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Objective To explore the effects of oscillating blood glucose on apoptosis in renal tubular epithelial cells of diabetic rats and the role of oxidative stress.Methods Renal tubular epithelial cells (HK-2) were cultured in vitro with stable high glucose or oscillating high glucose,and MTF assay was applied to the neasurement of cell proliferation.Streptozotocin induced diabetic model was established with SD rats and the oscillating high blood glucose animal model was induced by intraperitoneal injection of insulin and glucose at different time points every.day.12 weeks later 24 h urine protein (24hUP),blood urea nitrogen (BUN),serum creatinine (Scr),superoxide dismutase (SOD), and malondialdehyde (MDA)were determined. Apoptosis was assessed by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL),and immunohisto-chemistry was used to detect apoptosis associated gene Bax,Bcl-2,and NOX-4 expression in kidney.Changes in ultrastructure were observed.Results Oscillating high glucose may inhibit renal cells proliferation obviously when comparing with stable high glucose.In the rats with oscillating blood glucose rather than those with stable high blood glucose,there was a significant increase of BUN,Scr,24hUP,and MDA and a decrease of SOD[ (21.50 ± 1.72 vs 12.50 ± 1.85 )mmol/L,(97.51 ± 7.84 vs 82.12 ± 11.48 ) μmol/L,( 1.57 ± 0.09 vs 1.04 ± 0.12 ) mg/24 h,( 23.50 ± 1.87 vs 14.82 ± 2.96) nmol/ml,( 17.22 ± 1.12 vs 21.11 ± 1.80) U/ml,all P＜0.05 ] ; cell apoptosis was intensified with the up-regulation of Bax and NOX-4 protein expression and down-regulation of Bcl-2 in glomerular endothelial cells; and more severe pathological damages were observed.ConclusionComparing with stable high blood glucose,oscillating high blood glucose induces more apoptosis and less proliferation of renal tubular epithelial cells and the mechanism may be related to the increased oxidative stress.
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Objective To investigate the correlations of extracellular matrix and hepatic ultramicrostructural changes with clinical manifestations in patients with mild chronic hepatitis B (CHB).Methods Patients with chronic HBV infections were enrolled and were divided into mild CHB group (n=66) and HBV carrier group (n=10).Serum samples were collected from patients, and serum HBV markers, HBV DNA load and liver fibrosis indexes were measured.All subjects received liver biopsy, and the tissue samples were observed by light microscope and electron microscope.T test and χ2 test were performed for measurement data and enumeration data, respectively.Spearman test was used for ranked data.Results The differences on ALT and AST levels between mild CHB group and HBV carrier group were significant (t=12.42, 7.06, P＜0.05), but there was no significant difference on HBV DNA load between two groups (t=0.24, P ＞ 0.05).Serum liver fibrosis indexes (hyaluronic acid, type Ⅲ collagen,type Ⅳ collagen and laminin protein) in mild CHB group were not significantly higher than those in HBV carrier group (t=0.45, 0.95, 0.76 and 1.21, P ＞0.05).In mild CHB group, there were 33 patients with ≥G2 and ≥S2, but in HBV carrier group were only 2 patients (χ2=4.17, P ＜ 0.05).Seventeen patients in mild CHB group were with S3-4, while that was not observed in HBV carrier group (χ2=4.75, P ＜0.05).In mild CHB group, hepatic ultramicrostrutural changes on fat storing cell, collagen protein and portal area were correlated with fibrosis grades, and the correlation coefficients were 0.351, 0.675 and 0.301, respectively (P=0.004, 0.000 and 0.014).Conclusion Electron microscope is of higher sensitivity than light microscope in observing hepatic ultramicrostructural changes, which is effective in evaluating the severity of mild CHB.
