Changes of serum levels of nitric oxide and nitric oxide synthase in patients during liver transplantation

AIM: To study the changes of serum levels of nitric oxide (NO) and nitric oxide synthase (NOS) in patients during liver transplantation. METHODS: Samples were obtained from 30 patients in end liver disease at five time points during liver transplantation. NO level and NOS activity were measured by radioimmunoassay and colorimetry, respectively. Arterial and mixed venous blood samples used for blood gas analysis were taken at the same time. Intrapulmonary shunt (Qs/Qt) was calculated according to the standard formula. The hemodynamics parameters including continuous cardic output (CO), HR, MABP, CVP, SVR were measured during liver transplantation. RESULTS: (1) NO2-/NO3- level at 10 min before anhepatic period was significantly higher than the baseline level. Compared with NO2-/NO3- level at 10 min before anhepatic period, NO2-/NO3- level at 30 min after anhepatic period was significantly decreased. NO2-/NO3- level at 30 min after neohepatic period was significantly higher than the baseline level and at 30 min after anhepatic period. (2) No significant change of tNOS activity was observed. Compared with the baseline activity of inducible nitric oxide synthase (iNOS), the activity at 10 min before anhepatic period and at 30 min after neohepatic period was significantly increased. The activity at 30 min after neohepatic period was significantly higher than that at 30 min after anhepatic period. (3) MABP decreased significantly when opening the inferior vena cava. CO and CVP decreased in the anhepatic stage and increased in the reperfusion stage. SVR increased during anhepatic stage and decreased significantly during neohepatic period. (4) Qs/Qt decreased significantly during anhepatic stage and increased significantly at 30 min after neohepatic period. CONCLUSIONS: Serum level of NO and NOS activity are significantly changed during liver transplantation. High level of NO may result in low systemic vascular resistance and increasing in intrapulmonary shunt.     

Influence of MTHFR genetic polymorphism on the indexes of vascular endothelial functions

AIM: To investigate the influence of methylenetetrahydrofolate reductase (MTHFR) 677 C→T mutation on angiotensin Ⅱ (Ang II), prostacyclin (PGI₂) and nitric oxide (NO). METHODS: By cluster sampling, 1146 adult Han people were selected from the residential communities. MTHFR 677 genotypes were identified by polymerase chain reaction-restriction fragment length polymorphism for each sample. Plasma levels of homocysteine were determined by fluorescence ration biochemical assay. Serum NO levels were determined by cadmium reduction method. Plasma AngⅡ and PGI₂ concentrations were determined by radioimmunoassay. SPSS 13.0 was used for data analysis. RESULTS: Total samples were divided into three groups according to the genotypes. No significant difference in PGI₂ and AngⅡamong the three groups was observed. The difference of serum NO level between the C/C and T/C genotypes was not significant (P>0.05). The serum concentration of NO of T/T genotype was significantly lower than that of T/C and C/C genotypes (P<0.01). CONCLUSION: The influence of MTHFR 677 C→T mutation on Ang II and PGI₂ is not significant in the people from the residential communities. The decrease in serum NO level might be one of the underlying mechanisms of MTHFR 677 C→T mutation causing myocardial infarction and ischemic stroke.     

Effects of different doses of L-dopa on rotational behavior and amounts of cells expressing D2 receptors in hemiparkinsonian rats

AIM：To observe the effects of different doses of L-dopa on the rotational behavior and amounts of cells expressing D₂ receptors in striatum in hemiparkinsonian rats.METHODS：A hemiparkinsonian model was established in rats by pretreatment with 6-hydroxydopamine.The D₂ receptor expression were detected by immunohistochemical staining.The numbers of rotations induced by apomorphine was counted within 30 min before and after L-dopa (10 mg·kg^-1·d^-1,50 mg·kg^-1·d-^-1 or 100 mg·kg^-1·d^-1,ip) was introduced to Parkinson’s disease (PD) model rats for 15 days.RESULTS：In successful PD model rats,the increased percentage of D₂ receptor in lesioned side compared with intact side was associated linearly with the numbers of rotations within 30 min (r=0.927,P<0.01).After high dose of L-dopa intervention to PD model,the numbers of rotations decreased significantly (P<0.05),the amounts of cells expressing D₂ receptor at the lesioned side striatum decreased significantly (P<0.01).CONCLUSION：After high dose of L-dopa intervention,rotation behavior of PD rats improves,and D₂ receptor is down-regulated significantly.     

