Objective To explore the effects of endoplasmic reticulum stress response(ERS) and its related apoptosis on dopaminergic neurons death.Methods NGF treated-PC12 cells were treated with 6-OHDA,MPP+ and rotenone.MTT assay and flow cytometry were used to measure the cell viability and the rate of cells apoptosis induced by those neurotoxins at different concentrations and times.The expressions of ERS-related gene XBP1,Grp78,CHOP,caspase-12 in drug-treated group and reserpine preincubation group were determined by RT-polymerase chain reaction(RT-PCR) and immunohistochemistry.Results After exposing to different concentration toxins,the vitality of PC12 cells was decreased by 52% at 100?mol/L 6-OHDA,by 44% at 75?mol/L MPP~+,and by 40% at 20 nmol/L rotenone for 24 hours respectively and ws decreased in a dose dependent manner. FCM assay confirmed time-dependent cell apoptosis.The apoptotic cells ratio of 24 h groups were (31.22?3.21)%,(27.46?2.35)%,(29.26?2.53)%,respectively(P＜0.01).In 6-OHDA groups,the gene expressions of XBP1,Grp78 were approximately 2-fold increased after 8 h exposure, CHOP reached peak level at 16 h(149.5?3.3% vs 35.9?1.8%,P＜0.01).The transcription level of caspase-12 was significantly higher than normal control at 16h[(95.4?2.8% vs(23.8?3.0)%, P＜0.01],but was alleviated by reserpine prcincubation(62.15?4.3%,P＜0.05).The increased expressions of Grp78 and CHOP after drug exposure were confirmed by immunochemistry stain.The similar results were observed in MPP~+ and rotenone groups.Conclusions The excessive ERS and ERS-activated cell apoptosis pathway may be involved in selective death of dopaminergic neurons.

Objective To explore the relationship between pulse pressure (PP) and artery atherosclerosis in elders. Methods Totally 2358 eiders( > 60y) with the risk factors of artery atherosclerosis from Yangpu District DOI:10.3760/cma.j.issn.1008-6315.2009.09.006Central Hospital and 11 Community Health Service Centers were enrolled . The basic information includes age, sex, height,body mass, history of smoking, blood pressure, heart rate, history of coronary heart disease, ischemic stoke, chronic kidney diseases and diabetes,etc. The fasting blood sugar(FBS) ,blood lipid( total cholesterel,triglyceride, low-density lipreprotein, high-deusity liproprotein), serum creatinine, serum uric acid, serum alanine aminotransferase were measured. The elders were divided into two groups according to the PP( PP <60 mm Hg and PP≥60 mm Hg) to analyse the relationship between PP and age, BMI, SBP, DBP, HR, FBS, TC, TG, LDL-C, HDL-C, Cr, Ccr, UA, AST and history of coronary heart disease,ischemic stoke,chronic renal disease, and diabetes. Results Serum levels of FBS,TC,TG, LDL-C, UA in the group with PP less than 60 nun Hg were ( 5.6±1.7 ) mmol/L, (4.3±1.3 ) mmol/L, ( 1.5±1.2) mmol/L, ( 2.5±1.1 ) mmol/L, UA ( 246.8±131.2 ) μmol/L, that in the group with PP over or equal to 60 mm Hg group were (6.3±2.6) mmol/L, (4.8±1.2) mmol/L, ( 1.9±1.1 ) mmol/L, (2.9±1.2) mmol/L, (291.4±133.6) μmol/L, Serum levels of FBS, TC, TG, LDL-C, UA in the group with PP over 60 mm Hg were obviously elevated as compared with that in the group with PP lower than 60 ram Hg(P <0.01 ) ; The preven-lence rates of coronary heart disease ischemic stoke,chronic,renal disease, diabetes in the group with PP less than 60 mm Hg were 10.8% ,26.0%, 12.4%, 17.6%, that in the group with PP over or equal to 60 mm Hg group were 17.8% 、31.7% ,16.9% ,23.8% (P <0.01 ). The preveulence of coronary heart disease ischemic stoke,chronic re-nal disease, diabetes were significandy increased in the group with PP over or equal to 60 mm Hg group. Logistic re-gression analysis showed PP was closely related to FBS, BMI, age, LDL-C, UA, HDL-C ( β = 0.103,0.093.0.097, 0.089 ,0.076 , - 0.057, P < 0.05 ). Conclusions For those elders with the basis of artery atherosclerosis, high PP can inhance the development of artery atheresclerosis and also inhance the occurance and development of cardio-ce-rebrovascular and renal diseases.

