Characteristics and pathomechanisms of hyperuricemia combined with hypertriglyceridemia and hyperglycemia in a coturnix model induced by high-purine diet

AIM: To investigate the pathomechanisms in a coturnix model of high-purine diet and the metabolic characteristics of glucose and lipids. METHODS: Twenty-four French male quails were randomly divided into 2 groups: control group and model group. The animals in control group were fed with normal diet and the quails in model group were fed with high-purine diet. The body weight, serum uric acid (UA), triglyceride (TG), glucose (GLU), the activity of xanthine oxidase (XOD), guanine deaminase (GuDa) and glyceraldehyde phosphate dehydrogenase (GAPDH), and the level of insulin (Ins) were determined. RESULTS: No change of body weight in model group was observed. In model group, the serum levels of UA,TG and GLU were significantly increased from 10 d to 140 d, 60 d to 140 d and 90 d to 140 d, respectively. At 10 d, 60 d and 140 d, the activity of XOD in model group was significantly higher than that in control group. From 30 d to 140 d, the activity of GAPDH was significantly decreased. From 60 d to 140 d, the level of Ins was significantly increased. CONCLUSION: (1) High-purine diet induces multiple metabolic disorders of UA, TG and GLU. (2) The pathologic processes can be divided into three stages: simple hyperuricemia in the first stage, hyperuricemia combined with hypertriglyceridemia in the second stage and hyperuricemia combined with hypertriglyceridemia and hyperglycemia in the third stage. (3) The pathomechanisms may relate to the increased activity of XOD, decreased activity of GAPDH and increased level of Ins.     

Effect of renal transporter Glut9 on hyperuricemia in rats induced by fructose

AIM: To explore the effect of renal transporter glucose transporter 9 (Glu9) on hyperuricemia in the rats induced by fructose.METHODS: SD male rats (n=30) were randomly divided into normal group, model group and benzbromarone group, according to the weight. The rats in normal group was given water, while the rats in model group and benzbromarone group were given 10% fructose solution to establish hyperuricemia model. At the same time, the rats in normal group and model group were given a gavage of distilled water, while the rats in benzbromarone group were given benzbromarone at the dose of 20 mg/kg. The rats were sacrificed on the 40th day. The serum uric acid (SUA) and urinary uric acid (UUA) were detected to calculate the clearance rate of uric acid (CUA) in the kidney. The activity of hepatic xanthine oxidase (XOD) was also measured. The expression of renal Glut9 at mRNA and protein levels was determined by RT-qPCR and immunohistochemical staining. RESULTS: From the 20th day to the 40th day, the SUA in model group was significantly higher than that in normal group, but the UUA and CUA had no difference. On the 20th day, the SUA in benzbromarone group was markedly decreased as compared with model group, but UUA and CUA had no significant difference. On the 40th day, the hepatic XOD activity in model group was significantly elevated, and no difference of XOD between model group and the benzbromarone group was observed. Compared with normal group, the protein expression of Glut9 in the renal tissues of model group were markedly increased, and that in benzbromarone group was significantly lower than that in model group. However, no difference of the Glut9 mRNA expression was observed among groups. CONCLUSION: Fructose drinking induces hyperuricemia in rats, which is probably related to the up-regulation of renal Glut9 expression at protein level, and the increase in the reabsorption of uric acid in the kidneys.     

Mechanisms of hyperglycemia induced by immunosuppressant FK506

AIM：To investigate the effect of immunosuppressant FK506 on serum glucose in rats and to explore its mechanism. METHODS：Sprague-Dawley rats (n=12) were randomly divided into drug group and normal group. The rats in drug group were intraperitoneally injected with FK506 at dose of 1 mg·kg-1·d-1 and the rats in normal group received saline (1 mL·kg-1·d-1, ip) for 14 d. The fasting weight and fasting glucose were regularly measured every 2 d. Visceral fat was isolated from the rats at the end of experiment. The mRNA expression of adiponectin, leptin, visfatin, resistin, retinol-binding protein 4 (RBP4) and peroxisome proliferator-activated receptors γ (PPAR-γ) was determined by real-time fluorescence quantitative PCR. The protein expression of PPAR-γ and adiponectin was measured by Western blotting. RESULTS：Compared with normal group, the concentration of fasting blood glucose in model group was significantly increased from the 10th day (P<0.05). At day 14, the fasting blood glucose of the model group increased from (5.10±062) mmol/L to (7.73 ± 0.73) mmol/L. No significant change of blood glucose in normal group between the 10th day and the 14th day ［from (4.66 ± 0.32) mmol/L to (5.80±0.10) mmol/L］ was observed. Compared with normal group, the mRNA expression of PPAR-γ, adiponectin and leptin in the adipose tissue of model group was significantly decreased (P<001), whereas the expression of visfatin, resistin and RBP4 was significantly increased (P<005). Compared with normal group, the expression of PPAR-γ and adiponectin in model group was decreased (P<001). CONCLUSION：FK506 may decrease the expression of PPAR-γ to change the expression of adipocytokines and induce hyperglycemia in rats.     

