Expression of cathepsin B and cystatin C and its potential role in diabetic rats

AIM: To observe the expressions of cathepsin B (CB) and cystatin C (CC) in different stage of diabetic rats and to investigates their potential roles.METHODS: Sixty rats were divided into diabetes mellitus group induced by intravenous injection of streptozotocin (55 mg/kg) and normal group injected with citrate buffer. Ten rats were sacrificed respectively at the end of fourth week, eighth week and sixteenth week in both groups. 24 h urine excretion was collected in rats before sacrifice. The blood and the kidney were also collected. The mRNA and protein expressions of CB and CC in kidney were detected by real time PCR and immunohistochemical staining, respectively.RESULTS: At the end of eighth week, the expression of Ccr, 24 h urinary protein excretion, CB, CC in diabetic rats increased significantly, compared to the results at the fourth week (P<0.01 or P<0.05). With the aggravation of diabetic nephropathy, the expressions of CC, colⅣ, FN and 24 h urinary protein excretion were up-regulated significantly (P<0.01 or P<0.05). The expression of CB in diabetic rats was up-regulated at eighth week significantly (P<0.01), whereas, at the end of sixteenth week it was down-regulated significantly (P<0.01). The 24 h urinary protein excretion, the expressions of colⅣ at protein and mRNA levels and FN were negatively correlated with CB (P<0.01).CONCLUSION: The unbalance of CB and CC exists in diabetic nephropathy renal tissue, which is likely to lead to the accumulation of extracellular matrix.     

Role of Rho kinase in the dysfunction of myocardium contraction in hemorrhagic shock rats

AIM: To explore the role of Rho kinase in myocardium contraction in hemorrhagic shock rats. METHODS: The rat cardiac papillary muscle and rat heart at different time points during hemorrhagic shock were isolated. The contraction of papillary muscle in response to isoprenaline and the hemodynamic parameters of the isolated hearts in vitro including left intraventricular systolic pressure (LVSP), the maximal change rate of left intraventricular pressure (±dp/dtmax) were measured. RESULTS: The activity of Rho kinase, the contractile response of cardiac papillary muscle to isoprenaline and the hemodynamic parameters of the hearts were decreased gradually at the 1 h, 2 h and 4 h after shock. Compared to the normal controls, the activity of Rho kinase, the contractile response of cardiac papillary muscle and the hemodynamic parameters of the hearts were decreased significantly. The change of Rho kinase was positively correlated with the contractile response and the hemodynamic parameters of the heart. Stimulation with U-46619 at concentration of 10-8 mmol/L increased the activity of Rho kinase and improved the contractile response of cardiac papillary muscle to isoprenaline and the hemodynamic parameters of the isolated heart. The effects of U-46619 on the above parameters were abolished by Rho kinase specific inhibitor Y-27632 at concentration of 10-5 mol/L. CONCLUSION: Rho kinase plays an important role in the regulation of cardiac contractility during hemorrhagic shock.     

Positive inotropic action of urocortin on isolated heart tissues of the spontaneously hypertensive rat

AIM:To study the effects of urocortin (Ucn) on the isolated heart tissues of spontaneously hypertensive rat (SHR). METHODS:Effects of Ucn on contractile force and heart rate were observed in the SHR and Wistar rat right atrium, left atrium and right ventricle strip. RESULTS:Ucn (1-10 nmol/L) concentration-dependently increased the contractile force in the SHR and Wistar rat isolated right atrium. Ucn increased the contractile force in the SHR by (31.1±14.9)% at 3 nmol/L and by (65.7±22.4)% at 10 nmol/L, and its inotropic effect was significantly greater than that in Wistar rat (P<0.01) at the same concentration. Ucn (1-10 nmol/L) concentration-dependently increased the contractile force in the SHR and Wistar rat isolated left atrium, this effect had no difference between SHR and Wistar rat. No effect of Ucn on the heart rate of the isolated right atrium and on the contractile force of the isolated right ventricle strip was observed. CONCLUSION:The results indicate that the inotropic responses to Ucn in SHR are significantly greater than those in Wistar rats, suggesting that the change in heart function related with Ucn in SHR may have an important role in the pathophysiology of the hypertension.     