Role of ERK1/2-STAT3 pathway in adaptive cytoprotection induced by H2O2 preconditioning

AIM:To explore the role of extracellular signal-regulated kinases ERK1/2-STAT3 pathway in adaptive cytoprotection induced by H₂O₂ preconditioning in PC12 cells. METHODS:In PC12 cells, the experimental model of cytoprotection by H₂O₂ preconditioning against oxidative stress-induced injury was set up. The morphological changes in the apoptotic cells were observed by using of chromatin dye Hoechst 33258. The percent of apoptotic cells was determined by flow cytometry (FCM) with propidium iodide staining. The levels of p-ERK1/2 and p-STAT3 expression were detected by Western blotting assay. RESULTS:Preconditioning with H₂O₂ at concentration of 100 μmol/L for 90 min obviously inhibited apoptosis induced by 300 μmol/L H₂O₂, and both ERK1/2 and STAT3 were activated. UO126 (10 μmol/L, an inhibitor of ERK1/2) or AG-490 (10μmol/L, an inhibitor of JAK2) significantly blocked the cytoprotection effect of H₂O₂ preconditioning. Moreover, UO126 (10 μmol/L) also markedly inhibited the up-regulation of p-STAT3 expression by H₂O₂ preconditioning. CONCLUSION:H₂O₂ preconditioning activates ERK1/2-STAT3 signal pathway, which may be one of the mechanisms underlying H₂O₂ preconditioning-induced cytoprotection.     

Effects of luteolin on endothelial cell injury induced by H2O2

AIM: To study the effect of luteolin on endothelial cell injuries induced by H₂O₂. METHODS: Endothelial cells were cultured in vitro and divided into seven groups: control; solvent; H₂O₂; quercetin+H₂O₂; luteolin-L+H₂O₂; luteolin-M+H₂O₂; luteolin-H+H₂O₂ group, respectively. Endothelial cells were incubated with 750 μmol/L H₂O₂ for 18 h or 14 h in the absence or presence of various concentrations of luteolin and quercetin. The expression of caspase-3, the pro-apoptotic factors, was detected by immunohistochemical technique, and the apoptotic index was detected by flow cytometry. Meanwhile, the amount of malondialdehyde (MDA), nitric oxide (NO), lacate dehydrogenase (LDH) in serum, and cell viability were measured. RESULTS: Incubation of endothelial cells with H₂O₂ for 18 h elicited the increase in the extent of immunostaining for caspase-3, the rate of apoptosis, the release of LDH, and the number of dead cells accompanied by the decrease in the content of NO in the medium, which were inhibited by pretreatment of luteolin at concentrations of 0.5-50 μmol/L in a concentration-dependent manner (P<0.01). CONCLUSION: Luteolin possesses a protective effect against endothelial cell injuries induced by H₂O₂, which may be related with anti-oxidation, increasing the concentration of NO and inhibiting the activation of caspase-3.     

