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Cellutron Inc neomyt isolation kit nc-6031
Neomyt Isolation Kit Nc 6031, supplied by Cellutron Inc, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/isolation+kit+nc-6031/neonatal+rat+mouse+cardiomyocyte+isolation+kit/pm37972593-200-15-15
Average 90 stars, based on 1 article reviews

neomyt isolation kit nc-6031 - by Bioz Stars, 2026-09

90/100 stars

Images

Isolation:

Article Title: GIP and exendin hybrid polypeptides
Article Snippet: For 15 rat hearts, 8 digestions with 6 mls of buffer (Cellutron D2 buffer) were sufficient.

Article Title: GIP analog and hybrid polypeptides with selectable properties
Article Snippet: The Cellutron Isolation kit (nc-6031) (Cellutron, NJ) and protocol was used for cell isolation.

Cell Isolation:

Article Title: GIP and exendin hybrid polypeptides
Article Snippet: For 15 rat hearts, 8 digestions with 6 mls of buffer (Cellutron D2 buffer) were sufficient.

Article Title: GIP analog and hybrid polypeptides with selectable properties
Article Snippet: The Cellutron Isolation kit (nc-6031) (Cellutron, NJ) and protocol was used for cell isolation.

Image Search Results

(A) Representative live and dead staining images of cardiomyocytes and cardiac fibroblasts cultured on different membranes for one and 7 days. Scale bars = 200 μm. (B) Seeding efficiencies of cardiomyocytes and cardiac fibroblasts on different membranes.

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: Gelatin/Polycaprolactone Electrospun Nanofibrous Membranes: The Effect of Composition and Physicochemical Properties on Postoperative Cardiac Adhesion

doi: 10.3389/fbioe.2021.792893

Figure Lengend Snippet: (A) Representative live and dead staining images of cardiomyocytes and cardiac fibroblasts cultured on different membranes for one and 7 days. Scale bars = 200 μm. (B) Seeding efficiencies of cardiomyocytes and cardiac fibroblasts on different membranes.

Article Snippet: The neonatal rat/mouse cardiomyocyte isolation kit (nc-6031) was purchased from Cellutron Life Technologies (United States).

Techniques: Staining, Cell Culture

(A) Representative immunofluorescence staining images of cardiomyocytes and cardiac fibroblasts cultured on different membranes for one and 5 days. Blue: nucleus. Red: cTNT. Green: vimentin. Scale bars = 100 μm. (B) Proliferation of cardiomyocytes and cardiac fibroblasts cultured on different nanofibrous membranes for one, three, five, and 7 days. (*** p < 0.001).

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: Gelatin/Polycaprolactone Electrospun Nanofibrous Membranes: The Effect of Composition and Physicochemical Properties on Postoperative Cardiac Adhesion

doi: 10.3389/fbioe.2021.792893

Figure Lengend Snippet: (A) Representative immunofluorescence staining images of cardiomyocytes and cardiac fibroblasts cultured on different membranes for one and 5 days. Blue: nucleus. Red: cTNT. Green: vimentin. Scale bars = 100 μm. (B) Proliferation of cardiomyocytes and cardiac fibroblasts cultured on different nanofibrous membranes for one, three, five, and 7 days. (*** p < 0.001).

Article Snippet: The neonatal rat/mouse cardiomyocyte isolation kit (nc-6031) was purchased from Cellutron Life Technologies (United States).

Techniques: Immunofluorescence, Staining, Cell Culture

Inhibition of miR-199a attenuated hypertrophy of neonatal rat ventricular cardiomyocytes (A) The expression of miR-199a in NRVMs with or without miR-199a inhibitor treatment (n = 3). (B) Representative confocal images of sarcomeric alpha actinin (SAA, red) and DAPI (nuclei, blue) staining of NRVMs in different groups. NRVM size in each group was calculated by ImageJ. Scale bar, 20 μm. Approximately 200 counted cells in each group. (C) The expression of hypertrophic markers ANP, BNP, β-MHC, and Acta1 in each NRVM group (n = 3). ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001.

Journal: Molecular Therapy. Nucleic Acids

Article Title: Adeno-associated virus-mediated delivery of anti-miR-199a tough decoys attenuates cardiac hypertrophy by targeting PGC-1alpha

doi: 10.1016/j.omtn.2020.11.007

Figure Lengend Snippet: Inhibition of miR-199a attenuated hypertrophy of neonatal rat ventricular cardiomyocytes (A) The expression of miR-199a in NRVMs with or without miR-199a inhibitor treatment (n = 3). (B) Representative confocal images of sarcomeric alpha actinin (SAA, red) and DAPI (nuclei, blue) staining of NRVMs in different groups. NRVM size in each group was calculated by ImageJ. Scale bar, 20 μm. Approximately 200 counted cells in each group. (C) The expression of hypertrophic markers ANP, BNP, β-MHC, and Acta1 in each NRVM group (n = 3). ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001.