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AIM: To investigate the effect of puerarin on diabetic myocardial damage and to explore its possible mechanisms in rats. METHODS: The male Sprague-Dawley rats were randomly divided into normal control group, diabetic group, high dose puerarin (160 mg/kg), middle dose puerarin (120 mg/kg), low dose puerarin (80 mg/kg) treatment groups and aminoguanidine (100 mg/kg) treatment group. Corresponding drugs were intraperitoneally injected once a day. The animals in normal control group and diabetic group were given equal propylene glycol. 12 weeks later, the rats were sacrificed and their cardiac muscles were collected. Myocardial structure was observed under transmission electron microscope (TEM). The blood glucose concentration and the activities of superoxide dismutase (SOD), Ca~(2+)-ATPase, Na~+-K~+-ATPase and the content of malondialdehyde (MDA) in myocardial homogenate were measured biochemically. In addition, the mRNA expressions of peroxisome proliferator activated receptor γ (PPAR-γ), glucose transporter-4 (GLUT-4) and aldose reductase (AR) in myocardial tissues were determined by reverse transcription polymerase chain reaction (RT-PCR). RESULTS: It is observed that the myofibrils were diminished, broken or fused, some lipid droplets deposited in the cytoplasm of cardiocytes and part of cristae of mitochondria were broken or disappeared under TEM in diabetic group. The activities of SOD, Ca~(2+)-ATPase, Na~+-K~+-ATPase as well as the mRNA expressions of PPAR-γ and GLUT-4 decreased significantly (all P<0.01). The blood glucose concentration and the MDA level and the AR mRNA expression increased obviously (all P<0.01) in diabetic group as compared to those in normal control group. How-ever, in puerarin treatment groups, the above changes were reversed, a significant differences of those were found as compared to those in diabetic group (P<0.05 or P<0.01, respectively). The pathological change of cardiac muscles was relieved. It showed that myofibrils were well-arranged and only few lipid droplets deposited in the cytoplasm of cardiocytes and most mitochondria had clear and regular cristae under TEM in puerarin treatment groups. CONCLUSION: Puerarin exerts preventive and remedial effects on the diabetic myocardium, which may be related to up-regulating the mRNA expressions of PPAR-γ and GLUT-4, promoting glucose uptake and relieving oxidative stress damage.
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AIM: To study the protective effects of the extract of Ginkgo biloba (EGB) on brain tissues in experimental diabetic rats and explore its possible mechanisms. METHODS: Thirty male Sprague-Dauley rats were divided into three groups: normal control group, diabetic group and EGB-treated group. Strepozotocin were injected intraperitoneally on the later two groups to induce diabetes, EGB-treated group was injected intraperitoneally with EGB, and the others were treated with normal saline at the same volume. After five weeks, the content of endothelin (ET), malondial dehyde (MDA), NO2~-/NO 3~-, and the activity of superoxide dismutase (SOD), nitric oxide synthase (NOS) were determined in brain homogenate, and the level of blood glucose, insulin and ET were measured respectively. In addition, the morphologic changes of the brain tissue were studied by light microscopy and electron microscopy, respectively. RESULTS: In diabetic group, there were degeneration of neuron, brain edema, softened focus and demyelination in the white matter of brain by light microcopy. There were expansion of mitochondria of neuron and gliocyte, the shortened of crista, demyelination of neurofiber and injury of blood-brain-barrier by the electron microscopy. After treated with EGB, the pathological changes decreased in brain under light microcopy and electron microcopy. Compared with diabetic rats, the activity of SOD and the level of serum insulin increased, while the level of blood ET, the activity of NOS, the content of ET, MDA, NO2~-/NO3~- decreased in EGB-treated rats (P
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Objective To identify the histopathological characteristics of multiple organ dysfunction syndrome (MODS) with V. vulnificus in mice. Methods Sixteen healthy KM mice (6～8 week old ) divided randomly into two groups, study group ( n =12) and control group ( n =4) The animal model of MODS was established by received either an intraperitioneal, intramuscular subcuneous inoculum of 4.34?10 6 cfu/0.2 ml of V.vulnificus or intraperitioneal injection of a sterile physiological salt solution (control group). Pathological changes of the man organs were individually obsenved in under election microscope (EM). Results Mortality rates exced 100%, 12/12) in study group after inoculums of mice within 4～8 h, while in 0% (0/0) in the control . The detection rate of V. vulnificus were in 100%(12/12) from blood, hearts, lungs, livers, intramuscularly, subcutaneous in the study group, while in 0% (0/4) after sterile saline intraperitioneal injection. The man histopathological changes were :degeneration and necrosis of the parenchyma cells in the different organs;interstitial swelling, mitochondriondrial injure of multiple organs.These changes were especially obvious in the lungs and myocardeum. Conclusions Above pathological changes suggested that results of MODS caused by V. vulnificus septicemia,the multiple organs failure as an important feature of the fatality of V. vulnificus infections,and my be helpful for researchers investigating of V.vulnificus.