Effect of ERK1/2 on ischemic preconditioning in diabetic rat hearts

AIM: To test the effect of ERK1/2 on ischemic preconditioning (IPC) in diabetic rat hearts. METHODS: The diabetic rat model was made with alloxan. After eight weeks, 24 rats were divided into 4 groups: non-diabetic IPC rats (group A); non-diabetic non-IPC rats (group B); diabetic IPC rats (group C); diabetic non-IPC rats (group D). ECGⅡ lead, left ventricular development pressure (LVDP), and first derivative of LVDP (±dp/dtmax) were recorded. Myocardial phosphorylation of extracellular signal regulated kinases1/2 (ERK1/2) was detected by Western-blotting. RESULTS: (1) The ventricular arrythmia score was significantly lower in group A than that in group C (P<0.01). (2) LVDP and +dp/dtmax % were significantly higher in group A than those in group B and group C (P<0.01). -dp/dtmax% was significantly higher in group A than that in group B. (3) Phosphorylation of myocardial ERK1/2 was significantly higher in group A than those in group B and group C. CONCLUSIONS: IPC reduced the severity of ischemia/rerperfusion- induced arrhythmias, improved cardiac function in non-diabetic rats. However, such beneficial effects of IPC were not observed in diabetic rats. The attenuation of cardioprotection by IPC in diabetic animals may be due to the decrease in the activation of myocardial ERK1/2.     

Effect of β3-adrenoceptor activation on ventricle fibrillation threshold and effective refractory period in rats with heart failure

AIM: To observe the effect of β₃-adrenoceptor (AR) on ventricle fibrillation threshold (VFT) and effective refractory period (ERP) in rats with heart failure.METHODS: Rats were randomized into control group and heart failure group. The expression of β₃- AR mRNA was detected with RT-PCR. The VFT, ERP, left ventricle end-systolic pressure(LVESP)，left ventricle end-diastolic pressure(LVEDP), +dp/dtmax and -dp/dtmax were measured at the same time with administration of BRL37344 (β₃-AR agonist).RESULTS: ① Both the expression of β₃-AR mRNA and the proportion (β₃/β₁+β₂+β₃) were increased in failure rats comparied with those in control rats (0.028 vs 0.011 and 5.4% vs 1.2%, P<0.05). ② ERP was longer in rats with heart failure than that in control group (70.5 ms±5.5 ms vs 59.5 ms±6.4 ms, P<0.05). No difference in ERP in rats with heart failure was observed before and after administration of BRL37344 (73.0 ms±4.8 ms vs 70.5 ms±5.5 ms, P>0.05). ③ VFT was lower in rats with heart failure than that in control group (10.9 mV±0.8 mV vs 30.5 mV±1.3 mV, P<0.05) and decreased obviously in rats with heart failure after administration of BRL37344 (7.1 mV±0.6 mV vs 10.9 mV±0.8 mV, P<0.05). The decrease in VFT correlated with the effect of LVESP, +dp/dtmax, -dp/dtmax with BRL37344 and the expression of β₃-AR mRNA (correlation coefficient: 0.788, 0.708, 0.759, 0.787; P<0.05). CONCLUSION: The expression of β₃-AR mRNA in left ventricle is obviously increased in rats with heart failure. The activation of β₃-AR has no effect on ERP but can decrease VFT which correlates with the effect of β₃-AR on LVESP, +dp/dtmax, -dp/dtmax and the expression of β₃-AR mRNA.     