Effect of diazoxide on change of H2O2 in rat pulmonary artery smooth muscle cells and proliferation of hypoxic rat pulmonary artery smooth muscle cells

AIM: To investigate the contribution of diazoxide,an opener of mitochondrial ATP-sensitive K+ channel (MitoK_ATP),and mitochondrial membrane potential (ΔΨm) to change of H₂O₂ in rat pulmonary artery smooth muscle cells (PASMCs) and to unbalance between cell proliferation and apoptosis of PASMCs induced by hypoxia.METHODS: The rat PASMCs were isolated from fresh normal lung tissues and cultured,which were divided into 6 groups,as follows: ① control group;② diazoxide group;③ 5-HD group;④chronic hypoxia group;⑤ chronic hypoxia+diazoxide group;⑥ chronic hypoxia +5-HD group.The relative change in mitochondrial potential was detected with rhodamine fluorescence (R-123) technique.The level of H₂O₂ in rat PASMCs was detected with chemiluminescence method.The proliferation of rat PASMCs was examined by cell cycle analysis and MTT colorimetric assay.RESULTS: After exposed to diazoxide for 24 h,the intensity of R-123 fluorescence,the level of H₂O₂ and the A value in normoxic rat PASMCs were significantly increased,and the apoptosis of rat PASMCs was significantly decreased as compared with control group (P<0.05).However,there were no significant changes in these tests after the rat PASMCs had been exposed to 5-HD for 24 h.Chronic hypoxia or chronic hypoxia+diazoxide markedly increased the intensity of R-123 fluorescence,the level of H₂O₂ and the A value in rat PASMCs,and also markedly decreased the apoptosis of rat PASMCs as compared with control group (P<0.05),and these changes were more significant in chronic hypoxia +diazoxide group than those in chronic hypoxia group (P<0.05).5-HD partly weakened the effect of hypoxia on the intensity of R-123 fluorescence,the level of H₂O₂,the A value and the apoptosis of rat PASMCs (P<0.05).Significant and positive correlations were found between the intracellular H₂O₂ and the R-123 fluorescence or the A value.Significant and negative correlation was found between the intracellular H₂O₂ and the apoptosis of rat PASMCs.CONCLUSION: The results suggest that the opening of MitoK_ATP followed by a depolarization of ΔΨm can contribute to the increase in the level of H₂O₂ in rat PASMCs and to the proliferation of rat PASMCs induced by hypoxia.This might be a mechanism of the development of hypoxic pulmonary hypertension.     

Effects of olaquindox on PLA2 activity and phospholipid metabolism of biomembrane in chicks

AIM: To study the effect of olaquindox on phospholipase A₂ (PLA₂) activity and phospholipid metabolism in biomembrane.METHODS: 150 healthy avin chicks were divided into normal control group and olquindox group.The two groups were fed with same feedstuff without olaquindox.The animals in olquindox group were fed with olquindox at dose of 15mg/kg body weight everyday.The experimental period lasted six weeks.Every week 6 vein blood samples were collected randomly from every group for preparing erythrocyte membrane (ECM) and hepatic mitochondria membrane (MiM).RESULTS: PLA₂ activities of ECM and MiM in olquindox group were significant higher than those in control group from 3 to 6 weeks of experiment (P<0.05，P<0.01).The level of major membrane phospholipids in olquindox group was lower than that in control group (P<0.05,P<0.01).CONCLUSION: Olaquindox affects PLA₂ activity and major phospholipid concentration in ECM and MiM,indicating that olaquindox may induce biomembrane injury.     

VEGF induces HUVECs to produce extracellular H2O2 and its proliferation role

AIM:To study the effect of VEGF on extracellular H₂O₂ production in HUVECs and the role of H₂O₂ in the VEGF-induced proliferation. METHODS:HUVECs was stimulated with 500 μg/L VEGF. Products of extracellular H₂O₂ was detected by H₂DCFDA staining. MTT method was used to value the influences of 3×10⁶ U/L catalase and 5-20 mmol/L H₂O₂ to VEGF function. RESULTS:After treatment for 15 min with VEGF, HUVECs appeared fluorescence, and continued to become stronger, peaked at 45 min then decreased. HUVECs, which was treated simultaneity with VEGF and 3×10⁶ U/L catalase, only appeared very faint fluorescence. The proliferation of HUVECs by VEGF was restrained when treated with 3×10⁶ U/L catalase. The extrinsic H₂O₂ at concentration of 5-10 mmol/L promoted the proliferation of HUVECs but inhibited the proliferation effect of VEGF on HUVECs (P<0.01). CONCLUSION:These findings indicate that VEGF induces HUVECs to produce extracellular H₂O₂ and plays role in proliferation, but extrinsic H₂O₂ restrains VEGF function.     

