Trimetazidine inhibits cardiac fibrosis by reducing reactive oxygen species and downregulating connective tissue growth factor in streptozotocin-induced diabetic rats

Abstract

Diabetes may affect myocardial fibrosis through oxidative stress. Trimetazidine (TMZ) is an anti-anginal agent. The present study aimed to determine the modulatory effect of TMZ on reactive oxygen species (ROS) and connective tissue growth factor (CTGF) expression and to evaluate the potential of TMZ to improve diastolic function in streptozotocin (STZ)-induced diabetic rats. After treating STZ-induced diabetic rats with TMZ for 16 weeks, a decrease in malondialdehyde levels, cardiac collagen volume fraction, left ventricular (LV) end-diastolic pressure and protein expression of collagen-I (Col I), Col III and CTGF compared with those in diabetic control rats was observed. In vitro, TMZ inhibited Col I, Col III and CTGF protein expression in cardiac fibroblasts treated with high glucose and decreased intracellular ROS generation and hydroxyproline content in the cell culture medium of cardiac fibroblasts. TMZ markedly improved cardiac fibrosis and diastolic function in diabetic rats. This effect was associated with a reduction in ROS production and CTGF expression in cardiac fibroblasts. The present study suggests that TMZ may be beneficial for protecting the hearts of diabetic patients.

Keywords: cardiac fibrosis; connective tissue growth factor; diabetic rat; reactive oxygen species; trimetazidine.

Figure 1.

TMZ inhibits cardiac fibrosis in diabetic rats. (A) Masson staining of myocardial tissue from the different groups (magnification, ×200; scale bar, 50 µm); (B) TMZ reduced the content of hydroxyproline in myocardial tissue. (C) Western blot analysis indicated that TMZ reduced the protein expression of collagen-I and collagen-III in myocardial tissue from diabetic rats. Values are expressed as the mean ± standard deviation (n=8 animals/group). **P<0.01 and ***P<0.01 vs. N; ^#P<0.05 vs. C. C, vehicle-treated diabetic group; CVF, cardiac collagen volume fraction; N, normal group; TMZ, trimetazidine.

Figure 2.

(A) TMZ reduced the protein expression of CTGF in myocardial tissue from diabetic rats. (B) Effects of TMZ on the protein expression of SOD-2 and on MDA content in myocardial tissue in each group. Values are expressed as the mean ± standard deviation (n=8 animals/group). *P<0.05, **P<0.01 and ***P<0.01 vs. N; ^#P<0.05 vs. C. C, vehicle-treated diabetic group; CFs, cardiac fibroblasts; CTGF, connective tissue growth factor; MDA, malondialdehyde; N, normal group; TMZ, trimetazidine; SOD, superoxide dismutase.

Figure 3.

(A) Immunochemical stains for DAPI (blue), von Willebrand factor (red) and vimentin (green) for isolated CFs from 1–3 day old neonatal SD rats (fluorescence microscope, ×200; scale bar, 50 µm). CFs were determined by positive staining for vimentin and negative staining for von Willebrand factor. The purity of the CFs was >95%. (B) TMZ reduced the protein expression of collagen-I and collagen-III in cardiac fibroblasts. (C) TMZ reduced the content of hydroxyproline in the cell supernatant. Values are expressed as the mean ± standard deviation (n=3). *P<0.05, **P<0.01 and ***P<0.001 vs. NG; ^#P<0.05, ^##P<0.01 and ^###P<0.001 vs. HG. NG, normal glucose; HG, high glucose; TMZ, trimetazidine.

Figure 4.

Trimetazidine reduced the protein expression of CTGF in cardiac fibroblasts. Values are expressed as the mean ± standard deviation (n=3). *P<0.05, ***P<0.001 vs. NG; ^#P<0.05, ^##P<0.01 vs. HG. CTGF, connective tissue growth factor; HG, high glucose; NG, normal glucose; TMZ, trimetazidine.

Figure 5.

Effect of TMZ on reactive oxygen species production in neonatal rat cardiac fibroblasts. Values are expressed as the mean ± standard deviation (n=3). **P<0.01 vs. NG; ^#P<0.05 vs. HG. HG, high glucose; MFI, mean fluorescence intensity; NG, normal glucose; TMZ, trimetazidine.