Abstract:Objective
To observe the changes of neuronal apoptosis  and expression levels of  Caspase-9  in hippocampus and amygdala of posttraumatic stress disorder(PTSD)  rats,and to disscus the mechanism of the volume abnormalities of hippocampus and amygdala. Methods 100 male Wistar rats were divided into control group (n=20) and PTSD group (n=80). The PTSD rat model was produced by giving the rats single-prolonged stresS followed a single inescapable electric foot shock (SPS & S). The behavior changes of rats were examined using Morris water maze test and freezing behavior test. The positive apoptotic rates of  cells and apoptotic rates in hippocampus and amygdalawere detected by TUNEL-staining and  flow  cytometry (FCM). The expression level of Caspase-9 was detected by Western blotting method. Results The excape latency (EL) time and percentage of freezing behavior in PTSD group were higher than those in  control group (P<0.05). The percentages of TUNEL-positive cells and the apoptotic  rates in hippocampus and amygdala of PTSD  1,4,7 and 14 d rats were higher than those in control group (P<0.05),but there was no significant difference between PTSD 28 d rats and control rats (P>0.05). The expression levels of Caspase-9 in hippocampus and amygdala of  PTSD 1,4,7 and 14 d rats were higher than those in control group(P<0.05). Conclusion The apoptosis of neurons and expression levels of Caspase-9 in hippocampus and amygdala  of PTSD rats are increased,which indicates that apoptosis may be duced to the volume abnormalities of hippocampus and amygdala of PTSD.

To investigate the effects of rhein lysinate (RHL),taxol and their combination on the proliferation and apoptosis of lung cancer H460 cells,and to clarify their mechanisms. Methods The lung cancer H460 cells in logarithm growth phase were selected and randomly divided into   control group(adding no drug),taxol group(1 μmol•L^-1 taxol),RHL group(100 μmol•L^-1 RHL)  and taxol combined with RHL group;at the same time,blank control group was set up. MTT assay was used to detect the proliferation of H460 cells and flow cytometry method was used to analyze the apoptotic rates in various groups 48 h after treatment. The expression levels of apoptosis-related  proteins  and MEK/ERK proteins were detected  by Western blotting method. Results
After culture for 48 h,the cell proliferation rate in taxol combined with RHL group was lower than those in control,taxol,and RHL groups (P<0.05); the apoptotic rate in taxol combined with RHL  group was higher than those in control,taxol,and RHL groups (P<0.05);the expressions of  caspase-3 and poly ADP-ribose polymerase (PARP) fragment protein  in  taxol combined with RHL  group were higher than those in control,taxol,and RHL groups;whereas the expressions of Bcl-2 and NF-κB proteins were lower than those in control,taxol,and RHL groups. Moreover,the phosphorylation levels of MEK and ERK proteins up-regulated  by taxol in RHL group were decreased.  Conclusion Both taxol and RHL can inhibit cell proliferation and induce apoptosis of lung cancer H460 cells,and RHL can enhance the taxol-induced cytotoxicity and apoptosis by reducing the activity  of ERK and increasing the expression levels of  caspase-3 and PARP fragment protein.

Abstract:Objective
To construct the prokaryotic expression plasmids of   truncated regions of human  p21-activated kinase 6(PAK6) to induce and identify their recombinant proteins,and to provide basis for discussing the biological function of PAK6 gene.Methods The eukaryotic expression plasmid pcDNA3.1-GFP-PAK6  was used as the template,and the truncated segments  of PAK6 gene were amplified by PCR method. The truncated segments were cloned into GST-tagged prokaryotic expression vector pGEX-5X-1 after do uble enzyme digestion of  EcoRⅠ/XhoⅠ.The truncated plasmids were transfect
ed into E.coli BL21 and the  PAK6 gene truncated fusion proteins were induced by IPTG and verified by Western blotting method.Results The vector fragment (5 000 bp) and PAK6 1-55（165 bp），PAK6 56-210（465 bp），PAK6 211-410（600 bp），and PAK6 385-681（891 bp）  vector fragments being consistent with the expected fragments were obtained after  double enzyme digestion of EcoRⅠ/XhoⅠ.The Western blotting  results showed that the relative molecular mass of GST-PAK6 truncated fusion proteins of PAK6 truncated region plasmids were 32 000,43 000,48 000,and 60 000,respectively.
Conclusion GST-tagged prokaryotic expression plasmids of different truncated regions of  PAK6 gene are constructed and their recombinant proteins are expressed successfully.

