Objective To study the gene mutation and clinical characteristic of hereditary spinocerebellar ataxia type 7 (SCA7).Methods The SCA7 (CAG) trinucleotide repeat mutations were detected by polymerase chain reaction(PCR) and polyacrylamide gel electrophoresis technique in 24 patients with autosomal dominant SCA from 15 families, 20 sporadic SCA patients and 41 normal persons from the same family and 30 healthy persons from different family,the abnormal allele fragments were sequenced by ABI 373 DNA sequencing machine.Results 24 patients with SCA had CAG repeat numbers of SCA 7 allele from 9~18.Normal alleles of SCA 7 had CAG repeat number from 9 to 19. One sporadic SCA patient had one abnormal SCA 7 allele with the CAG repeat expanded to 63 repeats, being confirmed by DNA sequencing.Conclusion CAG expansions were pathogenic cause of SCA 7. The technique of gene mutation detection could provide an effective way for the prediction of asymptomatic and genetic counseling,which was a basis for gene typing.

Objective To investigate the protective effect of c-jun N-terminal kinase (JNK)inhibitor SP600125 against acute liver failure in mice.Methods Fifty-five male C57/BL6 mice were divided into control group (n =30) and SP600125 group (n =25).The animals were given an intraperitoneal injection of D-galactosamine (D-GalN,400 mg/kg body weight)/lipopolysaccharide (LPS,30 μg/kg body weight).The control group and SP600125 group were given 10％ dimethyl sulfoxide (15 mL/kg body weight) or SP600125 (75 mg/kg body weight) subcutaneously 12 h and 1 h before D-GalN/LPS administration,respectively.D GalN/LPS induced mouse JNK activation was detected by immunohistochemistry for phospho JNK (p-JNK).D-GalN/LPS induced mouse liver cell apoptosis was detected by immunohistochemistry for Caspase-3 and TdT-mediated-dUTP nick endlabeling (TUNEL).Serum alanine transaminase (ALT) level was tested to assess liver injury.Survival rate of mice within 24 h after D-GalN/LPS administration was observed.The comparison between groups was done by t test and survival rate was analyzed by Kaplan-Meier method.Results JNK activity in liver tissues,as indicated by observation of p-JNK positive cells by immunohistochemistry,was diminished 4 h after D-GalN/LPS administration in SP600125 group.Reduced Caspase-3 activity was observed 6 h after D-GalN/LPS administration in SP600125 group (as indicated in immunohistochemistry by Caspase-3 positive cells).Mice in SP600125 group showed significantly lower TUNEL-positive cell count than control group (43.0±24.5 vs 194.7±73.8; t=9.743,P=0.000).Serum ALT level 6 h after D-GalN/LPS administration was (24.0±54.7) U/L in SP600125 group,which was significantly lower than that in control group [(1234.4±478.4) U/L; t=4.734,P=0.0015].SP600125 also significantly improved the survival rate within 24 h after D-GalN/LPS administration (4/5 vs 1/10； x2=5.225,P=0.0223).Conclusions JNK inhibitor SP600125 exerts protective effects against D-GalN/LPS induced acute liver failure in mice by suppressing JNK activation and hepatocyte apoptosis.

Objective: To investigate chronic kidney disease (CKD) and its risk factors in people receiving physical examination. Methods: hTis retrospective study included people over 20 years old who had physical examination in the Health Management Center of Third Xiangya Hospital from Janurary 2008 to June 2011. CKD and its risk factors as well as questionnaire were recorded. hTe risk factors were analyzed by multivariate logistic analysis. CKD was deifned by kidney damage (microalbuminuria≥30 mg/L) and/or hematuria and/or reduced kidney function [evaluate glomerular ifltration rate (eGFR)<60mL/(min.1.73 m2)]. We counted eGFR according to the modiifcation of diet in renal disease (MDRD). Results: A total of 5 708 physical examination reports were included. The detection rate of albuminuria, reduced renal function and hematuria was 25.0%, 1.7% and 1.1%. hTe detection rate of CKD was 25.6%, and detection rate of CKD stage 1-5 was 17.8%, 6.7%, 1.1%, 0 and 0, respectively. Multivariate logistic analysis indicated that diabetes mellitus, hypertension, hypercholesterolemia, male, age, and smoking were the risk factors for CKD. Increasing physical activity was the protective factor against CKD. Conclusion: High prevalence of CKD in people receiving physical examination is found in Changsha, especially stage 1 and 2 CKD. Physical examination is important to screen CKD. Stopping smoking, control of blood glucose, blood pressure, blood lipids and increasing physical activity may help reduce the prevalence of CKD.

