Journal: Chembiochem

Article Title: Functional Heterogeneity of the Autism Spectrum Disorder‐Associated Gut Microbial Ecosystem Revealed by Fecal Lipopolysaccharides and Bacterial Proteomic Profiling

doi: 10.1002/cbic.70557

Figure Lengend Snippet: Effects of conditioned media (CM) from fecal LPS‐treated HMC3 microglial on SH‐SY5Y viability and differentiation. (A) Bright‐field image of differentiated SH‐SY5Y cells (40× magnification, scale bar: 20 µm). (B) Cell viability of SH‐SY5Y neurons after 24 h exposure to CM from HMC3 microglia previously stimulated with 0.1, 1, 10, 100 ng/mL of the pooled fecal LPS preparations obtained from the ASD and NASD cohorts. Cell viability was expressed as % relative to non‐stimulated (NS) cells and assessed by MTT assay (570 nm). (C) Representative integrated immunofluorescence images combining MAP2 (red) and DAPI (blue) channels, showing MAP2 immunoreactivity in SH‐SY5Y cells exposed to CM from HMC3 cells treated with 100 ng/mL fecal. The integration of MAP2 and DAPI channels provides both neuronal morphology and nuclear localization within the same composite image. Original magnification: 10× (scale bar, 100 µm); zoomed‐in image: 40× (scale bar, 20 µm). (D) Quantitative MAP2 immunofluorescence analysis performed using ImageJ software (version 1.53t), MAP2 cells integrated density was expressed as % relative to NS. (E) Western blot analysis of MAP2 protein levels in SH‐SY5Y cells exposed to CM from HMC3 cells treated with 100 ng/mL of fecal LPS. Data were presented as mean ± SEM and NS cells served as negative control. Three independent experiments ( n = 3) were performed, with each experimental condition tested in triplicate wells within each experiment. Statistical significance was assessed by ordinary one‐way ANOVA followed by Tukey's multiple‐comparison test.

Article Snippet: Human embryonic microglial cells (HMC3, #300102) and human neuroblastoma cells (SH‐SY5Y, #300154) were purchased from Cytion Biosciences (Heidelberg, Germany).

Techniques: MTT Assay, Immunofluorescence, Morphology, Analysis, Software, Western Blot, Negative Control, Comparison

Journal: Genes

Article Title: Transcriptional Activation of the TREM2 Gene by ZEB2 in a Zinc Finger-Dependent Manner

doi: 10.3390/genes16111329

Figure Lengend Snippet: The 5 kb upstream region and 5′-UTR of TREM2 recapitulate its cell type-specific expression. ( A ) Schematic representation of the reporter construct ( T2-5k-u-mCherry ). The CMV promoter in the mCherry-N3 vector was replaced with a fragment containing the 5 kb sequence upstream of the TREM2 transcription start site and its 5′-UTR. ( B ) T2-5k-u-mCherry was transfected into HEK293, THP-1, and HMC3 cells. Representative fluorescence images of mCherry expression are shown. Nuclei were counterstained with Hoechst33342. Scale bars, 50 μm. ( C ) Schematic diagram of the T2-5k-mCherry construct lacking the TREM2 5′-UTR. ( D ) Fluorescent images of THP-1 cells transfected with T2-5k-u-mCherry or T2-5k-mCherry . Scale bars, 50 μm. ( E ) Quantification of mCherry-positive cells relative to Hoechst-positive cells using the IN Cell Analyzer. Error bars represent SDs; n = 3; Welch’s t -test. ( F ) Fluorescence images of HMC3 cells transfected with T2-5k-u-mCherry or T2-5k-mCherry . Scale bars, 50 μm. ( G ) Quantification of mCherry-positive cells relative to Hoechst-positive cells analyzed as in ( E ). Error bars represent SDs; n = 3; Welch’s t -test.

Article Snippet: The HMC3 cell line (EP-CL-0620) was obtained from Elabscience (Houston, TX, USA).