Article Snippet: NRVMs were prepared according to the instructions of the neonatal cardiomyocyte isolation kit (nc-6031, Cellutron, Baltimore, MD, USA).

Techniques: Inhibition, Expressing, Staining

AAV-mediated miR-199a inhibition in vivo (A and B) Representative confocal images of wheat germ agglutinin (WGA, red) and DAPI (nuclei, blue) staining of hearts in 8-day-old, 4-week-old, and 13-week-old mice injected with AAV9-anti-miR-199a TuDs or AAV9-EGFP. Green, autofluorescence of cardiomyocytes with AAV9 virus infection. Scale bars, 200 μm; n = 4–5. (C) Calculated relative percentage of GFP-positive cells in the hearts of 8-day-old, 4-week-old, and 13-week-old mice injected with AAV9-anti-miR-199a TuDs or AAV9-EGFP. n = 4–5 different sections in each group. NS represents not significant compared with that in 8-day-old mice. (D) The expression of miR-199a in the hearts of 8-day-old, 4-week-old, and 13-week-old mice injected with AAV9-anti-miR-199a TuDs or AAV9-EGFP (n = 6). (E) Representative confocal images of WGA (red) and DAPI (nuclei, blue) staining of skeletal muscle, liver, and brain tissue in 13-week-old mice injected with AAV9-anti-miR-199a TuDs or AAV9-EGFP. Arrow, green autofluorescence of cardiomyocytes with AAV9 infection. Scale bars, 200 μm; n = 3. (F) The expression of miR-199a in liver, brain, and skeletal muscle of 13-week-old mice with AAV9-anti-miR-199a TuDs or AAV9-EGFP injection (n = 5). NS, not significant. (G) The plasma cytokine levels of IL-1β, IL-6, and IFN-γ in AAV-injected mice and wild-type control mice (n = 6). IFN-γ in each group was lower than 7.8 pg/mL. ∗p < 0.05, ∗∗p < 0.01.

Journal: Molecular Therapy. Nucleic Acids

Article Title: Adeno-associated virus-mediated delivery of anti-miR-199a tough decoys attenuates cardiac hypertrophy by targeting PGC-1alpha

doi: 10.1016/j.omtn.2020.11.007

Figure Lengend Snippet: AAV-mediated miR-199a inhibition in vivo (A and B) Representative confocal images of wheat germ agglutinin (WGA, red) and DAPI (nuclei, blue) staining of hearts in 8-day-old, 4-week-old, and 13-week-old mice injected with AAV9-anti-miR-199a TuDs or AAV9-EGFP. Green, autofluorescence of cardiomyocytes with AAV9 virus infection. Scale bars, 200 μm; n = 4–5. (C) Calculated relative percentage of GFP-positive cells in the hearts of 8-day-old, 4-week-old, and 13-week-old mice injected with AAV9-anti-miR-199a TuDs or AAV9-EGFP. n = 4–5 different sections in each group. NS represents not significant compared with that in 8-day-old mice. (D) The expression of miR-199a in the hearts of 8-day-old, 4-week-old, and 13-week-old mice injected with AAV9-anti-miR-199a TuDs or AAV9-EGFP (n = 6). (E) Representative confocal images of WGA (red) and DAPI (nuclei, blue) staining of skeletal muscle, liver, and brain tissue in 13-week-old mice injected with AAV9-anti-miR-199a TuDs or AAV9-EGFP. Arrow, green autofluorescence of cardiomyocytes with AAV9 infection. Scale bars, 200 μm; n = 3. (F) The expression of miR-199a in liver, brain, and skeletal muscle of 13-week-old mice with AAV9-anti-miR-199a TuDs or AAV9-EGFP injection (n = 5). NS, not significant. (G) The plasma cytokine levels of IL-1β, IL-6, and IFN-γ in AAV-injected mice and wild-type control mice (n = 6). IFN-γ in each group was lower than 7.8 pg/mL. ∗p < 0.05, ∗∗p < 0.01.

Article Snippet: NRVMs were prepared according to the instructions of the neonatal cardiomyocyte isolation kit (nc-6031, Cellutron, Baltimore, MD, USA).

