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    Name:
    Bovine Serum Albumin
    Description:

    Catalog Number:
    a9306
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    Structured Review

    Millipore ffa bsa
    Bovine Serum Albumin

    https://www.bioz.com/result/ffa bsa/product/Millipore
    Average 99 stars, based on 2 article reviews
    Price from $9.99 to $1999.99
    ffa bsa - by Bioz Stars, 2020-10
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    Images

    1) Product Images from "Antilipotoxicity Activity of Osmanthus fragrans and Chrysanthemum morifolium Flower Extracts in Hepatocytes and Renal Glomerular Mesangial Cells"

    Article Title: Antilipotoxicity Activity of Osmanthus fragrans and Chrysanthemum morifolium Flower Extracts in Hepatocytes and Renal Glomerular Mesangial Cells

    Journal: Mediators of Inflammation

    doi: 10.1155/2017/4856095

    Effects of OF and CM on the mRNA expressions of lipogenesis-related genes. Real-time RT-PCR analysis of sterol regulatory element-binding protein-1 ( SREBP-1c ) (a) and glycerol-3-phosphate acyltransferase ( GPAT ) (b) mRNA levels in 1 mM FFA/BSA-treated HepG2 cells. All data were normalized to GAPDH mRNA, and the fold changes in expression were calculated relative to control cells (treated with 1% BSA). Each experiment was independently performed three times. Data were presented as the mean ± SD. Values not sharing common superscripts are significantly different ( p
    Figure Legend Snippet: Effects of OF and CM on the mRNA expressions of lipogenesis-related genes. Real-time RT-PCR analysis of sterol regulatory element-binding protein-1 ( SREBP-1c ) (a) and glycerol-3-phosphate acyltransferase ( GPAT ) (b) mRNA levels in 1 mM FFA/BSA-treated HepG2 cells. All data were normalized to GAPDH mRNA, and the fold changes in expression were calculated relative to control cells (treated with 1% BSA). Each experiment was independently performed three times. Data were presented as the mean ± SD. Values not sharing common superscripts are significantly different ( p

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

    Effect of OF and CM on FFA-induced ROS production. HepG2 cells were incubated with 1 mM FFAs/BSA for 24 h in the presence of OF or CM. Vehicle cells were incubated with 0.1% DMSO in the presence of FFAs/BSA. Control cells were incubated with 1% BSA. Intracellular ROS production was quantified using the fluorescent probe DCFDA. Data were presented as mean ± SD of three independent experiments. Values not sharing common superscripts are significantly different ( p
    Figure Legend Snippet: Effect of OF and CM on FFA-induced ROS production. HepG2 cells were incubated with 1 mM FFAs/BSA for 24 h in the presence of OF or CM. Vehicle cells were incubated with 0.1% DMSO in the presence of FFAs/BSA. Control cells were incubated with 1% BSA. Intracellular ROS production was quantified using the fluorescent probe DCFDA. Data were presented as mean ± SD of three independent experiments. Values not sharing common superscripts are significantly different ( p

    Techniques Used: Incubation

    Effects of OF and CM on lipid accumulation in free fatty acid-overloaded HepG2 cells. HepG2 cells were incubated with 1 mM FFAs/BSA and cotreated with various concentrations of OF and CM for 24 h. Vehicle cells were incubated with 0.1% DMSO in the presence of FFAs/BSA. Control cells were incubated with 1% BSA. Cell viability was measured by the MTT assay (a). Quantitative analysis of lipid deposition in cells by the OD 500 nm values using Oil Red O staining (b). Intracellular triglyceride (c) and cholesterol (d) contents were determined in cell lysates by an enzymatic colorimetric method using a commercially available kit. Total cholesterol and TG levels of the control cells were 25.3 ± 7.1 and 28.7 ± 2.7 μ g/mg of cellular protein, respectively. Data were presented as mean ± SD of three independent experiments. Values not sharing common superscripts are significantly different ( p
    Figure Legend Snippet: Effects of OF and CM on lipid accumulation in free fatty acid-overloaded HepG2 cells. HepG2 cells were incubated with 1 mM FFAs/BSA and cotreated with various concentrations of OF and CM for 24 h. Vehicle cells were incubated with 0.1% DMSO in the presence of FFAs/BSA. Control cells were incubated with 1% BSA. Cell viability was measured by the MTT assay (a). Quantitative analysis of lipid deposition in cells by the OD 500 nm values using Oil Red O staining (b). Intracellular triglyceride (c) and cholesterol (d) contents were determined in cell lysates by an enzymatic colorimetric method using a commercially available kit. Total cholesterol and TG levels of the control cells were 25.3 ± 7.1 and 28.7 ± 2.7 μ g/mg of cellular protein, respectively. Data were presented as mean ± SD of three independent experiments. Values not sharing common superscripts are significantly different ( p

    Techniques Used: Incubation, MTT Assay, Staining

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    Transfection:

    Article Title: A Novel Cellular Defect in Diabetes
    Article Snippet: .. Cells were treated for 20 h with 5 mmol/L of AGE-modified BSA (AGE-BSA) (Sigma) 4 h after transfection. ..

    Mouse Assay:

    Article Title: Diagnosis of hepatocellular carcinoma using a novel anti-glycocholic acid monoclonal antibody-based method
    Article Snippet: .. As previously described , mice were subcutaneously immunized with 100 µg GCA-BSA emulsified with Freund's complete adjuvant (Sigma-Aldrich; Merck KGaA) with a volume ratio of 1:1. ..

