Aim To investigate the effects of Tongxin-luo capsule on oxidative stress in diabetic peripheral neuropathy (DPN )mice and its mechanisms.Meth-ods KK/Upj-Ay mice were divided into model, Tongxinluo low-dose group, Tongxinluo middle-dose group and Tongxinluo high-dose group.C57BL/6 mice were selected as control group.Mice were given drugs intragastrically for 12 weeks.Paw withdrawal latency, motor nerve conduction velocity (MNCV)were detec-ted.Activity of superoxide dismutase (SOD),gluta-thione peroxidase (GSH-Px)and content of malondial-dehyde (MDA)in blood were detected by colorimetric method.The expression of heme oxygenase-1 (HO-1 ),γ-glutamyl cysteine synthetase (γ-GCS ) of sciatic nerve was examined by real time PCR and Western blot.The protein expression of p38 MAPK,p-p38 MAPK,JNK,p-JNK,ERK,p-ERK was examined by Western blot.Results Compared with model group, paw withdrawal latency was increased and MNCV was faster in Tongxinluo group (P <0.05 ,P <0.0 1 ). SOD and GSH-Px contents significantly increased, MDA content decreased (P <0.01 ).HO-1,γ-GCS mRNA and protein expression significantly increased (P<0.05,P <0.01 )in Tongxinluo group.p-p38 MAPK protein expression decreased in Tongxinluo group (P<0.05 ).Conclusion Tongxinluo can in-hibit oxidative stress in DPN of mice via suppressing the phosphorylation of p38 MAPK.

OBJECTIVE:To investigate the effects of Tongxinluo capsules on inflammatory cytokines of mice with diabetic pe-ripheral neuropathy. METHODS:40 KK/Upj-Ay mice were randomly divided into model group(pure water),and Tongxinluo low, medium and high dose groups [1,2 and 4 g (medicinial materials)/kg]. 10 C57BL/6 mice were selected into the control group (pure water). The drugs were given once a day for consecutive 12 weeks,ig. The pain perception threshold,motor nerve conduc-tion velocity(MNCV)and sensory nerve conduction velocity(SNCV)were detected for the mice 1 h after the last administration of drugs. The flow cytometry was used to detect the levels of tumor necrosis factor α(TNF-α),interleukin-1β(IL-1β),cyclooxy-genase 2(COX-2)and monocyte chemoattractant protein 1(MCP-1)in serum. Real-time fluorescent quantitative polymerase chain reaction(RT-PCR)and Western blot were respectively employed to detect the mRNA and protein expressions of TNF-α,IL- 1β, COX-2 and MCP-1 in the mice’s sciatic nerves. RESULTS:Compared with control group,pain perception threshold in model group was decreased;MNCV and SNCV were slowed;the levels of TNF-α,IL-1β,COX-2 and MCP-1 in serum were increased;the expressions of mRNA and protein of TNF-α,IL-1β,COX-2,MCP-1 were increased,with significant differences(P<0.01 or P<0.05). Compared with model group,pain perception threshold in Tongxinluo medium and high dose groups were increased;MNCV and SNCV were increased;the levels of TNF-α,IL-1β,COX-2 and MCP-1 in serum were decreased;the expressions of mRNA and protein of TNF-α,IL-1β,COX-2,MCP-1 in sciatic nerves were decreased;the expressions of mRNA of COX-2, MCP-1 and protein of IL-1β,COX-2,MCP-1 in sciatic nerves in Tongxinluo low dose group were decreased,there were statistical significant difference(P<0.01 or P<0.05). CONCLUSIONS:Tongxinluo capsules have certain protective effects on the mice with peripheral nerve in case of diabetes by a mechanism that may be associated with the inhibition of inflammatory cytokines.

