells at indicated E/T ratios over 4 h. ( E ) CD73 activity of GBM43 cells after incubation with cleaved aCD73 scFv following release from uPA-treated CD73.mCAR-NK92 cells. ( F-H ) In vitro cytotoxicity of pNK and CD73.mCAR-pNK cells against different GBM cells, including SJ-GBM2, GBM43 and GBM10 cells, at indicated E/T ratios over 4 h. ( I ) Degranulation of pNK and CD73.mCAR-pNK cells (% CD107) after 4 h coculture with GBM43 cells (E/T ratio, 5:1). NK cells were analyzed by flow cytometry for surface CD107a expression as a marker of degranulation. ( J ) IFN-γ production of pNK and CD73.mCAR-pNK cells (% IFN-γ) after 4 h coculture with GBM43 cells (E/T ratio, 5:1). ( K ) In vitro cytotoxicity of pNK and CD73.mCAR-pNK (following aCD73 scFv cleavage) cells against GBM43 cells at indicated E/T ratios over 4 h. ( L ) CD73 activity of GBM43 cells after incubation with cleaved aCD73 scFv following cleavage from uPA-treated CD73.mCAR-NK cells. ( M ) Relative decrease in CD16 expression on pNK and CD73.mCAR-pNK cells (% of MFI) after 12 h coculture with GBM43 cells (E/T ratio, 5:1). ( N ) Relative increase in NKG2A expression on pNK and CD73.mCAR-pNK cells (% of MFI) after 12 h coculture with GBM43 cells (E/T ratio, 5:1). ( O and P ) In vitro cytotoxicity of pNK and CD73.mCAR-pNK cells against nonmalignant neural cell lines hCMEC/D3 and HCN-2 at indicated E/T ratios over 4 h. Note: the data shown through ( F )-( P ) is for isolated pNK cells in at least triplicates from a representative donor. Data are shown as mean ± SEM. * p
    Figure Legend Snippet: In vitro effector activity of multifunctional genetically-engineered NK cells against patient-derived GBM. ( A ) In vitro cytotoxicity of NK-92 and CD73.mCAR-NK92 cells against GBM43 cells at indicated E/T ratios over 4 h. ( B ) Degranulation of NK-92 and CD73.mCAR-NK92 cells [% CD107 and (MFI) CD107] after 4 h coculture with GBM43 cells (E/T ratio, 5:1). NK cells were analyzed by flow cytometry for surface CD107a expression as a marker of degranulation. ( C ) IFN-γ production by NK-92 and CD73.mCAR-NK92 cells (% IFN-γ) after 4 h coculture with GBM43 cells (E/T ratio, 5:1). ( D ) In vitro cytotoxicity of CD73.mCAR-NK92 and CD73.mCAR-NK92 (following aCD73 scFv cleavage) cells against GBM43 cells at indicated E/T ratios over 4 h. ( E ) CD73 activity of GBM43 cells after incubation with cleaved aCD73 scFv following release from uPA-treated CD73.mCAR-NK92 cells. ( F-H ) In vitro cytotoxicity of pNK and CD73.mCAR-pNK cells against different GBM cells, including SJ-GBM2, GBM43 and GBM10 cells, at indicated E/T ratios over 4 h. ( I ) Degranulation of pNK and CD73.mCAR-pNK cells (% CD107) after 4 h coculture with GBM43 cells (E/T ratio, 5:1). NK cells were analyzed by flow cytometry for surface CD107a expression as a marker of degranulation. ( J ) IFN-γ production of pNK and CD73.mCAR-pNK cells (% IFN-γ) after 4 h coculture with GBM43 cells (E/T ratio, 5:1). ( K ) In vitro cytotoxicity of pNK and CD73.mCAR-pNK (following aCD73 scFv cleavage) cells against GBM43 cells at indicated E/T ratios over 4 h. ( L ) CD73 activity of GBM43 cells after incubation with cleaved aCD73 scFv following cleavage from uPA-treated CD73.mCAR-NK cells. ( M ) Relative decrease in CD16 expression on pNK and CD73.mCAR-pNK cells (% of MFI) after 12 h coculture with GBM43 cells (E/T ratio, 5:1). ( N ) Relative increase in NKG2A expression on pNK and CD73.mCAR-pNK cells (% of MFI) after 12 h coculture with GBM43 cells (E/T ratio, 5:1). ( O and P ) In vitro cytotoxicity of pNK and CD73.mCAR-pNK cells against nonmalignant neural cell lines hCMEC/D3 and HCN-2 at indicated E/T ratios over 4 h. Note: the data shown through ( F )-( P ) is for isolated pNK cells in at least triplicates from a representative donor. Data are shown as mean ± SEM. * p

    Techniques Used: In Vitro, Activity Assay, Derivative Assay, Flow Cytometry, Expressing, Marker, Incubation, Isolation

    10) Product Images from "MicroRNA-325-3p prevents sevoflurane-induced learning and memory impairment by inhibiting Nupr1 and C/EBPβ/IGFBP5 signaling in rats"

    Article Title: MicroRNA-325-3p prevents sevoflurane-induced learning and memory impairment by inhibiting Nupr1 and C/EBPβ/IGFBP5 signaling in rats

    Journal: Aging (Albany NY)

    doi: 10.18632/aging.102942

    Nupr1 knockdown attenuated sevoflurane-induced apoptosis and decreased C/EBPβ and IGFBP5 expression in HCN-2 neuronal cells. HCN-2 neuronal cells were transfected with NC or Nupr1 siRNA and then exposed to fresh gas (21% O 2 , 5% CO 2 , remainder N 2 ) alone or with the addition of 3.4% sevoflurane for 6 h. ( A , B ) Western blotting for Nupr1, C/EBPβ, and IGFBP5 in 3.4% SEVO-exposed vs. control HCN-2 cells; representative images ( A ) and quantification ( B ). ( C , D ) Western blotting for Cleaved-Caspase-3, Bax, and Bcl-2 in 3.4% SEVO-exposed vs. control HCN-2 cells; representative images (C) and quantification ( D ). ( E , F ) Annexin V-FITC/PI staining and flow cytometry analysis; representative images ( E ) and quantification ( F ). **p
    Figure Legend Snippet: Nupr1 knockdown attenuated sevoflurane-induced apoptosis and decreased C/EBPβ and IGFBP5 expression in HCN-2 neuronal cells. HCN-2 neuronal cells were transfected with NC or Nupr1 siRNA and then exposed to fresh gas (21% O 2 , 5% CO 2 , remainder N 2 ) alone or with the addition of 3.4% sevoflurane for 6 h. ( A , B ) Western blotting for Nupr1, C/EBPβ, and IGFBP5 in 3.4% SEVO-exposed vs. control HCN-2 cells; representative images ( A ) and quantification ( B ). ( C , D ) Western blotting for Cleaved-Caspase-3, Bax, and Bcl-2 in 3.4% SEVO-exposed vs. control HCN-2 cells; representative images (C) and quantification ( D ). ( E , F ) Annexin V-FITC/PI staining and flow cytometry analysis; representative images ( E ) and quantification ( F ). **p

    Techniques Used: Expressing, Transfection, Western Blot, Staining, Flow Cytometry

    Sevoflurane increased Nupr1, C/EBPβ, and IGFBP5 expression and induced apoptosis in HCN-2 neuronal cells. Identical numbers of HCN-2 neuronal cells were exposed to fresh gas (21% O 2 , 5% CO 2 , remainder N 2 ) alone or with the addition of 3.4% sevoflurane for 6 h and then subjected to analysis. ( A , B ) Western blotting for Cleaved-Caspase-3, Bax, and Bcl-2 in 3.4% SEVO-exposed vs. control HCN-2 cells; representative images ( A ) and quantification ( B ). ( C , D ) Annexin V-FITC/PI staining and flow cytometry analysis; representative images ( C ) and quantification ( D ). ( E , F ) Western blotting for Nupr1, C/EBPβ, and IGFBP5 in 3.4% SEVO-exposed vs. control HCN-2 cells; representative images ( E ) and quantification ( F ). **p
    Figure Legend Snippet: Sevoflurane increased Nupr1, C/EBPβ, and IGFBP5 expression and induced apoptosis in HCN-2 neuronal cells. Identical numbers of HCN-2 neuronal cells were exposed to fresh gas (21% O 2 , 5% CO 2 , remainder N 2 ) alone or with the addition of 3.4% sevoflurane for 6 h and then subjected to analysis. ( A , B ) Western blotting for Cleaved-Caspase-3, Bax, and Bcl-2 in 3.4% SEVO-exposed vs. control HCN-2 cells; representative images ( A ) and quantification ( B ). ( C , D ) Annexin V-FITC/PI staining and flow cytometry analysis; representative images ( C ) and quantification ( D ). ( E , F ) Western blotting for Nupr1, C/EBPβ, and IGFBP5 in 3.4% SEVO-exposed vs. control HCN-2 cells; representative images ( E ) and quantification ( F ). **p