BACKGROUND: In the treatment of unicompartmental knee osteoarthritis, unicompartmental knee arthroplasty has the advantages of less blood loss, less trauma, quick recovery, maximum retention of bone mass, low cost, fewer complications and higher patient satisfaction. However, for the mobile bearing and the fixed bearing, the choice of single condylar prosthesis is till controversial.OBJECTIVE: To investigate the difference of LINK fixed-bearing and Oxford mobile-bearing unicompartment arthroplasty in relieving knee joint pain, reconstruction of the knee joint function, correct the knee joint deformity for medial unicompartment knee osteoarthritis, thus providing the basis for selecting a better prosthesis.METHODS: Ninety patients with medial unicompartment knee osteoarthritis undergoing unicompartmental knee arthroplasty in the Department of Joint Surgery, Guangdong Provincial Hospital of TCM from December 2014 to June 2016 were analyzed retrospectively, followed by allotted into Oxford mobile-bearing and LINK fixed-bearing groups (n=45 per group). The preoperative and postoperative pain level, range of motion of the knee, complications, and limb alignment were assessed. The range of motion and function of the knee were evaluated by Knee Society Score system.RESULTS AND CONCLUSION: (1) The two kinds of prosthesises both could obtain good clinical effectiveness. (2)There was no significant difference in the Knee Society Score, Visual Analogue Scale scores, or the range of motion of the knee between two groups (P > 0.05). (3) No complications such as deep vein thrombosis, prosthesis loosening and joint revision were found in both groups. (4) The correction of limb alignment showed significant difference between two groups (P < 0.05). The corrective femorotibial angle in the Oxford mobile-bearing group was larger than that in the LINK fixed-bearing group. (5) These findings manifest that under the precondition of strictly mastering the indications, both fixed-bearing and Oxford mobile-bearing unicompartment arthroplasties can effectively relieve the pain and improve the function of knee joint in the treatment of unicompartmental knee osteoarthritis. Oxford mobile-bearing prosthesis exhibits advantage in restoring the limb alignment; the movable platform lower limb alignment correction angle is often bigger than fixed platform correction angle, but may accelerate the lateral cartilage wear.

Aged ; Carcinoma, Small Cell ; genetics ; Case-Control Studies ; Cytochrome P-450 CYP2E1 ; genetics ; Female ; Humans ; Lung Neoplasms ; genetics ; Male ; Middle Aged ; Polymorphism, Single Nucleotide

OBJECTIVETo study the influence of alcohol on the liver sinusoids endothelial cell (LSEC) fenestrae of rats.

METHODSSetting up the rat model of alcoholic liver disease by orogastric administration of alcohol, then kill the experimental and control groups of rats at the end of 4 weeks, 8 weeks and 12 weeks after alcohol feeding, and also at the end of another 12 weeks after balance foods feeding succeeding with alcohol feeding for 12 weeks. Staining the liver tissue by means of HE method and observing the successive change of LSEC fenestrae by transmission electron microscope.

RESULTSThe normal LSEC was flat with nucleus and organelle arranged regularly. The distal cytoplasm displayed as lamina with many fenestrae, not accompanied by basement membrane (BM) formation under the endothelial cell. At the end of 4 weeks of alcohol feeding, fenestrae decreased at the partial distal LSEC cytoplasm, but no BM developed. At the end of 8 weeks, fenestrae decreased significantly, even disappeared, with the BM developed incompletely under the endothelial cell. Concomitantly, fibroblast with active function developed. At the end of 12 weeks, the changes became more obvious; the complete BM could even be seen. However, this kind of changes was mostly limited in the single or adjoining sinusoids, as well as with little widespread formation of fibrosis. At the end of 12 weeks of stopping alcohol feeding, defenestrae and development of BM attenuated obviously.

CONCLUSIONThe defenestrae and BM of LSEC develop gradually with the chronic alcohol stimulation. Sinusoid capillarization and liver fibrosis even form when significant changes happen. The early change of the limited defenestrae and capillarization may be the basis of alcohol periportal fibrosis formation. This kind of liver fibrosis can be reversible after stopping alcohol feeding.

