Effects of EGb761 on synaptophysin expression in hippocampus of vascular dementia rats

AIM: To investigate the effects of Ginkgo biloba extract 761 (Egb761) on synaptophysin (SYN) expression in hippocampus of vascular dementia (VD) rats.METHODS: VD rat models, established by repeatedly cerebral ischemia/reperfusion, were randomly divided into two groups: model group and EGb761 treated group (both n=30), and another 30 condition-matched rats were selected as the sham-operated controls. Spatial learning and memory abilities of rats were assessed by Morris water maze (MWM) task, and SYN expression in hippocampal formation of rats in different groups was detected by immunohistochemical technique and image analysis.RESULTS: The MWM escape latency (EL) in model group was highly longer than that in the sham-operated group (P<0.01), while the EL of EGb761-treated group was significantly shorter than that in model group, but still longer than that in the sham-operated group at 1 m, 2 m and 4 m after VD modeling operation (P<0.05 or P<0.01). Immunohistochemical analysis showed that the SYN immunoreactive expression in hippocampal formation in model group greatly decreased and mean optical density of SYN expression highly increased compared with both sham-operated group and EGb761-treated group at three time points (P<0.01).CONCLUSION: EGb761 can increase the expression of SYN in hippocampus, which may be one of important mechanisms of EGb761 in improving learning and memory dysfunction of VD rats.     

Expression of GAP-43 mRNA and protein following brachial plexus avulsion injury

AIM:To investigate the expression of GAP-43 mRNA and protein of the motor neurons in spinal cord following the brachial plexus avulsion injury. METHODS:In the present study, three kinds of models of brachial plexus avulsion injury were made: right C₇ anterior root avulsion (group A), C₇ anterior root avulsion with right C₅-T₁ posterior roots breaking (group B), right C₇ anterior root avulsion with hemi-transect between C₅ and C₆ segment of spinal cord (group C). The expression of GAP-43 mRNA in anterior horn of spinal cord was detected at 14 days after operation by SYBR green quantification RT-PCR technique. The amounts of GAP-43 positive neurons in spinal cord were detected at 1, 3, 7 and 14 days after operation by immunohistochemistry technique. RESULTS:In control group, the expression of GAP-43 mRNA was very low in anterior horn. By 14 days after operation, the expression of GAP-43 mRNA was evidently up-regulated compared with control group. GAP-43 positive neuron was observed in control group at 1st day and 3rd day after operation. GAP-43 positive neurons appeared at 7th day and peaked at 14th day after operation. The expression of GAP-43 mRNA and protein were maximum in group C, group B was the lowest. CONCLUSION:The expression of GAP-43 mRNA and GAP-43 protein were up-regulated following brachial plexus injury. The expression of GAP-43 protein results from the recombination of proteins. GAP-43 is closely related to the axon regeneration and functional reconstruction.     

Protective effects of extract of Ginkgo biloba on diaphragm of diabetic rats

AIM: To investigate the effects of extract of Ginkgo biloba (EGb) on diaphragm from diabetic rats. METHODS: Sprague-Dauley rats were divided into three groups: normal control, diabetic group and EGb treatment group. The morphologic changes of diaphragm tissues were studied by light and electron microscopy, the activities of succinate dehydrogenase (SDH), superoxide dismutase (SOD), nitric oxide synthase (NOS) and contents of malondialdehyde (MDA), nitric oxide (NO₂^-/NO₃-) in the diaphragm mitochondria were assayed by spectophotometer, respectively. RESULTS: The activities of SOD, SDH decreased in diabetic diaphragm mitochondria, but the activitiy of NOS, the contents of NO₂^-/NO₃^-, MDA increased compared with control group. The activities of SOD, SDH were increased as well as NOS were decreased and the contents of NO₂^-/NO₃^-, MDA decreased in EGb treatment group compared with the diabetic group. CONCLUSION: EGb may protects the diaphragm mitochondria of diabetic rats by enhancing the function of respiratory chain, anti-oxidation and decreasing NO level.     