    Techniques Used: Expressing, Western Blot, Staining, Flow Cytometry

    Overexpression of Mir-325-3p inhibited sevoflurane-induced apoptosis in HCN-2 neuronal cells. HCN-2 neuronal cells were transfected with miR-325 mimic or NC mimic and then exposed to fresh gas (21% O 2 , 5% CO 2 , remainder N 2 ) alone or with the addition of 3.4% sevoflurane for 6 h. ( A , B ) Western blotting for Cleaved-Caspase-3, Bax, and Bcl-2 in 3.4% SEVO-exposed vs. control HCN-2 cells; representative images ( A ) and quantification ( B ). ( C , D ) Annexin V-FITC/PI staining and flow cytometry analysis; representative images ( C ) and quantification ( D ). **p
    Figure Legend Snippet: Overexpression of Mir-325-3p inhibited sevoflurane-induced apoptosis in HCN-2 neuronal cells. HCN-2 neuronal cells were transfected with miR-325 mimic or NC mimic and then exposed to fresh gas (21% O 2 , 5% CO 2 , remainder N 2 ) alone or with the addition of 3.4% sevoflurane for 6 h. ( A , B ) Western blotting for Cleaved-Caspase-3, Bax, and Bcl-2 in 3.4% SEVO-exposed vs. control HCN-2 cells; representative images ( A ) and quantification ( B ). ( C , D ) Annexin V-FITC/PI staining and flow cytometry analysis; representative images ( C ) and quantification ( D ). **p

    Techniques Used: Over Expression, Transfection, Western Blot, Staining, Flow Cytometry

    miR-325-3p, which suppressed Nupr1 translation, was downregulated by sevoflurane in neuronal cells. ( A ) RT-qPCR for 10 Nupr1-targeting miRNAs in HCN-2 cells exposed to fresh gas (21% O 2 , 5% CO 2 , remainder N 2 ) alone or with the addition of 3.4% sevoflurane for 6 h. ( B , C ) Bioinformatics analysis showing predicted binding of miR-325-3p to the 3'-UTR of rat ( B ) and human ( C ) Nupr1 mRNA. ( D ) Wild-type Nupr1 mRNA (Nupr1 3'-UTR) and Nupr1 mRNA with a mutation in the 3'-UTR miR-325-3p-binding site (Nupr1 3'-UTR mut) were prepared. A dual-luciferase reporter assay was performed using all combinations of miR-325-3p-modifying and Nupr1 3'-UTR plasmids. ( E ) MiR-325-3p was overexpressed using an miR-325-3p mimic in HCN-2 neuronal cells. Controls were transfected with NC mimic. miR-325-3p levels in these cells were measured using RT-qPCR. ( F , G ) Western blot ( F ) and RT-qPCR ( G ) for Nupr1 levels in miR-325-3p-modified HCN-2 cells. **p
    Figure Legend Snippet: miR-325-3p, which suppressed Nupr1 translation, was downregulated by sevoflurane in neuronal cells. ( A ) RT-qPCR for 10 Nupr1-targeting miRNAs in HCN-2 cells exposed to fresh gas (21% O 2 , 5% CO 2 , remainder N 2 ) alone or with the addition of 3.4% sevoflurane for 6 h. ( B , C ) Bioinformatics analysis showing predicted binding of miR-325-3p to the 3'-UTR of rat ( B ) and human ( C ) Nupr1 mRNA. ( D ) Wild-type Nupr1 mRNA (Nupr1 3'-UTR) and Nupr1 mRNA with a mutation in the 3'-UTR miR-325-3p-binding site (Nupr1 3'-UTR mut) were prepared. A dual-luciferase reporter assay was performed using all combinations of miR-325-3p-modifying and Nupr1 3'-UTR plasmids. ( E ) MiR-325-3p was overexpressed using an miR-325-3p mimic in HCN-2 neuronal cells. Controls were transfected with NC mimic. miR-325-3p levels in these cells were measured using RT-qPCR. ( F , G ) Western blot ( F ) and RT-qPCR ( G ) for Nupr1 levels in miR-325-3p-modified HCN-2 cells. **p

    Techniques Used: Quantitative RT-PCR, Binding Assay, Mutagenesis, Luciferase, Reporter Assay, Transfection, Western Blot, Modification

    11) Product Images from "MicroRNA-325-3p prevents sevoflurane-induced learning and memory impairment by inhibiting Nupr1 and C/EBPβ/IGFBP5 signaling in rats"

    Article Title: MicroRNA-325-3p prevents sevoflurane-induced learning and memory impairment by inhibiting Nupr1 and C/EBPβ/IGFBP5 signaling in rats

    Journal: Aging (Albany NY)

    doi: 10.18632/aging.102942

    Nupr1 knockdown attenuated sevoflurane-induced apoptosis and decreased C/EBPβ and IGFBP5 expression in HCN-2 neuronal cells. HCN-2 neuronal cells were transfected with NC or Nupr1 siRNA and then exposed to fresh gas (21% O 2 , 5% CO 2 , remainder N 2 ) alone or with the addition of 3.4% sevoflurane for 6 h. ( A , B ) Western blotting for Nupr1, C/EBPβ, and IGFBP5 in 3.4% SEVO-exposed vs. control HCN-2 cells; representative images ( A ) and quantification ( B ). ( C , D ) Western blotting for Cleaved-Caspase-3, Bax, and Bcl-2 in 3.4% SEVO-exposed vs. control HCN-2 cells; representative images (C) and quantification ( D ). ( E , F ) Annexin V-FITC/PI staining and flow cytometry analysis; representative images ( E ) and quantification ( F ). **p
    Figure Legend Snippet: Nupr1 knockdown attenuated sevoflurane-induced apoptosis and decreased C/EBPβ and IGFBP5 expression in HCN-2 neuronal cells. HCN-2 neuronal cells were transfected with NC or Nupr1 siRNA and then exposed to fresh gas (21% O 2 , 5% CO 2 , remainder N 2 ) alone or with the addition of 3.4% sevoflurane for 6 h. ( A , B ) Western blotting for Nupr1, C/EBPβ, and IGFBP5 in 3.4% SEVO-exposed vs. control HCN-2 cells; representative images ( A ) and quantification ( B ). ( C , D ) Western blotting for Cleaved-Caspase-3, Bax, and Bcl-2 in 3.4% SEVO-exposed vs. control HCN-2 cells; representative images (C) and quantification ( D ). ( E , F ) Annexin V-FITC/PI staining and flow cytometry analysis; representative images ( E ) and quantification ( F ). **p

    Techniques Used: Expressing, Transfection, Western Blot, Staining, Flow Cytometry

    Sevoflurane increased Nupr1, C/EBPβ, and IGFBP5 expression and induced apoptosis in HCN-2 neuronal cells. Identical numbers of HCN-2 neuronal cells were exposed to fresh gas (21% O 2 , 5% CO 2 , remainder N 2 ) alone or with the addition of 3.4% sevoflurane for 6 h and then subjected to analysis. ( A , B ) Western blotting for Cleaved-Caspase-3, Bax, and Bcl-2 in 3.4% SEVO-exposed vs. control HCN-2 cells; representative images ( A ) and quantification ( B ). ( C , D ) Annexin V-FITC/PI staining and flow cytometry analysis; representative images ( C ) and quantification ( D ). ( E , F ) Western blotting for Nupr1, C/EBPβ, and IGFBP5 in 3.4% SEVO-exposed vs. control HCN-2 cells; representative images ( E ) and quantification ( F ). **p
    Figure Legend Snippet: Sevoflurane increased Nupr1, C/EBPβ, and IGFBP5 expression and induced apoptosis in HCN-2 neuronal cells. Identical numbers of HCN-2 neuronal cells were exposed to fresh gas (21% O 2 , 5% CO 2 , remainder N 2 ) alone or with the addition of 3.4% sevoflurane for 6 h and then subjected to analysis. ( A , B ) Western blotting for Cleaved-Caspase-3, Bax, and Bcl-2 in 3.4% SEVO-exposed vs. control HCN-2 cells; representative images ( A ) and quantification ( B ). ( C , D ) Annexin V-FITC/PI staining and flow cytometry analysis; representative images ( C ) and quantification ( D ). ( E , F ) Western blotting for Nupr1, C/EBPβ, and IGFBP5 in 3.4% SEVO-exposed vs. control HCN-2 cells; representative images ( E ) and quantification ( F ). **p

    Techniques Used: Expressing, Western Blot, Staining, Flow Cytometry

    Overexpression of Mir-325-3p inhibited sevoflurane-induced apoptosis in HCN-2 neuronal cells. HCN-2 neuronal cells were transfected with miR-325 mimic or NC mimic and then exposed to fresh gas (21% O 2 , 5% CO 2 , remainder N 2 ) alone or with the addition of 3.4% sevoflurane for 6 h. ( A , B ) Western blotting for Cleaved-Caspase-3, Bax, and Bcl-2 in 3.4% SEVO-exposed vs. control HCN-2 cells; representative images ( A ) and quantification ( B ). ( C , D ) Annexin V-FITC/PI staining and flow cytometry analysis; representative images ( C ) and quantification ( D ). **p
    Figure Legend Snippet: Overexpression of Mir-325-3p inhibited sevoflurane-induced apoptosis in HCN-2 neuronal cells. HCN-2 neuronal cells were transfected with miR-325 mimic or NC mimic and then exposed to fresh gas (21% O 2 , 5% CO 2 , remainder N 2 ) alone or with the addition of 3.4% sevoflurane for 6 h. ( A , B ) Western blotting for Cleaved-Caspase-3, Bax, and Bcl-2 in 3.4% SEVO-exposed vs. control HCN-2 cells; representative images ( A ) and quantification ( B ). ( C , D ) Annexin V-FITC/PI staining and flow cytometry analysis; representative images ( C ) and quantification ( D ). **p