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AIM:To investigate the protective effect of puerarin on testis in diabetic rats.METHODS:The male Sprague-Dawley rats were randomly divided into normal control group,diabetic group,high dose puerarin(160 mg/kg),middle dose puerarin(120 mg/kg),low dose puerarin(80 mg/kg)treatment groups and aminoguanidine(100 mg/kg)treatment group.Corresponding drugs were intraperitoneal injected once a day.The animals in normal control group and diabetic group were given equal propylene glycol.12 weeks later,the rats were sacrificed and their testes were collected.Testicular structure was observed under transmission electronic microscope(TEM).The contents of nitric oxide(NO),malondialdehyde(MDA)and the activity of nitric oxide synthase(NOS),inducible nitric oxide synthase(iNOS),superoxide dismutase(SOD),Ca2+-ATPase and Na+-K+-ATPase in testis homogenate were detected biochemically.The levels of endothelin-1(ET-1)and calcitonin gene-related peptide(CGRP)were also measured by radioimmunoassay.In addition,the mRNA expressions of eNOS,iNOS,CGRP,ET-1 and AR in testis tissues were determined by reverse transcription polymerase chain reaction(RT-PCR).RESULTS:The pathologic changes such as cytoplasmic inclusion of sertoli cell and obvious vacuoles between sertoli cells and basal membrane of the contorted seminiferous tubules were observed under TEM in diabetic group.The contents of NO and CGRP,the activities of SOD,Ca2+-ATPase,Na+-K+-ATPase as well as the mRNA expressions of eNOS and CGRP decreased significantly(P
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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 11?-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
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AIM: To investigate the effect of puerarin (Pur) on expression of Fas/FasL mRNA in lung tissue during pulmonary ischemia and reperfusion injury(PIRI) in rabbits.METHODS: Single lung ischemia and reperfusion animal model was used.The rabbits were randomly divided into three groups,sham operated group(sham,n=10),PIR group(I-R,n=30) and PIR+ Pur group(Pur,n=30).Changes of several parameters included apoptotic index(AI),wet to dry ratio of lung tissue weight(W/D) and index of quantitative assessment of histologic lung injury(IQA) were measured at 60,180 and 300 minutes after reperfusion in lung tissue.Meanwhile,the location and expression of Fas/FasL mRNA were observed.Lung tissue was prepared for light microscopic and electron microscopic observation at 60,180,300 minutes after reperfusion.RESULTS: As compared with group I-R,Fas/FasL mRNA slightly expressed in intima and extima of small pulmonary artery,alveoli,and bronchiole epithelia in group Pur.The values of AI,W/D and IQA showed significantly lower than that in group I-R at 60,180,300 minutes after reperfusion in lung tissue(P
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AIM: To clarify the role of eNOS and ET-1 in development of pulmonary hypertension (PH) associated with congenital heart diseases. METHODS: 40 patients were randomly divided into three groups: severe or moderate PH group (group A, 12 cases), slight PH group (group B, 14 cases) and normal group (group C, 14 cases). ET-1 and eNOS were examined by using the technique of immunohistochemistry. RESULTS: ① Plasma ET-1 concentration was significantly higher in group A and B than that in group C (P
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AIM: To study the effect of L-arginine (L-Arg) on function and structure of mitochondria in ischemia-reperfusion (MRI) myocardial cells. METHODS: Thirty rabbits were randomly divided into three groups (n=10 in each), control group, MIR group and MIR+L-Arg group. The mitochondrial respiratory function, Ca~(2+) concentration ([Ca~(2+)]m), malondialdehyde (MDA) content and superoxide dismutase (SOD) activity were determined. Meanwhile, the contents of adenosine triphosphate (ATP), adenosine diphosphate (ADP), adenosine monophosphate (AMP), total adenylic acid number (TAN) and energy charge (EC) in the myocardial tissue were respectively measured. Moreover, the ultrastructure changes in myocardial mitochondria were observed during MIR. RESULTS: The mitochondrial respiratory control rate (RCR), velocity 3 (V_3), SOD, surface density (Sv) and specific surface (?) in MIR+L-Arg group were higher than those in MIR group, velocity 4 (V_4), [Ca~(2+)]m, MDA, volume density (Vv), horizental diameter (Hd) were lower than those in MIR group. ATP, ADP, TAN and EC levels of myocardial tissue were higher than those in MIR group. There was no significant difference between MIR+L-Arg and control group in V_3, V_4, SOD, MDA, Vv, Sv, ?, Nv, Vd, AMP and TNA. CONCLUSION: It is suggested that L-Arg improves the function and structure of mitochondria in myocardial cells in the reperfusion injury after myocardial ischemia by decreasing oxygen free radical level and Ca~(2+) overload in the mitochondria. [