Techniques: Inhibition, In Vivo, Staining, Injection, Virus, Infection, Expressing, Clinical Proteomics, Control

$AAV9-anti-miR-199a TuDs virus attenuated cardiac hypertrophy (A) The representative gross morphology (scale bars, 1 mm) and H&E staining (scale bars, 400 μm) of heart sections in control and cardiac hypertrophy mice injected with AAV9-anti-miR-199a TuDs or AAV9-EGFP. n = 8. (B) The ratio of heart weight (HW, mg) and tibia length (TL, mm) in different mouse groups. n = 8. (C) Representative images of WGA (green) and DAPI (nuclei, blue) staining of hearts in different mouse groups (left). Scale bars, 20 μm. Calculated relative cell areas of cardiomyocytes in different mouse groups (right). n = 8. (D) Representative M-mode echocardiography images in different mouse groups (top). The percentages of left ventricular ejection fraction (LVEF) and left ventricular fractional shortening (LVFS) in the different mouse groups (bottom). n = 8. (E) The expression of hypertrophic markers ANP, BNP, β-MHC, and Acta1 in different mouse groups. n = 8. (F) The expression of miR-199a in different mouse groups. n = 8 in each group. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001.$

Journal: Molecular Therapy. Nucleic Acids

Article Title: Adeno-associated virus-mediated delivery of anti-miR-199a tough decoys attenuates cardiac hypertrophy by targeting PGC-1alpha

doi: 10.1016/j.omtn.2020.11.007

Figure Lengend Snippet: AAV9-anti-miR-199a TuDs virus attenuated cardiac hypertrophy (A) The representative gross morphology (scale bars, 1 mm) and H&E staining (scale bars, 400 μm) of heart sections in control and cardiac hypertrophy mice injected with AAV9-anti-miR-199a TuDs or AAV9-EGFP. n = 8. (B) The ratio of heart weight (HW, mg) and tibia length (TL, mm) in different mouse groups. n = 8. (C) Representative images of WGA (green) and DAPI (nuclei, blue) staining of hearts in different mouse groups (left). Scale bars, 20 μm. Calculated relative cell areas of cardiomyocytes in different mouse groups (right). n = 8. (D) Representative M-mode echocardiography images in different mouse groups (top). The percentages of left ventricular ejection fraction (LVEF) and left ventricular fractional shortening (LVFS) in the different mouse groups (bottom). n = 8. (E) The expression of hypertrophic markers ANP, BNP, β-MHC, and Acta1 in different mouse groups. n = 8. (F) The expression of miR-199a in different mouse groups. n = 8 in each group. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001.

Article Snippet: NRVMs were prepared according to the instructions of the neonatal cardiomyocyte isolation kit (nc-6031, Cellutron, Baltimore, MD, USA).

Techniques: Virus, Staining, Control, Injection, Expressing

a Neonatal rat ventricular myocytes (NRVMs) were treated with phenylephrine (PE, 50 μM) for 24 and 48 h, respectively. Representative immunoblots of the hexosamine biosynthetic pathway (HBP) enzymes (Gfat1, Gnpnat1, Pgm3, and GalE) are shown. Quantification (right) indicates significant upregulation of the HBP by PE treatment in cultured cardiomyocytes. N = 6 for GalE quantification and n = 9 for all other proteins. b PE treatment led to strong increases of the HBP enzymes at the mRNA level. N = 3–6. Data are shown as mean ± SEM. Significance was calculated by two-way ANOVA, followed by Tukey’s test. *** p < 0.001. Source data are provided as a Source Data file.

Journal: Nature Communications

Article Title: Chronic activation of hexosamine biosynthesis in the heart triggers pathological cardiac remodeling

doi: 10.1038/s41467-020-15640-y

Figure Lengend Snippet: a Neonatal rat ventricular myocytes (NRVMs) were treated with phenylephrine (PE, 50 μM) for 24 and 48 h, respectively. Representative immunoblots of the hexosamine biosynthetic pathway (HBP) enzymes (Gfat1, Gnpnat1, Pgm3, and GalE) are shown. Quantification (right) indicates significant upregulation of the HBP by PE treatment in cultured cardiomyocytes. N = 6 for GalE quantification and n = 9 for all other proteins. b PE treatment led to strong increases of the HBP enzymes at the mRNA level. N = 3–6. Data are shown as mean ± SEM. Significance was calculated by two-way ANOVA, followed by Tukey’s test. *** p < 0.001. Source data are provided as a Source Data file.

Article Snippet: Isolation was done in accordance with the instruction of the cardiomyocyte isolation kit (Cellutron, #NC-6031).