Effect of taurine on L-arginine/NO pathway in erythrocytes of rats of hypertension with insulin resistance induced by fructose

AIM and METHODS:The present study observed the change of L-arginine(L-Arg)/Nitric oxide(NO)pathway in ergthrocytes in hypertension with insulin resistance rat induced by fructose and the effect of taurine on L-Arg/NO pathway.RESULTS:Drinking 4%fructose, while inducing blood pressure, glucose and plasma insulin contents increase, obviously decreased the maximal velocity of L-Arg transport about 31%and 37%(P<0.01), more than that of control group in total and Y⁺ carrier, the NO synthase(NOS)activity, nitrite(NO₂^-)content and cyclic guanylate monophosphate(cGMP)level more than that of control group, but obviously enhanced Michaelis Constant(Km)about 35%and 30%(P<0.01)more than that of control group in total and Y⁺ carrier transport.The taurine treatment significantly counteracted the above changes.CONCLUSION:There exists a functional disturbance in L-Arg/NO system in the erythrocyte of hypertension rats with insulin resistance, but taurine can obviously enhanced the maximal velocity of L-Arg transport and NOS activity.Thus, it appears that taurinemay have vital value in the treatment of hypertension with insulin resistance.     

Role of TRIM8 in mouse cardiomyocyte apoptosis induced by high glucose and high free fatty acid

AIM: To investigate the effects of tripartite motif-containing protein 8 (TRIM8) on the apoptosis of mouse cardiomyocytes (MCMs) induced by high glucose and high free fatty acid (HGHF) and the underlying mechanism. METHODS: The MCMs were divided into normal glucose (NG) group (glucose at 5.5 mmol/L), high glucose (HG) group (glucose at 33 mmol/L), high free fatty acid (HF) group (sodium palmitate at 300 μmol/L) and HGHF group (glucose at 33 mmol/L and sodium palmitate at 300 μmol/L). The expression of TRIM8 in the MCMs was knocked down by siRNA, and the MCMs was further divided into control group, scrambled siRNA (Scra-siRNA)/PBS group, TRIM8-siRNA/PBS group, Scra-siRNA/HGHF group and TRIM8-siRNA/HGHF group. To further confirm the specific mechanism of TRIM8 in the MCM injury induced by HGHF, the MCMs were subgrouped into HGHF/DMSO group, HGHF+TRIM8-siRNA+DMSO (HGHF+Ts/DMSO) group, HGHF/ML385 group and HGHF+Ts/ML385 group. Accordingly, apoptosis was analyzed by flow cytometry, and the levels of reactive oxygen species (ROS) were measured by flow cytometry and DHE staining. The expression of TRIM8, nuclear factor E2-related factor 2 (Nrf2), glutamate-cysteine ligase catalytic subunit (GCLC), heme oxygenase-1 (HO-1) and NAD(P)H:quinone oxidoreductase 1 (NQO-1) at mRNA and protein levels was determined by qPCR and Western blot. RESULTS: HGHF increased the expression of TRIM8, and suppressed the expression of Nrf2, GCLC, HO-1 and NQO-1 in the MCMs (P < 0.05). Compared with Scra-siRNA/HGHF group, the intracellular ROS content and apoptotic rate were decreased in TRIM8-siRNA/HGHF group (P < 0.05). Correspondingly, the expression of the antioxidant molecule Nrf2 and its downstream genes GCLC, HO-1 and NQO-1 was increased (P < 0.05). In contrast, the addition of Nrf2 inhibitor ML385 partially reversed the inhibitory effect of TRIM8 expression knock-down on HGHF-induced apoptosis of MCMs. CONCLUSION: TRIM8 exacerbates the HGHF-induced cardiomyocyte apoptosis by modulating Nrf2 antioxidative pathway.     