Role of injury and phenotype shift of liver sinusoidal endothelial cells in the development of portal hypertension of cirrhosis in rats

AIM: To study the role of injury and phenotype shift of liver sinusoidal endothelial cells in the development of portal hypertension of liver cirrhosis in rats. METHODS: The rat liver cirrhosis model was established by peritoneal injection of dimethylnitrosamine (DMN) (at a dose of 10 mg·kg^-1, 3 times a week, for 4 weeks). The dynamic changes of liver cirrhosis were observed at different time points (1 day, 2 days, 3 days, 1 week, 2 weeks, 4 weeks, 6 weeks and 8 weeks). The pressure of portal vein (Ppv), the expression of CD44, von Willebrand factor (vWF), endothelin-1 (ET-1) mRNA and endothelial nitric oxide synthase (eNOS) mRNA, the serum hyaluronic acid (HA) content and liver ET-1 content were measured. RESULTS: Compared with the normal control rats, CD44 positive staining was weak in the 1 day model rats, and the numbers of fenestrae of sinusoidal endothelial cells (SECs) rapidly decreased, but serum HA content rapidly increased (P<0.05). vWF positive staining in the 2-day model rats was stronger than that in normal control rats (P<0.05). There was a positive correlation between the Ppv and the vWF expression, serum HA content in the DMN-induced liver cirrhosis rats (P<0.05). Compared with the normal control rats, ET-1 mRNA expression increased in the 2-day and 3-day model rats, and ET-1 content lightly increased. eNOS mRNA expression was stronger in the 1-day, 2-day and 3-day model rats than that in normal control rats, meanwhile eNOS always expressed at a low level. CONCLUSION: The injury and phenotype shift of SECs is a pathological basis in the development of portal hypertension of DMN-induced liver cirrhosis in rats. Imbalance of ET-1 and NO production increases intrahepatic resistance, which plays an important role in the development of portal hypertension.     

Astragalus injection inhibits the expression of JNK3 mRNA after hypoxia/hypoglycemia and reoxygenation in hippocampal neurons of rats

AIM: To investigate the effect of astragalus injection on the expression of c-jun N terminal kinase (JNK3) mRNA interrelated with apoptosis after hypoxia/hypoglycemia and reoxygenation in hippocampal neurons of rats. METHODS: The hippocampal neurons cultured for eight days were divided into 4 groups: normal control group, original astragalus injection group, hypoxia/hypoglycemia and reoxygenation group, astragalus injection group. Hypoxia/ hypoglycemia and reoxygenation group, astragalus injection group and original astragalus injection group were treated with hypoglycemia and reoxygenation after deprived of oxygen and glucose for 30 min. Methods of in situ hybridization and RT-PCR were used respectively to measure the expression of JNK3 mRNA after hypoxia/hypoglycemia and reoxygenation 0 h, 0.5 h, 2 h, 6 h, 24 h, 72 h and 120 h. RESULTS: In normal control group the volume of hippocampal neuronal nucleolus was accretion, cellular tuber was distinct and cytokinesis was dyed by yellow a lot. In hypoxia/hypoglycemia and reoxygenation group the hippocampal neuronal nucleolus was crimple, cellular tuber was shrinked, large number of cytokinesis was dyed by yellow and yellow granule was observed. Compared with normal control group, the numbers of JNK3 mRNA positive neuronal cells at each time point increased obviously in the hypoxia/hypoglycemia and reoxygenation group (P<0.05). The change of neuronal configuration and the numbers of JNK3 mRNA positive neuronal cells in original astragalus injection group accorded with hypoxia/ hypoglycemia and reoxygenation group (P>0.05). In astragalus injection group the hippocampal neuronal nucleolus was crimple slightly and segmental cytokinesis was dyed by yellow.Compared to hypoxia/hypoglycemia and reoxygenation group, the numbers of JNK3 mRNA positive neuronal cells at each time point were less obviously in the astragalus injection group besides 120 h (P<0.05). Compared to normal control group, the mean optic density of expression of JNK3 mRNA in hippocampal neurons of rats at each time point increased obviously in hypoxia/ hypoglycemia and reoxygenation group (P<0.05). Compared to hypoxia/hypoglycemia and reoxygenation group, the mean optic density of JNK3 mRNA expression at each time point in original astragalus injection group had no obvious change (P>0.05), however the mean optic density of JNK3 mRNA expression in hippocampal neurons of rats at each time point decreased obviously in the astragalus injection group besides 120 h (P<0.05). CONCLUSION: Astragalus injection inhibits the expression of JNK3 mRNA after hypoxia/hypoglycemia and reoxygenation, accordingly inhibits hippocampal neuronal apoptosis.     