Techniques: Expressing, Construct, Plasmid Preparation, Sequencing, Transfection, Fluorescence

Journal: Genes

Article Title: Transcriptional Activation of the TREM2 Gene by ZEB2 in a Zinc Finger-Dependent Manner

doi: 10.3390/genes16111329

Figure Lengend Snippet: ZEB2 overexpression or knockdown modulates TREM2 mRNA and protein expression. ( A ) RT-qPCR analysis of ZEB2 mRNA in doxycycline-treated inducible EGFP and EGFP-ZEB2 cells. Expression was normalized to ACTB . Error bars represent SDs; n = 3; Welch’s t -test. ( B ) TREM2 mRNA levels normalized to ACTB , quantified from the same samples as in ( A ). Error bars indicate SDs; n = 3; Welch’s t -test. ( C ) ZEB2 siRNAs were transfected into HMC3 cells. ZEB2 mRNA levels were measured by RT-qPCR and normalized to B2M . Error bars indicate SDs; n = 5; Tukey’s test. ( D ) TREM2 mRNA levels quantified as in ( C ). Error bars indicate SDs; n = 5; Tukey’s test. ( E ) Membrane-bound protein fractions from HMC3 cells were analyzed for TREM2 protein levels following ZEB2 knockdown. APP served as a loading control. ( F ) Quantification of ( E ). Error bars indicate SDs; n = 5; Welch’s t -test. ( G ) Relative YY1 mRNA expression in HMC3 cells transfected with YY1 siRNA. YY1 mRNA levels were normalized to B2M. Error bars indicate SDs; n = 3; Welch’s test. ( H ) TREM2 mRNA levels in HMC3 cells transfected with siZEB2 or siYY1. Error bars indicate SDs; n = 5; Tukey’s t-test. ( I ) Relative luciferase activity induced by EGFP-ZEB1. Error bars indicate SDs; n = 3; Welch’s test.

Article Snippet: The HMC3 cell line (EP-CL-0620) was obtained from Elabscience (Houston, TX, USA).

Techniques: Over Expression, Knockdown, Expressing, Quantitative RT-PCR, Transfection, Membrane, Control, Luciferase, Activity Assay

Journal: Cell Proliferation

Article Title: Microglia and glioblastoma heterocellular interplay sustains tumour growth and proliferation as an off‐target effect of radiotherapy

doi: 10.1111/cpr.13606

Figure Lengend Snippet: Irradiated HMC3 conditioned media (CM) preserve radiation‐naïve U‐87 MG and U‐251 MG cell lines viability. (A) Experimental workflow and CM treatment. (B–I) Cytofluorimetric analysis of viability evaluated with Annexin V/propidium iodide (PI) assay on U‐87 MG treated with U‐87 MG CM (B,C), U‐251 MG treated with U‐251 MG CM (D,E), U‐87 MG treated with HMC3 CM (F,G) and U‐251 MG treated with HMC3 CM (H,I). Data are shown as standard box and whiskers and viability is expressed as the percentage of gated cells, n = 4 independent replicates for each experimental condition. * p ‐value <0.05; *** p ‐value <0.001; **** p ‐value <0.0001. GBM, glioblastoma; Mφ, microglia; IR, irradiated.

Article Snippet: HMC3, were purchased from European Collection of Authenticated Cell Cultures (ECACC, Public Health England, Porton Down Salisbury, UK) and cultured with HMC3 growth medium [Minimum Essential Medium (MEM) supplemented with 10% FBS, Penicillin–Streptomycin 100 IU/mL and L‐glutamine 2 mmol/L].

Techniques: Irradiation

Journal: Cell Proliferation

Article Title: Microglia and glioblastoma heterocellular interplay sustains tumour growth and proliferation as an off‐target effect of radiotherapy

doi: 10.1111/cpr.13606

Figure Lengend Snippet: Irradiated HMC3 conditioned media (CM) stimulate naïve U‐87 MG and U‐251 MG clone formation. (A,B) Surviving fraction and representative pictures of U‐87 MG treated with 0 Gy and 15 Gy U‐87 MG CM (A) or 0 Gy and 15 Gy HMC3 CM (B). (C,D) Surviving fraction and representative pictures of U‐251 MG treated with 0 Gy and 15 Gy U‐251 MG CM (C) or 0 Gy and 15 Gy HMC3 CM (D). Data are expressed as scattered dot‐plot and mean ± SEM of n ≥ 3 independent experiments. *** p ‐value <0.001 and **** p ‐value <0.0001.