Protective effect of aFGF delivery by ultrasound-targeted microbubble destruction on left ventricular function in diabetic cardiomyopathy rats

AIM：To observe the protective effect of delivery of acidic fibroblast growth factor (aFGF) to myocardium by ultrasound-targeted microbubble destruction (UTMD) on left ventricular function in diabetic cardiomyopathy (DCM) rats and to investigate the possible mechanisms. METHODS：Twenty-four rats were intraperitoneally injected with streptozocin to induce DCM and were randomly divided into DCM group and aFGF treatment group. Twelve healthy rats served as normal controls. The rats in aFGF treatment group were infused with SonoVue-aFGF mixed fluid through tail vein and UTMD was simultaneously performed. Four weeks after intervention, all rats underwent cardiac catheterization to mea-sure left ventricular end-systolic pressure (LVESP), left ventricular end-diastolic pressure (LVEDP) and the maximal increase/decrease rate of left ventricular pressure (LV±dp/dtmax). The microvessel density (MVD) of rat myocardial tissues was measured by immunohistochemical staining for CD31. The myocardial collagen volume fraction (CVF) was determined by improved Masson staining. The apoptotic index (AI) was detected by TUNEL method. RESULTS：Four weeks after intervention, the LVESP and LV±dp/dtmax in aFGF treatment group were significantly increased compared with DCM group (P<0.01), while the LVEDP in aFGF treatment group was significantly lower than that in DCM group (P<0.01). The MVD in aFGF treatment group was significantly increased compared with DCM group (P<0.01), but the CVF and AI in aFGF treatment group were significantly lower than those in DCM group (P<0.01). CONCLUSION: Delivery of aFGF to diabetic myocardium by UTMD could improve the left ventricular function of DCM rats and may be a new feasible therapeutic method for DCM.     

Effects of lipoteichoic acid-induced delayed preconditioning on cardioplegic arrest/reperfusion injury in donor rat heart

AIM: To study the potential effects of lipoteichoic acid (LTA)-induced delayed preconditioning (PC) on cardioplegic arrest/reperfusion injury in donor rat heart. METHODS: The rats were pretreated with LTA (1 mg/kg, ip) 24 h before the experiment, and the isolated hearts were subjected to arrested by cardioplegic solution and stored in Eurocollin's solution for 4 h by the Langendorff method, and to evaluate the changes of cardiac function at the reperfusion for 30 min and 60 min, to measure the amounts of MB isoenzyme of creatine kinase (CK-MB), lactate dehydrogenase (LDH) and total nitric oxide (NO) oxidation products in the coronary effluent, and to detect myocardial apoptosis on tissue samples of left ventricle at the end of reperfusion by TUNEL staining. RESULTS: Pretreated with LTA significantly improved the recovery of cardiac function with a significant increase in coronary flow (CF), left ventricular developed pressure (LVDP), maximal rate of left ventricular developed pressure (+dp/dtmax), and minimal rate of left ventricular decline pressure (-dp/dtmax) at 30 min and 60 min of reperfusion (all P<0.01), reduced CK-MB (P<0.01) and LDH (P<0.01) release and raised the concentrations of NO2-/NO3- in coronary effluent. In addition, LTA pretreatment obviously decreased myocardial apoptosis in left ventricle at the end of reperfusion (P<0.01). The protective effects were abrogated by pretreatment of the rats with L-nitroarginine methyl eater (L-NAME, 10 mg/kg, ip). CONCLUSION: LTA pretreatment significantly improves cardiac function after cardioplegic arrest/reperfusion injury in donor heart of rats, and endogenous NO plays an essential role as an effector of delayed PC induced by LTA.     