    Enzyme-linked Immunosorbent Assay:

    Article Title: An Essential Role for RGS Protein/Gαi2 Interactions in B Lymphocyte Directed Cell Migration and Trafficking
    Article Snippet: .. Briefly, 96 well ELISA plates (Nunc) were coated with TNP-BSA (Biosearch Technology) overnight at 4°C, washed and blocked with 1% BSA fraction V (Sigma-Aldrich), serum titers were then added to the plates and incubated 4h at 4°C. .. After washing alkaline phosphatase-labeled goat anti-mouse Ig isotype specific antibodies were added for 2h at RT (SouthernBiotech).

    Incubation:

    Article Title: Impaired Fc?RI stability, signaling, and effector functions in murine mast cells lacking glycosylphosphatidylinositol-anchored proteins
    Article Snippet: .. After stimulation with IgE anti-DNP (IGEL-A2) and DNP-BSA, BMMC were lysed by incubation for 30 minutes on ice in lysis buffer (20mM Tris-HCl, at pH 7.4 containing 0.5% Triton X-100, 150mM NaCl, 2mM EDTA, 50mM NaF, 1mM Na3 VO4 , 1mM PMSF, 10 μg aprotinin/mL, and 10 μg leupeptin/mL). ..

    Article Title: An Essential Role for RGS Protein/Gαi2 Interactions in B Lymphocyte Directed Cell Migration and Trafficking
    Article Snippet: .. Briefly, 96 well ELISA plates (Nunc) were coated with TNP-BSA (Biosearch Technology) overnight at 4°C, washed and blocked with 1% BSA fraction V (Sigma-Aldrich), serum titers were then added to the plates and incubated 4h at 4°C. .. After washing alkaline phosphatase-labeled goat anti-mouse Ig isotype specific antibodies were added for 2h at RT (SouthernBiotech).

    other:

    Article Title: Low-Temperature Storage Improves the Over-Time Stability of Implantable Glucose and Lactate Biosensors
    Article Snippet: Ascorbic acid, D-(+)-glucose, L-lactate, glucose oxidase from Aspergillus Niger (EC 1.1.3.4), lactate oxidase from Pediococcus species (EC 1.1.3.2), albumin from bovine serum (BSA) , o -phenylenediamine (OPD), polyethylenimine (PEI), glutaraldehyde (GTA) , polyurethane (PU) and tetrahydrofuran (THF) were purchased from Sigma-Aldrich (Milano, Italy).

    Lysis:

    Article Title: Impaired Fc?RI stability, signaling, and effector functions in murine mast cells lacking glycosylphosphatidylinositol-anchored proteins
    Article Snippet: .. After stimulation with IgE anti-DNP (IGEL-A2) and DNP-BSA, BMMC were lysed by incubation for 30 minutes on ice in lysis buffer (20mM Tris-HCl, at pH 7.4 containing 0.5% Triton X-100, 150mM NaCl, 2mM EDTA, 50mM NaF, 1mM Na3 VO4 , 1mM PMSF, 10 μg aprotinin/mL, and 10 μg leupeptin/mL). ..

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    Millipore fatty acid free bsa
    PA-induced lipotoxicity and DHA neuroprotective action on primary rat cortical cells <t>(RCC).</t> A : Cellular morphology of control and treated <t>(PA/BSA,</t> 2:1, 48 hr) RCC. B : Nuclear morphology of control and treated RCC. Arrows indicate fragmented nuclei. C : Viability of RCC treated for 48 hr with PA/BSA (2:1 ratio; PA) or in cotreatment with DHA (PA + DHA). Viability was determined using the WST-1 assay. Data represent mean ± SEM of three independent experiments. ★ P
    Fatty Acid Free Bsa, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 507 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/fatty acid free bsa/product/Millipore
    Average 99 stars, based on 507 article reviews
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    99
    Millipore ffa bsa
    Effects of OF and CM on the mRNA expressions of lipogenesis-related genes. Real-time RT-PCR analysis of sterol regulatory element-binding protein-1 ( SREBP-1c ) (a) and glycerol-3-phosphate acyltransferase ( GPAT ) (b) mRNA levels in 1 mM <t>FFA/BSA-treated</t> HepG2 cells. All data were normalized to GAPDH mRNA, and the fold changes in expression were calculated relative to control cells (treated with 1% BSA). Each experiment was independently performed three times. Data were presented as the mean ± SD. Values not sharing common superscripts are significantly different ( p
    Ffa Bsa, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/ffa bsa/product/Millipore
    Average 99 stars, based on 2 article reviews
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    99
    Millipore cardiomyocyte fraction
    Targeted <t>cardiomyocyte</t> deletion of growth differentiation factor 11 ( Gdf11 ) does not decrease total Gdf11 mRNA expression in mouse hearts. A : representative agarose gel image depicting Myh6 (wild-type band at 894 bp, Myh6-cre band at 300 bp) and Gdf11 [wild-type band at 359 bp, flox band at 393 bp, and ∆2–3 (postrecombination) band at 300 bp] alleles seen in the heart on day of life 0 – 1 by polymerase chain reaction (PCR) in Myh6 cre/wt , Gdf11 fl/fl , and Myh6 cre/wt ; Gdf11 fl/fl pups. B : recombination of Gdf11 at 6 mo of age in females and males in the heart but not in lung, liver, kidney, spleen, or skeletal muscle by PCR. C and D : Gdf11 expression in RNA extracted from whole heart in 3 ( n = 4/group, C )- or 6 ( n = 3–4/group, D )-mo-old female and male mice by quantitative PCR (qPCR). Data are normalized to TATA-binding protein ( Tbp ) and then to female Myh6 cre/wt control. E and F : Gdf11 expression in RNA extracted from isolated adult ( > 2 mo old) cardiomyocytes ( E ) or noncardiomyocytes ( F ), with each data point (shown with open symbols) representing isolated cells from a single mouse of the same genotype, n = 3/group (2 males, 1 female). Data were normalized to Tbp and then Myh6 cre/wt control. Cardiomyocytes ( E ), P value = not significant (ns, 0.05); noncardiomyocytes ( F ), P value = ns (0.3) by Kruskal-Wallis test. G and H : serum levels of GDF11 ( G ) and myostatin ( H ) as determined by quantitative mass spectrometry in 6-mo-old mice; n = 3–4/group. * P
    Cardiomyocyte Fraction, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 0 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/cardiomyocyte fraction/product/Millipore
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    PA-induced lipotoxicity and DHA neuroprotective action on primary rat cortical cells (RCC). A : Cellular morphology of control and treated (PA/BSA, 2:1, 48 hr) RCC. B : Nuclear morphology of control and treated RCC. Arrows indicate fragmented nuclei. C : Viability of RCC treated for 48 hr with PA/BSA (2:1 ratio; PA) or in cotreatment with DHA (PA + DHA). Viability was determined using the WST-1 assay. Data represent mean ± SEM of three independent experiments. ★ P