Article Snippet: HMC3, were purchased from European Collection of Authenticated Cell Cultures (ECACC, Public Health England, Porton Down Salisbury, UK) and cultured with HMC3 growth medium [Minimum Essential Medium (MEM) supplemented with 10% FBS, Penicillin–Streptomycin 100 IU/mL and L‐glutamine 2 mmol/L].

Techniques: Irradiation

Journal: Cell Proliferation

Article Title: Microglia and glioblastoma heterocellular interplay sustains tumour growth and proliferation as an off‐target effect of radiotherapy

doi: 10.1111/cpr.13606

Figure Lengend Snippet: Irradiated HMC3 conditioned media (CM) do not influence mitochondrial mass, fragmentation and integrity in naïve U‐87 MG and U‐251 MG cells. (A,B) Representative pictures of immunofluorescence staining with Mitotracker of U‐87 MG treated with 0 or 15 Gy HMC3 CM (A) and high‐content analysis of Mitotracker mean fluorescence intensity (MFI), mitochondrial fragmentation and percentage of mitochondrial integrity (B). (C,D) Representative pictures of immunofluorescence staining with Mitotracker of U‐251 MG treated with 0 or 15 Gy HMC3 CM (C) and high‐content analysis of Mitotracker MFI, mitochondrial fragmentation and percentage of mitochondrial integrity (D). Data are shown as bar plot and expressed as mean ± SEM of n = 4 independent replicates.

Article Snippet: HMC3, were purchased from European Collection of Authenticated Cell Cultures (ECACC, Public Health England, Porton Down Salisbury, UK) and cultured with HMC3 growth medium [Minimum Essential Medium (MEM) supplemented with 10% FBS, Penicillin–Streptomycin 100 IU/mL and L‐glutamine 2 mmol/L].

Techniques: Irradiation, Immunofluorescence, Staining, High Content Screening, Fluorescence

Journal: Cell Proliferation

Article Title: Microglia and glioblastoma heterocellular interplay sustains tumour growth and proliferation as an off‐target effect of radiotherapy

doi: 10.1111/cpr.13606

Figure Lengend Snippet: Radiation increases mitochondrial reactive oxygen species and irradiated HMC3 conditioned media (CM) preserve mitochondrial oxidative state of U‐87 MG and U‐251 MG cells. (A) qRT‐PCR analysis of mRNA expression levels of DNM1L, FIS1, MNF1, OPA1, MNF2, CYTB, ND4, TFAM, and ATP5F1A in mock‐IR U‐87 MG, 15 Gy U‐87 MG and mock‐IR treated with 15 Gy U‐87 MG CM or 15 Gy HMC3 CM; data are shown as Log 2 FC over mock‐IR; (B) qRT‐PCR analysis of mRNA expression levels of DNM1L, FIS1, MNF1, OPA1, MNF2, CYTB, ND4, TFAM and ATP5F1A in mock‐IR U‐251 MG, 15 Gy U‐251 MG and mock‐IR treated with 15 Gy U‐251 MG CM or 15 Gy HMC3 CM; data are shown as Log 2 FC over mock‐IR; (C–E) Cytofluorimetric analysis of MitoSOX positive cells in 0 or 15 Gy U‐87 MG (C), 0 or 15 Gy U‐251 MG (D) and 0 or 15 Gy HMC3 (E). (F,G) Cytofluorimetric analysis of MitoSOX positive cells in U‐87 MG cultured with 0 or 15 Gy HMC3 CM (F) and in U‐251 MG cultured with 0 or 15 Gy HMC3 CM (G). Data in (C–G) are shown via standard box and whiskers and are expressed as percentage of MitoSOX positive cells of n = 4 replicates for each experimental condition. * p ‐value <0.05; **** p ‐value <0.0001.