Potassium antagonizes damage of coronary artery induced by high salt intake

AIM：To investigate the effect of potassium treatment on coronary arterial impairment induced by high salt intake. METHODS：Sprague-Dawley rats (4-week-old, n=10 in each group) received distilled water (NS), water containing 1.5% NaCl (HS), or 1.5% NaCl and 0.5% KCl (HS+HP) for 16 weeks. Systolic blood pressure (SBP) was determined by tail plethysmography every 2 weeks. After 16 weeks of treatment, vascular remodeling, superoxide production, malondialdehyde (MDA) content, and endothelial nitric oxide synthase (eNOS) and gp91 expression in the coronary arteries were detected. RESULTS：After 16 weeks of salt loading, the rats in HS group was divided into salt sensitive subgroup and salt resistance subgroup according to the tail-cuff blood pressure. In this experiment, the salt-sensitive rats were selected as HS group. In HS group, salt loading significantly increased SBP, serum MDA and gp91 expression, decreased serum NO and eNOS expression in the coronary arteries, and induced the coronary artery remodeling compared with NS group. In salt-loaded SD rats, 16-week potassium treatment abrogated the effects induced by salt loading. CONCLUSION：High salt may affect structural and functional changes in coronary arteries by activating oxidative stress. Potassium treatment antagonizes the effect of high salt intake.     

Alteration of pulmonary vascular structure and gaseous molecules in rats with pulmonary hypertension induced by high pulmonary blood flow

AIM: To examine the alteration of pathologic structure and gaseous molecules in rats with pulmonary hypertension induced by high pulmonary blood flow.METHODS: Aortocaval shunting was produced for 11 weeks in rats, and pulmonary hemodynamics was evaluated.Pulmonary vascular micro- and ultra- structure was also examined.Meanwhile,the concentration of plasma nitric oxide (NO) and carbon monoxide (CO) was measured by spectrophotometry.The expression of endothelial nitric oxide synthase (eNOS) and heme oxygenase-1 (HO-1) in pulmonary arteries was detected by immunohistochemistry.RESULTS: After 11- week aortocaval shunting,pulmonary artery mean pressure was significantly increased.Muscularization of small pulmonary vessels and relative medial thickness and area of pulmonary arteries were obviously increased in shunting rats compared with controls.Ultrastructure of intrapulmonary arteries changed obviously in shunting rats.Meanwhile,plasma NO concentration was increased and eNOS expression in pulmonary artery endothelial cells was significantly augmented in rats of shunting group.Plasma carbon monoxide level and HO-1 expression in puomonary artery smooth muscle cells,however,were not altered in shunting rats.CONCLUSIONS: Pulmonary vascular structural remodeling is the important pathologic basis of pulmonary hypertension induced by a left-to-right shunt,and NO other than CO might play an important regulating role in the development of high pulmonary blood flow-induced pulmonary hypertension.     

Intermedin inhibits pulmonary collagen synthesis in rats with pulmonary hypertension induced by high pulmonary blood flow

AIM: To explore the regulatory effect of intermedin (IMD) on pulmonary collagen synthesis and accumulation in rats with pulmonary hypertension induced by high pulmonary blood flow.METHODS: Healthy male SD rats (n=20) were randomly divided into control group (n=7), shunt group (n=7) and shunt with IMD group (n=6). The shunting of abdominal aorta and inferior vena cava was produced in rats of shunt group and shunt with IMD group. After 8 weeks, IMD was administered into the rats of shunt with IMD group subcutaneously by mini-osmotic pump for 2 weeks. Mean pulmonary artery pressure (mPAP), relative medial thickness (RMT) of pulmonary arteries, contents of hydroxyproline, collagen type I and III, bone morphogenetic protein-2 (BMP-2), and the mRNA expression of procollagen I and III in lung tissues were measured and compared. RESULTS: Compared with control group, mPAP and RMT of medium and small pulmonary arteries in the rats of shunt group were significantly increased. Meanwhile, the lung hydroxyproline, collagens I and III and BMP-2 contents, and the mRNA expression of lung procollagen I and III were all significantly increased compared with control group. However, IMD significantly decreased mPAP, alleviated the changes of pulmonary vascular micro-structure, decreased the collagen accumulation and pulmonary tissue homogenate BMP-2 contents, and inhibited the mRNA expression of procollagen I and III in the lung tissue of shunting rats.CONCLUSION: IMD plays a protective role in the development of pulmonary hypertension and pulmonary vascular structural remodeling induced by high blood flow by inhibiting pulmonary collagen synthesis and accumulation, possibly in association with the BMP-2 pathway.     