To observe the influence of different acupoints of preventive acupuncture on gastric tissue damage of stress ulcer rat and  protein mass spectrometry,and to clarify the effect of different acupoints of preventive acupuncture on preventing stress ulcer,and  to obtain the related diffierential  expression proteins.Methods 120 male Wistar rats were randomly divided into normal control group (n=15),bundled control group (n=15),stress ulcer model group(n=15),Mu point group(n=15),Xiahe point group (n=15),He and Mu points group(n=15),Back-shu group(n=15),and Back-shu and Mu points group(n=15).The stress ulcer model was constructed by water-immersion method after acupuncture.The morphological changes of the damage of the rat gastric mucosa were observed,and the ulcer index was assessed in various groups.The stomach tissue proteins were extracted,and the proteins in supernatant of stomach tissue homogenate were separated  by SDS-PAGE gel electrophoresis method. Nanoliter electrospray mass high-performance liquid  instrument combined with  LTQ linear ion trap mass spectrometer (Eksigent-Thermo LTQ-MS)  was used to detect the expression levels of proteins. Results The ulcer index of the rats in  He and Mu points group was lower than those  in Mu point group, Xiahe point group,Back-shu point  group, Back-shu and Mu points group(P<0.05). After screening,the interesting differential expression proteins were obtained as follows:①Enzymes,including glyceraldehyde 3-phosphate dehydrogenase (GAPDH),carbonic anhydrase 1 (Car1),carbonic anhydrase 3 (Car3),aconitum synthase (Aco2movement),alpha-enolase (of Eno1),beta enolase (Eno3),and gamma enolase;②Skeleton proteins,including alpha1 actin protein (Actc1),beta actin (Actb),α-actin protein (Acta1),and 43 000 protein (Actg2);
③ Transport proteins,including serum transferrin (TF subtype 1) and calreticulin (Calr);④Heat shock protein,including 10 000 heat shock protein (Hspe1).Conclusion The preventive  effect of  He and  Mu points on stress ulcer  is better than those of single point  and Back-shu and Mu points,its mechanism may be associated with a variety of protein expression changes.

Abstract:Objective
To investigate the effect of urea transporter B(UT-B) gene on testosterone secreted by Leydig cells of male mice  by using UT-B gene knockout mouse models.Methods The UT-B gene knockout (UT-B－／－) mice were fed,bred and identified.5  UT-B wild type (UT-B+/+) male mice  and 5 UT-B－／－ male mice from the same genetic background aged  4 weeks were selected.The levels  of the serum testosterone and luteotropic hormone(LH) of two kinds of mice were detected by radioimmunoassay;the Leydig cells of two kinds of mice were isolated and cultured and then identified;the level of testosterone in culture supernatants was detected.Results The UT-B+/+ male mice and UT-B－／－ male mice were obtained  successfully;the primary Leydig cells were isolated and cultured and the cell purity was up to 80% identified by 3β-HSD staining method.The serum testosterone level  of    UT-B－／－  male mice aged 4 weeks ［（7.29±1.27）×10-2 μg•L^-1］was higher than that of UT-B+/+ male mice［（5.53±1.58）×10-2 μg•L^-1］（P<0.05），but there was no significant difference of serum LH levels  between two kinds of mice (P>0.05).The testosterone level  in the Leydig cells supernatants of  UT-B－／－ male mice［（17.300±1.179 ） nmol•L^-1］　was higher than that of UT-B+/+ male mice［(12.300±0.916 ) nmol•L^-1］（P<0.01）.
Conclusion The ability of Leydig cells secreting testosterone of UT-B－／－ male mice aged 4 week  is stronger than that of UT-B+/+ mice,which might be one of the reasons that the reproductive age of  UT-B－／－ male mice   is earlier than that of UT-B+/+ male mice.