Relationship between expression of heme oxygenase-1 mRNA and pulmonary hypertention induced by hypoxic hypercapnia

AIM: To study the effect of chronic hypoxic hypercapnia on expression of heme oxygenase-1 (HO-1). METHODS: Sprague-Dawley rats were randomly divided into three groups: control group(A),hypoxic hypercapnic group(B), hypoxic hypercapnia+hemin group(C). HO-1 and HO-1 mRNA were observed in pulmonary arterioles by the technique of immunohistochemistry and in situ hybridization. RESULTS: ① mPAP and weight ratio of right ventricle (RV) to left ventricle plus septum (LV+S) were significantly higher in rats of B group than those of A and C group (P<0.01). Differences of mCAP were not significant in three groups(P>0.05). ② Blood CO concentration was significantly higher in rats of B group than that of A group (P<0.01), it was much higher in C group than that of B group(P<0.01). ③ Light microscopy showed that vessel well area/total area (WA/TA), density of medial smooth muscle cell (SMC) and media thickness of pulmonary arterioles were much higher in rats of B group than those of A and C group (P<0.01). ④ The observation by electron microscopy showed proliferation of medial smooth muscle cells and collageous fibers of pulmonary arterioles in rats of B group, hemin could reverse the changes mentioned above. ⑤ HO-1 and HO-1 mRNA in pulmonary arterioles was significantly higher in rats of B group than those of A group(P<0.01), and they were significantly higher in rats of C group than those of B group (P<0.01). CONCLUSION: Expression of HO-1 mRNA and HO-1 in pulmonary arterioles was enhanced by hypoxic hypercapnia. Hemin partly inhibited pulmonary hypertension and pulmonary vessel remodeling by enhancing the expression of HO-1 mRNA and HO-1.     

Expression of zinc-finger transcription factor Snail 1 in the kidney of diabetic rats

AIM: To explore the expression of Snail 1 in renal tissues of diabetic rats, and to investigate its contribution to the progression of diabetic nephropathy. METHODS: Streptozotocin-induced diabetic rats were randomly divided into 2, 4, 8, 12, 16, 20, 24 weeks groups and 16 week A, 20 week A and 24 week A groups. A groups were treated with insulin to control blood glucose to normal level from the 13th week. Control groups were set up in age-matched time points. Blood glucose, 24 h urine protein, serum creatinine (Scr) and kidney index of rats were measured. Periodic acid-silver (PAS) staining was used to observe the renal pathological changes. The mRNA and protein expressions of Snail 1 and FN in renal cortex were detected by RT-PCR and immunohistochemical staining, respectively. Western blotting was employed to detect the expression of Snail 1 protein in the renal cortex. RESULTS: The levels of blood glucose, Scr, kidney weight index were increased remarkably in diabetic rats as compared with those in control groups (P<0.05, P<0.01), and decreased remarkably in the insulin-treated rats as compared with those in the diabetic rats (P<0.05, P<0.01). The Snail 1 protein was not detected by immunohistochemical staining in normal renal tissues. However, strongly positive staining was observed in renal tubules of diabetic rats. A time-dependent loss of Snail 1 expression was detected in the kidney in insulin-treated rats. The Snail 1 protein and mRNA of Snail 1 and FN were significantly up-regulated in the diabetic rats as compared with those in controls (P<0.01), while down-regulated in the insulin-treated diabetic rats (P<0.01). A close positive relationship existed between the mRNA expression of Snail 1 and FN (r=0.800, P<0.01). The level of Snail 1 protein expression was positively correlated with blood glucose, urine protein, Scr, kidney index (r=0.877, 0.694, 0.522, 0.875, P<0.01).CONCLUSION: These findings suggest that Snail 1 gene and protein expression are up-regulated in the kidney of rats with diabetes and may be involved in the pathogenesis of diabetic nephropathy.     