Effects of Snail1 siRNA on tubular epithelial-to-mesenchymal transition induced by high glucose

AIM: To explore the effect of Snail1 siRNA on high-glucose induced tubular epithelial-to-mesenchymal transition (TEMT). METHODS: Subconfluent renal tubular epithelial cells were incubated in serum-free DMEM for 24 h to arrest and synchronize the cell growth. Then cells were treated with normal glucose (5.5 mmol/L D-glucose) or high glucose (25 mmol/L D-glucose) for 72 h. Meanwhile 19.5 mmol/L D-manntiol was used as high osmotic control. Snail1 siRNA was transfected into tubular epithelial cells. In parallel, cells were transfected with non-specific siRNA which served as the control data sets. Cells were then treated with 25 mmol/L D-glucose for 72 h. RNA and cell lysates were collected to determine the protein and mRNA levels of Snail1, TGF-β1, α-SMA, vimentin and E-cadherin. RESULTS: Transfection caused the decreases in Snail1 at mRNA and protein levels by 62% and 68% respectively as compared to those in untransfected cells cultured in high glucose medium. Western blotting exhibited that Snail1 siRNA transfection restored E-cadherin protein expression by 61% compared to that in high-glucose-treatment cells, whereas it inhibited high-glucose-induced induction of α-SMA protein by 58%. Similarly, RT-PCR revealed that Snail1 siRNA transfection dramatically suppressed the high-glucose-induced mRNA expressions of α-SMA and vimentin by 72% and 61%, respectively, while E-cadherin mRNA increased by 53%. CONCLUSION: Our study provides direct evidence that Snail1 is able to control TEMT.     

Role of EETs downregulation in obesity-related hypertension induced by a high fat diet

AIM: To study the effect of renal epoxyeicosatrienoic acids (EETs)on juvenile rats with obesity related hypertension induced by high fat diet.METHODS: Sprague-Dawley male rats were fed with high fat diet from 3-week old. The changes of weight and sBP between the rats of high fat diet and normal diet were compared. EETs activity was analyzed with RP HPLC and Western blotting in different parts of kidney.RESULTS: Weight and sBP increased in high fat diet group at the eighth and eleventh weeks ［(328±23)g vs (273.0±21.0)g, (153.0±8.6)mmHg vs (134.0±7.7)mmHg, P<0.05］. No significant change of the EETs activity of renal microvessels between two groups was observed. The EETs activity in cortex and papilla decreased in high fat diet group compared with that in normal diet group ［(75.4±9.2)nmol·g-1·min-1 vs (138.1±10.3)nmol·g-1·min-1, (55.8±6.2)nmol·g-1·min-1 vs (121.6±11.3)nmol·g-1·min-1, P<0.05］, and this was confirmed by Western blotting.CONCLUSION: These results demonstrate that juvenile rats with obesity related hypertension induced by high fat diet might be related to the downregulation of EETs activity in cortex and papilla.     

Changes of immune function and mortality in patients with sepsis-induced stress hyperglycemia

AIM: To investigate the role of stress hyperglycemia on condition assessment and predicting prognosis in patients with sepsis. METHODS: The study included 44 patients with sepsis, divided into three groups according to their blood glucose profile within 24 h after admission: patients with stress hyperglycemia (group SH, n=15), diabetes mellitus type 2 (group DM, n=10), and normal glucose levels (group NG, n=19). CD4+/CD8+ ratio, Th1/Th2 ratio and HLA-DR% of the patients were measured within 24 h after admission by flow cytometry as assessment of their immune function. The sepsis-related organ failure assessment (SOFA) scores and acute physiology and chronic health evaluation II (APACHE II) scores of patients were recorded at the same time to analyze whether stress hyperglycemia affects the immune function and the 28 d mortality in patients with sepsis. RESULTS: A higher mortality rate of septic patients with stress hyperglycemia was observed compared to diabetic patients (53.3% vs 10.0%) and group NG (53.3% vs 21.1%). SOFA score and APACHE II score were higher in group SH than those in group DM and group NG while lower levels of human leucocyte antigen DR (HLA-DR) expression and CD4+/CD8+ ratio was found in group SH than those in group DM and group NG. No difference in the levels of Th1/Th2 among the three groups was observed. Non-survivors had higher levels of SOFA score, APACHEⅡ score, Th1/Th2 ratio and HLA-DR%. No difference was detected for CD4+/CD8+ ratio, mean glucose values and age. CONCLUSION: Stress hyperglycemia is associated with decreased immune function and an adverse clinical outcome in patients with sepsis.     