Advances in experimental research in HCV cell model and anti-HCV effect of As2O3

Hepatitis C virus (HCV) is a human pathogen responsible for liver diseases including acute and chronic hepatitis and hepatocellular carcinoma. However, the high prevalence, the absence of antiviral drugs and vaccines for prevention and treatment are the difficult medical problems. Lacking appropriate culture method and small animal model have severely limited investigation of the HCV infection mechanism and the development of the therapeutic strategy. Recently, the in vitro culture system develops rapidly, and provides a powerful tool for HCV related research. Despite the well known toxicity of the chemical, arsenic trioxide (As₂O₃) is effective for treating the patients with refractory or relapsed acute promyelocytic leukemia and many other cancers. Interestingly, As₂O₃ shows the ability of inhibiting HCV RNA replication and infection. This review describes the different types of in vitro HCV models developed, and many studies of the potent effect of As₂O₃ against HCV and its associated molecular mechanisms.     

Calcineurin signal transduction pathway involves in cardiomyocyte hypertrophy induced by prostaglandin F2α

AIM: To study the role of prostaglandin F₂α (PGF₂α) in cardiac hypertrophy and its relation with calcineurin (CaN) signal transduction pathway in vitro. METHODS: The cultured neonatal rat cardiomyocyte was used to observe the hypertrophic effect of PGF2α, and the hypertrophic response was assayed by measuring the cell diameter, protein content and atrial natriuretic factor (ANF) mRNA expression. For mechanism studies, the intracellular free calcium concentration (［Ca²⁺］_i) in cultured cardiomyocytes was measured by using Fura-2/AM as a fluorescent indicator. ANF and CaN mRNA expressions, and the expressions of CaN and its downstream effectors, NFAT₃ and GATA₄ proteins were assayed by RT-PCR and Western blotting, respectively.RESULTS: In cultured cardiomyocytes, PGF₂α induced profound hypertrophic morphology change, the significant increase in cell diameter and protein content in a concentration-dependent manner compared with those in vehicle control (P<0.01). The same result was found in measuring the ［Ca²⁺］_i in cardiomyocytes (P<0.01). PGF₂α at concentration of 10-7 mol/L significantly promoted ANF and CaN mRNA expressions and the protein expressions of CaN/NFAT₃/GATA₄ compared with those in the vehicle control (P<0.05). Cyclosporin A, a CaN inhibitor, markedly inhibited the myocyte hypertrophy (P<0.01), reduced the increased ［Ca²⁺］_i(P<0.01) and decreased the expressions of CaN mRNA and CaN/NFAT₃/GATA₄ proteins (P<0.05) compared with those of only PGF₂α　10^-7 mol/L treatment. CONCLUSION: Cardiomyocyte hypertrophy induced by PGF2α may be, at least in part, mediated by CaN signal transduction pathway activated by increasing ［Ca²⁺］_i.     