Animals ; Basement Membrane ; pathology ; Endothelium ; drug effects ; pathology ; Ethanol ; Liver ; blood supply ; pathology ; Liver Cirrhosis, Experimental ; pathology ; Liver Diseases, Alcoholic ; pathology ; Male ; Rats ; Rats, Wistar

There are many E. coli rare codons in the gene of porcine interferon alpha-1. In order to obtain high expression of poIFN-alpha1 in E. coli, the cDNA encoded poIFN-alpha1 mature protein was synthesized using biased codons of E. coli without changing the original amino acid sequence and the terminator was changed as TAA. At the same time, Adenine and Thymine were used to the largest extent near the 5' terminus of poIFN-alpha1 mature protein gene. The synthesized gene was inserted into the Eco RI and Sal I site of the expression vector pRLC resulting pRLC-poIFN-alpha1. The poIFN-alpha1 is highly expressed in E. coli DH5alpha when the induction was carried out at 42 degrees C . The expressed poIFN-alpha1 account for 24.5% of the total cellular proteins and existed as inclusion body. The poIFN-alpha1 inclusion body was dissolved in 6mol/L guanidine chloride contained DTT and subsequently the denatured poIFN-alpha1 was re-natured by dilution in refolding buffer containing GSH and GSSH. In the present study it was found that the denatured poIFN-alpha1 was most efficiently re-natured in refolding buffer containing 1 mol/L guanidine chloride. In order to obtain pure protein, the concentrated re-natured poIFN-alpha1 was purified by Sephacryl S-200 chromatography. As a result, the purified poIFN-alpha1 is verified to be of high cytokine activity by inhibiting the cytopathic effect of vesicular stomatitis virus in MDBK cells, which is about 6.4 x 10(6) u/mg. This study paved the way for large-scale production of recombinant poIFN-alpha1 and its usage in virus disease control of pigs.
Animals ; Codon ; genetics ; Escherichia coli ; genetics ; metabolism ; Genetic Vectors ; Interferon-alpha ; biosynthesis ; genetics ; Recombinant Proteins ; biosynthesis ; genetics ; Swine ; Transduction, Genetic

Objective Transcription factor forkhead box L 2 (FOXL2) is a key regulator of granulosa cells (GCs) estrogen syn-thesis and function maintenance .However, the FOXL2 protein expres-sion and function regulation mechanism are unknown .We explored how DNAJB11 regulates estrogen synthesis of granulosa cells with immunoprecipitation , immunofluorescent staining and luciferase re-porter gene. Methods The expression and localization of DNAJB 11 was detected by immunohistochemistry staining in isolated mouse ovary tissues .we use immunoprecipitation , immunofluorescence staining and luciferase reporter gene assay to investigate the mechanism of DNAJB11, a member of the endoplasmic reticulum Hsp 40 /DnaJ family, regulating the estrogen synthesis in granulosa cells . Results DNAJB11 is expressed in the mouse ovary and granulosa cells .Follicle-stimulating hormone (FSH) promotes DNAJB11 ex-pression in a time and concentration dependent manner and induces endogenous DNAJB 11 protein translocation from the ER to the nu-cleus in KGN cells.Moreover, Adenovirus-mediated overexpression of DNAJB11 did not affect the proliferation of granulosa cells .How-ever, the concentration of estrogen in granulosa cells was affected by concentration -dependent and subcellular localization-dependent manner (10749±801.7 pg/mL vs 14217±1218.0 pg/mL P<0.01).Immunoprecipitation assay confirmed that DNAJB11 binds to FOXL2 in granulosa cells .When overexpressed in the nucleus of granulosa cells , DNAJB11 could significantly enhance the stability of FOXL2 (P<0.05) and promote FOXL2-mediated activity of Cyp19A1 promoter (P<0.01), while the expression of DNAJB11 in the nucleus increased the expression of Cyp 19A1 protein by 1.5 times ( P<0.05) . Conclusion These results demonstrate that DNAJB 11 was a new binding molecule of transcription factor FOXL 2 and regulated FOXL 2 protein stability and transcription activity .

OBJECTIVETo explore the effects of exogenous carbon monoxide-releasing molecules 2 (CORM-2) on the vitality and toxicity of E. coli ATCC 25922, and to analyze the potential mechanism.