Objective: To investigate the effects of dexmedetomidine (DEX) on oxidative stress and systemic inflammatory response syndrome(SIRS) during cardiopulmonary bypass (CPB).Methods: Totally 62 patients scheduled for mitral valve replacement surgery were enrolled in this study and randomly assigned to DEX group with constant infusion of DEX (0.5 μg·kg-1 ·h-1 , n =31) and the control group with the same amount of 0.9% sodium chloride injection(n =31).A volume of 4 ml of jugular vein blood was respectively withdrawn before the induction of anesthesia (T0), at 10 min after aortic-off clamping(T1), the end of bypass(T2), 4h after bypass(T3),12h after bypass(T4) and 24h after bypass(T5).The serum C-reactive protein (CRP), tumor necrosis factor-α (TNF-α), interleukin-6(IL-6), interleukin-8(IL-8) and interleukin-10(IL-10) were determined.Results: The serum CRP in both groups increased remarkably at T1-T5 , while the serum CRP in DEX group was significantly lower than those in the control group at T3-T5.The serum TNF-α increased remarkably in control group at T2-T4, while the serum TNF-α increased remarkably in DEX group at T2-T3;the serum TNF-α in DEX group was significantly lower than those in the control group at T3-T4.The serum IL-6, IL-8, IL-10 in both groups were increased remarkably at T2-T5, T1-T5,T1-T3, respectively.The serum IL-6 and IL-8 in the DEX group were significantly lower than those in the control group at T2-T4 while the serum IL-10 in the DEX group was significantly higher than that in the control group at T1-T3.The serum WBC in both groups at T3-T5 were higher than those at T0, while the serum WBC in the control group was significantly higher than that in the DEX group at T3-T5.However, the differences between the two groups at mechanical ventilation time, ICU observation time, the incidence of postoperative SIRS after 24h, hospitalization days, and the adverse reactions were not statistically significant.Conclusion: DEX has notable protective effects on systemic inflammatory response syndrome during extracorporeal circulation.

Objective 　Determination of Wilson disease gene mRNA expression in human fibroblast cell strain (Me32aT22/2L) by reverse transcription-polymerase chain reaction (RT-PCR). Methods　Using lipofection reagent, the plasmid vector carrying the Wilson disease gene (pRc/CMV-WD) was transferred into Me32aT22/2L cultured in serum free complement medium. RT-PCR was used to determine WD mRNA expression in Me32aT22/2L. Results　 Wilson disease gene expression was detected in Me32aT22/2L, while no specific signals were detected in untransfected fibroblast. Conclusions　It demonstrated that Me32aT22/2L strain could express the Wilson disease gene, suggesting that Wilson disease gene transfer might develop a new approach to study Wilson disease.

Objective To investigate the effect of analgesia with dezocine on CD4+ and CD8+ T lymphocytes in patients after pulmonary lobectomy.Methods Seveuty patients with early non-small-cell lung cancer were selected,and they were scheduled the pulmonary lobectomy.The patients were divided into control group and dezocine group by random digits table method with 35 cases each.The patients in dezocine group were given 0.5 mg/ml dezocine by patient-controlled analgesia (PCA) pump,and the patients in control group were given 0.9％ sodium chloride by PCA pump.The peripheral blood lymphocytes count and percentages of CD4+ T lymphocytes,CD8+ T lymphocytes,natural killer cell (NK cell) before anesthesia induction and 4,24,48 h after operation were detected.Results In control group,the peripheral blood lymphocytes count and NK cell 24 h after operation were (1.08 ± 0.21) × 109/L and 0.141 4 ± 0.021 8,which before anesthesia induction were (1.71 ± 0.33) × 109/L and 0.190 9 ± 0.022 8,and there were statistical differences (P ＜ 0.05).In dezocine group,the peripheral blood lymphocytes count and NK cell 24 h after operation were (1.14 ±0.28) × 109/L and 0.124 9 ± 0.027 6,which before anesthesia induction were (1.69 ± 0.28) × 109/L and 0.198 6 ± 0.027 5,and there were statistical differences (P ＜ 0.05).The CD4+ T lymphocytes 24 h after operation in dezocine group was significantly higher than that in control group (0.355 6 ±0.031 1 vs.0.273 5 ±0.029 4),and there was statistical difference (P ＜0.05).There was no statistical difference in CD8+ T lymphocytes between the 2 groups (P ＞ 0.05).Conclusion Analgesia with dezocine can notably improve immunosuppression in patients after pulmonary lobectomy.