    Journal: Journal of neuroscience research

    Article Title: Activation and Reversal of Lipotoxicity in PC12 and Rat Cortical Cells Following Exposure to Palmitic Acid

    doi: 10.1002/jnr.21918

    Figure Lengend Snippet: PA-induced lipotoxicity and DHA neuroprotective action on primary rat cortical cells (RCC). A : Cellular morphology of control and treated (PA/BSA, 2:1, 48 hr) RCC. B : Nuclear morphology of control and treated RCC. Arrows indicate fragmented nuclei. C : Viability of RCC treated for 48 hr with PA/BSA (2:1 ratio; PA) or in cotreatment with DHA (PA + DHA). Viability was determined using the WST-1 assay. Data represent mean ± SEM of three independent experiments. ★ P

    Article Snippet: PA and/or DHA (Sigma, St. Louis, MO) was first dissolved/diluted in 100% ethanol and further diluted in warm low-serum medium for NGFDPC12 cells, or serum-free medium for RCC, with fatty acid-free BSA (EMD Biosciences, La Jolla, CA).

    Techniques: WST-1 Assay

    Palmitic acid-induced lipotoxicity in NGFDPC12 cells. A : NGFDPC12 cells were exposed to PA/BSA (2:1 ratio), and viability was determined using WST-1 assay at the indicated times. B : NGFDPC12 cells were exposed to different PA/BSA ratios (0.25:1, 0.5:1, 1:1, and 2:1 with BSA at a concentration of 0.150 mM) for 24 hr. Cell viability was measured by the WST-1 assay. C : NGFDPC12 cells were exposed to 0.150 mM BSA without PA (control) or PA/BSA (2:1; PA) for 12 hr. Cells were stained with Hoechst (10 ng/liter), and nuclei were visualized under fluorescent microscopy. Arrows indicate nuclei showing chromatin condensation and fragmentation. D : Regulation of apoptosis-asociated genes in NGFDPC12 cultures exposed to PA/BSA (2:1). Quantitative RT-PCR experiments show the time-dependent mRNA expression of Bim, Bad, AIF, BNIP-3, 14.3.3, and FAS-R. GAPDH mRNA expression was used as the internal control. Data represent mean 6 SEM of three independent experiments. ★ P

    Journal: Journal of neuroscience research

    Article Title: Activation and Reversal of Lipotoxicity in PC12 and Rat Cortical Cells Following Exposure to Palmitic Acid

    doi: 10.1002/jnr.21918

    Figure Lengend Snippet: Palmitic acid-induced lipotoxicity in NGFDPC12 cells. A : NGFDPC12 cells were exposed to PA/BSA (2:1 ratio), and viability was determined using WST-1 assay at the indicated times. B : NGFDPC12 cells were exposed to different PA/BSA ratios (0.25:1, 0.5:1, 1:1, and 2:1 with BSA at a concentration of 0.150 mM) for 24 hr. Cell viability was measured by the WST-1 assay. C : NGFDPC12 cells were exposed to 0.150 mM BSA without PA (control) or PA/BSA (2:1; PA) for 12 hr. Cells were stained with Hoechst (10 ng/liter), and nuclei were visualized under fluorescent microscopy. Arrows indicate nuclei showing chromatin condensation and fragmentation. D : Regulation of apoptosis-asociated genes in NGFDPC12 cultures exposed to PA/BSA (2:1). Quantitative RT-PCR experiments show the time-dependent mRNA expression of Bim, Bad, AIF, BNIP-3, 14.3.3, and FAS-R. GAPDH mRNA expression was used as the internal control. Data represent mean 6 SEM of three independent experiments. ★ P

    Article Snippet: PA and/or DHA (Sigma, St. Louis, MO) was first dissolved/diluted in 100% ethanol and further diluted in warm low-serum medium for NGFDPC12 cells, or serum-free medium for RCC, with fatty acid-free BSA (EMD Biosciences, La Jolla, CA).

    Techniques: WST-1 Assay, Concentration Assay, Staining, Microscopy, Quantitative RT-PCR, Expressing

    DNA fragmentation and caspase-3 activity of NGFDPC12 cells exposed to PA. A : Representative experimental results of BRDU-FITC plots (apoptotic cells) vs. PI (total DNA marker) for controls and cells exposed to PA/BSA (2:1) for 12 hr. The R2 quadrant shows cells exhibiting increased BrdU-FITC fluorescence (green). Nonapoptotic cells are indicated in red. B : Representative distribution diagram of positive apoptotic NGFDPC12 cells. Cells were treated with BSA for negative control (green), PA/BSA (2:1) for 12 hr (red) or 24 hr (blue), or staurosporine for positive control (orange). C : Quantitative analysis of positive apoptotic NGFDPC12 cells as shown in B. The data represent the mean ± SEM of three independent experiments done in triplicate. D : Caspase-3 activity in cell extracts was determined in control or NGFDPC12 cells treated with PA/BSA for 12 hr. E : Early posttreatment with BSA protects NGFDPC12 cells from PA-induced lipotoxicity. Additional BSA (0.6 mM final concentration) was added to NGFDPC-12 cells at 2 or 6 hr after initial PA/BSA (2:1 ratio) treatment. Percentage of viable cells was measured by the WST-1 assay at the end of a 24-hr incubation period. Nuclei in bottom panel were visualized with Hoechst staining. Data represent mean ± SEM of three independent experiments. ★ P