Techniques: Western Blot, Cell Culture

a NRVMs were infected with adenovirus expressing either control GFP (Ad-GFP) or Gfat1 (Ad-Gfat1). Gfat1 protein level was examined by immunoblotting and quantified. GAPDH was used as a loading control. N = 5 for Ad-GFP; n = 6 for Ad-Gfat1. b Representative images of NRVMs stained for α-actinin (red) and the nucleus (DAPI, blue) are shown. Scale: 20 μm. Bar graph depicts relative fold changes in cardiomyocyte surface area normalized to the cells infected by Ad-GFP. N = 91 cells for Ad-GFP; n = 86 cells for Ad-Gfat1. At least three independent experiments were conducted with two to three samples/group/experiment. c Radioactive leucine was included in culture media after Gfat1 overexpression. Leucine incorporation was quantified as indication of protein synthesis. N = 5 for Ad-GFP; n = 6 for Ad-Gfat1. d Overexpression of Gfat1 in NRVMs led to upregulation of hypertrophic marker genes (Anf, Rcan1.4) at the protein levels. N = 5 for Anf; n = 4 for Rcan1. e The hypertrophic marker gene expression was augmented by Gfat1 overexpression at the mRNA levels, as examined by real-time PCR. N = 8 for Anf ; n = 3 for Rcan1 . Data are shown as mean ± SEM. Student’s t test (two-tailed) was conducted to calculate significance. *** p < 0.001. Source data are provided as a Source Data file.

Journal: Nature Communications

Article Title: Chronic activation of hexosamine biosynthesis in the heart triggers pathological cardiac remodeling

doi: 10.1038/s41467-020-15640-y

Figure Lengend Snippet: a NRVMs were infected with adenovirus expressing either control GFP (Ad-GFP) or Gfat1 (Ad-Gfat1). Gfat1 protein level was examined by immunoblotting and quantified. GAPDH was used as a loading control. N = 5 for Ad-GFP; n = 6 for Ad-Gfat1. b Representative images of NRVMs stained for α-actinin (red) and the nucleus (DAPI, blue) are shown. Scale: 20 μm. Bar graph depicts relative fold changes in cardiomyocyte surface area normalized to the cells infected by Ad-GFP. N = 91 cells for Ad-GFP; n = 86 cells for Ad-Gfat1. At least three independent experiments were conducted with two to three samples/group/experiment. c Radioactive leucine was included in culture media after Gfat1 overexpression. Leucine incorporation was quantified as indication of protein synthesis. N = 5 for Ad-GFP; n = 6 for Ad-Gfat1. d Overexpression of Gfat1 in NRVMs led to upregulation of hypertrophic marker genes (Anf, Rcan1.4) at the protein levels. N = 5 for Anf; n = 4 for Rcan1. e The hypertrophic marker gene expression was augmented by Gfat1 overexpression at the mRNA levels, as examined by real-time PCR. N = 8 for Anf ; n = 3 for Rcan1 . Data are shown as mean ± SEM. Student’s t test (two-tailed) was conducted to calculate significance. *** p < 0.001. Source data are provided as a Source Data file.

Article Snippet: Isolation was done in accordance with the instruction of the cardiomyocyte isolation kit (Cellutron, #NC-6031).

Techniques: Infection, Expressing, Control, Western Blot, Staining, Over Expression, Marker, Gene Expression, Real-time Polymerase Chain Reaction, Two Tailed Test

a NRVMs were first transfected with either control (ctrl si) or Gfat1 (Gfat1 si) siRNA. PE was then used to trigger cardiomyocyte hypertrophic growth. The cells were harvested for immunofluorescence staining with α-actinin antibody (red) and DAPI (blue). Scale: 20 μm. Quantification showed a significant decrease of cardiomyocyte surface area by Gfat1 silencing. N = 160 cells for ctrl si/veh; n = 102 cells for ctrl si/PE; n = 108 cells for Gfat1 si/veh; n = 140 cells for Gfat1 si/PE. At least three independent experiments were conducted with two to three samples/group/experiment. b Leucine incorporation assay showed Gfat1 knockdown led to strong suppression of cell growth compared with control siRNA. N = 4 for ctrl si/PE group; n = 6 for all other groups. c Representative immunoblots of Gfat1, Rcan1.1, Rcan1.4, and Anf after PE treatment are shown. Quantification is at the right. N = 3–8. d Gfat1 silencing in NRVMs inhibited hypertrophic marker expression at the mRNA levels, as examined by real-time PCR analysis. N = 3–6. Data are shown as mean ± SEM. Significance was calculated by two-way ANOVA, followed by Tukey’s test. *** p < 0.001. Source data are provided as a Source Data file.