Expression of angiogenic factors in myocardial tissue of diabetic rats

AIM：To observe the expression of angiogenesis factors in the myocardial tissue of streptozotocin-induced diabetic rats. METHODS：The diabetic rat model was induced by intraperitoneal injection of streptozotocin. After 12 weeks, the cardiac function was measured by MPA cardiac function analysis system. The myocardial collagen volume fraction (CVF) was assessed by Masson staining. The capillary vessels was quantified as the ratio of capillary to myocyte (C/M) using CD31 immunostaining. The expression levels of vascular endothelial growth factor (VEGF), angiopoietin (Ang)-1, endostatin and Ang-2 were observed by Western blotting. RESULTS：Compared with normal control group, the left ventricular end-diastolic pressure (LVEDP) was evidently increased (P<0.01), but left ventricular pressure rise maximum rate (+dp／dtmax), left ventricular pressure decrease maximum rate (-dp／dtmax) and the ratio of capillary/myocyte (C/M) were significantly decreased (P<0.05). The CVF and the expression level of endostatin were significantly increased, whereas the expression levels of VEGF and Ang-1 evidently decreased (both P<0.05) in diabetic rats. However, no marked difference in the expression of Ang-2 between the 2 groups was observed (P>0.05). CONCLUSION：Imbalances between the angiogenic factors (VEGF and Ang-1) and anti-angiogenic factors (endostatin) may play an important role in the pathogenesis of diabetic cardiomyopathy.     

Changes of diaphragmatic function and morphologic features in diabetic rats

AIM:To evaluate the changes of diaphragm contractile properties and morphologic features in experimental diabetic rats.METHODS:Diaphragm contractility was assessed by twitch kinetics,maximal tetanic force (P₀),tension-frequency relationship and fatigue index(FI)in vitro diaphragm strip preparations of 4th week diabetic Wistar rats,and compared with that of the control group.The structure of diaphragm was analysed in diabetic rats and control with Hematoxyline and eosin staining,Heidenhain staining and enzymatic histochemistry.RESULTS:The twitch (Pt),contraction time(CT),half relaxation time(RT_1/2)and FI decreased significantly in diabetic group as compared with the control group.There was no difference in Po between the two groups.The tension of diabetic diaphragm at frequencies of 25,50,75,100 and 125 Hz was significantly decreased compared with that of the normal rats.After diaphragm bundles were subjected to fatiguing stimulations and incubated with aminophylline at 250 mg/L for 30 minutes, the tension of diabetic diaphragm decreased remarkably at both high and low frequencies in comparison with that of the normal rats.In addition,the activity and optical density ofα-phosphoglycerol dehydrogenase in diaphragm from diabetic rats were significantly lower than that from control rats(P<0101).CONCLUSION:The reduction of contractile function,decreased diaphragm fatigue resistance,and damaged structure occurred in 4th week diabetic rat diaphragm.     

X-ray irradiation increases sensitivity of myocardium to ischemia/reperfusion injury in rats

AIM：To observe the sensitivity of myocardium to ischemia/reperfusion (I/R) injury in the rats with chest radiotherapy. METHODS：The radiation-induced heart disease model was established by local 20 Gy of X-ray irradiation in the chest. Male Wistar rats (n=42) were randomly divided into 6 groups:sham trauma group, trauma group, sham trauma+sham operation group, sham trauma +I/R group, trauma+sham operation group and trauma+I/R group. The rats were subjected to 30 min of ischemia and 1 h of reperfusion 2 week after trauma. The left ventricular developed pressure (LVDP) and ±dp/dtmax were recorded by BL-410 biological signal recording and analysis system. The serum cardiac troponin I (cTnI) and creatine kinase isoenzyme (CK-MB) were measured by ELISA. The myocardial infarct size was determined by nitroblue tetrazolium(NBT) staining method and BI2000 image analysis software. RESULTS：Compared with sham trauma+I/R group, significant decreases in LVDP and ±dp/dtmax were observed in trauma+I/R group (P<0.01) with significant increases in the infarct size and the concentrations of cTnI and CK-MB (P<0.01). CONCLUSION:Chest X-ray irradiation increases the sensitivity of myocardium to I/R injury in rats.     