    Journal: Journal of neuroscience research

    Article Title: Activation and Reversal of Lipotoxicity in PC12 and Rat Cortical Cells Following Exposure to Palmitic Acid

    doi: 10.1002/jnr.21918

    Figure Lengend Snippet: DNA fragmentation and caspase-3 activity of NGFDPC12 cells exposed to PA. A : Representative experimental results of BRDU-FITC plots (apoptotic cells) vs. PI (total DNA marker) for controls and cells exposed to PA/BSA (2:1) for 12 hr. The R2 quadrant shows cells exhibiting increased BrdU-FITC fluorescence (green). Nonapoptotic cells are indicated in red. B : Representative distribution diagram of positive apoptotic NGFDPC12 cells. Cells were treated with BSA for negative control (green), PA/BSA (2:1) for 12 hr (red) or 24 hr (blue), or staurosporine for positive control (orange). C : Quantitative analysis of positive apoptotic NGFDPC12 cells as shown in B. The data represent the mean ± SEM of three independent experiments done in triplicate. D : Caspase-3 activity in cell extracts was determined in control or NGFDPC12 cells treated with PA/BSA for 12 hr. E : Early posttreatment with BSA protects NGFDPC12 cells from PA-induced lipotoxicity. Additional BSA (0.6 mM final concentration) was added to NGFDPC-12 cells at 2 or 6 hr after initial PA/BSA (2:1 ratio) treatment. Percentage of viable cells was measured by the WST-1 assay at the end of a 24-hr incubation period. Nuclei in bottom panel were visualized with Hoechst staining. Data represent mean ± SEM of three independent experiments. ★ P

    Article Snippet: PA and/or DHA (Sigma, St. Louis, MO) was first dissolved/diluted in 100% ethanol and further diluted in warm low-serum medium for NGFDPC12 cells, or serum-free medium for RCC, with fatty acid-free BSA (EMD Biosciences, La Jolla, CA).

    Techniques: Activity Assay, Marker, Fluorescence, Negative Control, Positive Control, Concentration Assay, WST-1 Assay, Incubation, Staining

    ROS generation and changes in mitochondrial membrane permeability in NGFDPC12 cells undergoing PA-induced lipotoxicity. A : ROS generation in NGFDPC12 cells was detected by the 2′,7′ -dichlorofluorescein diacetate (DCF) method at 12 hr after the initial exposure to PA using flow cytometry. NGFDPC12 cells were treated with no DCF (light green), DCF (dark green), PA + DCF (blue), or hydrogen peroxide + DCF (red). B : Quantification of ROS production in NGFDPC12 cells at 3, 6, 9, and 12 hr after PA/BSA treatment. C : ROS modulation after addition of DHA, MCI-186, or BSA to NCGDPC12 cells treated with PA for 6 hr. D: Mitochondrial depolarization in NGFDPC12 cells, control or PA/BSA treated, detected by JC-1 flow cytometric analysis. JC-1 was detected in the FL1 or FL2 channel depending on its aggregation status (see Mqaterials and Methods). The R2 quadrant defines cells with leaky mitochondria showing reduced FL2 readings (green). Cells with intact mitochondria are indicated in R1 (red). Data represent mean ± SEM sent of at least three independent experiments. ★ P

    Journal: Journal of neuroscience research

    Article Title: Activation and Reversal of Lipotoxicity in PC12 and Rat Cortical Cells Following Exposure to Palmitic Acid

    doi: 10.1002/jnr.21918

    Figure Lengend Snippet: ROS generation and changes in mitochondrial membrane permeability in NGFDPC12 cells undergoing PA-induced lipotoxicity. A : ROS generation in NGFDPC12 cells was detected by the 2′,7′ -dichlorofluorescein diacetate (DCF) method at 12 hr after the initial exposure to PA using flow cytometry. NGFDPC12 cells were treated with no DCF (light green), DCF (dark green), PA + DCF (blue), or hydrogen peroxide + DCF (red). B : Quantification of ROS production in NGFDPC12 cells at 3, 6, 9, and 12 hr after PA/BSA treatment. C : ROS modulation after addition of DHA, MCI-186, or BSA to NCGDPC12 cells treated with PA for 6 hr. D: Mitochondrial depolarization in NGFDPC12 cells, control or PA/BSA treated, detected by JC-1 flow cytometric analysis. JC-1 was detected in the FL1 or FL2 channel depending on its aggregation status (see Mqaterials and Methods). The R2 quadrant defines cells with leaky mitochondria showing reduced FL2 readings (green). Cells with intact mitochondria are indicated in R1 (red). Data represent mean ± SEM sent of at least three independent experiments. ★ P

    Article Snippet: PA and/or DHA (Sigma, St. Louis, MO) was first dissolved/diluted in 100% ethanol and further diluted in warm low-serum medium for NGFDPC12 cells, or serum-free medium for RCC, with fatty acid-free BSA (EMD Biosciences, La Jolla, CA).