Effect of NPPB and niflumic acid on expression of GCL and CLIC4 in H2O2 injured C6 glioma cells

AIM: To observe the effect of 5-nitro-2- (3-phenylpropylamino) benzoate acid (NPPB), niflumic acid (NFA) (chloride channel blockers) on malignant glioma C6 cells injured by hydrogen peroxide (H₂O₂). METHODS: The viability of C6 cells treated with NPPB, NFA and H₂O₂ was measured by MTT assay. LDH release rate and GSH contents were detected by ultraviolet spectrophotometry. mRNA levels of GCLC, GCLM and CLIC4 were determined by RT-PCR. CLIC4 protein level was detected by Western blotting. RESULTS: Compared to the control group, H₂O₂ treatment induced the decrease in cell viability and GSH contents, the increase in LDH release rate, the decrease in the expression of GCLC, GCLM and CLIC4 mRNA and the increase in CLIC4 protein level (P<0.05 ), respectively. Compared with the H₂O₂ group, H₂O₂ combined with NPPB or NFA treatment did not change the cell viability, the GSH contents and the GCLC, GCLM mRNA expression. However, the LDH release rate and CLIC4 protein level decreased (P<0.05). CONCLUSION: The chloride channel blockers NPPB or NFA lessen the oxidative injury of C6 cells through modulating the function of membrane and down-regulating the protein expression of CLIC4.     

Effect of H2O2 on voltage-gated potassium channels in rat pulmonary artery smooth muscle cells

AIM:To determine the effect of hydrogen peroxide (H₂O₂) on voltage-gated potassium channel currents (IKv) in pulmonary vascular smooth muscle cells (PASMCs). METHODS:Using whole cell patch-clamp technique, IKv was recorded in freshly isolated rat PASMCs with acute enzymatic digestion method. The effect of hydrogen peroxide on IKv in PASMCs was investigated in normoxia. RESULTS:IKv in PASMCs was increased significantly by H₂O₂ and the increase depended on the concentration in normoxia. Current-voltage relationship curve shifted to the left. CONCLUSION:Hydrogen peroxide is an important K+ channel opener.     

Mechanisms of PGF2α on the glucose-induced insulin secretion in NIT-1β cells

AIM:To investigate the cellular mechanisms by which PGF₂α promotes glucose-stimulated insulin secretion in NIT-1 beta cells. METHODS:Using the radioimmunoassay (RIA), the amount of the PGF₂α augmentation of glucose stimulated insulin secession was determined in different conditions, and the confocal laser scanning methods by Fluo-3AM as a fluorescent probe were used to analyze the changes of intracellular calcium in NIT-1β cells. RESULTS:At the lower glucose (0, 5.5 mmol/L), PGF₂α (5 μmol/L) failed to potentiate insulin secretion (P>0.05). However, in the presence of 16.5 mmol/L glucose, PGF₂α increased significantly in insulin secretion (P<0.05). Neither the AC inhibitor ddA nor the GC inhibitor Ly-83583 altered PGF₂α-potentiated insulin secretion in the presence of 16.5 mmol/L (P<0.05 or P<0.01). Otherwise, the PLC inhibitor U-73122 and the PKC blocker calphostin C both counteracted the insulinotropic of PGF₂α (P<0.01 or P<0.05). Moreover, exposure of the NIT-1β cells to 5 μmol/L PGF₂α induced a rapid increase of intracellular calcium (P<0.01). The inhibitor, ddA or Ly-83583 had no impact on PGF₂α-induced elevation of the intracellular calcium (P<0.01). Pretreatment of the cells with U-73122 completely prevented the calcium response induced by PGF₂α (P<0.01). CONCLUSION:Efects of PGF₂α was independent of cAMP or cGMP, potentiated glucose (16.5 mmol/L)-induced insulin secretion in NIT-1β cells through stimulation of phospholipase C, which subsequently mediated the elevation of intracellular calcium and activation of protein kinase C.     