    Techniques Used: Over Expression, Transfection, Western Blot, Staining, Flow Cytometry

    miR-325-3p, which suppressed Nupr1 translation, was downregulated by sevoflurane in neuronal cells. ( A ) RT-qPCR for 10 Nupr1-targeting miRNAs in HCN-2 cells exposed to fresh gas (21% O 2 , 5% CO 2 , remainder N 2 ) alone or with the addition of 3.4% sevoflurane for 6 h. ( B , C ) Bioinformatics analysis showing predicted binding of miR-325-3p to the 3'-UTR of rat ( B ) and human ( C ) Nupr1 mRNA. ( D ) Wild-type Nupr1 mRNA (Nupr1 3'-UTR) and Nupr1 mRNA with a mutation in the 3'-UTR miR-325-3p-binding site (Nupr1 3'-UTR mut) were prepared. A dual-luciferase reporter assay was performed using all combinations of miR-325-3p-modifying and Nupr1 3'-UTR plasmids. ( E ) MiR-325-3p was overexpressed using an miR-325-3p mimic in HCN-2 neuronal cells. Controls were transfected with NC mimic. miR-325-3p levels in these cells were measured using RT-qPCR. ( F , G ) Western blot ( F ) and RT-qPCR ( G ) for Nupr1 levels in miR-325-3p-modified HCN-2 cells. **p
    Figure Legend Snippet: miR-325-3p, which suppressed Nupr1 translation, was downregulated by sevoflurane in neuronal cells. ( A ) RT-qPCR for 10 Nupr1-targeting miRNAs in HCN-2 cells exposed to fresh gas (21% O 2 , 5% CO 2 , remainder N 2 ) alone or with the addition of 3.4% sevoflurane for 6 h. ( B , C ) Bioinformatics analysis showing predicted binding of miR-325-3p to the 3'-UTR of rat ( B ) and human ( C ) Nupr1 mRNA. ( D ) Wild-type Nupr1 mRNA (Nupr1 3'-UTR) and Nupr1 mRNA with a mutation in the 3'-UTR miR-325-3p-binding site (Nupr1 3'-UTR mut) were prepared. A dual-luciferase reporter assay was performed using all combinations of miR-325-3p-modifying and Nupr1 3'-UTR plasmids. ( E ) MiR-325-3p was overexpressed using an miR-325-3p mimic in HCN-2 neuronal cells. Controls were transfected with NC mimic. miR-325-3p levels in these cells were measured using RT-qPCR. ( F , G ) Western blot ( F ) and RT-qPCR ( G ) for Nupr1 levels in miR-325-3p-modified HCN-2 cells. **p

    Techniques Used: Quantitative RT-PCR, Binding Assay, Mutagenesis, Luciferase, Reporter Assay, Transfection, Western Blot, Modification

    12) Product Images from "Disrupting the leukemia niche in the central nervous system attenuates leukemia chemoresistance"

    Article Title: Disrupting the leukemia niche in the central nervous system attenuates leukemia chemoresistance

    Journal: bioRxiv

    doi: 10.1101/750547

    Leukemia cells exhibit increased chemoresistance when co-cultured with meningeal cells. (A) Percent of NALM-6 leukemia cells adherent to primary human meningeal cells, retronectin (recombinant fibronectin fragment) positive control, or non-tissue culture treated well after 2 hours. (B-C) NALM-6 and Jurkat leukemia cells (B) or primary B-cell ALL cells (C) cultured in suspension or adherent to CNS-derived cells (primary human meningeal cells or the HCN-2 neuronal cell line) were treated with cytarabine 500 nM or methotrexate 500 nM for 48 hours and then apoptosis measured using annexin-V and viability staining and flow cytometry. (D) NALM-6 and Jurkat cells were cultured in either regular media or meningeal conditioned media (CM) and treated with cytarabine 500 nM or methotrexate 500 nM for 48 hours before apoptosis was measured using annexin-V staining and flow cytometry. For all graphs, data are the mean +/- SEM from three independent experiments and P : **,
    Figure Legend Snippet: Leukemia cells exhibit increased chemoresistance when co-cultured with meningeal cells. (A) Percent of NALM-6 leukemia cells adherent to primary human meningeal cells, retronectin (recombinant fibronectin fragment) positive control, or non-tissue culture treated well after 2 hours. (B-C) NALM-6 and Jurkat leukemia cells (B) or primary B-cell ALL cells (C) cultured in suspension or adherent to CNS-derived cells (primary human meningeal cells or the HCN-2 neuronal cell line) were treated with cytarabine 500 nM or methotrexate 500 nM for 48 hours and then apoptosis measured using annexin-V and viability staining and flow cytometry. (D) NALM-6 and Jurkat cells were cultured in either regular media or meningeal conditioned media (CM) and treated with cytarabine 500 nM or methotrexate 500 nM for 48 hours before apoptosis was measured using annexin-V staining and flow cytometry. For all graphs, data are the mean +/- SEM from three independent experiments and P : **,

    Techniques Used: Cell Culture, Recombinant, Positive Control, Derivative Assay, Staining, Flow Cytometry

    13) Product Images from "Disrupting the leukemia niche in the central nervous system attenuates leukemia chemoresistance"

    Article Title: Disrupting the leukemia niche in the central nervous system attenuates leukemia chemoresistance

    Journal: Haematologica

    doi: 10.3324/haematol.2019.230334

    Leukemia cells exhibit increased chemoresistance when co-cultured with meningeal cells. (A) Percent of NALM-6 leukemia cells adherent to primary human meningeal cells, retronectin (recombinant fibronectin fragment) positive control, or non-tissue culture treated well after 2 h. (B, C) NALM-6 and Jurkat leukemia cells (B) or primary B-cell acute lymphocytic leukemia cells (C) cultured in suspension or adherent to central nervous system-derived cells (primary human meningeal cells or the HCN-2 neuronal cell line) were treated with cytarabine 500 nM or methotrexate 500 nM for 48 h and then apoptosis was measured using annexin-V and viability staining and flow cytometry. (D) NALM-6 and Jurkat cells were cultured in either regular media or meningeal conditioned media (CM; 100%) and treated with cytarabine 500 nM or methotrexate 500 nM for 48 h before apoptosis was measured using annexin-V staining and flow cytometry. For all graphs, data are the mean ± standard error of mean from three independent experiments. ** P
    Figure Legend Snippet: Leukemia cells exhibit increased chemoresistance when co-cultured with meningeal cells. (A) Percent of NALM-6 leukemia cells adherent to primary human meningeal cells, retronectin (recombinant fibronectin fragment) positive control, or non-tissue culture treated well after 2 h. (B, C) NALM-6 and Jurkat leukemia cells (B) or primary B-cell acute lymphocytic leukemia cells (C) cultured in suspension or adherent to central nervous system-derived cells (primary human meningeal cells or the HCN-2 neuronal cell line) were treated with cytarabine 500 nM or methotrexate 500 nM for 48 h and then apoptosis was measured using annexin-V and viability staining and flow cytometry. (D) NALM-6 and Jurkat cells were cultured in either regular media or meningeal conditioned media (CM; 100%) and treated with cytarabine 500 nM or methotrexate 500 nM for 48 h before apoptosis was measured using annexin-V staining and flow cytometry. For all graphs, data are the mean ± standard error of mean from three independent experiments. ** P

    Techniques Used: Cell Culture, Recombinant, Positive Control, Derivative Assay, Staining, Flow Cytometry

    14) Product Images from "Chronic hyperglycemia regulates microglia polarization through ERK5"

    Article Title: Chronic hyperglycemia regulates microglia polarization through ERK5

    Journal: Aging (Albany NY)

    doi: 10.18632/aging.101770

    Concomitant alteration of ERK5 activation with polarization of HG-cultured microglia, which affects the apoptosis of neuronal cells cultured in hypoxic conditions. ( A ) Western blot for phosphorylated ERK5 (pERK5), compared to the total ERK5 levels in HG- or NG-cultured microglia in different time courses. ( B ) Western blot for phosphorylated Stat3 (pStat3), compared to the total ERK5 levels in HG- or NG-cultured microglia in different time courses. ( C ) Conditioned media (CM) from polarized microglia in different conditions and different time courses were added into a neuronal cell line, HCN-2, cultured in hypoxic conditions. ( D ) TUNEL assay on HCN-2 cells. N=5. *p
    Figure Legend Snippet: Concomitant alteration of ERK5 activation with polarization of HG-cultured microglia, which affects the apoptosis of neuronal cells cultured in hypoxic conditions. ( A ) Western blot for phosphorylated ERK5 (pERK5), compared to the total ERK5 levels in HG- or NG-cultured microglia in different time courses. ( B ) Western blot for phosphorylated Stat3 (pStat3), compared to the total ERK5 levels in HG- or NG-cultured microglia in different time courses. ( C ) Conditioned media (CM) from polarized microglia in different conditions and different time courses were added into a neuronal cell line, HCN-2, cultured in hypoxic conditions. ( D ) TUNEL assay on HCN-2 cells. N=5. *p