Objective To investigate the effect of down-regulated Blimp1 gene expression on differenetiation of bone marrow cells into dendritic cells (DCs).Methods Blimp1-shRNA was constructed and then loaded into lentivirus vector as lenti-blimp1-shRNA.Bone marrow cells from Balb/c mice were induced differentiation to DCs in an 8-day cell culture system with GM-CSF/IL-4 incubation and LPS stimulation at day 7.The cells were divided into groups as empty control (no treatment),lenti-control (transfected by lentivirus empty vector at day 1),and lenti-Blimp (transfected by lenti-blimp1-shRNA at day 1).The transfection efficiency was evaluated by GFP fluorescence for one week.The morphology and growth curve were analyzed.Real-time PT-PCR and Western blotting were used to evaluate mRNA and protein expression of Blimp1.At day 8,CD11 c and CD86/MHC-Ⅱ were quatitified using flow cytometry.Results GFP fluorescence emerged 3 days after transfection and was continuously expressed.Classic DC morphology was shown in no treatment cells,while damaged morphology presented in the cells with lentivirus transfection.The empty control cells proliferated from day 3,peaked as (2.45 ± 0.26) 106/well at day 4,and kept at (2.27 ± 0.19) 106/ well at day 8,The cells receiving lentivirus presented (1.69 ± 0.39) 106/well.The expression of Blimp1 mRNA and protein in the lenti-Blimp1 group was 76％/1％ and 1.0％/74.0％ of the empty control group.At day 8,CD11c,CD86 and MHC-Ⅱ expression in the empty control group was (69.2 ±5.0)％,(51.1± 4.9) ％ and (56.3 ± 7.3) ％,while (68.6±5.9)％,(49.5±4.3)％ and (69.4±4.5)％ in the lenti-control group,and (72.8 ± 5.5)％,(50.2 ± 6.0)％ and (46.5 ± 5.7)％ in the lenti Blimp1 group.Conclusion Lentivirus-mediated Blimp1-shRNA gene therapy modulates blimp1 expression of DC precursors.Down-regulation of Blimp1 fails to interrupt the differentiation of DCs but inhibits the maturation.

Objective To investigate the effect of oxymatrine on liver triglyceride metabolism enzymes of apolipoprotein E(ApoE)－/－mice with fat-induced insulin resistance. Methods A total of 72 ApoE－/－mice fed with a high-fat diet for 16 weeks were randomly assigned into a model group ,an oxymatrine 25 mg/kg group ,an oxymatrine 50 mg/kg group ,and an oxymatrine 100 mg/kg group , and oxymatrine was administered p.o.for 8 weeks.C57BL/6J mice were selected to serve as the control group.Serum biochemical parameters were assessed ;Insulin resistance was assessed with the Hyper insulinemic-euglycemic clamp test ;Pathological changes in the liver were visualized by hematoxylin ( HE ) staining ;levels of gene expression of triglyceride metabolism enzymes were examined by real-time polymerase chain reaction ( PCR ) and Western-blotting. Results Administration of oxymatrine reduced body weight ,fasting blood glucose , cholesterol ,and triglyceride to varying degrees and alleviated pathological changes in the liver.Glucose infusion rates were(18.5 ± 1.6)mU · kg －1· min－1,(20.1 ± 1.8)mU · kg －1· min－1,and(21.3 ± 2.3)mU · kg －1· min－1in the oxymatrine 25 ,50 ,and 100 mg/kg groups ,respectively ,and were significantly higher than that in the model group (16.5 ± 1.6)mU · kg －1· min－1(all P ＜ 0.05) .mRNA expression levels of hormone-sensitive lipase(HSL) ,adipose triglyceride lipase(ATGL) ,and peroxisome proliferator-activated receptor γ(PPARγ)were higher in the three oxymatrine groups than in the model group ,while levels of diacylglycerol acyltransferase (DGAT1 ) were lower in the oxymatrine groups(F＝53.81 ,21.06 ,23.67 ,35.37 ;all P＜0.05) ;Levels of HSL ,ATGL ,and PPARγ were higher ,while levels of DGAT1 were lower in the oxymatrine 50 and 100 mg·kg －1groups than in the model group ( F ＝ 53.62 ,22.87 ,28.13 ,33.54 ;all P ＜ 0.05 ). Conclusions Oxidative can mitigate insulin resistance in mice fed with a high fat diet through regulating the expression of liver triglyceride metabolism enzymes.

