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Targeted inhibition of calpain reduces myocardial hypertrophy and fibrosis in mouse models of type 1 diabetes.

Li Y, Ma J, Zhu H, Singh M, Hill D, Greer PA, Arnold JM, Abel ED, Peng T - Diabetes (2011)

Bottom Line: Calpain activity, cardiomyocyte cross-sectional areas, and myocardial collagen deposition were significantly increased in both STZ-induced and OVE26 diabetic hearts, and these were accompanied by elevated expression of hypertrophic and fibrotic collagen genes.These effects were associated with a normalization of the nuclear factor of activated T-cell nuclear factor-κB and matrix metalloproteinase (MMP) activities in diabetic hearts.Thus targeted inhibition of calpain represents a potential novel therapeutic strategy for reversing diabetic cardiomyopathy.

View Article: PubMed Central - PubMed

Affiliation: Critical Illness Research, Lawson Health Research Institute, University of Western Ontario, London, Ontario, Canada.

ABSTRACT

Objective: Recently we have shown that calpain-1 activation contributes to cardiomyocyte apoptosis induced by hyperglycemia. This study was undertaken to investigate whether targeted disruption of calpain would reduce myocardial hypertrophy and fibrosis in mouse models of type 1 diabetes.

Research design and methods: Diabetes in mice was induced by injection of streptozotocin (STZ), and OVE26 mice were also used as a type 1 diabetic model. The function of calpain was genetically manipulated by cardiomyocyte-specific knockout Capn4 in mice and the use of calpastatin transgenic mice. Myocardial hypertrophy and fibrosis were investigated 2 and 5 months after STZ injection or in OVE26 diabetic mice at the age of 5 months. Cultured isolated adult mouse cardiac fibroblast cells were also investigated under high glucose conditions.

Results: Calpain activity, cardiomyocyte cross-sectional areas, and myocardial collagen deposition were significantly increased in both STZ-induced and OVE26 diabetic hearts, and these were accompanied by elevated expression of hypertrophic and fibrotic collagen genes. Deficiency of Capn4 or overexpression of calpastatin reduced myocardial hypertrophy and fibrosis in both diabetic models, leading to the improvement of myocardial function. These effects were associated with a normalization of the nuclear factor of activated T-cell nuclear factor-κB and matrix metalloproteinase (MMP) activities in diabetic hearts. In cultured cardiac fibroblasts, high glucose-induced proliferation and MMP activities were prevented by calpain inhibition.

Conclusions: Myocardial hypertrophy and fibrosis in diabetic mice are attenuated by reduction of calpain function. Thus targeted inhibition of calpain represents a potential novel therapeutic strategy for reversing diabetic cardiomyopathy.

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Proinflammatory cytokine expression in diabetic Capn4 knockout hearts. Five months after STZ injection, the mRNA levels of TGF-β1 (A) and TNF-α (B) in Capn4−/− (knockout; KO) and Capn4+/+ (wild-type; WT) hearts were determined by real-time RT-PCR. Data are mean ± SD from six different hearts in each group. *P < 0.05 vs. WT. Effects of calpain inhibition on MMP activity and cell proliferation. Cardiac fibroblast cells were isolated from WT and Tg-CAST mice and were incubated with normal (NG; 5.5 mmol/L) and high glucose (HG; 30 mmol/L) in the presence and absence of calpain inhibitor-III (10 μmol/L) for 24 h. Calpain activity in fibroblast cells (C) and MMP activity (D) in culture media were measured. Cell proliferation was determined by the 3-(4, 5-dimethyl thiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay (E) and BrdU incorporation assay (F). NF-κB activity was measured in fibroblasts (G; see details in research design and methods). Data are mean ± SD from at least three different cultures. *P < 0.05 vs. NG; #P < 0.05 vs. WT cells cultured in HG.
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Figure 7: Proinflammatory cytokine expression in diabetic Capn4 knockout hearts. Five months after STZ injection, the mRNA levels of TGF-β1 (A) and TNF-α (B) in Capn4−/− (knockout; KO) and Capn4+/+ (wild-type; WT) hearts were determined by real-time RT-PCR. Data are mean ± SD from six different hearts in each group. *P < 0.05 vs. WT. Effects of calpain inhibition on MMP activity and cell proliferation. Cardiac fibroblast cells were isolated from WT and Tg-CAST mice and were incubated with normal (NG; 5.5 mmol/L) and high glucose (HG; 30 mmol/L) in the presence and absence of calpain inhibitor-III (10 μmol/L) for 24 h. Calpain activity in fibroblast cells (C) and MMP activity (D) in culture media were measured. Cell proliferation was determined by the 3-(4, 5-dimethyl thiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay (E) and BrdU incorporation assay (F). NF-κB activity was measured in fibroblasts (G; see details in research design and methods). Data are mean ± SD from at least three different cultures. *P < 0.05 vs. NG; #P < 0.05 vs. WT cells cultured in HG.

Mentions: Inflammation plays an important role in myocardial fibrosis. Our recent study demonstrated that expression of proinflammatory cytokines correlates with myocardial fibrosis in the diabetic heart (25). In line with the inhibition of myocardial fibrosis, deletion of Capn4 significantly decreased TNF-α and TGF-β1 expression in diabetic Capn4−/− compared with Capn4+/+ mice (Fig. 7A and B). In addition, the number of mast cells was increased in the diabetic heart, which was reduced in Capn4−/− mice (Supplementary Fig. 7A and B).