Objective To study the effects of body weight support treadmill training (BWSTT) or Du channel electroacupuncture (DcEA) on locomotion function and expressions of nerve growth factor (NGF) and growth-associated protein-43 (GAP-43) in rats with spinal cord transection. Methods 54 adult SD rats were divided into control group (n=18), BWSTT group (n=18) and DcEA group (n=18). Each group was divided into 3 subgroups (n=6) for 8 d, 15 d and 30 d respectively. Spinal cord transection was performed at T10 level. The locomotion function was assessed with Bosso-Beattie-Bresnahan locomotor rating scale for open field (BBB scale) and the expressions of NGF and GAP-43 in spinal cord tissues to the injury caudal ends were measured with immunohistochemistry. Results The scores of BBB scales and expressions of NGF and GAP-43 were higher in BWSTT group and DcEA group than those in the control group on the 8th d,15th d and 30th d respectively (P<0.01); and were higher in DcEA group than in BWSTT group on the 15th d and 30th d respectively (P<0.01). Conclusion Both BWSTT and DcEA can improve the locomotion function in rats with spinal cord transection and increase expressions of NGF and GAP-43 in spinal cord tissues, and DcEA is more effective.

Objective To investigate the expression of DRAM in breast cancer cell line MCF-7 in radiation-induced autophagy. Methods GFP-LC3 transfection method was used to observe autophagy bubble.Real time-PCR was used to detect DRAM and MAPLC3 from transcriptional and translational level,respectively. The silencing vector from gene engineering was introduced by calcium phosphate transfection.Results Compared with the control group,GFP-LC3 increased significantly after 8 Gy irradiation by immunofluorescent assay,and the level of MAP-LC3 expression was higher than control group after 8 Gy irradiation by Western blot ( F =5.38,8.72,10.63,15.23,20.78 and 55.23,P ＜ 0.05 ).DRAM protein expression increased significantly at 2 h in the 8 Gy time-dose study,up to maximum at the 32 h( F =116.34,P ＜ 0.05 ).In DRAM model,the expression of LC3 and DRAM decreased significantly (F =20.36 and 40.35,P ＜ 0.05 ) and DRAM expression increased 8 Gy post-irradiation,but still lower than that in 8 Gy irradiation wild-type group.The LC3 expression also decreasaed 8 Gy post-irradiation(F =50.34,P ＜ 0.05 ).Conclusions DRAM plays an important role in irradiation-induced autophagy in breast cancer cell.DRAM might participate in the process and serve as a theoretical target for clinical treatment of breast cancer.