    Techniques Used: Activation Assay, Cell Culture, Western Blot, TUNEL Assay

    Suppressing ERK5 activation prevents HG-induced M2a polarization of microglia and induces earlier polarization of M2b-like followed by M1-like microglia. ( A ) BIX021895 (BIX) was applied to the HG-cultured microglia. ( B ) Western blot for pERK5, compared to the total ERK5 levels in HG- cultured microglia at different time courses, with or without BIX. ( C – I ) RT-qPCR for iNOS ( C ), TNF-α ( D ), IL-6 ( E ), IL-12 ( F ), IL-10 ( G ), Arg-1 ( H ) and CD206 ( I ) mRNA levels in HG- cultured microglia at different time points. ( J ) TUNEL assay on HCN-2 cells. N=5. *p
    Figure Legend Snippet: Suppressing ERK5 activation prevents HG-induced M2a polarization of microglia and induces earlier polarization of M2b-like followed by M1-like microglia. ( A ) BIX021895 (BIX) was applied to the HG-cultured microglia. ( B ) Western blot for pERK5, compared to the total ERK5 levels in HG- cultured microglia at different time courses, with or without BIX. ( C – I ) RT-qPCR for iNOS ( C ), TNF-α ( D ), IL-6 ( E ), IL-12 ( F ), IL-10 ( G ), Arg-1 ( H ) and CD206 ( I ) mRNA levels in HG- cultured microglia at different time points. ( J ) TUNEL assay on HCN-2 cells. N=5. *p

    Techniques Used: Activation Assay, Cell Culture, Western Blot, Quantitative RT-PCR, TUNEL Assay

    Sustained ERK5 activation maintains HG-induced M2a polarization of microglia. ( A ) HG-culturing of MEK5DD-transfected and control microglia. ( B ) Western blot for pERK5, compared to the total ERK5 levels in HG- cultured, MEK5DD-transfected and control microglia at different time courses. ( C – I ) RT-qPCR for iNOS ( C ), TNF-α ( D ), IL-6 ( E ), IL-12 ( F ), IL-10 ( G ), Arg-1 ( H ) and CD206 ( I ) mRNA levels in HG- cultured, MEK5DD-transfected and control microglia at different time points. ( J ) TUNEL assay on HCN-2 cells. N=5. *p
    Figure Legend Snippet: Sustained ERK5 activation maintains HG-induced M2a polarization of microglia. ( A ) HG-culturing of MEK5DD-transfected and control microglia. ( B ) Western blot for pERK5, compared to the total ERK5 levels in HG- cultured, MEK5DD-transfected and control microglia at different time courses. ( C – I ) RT-qPCR for iNOS ( C ), TNF-α ( D ), IL-6 ( E ), IL-12 ( F ), IL-10 ( G ), Arg-1 ( H ) and CD206 ( I ) mRNA levels in HG- cultured, MEK5DD-transfected and control microglia at different time points. ( J ) TUNEL assay on HCN-2 cells. N=5. *p

    Techniques Used: Activation Assay, Transfection, Western Blot, Cell Culture, Quantitative RT-PCR, TUNEL Assay

    15) Product Images from "Sevoflurane impairs learning and memory of the developing brain through post-transcriptional inhibition of CCNA2 via microRNA-19-3p"

    Article Title: Sevoflurane impairs learning and memory of the developing brain through post-transcriptional inhibition of CCNA2 via microRNA-19-3p

    Journal: Aging (Albany NY)

    doi: 10.18632/aging.101673

    SEVO upregulates miR-19-3p in neuron cells. ( A ) RT-qPCR for 18 CCNA2-targeting miRNAs in HCN-2 cells exposed to either 5% SEVO or CTL gas. ( B - C ) Bioinformatics analysis showed predictive binding of miR-19-3p onto 3’-UTR of CCNA2 mRNA of rat ( B ) and human ( C ). N=5.
    Figure Legend Snippet: SEVO upregulates miR-19-3p in neuron cells. ( A ) RT-qPCR for 18 CCNA2-targeting miRNAs in HCN-2 cells exposed to either 5% SEVO or CTL gas. ( B - C ) Bioinformatics analysis showed predictive binding of miR-19-3p onto 3’-UTR of CCNA2 mRNA of rat ( B ) and human ( C ). N=5.

    Techniques Used: Quantitative RT-PCR, CTL Assay, Binding Assay

    SEVO impairs neuron cell proliferation in vitro likely through decreasing CCNA2. Identical number of HCV-2 neural cells was exposed to different doses of SEVO for 48 hours and then subjected to analysis. ( A - B ) Ki-67+ staining on HCN-2 cells, shown by representative images ( A ), and by quantification ( B ). ( C ) The total cell number based on DAPI staining. ( D ) Western blotting for CDKs and cyclins in 5% SEVO-exposed HCN-2 cells vs gas-exposed HCN-2 cells. ( E ) Quantification of changes in CCNA2 protein. ( F ) RT-qPCR for CCNA2 mRNA. *p
    Figure Legend Snippet: SEVO impairs neuron cell proliferation in vitro likely through decreasing CCNA2. Identical number of HCV-2 neural cells was exposed to different doses of SEVO for 48 hours and then subjected to analysis. ( A - B ) Ki-67+ staining on HCN-2 cells, shown by representative images ( A ), and by quantification ( B ). ( C ) The total cell number based on DAPI staining. ( D ) Western blotting for CDKs and cyclins in 5% SEVO-exposed HCN-2 cells vs gas-exposed HCN-2 cells. ( E ) Quantification of changes in CCNA2 protein. ( F ) RT-qPCR for CCNA2 mRNA. *p

    Techniques Used: In Vitro, Staining, Western Blot, Quantitative RT-PCR

    Generation of AAVs that express as-miR-19-3p. ( A ) Schematic showing AAVs carrying as-miR-19-3p under the control of a CMV promoter and AAV carrying null under the control of a CMV promoter as a control. The viral backbone had a GFP reporter, which was co-expressed with the transgene, which allowed the transduced cells to be visualized by green fluorescence. ( B ) Transduced HCN-2 cell in culture. ( C ) RT-qPCR for miR-19-3p in AAV-as-miR-19-3p-transduced cells, un-transduced cells (UnT) and null-transduced cells. *p
    Figure Legend Snippet: Generation of AAVs that express as-miR-19-3p. ( A ) Schematic showing AAVs carrying as-miR-19-3p under the control of a CMV promoter and AAV carrying null under the control of a CMV promoter as a control. The viral backbone had a GFP reporter, which was co-expressed with the transgene, which allowed the transduced cells to be visualized by green fluorescence. ( B ) Transduced HCN-2 cell in culture. ( C ) RT-qPCR for miR-19-3p in AAV-as-miR-19-3p-transduced cells, un-transduced cells (UnT) and null-transduced cells. *p

    Techniques Used: Fluorescence, Quantitative RT-PCR

    MiR-19-3p targets CCNA2 to suppress its translation in neural cells. ( A ) MiR-19-3p was overexpressed by a plasmid carrying miR-19-3p or knocked down by a plasmid carrying as-miR-19-3p in a human neural cell line, HCN-2. Transfection with a null sequence was used as a control. RT-qPCR for miR-19-3p levels in these cells. ( B ) An intact 3'-UTR of CCNA2 mRNA (CCNA2 3'-UTR) and an 3'-UTR of CCNA2 mRNA with a mutant at miR-19-3p-binding site (CCNA2 3'-UTR mut) were prepared. A dual luciferase reporter assay was performed using combinations of one miR-19-3p-modifying plasmid and one CCNA2 3’-UTR plasmid. ( C - D ) RT-qPCR ( C ) and Western blot ( D ) for CCNA2 levels in miR-19-3p-modified HCN-2 cells. *p
    Figure Legend Snippet: MiR-19-3p targets CCNA2 to suppress its translation in neural cells. ( A ) MiR-19-3p was overexpressed by a plasmid carrying miR-19-3p or knocked down by a plasmid carrying as-miR-19-3p in a human neural cell line, HCN-2. Transfection with a null sequence was used as a control. RT-qPCR for miR-19-3p levels in these cells. ( B ) An intact 3'-UTR of CCNA2 mRNA (CCNA2 3'-UTR) and an 3'-UTR of CCNA2 mRNA with a mutant at miR-19-3p-binding site (CCNA2 3'-UTR mut) were prepared. A dual luciferase reporter assay was performed using combinations of one miR-19-3p-modifying plasmid and one CCNA2 3’-UTR plasmid. ( C - D ) RT-qPCR ( C ) and Western blot ( D ) for CCNA2 levels in miR-19-3p-modified HCN-2 cells. *p

    Techniques Used: Plasmid Preparation, Transfection, Sequencing, Quantitative RT-PCR, Mutagenesis, Binding Assay, Luciferase, Reporter Assay, Western Blot, Modification

    16) Product Images from "Sevoflurane impairs learning and memory of the developing brain through post-transcriptional inhibition of CCNA2 via microRNA-19-3p"

    Article Title: Sevoflurane impairs learning and memory of the developing brain through post-transcriptional inhibition of CCNA2 via microRNA-19-3p

    Journal: Aging (Albany NY)

    doi: 10.18632/aging.101673

    SEVO upregulates miR-19-3p in neuron cells. ( A ) RT-qPCR for 18 CCNA2-targeting miRNAs in HCN-2 cells exposed to either 5% SEVO or CTL gas. ( B - C ) Bioinformatics analysis showed predictive binding of miR-19-3p onto 3’-UTR of CCNA2 mRNA of rat ( B ) and human ( C ). N=5.
    Figure Legend Snippet: SEVO upregulates miR-19-3p in neuron cells. ( A ) RT-qPCR for 18 CCNA2-targeting miRNAs in HCN-2 cells exposed to either 5% SEVO or CTL gas. ( B - C ) Bioinformatics analysis showed predictive binding of miR-19-3p onto 3’-UTR of CCNA2 mRNA of rat ( B ) and human ( C ). N=5.