Overexpression of integrin-linked kinase improves survival and proliferation of cardiac c-Kit+ cells

AIM: To investigate the effects of integrin-linked kinase (ILK) overexpression on survival and proliferation of cardiac c-Kit⁺ cells, and the role of ILK-overexpressing c-Kit⁺ cell transplantation in cardiac function in a rat myocardial infarction (MI) model.METHODS: Cardiac c-Kit⁺ cells were isolated from the hearts of neonatal Sprague-Dawley (SD) rats and cultured to prepare the ILK-c-Kit⁺ cells by infected with recombinant adenoviral vector harboring human wild-type ILK cDNA. The survival and proliferation of cardiac c-Kit⁺ cells were detected by cell counting and CCK-8 assay at 48 h after infection, respectively. The protein levels of cyclin D1 and proliferating cell nuclear antigen (PCNA) in the cardiac c-Kit⁺ cells were examined by Western blot. MI was induced by coronary artery ligation in 40 adult rats. After 15 min, ILK-c-Kit⁺ cells were transplanted into the hearts by myocardial injection at 3 different sites in the infracted zone and border zone. All rats were randomly divided into 4 groups:sham group, MI plus saline injection group (MI group), MI plus null vector-infected cardiac c-Kit⁺ cell injection group (Ad-null-c-Kit⁺ cell group), and MI plus ILK-overexpressing cardiac c-Kit⁺ cells injection group (ILK-c-Kit⁺ cell group), with 10 rats in each group. At 2 weeks after MI, the protein levels of c-Kit in MI hearts were investigated by immunohistochemical assay. At 4 weeks, left ventricular function was examined by hemodynamic measurement.RESULTS: The survival and proliferation of cardiac c-Kit⁺ cells and the protein levels of cyclin D1 and PCNA were enhanced by ILK overexpression compared with Ad-null group. In MI rat model, the number of c-Kit⁺ cells was increased by ILK-c-Kit⁺ cell injection compared with Ad-null-c-Kit⁺ cell group at 2 weeks after MI. Cardiac function was significantly improved in ILK-c-Kit⁺ cell-transplanted rats.CONCLUSION: ILK overexpression improves survival and proliferation of cardiac c-Kit⁺ cells by increasing the protein levels of cyclin D1 and PCNA. ILK-c-Kit⁺ cell transplantation enhances the therapeutic efficiency of cardiac c-Kit⁺ cells in the post-MI hearts of rats.     

Correlation between intraventricular pressure and early phase proto-oncogene expression in volume overload rats

AIM: To investigate the effect of intraventricular pressure change by volume overload (VOL) on expression of proto-oncogene c-fos,c-jun,c-myc and egr-1. METHODS: Left ventricular systolic pressure(LVSP) and left ventricular end diastolic pressure (LVEDP) of rat with VOL induced by aortacaval fistula operation and rats of control group were measured at 30 min, 1, 4, 6, 12 and 48 h after the operation,these mRNAs at the foregoing time points were measured by slot bloting method and quantified with densitometry. RESULTS: Be compared with the control group, VOL rats LVSP decreased (P＜0.05) and declined most remarkably at 48 h,LVEDP elevated significantly at 30 min (P＜0.05), reached a maximal value at 12 h and the levels of control group at 48 h (P＞0.05) after the operation.The proto-oncogene expression signals were not detected in the control,negative controls and VOL rats at 30 min after the operation. The c-fos,c-jun and egr-1 mRNA signals appeared earlier,at 1 h, and c-myc mRNA increased later at 4 h.All reached peak value at 4 h and then declined gradually.The c-fos mRNA were not detected at 48 h. The c-myc,c-0jun and egr-1 mRNA persisted throught the entire observation period from 1 h to 48 h. CONCLUSIONS: During VOL early phase the overload have effect on expression of the proto-oncogene mRNA,c-fos,c-jun and egr-1 mRNA appear earlier, c-myc later,egr-1,c-jun and c-myc persist longer period, but the expressions do not strengthen with the ventricule wall load increase.This sequential induction pattern may reflect the time course regularity of the proto-oncogenes expression induced by VOL,and indicate the proto-oncogenes expression initiate while the heart load accumulate some extent and duration and the load magnitude may not play a critical role.     