Journal: Nature Communications

Article Title: Chronic activation of hexosamine biosynthesis in the heart triggers pathological cardiac remodeling

doi: 10.1038/s41467-020-15640-y

Figure Lengend Snippet: a NRVMs were first transfected with either control (ctrl si) or Gfat1 (Gfat1 si) siRNA. PE was then used to trigger cardiomyocyte hypertrophic growth. The cells were harvested for immunofluorescence staining with α-actinin antibody (red) and DAPI (blue). Scale: 20 μm. Quantification showed a significant decrease of cardiomyocyte surface area by Gfat1 silencing. N = 160 cells for ctrl si/veh; n = 102 cells for ctrl si/PE; n = 108 cells for Gfat1 si/veh; n = 140 cells for Gfat1 si/PE. At least three independent experiments were conducted with two to three samples/group/experiment. b Leucine incorporation assay showed Gfat1 knockdown led to strong suppression of cell growth compared with control siRNA. N = 4 for ctrl si/PE group; n = 6 for all other groups. c Representative immunoblots of Gfat1, Rcan1.1, Rcan1.4, and Anf after PE treatment are shown. Quantification is at the right. N = 3–8. d Gfat1 silencing in NRVMs inhibited hypertrophic marker expression at the mRNA levels, as examined by real-time PCR analysis. N = 3–6. Data are shown as mean ± SEM. Significance was calculated by two-way ANOVA, followed by Tukey’s test. *** p < 0.001. Source data are provided as a Source Data file.

Article Snippet: Isolation was done in accordance with the instruction of the cardiomyocyte isolation kit (Cellutron, #NC-6031).

Techniques: Transfection, Control, Immunofluorescence, Staining, Knockdown, Western Blot, Marker, Expressing, Real-time Polymerase Chain Reaction

$a Schematic representation of TRE-Gfat1 and αMHC-tTA models for generation of the cardiac-specific Gfat1 transgenic mouse model (TG). Control (either TRE-Gfat1 only or αMHC-tTA only) and TG (TRE-Gfat1;αMHC-tTA double transgenic) mice were maintained on doxycycline water to suppress the transgene Gfat1 expression. These mice were switched to regular water for 2 weeks to induce Gfat1 expression and then subjected to surgery. b Representative images of mouse heart sections stained with either hematoxylin & eosin or Masson’s trichrome at 3 weeks post sham or TAC. Scale: 1 mm. c Quantification showed a significant increase of fibrosis in the Gfat1 TG heart after TAC. N = 5 for ctrl/sham; n = 4 for TG/sham; n = 9 for ctrl/TAC; n = 7 for TG/TAC. d Wheat germ agglutinin (WGA) staining of cardiac sections (left) showed a significant increase in cardiomyocyte size (quantified at right) in the TG mice after TAC. Scale: 20 μm. N = 6 for ctrl/sham; n = 4 for TG/sham; n = 9 for ctrl/TAC; n = 7 for TG/TAC. e Measurements of heart weight/body weight ratio (HW/BW) indicate significant cardiac hypertrophic growth in the TG mice. f Heart weight/tibia length ratio (HW/TL) suggests that Gfat1 overexpression in the heart led to more profound cardiac hypertrophy by pressure overload. g Representative images of M-mode echocardiography. h Gfat1 overexpression exacerbated cardiac response by pressure overload as shown by a significant decrease in ejection fraction. i Fractional shortening measurement indicates that Gfat1 overexpression led to cardiac dysfunction after TAC in TG mice compared to control animals. N = 19 for ctrl/sham; n = 7 for TG/sham; n = 13 for ctrl/TAC; n = 10 for TG/TAC ( e – i ). Data are shown as mean ± SEM. Significance was calculated by two-way ANOVA, followed by Tukey’s test. *** p < 0.001. Source data are provided as a Source Data file.$

Journal: Nature Communications

Article Title: Chronic activation of hexosamine biosynthesis in the heart triggers pathological cardiac remodeling

doi: 10.1038/s41467-020-15640-y

Figure Lengend Snippet: a Schematic representation of TRE-Gfat1 and αMHC-tTA models for generation of the cardiac-specific Gfat1 transgenic mouse model (TG). Control (either TRE-Gfat1 only or αMHC-tTA only) and TG (TRE-Gfat1;αMHC-tTA double transgenic) mice were maintained on doxycycline water to suppress the transgene Gfat1 expression. These mice were switched to regular water for 2 weeks to induce Gfat1 expression and then subjected to surgery. b Representative images of mouse heart sections stained with either hematoxylin & eosin or Masson’s trichrome at 3 weeks post sham or TAC. Scale: 1 mm. c Quantification showed a significant increase of fibrosis in the Gfat1 TG heart after TAC. N = 5 for ctrl/sham; n = 4 for TG/sham; n = 9 for ctrl/TAC; n = 7 for TG/TAC. d Wheat germ agglutinin (WGA) staining of cardiac sections (left) showed a significant increase in cardiomyocyte size (quantified at right) in the TG mice after TAC. Scale: 20 μm. N = 6 for ctrl/sham; n = 4 for TG/sham; n = 9 for ctrl/TAC; n = 7 for TG/TAC. e Measurements of heart weight/body weight ratio (HW/BW) indicate significant cardiac hypertrophic growth in the TG mice. f Heart weight/tibia length ratio (HW/TL) suggests that Gfat1 overexpression in the heart led to more profound cardiac hypertrophy by pressure overload. g Representative images of M-mode echocardiography. h Gfat1 overexpression exacerbated cardiac response by pressure overload as shown by a significant decrease in ejection fraction. i Fractional shortening measurement indicates that Gfat1 overexpression led to cardiac dysfunction after TAC in TG mice compared to control animals. N = 19 for ctrl/sham; n = 7 for TG/sham; n = 13 for ctrl/TAC; n = 10 for TG/TAC ( e – i ). Data are shown as mean ± SEM. Significance was calculated by two-way ANOVA, followed by Tukey’s test. *** p < 0.001. Source data are provided as a Source Data file.