Effects of apelin-13 on oxidative stress induced by high uric acid in adipocytes

AIM: To study the effects of apelin-13 on oxidative stress induced by high uric acid in 3T3-L1 adipocytes and its underlying mechanisms. METHODS: 3T3-L1 adipocytes were stimulated with uric acid at 10 mg/dL for 48 h. Some of the adipocytes were administered with 1 μmol/L apelin-13 in the presence of uric acid at 10 mg/dL. The adipocytes stimulated with 100 μmol/L H₂O₂ were served as positive controls. The intracellular reactive oxygen species (ROS) concentrations were detected by flow cytometry. The biochemical kits were used to measure the activities of superotide dismutase (SOD), glutathione peroxidase (GSH-Px), catalase (CAT) and NADPH oxidase (NOX) activity, and the content of malondialdehyde (MDA) in the cell lysate and the supernatant. The mRNA levels of renin-angiotensin system (RAS) components, including angiotensinogen (AGT), angiotensin-converting enzyrne1 (ACE1), angiotensin II type 1 receptor (AT1R) and AT2R, as well as angiotensin II receptor -like 1 (APJ) were measured by real-time PCR. The concentrations of angiotensin II (AngⅡ) in the cell lysate and the supernatant were measured by ELISA. RESULTS: Adipocytes stimulated with uric acid at 10 mg/dL had lower activities of antioxidant enzymes (SOD, GSH-PX and CAT) and higher levels of NOX activity and MDA content (P < 0.05). Accordingly, the intracellular ROS levels were found to be dramatically increased. However, apelin-13 administration attenuated uric acid-induced oxidative stress in the 3T3-L1 adipocytes. Uric acid at 10 mg/dL upregulated the mRNA expression of local RAS, enhanced AngⅡ concentrations both in the cell lysate and the supernatant, and down-regulated the mRNA level of APJ in the adipocytes (P < 0.05). Conversely, apelin-13 partially reversed these parameters. CONCLUSION: Apelin-13 attenuates oxidative stress induced by uric acid, may be via down-regulation of local RAS expression in the 3T3-L1 adipocytes.     

Changes of aryl hydrocarbon receptor in cardiac hypertrophy induced by high glucose in vitro

AIM: To investigate the changes of aryl hydrocarbon receptor (AhR) in the process of cardiomyocyte hypertrophy induced by high glucose, and to explore its potential mechanisms. METHODS: The rat cardiomyocytes (H9c2 cells) were divided into normal glucose group, high glucose group, DMSO group and resveratrol (an AhR antagonist) group. The content and distribution of AhR were observed with immunofluorescence staining. The myocardial cells were stained with rhodamine-labeled phalloidin to visualize cytoskeleton, and the cell surface area were determined after imaging by fluorescence microscopy. The generation of reactive oxygen species (ROS) in the cardiomyocytes was measured using a fluorescent probe DCFH-DA. The mRNA expression of AhR, CYP1A1, atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) were evaluated by real-time quantitative PCR (RT-qPCR). The protein levels of AhR, CYP1A1, ANP and BNP were assessed by Western blot. RESULTS: AhR was constitutively presented in the cytosol under normal-glucose condition and was translocated to the nuclei under high-glucose condition. High glucose induced cardiac hypertrophy, and increased ROS generation. Significant reductions in the cell size and ROS generation were observed after treated with resveratrol. The expression of AhR, CYP1A1, ANP and BNP at mRNA and protein levels in high glucose group was increased as compared with normal glucose group and resveratrol group, and the above-mentioned indexes significantly decreased in resveratrol group as compared with DMSO group. CONCLUSION: High glucose-induced cardiac hypertrophy increases AhR expression, which may be involved in the maintenance of glucose homeostasis in the cardiomyocytes. AhR translocation to the nucleus induced by high glucose results in the increases in CYP1A1 expression and ROS generation, which may be an important mechanism of high glucose-induced cardiomyocyte hypertrophy.     