Vasodilatatory effect and mechanism of CH2Cl2 extract of flos magnoliae on isolated rat thoracic aorta

AIM: To characterize the vasodilatatory effect of CH₂Cl₂ extract of flos magnoliae (CEF) on isolated rat thoracic aorta and to elucidate its possible mechanism. METHODS: The thoracic aorta was isolated from male Sprague-Dawley (SD) rats and the isometric tension of aortic rings with or without endothelium was measured. RESULTS: CEF (0.1-1 000 mg/L) produced concentration-dependent, endothelium-independent relaxations in phenylephrine (PE)-contracted aortic rings. The maximum relaxation induced by CEF was 78.68%±6.03% in endothelium intact rings and 64.98%±13.90% in endothelium removed rings while the forskolin (1 μmol/L)-induced vasodilation was obtained as 100%. The vasodilatatory effect of CEF was not statistically inhibited by 10 μmol/L glibenclamide (Glib), 3 mmol/L tetraethylammonium (TEA), 100 μmol/L BaCl₂ and 10 μmol/L 1H- -oxadiazole- -quinoxalin-1-one (ODQ) in the preparations without endothelium. The CEF pre-treatment significantly inhibited vasoconstrictions to angiotensin Ⅱ (AngⅡ), prostaglandin F_2α, (PGF_2α), dopamine (Dopa), vasopressin (Vaso), 5-hydroxytryptamine (5-HT) and PE by 91.31%, 82.11%, 95.32%, 90.53%, 72.22% and 83.63%, respectively (P<0.01). In Ca²⁺-free medium treated endothelium removed aortic ring, incubation with CEF at concentration of 82 mg/L significantly attenuated intracellular Ca²⁺ release by PE. In Ca²⁺-free + high potassium medium incubated aortic rings without endothelium, CEF (82 mg/L) markedly inhibited potassium-stimulated Ca²⁺-dependent contraction which was mainly due to Ca²⁺ influx (P<0.01).CONCLUSION: CEF induced vasorelaxation is mainly related to interfering intracellular calcium homeostasis by blocking Ca²⁺ influx and intracellular Ca²⁺ release.     

Mcl-1 involves in G2/M arrest of HL-60 cells induced by diallyl disulfide

AIM: To investigate the role of Mcl-1 in the G₂/M arrest induced by diallyl disulfide (DADS) in leukemic HL-60 cells.METHODS: The inhibitory effect of DADS on human leukemic HL-60 cells was detected by CCK-8 method in vitro. Flow cytometry analysis was employed to observe the cycle arrest in HL-60 cells and the effect of DADS-induced G₂/M arrest on HL-60 cells with Mcl-1 gene knockdown by RNAi silencing. The expression of Mcl-1, PCNA and CDK1 in HL-60 cells treated with DADS was determined by Western blotting. The binding of Mcl-1 with PCNA and CDK1 was detected by coimmuno-precipitation. RESULTS: HL-60 cells were treated with DADS at concentration of 15, 30, 60, 120 or 240 μmol/L for 48 h. The inhibition rates of HL-60 cell proliferation were 31.15%, 55.88%, 66.14%, 75.29% and 80.35%, respectively, and gradually enhanced with the increase in the concentration of DADS (P<0.05). Flow cytometry analysis revealed that the proliferation of HL-60 cells was blocked by DADS in the G₂/M phase. Treatment with DADS for 24 h and 48 h at concentrations of 60 μmol/L and 120 μmol/L, the percentage of G₂/M phase cells increased as compared to the untreated cells (P<0.05). DADS induced arrest of HL-60 cells in G₂/M phase in a time- and dose- dependent manner (P<0.05). The results of Western blot analysis indicated that Mcl-1, PCNA and CDK1 in HL-60 cells were significantly reduced after treated with DADS (P<0.05). HL-60 cell cycle progression delayed by silencing Mcl-1 gene with siRNA technique, suggesting that silence of Mcl-1 gene led to G₂/M arrest. Compared to the cells treated with DADS only, the percentage of G₂/M cells raised in the cells with Mcl-1 gene silencing and treated with DADS (P<0.05), indicating that Mcl-1 gene silencing enhanced the effect of DADS-induced G₂/M arrest in HL-60 cells. The binding of Mcl-1 with PCNA and CDK1 was detected by coimmuno-precipitation and the formation of heterodimers was observed, which was decreased after treated with DADS for 4 h.CONCLUSION: DADS inhibits the proliferation of HL-60 cells and induces its G₂/M phase arrest. The decreased expression of PCNA is related to inhibiting the proliferation of leukemic cells. Knockdown of Mcl-1 gene enhances the effect of DADS-induced G₂/M arrest.     