Effects of extract of Ginkgo biloba on cognitive function and apoptosis of hippocampus neuronal cells in type 1 diabetic rats

AIM: To investigate the protective mechanism of extract of Ginkgo biloba (EGB) on apoptosis of hippocampus neuronal cells in type 1 diabetic encephalopathic rats. METHODS: Thirty-six male Sprague-Dauley rats were divided into 3 groups: control group, diabetic group and EGB-treated group. Streptozotocin was injected intraperitoneally to the animals in later two groups to induce diabetes. The rats in EGB-treated group were injected intraperitoneally with EGB, and the same volume of normal saline was injected to the rats in other groups. At the end of the 12th week, the spatial learning and memory abilities of rats in each group were examined by Morris water maze test. Blood glucose and serum insulin concentration were measured. The neuron densities in hippocampus were measured by Image-Pro Plus 6.0 software. The expressions of Bax, Bcl-2, caspase-3 were assayed by Western blotting and immunohistochemistry. RESULTS: Compared to control group, the level of blood glucose (P<0.01), the protein expression of Bax (P<0.01) and caspase-3 (P<0.01) in hippocampus neuronal cells, and the ratio of Bax/Bcl-2 (P<0.01) and the escape latency (P<0.01) in diabetic group, were significantly increased, while the serum insulin concentration (P<0.01), the neuronal density (P<0.05) in CA1,CA2 hippocampal regions and the platform searching score (P<0.01) were significantly deceased. After treated with EGB, the serum insulin concentration (P<0.05), the neuronal density (P<0.05) in CA1,CA2 hippocampal regions and the platform searching score (P<0.01) were significantly increased, while the level of blood glucose (P<0.01), the protein expression of Bax (P<0.05), caspase-3 (P<0.05) in hippocampus neuronal cells, the ratio of Bax/Bcl-2 (P<0.01) and the escape latency (P<0.05) were significantly deceased than those in diabetic group. The protein expression of Bcl-2 in hippocampus neuronal cells did not alter in any experimental rats. CONCLUSION: EGB improves the spatial learning and memory capacity in diabetic rats by decreasing the expression of Bax, Bax/Bcl-2 ratio and down-regulating caspase-3 to reduce neurocyte apoptosis and increase the neuron density in CA1, CA2 hippocampal regions, suggesting that effective regulation of neuron apoptosis associated genes may be one of the mechanisms for EGB to treat diabetic encephalopathy.     

Intervention of gossypol and relationship between cognitive function and expressions of 11β-HSD1 and GR in brain of type 2 diabetic rats

AIM: To study the effect of gossypol on the cognitive function of type 2 diabetic rats, and to explore its mechanism. METHODS: Thirty male Sprague-Dawley rats were divided into three groups randomly: normal group, type 2 diabetic group and gossypol treated group. After fed with high-fat diet for 4 weeks, the later two groups were injected with streptozotocin intraperitoneally to establish type 2 diabetic rat model. The animals in gossypol treated group were given gossypol at dosage of 15 mg/kg once per day for 4 weeks by gavage. Since 5th week, the times of gavages were changed into once per week at the same dosage and lasted to 12th week. Learning and memory abilities of rats were assayed with Morris water maze test. The concentration of blood glucose was measured by biochemical method. The levels of serum corticosterone and insulin were detected by enzyme-linked immunosorbent assay and radioimmunoassay, respectively. The protein expressions of 11β-HSD1 and GR in cerebral cortex and hippocampus were determined by Western blotting. The morphological changes of cerebral cortex and hippocampus were observed under light microscope and transmission electronic microscope, respectively. RESULTS: Compared to normal group, the karyopyknosis, dilation of golgiosome and mitochondria swelling of neuron from cerebral cortex and hippocampus were prominent in diabetic group. The concentrations of blood glucose, serum corticosterone and insulin increased significantly (P<0.01). Protein expression of GR decreased (P<0.05), 11β-HSD1 protein tended to increase. Platform searching score was lower (P<0.01) and escape latency was longer (P<0.01) in diabetic group. After treated with gossypol, the concentrations of blood glucose, serum corticosterone and insulin declined (P<0.01). The protein expression of 11β-HSD1 was decreased (P<0.05) and GR was increased (P<0.05). Escape latency was shorter (P<0.01) and platform searching score was increased (P<0.01). CONCLUSION: Gossypol may improve the cognitive function of type 2 diabetic rats. Decreasing the level of 11β-HSD1 and increasing GR protein in the brain may be involved in the mechanism.     