Effect of GLP-1 on insulin resistance and PKCε in rats with nonalcoholic fatty liver disease induced by high fat diet

AIM: To observe the therapeutic effect of glucagon-like peptide 1 (GLP-1) analog on nonalcoholic fatty liver disease of rats and to investigate the underlying mechanism.METHODS: SD rats (n=21) were used to establish a nonalcoholic fatty liver disease model by feeding a high fat diet for 12 weeks, and other 11 rats were fed with a normal diet for 16 weeks. The model rats were randomly divided into 2 equal groups:one group was treated with glucagon-like peptide 1 analog (0.6 mg·kg^-1·d^-1) by intraperitoneal injection for 4 weeks, the other group using saline as a control. After treatment, fasting blood glucose, serum insulin, blood lipids, liver function and the pathological changes of the hepatic tissues were evaluated and the expression of PKCε at mRNA and protein levels in the liver tissues was detected by real-time PCR and Western blot, respectively.RESULTS: Compared with model group, the intervention of GLP-1 significantly reduced insulin resistance index (HOMA-IR), improved the liver function (P<0.05), decreased the liver index and blood lipids (P<0.05). HE staining showed obvious pathological changes of the hepatic tissues in model group, and the intervention of GLP-1 significantly reduced lipid droplets in the hepatocytes and improved the structural damage of the liver. The expression of hepatic protein kinase Cε (PKCε) at mRNA and protein levels significantly decreased which were reversed by treating with GLP-1.CONCLUSION: GLP-1 shows good therapeutic effect on nonalcoholic fatty liver disease of rats, possibly by controlling lipid metabolism and reducing insulin resistance, which may be related to PKCε expression.     

Effect of VDUP-1 on apoptosis of renal tubular epithelial cells induced by high glucose

AIM: To investigate the effect of vitamin D3 up-regulated protein 1 (VDUP-1) on apoptosis of renal tubular epithelial cells induced by high glucose and its mechanism. METHODS: Human renal proximal tubular epithelial cell line HK-2 was treated with high glucose. The mRNA and protein levels of VDUP-1 in HK-2 cells were detected by real-time PCR and Western blot. HK-2 cells were transfected with VDUP-1 small interfering RNA (siRNA). Real-time PCR and Western blot were used to detect the inhibitory effect. The HK-2 cells were treated with high glucose, and the change of VDUP-1 expression was detected. The apoptosis was analyzed by flow cytometry. The activities of caspase-3 and caspase-9 in the cells were measured. The tumor necrosis factor-α (TNF-α) content in the culture supernatant was examined by ELISA. The key proteins of Sonic hedgehog (Shh) signaling pathway, Patched 1 (Ptch1), Smoothened (Smo), zinc finger protein Gli2 and Shh, were determined by Western blot. The HK-2 cells were treated with exogenous Shh, and the levels of Ptch1, Smo and Gli2 were detected by Western blot. After the HK-2 cells with VDUP-1 silencing were treated with exogenous Shh and high glucose, the apoptosis was analyzed by flow cytometry, the activities of caspase-3 and caspase-9 in the cells were examined, and the TNF-α content in culture supernatant was measured by ELISA. RESULTS: High levels of VDUP-1 mRNA and protein were observed in the HK-2 cells treated with high glucose. The mRNA and protein levels of VDUP-1 were decreased in the HK-2 cells transfected with VDUP-1 siRNA(P<0.05). Compared with the normally cultured cells, the apoptotic rate of HK-2 cells was increased after high glucose treatment, and the activities of caspase-3 and caspase-9 and the content of TNF-α were also significantly increased (P<0.05). After down-regulation of VDUP-1 expression by siRNA transfection, the apoptotic rate of HK-2 cells decreased after high glucose treatment, and the activities of caspase-3 and caspase-9, and the content of TNF-α were also significantly decreased (P<0.05). The protein levels of Ptch1, Smo, Gli2 and Shh were decreased after high glucose culture, while down-regulation of VDUP-1 partly antagonized the effect of high glucose on the expression of Ptch1, Smo, Gli2 and Shh in the HK-2 cells. Exogenous Shh promoted the expression of Ptch1, Smo and Gli2, and inhibited the apoptosis of the HK-2 cells induced by high glucose. Exogenous Shh and down-regulation of VDUP-1 synergistically inhibited high glucose-induced apoptosis of the HK-2 cells. CONCLUSION: Down-regulation of VDUP-1 expression inhibits high glucose-induced apoptosis and release of TNF-α in renal tubular epithelial cells by activating Shh signaling pathway.     