Cyclosporine A suppresses the hypertrophic effect of neuropeptide Y in rat cardiomyocytes

AIM:To investigate the effect of cyclosporine A on rat cardiomyocyte hypertrophy caused by neuropeptide Y (NPY).METHODS:Cardiomyocytes of neonatal Wistar rats were cultured with NPY or NPY together with CsA. For assessing protein synthesis rate and c-jun mRNA expression in cardiomyocytes, the methods of [³H]-Leu incorporation and RT-PCR were used.RESULTS:(1) [³H]-Leu incorporation in cardiomyocytes: [³H]-Leu incorporation in NPY (10 nmol/L) group was higher than that in control group, but there were no distinct changes between two groups. To compare with control group, [³H]-Leu incorporation in NPY (100 nmol/L) group were increased significantly (P＜0.05). There was no significant change between control group and CsA group; (2) c-jun mRNA expression in cardiomyocytes: RT-PCR production of c-jun mRNA in NPY group was enhanced considerably compared with CsA group and control group (P＜0.01). There was no significant change between CsA group and control group.CONCLUSIONS:NPY can induce cardiomyocyte hypertrophy. Cyclosporine A (inhibitor of CaN) can blunt the effect of NPY, suggesting that the Ca²⁺/CaM-dependent calcineurin (CaN) signaling pathway plays an important role in it.     

Expression of ROCK I and TGF-β1 in pulmonary arterioles of rat with chronic thromboembolism pulmonary hypertension

AIM:To investigate the dynamic expression of Rho kinase (ROCK I) and transforming growth factor β₁ (TGF-β₁) in pulmonary arterioles of rat with chronic thromboembolic pulmonary hypertension. METHODS: Sixty-four male Wister rats were randomly divided into eight groups: beginning control group, embolism for 3 d, 1 week, 2 weeks, 4 weeks, 8 weeks, 12 weeks groups and end control group. The pulmonary thromboembolism (PTE) model was established by injecting thrombin into jugular vein two times in two weeks and each rat underwent peritoneal injection with tranexamic acid one time a day during experiment to prevent thrombolysis. The mean pulmonary artery pressure (mPAP), right ventricular hypertrophy index (RVHI), relative medial thickness of small pulmonary arteries (PAMT) and vessel wall area/total area (WA/TA) were measured. The levels of ROCK I mRNA and TGF-β₁ protein in rat pulmonary artery were determined by in situ hybridization, immunohistochemistry and image analysis, respectively. RESULTS: mPAP, PAMT and WA/TA were higher respectively in embolism from 4 weeks group to 12 weeks group than those in beginning control group (mPAP: all P<0.01, PAMT and WA/TA: 4 weeks group P<0.05, 8 weeks group and 12 weeks group P<0.01). RVHI was elevated in 8 weeks group P<0.05, in 12 weeks group P<0.01. ROCK I mRNA and TGF-β₁ protein in pulmonary arterioles got the enhanced positive signals of in situ hybridization or immunohistochemistry staining with prolonging the time of rats with pulmonary thromboembolism. ROCKⅠ mRNA: embolism from 3 d group to 2 weeks group P<0.05, 4 weeks group to 12 weeks group P<0.01, TGF-β₁ protein: 1 week group and 2 weeks group P<0.05, 4 weeks group to 12 weeks group P<0.01. Linear correlation analysis showed that ROCK I mRNA and TGF-β₁ protein were positively correlated with mPAP, RVHI and vessel remodeling index (all P<0.01), ROCK I mRNA were positively correlated with TGF-β₁ protein (P<0.01). CONCLUSION:ROCK I and TGF-β₁ play a role in the pathogenesis of chronic thromboembolic pulmonary hypertension and pulmonary vessel remodeling. TGF-β₁ produces biological effect by active ROCK signal pathway.     