Article Snippet: Isolation was done in accordance with the instruction of the cardiomyocyte isolation kit (Cellutron, #NC-6031).

Techniques: Transgenic Assay, Control, Expressing, Staining, Over Expression

$a Schematic representation of Gfat1 fl/fl and α-MHC-MCM models for generation of cardiac-specific inducible Gfat1 conditional knockout mouse model. Both control (Gfat1 fl/fl only) and cKO (Gfat1 fl/fl ;αMHC-MCM) animals were injected with tamoxifen for 5 consecutive days, followed by sham or TAC surgery. b Representative images of mouse heart sections stained with either hematoxylin & eosin or Masson’s trichrome at 3 weeks after surgery. Scale: 1 mm. Relative fibrosis was quantified at the right. N = 6 for cKO/TAC; n = 5 for all other groups. c Cardiomyocyte WGA staining was conducted using cardiac tissue sections (left). Scale: 20 μm. Bar graph depicts quantification of cross-sectional area normalized to the ctrl/sham group (right). N = 5 for Gfat1 fl/fl ; n = 6 for cKO. d Ratios of heart weight/body weight (HW/BW) and heart weight/tibia length (HW/TL) indicate that Gfat1 deletion suppressed cardiac hypertrophic growth. N = 6 for Gfat1 fl/fl /sham; n = 6 for cKO/sham; n = 5 for Gfat1 fl/fl /TAC; n = 7 for cKO/TAC. e Representative images of M-mode echocardiography. f Quantification of ejection fraction showed an improvement in cardiac systolic function in the cKO mice after TAC. N = 6 for Gfat1 fl/fl /sham; n = 6 for cKO/sham; n = 5 for Gfat1 fl/fl /TAC; n = 7 for cKO/TAC. g Fractional shortening measurement indicates that Gfat1 deletion in the heart improved cardiac response by pressure overload. N = 6 for Gfat1 fl/fl /sham; n = 6 for cKO/sham; n = 5 for Gfat1 fl/fl /TAC; n = 7 for cKO/TAC. Data are shown as mean ± SEM. Significance was calculated by two-way ANOVA, followed by Tukey’s test. *** p < 0.001. Source data are provided as a Source Data file.$

Journal: Nature Communications

Article Title: Chronic activation of hexosamine biosynthesis in the heart triggers pathological cardiac remodeling

doi: 10.1038/s41467-020-15640-y

Figure Lengend Snippet: a Schematic representation of Gfat1 fl/fl and α-MHC-MCM models for generation of cardiac-specific inducible Gfat1 conditional knockout mouse model. Both control (Gfat1 fl/fl only) and cKO (Gfat1 fl/fl ;αMHC-MCM) animals were injected with tamoxifen for 5 consecutive days, followed by sham or TAC surgery. b Representative images of mouse heart sections stained with either hematoxylin & eosin or Masson’s trichrome at 3 weeks after surgery. Scale: 1 mm. Relative fibrosis was quantified at the right. N = 6 for cKO/TAC; n = 5 for all other groups. c Cardiomyocyte WGA staining was conducted using cardiac tissue sections (left). Scale: 20 μm. Bar graph depicts quantification of cross-sectional area normalized to the ctrl/sham group (right). N = 5 for Gfat1 fl/fl ; n = 6 for cKO. d Ratios of heart weight/body weight (HW/BW) and heart weight/tibia length (HW/TL) indicate that Gfat1 deletion suppressed cardiac hypertrophic growth. N = 6 for Gfat1 fl/fl /sham; n = 6 for cKO/sham; n = 5 for Gfat1 fl/fl /TAC; n = 7 for cKO/TAC. e Representative images of M-mode echocardiography. f Quantification of ejection fraction showed an improvement in cardiac systolic function in the cKO mice after TAC. N = 6 for Gfat1 fl/fl /sham; n = 6 for cKO/sham; n = 5 for Gfat1 fl/fl /TAC; n = 7 for cKO/TAC. g Fractional shortening measurement indicates that Gfat1 deletion in the heart improved cardiac response by pressure overload. N = 6 for Gfat1 fl/fl /sham; n = 6 for cKO/sham; n = 5 for Gfat1 fl/fl /TAC; n = 7 for cKO/TAC. Data are shown as mean ± SEM. Significance was calculated by two-way ANOVA, followed by Tukey’s test. *** p < 0.001. Source data are provided as a Source Data file.