Targeted inhibition of calpain reduces myocardial hypertrophy and fibrosis in mouse models of type 1 diabetes.

Li Y, Ma J, Zhu H, Singh M, Hill D, Greer PA, Arnold JM, Abel ED, Peng T - Diabetes (2011)

Proinflammatory cytokine expression in diabetic Capn4 knockout hearts. Five months after STZ injection, the mRNA levels of TGF-β1 (A) and TNF-α (B) in Capn4−/− (knockout; KO) and Capn4+/+ (wild-type; WT) hearts were determined by real-time RT-PCR. Data are mean ± SD from six different hearts in each group. *P < 0.05 vs. WT. Effects of calpain inhibition on MMP activity and cell proliferation. Cardiac fibroblast cells were isolated from WT and Tg-CAST mice and were incubated with normal (NG; 5.5 mmol/L) and high glucose (HG; 30 mmol/L) in the presence and absence of calpain inhibitor-III (10 μmol/L) for 24 h. Calpain activity in fibroblast cells (C) and MMP activity (D) in culture media were measured. Cell proliferation was determined by the 3-(4, 5-dimethyl thiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay (E) and BrdU incorporation assay (F). NF-κB activity was measured in fibroblasts (G; see details in research design and methods). Data are mean ± SD from at least three different cultures. *P < 0.05 vs. NG; #P < 0.05 vs. WT cells cultured in HG.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

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Figure 7: Proinflammatory cytokine expression in diabetic Capn4 knockout hearts. Five months after STZ injection, the mRNA levels of TGF-β1 (A) and TNF-α (B) in Capn4−/− (knockout; KO) and Capn4+/+ (wild-type; WT) hearts were determined by real-time RT-PCR. Data are mean ± SD from six different hearts in each group. *P < 0.05 vs. WT. Effects of calpain inhibition on MMP activity and cell proliferation. Cardiac fibroblast cells were isolated from WT and Tg-CAST mice and were incubated with normal (NG; 5.5 mmol/L) and high glucose (HG; 30 mmol/L) in the presence and absence of calpain inhibitor-III (10 μmol/L) for 24 h. Calpain activity in fibroblast cells (C) and MMP activity (D) in culture media were measured. Cell proliferation was determined by the 3-(4, 5-dimethyl thiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay (E) and BrdU incorporation assay (F). NF-κB activity was measured in fibroblasts (G; see details in research design and methods). Data are mean ± SD from at least three different cultures. *P < 0.05 vs. NG; #P < 0.05 vs. WT cells cultured in HG.
Mentions: Inflammation plays an important role in myocardial fibrosis. Our recent study demonstrated that expression of proinflammatory cytokines correlates with myocardial fibrosis in the diabetic heart (25). In line with the inhibition of myocardial fibrosis, deletion of Capn4 significantly decreased TNF-α and TGF-β1 expression in diabetic Capn4−/− compared with Capn4+/+ mice (Fig. 7A and B). In addition, the number of mast cells was increased in the diabetic heart, which was reduced in Capn4−/− mice (Supplementary Fig. 7A and B).

Bottom Line: Calpain activity, cardiomyocyte cross-sectional areas, and myocardial collagen deposition were significantly increased in both STZ-induced and OVE26 diabetic hearts, and these were accompanied by elevated expression of hypertrophic and fibrotic collagen genes.These effects were associated with a normalization of the nuclear factor of activated T-cell nuclear factor-κB and matrix metalloproteinase (MMP) activities in diabetic hearts.Thus targeted inhibition of calpain represents a potential novel therapeutic strategy for reversing diabetic cardiomyopathy.

View Article: PubMed Central - PubMed

Affiliation: Critical Illness Research, Lawson Health Research Institute, University of Western Ontario, London, Ontario, Canada.

ABSTRACT

Objective: Recently we have shown that calpain-1 activation contributes to cardiomyocyte apoptosis induced by hyperglycemia. This study was undertaken to investigate whether targeted disruption of calpain would reduce myocardial hypertrophy and fibrosis in mouse models of type 1 diabetes.

Research design and methods: Diabetes in mice was induced by injection of streptozotocin (STZ), and OVE26 mice were also used as a type 1 diabetic model. The function of calpain was genetically manipulated by cardiomyocyte-specific knockout Capn4 in mice and the use of calpastatin transgenic mice. Myocardial hypertrophy and fibrosis were investigated 2 and 5 months after STZ injection or in OVE26 diabetic mice at the age of 5 months. Cultured isolated adult mouse cardiac fibroblast cells were also investigated under high glucose conditions.

Results: Calpain activity, cardiomyocyte cross-sectional areas, and myocardial collagen deposition were significantly increased in both STZ-induced and OVE26 diabetic hearts, and these were accompanied by elevated expression of hypertrophic and fibrotic collagen genes. Deficiency of Capn4 or overexpression of calpastatin reduced myocardial hypertrophy and fibrosis in both diabetic models, leading to the improvement of myocardial function. These effects were associated with a normalization of the nuclear factor of activated T-cell nuclear factor-κB and matrix metalloproteinase (MMP) activities in diabetic hearts. In cultured cardiac fibroblasts, high glucose-induced proliferation and MMP activities were prevented by calpain inhibition.

Conclusions: Myocardial hypertrophy and fibrosis in diabetic mice are attenuated by reduction of calpain function. Thus targeted inhibition of calpain represents a potential novel therapeutic strategy for reversing diabetic cardiomyopathy.

Show MeSH
Related in: MedlinePlus