Blockade of L-type Ca2+ channels suppresses activation of calcineurin and development of right ventricle cardiac hypertrophy induced by chronic hypoxia

AIM: To study the role of calcineurin in the progression of right ventricle cardiac hypertrophy in the chronic hypoxia rats and examine the effect of Ca²⁺ channel blockers on the activation of calcineurin. METHODS: Sixty rats were divided into three groups: treatment group with amlodipine besylate ablets, chronic hypoxia group, normal control group with normal oxygen. The rats in treatment group and chronic hypoxia group were exposed to normobaric chronic hypoxia(10±0.5)% O₂ for 21 days. All hearts were removed immediately after dissection for further investigation. RESULTS: (1)The RV/(LV+S),RV/BW were significantly higher in hypoxia group than that of control group and treatment group(P<0.01,respectively); (2) Right ventricular cardiomyocytes [Ca²⁺]_i in treatment group were significantly higher than that of control group and lower that that of hypoxia group(P<0.01,respectively); (3) The activity of calcineurin of the heart in hypoxia group were significantly increased when compared with control group. Amlodipine besylate ablets apparently suppressed the activity of calcineurin(P<0.01). CONCLUSION: Calcineurin possibly plays a role in the progression of right ventricle cardiac hypertrophy in the chronic hypoxia rats;Blockade of L-type Ca²⁺ channels with amlodipine besylate ablets effectively prevents its development possibly by inhibition of calcium inflow and suppression of the calcineurin activity.     

An enzyme histochemical study of diaphragm in diabetic rats

AIM: The aim of this research is to study the earlier enzyme activity changes of the diaphragm in diabetic rats. METHODS: An enzyme histochemical method was used to observe the changes in the enzyme activities of dehydrogenases,hydrolases and oxidases in 4th week diabetic rat diaphragm. RESULTS: The activites of enzymes including SDH(Succinate dehydrogenase),MDH(Malate dehydrogenase), GDH(Glutamate dehydrogenase), ICDH(Isocitrate dehydrogenase), NADHD(NADH diaphorase), G-6-PD(Glucose-6-phosphate dehydrogenase), ACP(Acid phosphatase) and ANAE(Acid α-naphtyl acid esterase) were increased in diabetic diaphragm compared with the control. LDH (Lactate dehydrogenase)and CCO(Cytochrome oxidase) activities were decreased, whereas NADPHD(NADPH diaphorase) showed no changes in diabetic rats. Eleven kinds of enzyme were analysed with image analysis.Optical density (A) of SDH, MDH, GDH, ICDH, NADHD, G-6-PD, ACP and ANAE in diaphragm of diabetic rats were significantly higher than that of control rats (P<0.01). A value of LDH and CCO in diaphragm from diabetic rats were significantly lower than that of control rats (P<0.01). A value of NADPHD in diaphragm from diabetic rats showed no apparent alteration compared with the control rats(P>0.05). CONCLUSION: Increase in the aerobic capacity, decrease in the glycolytic capacity, and disturbance of lipid and energy metabolism were found in diaphragm of 4th week diabetic rats.     