PDL1Ig gene-modified BMSCs induce immune tolerance in rat liver transplantation

AIM: To investigate the effects of bone marrow mesenchymal stem cells(BMSCs) modified by programed death ligand-1 immunoglobulin(PDL₁Ig) gene on immune rejection of orthotopic liver transplantation in rats. METHODS: Rat BMSCs were cultured and identified. The protein expression of PDL₁Ig in the BMSCs 72 h after infection with pAdEasy-1/PDL₁Ig was detected by Western blot. Mixed lymphocyte reaction was used to detect the inhibitory effect of BMSCs on the viability of T-lymphocytes in peripheral blood. The male Wistar rats were used as donors(n=40), and the male SD rats were used as recipients(n=40). The rat model of orthotopic liver transplantation was established by improved cuff method for observing acute rejection. The rats were randomly divided into control group, BMSCs treatment group, BMSCs/GFP treatment group and BMSCs/PDL₁Ig treatment group with 10 pairs each. Five rats were executed at the 7th day and the remains were used for measuring the survival time. RESULTS: The expression of PDL₁Ig in the BMSCs was detected after pAdEasy-1/PDL₁Ig infection. The effect of BMSCs/PDL₁Ig on the viability of the lymphocytes was stronger than that of BMSCs/GFP. The level of IL-4 in BMSCs/PDL₁Ig group was significantly higher than that in the other 3 groups, while the levels of IFN-γ and IL-2 were significantly decreased. The liver function in BMSCs/PDL₁Ig group was significantly improved and the levels of ALT, AST and TBil were almost recovered to normal at the 7th day after transplantation. Severe rejection reaction was observed in control group, and rejection reactions were decreased with different degrees in BMSCs treatment group and BMSCs/GFP treatment group. Much slighter rejection reaction and significantly longer survival time were showed in BMSCs/PDL₁Ig group than those in the other 3 groups. CONCLUSION: PDL₁Ig-modified BMSCs inhibit the rejection of liver transplantation in rats and induce the immune tolerance, and the effect is better than that of BMSCs alone.     

Long-term effects of TCV116 on left ventricular remodeling and heart function after myocardial infarction in rats

AIM: To investigate the effects of long-term TCV116 on left ventricular remodeling and heart function after myocardial infarction. METHODS: Myocardial infarction (MI) was caused by ligation of the left anterior descending coronary artery in rats. One week after the surgical performance, the surviving rats were randomly assigned to the following treatment protocols: (1) MI rats with no therapy; (2) MI rats treated with TCV116 2 mg/kg per day; (3) Sham-operated control; (4) Sham-operated rats, treated with TCV116 2 mg/kg per day. At 22 weeks, cardiac hemodynamic parameters such as MAP, LVSP, dp/dt_max and LVEDP, and histomorphometric parameters such as LVW/BW and LVCA/BW were measured, mRNA of cardiac genes such as βMHC, BNP, TGF-β₁, collagen I and III were quantified, and survival rates were calculated. RESULTS: Compared with sham-operated rats, MI rats without therapy showed significant increases in histomorphometric parameters as well as in mRAN expressions of cardiac genes (P<0.01); While their hemodynamic parameters were significantly impaired (P<0.01), and survival duration shortened (P<0.05). Compared with MI rats without therapy, MI rats treated with TCV116 showed significant attenuation of mRAN expression of cardiac genes (P<0.01); While their hemodynamic parameters were significantly improved (P<0.05 or P＜0.01), and survival duration extended (P<0.05). CONCLUSION: Treatment with long-term angiotensin II type 1 receptor antagonist may improve left ventricular remodeling and cardiac function after MI in rats.     

Influence of chronic psychological stress on periodontitis and serum isoenzyme LDH1 in rats

AIM: To investigate the role of chronic psychological stress on periodontitis and serum level of isoenzyme LDH1 in rats. METHODS: Eighty female special pathogen-free Wistar rats were randomly divided into 4 groups:(1) normal control group; (2) experimental periodontitis group: the periodontitis model was induced by wrapping 3/0 silk ligature inoculated with putative periodontopathic bacteria around the left maxillary second molar of the rats; (3) psychological stress stimulation group: the rats were treated with stress stimulation alone; (4) periodontitis model with stress stimulation group: the periodontitis models were induced by wrapping 3/0 silk ligature inoculated with putative periodontopathic bacteria around the left maxillary second molar of the rats, and then treated with stress stimulation. The rats were sacrificed at 2nd, 4th, 6th and 8th weeks after ligature. The levels of blood glucose and adrenocorticotropic hormone (ACTH) were measured as the stress markers, and the serum isoenzyme lactate dehydrogenase 1(LDH1) was used to evaluate the severity of hypoxia. The histological changes of periodontal tissues were observed under microscope with HE staining. RESULTS: The levels of blood glucose and ACTH in stress group and periodontitis with stress group were significantly higher than those in control group and experimental periodontitis group at the 2nd and 4th weeks after ligature (P<0.01). The level of serum LDH in periodontitis with stress group was significantly higher than that in control group. The level of serum LDH1 in periodontitis with stress group was significantly lower than that in control group (P<0.01). No difference of histological change in periodontal tissues was observed in control group and stress group. Severer inflammatory changes and alveolar bone destruction were observed in periodontitis with stress group than those in experimental periodontitis group. CONCLUSION: Stress stimulation is one of the inducing factors of periodontitis in rats that aggravates periodontitis by decreasing the tissue oxygenation.     