    Techniques: Permeability, Flow Cytometry, Cytometry

    Inhibition of fatty acid-induced lipotoxicity by DHA. A : DHA inhibits induction of cell death by PA in NGFDPC12 cells. Cell viability was determined after treatment for 24 hr as follows: 0.150 mM BSA alone (control), PA/BSA 2:1 ratio (PA), PA/BSA 2:1 cotreated with DHA followed by an increase in BSA final concentration to 0.6 mM at 6 hr (PA + DHA + BSA at 6 hr), and DHA pretreatment for 12 hr followed by treatment with PA/BSA 2:1 (DHA 12 hr pre + PA). B : DHA treatment inhibits apoptotic features of NGFDPC12 cells treated with PA/BSA. Morphology of NGFDPC12 cells treated with BSA alone (control), PA/BSA 2:1 cotreated with DHA, or PA/BSA 2:1 for 24 hr. C : Nuclear morphology of NGFDPC12 cells treated with BSA alone (control), PA/BSA 2:1 cotreated with DHA or PA/BSA 2:1 for 24 hr. Arrows indicate nuclei exhibiting fragmentation. D : DHA cotreatment produced a significant inhibitory effect of caspase-3 activity in cellular extracts of NGFDPC12 cells exposed to PA/BSA (2:1). E : Cleavage of PARP and lamin B in NGFDPC-12 cells exposed to PA/BSA (2:1), as assessed by Western blotting. PA treatment induced the appearance of the signature apoptotic fragments of PARP (85 kDa) or lamin B (46 kDa). DHA inhibited PARP and lamin B cleavage. Data in A and D represent mean ± SEM of three independent experiments. ★ P

    Journal: Journal of neuroscience research

    Article Title: Activation and Reversal of Lipotoxicity in PC12 and Rat Cortical Cells Following Exposure to Palmitic Acid

    doi: 10.1002/jnr.21918

    Figure Lengend Snippet: Inhibition of fatty acid-induced lipotoxicity by DHA. A : DHA inhibits induction of cell death by PA in NGFDPC12 cells. Cell viability was determined after treatment for 24 hr as follows: 0.150 mM BSA alone (control), PA/BSA 2:1 ratio (PA), PA/BSA 2:1 cotreated with DHA followed by an increase in BSA final concentration to 0.6 mM at 6 hr (PA + DHA + BSA at 6 hr), and DHA pretreatment for 12 hr followed by treatment with PA/BSA 2:1 (DHA 12 hr pre + PA). B : DHA treatment inhibits apoptotic features of NGFDPC12 cells treated with PA/BSA. Morphology of NGFDPC12 cells treated with BSA alone (control), PA/BSA 2:1 cotreated with DHA, or PA/BSA 2:1 for 24 hr. C : Nuclear morphology of NGFDPC12 cells treated with BSA alone (control), PA/BSA 2:1 cotreated with DHA or PA/BSA 2:1 for 24 hr. Arrows indicate nuclei exhibiting fragmentation. D : DHA cotreatment produced a significant inhibitory effect of caspase-3 activity in cellular extracts of NGFDPC12 cells exposed to PA/BSA (2:1). E : Cleavage of PARP and lamin B in NGFDPC-12 cells exposed to PA/BSA (2:1), as assessed by Western blotting. PA treatment induced the appearance of the signature apoptotic fragments of PARP (85 kDa) or lamin B (46 kDa). DHA inhibited PARP and lamin B cleavage. Data in A and D represent mean ± SEM of three independent experiments. ★ P

    Article Snippet: PA and/or DHA (Sigma, St. Louis, MO) was first dissolved/diluted in 100% ethanol and further diluted in warm low-serum medium for NGFDPC12 cells, or serum-free medium for RCC, with fatty acid-free BSA (EMD Biosciences, La Jolla, CA).

    Techniques: Inhibition, Concentration Assay, Produced, Activity Assay, Western Blot

    DHA reduces mitochondrial depolarization of NGFDPC12 cells undergoing PA-induced lipotoxicity. A : NGFDPC12 cells were treated with PA/BSA alone (PA) or in the presence DHA (PA + DHA) for 9 or 12 hr, and mitochondrial depolarization was determined by JC-1 flow cytometry assays. Representative flow cytometric plots are shown. B : Quantification of mitochondria depolarization (R2 in JC-1 flow cytometry) in NGFDPC12 cells undergoing PA-induced lipotoxicity. Data represent mean ± SEM of three independent experiments. ★ P

    Journal: Journal of neuroscience research

    Article Title: Activation and Reversal of Lipotoxicity in PC12 and Rat Cortical Cells Following Exposure to Palmitic Acid

    doi: 10.1002/jnr.21918

    Figure Lengend Snippet: DHA reduces mitochondrial depolarization of NGFDPC12 cells undergoing PA-induced lipotoxicity. A : NGFDPC12 cells were treated with PA/BSA alone (PA) or in the presence DHA (PA + DHA) for 9 or 12 hr, and mitochondrial depolarization was determined by JC-1 flow cytometry assays. Representative flow cytometric plots are shown. B : Quantification of mitochondria depolarization (R2 in JC-1 flow cytometry) in NGFDPC12 cells undergoing PA-induced lipotoxicity. Data represent mean ± SEM of three independent experiments. ★ P

    Article Snippet: PA and/or DHA (Sigma, St. Louis, MO) was first dissolved/diluted in 100% ethanol and further diluted in warm low-serum medium for NGFDPC12 cells, or serum-free medium for RCC, with fatty acid-free BSA (EMD Biosciences, La Jolla, CA).