Abstract:Objective To treat the titanium alloy serface with femto-second lasers with  different energy densities,and   compare their influence  in adhesion and proliferation of osteoblasts, and to discuss the effect of  femto-second laser on the implant material surface modification. Methods Disk-like Ti-6Al-4V titanium alloy was polished and processed by femto-second lasers with different energy densities   and then  divided into low-energy group（0.07 J•cm^-2）　and high-energy group （1.40 J•cm^-2）, and the untreated disk-like Ti-6Al-4V titanium alloy   was regarded as control group; the morphology of the material surface  in  each group was observed by scanning   electron microscope. The human osteoblast-like cell line MG63 were co-cultured with different materials, the morphology of the adherent cells in  each group was observed  by scanning electron microscope, and the quantity of the adherent cells on the surface was detected  by acridine  orange staining and fluorescence microscope, and the optical density(A) value of MG63 cells was dete
rmined by methylthiazolyldiphenyl-tetrazolium bromide(MTT) colorimetric analysis, and the cell proliferation rates at different time were analyzed. Results Two
kinds of micro-morphology structure were formed on the titanium alloy surface modified by femto-second laser , one of which was  nanoscaled paralleled stripe structure and the other was micro-nano composite structure of micron-sized protrudes combined with the nanoscaled stripes structure . The amounts of the adheren cells in different groups were close at 24 h. After 48 h, the morphology of single cell on material surface in each group was not obvious, however, the cell cover area of micro-nano composite structure surface was bigger  than those  of  nanoscaled composite structure surface and the smooth surface in control group.The cell proliferation rate in high-energy group was higher than those in  low-energy group and control group(P＜0.05).  Conclusion The micro-nano composite structure on the titanium alloy surface induced  by femto-second laser can promote the  adhesion and proliferation of MG63 osteoblasts.

Abstract:Objective
To investigate the influence of garlic polysaccharide(GPL) in the  learning and memory abilities  of mice with chronic alcoholism, and to explain the protective effect and its mechanism,and to provide experimental basis for developing natural medicine for prevention and treatment for  alcohol-related brain damage.Methods 94 Kunming mice were randomly divided into normal group,model group,GPL low　dose,GPL medium dose and GPL  high dose(150,200,and 250 mg•kg^-1•d^-1)  groups;there were 14 mice in normal group and 20 mice in the other each group. The mouse model of brain damage induced by  chronic alcoholism was  constructed  by gastric perfusion of  56°spirit and then the mice were treated with different doses of GPL.Place navigation and spatial probe tests in Morris water maze(MWM) were conducted to detect the learning and memory abilities of mice according to their escape latencies,staying  time in target quadrant,and times of crossing platform in the 6th and  9 th weeks. After ten weeks,the activities of acetylcholinesterase (TChE) and monoamine oxidase(MAO) in the   brain tissue of mice were detected  and  the morphological changes of mouse  brain tissue were observed by  light microscope.  Results Compared  with normal group,the mice in model group had longer escape latency in the  6th and 9th weeks (P<0.05),less time in target quadrant and fewer times of crossing the platform in the 9th week (P<0.05). Compared with normal group,the activities of TChE and MAO in the  brain tissue of the mice in  model group were increased statistically (P<0.05) and the cerebral hippocampal C1 area in model group showed more cell necrosis and edema after 10 weeks.Compared  with  normal group,the mice in  GPL high dose group had longer escape latency,less time in target quadrant,fewer times of crossing the platform (P<0.05) on the 2nd day in the 9th week  while the mice in the other two GPL groups had no statistical differences (P>0.05). Compared with  model group,the mice in three GPL groups had shorter escape latencies in the  6th and 9th weeks(P<0.05) and the mice in GPL  low and medium doses  groups had longer time in target quadrant and more times of crossing the platform in the  9th week (P<0.05). Compared with model group,the activities of TChE and MAO in the brain tissue of the mice in  three GPL groups were decreased statistically (P<0.05) and  after 10 weeks the cerebral hippocampal C1 area of the mice in these groups showed fewer necrotic cells,especially in GPL  medium dose group.  Conclusion GPL has anti-agonistic effect on the   injury of brain cells and the decresing of learning and  memory abilities induced by chronic alcoholism and its mechanism may be related with regulating and improving the function of cholinergic and nervous system.

