Effect of Astragalus Polysaccharides on Cardiac Dysfunction in db/db Mice with Respect to Oxidant Stress

Figure 1. Effects of APS on left ventricular hematodynamics in db/db mice. After anesthetization, two-dimensional targeted M-mode echocardiography images were taken, and left ventricular hemodynamic parameters were measured in C57BJ/6J mice and db/db mice with or without APS treatment (n=6 per group). (a)~(c) Representative M-mode echocardiographic images. (d)~(f) Left ventricular hematodynamic parameters. LVEDD: left ventricular end diastolic diameter; LVESD: left ventricular end-systolic diameter; LVEF: left ventricular ejection fraction. Values were presented as mean ± standard error of the mean (SEM). ∗P<0.05 versus C57BJ/6J control mice, and #P<0.05 versus db/db mice.

Figure 2. Effects of APS on ultra-structural pathological changes of myocardium in db/db mice. Left ventricular samples from C57BJ/6J mice and db/db mice with or without APS treatment (n=6 per group) were prepared and evaluated by transmission electron microscopy (TEM) analysis. (a)~(c) Representative transmission electron microscopy photographs of cardiomyocytes. Disrupted tubes, sarcomeres, and mitochondrial cristae were noted in cardiac myocytes from db/db mice (b), whereas TEM Features in hearts from APS-treated db/db mice (c) showed nearly normal-appearing mitochondrial cristae, tubes, and sarcomeres that resembled those from normal control (a) (original magnification:×22500).

Figure 3. Effects of APS on myocardial cell apoptosis in db/db mice. Left ventricle was harvested from db/db mice with or without APS treatment (n=6 per group) after sacrifice. C57BJ/6J mice were taken as the normal control (n=6 per group). Sections of myocardium were subjected to the TUNEL assay for evaluation of apoptosis. (a)~(c) The microphotographs of TUNEL analysis in myocardium (brown stained nuclei: TUNEL staining positive, showing apoptosis; blue stained nuclei: TUNEL staining negative, showing normal). (d) The illustration for the percentage of apoptotic cardiomyocytes.

Figure 4. Effects of APS on protein levels of apoptosis target genes in myocardium of db/db mice. Left ventricular tissues obtained from C57BJ/6J mice and db/db mice with or without APS treatment (n=6 per group) were prepared and subjected to Western blotting of apoptosis target genes. Membranes were stripped and reprobed for GAPDH as a loading control. Protein content was measured respectively and normalized as a ratio relative to the normal control. (a)~(c) Protein levels of Bax, Bcl-2, and caspase-3. (d) Representative autoradiographs of Western blots. Values were presented as mean ± standard error of the mean (SEM). ∗P<0.05 versus C57BJ/6J control mice, and #P<0.05 versus db/db mice.

Figure 5. APS inhibited ROS formation in myocardium of db/db mice. Tissue sections of the left ventricle myocardium were obtained from C57BJ/6J mice and db/db mice with or without APS treatment (n=6 per group). CM-H2DCFDA was employed to measure the ROS formation, and the myocardial H2O2 and ·OH concentration were analyzed utilizing InSpeak Microscopy Image Intensity Calibration Microspheres and ImagePro analysis software. (a)~(c) The fluorescence microphotographs showing ROS formation in myocardium (red: fluorescence for H2O2 and ·OH; blue: cardiomyocytes). (d) ROS formation in myocardium from left ventricle. Values were presented as mean ± standard error of the mean (SEM). ∗P<0.05 versus C57BJ/6J control mice, and #P<0.05 versus db/db mice.

Figure 6. Effects of APS on protein levels of oxidation target genes in myocardium of db/db mice. Left ventricular tissues obtained from C57BJ/6J mice and db/db mice with or without APS treatment (n=6 samples per group) were prepared and subjected to Western blotting of oxidation target genes. Membranes were stripped and reprobed for GAPDH as a loading control. Protein content was measured respectively and normalized as a ratio relative to the normal control. (a)~(f) Protein levels of Gpx, SOD2, t-JNK, p-JNK, catalase, p-p38 MAPK, t-p38 MAPK, t-ERK, and p-ERK. (g) Representative autoradiographs of Western blots. Values were presented as mean ± standard error of the mean (SEM). ∗P<0.05 versus C57BJ/6J control mice, and #P<0.05 versus db/db mice. Gpx: Glutathione Peroxidase; t/p-JNK: total/phosphorylated c-Jun N-terminal kinase; t/p-p38 MAPK: total/phosphorylated p38 Mitogen-activated protein kinase; t/p-ERK: total/phosphorylate extracellular signal-regulated kinase.