Objective To investigate the value of combined detection of serum cystatin C(Cys-C),alpha 1- microspheres(α1-MG)and urinary microalbumin(mAlb)in the early stage of renal damage in patients with primary hypertension(PH).Methods One hundred and twenty-eight patients with PH were divided into 3 groups according to urinary albumin excretion rate(UAER),group I,normal proteinuria,47 cases,group II microalbuminuria,43 cases,group III,massive proteinuria,38 cases and 50 healthy subjects(the control group)served as research object.Hitachi 7180 automatic biochemical analyzer was applied with latex turbidimetric immunoassay to detect serum Cys-C,α1-MG and mAlb,serum creatinine(Crea)and serum urea (Urea)were determined by enzyme method.Comparative analysis on the early diagnosis efficiency of renal damage was made o based on the combined detection results.Results PH patients were tested by Cys-C( (0.98±0.13)mg/L vs.(1.62±0.55)mg/L vs.(3.17±1.04)mg/L),α1-MG((29.32±6.46)mg/L vs. (38.58±11.79)mg/L vs.(61.42±22.71)mg/L),mAlb((26.35±6.53)mg/24 h vs.(54.24±12.78)mg/24 h vs.(373.6±98.40)mg/24 h),Crea((5.06±0.67)μmol/L vs.(89.64±28.84)μmol/L vs.(164.16 ±58.77)μmol/L),Urea((5.06 ± 0.67)mmol/L vs.(7.87 ± 2.95)mmol/L vs.(15.10 ± 7.41)mmol/L, compared with those in the control group((0.69±0.08)mg/L,(17.15±4.30)mg/L,(19.55±4.28)mg/24 h,(68.39±20.11)μmol/L,(5.05±0.78)mmol/L,there were significant differences among the four groups(F=164.74,93.01,553.27,61.38,65.24,P=0).The results of Cys-C,α1-MG and mAlb in patients with PH were higher than those in the control group(P<0.05,P<0.01).The results of Cys-C,α1-MG and mAlb in group II were higher than those in group I,but lower than those in group III(P<0.01).The detection results of Crea in group I and group II were higher than those in the control group(P>0.05),but lower than that in group III(P<0.05).The result of Urea in group I was higher than that in the control group(P>0.05),but lower than those in group II and group III(P<0.05).Crea Cys-C and Urea were positively correlated(r=0.385,r=0.310,P=0.006,P=0.025),α1-MG was positively correlated with Crea,Urea(r=0.310,r=0.228,P=0.028,P=0.043); mAlb was positively correlated with Crea and Urea(r=0.382,r=0.302,P=0.006,P=0.033). The specificity of combined detection of three markers(55.3%)was lower than the individual detection(74.5%,68.1%,80.9%)and also lower than the two-unit(70.2%,63.8%,index 78.7%); its sensitivity(90.7%)was significantly higher than that of the individual detection(65.1%,58.1%,67.4%)and the two-unit(79.1%,74.4%,69.8%).Conclusion Cys-C,α1-MG and mAlb can be used as important indicators in the diagnosis in the patients with PH renal damage,which can increase the effectiveness PH early renal damage diagnosis,can be used for early detection of renal damage in PH patients,with the disease monitoring.

Sequential activation of the JNK pathway components, including Rac1/Cdc42, MLKs (mixed-lineage kinases), MKK4/7 and JNKs, plays a required role in many cell death paradigms. Those components are organized by a scaffold protein, POSH (Plenty of SH3's), to ensure the effective activation of the JNK pathway and cell death upon apoptotic stimuli. We have shown recently that the expression of POSH and MLK family proteins are regulated through protein stability. By generating a variety of mutants, we provide evidence here that the Nterminal half of POSH is accountable for its stability regulation and its over-expression-induced cell death. In addition, POSH's ability to induce apoptosis is correlated with its stability as well as its MLK binding ability. MLK family's stability, like that of POSH, requires activation of JNKs. However, we were surprised to find out that the widely used dominant negative (d/n) form of c-Jun could down-regulate MLK's stability, indicating that peptide from d/n c-Jun can be potentially developed into a therapeutical drug.
Adaptor Proteins, Signal Transducing ; genetics ; metabolism ; Animals ; Apoptosis ; physiology ; Base Sequence ; Cell Line ; DNA Primers ; genetics ; Humans ; JNK Mitogen-Activated Protein Kinases ; antagonists & inhibitors ; genetics ; metabolism ; MAP Kinase Kinase Kinases ; genetics ; metabolism ; Mutant Proteins ; genetics ; metabolism ; Nuclear Proteins ; genetics ; metabolism ; PC12 Cells ; Peptide Fragments ; genetics ; metabolism ; Protein Stability ; Rats ; Recombinant Proteins ; genetics ; metabolism ; Signal Transduction ; physiology ; Transfection ; Ubiquitin-Protein Ligases ; genetics ; metabolism