    Techniques: Flow Cytometry, Cytometry

    Analysis of de novo sphingolipid biosynthesis in RAW264.7 cells by [ 13 C]palmitate labeling. RAW264.7 cells were incubated with 0.1 m m [U- 13 C]palmitic acid (as the BSA complex) with vehicle control (PBS), KLA (100 ng/ml), ISP1 (1 μ m ), or KLA+ ISP1. For cells treated with KLA + ISP1, ISP1 was added 1 h prior to the addition of KLA for the times shown. The appearance of 13 C in newly synthesized sphingolipids was quantified by mass spectrometry as described under supplemental “Materials and Methods” . A , summation of base (backbone)-labeled and dual (backbone and fatty acid)-labeled Cer subspecies. Data represent the means ± S.E. ( n = 3). B , summation of base (backbone)-labeled and dual (backbone and fatty acid)-labeled Cer species in cells treated with control ( left ) or KLA ( right ), Data represent the means ( n = 3). Because of the difficulties in visualization, the quantities of ceramide subspecies in control treated cells at the 24-h time point are as follows: C16 (7.6 pmol/μg DNA); C18 (0.7 pmol/μg DNA); C20 (0.3 pmol/μg DNA); C22 (3.2 pmol/μg DNA); C24:1 (6 pmol/μg DNA); C24 (6.4 pmol/μg DNA); C26:1 (0.2 pmol/μg DNA); C26 (0.1 pmol/μg DNA). C , summation of base (backbone)-labeled and dual (backbone and fatty acid)-labeled Cer species ( left ) and 12 C (unlabeled) Cer species ( right ) in cells treated with KLA + ISP1. Data represent the means ( n = 3).

    Journal: The Journal of Biological Chemistry

    Article Title: Kdo2-Lipid A, a TLR4-specific Agonist, Induces de Novo Sphingolipid Biosynthesis in RAW264.7 Macrophages, Which Is Essential for Induction of Autophagy *

    doi: 10.1074/jbc.M110.170621

    Figure Lengend Snippet: Analysis of de novo sphingolipid biosynthesis in RAW264.7 cells by [ 13 C]palmitate labeling. RAW264.7 cells were incubated with 0.1 m m [U- 13 C]palmitic acid (as the BSA complex) with vehicle control (PBS), KLA (100 ng/ml), ISP1 (1 μ m ), or KLA+ ISP1. For cells treated with KLA + ISP1, ISP1 was added 1 h prior to the addition of KLA for the times shown. The appearance of 13 C in newly synthesized sphingolipids was quantified by mass spectrometry as described under supplemental “Materials and Methods” . A , summation of base (backbone)-labeled and dual (backbone and fatty acid)-labeled Cer subspecies. Data represent the means ± S.E. ( n = 3). B , summation of base (backbone)-labeled and dual (backbone and fatty acid)-labeled Cer species in cells treated with control ( left ) or KLA ( right ), Data represent the means ( n = 3). Because of the difficulties in visualization, the quantities of ceramide subspecies in control treated cells at the 24-h time point are as follows: C16 (7.6 pmol/μg DNA); C18 (0.7 pmol/μg DNA); C20 (0.3 pmol/μg DNA); C22 (3.2 pmol/μg DNA); C24:1 (6 pmol/μg DNA); C24 (6.4 pmol/μg DNA); C26:1 (0.2 pmol/μg DNA); C26 (0.1 pmol/μg DNA). C , summation of base (backbone)-labeled and dual (backbone and fatty acid)-labeled Cer species ( left ) and 12 C (unlabeled) Cer species ( right ) in cells treated with KLA + ISP1. Data represent the means ( n = 3).

    Article Snippet: Materials The suppliers for the reagents were as follows: Kdo2 -lipid A and the internal standard mixture for SL analysis by LC ESI-MS/MS (ceramide/sphingoid internal standard mixture II, LM-6005) (Avanti Polar Lipids, Alabaster, AL); [U-13 C]palmitic acid (98%) (Cambridge Isotopes, Andover, MA); fatty acid-free BSA (Calbiochem) ; Hoechst 33342 (Invitrogen); ISP1 (Biomol, Plymouth Meeting, PA); mouse monoclonal antibody to SPT1 (anti-LCB1), anti-BiP/GRP78, and anti-GM130 (BD Biosciences); rabbit monoclonal anti-HA tag and rabbit polyclonal LC3B antibody (Cell Signaling Technology, Boston); mouse monoclonal anti-Cer antibody clone 15B4 (Alexis Biochemicals, San Diego) This antibody clone has been successfully used in immunocytochemistry applications with no indication of nonspecific reactions with other lipids ( ) The secondary Alexa Fluor-conjugated F(ab)2 fragment goat anti-mouse antibody and goat anti-rabbit antibody were from Molecular Probes, Inc. (Eugene, OR.).

    Techniques: Labeling, Incubation, Synthesized, Mass Spectrometry

    Effects of OF and CM on the mRNA expressions of lipogenesis-related genes. Real-time RT-PCR analysis of sterol regulatory element-binding protein-1 ( SREBP-1c ) (a) and glycerol-3-phosphate acyltransferase ( GPAT ) (b) mRNA levels in 1 mM FFA/BSA-treated HepG2 cells. All data were normalized to GAPDH mRNA, and the fold changes in expression were calculated relative to control cells (treated with 1% BSA). Each experiment was independently performed three times. Data were presented as the mean ± SD. Values not sharing common superscripts are significantly different ( p

    Journal: Mediators of Inflammation

    Article Title: Antilipotoxicity Activity of Osmanthus fragrans and Chrysanthemum morifolium Flower Extracts in Hepatocytes and Renal Glomerular Mesangial Cells

    doi: 10.1155/2017/4856095

    Figure Lengend Snippet: Effects of OF and CM on the mRNA expressions of lipogenesis-related genes. Real-time RT-PCR analysis of sterol regulatory element-binding protein-1 ( SREBP-1c ) (a) and glycerol-3-phosphate acyltransferase ( GPAT ) (b) mRNA levels in 1 mM FFA/BSA-treated HepG2 cells. All data were normalized to GAPDH mRNA, and the fold changes in expression were calculated relative to control cells (treated with 1% BSA). Each experiment was independently performed three times. Data were presented as the mean ± SD. Values not sharing common superscripts are significantly different ( p

    Article Snippet: The FFA/BSA or oleic acid (OA)/BSA complex solution was sterile-filtered through 0.22 μ m sterile filters (Millipore S.A.S., Molsheim, France) and then stored at −20°C until use.