    Techniques Used: Quantitative RT-PCR, CTL Assay, Binding Assay

    SEVO impairs neuron cell proliferation in vitro likely through decreasing CCNA2. Identical number of HCV-2 neural cells was exposed to different doses of SEVO for 48 hours and then subjected to analysis. ( A - B ) Ki-67+ staining on HCN-2 cells, shown by representative images ( A ), and by quantification ( B ). ( C ) The total cell number based on DAPI staining. ( D ) Western blotting for CDKs and cyclins in 5% SEVO-exposed HCN-2 cells vs gas-exposed HCN-2 cells. ( E ) Quantification of changes in CCNA2 protein. ( F ) RT-qPCR for CCNA2 mRNA. *p
    Figure Legend Snippet: SEVO impairs neuron cell proliferation in vitro likely through decreasing CCNA2. Identical number of HCV-2 neural cells was exposed to different doses of SEVO for 48 hours and then subjected to analysis. ( A - B ) Ki-67+ staining on HCN-2 cells, shown by representative images ( A ), and by quantification ( B ). ( C ) The total cell number based on DAPI staining. ( D ) Western blotting for CDKs and cyclins in 5% SEVO-exposed HCN-2 cells vs gas-exposed HCN-2 cells. ( E ) Quantification of changes in CCNA2 protein. ( F ) RT-qPCR for CCNA2 mRNA. *p

    Techniques Used: In Vitro, Staining, Western Blot, Quantitative RT-PCR

    Generation of AAVs that express as-miR-19-3p. ( A ) Schematic showing AAVs carrying as-miR-19-3p under the control of a CMV promoter and AAV carrying null under the control of a CMV promoter as a control. The viral backbone had a GFP reporter, which was co-expressed with the transgene, which allowed the transduced cells to be visualized by green fluorescence. ( B ) Transduced HCN-2 cell in culture. ( C ) RT-qPCR for miR-19-3p in AAV-as-miR-19-3p-transduced cells, un-transduced cells (UnT) and null-transduced cells. *p
    Figure Legend Snippet: Generation of AAVs that express as-miR-19-3p. ( A ) Schematic showing AAVs carrying as-miR-19-3p under the control of a CMV promoter and AAV carrying null under the control of a CMV promoter as a control. The viral backbone had a GFP reporter, which was co-expressed with the transgene, which allowed the transduced cells to be visualized by green fluorescence. ( B ) Transduced HCN-2 cell in culture. ( C ) RT-qPCR for miR-19-3p in AAV-as-miR-19-3p-transduced cells, un-transduced cells (UnT) and null-transduced cells. *p

    Techniques Used: Fluorescence, Quantitative RT-PCR

    MiR-19-3p targets CCNA2 to suppress its translation in neural cells. ( A ) MiR-19-3p was overexpressed by a plasmid carrying miR-19-3p or knocked down by a plasmid carrying as-miR-19-3p in a human neural cell line, HCN-2. Transfection with a null sequence was used as a control. RT-qPCR for miR-19-3p levels in these cells. ( B ) An intact 3'-UTR of CCNA2 mRNA (CCNA2 3'-UTR) and an 3'-UTR of CCNA2 mRNA with a mutant at miR-19-3p-binding site (CCNA2 3'-UTR mut) were prepared. A dual luciferase reporter assay was performed using combinations of one miR-19-3p-modifying plasmid and one CCNA2 3’-UTR plasmid. ( C - D ) RT-qPCR ( C ) and Western blot ( D ) for CCNA2 levels in miR-19-3p-modified HCN-2 cells. *p
    Figure Legend Snippet: MiR-19-3p targets CCNA2 to suppress its translation in neural cells. ( A ) MiR-19-3p was overexpressed by a plasmid carrying miR-19-3p or knocked down by a plasmid carrying as-miR-19-3p in a human neural cell line, HCN-2. Transfection with a null sequence was used as a control. RT-qPCR for miR-19-3p levels in these cells. ( B ) An intact 3'-UTR of CCNA2 mRNA (CCNA2 3'-UTR) and an 3'-UTR of CCNA2 mRNA with a mutant at miR-19-3p-binding site (CCNA2 3'-UTR mut) were prepared. A dual luciferase reporter assay was performed using combinations of one miR-19-3p-modifying plasmid and one CCNA2 3’-UTR plasmid. ( C - D ) RT-qPCR ( C ) and Western blot ( D ) for CCNA2 levels in miR-19-3p-modified HCN-2 cells. *p

    Techniques Used: Plasmid Preparation, Transfection, Sequencing, Quantitative RT-PCR, Mutagenesis, Binding Assay, Luciferase, Reporter Assay, Western Blot, Modification

    17) Product Images from "Long non-coding RNA linc-ITGB1 promotes cell proliferation, migration, and invasion in human hepatoma carcinoma by up-regulating ROCK1"

    Article Title: Long non-coding RNA linc-ITGB1 promotes cell proliferation, migration, and invasion in human hepatoma carcinoma by up-regulating ROCK1

    Journal: Bioscience Reports

    doi: 10.1042/BSR20181289

    Effects of linc-ITGB1 knockdown on HCC cell proliferation, migration, and invasion This figure shows proliferation ( A ), migration ( B ), and invasion ( C ) of two HCC cell lines C3A and HEP G2 and a normal liver tissue cell line THLE-3 under linc-ITGB1 knockdown. Linc-ITGB1 knockdown significantly inhibited cancer cell proliferation, migration, and invasion, but showed no significant effects on cells of normal liver tissue cell line THLE-3. *, P
    Figure Legend Snippet: Effects of linc-ITGB1 knockdown on HCC cell proliferation, migration, and invasion This figure shows proliferation ( A ), migration ( B ), and invasion ( C ) of two HCC cell lines C3A and HEP G2 and a normal liver tissue cell line THLE-3 under linc-ITGB1 knockdown. Linc-ITGB1 knockdown significantly inhibited cancer cell proliferation, migration, and invasion, but showed no significant effects on cells of normal liver tissue cell line THLE-3. *, P

    Techniques Used: Migration

    Effects of linc-ITGB1 overexpression and knockdown on ROCK1 expression Linc-ITGB1 overexpression and knockdown were reached after transfection ( A ). Linc-ITGB1 overexpression significantly up-regulated ( B ), while linc-ITGB1 knockdown ( C ) significantly down-regulated the expression of ROCK1 in cells of HCC cell lines C3A and HEP G2 but not in normal liver tissue cell line THLE-3. *, P
    Figure Legend Snippet: Effects of linc-ITGB1 overexpression and knockdown on ROCK1 expression Linc-ITGB1 overexpression and knockdown were reached after transfection ( A ). Linc-ITGB1 overexpression significantly up-regulated ( B ), while linc-ITGB1 knockdown ( C ) significantly down-regulated the expression of ROCK1 in cells of HCC cell lines C3A and HEP G2 but not in normal liver tissue cell line THLE-3. *, P

    Techniques Used: Over Expression, Expressing, Transfection

    Effects of linc-ITGB1 overexpression on HCC cell proliferation, migration, and invasion This figure shows proliferation ( A ), migration ( B ), and invasion ( C ) of two HCC cell lines C3A and HEP G2 and a normal liver tissue cell line THLE-3 under linc-ITGB1 overexpression and ROCK1 inhibitor treatment. Linc-ITGB1 overexpression significantly promoted the proliferation, migration, and invasion of HCC cell lines C3A and HEP G2 but not normal cell line THLE-3. Treatment with Stemolecule™ ROCK I Inhibitor significantly reduced the enhancing effects of linc-ITGB1 overexpression on cancer cell proliferation, migration, and invasion of HCC cells. *, P
    Figure Legend Snippet: Effects of linc-ITGB1 overexpression on HCC cell proliferation, migration, and invasion This figure shows proliferation ( A ), migration ( B ), and invasion ( C ) of two HCC cell lines C3A and HEP G2 and a normal liver tissue cell line THLE-3 under linc-ITGB1 overexpression and ROCK1 inhibitor treatment. Linc-ITGB1 overexpression significantly promoted the proliferation, migration, and invasion of HCC cell lines C3A and HEP G2 but not normal cell line THLE-3. Treatment with Stemolecule™ ROCK I Inhibitor significantly reduced the enhancing effects of linc-ITGB1 overexpression on cancer cell proliferation, migration, and invasion of HCC cells. *, P

    Techniques Used: Over Expression, Migration

    18) Product Images from "Long non-coding RNA linc-ITGB1 promotes cell proliferation, migration, and invasion in human hepatoma carcinoma by up-regulating ROCK1"

    Article Title: Long non-coding RNA linc-ITGB1 promotes cell proliferation, migration, and invasion in human hepatoma carcinoma by up-regulating ROCK1