Article Snippet: HMC3, were purchased from European Collection of Authenticated Cell Cultures (ECACC, Public Health England, Porton Down Salisbury, UK) and cultured with HMC3 growth medium [Minimum Essential Medium (MEM) supplemented with 10% FBS, Penicillin–Streptomycin 100 IU/mL and L‐glutamine 2 mmol/L].

Techniques: Irradiation, Quantitative RT-PCR, Expressing, Cell Culture

Journal: Cell Proliferation

Article Title: Microglia and glioblastoma heterocellular interplay sustains tumour growth and proliferation as an off‐target effect of radiotherapy

doi: 10.1111/cpr.13606

Figure Lengend Snippet: Metformin administration reverses irradiated HMC3 conditioned media (CM)‐induced effects. (A,B) Surviving fraction and representative pictures of U‐87 MG (A) and U‐251 MG (B) cell lines ± metformin, cultured with 15 Gy GBM CM. (C,D) Surviving fraction and representative pictures of U‐87 MG (C) and U‐251 MG (D) cell lines ± metformin, cultured with standard growth medium. (E,F) Surviving fraction and representative pictures of U‐87 MG (E) and U‐251 MG (F) cell lines ± metformin, cultured with 15 Gy HMC3 CM. Data are expressed as scattered dot‐plot and mean ± SEM of n ≥ 4 independent experiments. * p ‐value <0.05 and ** p ‐value <0.01.

Article Snippet: HMC3, were purchased from European Collection of Authenticated Cell Cultures (ECACC, Public Health England, Porton Down Salisbury, UK) and cultured with HMC3 growth medium [Minimum Essential Medium (MEM) supplemented with 10% FBS, Penicillin–Streptomycin 100 IU/mL and L‐glutamine 2 mmol/L].

Techniques: Irradiation, Cell Culture

Journal: Frontiers in Immunology

Article Title: GPX8 regulates pan-apoptosis in gliomas to promote microglial migration and mediate immunotherapy responses

doi: 10.3389/fimmu.2023.1260169

Figure Lengend Snippet: In vitro validation on GPX8. (A) Quantitative Polymerase Chain Reaction (qPCR) assay revealing the expression of GPX8 in four groups. (B) Cell Counting Kit-8 (CCK8) assay revealing the proliferation ability of U251 cells in three groups. (C) EdU assay revealing the proliferation ability of U251 cells in three groups. (D) Transwell assay revealing the migration ability of U251 cells in three groups. ns, not statistically significant; *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001.

Article Snippet: The human U251 and HMC3 cell lines were acquired from iCell ( http://www.icellbioscience.com/search ).

Techniques: In Vitro, Biomarker Discovery, Real-time Polymerase Chain Reaction, Expressing, Cell Counting, CCK-8 Assay, EdU Assay, Transwell Assay, Migration

Journal: Journal of Neuroinflammation

Article Title: The human microglial HMC3 cell line: where do we stand? A systematic literature review

doi: 10.1186/s12974-018-1288-0

Figure Lengend Snippet: Historical reconstruction of the distribution process of the human microglial clone 3 cell line. The human microglial clone 3 cell line was developed in the laboratory of Prof. M Tardieu, Paris, in 1995 (red circle). As shown in the picture, clone 3 has been distributed worldwide, with the acronym of CHME3 cells (blue boxes) or HMC3 cells (green boxes). Distribution followed two main pathways, either directly from Prof. Tardieu’s laboratory (black thick arrows) or indirectly by the first recipient laboratory (black dotted arrows). A second main distributor of the CHME3 cell line is the laboratory of Prof. A Basu, National Brain Research Centre (NBRC), India (purple circle). Since 2014, this laboratory appears to be the main distributor of the CHME3 cells in India. However, we could not trace on the timeline when the cell line was transferred from the laboratory of Prof. Tardieu to NBRC. In addition, we identified several studies (not reported in the schematic), in which the CHME3 cells were used without any indication of the source, and one study in which the cell line was provided by an Academic institution without any link to published data. In 2016, the HMC3 cells were transferred to ATCC®, USA (orange box) and authenticated and distributed under the catalog designation of HMC3 (ATCC®CRL-3304)

Article Snippet: Of note, the HMC3 cells can grow also in different media, including MEM (Biochrom, Germany) and in DMEM-F12 (Biochrom), with the addition of 10% FCS (Biochrom) and antibiotics.