    Techniques: Quantitative RT-PCR, Binding Assay, Expressing

    Effect of OF and CM on FFA-induced ROS production. HepG2 cells were incubated with 1 mM FFAs/BSA for 24 h in the presence of OF or CM. Vehicle cells were incubated with 0.1% DMSO in the presence of FFAs/BSA. Control cells were incubated with 1% BSA. Intracellular ROS production was quantified using the fluorescent probe DCFDA. Data were presented as mean ± SD of three independent experiments. Values not sharing common superscripts are significantly different ( p

    Journal: Mediators of Inflammation

    Article Title: Antilipotoxicity Activity of Osmanthus fragrans and Chrysanthemum morifolium Flower Extracts in Hepatocytes and Renal Glomerular Mesangial Cells

    doi: 10.1155/2017/4856095

    Figure Lengend Snippet: Effect of OF and CM on FFA-induced ROS production. HepG2 cells were incubated with 1 mM FFAs/BSA for 24 h in the presence of OF or CM. Vehicle cells were incubated with 0.1% DMSO in the presence of FFAs/BSA. Control cells were incubated with 1% BSA. Intracellular ROS production was quantified using the fluorescent probe DCFDA. Data were presented as mean ± SD of three independent experiments. Values not sharing common superscripts are significantly different ( p

    Article Snippet: The FFA/BSA or oleic acid (OA)/BSA complex solution was sterile-filtered through 0.22 μ m sterile filters (Millipore S.A.S., Molsheim, France) and then stored at −20°C until use.

    Techniques: Incubation

    Effects of OF and CM on lipid accumulation in free fatty acid-overloaded HepG2 cells. HepG2 cells were incubated with 1 mM FFAs/BSA and cotreated with various concentrations of OF and CM for 24 h. Vehicle cells were incubated with 0.1% DMSO in the presence of FFAs/BSA. Control cells were incubated with 1% BSA. Cell viability was measured by the MTT assay (a). Quantitative analysis of lipid deposition in cells by the OD 500 nm values using Oil Red O staining (b). Intracellular triglyceride (c) and cholesterol (d) contents were determined in cell lysates by an enzymatic colorimetric method using a commercially available kit. Total cholesterol and TG levels of the control cells were 25.3 ± 7.1 and 28.7 ± 2.7 μ g/mg of cellular protein, respectively. Data were presented as mean ± SD of three independent experiments. Values not sharing common superscripts are significantly different ( p

    Journal: Mediators of Inflammation

    Article Title: Antilipotoxicity Activity of Osmanthus fragrans and Chrysanthemum morifolium Flower Extracts in Hepatocytes and Renal Glomerular Mesangial Cells

    doi: 10.1155/2017/4856095

    Figure Lengend Snippet: Effects of OF and CM on lipid accumulation in free fatty acid-overloaded HepG2 cells. HepG2 cells were incubated with 1 mM FFAs/BSA and cotreated with various concentrations of OF and CM for 24 h. Vehicle cells were incubated with 0.1% DMSO in the presence of FFAs/BSA. Control cells were incubated with 1% BSA. Cell viability was measured by the MTT assay (a). Quantitative analysis of lipid deposition in cells by the OD 500 nm values using Oil Red O staining (b). Intracellular triglyceride (c) and cholesterol (d) contents were determined in cell lysates by an enzymatic colorimetric method using a commercially available kit. Total cholesterol and TG levels of the control cells were 25.3 ± 7.1 and 28.7 ± 2.7 μ g/mg of cellular protein, respectively. Data were presented as mean ± SD of three independent experiments. Values not sharing common superscripts are significantly different ( p

    Article Snippet: The FFA/BSA or oleic acid (OA)/BSA complex solution was sterile-filtered through 0.22 μ m sterile filters (Millipore S.A.S., Molsheim, France) and then stored at −20°C until use.

    Techniques: Incubation, MTT Assay, Staining

    Targeted cardiomyocyte deletion of growth differentiation factor 11 ( Gdf11 ) does not decrease total Gdf11 mRNA expression in mouse hearts. A : representative agarose gel image depicting Myh6 (wild-type band at 894 bp, Myh6-cre band at 300 bp) and Gdf11 [wild-type band at 359 bp, flox band at 393 bp, and ∆2–3 (postrecombination) band at 300 bp] alleles seen in the heart on day of life 0 – 1 by polymerase chain reaction (PCR) in Myh6 cre/wt , Gdf11 fl/fl , and Myh6 cre/wt ; Gdf11 fl/fl pups. B : recombination of Gdf11 at 6 mo of age in females and males in the heart but not in lung, liver, kidney, spleen, or skeletal muscle by PCR. C and D : Gdf11 expression in RNA extracted from whole heart in 3 ( n = 4/group, C )- or 6 ( n = 3–4/group, D )-mo-old female and male mice by quantitative PCR (qPCR). Data are normalized to TATA-binding protein ( Tbp ) and then to female Myh6 cre/wt control. E and F : Gdf11 expression in RNA extracted from isolated adult ( > 2 mo old) cardiomyocytes ( E ) or noncardiomyocytes ( F ), with each data point (shown with open symbols) representing isolated cells from a single mouse of the same genotype, n = 3/group (2 males, 1 female). Data were normalized to Tbp and then Myh6 cre/wt control. Cardiomyocytes ( E ), P value = not significant (ns, 0.05); noncardiomyocytes ( F ), P value = ns (0.3) by Kruskal-Wallis test. G and H : serum levels of GDF11 ( G ) and myostatin ( H ) as determined by quantitative mass spectrometry in 6-mo-old mice; n = 3–4/group. * P