    Journal: Bioscience Reports

    doi: 10.1042/BSR20181289

    Effects of linc-ITGB1 knockdown on HCC cell proliferation, migration, and invasion This figure shows proliferation ( A ), migration ( B ), and invasion ( C ) of two HCC cell lines C3A and HEP G2 and a normal liver tissue cell line THLE-3 under linc-ITGB1 knockdown. Linc-ITGB1 knockdown significantly inhibited cancer cell proliferation, migration, and invasion, but showed no significant effects on cells of normal liver tissue cell line THLE-3. *, P
    Figure Legend Snippet: Effects of linc-ITGB1 knockdown on HCC cell proliferation, migration, and invasion This figure shows proliferation ( A ), migration ( B ), and invasion ( C ) of two HCC cell lines C3A and HEP G2 and a normal liver tissue cell line THLE-3 under linc-ITGB1 knockdown. Linc-ITGB1 knockdown significantly inhibited cancer cell proliferation, migration, and invasion, but showed no significant effects on cells of normal liver tissue cell line THLE-3. *, P

    Techniques Used: Migration

    Effects of linc-ITGB1 overexpression and knockdown on ROCK1 expression Linc-ITGB1 overexpression and knockdown were reached after transfection ( A ). Linc-ITGB1 overexpression significantly up-regulated ( B ), while linc-ITGB1 knockdown ( C ) significantly down-regulated the expression of ROCK1 in cells of HCC cell lines C3A and HEP G2 but not in normal liver tissue cell line THLE-3. *, P
    Figure Legend Snippet: Effects of linc-ITGB1 overexpression and knockdown on ROCK1 expression Linc-ITGB1 overexpression and knockdown were reached after transfection ( A ). Linc-ITGB1 overexpression significantly up-regulated ( B ), while linc-ITGB1 knockdown ( C ) significantly down-regulated the expression of ROCK1 in cells of HCC cell lines C3A and HEP G2 but not in normal liver tissue cell line THLE-3. *, P

    Techniques Used: Over Expression, Expressing, Transfection

    Effects of linc-ITGB1 overexpression on HCC cell proliferation, migration, and invasion This figure shows proliferation ( A ), migration ( B ), and invasion ( C ) of two HCC cell lines C3A and HEP G2 and a normal liver tissue cell line THLE-3 under linc-ITGB1 overexpression and ROCK1 inhibitor treatment. Linc-ITGB1 overexpression significantly promoted the proliferation, migration, and invasion of HCC cell lines C3A and HEP G2 but not normal cell line THLE-3. Treatment with Stemolecule™ ROCK I Inhibitor significantly reduced the enhancing effects of linc-ITGB1 overexpression on cancer cell proliferation, migration, and invasion of HCC cells. *, P
    Figure Legend Snippet: Effects of linc-ITGB1 overexpression on HCC cell proliferation, migration, and invasion This figure shows proliferation ( A ), migration ( B ), and invasion ( C ) of two HCC cell lines C3A and HEP G2 and a normal liver tissue cell line THLE-3 under linc-ITGB1 overexpression and ROCK1 inhibitor treatment. Linc-ITGB1 overexpression significantly promoted the proliferation, migration, and invasion of HCC cell lines C3A and HEP G2 but not normal cell line THLE-3. Treatment with Stemolecule™ ROCK I Inhibitor significantly reduced the enhancing effects of linc-ITGB1 overexpression on cancer cell proliferation, migration, and invasion of HCC cells. *, P

    Techniques Used: Over Expression, Migration

    19) Product Images from "MicroRNA-182 targets protein phosphatase 1 regulatory inhibitor subunit 1C in glioblastoma"

    Article Title: MicroRNA-182 targets protein phosphatase 1 regulatory inhibitor subunit 1C in glioblastoma

    Journal: Oncotarget

    doi: 10.18632/oncotarget.21309

    PPP1R1C is a bona-fide target of miR-182 in the glioblastoma cell line U87-MG (A) Steady state expression of miR-182 and PPP1R1C in U87-MG and HCN-2 cell lines were determined. Data was normalized to RNU6B and TBP expression, respectively. Fold expression in U87-MG cells was determined relative to expression in HCN-2 cell line. (B) Relative luciferase activity of transiently transfected luciferase reporter constructs containing either full-length or mutated (miR-182 binding site deleted) PPP1R1C 3′ UTR in U87-MG and HCN-2 cells, either mock transfected or transfected with miR-182 mimic or miR-182 antagomir. * p
    Figure Legend Snippet: PPP1R1C is a bona-fide target of miR-182 in the glioblastoma cell line U87-MG (A) Steady state expression of miR-182 and PPP1R1C in U87-MG and HCN-2 cell lines were determined. Data was normalized to RNU6B and TBP expression, respectively. Fold expression in U87-MG cells was determined relative to expression in HCN-2 cell line. (B) Relative luciferase activity of transiently transfected luciferase reporter constructs containing either full-length or mutated (miR-182 binding site deleted) PPP1R1C 3′ UTR in U87-MG and HCN-2 cells, either mock transfected or transfected with miR-182 mimic or miR-182 antagomir. * p

    Techniques Used: Expressing, Luciferase, Activity Assay, Transfection, Construct, Binding Assay

    20) Product Images from "MicroRNA-182 targets protein phosphatase 1 regulatory inhibitor subunit 1C in glioblastoma"

    Article Title: MicroRNA-182 targets protein phosphatase 1 regulatory inhibitor subunit 1C in glioblastoma

    Journal: Oncotarget

    doi: 10.18632/oncotarget.21309

    PPP1R1C is a bona-fide target of miR-182 in the glioblastoma cell line U87-MG (A) Steady state expression of miR-182 and PPP1R1C in U87-MG and HCN-2 cell lines were determined. Data was normalized to RNU6B and TBP expression, respectively. Fold expression in U87-MG cells was determined relative to expression in HCN-2 cell line. (B) Relative luciferase activity of transiently transfected luciferase reporter constructs containing either full-length or mutated (miR-182 binding site deleted) PPP1R1C 3′ UTR in U87-MG and HCN-2 cells, either mock transfected or transfected with miR-182 mimic or miR-182 antagomir. * p
    Figure Legend Snippet: PPP1R1C is a bona-fide target of miR-182 in the glioblastoma cell line U87-MG (A) Steady state expression of miR-182 and PPP1R1C in U87-MG and HCN-2 cell lines were determined. Data was normalized to RNU6B and TBP expression, respectively. Fold expression in U87-MG cells was determined relative to expression in HCN-2 cell line. (B) Relative luciferase activity of transiently transfected luciferase reporter constructs containing either full-length or mutated (miR-182 binding site deleted) PPP1R1C 3′ UTR in U87-MG and HCN-2 cells, either mock transfected or transfected with miR-182 mimic or miR-182 antagomir. * p

    Techniques Used: Expressing, Luciferase, Activity Assay, Transfection, Construct, Binding Assay

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    ATCC hcn2 neuronal cells
    GRP78 plays a key role in regulating the expression of APP and Tau after exposure of <t>neuronal</t> <t>cells</t> and microglia to AR12 and neratinib. <t>HCN2</t> and BV2 cells were transfected with a scrambled siRNA or with siRNA molecules to knock down the expression of GRP78, HSP70 or HSP90 and in parallel co-transfected to express Tau or APP. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of APP, Tau and ERK2. (n = 3 +/-SD) * p
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    GRP78 plays a key role in regulating the expression of APP and Tau after exposure of neuronal cells and microglia to AR12 and neratinib. HCN2 and BV2 cells were transfected with a scrambled siRNA or with siRNA molecules to knock down the expression of GRP78, HSP70 or HSP90 and in parallel co-transfected to express Tau or APP. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of APP, Tau and ERK2. (n = 3 +/-SD) * p

    Journal: Aging (Albany NY)

    Article Title: AR12 increases BAG3 expression which is essential for Tau and APP degradation via LC3-associated phagocytosis and macroautophagy

    doi: 10.18632/aging.204337

    Figure Lengend Snippet: GRP78 plays a key role in regulating the expression of APP and Tau after exposure of neuronal cells and microglia to AR12 and neratinib. HCN2 and BV2 cells were transfected with a scrambled siRNA or with siRNA molecules to knock down the expression of GRP78, HSP70 or HSP90 and in parallel co-transfected to express Tau or APP. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of APP, Tau and ERK2. (n = 3 +/-SD) * p

    Article Snippet: HCT116 colon cancer cells and HCN2 neuronal cells were purchased from the ATCC (Bethesda, MD).

    Techniques: Expressing, Transfection, Immunostaining

    Knock down of Beclin1 or ATG5 prevents autophagosome formation and autophagic flux in HCN2 cells. HCN2 neuronal cells were transfected with a scrambled siRNA or with an siRNA to knock down the expression of ATG5 or Beclin1 and were co-transfected with a plasmid to express LC3-GFP-RFP. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 4h and 8h. The mean number of intense GFP+RFP+ and RFP+ punctae per cell was determined (n = 3 +/-SD) # p

    Journal: Aging (Albany NY)

    Article Title: AR12 increases BAG3 expression which is essential for Tau and APP degradation via LC3-associated phagocytosis and macroautophagy

    doi: 10.18632/aging.204337

    Figure Lengend Snippet: Knock down of Beclin1 or ATG5 prevents autophagosome formation and autophagic flux in HCN2 cells. HCN2 neuronal cells were transfected with a scrambled siRNA or with an siRNA to knock down the expression of ATG5 or Beclin1 and were co-transfected with a plasmid to express LC3-GFP-RFP. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 4h and 8h. The mean number of intense GFP+RFP+ and RFP+ punctae per cell was determined (n = 3 +/-SD) # p

    Article Snippet: HCT116 colon cancer cells and HCN2 neuronal cells were purchased from the ATCC (Bethesda, MD).