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AIM: To investigate the protective effect of Ginkgo biloba extract (GBE) on diabetic testis and explore its possible mechanism. METHODS: Testicular structure of streptozotocin-induced diabetic rats was observed under light microscopy (LM) and transmission electron microscopy (TEM). Content of malondialdehyde (MDA), NO_2~-/NO_3~- and activity of superoxide dismutase (SOD), nitric oxide synthase (NOS) were determined in testicular homogenate. RESULTS: In diabetic rats, it was manifestated as deformation of seminiferous tubule, atrophy and shedding of germinal epithelium under LM, while expansion of smooth endoplasmic reticulum, formation of fatty vacuoles and decrease of lysosome obviously in the cytoplasm of sertoli cell under TEM, the injury of testicular tissue was improved by GBE. Compared with diabetic rats, activity of SOD increased while activity of tNOS and iNOS, content of MDA and NO_2~-/NO_3~- decreased in GBE-treated rats. CONCLUSION: GBE could effectively prevent the development of diabetic testis and the effect may be partly achieved by resisting lipid peroxidation,restraining the activity of testicular tissue iNOS and reducing the pathological alterations of NO. [
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AIM: To investigate the variations of heme oxygenase-1/carbon monoxide system in lung ischemia-reperfusion injury and effects of puerarin on the system.METHODS: The unilateral lung ischemia-reperfusion model was replicated in vivo.Rabbits were randomly divided into three groups(n=10 in each): control group,ischemia-reperfusion(I-R) group and puerarin group.The blood specimens collected at times before ischemia,ischemia 1 h,reperfusion 1 h,3 h and 5 h were tested for the contents of carboxyhemoglobin(COHb) and cyclic guanosine monophosphate(cGMP).The lung tissues sampled at 5 h after reperfusion were assayed for wet/dry weight ratio(W/T),the injured alveoli rate(IAR) and the activity of HO-1.The changes of ultrastructure were observed under electron microscope.The tissue slides were also stained by immunohistochemistry(IHC) and in situ hybridization(ISH) to detect the expression of HO-1.RESULTS: The plasma content of COHb and cGMP in I-R and puerarin group increased in a time-dependent manner after I-R compared with control group,but the increment in puerarin group was higher than that in I-R group(P
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AIM: To investigate the effect of heme oxygenase-1 (HO-1)/carbon monoxide (CO) system on pulmonary ischemia-reperfusion injury (PIRI) in rabbits. METHODS: Single lung ischemia and reperfusion animal model was used in vivo. The rabbits were randomly divided into three groups (n=10 in each), control group (C), PIR group (I-R), PIR+ hemin group (H) and PIR+zinc protoporphyrin IX (ZnPP) group (Z). Changes of several parameters which included plasma carboxyhemoglobin (COHb) at different time points, wet to dry ratio of lung tissue weight (W/D), the injured alveoli rate (IAR) and the HO-1 enzymatic activity were measured at 180 min after reperfusion in lung tissue. The tissue slides were also stained by immunohistochemistry (IHC) and in situ hybridization (ISH) for HO-1 to detect the expression of HO-1 in lung and to analyze the optical density. The lung tissue was prepared for electron microscopic observation at 180 min after reperfusion. RESULTS: The plasma content of COHb in I-R, H, and Z group increased in a time-dependent manner after I-R. But the increment of H group was higher than that of I-R group, while that of Z group was lower. The HO-1 activity in lung tissue was highest in H group, followed by IR group, Z group, and C group (P
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AIM:To observe the effect of the extract of Ginkgo biloba(EGB)on pituitary-testicular axis and the mRNA expressions of luteinizing hormone receptor(LHR)and steroidogenic acute regulatory protein(StAR).METHODS:Thirty male Sprague-Dauley rats were divided into three groups randomly:normal control group,type II diabetic group and EGB treatment group.After fed with high-fat diet for 4 weeks,the later two groups were injected with strepozotocin intraperitoneally to induce type II diabetes mellitus.The EGB treatment group was given EGB at the dose of 50 mg/kg once a day for 12 weeks by intragastric administration.The normal control and diabetic group were given normal saline of equal volume per day for 12 weeks.The indices of blood glucose,insulin and low-density lipoprotein-cholesterol(LDL-c)were measured.The morphologic change of testicular tissue was observed under light microscopy(LM)and transmission electron microscopy(TEM)respectively.The concentrations of blood luteinizing hormone(LH)and testosterone(T)were assayed by the technique of enzyme linked immunosorbent assay(ELISA).The mRNA expressions of LHR and StAR from Leydig cells were detected by RT-PCR.RESULTS:The concentrations of blood glucose,insulin and LDL-c increased obviously,and the testis weights lessened obviously in type II diabetic groups compared to those in normal control groups.Rare spermatogenic cells of seminiferous tubule and germinal arrest were observed in diabetic group under LM.Ultrastructural analysis of testicular tissue by TEM showed dilation of the endoplasmic reticulum and mitochondrial swelling in Leydig cell and sertoli cell in diabetic group.The level of blood LH and T decreased in typeⅡ diabetic groups in comparison with that in the normal control group.Compared to normal groups,the mRNA expression of StAR in type Ⅱ diabetic groups decreased,while the mRNA expression of LHR increased.After the treatment of EGB,the pathological change of testis was relieved,the concentrations of blood glucose,insulin and LDL-c were decreased,the level of blood LH and T,and the mRNA expression of StAR were increased,and the mRNA expression of LHR descended compared to type Ⅱ diabetic groups.CONCLUSION:EGB may increase the LH-induced testosterone production by correcting metabolic disorder of glucose and lipid,improving the function of pituitary-testicular axis and regulating the expression of LHR and StAR mRNA.