    Journal: American Journal of Physiology - Heart and Circulatory Physiology

    Article Title: Analysis of Cre-mediated genetic deletion of Gdf11 in cardiomyocytes of young mice

    doi: 10.1152/ajpheart.00615.2018

    Figure Lengend Snippet: Targeted cardiomyocyte deletion of growth differentiation factor 11 ( Gdf11 ) does not decrease total Gdf11 mRNA expression in mouse hearts. A : representative agarose gel image depicting Myh6 (wild-type band at 894 bp, Myh6-cre band at 300 bp) and Gdf11 [wild-type band at 359 bp, flox band at 393 bp, and ∆2–3 (postrecombination) band at 300 bp] alleles seen in the heart on day of life 0 – 1 by polymerase chain reaction (PCR) in Myh6 cre/wt , Gdf11 fl/fl , and Myh6 cre/wt ; Gdf11 fl/fl pups. B : recombination of Gdf11 at 6 mo of age in females and males in the heart but not in lung, liver, kidney, spleen, or skeletal muscle by PCR. C and D : Gdf11 expression in RNA extracted from whole heart in 3 ( n = 4/group, C )- or 6 ( n = 3–4/group, D )-mo-old female and male mice by quantitative PCR (qPCR). Data are normalized to TATA-binding protein ( Tbp ) and then to female Myh6 cre/wt control. E and F : Gdf11 expression in RNA extracted from isolated adult ( > 2 mo old) cardiomyocytes ( E ) or noncardiomyocytes ( F ), with each data point (shown with open symbols) representing isolated cells from a single mouse of the same genotype, n = 3/group (2 males, 1 female). Data were normalized to Tbp and then Myh6 cre/wt control. Cardiomyocytes ( E ), P value = not significant (ns, 0.05); noncardiomyocytes ( F ), P value = ns (0.3) by Kruskal-Wallis test. G and H : serum levels of GDF11 ( G ) and myostatin ( H ) as determined by quantitative mass spectrometry in 6-mo-old mice; n = 3–4/group. * P

    Article Snippet: The supernatant containing noncardiomyocytes and debris was plated in an uncoated tissue culture plate in DMEM–F-12–10% FBS for 3 h. The cardiomyocyte fraction in the pellet then underwent sequential gravity settling with low-speed centrifugation (12 g, 3 min) with calcium reintroduction followed by plating in laminin-coated plates for 3 h. After 3 h, both cardiomyocytes and noncardiomyocytes were washed and harvested in TRIzol.

    Techniques: Expressing, Agarose Gel Electrophoresis, Polymerase Chain Reaction, Mouse Assay, Real-time Polymerase Chain Reaction, Binding Assay, Isolation, Mass Spectrometry

    Cre mRNA expression is higher in Myh6 cre/wt compared with Myh6 cre/wt ; growth differentiation factor 11 ( Gdf11 ) fl/fl mice and is associated with myocardial fibrosis and increased cardiomyocyte size. A : representative histology sections of male and female Myh6 cre/wt , Gdf11 fl/fl , and Myh6 cre/wt ; Gdf11 fl/fl mice stained with Masson’s trichrome. B : global fibrosis (%blue pixels) shows increased fibrosis in male ( n = 5–9/group) Myh6 cre/wt mice compared with females ( n = 3–5/group). C : cardiomyocyte cross-sectional area is increased in male Myh6 cre/wt mice compared with male Gdf11 fl/fl and Myh6 cre/wt ; Gdf11 fl/fl mice. Cross-sectional area from 40 cardiomyocytes/mouse were averaged for each mouse, and then average cardiomyocyte cross-sectional area by mouse data was analyzed with n = 3–6 female mice and 5–10 male mice. * P

    Journal: American Journal of Physiology - Heart and Circulatory Physiology

    Article Title: Analysis of Cre-mediated genetic deletion of Gdf11 in cardiomyocytes of young mice

    doi: 10.1152/ajpheart.00615.2018

    Figure Lengend Snippet: Cre mRNA expression is higher in Myh6 cre/wt compared with Myh6 cre/wt ; growth differentiation factor 11 ( Gdf11 ) fl/fl mice and is associated with myocardial fibrosis and increased cardiomyocyte size. A : representative histology sections of male and female Myh6 cre/wt , Gdf11 fl/fl , and Myh6 cre/wt ; Gdf11 fl/fl mice stained with Masson’s trichrome. B : global fibrosis (%blue pixels) shows increased fibrosis in male ( n = 5–9/group) Myh6 cre/wt mice compared with females ( n = 3–5/group). C : cardiomyocyte cross-sectional area is increased in male Myh6 cre/wt mice compared with male Gdf11 fl/fl and Myh6 cre/wt ; Gdf11 fl/fl mice. Cross-sectional area from 40 cardiomyocytes/mouse were averaged for each mouse, and then average cardiomyocyte cross-sectional area by mouse data was analyzed with n = 3–6 female mice and 5–10 male mice. * P

    Article Snippet: The supernatant containing noncardiomyocytes and debris was plated in an uncoated tissue culture plate in DMEM–F-12–10% FBS for 3 h. The cardiomyocyte fraction in the pellet then underwent sequential gravity settling with low-speed centrifugation (12 g, 3 min) with calcium reintroduction followed by plating in laminin-coated plates for 3 h. After 3 h, both cardiomyocytes and noncardiomyocytes were washed and harvested in TRIzol.

    Techniques: Expressing, Mouse Assay, Staining