Abstract:Objective
To observe the expressions of nuclear factor E2-related factor 2(Nrf2) and NADPH quinine oxidoreductase(NQO1) in gastric cancer stem cells and normal gastric mucosa tissue,and to investigate the mechanism of Nrf2/NQO1 signal pathway in oxidative stress  of  gastric cancer stem cells.Methods Tumor spheroids suspended separation method was used to select stem cells in 60 gastric cancer tissue specimens. The stem cells were randomly divided into stem cells early group and activiated group. 30 cases of  normal gastric  mucosa tissues were regarded as normal control group. Western blotting method was applied to detect the expressions of Nrf2 and NQO1 protein in various groups. The activities of superoxide dismutase(SOD) and malondialdehyde(MDA) were observed. Results Compared with normal control group,the expression levels of Nrf2 and NQO1  in stem cells early group and activated group were increased(P<0.05),and the activity of SOD was decreased while the activity of MDA was increased significantly(P<0.05). Compared with  stem cells early group,the expression levels  of Nrf2 and  NQO1 in activated group were remarkably increased(P<0.05),and the activity of SOD was increased while the activity of MDA was decreased significantly(P<0.05). Conclusion Activating Nrf2/NQO1 signal pathway can adjust the activities of SOD and MDA in gastric mucosa tissue to enhance the tolerance of gastric mucosal cells against oxidative stress in order to protect the cells.

Abstract:Objective To isolate mesenchymal stem cells (MSCs) from the amniotic membrane of human term placenta or  the femur and tibia of Wistar rats and to study the methods of isolation and cultivation,biological characteristics,surface markers and multidirectional differentiation potential of human placenta-derived  MSCs(HPMSCs) and rat bone marrow mesenchymal stem cells(BMSCs). Methods The  HPMSCs were isolated from human term placenta tissue by digestion of collagenase Ⅱ.The BMSCs from the femur and tibia of Wistar rats aged 4  weeks were isolated,cultured and purified by the whole bone marrow adherence method. The proliferation ability was detected by living cell number counting method. The positive expression rates of the surface markers of two kinds of  cells were detected by flow cytometry. The cells were induced into osteoblasts with dexamethasone,vintamin C, and β sodium glycerophosphate; and they were also induced into adipocytes with insulin,dexamethasone,IBMX, and indometacin. After induction,the cells were observed by alizarin red staining and oil red O  staining. Results HPMSCs  were spindle-shaped adherent cells  with  strong proliferation ability.The positive expression rates of CD44 and CD34 were 94.45% and 1.67%，respectively.The MSCs isolated from the femur and tibia of rats were rounded,spindle-shaped and polygonal adherent cells with strong proliferation ability.The positive expression rates of CD44 and CD34 were 93.10% and 2.68%,respectively. After induction,the results of  alizarin red staining and oil red O staining  were both positive.Conclusion HPMSCs have similar biological characteristics to rat BMSCs. They also have multidirectional differentiation potential,but HPMSCs have stronger proliferation ability.