Article Snippet: Isolation was done in accordance with the instruction of the cardiomyocyte isolation kit (Cellutron, #NC-6031).

Techniques: Knock-Out, Control, Injection, Staining

a Knockdown of either Raptor or Rictor reduced Gfat1-mediated cardiomyocyte growth. NRVMs were transfected by siRNAs against Raptor or Rictor along with controls. Adenovirus expressing GFP or Gfat1 was then used to infect the cells. After immunofluorescent staining, cardiomyocyte size was quantified (right). Scale: 20 μM. N = 55–153/cells for each group. At least three independent experiments were conducted with 2–3 samples/group/experiment. b Protein expression of βMHC and Rcan1 was determined by Western blotting. Quantification at the right showed a significant decrease in expression by knockdown of either Raptor or Rictor, indicating both mTORC1 and mTORC2 contribute to Gfat1-mediated cardiomyocyte growth. N = 5–8. Data are shown as mean ± SEM. Significance was calculated by two-way ANOVA, followed by Tukey’s test. *** p < 0.001. Source data are provided as a Source Data file.

Journal: Nature Communications

Article Title: Chronic activation of hexosamine biosynthesis in the heart triggers pathological cardiac remodeling

doi: 10.1038/s41467-020-15640-y

Figure Lengend Snippet: a Knockdown of either Raptor or Rictor reduced Gfat1-mediated cardiomyocyte growth. NRVMs were transfected by siRNAs against Raptor or Rictor along with controls. Adenovirus expressing GFP or Gfat1 was then used to infect the cells. After immunofluorescent staining, cardiomyocyte size was quantified (right). Scale: 20 μM. N = 55–153/cells for each group. At least three independent experiments were conducted with 2–3 samples/group/experiment. b Protein expression of βMHC and Rcan1 was determined by Western blotting. Quantification at the right showed a significant decrease in expression by knockdown of either Raptor or Rictor, indicating both mTORC1 and mTORC2 contribute to Gfat1-mediated cardiomyocyte growth. N = 5–8. Data are shown as mean ± SEM. Significance was calculated by two-way ANOVA, followed by Tukey’s test. *** p < 0.001. Source data are provided as a Source Data file.

Article Snippet: Isolation was done in accordance with the instruction of the cardiomyocyte isolation kit (Cellutron, #NC-6031).

Techniques: Knockdown, Transfection, Expressing, Staining, Western Blot

a Pharmacological inhibition of mTOR by rapamycin or Torin 1 prevented cardiomyocyte growth from Gfat1 overexpression. Adenovirus-mediated overexpression of Gfat1 led to an increase in cardiomyocyte size, which was suppressed by either rapamycin (20 nM) or Torin 1 (50 nM) treatment in NRVMs. Scale: 20 μM. N = 80 cells for Ad-GFP/veh; n = 88 cells for Ad-Gfat1/veh; n = 143 cells for Ad-GFP/rapa; n = 104 cells for Ad-Gfat1/rapa; n = 170 cells for Ad-GFP/Torin 1; n = 120 cells for Ad-Gfat1/Torin 1. At least three independent experiments were conducted with two to three samples/group/experiment. b Inhibition of mTOR caused a decrease in Rcan1.4 expression in NRVMs. Western blotting was conducted to evaluate the expression Rcan1. In contrast to Rcan1.4, Rcan1.1 expression remained unchanged. N = 4. c Rapamycin treatment in vivo inhibited the hypertrophic growth by Gfat1 overexpression, as assessed by the ratios of heart weight/body weight (HW/BW) and heart weight/tibia length (HW/TL). Note that all mice underwent TAC surgery. N = 4 for control/veh; n = 3 for Gfat1-TG/veh; n = 7 for control/rapa; n = 4 for Gfat1-TG/rapa. d Inhibition of mTOR by rapamycin improved cardiac systolic performance in Gfat1 transgenic mice. N = 4 for control/veh; n = 3 for Gfat1-TG/veh; n = 7 for control/rapa; n = 4 for Gfat1-TG/rapa. Data are shown as mean ± SEM. Significance was calculated by two-way ANOVA, followed by Tukey’s test. *** p < 0.001. Source data are provided as a Source Data file.