The SDF-1/CXCR4 axis repairs hypoxia-ischemia brain damage via-regulating oriented differentiation of mesenchymal stem cells

AIM:To explore the effect of stromal cell-derived factor-1(SDF-1)/CXC chemokine receptor 4(CXCR4)axis on the neural differentiation of rat mesenchymal stem cells(rMSCs) and recovery from hypoxia-ischemia brain damage(HIBD). METHODS:The rMSCs were isolated from the rat bone marrow, and expanded in vitro. The mRNA and protein levels of CXCR4 in rMSCs treated with SDF-1α(10 μg/L) and hypoxia for 0 h, 6 h, 12 h, 24 h, 48 h and 72 h were detected by RT-PCR, Western blotting and flow cytometry. The mRNA and protein levels of SDF-1α in the hippocampus of the rats with hypoxia-ischemia for 1 d, 3 d, 5 d, 7 d, 14 d and 21 d were also detected by the same methods. The protein levels of neuron-specific enolase(NSE) and glial fibrillary acidic protein(GFAP), as well as the positive rate of neural-induced rMSCs pretreated with AMD3100(a CXCR4 antagonist) at dose of 5 mg/L were determined by Western blotting and immunocytochemistry. RESULTS:Compared with the normal controls, both mRNA and protein levels of CXCR4 increased in rMSCs exposed to hypoxia for 6 h and 12 h, and the results were also confirmed by flow cytometry. As expected, the mRNA level of SDF-1α in the hippocampus of HIBD rats was higher than that in normal control rats(P<0.01). Moreover, the mRNA expression of CXCR4 was extremely up-regulated in rMSCs treated with SDF-1α at concentration of 10 μg/L, and the results were also confirmed by Western blotting and flow cytometry analysis(P<0.01). The protein levels and positive cell numbers of NSE and GFAP were extremely decreased in rMSCs pretreated with AMD3100 at concentration of 5 mg/L. CONCLUSION:Compared with normal rats, SDF-1α level in the hippocampus of the rats with hypoxia-ischemia is increased. Hypoxia and micro-dose of SDF-1α induce the expression of CXCR4 in rMSCs, while CXCR4 antagonist reduces the neural differentiation of rMSCs, suggesting that SDF-1/CXCR4 axis may be deeply involved in the neural differentiation of rMSCs during the process of repairing HIBD.     

CX3CR1 mediates the neuroprotective effect of triptolide on 1-methyl-4-phenylpyridinium-induced hemiparkinson rats

AIM: To investigate the effect of triptolide on the inhibition of microglial activation in 1-methyl-4-phenyl pyridinium (MPP⁺)-induced hemiparkinson disease rats.METHODS: The rat model of Parkinson disease was established by intranigral injection of MPP⁺. The rats were randomly divided into sham group, MPP⁺ group, triptolide group and vehicle group. The survival of dopaminergic neurons was detected by the immunofluorescence of tyrosine hydroxylase (TH) in the substantia nigra (SN). The activation of microglia was determined by immunofluorescence of OX-42 (microglia marker) in the SN. The expression of chemokine receptor CX3CR1 in SN was measured by Western blotting.RESULTS: Intranigral injection of MPP⁺ increased the fluorescence intensity of the microglial marker, and promoted DA neuron degenerative death. Immunohistological analysis showed that the OX-42 density was decreased (P<0.01) and tyrosine hydroxylase (TH) positive neurons were increased in the triptolide group (P<0.01). The expression of CX3CR1 was lower in triptolide group than that in model group (P<0.05).CONCLUSION: Triptolide may improve PA neurons function in MPP⁺-induced rats through inhibiting CX3CR1 expression and microglial activation.     