Abstract:Objective
To separate and identify the exosomes  from colorectal carcinoma cells and to preliminarily study their  immunological characteristics,and to provide  basis for the application of exosomes in colorectal carcinoma immunotherapy.  Methods The normal exosomes and heat shock exosomes  of HCT116 and SW480 cells  were separated by ultracentrifugation and  purified by 220 nm microspore film. The morphology of exosomes  were observed under  transmission electron microscope. The protein compositions of cells and exosomes were analyzed by SDS-PAGE method. The  promoting  function of  exosomes in proliferation of peripheral blood monouclear cells (PBMCs)  was detected. Results Under  transmission electron microscope,the normal exosomes and heat shock exosomes  had similar appearance. They were either spherical or oval membrane vesicles,whose diameters were about 50-100 nm. The results of SDS-PAGE showed the differences of protein compositions between colorectal cell lysates  and exosomes derived from colorectal carcinoma cells. Compared with untreated and heat shock treated colorectal carcinoma cells,the  exosomes had stronger promotion effect on  PBMCs proliferation(P<0.05). The proliferaton of PBMCs could be reinforced with heat shock(P<0.05).   Conclusion For separation and identification of the exosomes derived from colorectal carcinoma cells,the method of ultracentrifugation and filtration is simple and feasible. The differences of protein composition between colorectal cell lysates  and exosomes may be the major reason of their differences in immunological  characteristics. The immuogenicity of exosomes could be enhanced  by treating the cells with heat shock. 

Abstract:Objective To explore the  methods to induce and expand the  NK and CIK cells in vitro and to compare the proliferation abilities and anti-tumor activitiesof NK and CIK cells. Methods Peripheral blood mononuclear cells(PBMCs) were separated from the peripheral blood of cancer patients. The PBMCs were induced by cytokines to culture NK and CIK cells. The morphological changes of NK and CIK cells were observed. The percentages of CD3－CD56+NK and CD3+CD56+ CIK cells were detected by flow cytometry. And the expansion folds of NK and CIK cells were calculated. The levels of interferon-γ(IFN-γ) secreted by NK and CIK cells were detected by ELISA method. And the killing activities of NK and CIK cells against K562 cell strain were detected by CCK-8 method. Results After induction and expansion in vitro for 14 d,the NK cells expanded (160.00±12.15) folds,and the CIK cells expanded (110.00±15.48) folds. The expansion ability of the  NK cells was higher than that of the  CIK cells（P<0.05）.The secretion levels of IFN-γ in the supernatant of NK and CIK culture system raised to (4 227.75±731.94) and (525.96±304.84)  μg/L,respectively. The secretion level of IFN-γ of the NK cells was higher than that of the CIK cells（P<0.01）.After induction and expansion,the cytotoxicities of NK and CIK cells against K562 cells were enhanced with the increase of effector-target  ratio.The killing rates of NK and CIK cells against K562 cells were 65.35% and 57.68% at 25∶1 effector-target  ratio,respectively.The killingactivity of NK cells was higher than that of CIK cells at the same effector-target  ratio（P<0.01）.Conclusion The induction and expansion system of NK and CIK cells  in vitro is established successfully,and the expansion capability,the secretion level of IFN-γ,and the cytotoxicity of the NK cells are higher than those of the CIK cells.

Objective To investigate the effect of 2% nano-silver base inorganic antibacterial agent on concentration of  residual methylmethacrylate (MMA) of room temperature curing polymethylmethacrylate(PMMA) material and to observe  the concentration changes of the residual MMA  after treated with  low temperature plasma argon (Ar) and tetrafluoromethane (CF4).Methods 2% nano-silver base inorganic antibacterial agent was added into the room temperature curing PMMA material through ball milling method to make specimen. Both antibacterial room temperature curing PMMA material group(antibacterial group) and conventional room temperature curing PMMA group(control group) were treated with Ar and CF4 plasma surface modification and surface bombardment processing. And then acetone and chloroform were used to extract the residual MMA of the specimens before and after treatment  and residual MMA released from the specimens into artificial saliva,separately. Gas chromatography(GC) method was used to determine the concentrations of residual MMA in tiye pumping and extract.Results Compared with control group,the addition of 2% nano-silver base inorganic antibacterial agent did not show statistical significance for the residual MMAconcentration in PMMA material in antibacterial group had no significant difference(P>0.05). There were siginficant differences of the concentration and release amount of MMA in the materials after treated with  low temperature plasma,and the concentration of the residual MMA in the materials  obtained in control group  after  treatment of CF4 plasma bombardment was the lowest.The content of MMA in the artificial saliva was also quite different   in the second day compared with the first day(P<0.05).  After 2 d,with the prolongation of   time,the amount of MMA released from polymers tended to be stable. In  antibacterial group,when the specimens were treated with  CF4 plasma surface bombardment and surface modification,the maximum amounts of MMA appeared in the second day and the fourth day,which were higher than those of the other time points,respectively(P<0.05).Conclusion The addition of  2% nano-silver base inorganic antibacterial agent will not affect the  residual amount of MMA in the PMMA material obviously. Low temperature plasma treatment technique can effectively reduce the amount of residual MMA and its emission in room temperature curing PMMA material.