METHODS(1) In vitro experiments. Standard strains of E. coli ATCC 25922 were divided into groups A (without addition), B, C, D, and E according to the random number table, and then the latter 4 groups were respectively cultured with 1.2 mmol/L CORM-2, 1.6 mmol/L CORM-2, 1.2 mmol/L inactive CORM-2 (iCORM-2), 1.6 mmol/L iCORM-2, with six samples in each group. After being cultured for 0, 3, 5, 8, 10, 12, 16, 20, 24, 27, 30, 48 hours, proliferative vitality of E. coli was examined (denoted as absorption value under 600 nm wavelength), and bacteria number was counted. Other standard strains of E. coli ATCC 25922 were divided into groups F (without addition) and G (cultured with 0.8 mmol/L CORM-2), the expressions of genes fliA, dnaK, marA, and waaQ related to E. coli were detected by quantitative real-time (qRT)-PCR. (2) In vivo experiments. Other standard strains of E. coli ATCC 25922 were grouped as A', B', C', D', and E' and treated with the same method as that in groups A, B, C, D, and E, and 0.5 mL bacterial liquid of each group were collected when the absorption value of bacterial liquid in group A' was equal to 0.4 (under 600 nm wavelength). Seventy-two C57BL/6 mice were divided into groups, namely blank control (without treatment), H, I, J, K, and L according to the random number table, with 12 mice in each group. The mice in the latter 5 groups were intraperitoneally injected with 0.5 mL bacterial suspension of groups A', B', C', D', and E' respectively. After injection, general condition of mice in groups H, I, J, K, and L was observed. The serum levels of TNF-α and IL-6 were determined at post injection hour (PIH) 6, 12. The liver and lung samples were harvested for determination of myeloperoxidase (MPO) activity at PIH 12. The same process was carried out in blank control group. Data were processed with repeated measure analysis of variance (ANOVA), factorial design ANOVA, one-way ANOVA, and t test.

RESULTS(1) In vitro experiments. Compared with those of groups A and D, the proliferative vitality and bacteria number of E. coli in group B were all decreased (with F values respectively 1170.80, 217.52, P values all below 0.01). Compared with those of groups A and E, the proliferative vitality and bacteria number of E. coli in group C were also obviously decreased (with F values respectively 7948.34, 14 432.85, P values all below 0.01). Compared with those in group F, the expression of fliA was downregulated, while the expressions of dnaK, marA, and waaQ were upregulated in group G (with t values 30.28, -165.54, -168.88, -187.28, P values all below 0.01). (2) In vivo experiments. Symptoms including listlessness and tachypnea were observed in mice in groups H, K, and L, and they were ameliorated or not obvious in groups I and J. At PIH 6, the serum levels of TNF-α and IL-6 in groups H and K were respectively (647.3 ± 3.8) pg/mL, (3.44 ± 0.22) ng/mL and (639.3 ± 0.8) pg/mL, (2.47 ± 0.32) ng/mL, which were obviously higher than those in group I [(124.6 ± 10.7) pg/mL, (1.03 ± 0.16) ng/mL, with t values from 15.22 to 84.03, P values all below 0.01]. The serum levels of TNF-α and IL-6 in group J at PIH 6, 12 were also obviously decreased as compared with those in groups H and L (with t values from 19.27 to 245.34, P values all below 0.01). MPO activity of liver and lung tissues were significantly attenuated in group I at PIH 12 as compared with those in groups H and K, and it was also attenuated in group J when compared with those in groups H and L (with t values respectively from 17.36 to 18.92 and 2.35 to 3.61, P values all below 0.01).

CONCLUSIONSCORM-2 can obviously inhibit the vitality and toxicity of E. coli, which might be attributable to regulation of expressions of genes fliA, dnaK, marA, and waaQ of E. coli.

Animals ; Carbon Monoxide ; metabolism ; DNA-Binding Proteins ; metabolism ; Escherichia coli ; drug effects ; metabolism ; physiology ; Escherichia coli Proteins ; metabolism ; Glycosyltransferases ; metabolism ; HSP70 Heat-Shock Proteins ; metabolism ; Interleukin-6 ; blood ; Liver ; enzymology ; Lung ; enzymology ; Mice ; Mice, Inbred C57BL ; Organometallic Compounds ; pharmacology ; Peroxidase ; metabolism ; Sigma Factor ; metabolism ; Tumor Necrosis Factor-alpha ; blood

OBJECTIVETo study the clinicopathologic features and prognostic factors of gastrointestinal stromal tumor (GIST).

