Progressive decay of Ca2+ homeostasis in the development of diabetic cardiomyopathy

Abstract

Background: Cardiac dysfunction in diabetic cardiomyopathy may be associated with abnormal Ca2+ homeostasis. This study investigated the effects of alterations in Ca2+ homeostasis and sarcoplasmic reticulum Ca2+-associated proteins on cardiac function in the development of diabetic cardiomyopathy.

Methods: Sprague-Dawley rats were divided into 4 groups (n = 12, each): a control group, and streptozotocin-induced rat models of diabetes groups, examined after 4, 8, or 12 weeks. Evaluations on cardiac structure and function were performed by echocardiography and hemodynamic examinations, respectively. Cardiomyocytes were isolated and spontaneous Ca2+ spark images were formed by introducing fluorescent dye Fluo-4 and obtained with confocal scanning microscopy. Expressions of Ca2+-associated proteins were assessed by Western blotting.

Results: Echocardiography and hemodynamic measurements revealed that cardiac dysfunction is associated with the progression of diabetes, which also correlated with a gradual but significant decline in Ca2+ spark frequency (in the 4-, 8- and 12-week diabetic groups). However, Ca2+ spark decay time constants increased significantly, relative to the control group. Expressions of ryanodine receptor 2 (RyR2), sarcoplasmic reticulum Ca2+-2ATPase (SERCA) and Na+/Ca2+ exchanger (NCX1) were decreased, together with quantitative alterations in Ca2+regulatory proteins, FKBP12.6 and phospholamban progressively and respectively in the diabetic rats.

Conclusions: Ca2+ sparks exhibited a time-dependent decay with progression of diabetic cardiomyopathy, which may partly contribute to cardiac dysfunction. This abnormality may be attributable to alterations in the expressions of some Ca2+-associated proteins.

Figure 1

Echocardiography of the study subjects with ejection fraction and fractional shortening. (a) Representative echocardiograph. (b) Representative measurement of the parameters. (c) Ejection fraction (EF) values of groups A-D. (d) Fractional shortening (FS) values of groups A-D. *P < 0.05, compared with group A.

Figure 2

Cardiac hemodynamics. The representative hemodynamic curves recorded by polygraphy: (a) Group A; (b) Group B; (c) Group C; (d) Group D. X-axes represent time in second. The parameters: left ventricular systolic peak pressure (LVPSP) (e), left ventricular end-diastolic pressure (LVEDP) (f), +dP/dt_max(g), −dP/dt_max(h) in groups A-D. ±dP/dt_max: maximal ascending and descending rates of left ventricular pressure. *P < 0.05, compared with group A; ^#P < 0.05, compared with group B; ^▲P < 0.05, compared with group C.

Figure 3

Ca²⁺ sparks in groups A-D. (a) Images of Ca²⁺ sparks; (b) Ca²⁺ spark frequency; (c) peak amplitude of Ca²⁺ sparks; (d) FWHM of Ca²⁺ sparks; (e) rise time; (f) Tau; (g) SR Ca²⁺ load. *P < 0.05, compared with group A; ^#P < 0.05, compared with group B; ^▲P < 0.05, compared with group C.

Figure 4

RyR2 and FKBP12.6 in groups A-D. (a) Western blots of RyR2, FKBP12.6, and β-actin; (b) ratios of RyR2 to β-actin; and (c) ratios of FKBP12.6 to β-actin; *P < 0.05, compared with group A; ^#P < 0.05, compared with group B; ^▲P < 0.05, compared with group C.

Figure 5

SERCA and NCX1 in groups A-D. (a) Western blots of SERCA, NCX1, and β-actin; (b) ratios of SERCA to β-actin; and (c) ratios of NCX1to β-actin. *P < 0.05, compared with group A; ^#P < 0.05, compared with group B; ^▲P < 0.05, compared with group C.

Figure 6

PLB, PLB-Thr17 and PLB-Ser16 in groups A-D. (a) Western blots of PLB, PLB-Thr17, PLB-Ser16 and β-actin; Relative ratios of PLB (b), PLB-Thr17 (c) and PLB-Ser16 (d) to β-actin. *P < 0.05, compared with group A; ^#P < 0.05, compared with group B; ^▲P < 0.05, compared with group C.