    Techniques: Transfection, Expressing, Plasmid Preparation

    AR12 and neratinib reduce chaperone expression in neuronal cells. HCN2 neuronal cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of GRP78 (total and cell surface), HSP70, HSP90, eIF2α and ERK2, and the phosphorylation of eIF2α S51. (n = 3 +/-SD) * p

    Journal: Aging (Albany NY)

    Article Title: AR12 increases BAG3 expression which is essential for Tau and APP degradation via LC3-associated phagocytosis and macroautophagy

    doi: 10.18632/aging.204337

    Figure Lengend Snippet: AR12 and neratinib reduce chaperone expression in neuronal cells. HCN2 neuronal cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of GRP78 (total and cell surface), HSP70, HSP90, eIF2α and ERK2, and the phosphorylation of eIF2α S51. (n = 3 +/-SD) * p

    Article Snippet: HCT116 colon cancer cells and HCN2 neuronal cells were purchased from the ATCC (Bethesda, MD).

    Techniques: Expressing, Immunostaining

    Enhanced BAG3 expression requires autophagy and ER stress signaling. HCN2 cells were transfected with a scrambled siRNA control or with siRNA molecules to knock down the expression of Beclin1, ATG5, ULK1, eIF2α or PERK. After 24h, cells were treated for 6h with vehicle control or with AR12 (2 μM). Cells were fixed in place and immunostaining performed to detect the expression of BAG3 and ERK2 (n = 3 +/-SD) # p

    Journal: Aging (Albany NY)

    Article Title: AR12 increases BAG3 expression which is essential for Tau and APP degradation via LC3-associated phagocytosis and macroautophagy

    doi: 10.18632/aging.204337

    Figure Lengend Snippet: Enhanced BAG3 expression requires autophagy and ER stress signaling. HCN2 cells were transfected with a scrambled siRNA control or with siRNA molecules to knock down the expression of Beclin1, ATG5, ULK1, eIF2α or PERK. After 24h, cells were treated for 6h with vehicle control or with AR12 (2 μM). Cells were fixed in place and immunostaining performed to detect the expression of BAG3 and ERK2 (n = 3 +/-SD) # p

    Article Snippet: HCT116 colon cancer cells and HCN2 neuronal cells were purchased from the ATCC (Bethesda, MD).

    Techniques: Expressing, Transfection, Immunostaining

    BAG3 is essential for drug-induced degradation of GRP78. ( A – C ) HCN2 cells were transfected with a scrambled siRNA or with an siRNA to knock down BAG3 expression. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of GRP78 (total and cell surface), HDAC6, HSP90, p62, LAMP2, PERK, P-PERK T980, eIF2α and P-eIF2α S51 and ERK2. (n = 3 +/-SD) * p

    Journal: Aging (Albany NY)

    Article Title: AR12 increases BAG3 expression which is essential for Tau and APP degradation via LC3-associated phagocytosis and macroautophagy

    doi: 10.18632/aging.204337

    Figure Lengend Snippet: BAG3 is essential for drug-induced degradation of GRP78. ( A – C ) HCN2 cells were transfected with a scrambled siRNA or with an siRNA to knock down BAG3 expression. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of GRP78 (total and cell surface), HDAC6, HSP90, p62, LAMP2, PERK, P-PERK T980, eIF2α and P-eIF2α S51 and ERK2. (n = 3 +/-SD) * p

    Article Snippet: HCT116 colon cancer cells and HCN2 neuronal cells were purchased from the ATCC (Bethesda, MD).

    Techniques: Transfection, Expressing, Immunostaining

    ER stress signaling plays a key role in facilitating autophagy and protein degradation in neuronal cells. Upper Graphs. HCN2 cells and BV2 cells were transfected with a scrambled siRNA or with an siRNA to knock down the expression of eIF2α and were co-transfected with a plasmid to express LC3-GFP-RFP. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 4h and 8h. The mean number of intense staining GFP+RFP+ and RFP+ punctae per cell was determined (n = 3 +/-SD) # p

    Journal: Aging (Albany NY)

    Article Title: AR12 increases BAG3 expression which is essential for Tau and APP degradation via LC3-associated phagocytosis and macroautophagy

    doi: 10.18632/aging.204337

    Figure Lengend Snippet: ER stress signaling plays a key role in facilitating autophagy and protein degradation in neuronal cells. Upper Graphs. HCN2 cells and BV2 cells were transfected with a scrambled siRNA or with an siRNA to knock down the expression of eIF2α and were co-transfected with a plasmid to express LC3-GFP-RFP. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 4h and 8h. The mean number of intense staining GFP+RFP+ and RFP+ punctae per cell was determined (n = 3 +/-SD) # p

    Article Snippet: HCT116 colon cancer cells and HCN2 neuronal cells were purchased from the ATCC (Bethesda, MD).

    Techniques: Transfection, Expressing, Plasmid Preparation, Staining

    Over-expression of GRP78 suppresses the drug-induced expression of BAG3. ( A ) HCN2 cells were transfected with a scrambled siRNA or with siRNA molecules to knock down the expression of GRP78, HSP70 or HSP90. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of BAG3 and ERK2. (n = 3 +/-SD) # p

    Journal: Aging (Albany NY)

    Article Title: AR12 increases BAG3 expression which is essential for Tau and APP degradation via LC3-associated phagocytosis and macroautophagy

    doi: 10.18632/aging.204337

    Figure Lengend Snippet: Over-expression of GRP78 suppresses the drug-induced expression of BAG3. ( A ) HCN2 cells were transfected with a scrambled siRNA or with siRNA molecules to knock down the expression of GRP78, HSP70 or HSP90. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of BAG3 and ERK2. (n = 3 +/-SD) # p

    Article Snippet: HCT116 colon cancer cells and HCN2 neuronal cells were purchased from the ATCC (Bethesda, MD).

    Techniques: Over Expression, Expressing, Transfection, Immunostaining

    ER stress signaling plays a key role in facilitating autophagy and HDAC6 protein degradation. ( A ) HCN2 cells were transfected with a scrambled siRNA or with an siRNA molecule to knock down the expression of eIF2α. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of HDAC6, LAMP2, p62 and ERK2. (n = 3 +/-SD) * p

    Journal: Aging (Albany NY)

    Article Title: AR12 increases BAG3 expression which is essential for Tau and APP degradation via LC3-associated phagocytosis and macroautophagy

    doi: 10.18632/aging.204337

    Figure Lengend Snippet: ER stress signaling plays a key role in facilitating autophagy and HDAC6 protein degradation. ( A ) HCN2 cells were transfected with a scrambled siRNA or with an siRNA molecule to knock down the expression of eIF2α. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of HDAC6, LAMP2, p62 and ERK2. (n = 3 +/-SD) * p

    Article Snippet: HCT116 colon cancer cells and HCN2 neuronal cells were purchased from the ATCC (Bethesda, MD).

    Techniques: Transfection, Expressing, Immunostaining

    Knock down of Rubicon suppresses drug-induced autophagosome formation but does not appear to alter autophagic flux. HCN2 neuronal cells and BV2 microglial cells were transfected with a scrambled siRNA or with an siRNA to knock down the expression of Rubicon and were co-transfected with a plasmid to express LC3-GFP-RFP. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 4h and 8h. The mean number of intense GFP+RFP+ and RFP+ punctae per cell was determined (n = 3 +/-SD) # p

    Journal: Aging (Albany NY)

    Article Title: AR12 increases BAG3 expression which is essential for Tau and APP degradation via LC3-associated phagocytosis and macroautophagy

    doi: 10.18632/aging.204337

    Figure Lengend Snippet: Knock down of Rubicon suppresses drug-induced autophagosome formation but does not appear to alter autophagic flux. HCN2 neuronal cells and BV2 microglial cells were transfected with a scrambled siRNA or with an siRNA to knock down the expression of Rubicon and were co-transfected with a plasmid to express LC3-GFP-RFP. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 4h and 8h. The mean number of intense GFP+RFP+ and RFP+ punctae per cell was determined (n = 3 +/-SD) # p

    Article Snippet: HCT116 colon cancer cells and HCN2 neuronal cells were purchased from the ATCC (Bethesda, MD).

    Techniques: Transfection, Expressing, Plasmid Preparation

    AR12 and the drug combination reduce the expression of mutant forms of TAU and APP. HCN2 and BV2 cells were transfected with plasmids to express wild type TAU or TAU 301L. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of TAU, TAU 301L and ERK2. (n = 3 +/-SD) * p

    Journal: Aging (Albany NY)

    Article Title: AR12 increases BAG3 expression which is essential for Tau and APP degradation via LC3-associated phagocytosis and macroautophagy

    doi: 10.18632/aging.204337

    Figure Lengend Snippet: AR12 and the drug combination reduce the expression of mutant forms of TAU and APP. HCN2 and BV2 cells were transfected with plasmids to express wild type TAU or TAU 301L. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of TAU, TAU 301L and ERK2. (n = 3 +/-SD) * p

    Article Snippet: HCT116 colon cancer cells and HCN2 neuronal cells were purchased from the ATCC (Bethesda, MD).