Expression of CTGF and its role in the streptozotocin-induced diabetic rat kidney

AIM:To investigate the change of connective tissue growth factor(CTGF) expression and its role in streptozotocin-induced diabetic rat kidney. METHODS:Male SD rats were randomly divided into two groups: control(sham operated rats, group C，n=32) and diabetic rats (group DN，n=35). Rats in each group were sacrificed at 1, 2, 4, and 8 weeks respectively after induction of diabetes. Body weight（BW）, blood glucose（BG）, 24-hour urine volumn（UV）, kidney weight, KW/BW,24-hour urinary albumin excretion (24Ualb), creatinine clearance (Ccr), kidney weight (KW), KW/BW, glomerular area (A_G), proximal tubular area (A_T) and the width of GBM、TBM at each time point were measured. Expression of CTGF and α-SMA were detected by immunostaining. RESULTS:There was a significant increase of 24 h Ualb, Ccr, KW/BW, A_G, V_G and the expression of CTGF in glomeruli and tubuli from week 1 onward in diabetic rats compared with those in group C (P<0.05, P<0.01, respectively), and an increasing expression of α-SMA mainly located in dilated tubuli from week 4 was found in group DN, which was more evident at week 8 accompanied by the decrease in A_T. Diabetic rats also had a significant increase in A_T from week 1 onward, which peaked at week 4. CONCLUSION:In the early stage of DN, the time-dependently upregulated CTGF might mediate the renal hypertrophy, which might be associated with the subsequent tubular epithelial-mesenchymal transdifferentiation (EMT) and renal fibrosis.     

Expression of TGF-β1 and apoptosis in myocardium of diabetic rats

AIM: To study the pathophysiological mechanism of cardiomyopathy, the expression of TGF-β₁ and apoptosis in myocardium of diabetic rats were investigated. METHODS:The diabetes models were established by single intravenous injection of streptozotocin (50 mg/kg) in rats. By the method of immunochemistry, the expression of TGF-β₁ in the cardiomyocytes was detected as the index to evaluate the degree of fibrosis. The method of TUNEL was used to measure the cardiomyocyte apoptosis as the index to explore its importance in process of diabetic cardiomyopathy. RESULTS:① The weight of diabetic rats was apparently lower than that in the rats before the diabetic model was built (P<0.01), and the increase in weight in diabetic rats within three month was less than that in normal group. ② Compared with control group, the concentration of blood glucose was continually elevated during the experiment. ③ The expression of TGF-β₁ in the diabetic cardiac muscle was much more than that in normal group (P<0.01). ④ The apoptosis of myocardium measured by the method of TUNEL was apparent in the diabetic groups than that in normal one (P<0.01). However, no significance was detected in the different courses of diabetic groups. CONCLUSIONS:The apoptosis might play an important role in leading the diabetic cardiomyopathy to heart failure. The expression of TGF-β₁ in the myocardium of diabetic rats was more than that in normal and had an increasing trend in the procession of diabetic cardiomyopathy. TGF-β₁ might be a significant factor in diabetic myocardium fibrosis. Apoptosis might play an important role in the initial stage of diabetes, which promotes the diabetic cardiomyopathy to heart failure.     

Early treatment with aminoguanidine on level of plasma and renal AngⅡ in diabetic rats

AIM: To investigate the effect of aminoguanidine (AG) on plasma and renal levels of angiogenesis Ⅱ (AngⅡ), and to identify the relationship of AGEs with AngⅡ in STZ-induced diabetic rats. METHODS: Wistar rats were randomly assigned to three groups. Diabetes was induced, rats were then received AG in treatment group. At the end of 12th week, urine albumin excretion rate (UAER) and calculate creatinine clearance (Ccr) were detected. Periodic acid-Schiff reagent was used to evaluate renal pathology. Plasma and renal AngⅡ were analyzed by radioimmunoassay and immunohistochemistry, respectively. RESULTS: AG treatment significantly prevented the increase in UAER (P<0.01), renal pathology (P<0.01), and level of renal AngⅡ (P<0.01). However, plasma concentration of AngⅡ was higher than that in diabetic rats without AG treatment (P<0.01). CONCLUSION: AG down-regulates renal Ang Ⅱ level, probably by reducing the formation of AGEs, which may be one of the renoprotective factors in diabetic nephropathy. More proofs are needed to identify the result that plasma AngⅡ concentration increases in DMA group.