Effect of vitamin K3 on apoptosis induced by androgen-independent prostate cancer cell PC-3M

AIM: To study the effect of vitamin K₃ (VK₃) on the induction of apoptosis in androgen-independent prostate cancer cell PC-3M in vitro.METHODS: Cell viability was estimated by MTT assay. AO/EB staining was performed to detect apoptotic cells. Apoptosis and the changes of cell cycle were detected by flow cytometry. NAC was used to observe the effect of growth inhibition by VK₃. RT-PCR was used to confirm the changes in gene expression. Levels of intracellular peroxides were estimated by using an oxidation-sensitive fluorescent probe DCFH-DA. RESULTS: PC-3M cells growth was significantly inhibited by VK₃ (≥60 μmol/L, P<0.05). The inhibitory effect was time and dosage dependent. The result of AO/EB staining showed that apoptosis of PC-3M cells were induced by VK₃. A typical subdiploid peak before G₀/G₁ phase was observed after treated for 12 h with VK₃ (60 μmol/L) by flow cytometry. The effect of growth inhibition treated with VK₃ was antagonized by antioxygen NAC (5, 10, 20, 40, 80 μmol/L). An increase in the level of DCF fluorescence after PC-3M cells were treated for 1-2 h with VK₃ was observed. Antioxidase GSH-Px and CAT were run-down after treated with VK₃. CONCLUSION: The results indicate that apoptosis in PC-3M cells is induced through oxidative stress by VK₃.     

Expression of TGF-β1 and Smad2/3 in kidney of STZ-induced diabetic nephropathy rats

AIM: To study the role of TGF-β/Smad pathway in the development of renal fibrosis in diabetic nephropathy.METHODS: Rats were induced to diabetic nephropathy by using tail intravenous injection of STZ.The expression of TGF-β₁, Smad2/3 protein and mRNA in kidney were examined at 2, 4, 8 and 16 weeks after STZ induction.CTGF, collagen-Ⅲ, PAI-1 mRNA expression in kidney at 16 weeks of STZ-induced diabetic nephropathy and normal rats were studied by RT-PCR.RESULTS: Weak TGF-β₁, Smad2/3 protein were detected in normal renal tissues while strong TGF-β₁, Smad2/3 staining were observed in renal tissues of diabetic nephropathy (0.057±0.030/0.223±0.040;0.017±0.010/0.153±0.010, respectively, P<0.05).The TGF-β₁, Smad2/3 protein expression were constantly high with the development of diabetic nephropathy and fibrosis (0.153±0.010, 0.122±0.050, 0.141±0.070 and 0.216±0.030 for 2, 4, 8 and 16 weeks, respectively).The TGF-β₁, Smad2 mRNA expression also increased with the development of diabetic nephropathy (2.86, 3.25 fold compared to control, respectively).The expression of TGF-β₁, Smad2, CTGF, collagen-Ⅲ and PAI-1 mRNA were significantly higher in kidney of 16 week diabetic nephropathy rats than that in normal ones (3.92, 2.95, 1.57, 1.95 and 1.97 folds compare to control, respectively, P<0.05).CONCLUSION: The results indicate that TGF-β₁/ Smad2 pathway activity might play an important role in pathophysiological process of diabetic nephropathy.It may be involved in diabetic renal fibrosis through up-regulation of CTGF and PAI-1 to promote extracellular matrix deposition.     