    Techniques: Expressing, Mutagenesis, Transfection, Immunostaining

    Degradation of APP and Tau by AR12 and neratinib requires LAP and macroautophagy in HCN2 neuronal cells and in BV2 microglia. ( A , B ) HCN2 cells were transfected with a scrambled siRNA or with siRNA molecules to knock down expression of Rubicon, Beclin1 or ATG5. In parallel, cells were transfected with an empty vector plasmid in ( B ), or transfected with plasmids to express APP or Tau in ( A ). After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of Tau, APP and ERK2. (n = 3 +/-SD) Endogenous expression of APP was 5% of the value for APP expressed from a plasmid. Endogenous expression of Tau was 6% of the value for Tau expressed from a plasmid. * p

    Journal: Aging (Albany NY)

    Article Title: AR12 increases BAG3 expression which is essential for Tau and APP degradation via LC3-associated phagocytosis and macroautophagy

    doi: 10.18632/aging.204337

    Figure Lengend Snippet: Degradation of APP and Tau by AR12 and neratinib requires LAP and macroautophagy in HCN2 neuronal cells and in BV2 microglia. ( A , B ) HCN2 cells were transfected with a scrambled siRNA or with siRNA molecules to knock down expression of Rubicon, Beclin1 or ATG5. In parallel, cells were transfected with an empty vector plasmid in ( B ), or transfected with plasmids to express APP or Tau in ( A ). After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of Tau, APP and ERK2. (n = 3 +/-SD) Endogenous expression of APP was 5% of the value for APP expressed from a plasmid. Endogenous expression of Tau was 6% of the value for Tau expressed from a plasmid. * p

    Article Snippet: HCT116 colon cancer cells and HCN2 neuronal cells were purchased from the ATCC (Bethesda, MD).

    Techniques: Transfection, Expressing, Plasmid Preparation, Immunostaining

    Expression of Tau or APP does not significantly alter the regulation of protein phosphorylation or protein expression caused by AR12 and neratinib. HCN2 cells were transfected with an empty vector plasmid or with plasmids to express Tau or APP. Twenty-four h afterwards, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the phosphorylation and expression of the indicated proteins (n = 3 +/-SD). * p

    Journal: Aging (Albany NY)

    Article Title: AR12 increases BAG3 expression which is essential for Tau and APP degradation via LC3-associated phagocytosis and macroautophagy

    doi: 10.18632/aging.204337

    Figure Lengend Snippet: Expression of Tau or APP does not significantly alter the regulation of protein phosphorylation or protein expression caused by AR12 and neratinib. HCN2 cells were transfected with an empty vector plasmid or with plasmids to express Tau or APP. Twenty-four h afterwards, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the phosphorylation and expression of the indicated proteins (n = 3 +/-SD). * p

    Article Snippet: HCT116 colon cancer cells and HCN2 neuronal cells were purchased from the ATCC (Bethesda, MD).

    Techniques: Expressing, Transfection, Plasmid Preparation, Immunostaining

    Control data showing protein expression knock down and protein over-expression. Left: BV2 and HCN2 cells as indicated were transfected with siRNA molecules to knock down the expression of the indicated proteins or transfected with plasmids to over-express the indicated proteins. The percentage remaining after knock-down or the percentage over-expression above basal levels is indicated. (n = 3 +/-SD) (total ERK2 is included as an invariant total protein loading control). Right: Images of HCN2 cells transfected to express TAU or APP.

    Journal: Aging (Albany NY)

    Article Title: AR12 increases BAG3 expression which is essential for Tau and APP degradation via LC3-associated phagocytosis and macroautophagy

    doi: 10.18632/aging.204337

    Figure Lengend Snippet: Control data showing protein expression knock down and protein over-expression. Left: BV2 and HCN2 cells as indicated were transfected with siRNA molecules to knock down the expression of the indicated proteins or transfected with plasmids to over-express the indicated proteins. The percentage remaining after knock-down or the percentage over-expression above basal levels is indicated. (n = 3 +/-SD) (total ERK2 is included as an invariant total protein loading control). Right: Images of HCN2 cells transfected to express TAU or APP.

    Article Snippet: HCT116 colon cancer cells and HCN2 neuronal cells were purchased from the ATCC (Bethesda, MD).

    Techniques: Expressing, Over Expression, Transfection

    Degradation of chaperones and eIF2α S51 phosphorylation requires LAP and macroautophagy. ( A ) HCN2 and ( B ) BV2 cells were transfected with a scrambled siRNA or with siRNA molecules to knock down the expression of Rubicon, Beclin1 or ATG5. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of GRP78 (total and cell surface / plasma membrane), HSP70, HSP90, eIF2α and ERK2, and the phosphorylation of eIF2α S51. (n = 3 +/-SD) * p

    Journal: Aging (Albany NY)

    Article Title: AR12 increases BAG3 expression which is essential for Tau and APP degradation via LC3-associated phagocytosis and macroautophagy

    doi: 10.18632/aging.204337

    Figure Lengend Snippet: Degradation of chaperones and eIF2α S51 phosphorylation requires LAP and macroautophagy. ( A ) HCN2 and ( B ) BV2 cells were transfected with a scrambled siRNA or with siRNA molecules to knock down the expression of Rubicon, Beclin1 or ATG5. After 24h, cells were treated with vehicle control, AR12 (2 μM), neratinib (50 nM) or the drugs in combination for 6h. Cells were fixed in place and immunostaining performed to determine the expression of GRP78 (total and cell surface / plasma membrane), HSP70, HSP90, eIF2α and ERK2, and the phosphorylation of eIF2α S51. (n = 3 +/-SD) * p

    Article Snippet: HCT116 colon cancer cells and HCN2 neuronal cells were purchased from the ATCC (Bethesda, MD).

    Techniques: Transfection, Expressing, Immunostaining

    AR12 and neratinib cause degradation of p70 S6K in HCN2 neuronal cells. ( A ) HCN2 cells were transfected with a scrambled siRNA control or with an siRNA to knock down expression of AMPKa. After 24h, cells were treated with vehicle control or with [AR12 (2 μM) + neratinib (50 nM)] for 6h. Cells were fixed in place and immunostaining performed to determine the phosphorylation and expression of the indicated proteins (n = 3 +/-SD). * p

    Journal: Aging (Albany NY)

    Article Title: AR12 increases BAG3 expression which is essential for Tau and APP degradation via LC3-associated phagocytosis and macroautophagy

    doi: 10.18632/aging.204337

    Figure Lengend Snippet: AR12 and neratinib cause degradation of p70 S6K in HCN2 neuronal cells. ( A ) HCN2 cells were transfected with a scrambled siRNA control or with an siRNA to knock down expression of AMPKa. After 24h, cells were treated with vehicle control or with [AR12 (2 μM) + neratinib (50 nM)] for 6h. Cells were fixed in place and immunostaining performed to determine the phosphorylation and expression of the indicated proteins (n = 3 +/-SD). * p

    Article Snippet: HCT116 colon cancer cells and HCN2 neuronal cells were purchased from the ATCC (Bethesda, MD).

    Techniques: Transfection, Expressing, Immunostaining

    Illustration and comparisons of the differentiation patterns of HCN-2 cells transfected with miRNA mimic. a HCN-2 cells were transfected with scrambled control mimic (control), hsa-miR-30e-5p mimic (miR-30e), hsa-miR-140-3p mimic (miR-140), or hsa-miR-126-5p mimic (miR-126) for 24 h. Then, they were seeded on new plates for monitoring and recording growth with a camera. Each transfection treatment had three independent wells, and three pictures were randomly taken from each well, resulting in 3*3 = 9 pictures. ( b – f ) We determined the differentiation patterns of HCN-2 cells in terms of different indices, including ( b ) vessel area, ( c ) vessel length, ( d ) junction number, and ( e ) mean lacunarity. The t -test p values were calculated by comparing the values of the control set on a specific day. f For a systematic comparison, the values were normalized to the average values of the control set at day 9. *, **, *** and **** denote p value

    Journal: Translational Psychiatry

    Article Title: MicroRNAs serve as prediction and treatment-response biomarkers of attention-deficit/hyperactivity disorder and promote the differentiation of neuronal cells by repressing the apoptosis pathway

    doi: 10.1038/s41398-022-01832-1

    Figure Lengend Snippet: Illustration and comparisons of the differentiation patterns of HCN-2 cells transfected with miRNA mimic. a HCN-2 cells were transfected with scrambled control mimic (control), hsa-miR-30e-5p mimic (miR-30e), hsa-miR-140-3p mimic (miR-140), or hsa-miR-126-5p mimic (miR-126) for 24 h. Then, they were seeded on new plates for monitoring and recording growth with a camera. Each transfection treatment had three independent wells, and three pictures were randomly taken from each well, resulting in 3*3 = 9 pictures. ( b – f ) We determined the differentiation patterns of HCN-2 cells in terms of different indices, including ( b ) vessel area, ( c ) vessel length, ( d ) junction number, and ( e ) mean lacunarity. The t -test p values were calculated by comparing the values of the control set on a specific day. f For a systematic comparison, the values were normalized to the average values of the control set at day 9. *, **, *** and **** denote p value

    Article Snippet: However, HCN-2 has scarcely been applied for an in vitro study on ADHD pathophysiology.

    Techniques: Transfection