Dynamical observation of the expression of TGF-β1 and MAPK1/3 in the renal tubules of rats with diabetes

AIM: To observe the expression of transforming growth factor β₁ (TGF-β₁), MAPK_1/3 and fibronectin (FN) in the development of renal tubulointerstitial disease. METHODS: Wistar male rats were randomly divided into normal control group, diabetic group of 1week, 2 weeks, 4 weeks and 8 weeks. Diabetic model was induced by peritoneal injection of streptozotocin. Immunohistochemistry was employed to detect the expression of TGF-β₁, MAPK_1/3 and FN in the kidney. TGF-β₁ protein in the renal cortex was checked by Western blot. BG, Scr and UP were analysed by biochemical methods, and the morphological changes in renal tubulointerstitium were also examined under microscopy on sections stained with HE and PAS. RESULTS: The expression of MAPK_1/3 and FN was observed, but not the expression of TGF-β₁ in normal renal tissue. Positive staining of TGF-β₁ was observed in the renal tubulo-interstitium in 1-week diabetic group and thereafter it increased in the course of diabetes. A continuous increase in the expression of MAPK_1/3 and FN was also observed in two - week diabetic rats. Chronologically the expression of TGF-β₁,MAPK_1/3 and FN and the ratio of KW/BW were positively correlative with each other in diabetic animals except one -week diabetic rats. There was also a positive correlation between MAPK_1/3 and FN in l -week diabetic rats. CONCLUSION: Our data suggest that TGF-β₁ appears in the renal tubulointerstitium in early period of diabetes and then its signal is mediated by MAPK_1/3 cascades to accelerate production of FN ,and in turn leads to renal hypertrophy and tubulointerstitial fibrosis.     

Effects of local application of simvastatin on osteoporotic fracture healing in rats

AIM: Many studies have documented an anabolic effect of hydroxymethylglutaryl coenzyme A (HMG-CoA) reductase inhibitors, statins, on undisturbed bone. Reports of their effects on fractured skeleton were limited. A study was therefore conducted to check the effects of statins on fracture healing. METHODS: Simvastatin (10 mg·kg^-1·d^-1) was injected subcutaneously to tissue overlying the site of fractured tibiae of ovariectomized rats for a treatment period of 5 d. Vehicle reagent was used as control. Healing quality was evaluated at 1, 2 and 4 weeks after fracture. RESULTS: Compared with vehicle group, callus cross section area in simvastatin treated rats were significantly enlarged by 21.3% (P<0.05) at 1 week and by 21.5% (P<0.05) at 2 weeks. New woven bone was relatively substantive and arranged more tightly and regularly at 2 and 4 weeks, and maximal load was increased by 57.5% (P<0.05) at 2 weeks and by 31.4% (P<0.05) at 4 weeks. Histomorphometrically, simvastatin was associated with a significant (P<0.05) increase in mineralization width (MLW), mineralization volume (MLV) and mineral apposition rate (MAR). CONCLUSION: Current study suggests that local application of simvastatin promotes fracture healing in ovariectomized rats.     

Effect of endoplasmic reticulum stress protein BiP on type 2 diabetic neuropathic pain in rats treated with curcumin