Techniques:

Journal: Journal of Neuroinflammation

Article Title: The human microglial HMC3 cell line: where do we stand? A systematic literature review

doi: 10.1186/s12974-018-1288-0

Figure Lengend Snippet: Antigenic profile of the human microglial clone 3 cell line

Article Snippet: Of note, the HMC3 cells can grow also in different media, including MEM (Biochrom, Germany) and in DMEM-F12 (Biochrom), with the addition of 10% FCS (Biochrom) and antibiotics.

Techniques: Western Blot

Journal: Journal of Neuroinflammation

Article Title: The human microglial HMC3 cell line: where do we stand? A systematic literature review

doi: 10.1186/s12974-018-1288-0

Figure Lengend Snippet: Production of cytokines, chemokines, and other inflammatory mediators

Article Snippet: Of note, the HMC3 cells can grow also in different media, including MEM (Biochrom, Germany) and in DMEM-F12 (Biochrom), with the addition of 10% FCS (Biochrom) and antibiotics.

Techniques:

Journal: Journal of Neuroinflammation

Article Title: The human microglial HMC3 cell line: where do we stand? A systematic literature review

doi: 10.1186/s12974-018-1288-0

Figure Lengend Snippet: HMC3 cell morphology and labeling of cytoskeletal F-actin filaments. a – b The human microglial cell line HMC3 as it was observed by phase-contrast microscopy at in vitro day 1 ( a ), and when cells reached the confluency ( b ). × 10 magnification, scale bar 100 μM. c – e A representative example of confocal images (1024 × 1024 pixels) acquired at × 20 magnification with a confocal laser scanning system (A1+, Nikon). Cells were grown on glass coverslips for 24 h, and their morphology was evaluated by labeling the cytoskeletal F-actin filaments with tetramethylrhodamine (TRITC)-conjugated phalloidin (red fluorescence). Cells were counterstained with the nuclear probe, 4′,6-damidino-2-phenylindole dihydrochloride (DAPI, blue fluorescence). The merged image is shown in ( e ). Scale bar 50 μM

Article Snippet: Of note, the HMC3 cells can grow also in different media, including MEM (Biochrom, Germany) and in DMEM-F12 (Biochrom), with the addition of 10% FCS (Biochrom) and antibiotics.

Techniques: Labeling, Microscopy, In Vitro, Fluorescence

Journal: Journal of Neuroinflammation

Article Title: The human microglial HMC3 cell line: where do we stand? A systematic literature review

doi: 10.1186/s12974-018-1288-0

Figure Lengend Snippet: HMC3 cell expression of microglial lineage markers. a – f Representative example of confocal images (1024 × 1024 pixels) acquired at × 20 magnification with a confocal laser scanning system (A1+, Nikon). HMC-3 cells immuno-labeled for IBA1 (green fluorescence, d , f ) and DAPI stained (blue fluorescence, a , b ) are shown. Merged images are shown in ( e , f ). Control experiments performed by omitting the primary antibody are shown in ( a , c, e ). No green fluorescence was present ( c , e ), indicating neither spontaneous fluorescence nor non-specificity of the secondary antibody. Scale bar 50 μM. g – h Total RNA was prepared from human microglial HMC3 cells 24 h after incubation in complete growth medium and retrotranscribed using random hexamers. Real time (Q)-PCR analysis for the mRNA levels of IBA1, CX3CR1, CCR2, P2RY12, TMEM119, and CSF1-R was carried out according to our standard protocols . g , h panels show a representative gel image of PCR products obtained at the end of the analysis from two different RNA samples for each condition. In the last line is shown the actin amplification, as positive control

Article Snippet: Of note, the HMC3 cells can grow also in different media, including MEM (Biochrom, Germany) and in DMEM-F12 (Biochrom), with the addition of 10% FCS (Biochrom) and antibiotics.