Comparison of immune-related pain induced by antigen-special complex with inflammatory pain induced by formalin in rats

AIM: To investigate the difference between immune-related pain induced by antigen-special complex and inflammatory pain induced by formalin, and to observe the differential expression of p38 mitogen-activated protein kinase in spinal cord. METHODS: Thirty adult health SD rats were randomly divided into control group, formalin group and immune complex group (10 rats in each group). After the baseline tests were finished, 5 rats in each group underwent intrathecal administration of p38 MAPK inhibitor SB203580. The right hindpaw of the rats were injected with PBS, formalin or rat IgG immune complex. The thickness of hindpaw and pain behaviors were observed at time points of 0 min, 30 min, 1 h, 2 h, 4 h, 8 h and 12 h after injection. The expression levels of total and activated p38 MAPK in spinal cord were determined by Western blotting analysis. RESULTS: The rats in formalin group showed significant nociceptive behaviors immediately, such as licking foot, and limping with highly swollen foot which could touch the ground. The pain threshold was decreased rapidly 30 min after injection and alleviated after then. The pain threshold of the rats in immune complex group obviously decreased 4 h after injection without red swollen hindpaw. The expression of activated p-p38 MAPK in spinal cord in formalin group was significantly higher than that in immune complex group and control group (P<0.01). No statistic difference of p-p38 expression between immune complex group and control group, also no significant effects of SB203580 on pain behaviors in immune complex group were observed. CONCLUSION: Activated p38 MAPK contributes to the pathogenesis of inflammatory pain, but not to the pathogenesis of immune-related pain. The mechanism of immune-related pain is different from inflammatory pain induced by formalin.     

Ubiquitin-dependent degradation of SnoN/Ski protein in glomerular mesangial cells induced by high glucose

AIM: To observe the protein expression of SnoN/Ski and ubiqutin ligase Arkadia in rat glomerular mesangial cells (GMCs) exposed to the high glucose. METHODS: Cultured rat glomerular mesangial HBZY-1 cells were divided into control group, 20 mmol/L glucose group, 30 mmol/L glucose group, 30 mmol/L glucose+MG132 group (culture medium containing 30 mmol/L glucose and 0.5 μmol/L specific proteasome inhibitor MG132), and mannitol group. The expression levels of SnoN, Ski and Arkadia were measured by Western blotting analysis, immunofluorescence and laser scanning confocal microscopy. RESULTS: In control GMCs, the expression of SnoN/Ski was rich and Arkadia was weak. After stimulated with high glucose, the expression of SnoN/Ski was decreased and Arkadia was gradually increased (P<0.05). Compared with high glucose group, the levels of SnoN/Ski and Arkadia were mostly reverted by adding the proteasome inhibitor MG132 at concentration of 0.5 μmol/L (P<0.01). The expression levels of SnoN/Ski and Arkadia were not significantly changed in mannitol group in comparison with control group (P>0.05). CONCLUSION: High glucose decreases the expression of SnoN/Ski through ubiquitin-dependent degradation of SnoN/Ski. The degradation of SnoN/Ski mediated by Arkadia may play an important role in the pathogenesis of diabetic nephropathy.     

Effects of berberine on the apoptosis of NIT-1 cells induced by high glucose and saturated fatty acids

AIM: To investigate the effects of berberine on the apoptosis of NIT-1 cells induced by high glucose and saturated fatty acid. METHODS: The influence of berberine at different concentrations on NIT-1 cells cultured with or without high glucose and saturated fatty acid were determined and compared using MTT colorimeric assay. The cell apoptotic rate was also determined by flow cytometry assay and in situ TUNEL method. RESULTS: The effects of berberine at different concentrations on NIT-1 cells showed dose-dependent, low dose (≤5 μmol/L) had dispensable cytotoxicity; meanwhile, high dose showed distinct effects. On the other hand, low dose of berberine alleviated the apoptosis in NIT-1 cells induced by high glucose and saturated fatty acid, when adding berberine to cell medium. CONCLUSION: Berberine inhibited the apoptosis of NIT-1 cells induced by high glucose and saturated fatty acid.