Objective To investigate the protective effect and mechanism of oleanolic acid(OA)on kidneys in rats with type 2 diabetes mellitus(T2DM).Methods A total of 35 Sprague-Dawley(SD)rats were enrolled in this study.25 SD rats were randomly selected to establish T2DM model,after modeling,20 rats remained and divided into T2DM group(n=6),low-dose oleanolic acid group(LOA,n=6)and high-dose oleanolic acid group(HOA,n=8).And ten rats were selected as normal control group(NC,n=10).The biochemical indicators,24 h urine volume and 24 h urinary microalbumin(UAlb)were compared among the four groups.Renal lipid deposition was evaluated by Oil red O staining.The protein expressions of Adenosine 5'-monophosphate-activated protein kinase(AMPK),p-AMPK and peroxisome proliferator-activated receptor γ coactivator-1 α(PGC-1 α)were detected by Western blot.Results Compared with the NC group,the levels of 24 h urine volume,fasting blood glucose(FPG),serum total cholesterol(TC),triglyceride(TG),low density lipoprotein cholesterol(LDL-C),serum creatinine(Scr),serum uric acid(SUA)and 24 hUAlb were increased(P<0.05),while the body weight,high density lipoprotein cholesterol(HDL-C),p-AMPK and PGC-1α were decrease in the T2DM group(P<0.05).Compared with the T2DM group,the expressions of HDL-C,p-AMPK and PGC-1α were increased(P<0.05),while the levels of 24 h urine volume,FPG,TG,TC and LDL-C,Scr,SUA and 24 hUAlb were decreased in the LOA and HOA groups(P<0.05).Compared with LOA group,the expressions of HDL-C,p-AMPK and PGC-1α were increased(P<0.05),while the levels of 24 h urine volume,FPG,TG,TC,LDL-C,Scr,SUA and 24 h UAlb were decreased in the HOA group(P<0.05).Compared with the NC group,a large number of red-stained lipid droplets were deposited in the renal tubular epithelial cells in the T2DM group.Compared with the T2DM group,the lipid droplet deposition was reduced in the LOA and HOA groups,and the improvement was more significant in the HOA group.Conclusion OA can alleviate renal injury in T2DM rats,which may be linked to activation of AMPK/PGC-1α pathway.

Objective To investigate the effects of activated protein kinase(AMPK)agonists on the expression of peroxisome proliferator-activated receptor γ coactivator 1α(PGC1a),mitofusin 2 (Mfn2) and nuclear respiratory factor1 (NRF1)in the process of lipid-induced mitochondrial dysfunction of skeletal muscles.Methods The expression of PGC1α,Mfn2 and NRF1 in C2C12 skeletal muscle cells after intervention with palmitic acid was detected using reverse transcription-polymerase chain reaction(RT-PCR)and Western blotting.The effect of 5-aminoimidazole-4-carboxamide ribonucleotide (AICAR) on Mfn2 and NRF1 and,the expression of Mfn2 and NRF1 in C2C12 cells induced by a PGC1α activator and PGC1α-siRNA were assessed by Western blotting.Results Palmitic acid decreased the mRNA and protein expression of PGC1α,Mfn2 and NRF1 in C2C12 cells (P＜0.05).Additionally,AICAR,rosiglitazone and metformin up-regulated PGC1α expression,regardless of the presence of palmitic acid and,AICAR reversed lipid-induced PGC1α,Mfn2 and NRF1 attenuation in C2C12 cells.Furthermore,Mfn2 and NRF1 protein expression increased with PGC1α over-expression,and decreased with down-regulated PGC1α expression.Conclusions AICAR can relieve the adverse effects of palmitic acid on PGC1α,Mfn2 and NRF1 in skeletal muscle cells.Moreover,it appears that AICAR can up-regulate Mfn2 and NRF1 expression through activating PGC1α.