Techniques: Expressing, Labeling, Fluorescence, Staining, Control, Incubation, Amplification, Positive Control

Journal: Journal of Neuroinflammation

Article Title: The human microglial HMC3 cell line: where do we stand? A systematic literature review

doi: 10.1186/s12974-018-1288-0

Figure Lengend Snippet: Phenotypical characterization of HMC3 cells under basal conditions and in response to IFNγ. HMC3 cells were plated at the density of 30,000 cells/cm 2 in T25 flasks, and grown for 3 days when cells were almost confluent. In the experiments in which INFγ was used, cells were stimulated for the last 36 h. Controls did not receive any stimulus for the same time period (“resting” HMC3 cells). For detection of intracellular antigens, aliquots of 1 × 10 6 cells in 100 μl were fixed using BD cytofix (BD Pharmingen) at 4 °C for 30 min, and permeabilized with BD FACS permeabilizing solution at 4 °C for 30 min. Cells were then stained using the following antibodies: PE-conjugated anti Human GFAP mouse Mab, BD Bioscience ( a ), and PE-CF594-conjugated anti Human CD68 mouse Mab, BD Bioscience ( e ), according to the manufacturer’s instructions. For the evaluation of surface antigens, aliquots of 5 × 10 5 cells in 100 μl were directly incubated in PBS buffer containing the following antibodies: PE-CF594-conjugated anti Human HLA-DR mouse Mab, BD Bioscience ( b ); FITC-conjugated anti Human CD14 mouse Mab, BD Pharmingen ( c ); and PE-conjugated anti Human CD11b mouse Mab, E-Bioscience ( d ). Cells were analyzed by the 6-parameter (2 scatter and 4 fluorescence signals) Coulter Epics XL flow cytometer (Beckman-Coulter). Control histograms (white histograms) indicate level of cell autofluorescence in the emission wavelength that pertains the fluorochrome-conjugated Mab. Panel ( a ) shows expression of GFAP on HMC3 cells at passage 4 (dark gray), and on the human glioblastoma U373 cell line (light gray) that constitutively expresses GFAP. As autofluorescence signal in the two cell lines was similar, only one representative histogram is plotted in panel ( a ) (white histogram). As shown in this panel, the HMC3 cells stain negatively for GFAP (the dark gray histogram completely overlaps the background histogram), whereas the U373 cells express the target antigen. Two different populations expressing GFAP at different levels were identified (light gray plot): 68% of the U373 cells express GFAP at low level (GFAP-dim), whereas 32% of the cells express GFAP at higher level (GFAP-high). Panels ( b – e ) show results from HMC3 cells obtained at passage 7. In these panels, white histograms indicate cell autofluorescence, light gray histograms and dark gray histograms are representative of control (“resting”) HMC3 cells and IFNγ-treated HMC3 cells, respectively. Gating strategy: all analyses were obtained after gating according to morphological characteristics (not shown). As levels of autofluorescence did not change according to cell treatment, one representative example is plotted for each emission wavelength. Linear regions are used for calculating percentage of positive cells (plots A–B). Central tendency of CD68 expression on HMC3 cells in the different experimental conditions is represented by the mean fluorescence intensity (MFI, see text)

Article Snippet: Of note, the HMC3 cells can grow also in different media, including MEM (Biochrom, Germany) and in DMEM-F12 (Biochrom), with the addition of 10% FCS (Biochrom) and antibiotics.