Journal: Nature Communications

Article Title: Chronic activation of hexosamine biosynthesis in the heart triggers pathological cardiac remodeling

doi: 10.1038/s41467-020-15640-y

Figure Lengend Snippet: a Pharmacological inhibition of mTOR by rapamycin or Torin 1 prevented cardiomyocyte growth from Gfat1 overexpression. Adenovirus-mediated overexpression of Gfat1 led to an increase in cardiomyocyte size, which was suppressed by either rapamycin (20 nM) or Torin 1 (50 nM) treatment in NRVMs. Scale: 20 μM. N = 80 cells for Ad-GFP/veh; n = 88 cells for Ad-Gfat1/veh; n = 143 cells for Ad-GFP/rapa; n = 104 cells for Ad-Gfat1/rapa; n = 170 cells for Ad-GFP/Torin 1; n = 120 cells for Ad-Gfat1/Torin 1. At least three independent experiments were conducted with two to three samples/group/experiment. b Inhibition of mTOR caused a decrease in Rcan1.4 expression in NRVMs. Western blotting was conducted to evaluate the expression Rcan1. In contrast to Rcan1.4, Rcan1.1 expression remained unchanged. N = 4. c Rapamycin treatment in vivo inhibited the hypertrophic growth by Gfat1 overexpression, as assessed by the ratios of heart weight/body weight (HW/BW) and heart weight/tibia length (HW/TL). Note that all mice underwent TAC surgery. N = 4 for control/veh; n = 3 for Gfat1-TG/veh; n = 7 for control/rapa; n = 4 for Gfat1-TG/rapa. d Inhibition of mTOR by rapamycin improved cardiac systolic performance in Gfat1 transgenic mice. N = 4 for control/veh; n = 3 for Gfat1-TG/veh; n = 7 for control/rapa; n = 4 for Gfat1-TG/rapa. Data are shown as mean ± SEM. Significance was calculated by two-way ANOVA, followed by Tukey’s test. *** p < 0.001. Source data are provided as a Source Data file.

Article Snippet: Isolation was done in accordance with the instruction of the cardiomyocyte isolation kit (Cellutron, #NC-6031).

Techniques: Inhibition, Over Expression, Expressing, Western Blot, In Vivo, Control, Transgenic Assay

a Inhibition of O-GlcNAcylation with alloxan led to a decrease in cardiomyocyte growth from Gfat1 overexpression. Adenovirus-mediated overexpression of Gfat1 increased cardiomyocyte size, which was reduced by alloxan treatment in NRVMs. Scale: 20 μM. N = 80 cells for Ad-GFP/veh; n = 88 cells for Ad-Gfat1/veh; n = 135 cells for Ad-GFP/alloxan; n = 118 cells for Ad-Gfat1/alloxan. At least three independent experiments were conducted with two to three samples/group/experiment. b Suppression of O-GlcNAcylation caused a decrease in the expression of molecular markers of cardiomyocyte hypertrophy in NRVMs. Western blotting was conducted. In contrast to Rcan1.4, Rcan1.1 expression remained unaltered. N = 8–29. c The mTOR signaling was significantly inhibited by alloxan treatment. N = 3–11. Data are shown as mean ± SEM. Significance was calculated by two-way ANOVA, followed by Tukey’s test. * p = 0.003; *** p < 0.001. Source data are provided as a Source Data file.

Journal: Nature Communications

Article Title: Chronic activation of hexosamine biosynthesis in the heart triggers pathological cardiac remodeling

doi: 10.1038/s41467-020-15640-y

Figure Lengend Snippet: a Inhibition of O-GlcNAcylation with alloxan led to a decrease in cardiomyocyte growth from Gfat1 overexpression. Adenovirus-mediated overexpression of Gfat1 increased cardiomyocyte size, which was reduced by alloxan treatment in NRVMs. Scale: 20 μM. N = 80 cells for Ad-GFP/veh; n = 88 cells for Ad-Gfat1/veh; n = 135 cells for Ad-GFP/alloxan; n = 118 cells for Ad-Gfat1/alloxan. At least three independent experiments were conducted with two to three samples/group/experiment. b Suppression of O-GlcNAcylation caused a decrease in the expression of molecular markers of cardiomyocyte hypertrophy in NRVMs. Western blotting was conducted. In contrast to Rcan1.4, Rcan1.1 expression remained unaltered. N = 8–29. c The mTOR signaling was significantly inhibited by alloxan treatment. N = 3–11. Data are shown as mean ± SEM. Significance was calculated by two-way ANOVA, followed by Tukey’s test. * p = 0.003; *** p < 0.001. Source data are provided as a Source Data file.

Article Snippet: Isolation was done in accordance with the instruction of the cardiomyocyte isolation kit (Cellutron, #NC-6031).

Techniques: Inhibition, Over Expression, Expressing, Western Blot

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