METHODSClinicopathologic data of 247 patients with GIST from January 2003 to November 2012 in the Affiliated Hospital of Qingdao University Medical College, and the prognostic factors were evaluated retrospectively by univariate and multivariate analysis with Log-rank test and Cox proportional hazard model.

RESULTSPatients were followed up with a median time of 26 months (1 to 113 months). Twenty-six patients developed recurrence or metastasis, and 18 died of GIST. The 1-, 3-, 5-year survival rates were 94%, 91% and 83% respectively. Univariate analysis showed that age, tumor location, tumor size, mitotic count and tumor rupture were predictive factors of survival after resection of primary GIST (all P<0.01). For patients at intermediate and high risk to relapse, imatinib group had a higher 5-year overall survival rate than non-imatinib group (85.7% vs. 81.0%, P<0.05). Multivariate analysis revealed that tumor size (RR=2.248, 95%CI:1.081-4.677, P=0.030), mitotic count (RR=2.220, 95%CI:1.032-4.776, P=0.041) and tumor rupture (RR=5.183, 95%CI:1.677-16.017, P=0.004) were independent prognostic factors.

CONCLUSIONSTumor size, mitotic count and tumor rupture affect the prognosis after resection of primary GIST independently. Imatinib adjuvant therapy can improve overall survival of patients at intermediate and high risk to relapse after surgery.

Adolescent ; Adult ; Aged ; Aged, 80 and over ; Female ; Follow-Up Studies ; Gastrointestinal Neoplasms ; diagnosis ; pathology ; surgery ; Gastrointestinal Stromal Tumors ; diagnosis ; pathology ; surgery ; Humans ; Male ; Middle Aged ; Multivariate Analysis ; Prognosis ; Proportional Hazards Models ; Retrospective Studies ; Young Adult

Objective To explore the effect of dendritic cells (DC) modified with transforming growth factor β1 (TGF-β1) gene on the expressions of CD28/CTLA-4:B7 costimulatory molecules in peripheral blood mononuclear cells (PBMC) in the Lewis rats with experimental autoimmune myasthenia gravis (EAMG). Methods Thirty inbreeding line, healthy, female Lewis rats were divided randomly into 6 groups: normal group, EAMG group, DC treatment group, pcDNA3-TGF-β1-DCtreatment group, pcDNA3-DC control group and normal saline group. The rats were immunized with the AChR protein extracted from electric organ of Narcine timilei and CFA in the groups except normal group. 2×106 pcDNA3-TGF-β1-DCs/rat were injected subcutaneously into the backs of the rats which had been immunized 5 d earlier with AChR+CFA. The rats in DC treatment group, pcDNA3-DC control group and normal saline group were injected in parallel with untreated DC, pcDNA3-DC and normal saline, respectively. Seven weeks after the first immunization, the expressions of CD28 mRNA and CTLA-4 mRNA were detected by reverse transcription-polymerase chain reaction (RT-PCR), and the levels of B7-1 and B7-2 on the surface of PBMC were examined using flow cytometry. Results (1)The low expression of CD28 mRNA and rare expression of CTLA-4 mRNA were found in the normal rats, and both expressions increased markedly in EAMG rats (P＜0.001). Compared to those in EAMG group, the expression of CD28 mRNA decreased and CTLA-4 mRNA was upregulated after the treatment with pcDNA3-TGF-β1-DC (P＜0.05). There was no significant difference in the expressions of CD28 mRNA and CTLA-4 mRNA among the EAMG group, DC treatment group, pcDNA3-DC control group and normal saline group (P＞0.05). (2) The expressions of CD28, CTLA-4, B7-1 and B7-2 on the surface of PBMC were rare in normal rats, which increased significantly in EAMG rats (P＜0.001). The levels of CD28, B7-1 and B7-2 in pcDNA3-TGF-β1-DC group were lower than those in EAMG group (P＜0.01), but the level of CTLA-4 was higher than that in EAMG group (P＜0.05). They showed no statistically difference among the EAMG group, DC treatment group, pcDNA3-DC control group and normal saline group (P＞0.05). Conclusions The expressions of CD28/CTLA-4:B7 costimulatory molecules are abnormal in the rats with EAMG. The regulation of CD28/CTLA-4:B7 costimulatory pathways may play a critical role in the mechanism of the treatment with DC transfected with pcDNA3-TGF-β1 in the incipient EAMG rats.