Techniques: Staining, Incubation, Fluorescence, Flow Cytometry, Control, Expressing

Journal: Journal of Neuroinflammation

Article Title: The human microglial HMC3 cell line: where do we stand? A systematic literature review

doi: 10.1186/s12974-018-1288-0

Figure Lengend Snippet: Expression of the human class I MHC antigens on HMC3 cells. a HMC3 were plated at the density 30,000 cells/cm 2 in T25 flasks, and grown for 3 days when cells were almost confluent. For the detection of class I MHC surface antigens, aliquots of 5 × 10 5 cells in 100 μl were directly incubated with FITC-conjugated anti human HLA-ABC mouse Mab, BD Bioscience. This antibody is specific for human MHCI antigens, and does not cross react with rat MHCI antigens. Cells were analyzed by the 6-parameter (2 scatter and 4 fluorescence signals) Coulter Epics XL flow cytometer (Beckman-Coulter). Results from HMC3 cells at passage 8 are shown. Gating strategy: the analysis was obtained after gating according to morphological characteristics (not shown). Background histogram (white) indicates level of cell autofluorescence. A linear region was used for calculating percentage of positive cells. b Gel image of PCR products obtained at the end of a HLA Locus B specific amplification protocol. The 922 bp amplicons were separated by electrophoresis through 1.5% agarose gels containing 0.1 μg/ml ethidium bromide. Lines 2–3 amplification products from genomic DNA extracted by HMC3 (ATCC®CRL-3304) cells; lines 4–5 amplification products from a positive control, i.e., an anonymous human genomic DNA sample provided by ViiV Healthcare Ldt

Article Snippet: Of note, the HMC3 cells can grow also in different media, including MEM (Biochrom, Germany) and in DMEM-F12 (Biochrom), with the addition of 10% FCS (Biochrom) and antibiotics.

Techniques: Expressing, Incubation, Fluorescence, Flow Cytometry, Amplification, Electrophoresis, Positive Control

Journal: Journal of Neuroinflammation

Article Title: The human microglial HMC3 cell line: where do we stand? A systematic literature review

doi: 10.1186/s12974-018-1288-0

Figure Lengend Snippet: Gene expression analysis of the HMC3 (ATCC®CRL-3304) cells

Article Snippet: Of note, the HMC3 cells can grow also in different media, including MEM (Biochrom, Germany) and in DMEM-F12 (Biochrom), with the addition of 10% FCS (Biochrom) and antibiotics.

Techniques: Gene Expression, Control

Journal: Frontiers in Immunology

Article Title: Identification of plasma inflammatory biomarkers for Alzheimer’s disease reveals IFN-γ as a regulator of ACSL1-mediated microglia phenotype

doi: 10.3389/fimmu.2026.1770509

Figure Lengend Snippet: IFN-γ enhances ApoE4-induced ACSL1 expression in HMC3 microglial cells. (A) Quantitative PCR (qPCR) analysis of mRNA expression levels of APOE, APOE2, APOE3, and APOE4 in control HMC3 cells and HMC3 cells overexpressing ApoE2 (HMC3-E2), ApoE3 (HMC3-E3), or ApoE4 (HMC3-E4). (B) qPCR analysis of ACSL1 mRNA expression levels in control HMC3 cells, HMC3-E2, HMC3-E3, and HMC3-E4 cells. (C) Western blot analysis of ACSL1 protein expression in the indicated groups. (D) Quantification of ACSL1 protein levels shown in (C) . (E, F) qPCR analysis comparing the transcriptional expression levels of IRF1 and CXCL10 in control HMC3 cells and HMC3-E4 cells in the presence or absence of IFN-γ stimulation. (G) qPCR analysis of ACSL1 mRNA expression in control HMC3 cells and HMC3-E4 cells with or without IFN-γ stimulation. (H) Western blot analysis of ACSL1 protein expression under the indicated conditions. (I) Quantification of ACSL1 protein levels shown in (H) . *P < 0.05; **P < 0.01; ***P < 0.001.

Article Snippet: HMC3 were purchased from the Procell Life Science & Technology Co. (Procell, Wuhan, China).

Techniques: Expressing, Real-time Polymerase Chain Reaction, Control, Western Blot