Alteration of cardiac M3 receptor and its relationship with arrhythmias in drug-induced and ischemia-induced arrhythmic models

AIM: To investigate the alteration of cardiac M₃ receptor and its relationship with arrhythmias in various arrhythmic models. METHODS: Forty Wistar rats were randomly divided into 4 groups: control, aconitine, BaCl₂ and ischemia. In the later three groups, arrhythmias were induced by treatment with aconitine, BaCl₂ and coronary artery occlusion, respectively. The arrhythmias were recorded for 1 h. Western blotting was then used to detect M₃ receptor contents. RESULTS: Arrhythmias were all induced in each group. In aconitine-induced arrhythmias, duration of arrhythmias and arrhythmia score were significantly increased than those in other two model groups. Western blotting showed that the expression of M₃ receptor upregulated 2.3, 1.4 and 1.3 folds respectively, more abundant in various arrhythmic groups than that in the normal control. Moreover, M3 receptor expression in aconitine group increased significantly than that in BaCl₂ and ischemia group. The arrhythmias and M₃ receptor protein expressions in myocytes were positively correlated. CONCLUSION: Arrhythmias upregulate the expression of cardiac M₃ receptor. The upregulating levels of M₃ receptor proteins diverge strikingly in different arrhythmic models. It is probably that the diversity of increase in M₃ receptor is positive related to severity of ventricular arrhythmias.     

Intervention of Lipo-PGE1 on liver blood perfusion by different time and medication: a empirical study

AIM: To explore the effect and mechanism of liposome prostaglandin E₁(Lipo-PGE₁) on liver blood perfusion by different time and medication.METHODS: Twelve healthy adult dogs were injected with Lipo-PGE₁₁ μg/kg via left small saphenous vein at speed of 0.05 μg·kg^-1·min^-1.Liver computed tomography perfusion imaging (CTPI) was performed on 0,5,15 and 30 min,and the value of hepatic arterial perfusion (HAP),portal vein perfusion (PVP) and total liver perfusion (TLP) among groups were compared.The impacts of Lipo-PGE₁ on liver haemodynamics at different time were investigated.Twenty-four health dogs were randomly divided into four groups: control group,peripheral vein group,hepatic artery group and superior mesenteric artery group.Liver CTPI was performed at 5 min after 1 μg/kg Lipo-PGE₁ administration in those groups.The values of HAP,PVP and TLP were compared and effects of Lipo-PGE₁ on liver blood flow by different medication were observed.RESULTS: The values of liver perfusion (mL·min^-1·mL^-1) at 0,5,15 and 30 min after 1 μg/kg Lipo-PGE₁ administration via vein were as follows: HAP: 0.22 ±0.65,0.24±0.65,0.22±0.69,0.22±0.06;PVP: 1.22±0.40,1.88±0.59,1.55±0.55,1.29 ±0.57;TLP: 1.44±0.42,2.12±0.61,1.77±0.56,1.51±0.58,respectively.No significant difference in HAP among groups was observed,but in PVP and TLP,significant differences (F=3.812,P<0.05;F=3.805,P<0.05) among groups were found.The values of PVP and TLP were most obviously increased at 5 min,and the values of PVP and TLP were still on the high level at 15 min and 30 min.The values of liver perfusion (mL·min^-1·mL^-1) by different medication were as fellows: HAP: 0.22±0.06,0.24±0.06,0.31±0.07,0.26±0.05;PVP: 1.28±0.38,2.33±0.41,2.37±0.55,2.83±0.94;TLP: 1.50±0.40,2.57±0.42,2.67± 0.58,3.09±0.94,respectively.No significant difference in HAP among groups (F=2.248,P>0.05) was found,but in PVP and TLP group,significant differences (F=6.892,P<0.01;F=7.802,P<0.01) among groups were observed.In addition,superior mesenteric artery group showed higher value of PVP and TLP than other methods.CONCLUSION: Lipo-PGE₁ obviously increases liver blood perfusion,especially for portal vein perfusion.Interventional technology provides an effective pathway to improve hepatic perfusion.