AIM: To investigate the effects of immunoglobulin heavy chain-binding protein (BiP),an endoplasmic reticulum stress protein, on mechanical withdrawal threshold (MWT), thermal withdrawal latency (TWL), spinal dorsal horn and dorsal root ganglion (DRG) in type Ⅱ diabetic neuropathic pain rats treated with curcumin. METHODS: The rats were fed with a high-fat and high-fructose diet for 8 weeks to induce insulin resistance, and then were intraperitoneally injected with streptozotocin (STZ, 35 mg/kg). Eighty-one rats were selected into experimental design as their blood glucose ≥ 16.7 mmol/L 3 d after STZ injection and their MWT and TWL were decreased to 85% of the baseline values 14 d after STZ injection. The rats were divided into 3 groups (n=27 each): DNP group: type 2 diabetic neuropathic pain; DCur group: type 2 diabetic neuropathic pain and intraperitonal injection of curcumin at a dose of 100 mg·kg^-1·d^-1; DSC group: type 2 diabetic neuropathic pain and intraperitonal injection of corn oil at a dose of 4 mL/kg. Another 27 normal SD male rats fed with normal forage were adopted as control group (C group). MWT and TWL were measured at the time points of 3 d, 7 d and 14 d after curcumin injection. The lumbar segment 4~6 of the spinal cord and the corresponding DRG were removed at the same time. The expression of BiP was determined by immunohistochemical staining and Western blotting. RESULTS: Compared with C group, the rats in DNP group developed hyperglycemia and a decrease in MWT and TWL, as well as an increase in the activity of BiP in spinal dorsal horn and DRG (P<0.05). Compared with DNP group, the rats in DCur group at the time point of 7 d significantly attenuated mechanical allodynia and thermal hyperalgesia, and these effects were correlated with the inhibition of BiP hyper-activation at the time point of 14 d after treatment with curcumin (P<0.05). No significant difference of MWT, TWL and the expression of BiP between DNP group and SC group was observed. CONCLUSION: BiP participates in the pathogenesis of type Ⅱ diabetic neuropathic pain. Curcumin attenuates the MWT and TWL in type 2 diabetic neuropathic pain rats. The mechanism may be involved in the inhibition of BiP expression by curcumin.     

Effects of hypoxia and hypercapnia on expression of soluble guanylate cyclase in rat pulmonary artery

AIM:To investigate the expression of soluble guanylate cyclase protein and its mRNA in rat pulmonary artery after exposure to hypoxia and hypercapnia.METHODS:Male Sprague-Dawley rats were randomly split into 4 group, which were hypoxic hypercapnic (HH 1 week, HH 2 weeks, HH 4 weeks) group and control group, to copy pulmonary hypertensive animal model. The expression of sGCα₁ and β₁ subunits protein of medial and small pulmonary artery was performed by immunohistochemistry with a polycolonal antibody. In situ hybridization was performed on the rat lung tissue using sGC oligonuclear probe to assay the expression of sGCα₁subunit mRNA.RESULTS:The sGCα₁ and β₁ subunits protein and sGCα₁ subunit mRNA were faint staining in the pulmonary small and medium artery in HH1 week, HH 2 weeks and HH 4 weeks groups compared with control group (all P＜0.01).CONCLUSION:sGC subunit mRNA and protein expression in pulmonary small and medium artery were decreased after exposure to hypoxia and hypercapnia, which took part in the development of the pulmonary hypertension.     

Effect of erythromycin on transforming growth factor-β1 and γ-glutamylcysteine synthetase in lung of smoking rats

AIM: To investigate the effect of erythromycin on the level of transforming growth factor-β₁ (TGF-β₁) and γ-glutaglutamylcysteine synthetase (γ-GCS) in smoking rats,and to explore the antioxidate therapeutic role of erythromycin in chronic obstructive pulmonary disease.METHODS: Wistar rats were exposed to cigarettes smoking to establish the model.After passive smoking for 4 weeks,erythromycin intragastric intervention was administered continuously for 8 weeks.The expiratory airway resistance and lung compliance were assessed and the expression levels of TGF-β₁ and γ-GCS proteins (and the mRNA) in airway endothelial cells and alveolar macrophages were observed respectively by immunohistochemical,immunocytochemical and (in situ) hybridization.RESULTS: The expiratory airway resistance was increased and the lung compliance was degraded significantly in smoking group and erythromycin group,compared to control group.In erythromycin group,the airway resistance was lower and the lung compliance was higher than that in smoking group (P<0.05).The levels of TGF-β₁ and γ-GCS in smoking group and erythromycin group was obviously increased in airway endothelial cells and alveolar macrophages in comparison with control group (P<0.05).The levels of TGF-β₁ and γ-GCS were inhibited by erythromycin (P<0.05).TGF-β₁ was obviously positive correlated with γ-GCS in smoking group,but this was not found in erythromycin group.CONCLUSION: Erythromycin therapy improves pulmonary function and relieves emphysema change induced by smoking in rats,and decreases the expression of TGF-β₁ and γ-GCS in alveolar macrophages and airway endothelial cells,suggesting that erythromycin may play a role in the antioxidate therapeutic in COPD.