Abstract:Objective To explore the therapeutic effect of suppository of Jiechang An combined with  Shuxue Ning on rats with ulcerative colitis(UC) induced with TNBS  and to clarify its mechanisms of Huoxue,anti-inflammtory,and immunomodulation.Methods 64 Wistar rats were randomly divided into control group(n=8),model group(n=8),mesalazine group(0.380 g/kg,n=8),Shuxue Ning group(1.8 mL?kg-1,n=8),suppository of Jiechang An  A and B  groups(1.215 g/kg，2.430 g/kg,n=8),suppository of Jiechang An combined Shuxue Ning A and B groups（1.215 g/kg+1.8 mL/kg and 2.430 g/kg+1.8 mL/kg,n=8).The UC model was induced by TNBS in rats.The drugs were administrated for 14 d after  modelling.The symptomatology,body weight,blood index,and proliferation ability of lymphocytes were observed.The expression levels of IFN-γ and TNF-α mRAN were detected with RT-PCR.The  pathological changes of colon tissue of rats  and the expression  of CD4+ in spleen tissue of rats were examined  by HE staining and  immunohistochemical staining. Results  Compared with model group,the symptoms  were improved  in combined drug groups,the body weights and stimulation index(SI) were increased;the platelet (PLT) amount(P<0.05 or P<0.01),the number of three kinds of inflammatory cells,the expression levels of IFN-γ，TNF-α mRNA  were decreased in combined drug groups;but the prothrombin time (PT)and partial thromboplastin time (APTT) were prolonged in combined drug groups（P<0.05 or P<0.01）.Obvious   colonic mucosa damages，such as  erosion and  ulcer formation of colonic mucosa and  a large number of infiltrated inflammatory cells  in model group were found. However,the colonic mucosa damage and inflammatory infiltrating were much slighter;parts of colonic mucosa ulcer mucous membrane epithelium was repaired,and the bodies of gland arranged in good order relatively, and the  damaged cyathiform cells were decreased in combined drug groups. Immunohistochemical staining showed that the numbers of cells with positive CD4+expression  in  combined drug groups were decreased.Conclusion Suppository of Jiechang An combined with  Shuxue Ning has therapeutic effect in UC rats,and its mechanism may be related to reducing inflammatory reaction through Huoxue and immunomodulation.

Objective To discuss the anti-fatigue effects of siraitia grosvenorii fruit extracts (SGFE) on the mice with exercise fatigue and to provide basis for clinical application of SGFE. Methods 144 male ICR mice were randomly divided into control group(n=36),SGFE low dosage group(n=36),SGFE middle dosage group(n=36) and SGFE high dosage  group(n=36).The mice in control group were received an oral administration of physiological saline in a volume of 1.0 mL,and the mice in other three groups were received 1.0 mL of SGFE (100,200 and 400 mg?kg-1,once a day) for 28 d. All mice were forced to swim for 15 min each day. After 28 d,the blood,liver and gastrocnemius were collected after forced  swimming test,and the biochemical parameters related to fatigue,including blood lactic acid(BLA),blood urea nitrogen(BUN),liver glycogen,and muscle glycogen were measured.Results Compared with control group,the exhautive swimming time of the mice in SGFE  low,middle and high dosage  groups was prolonged(P<0.05),but there were no significant differences between SGFE low dosage,middle dosage and high dosage  groups (P>0.05);the levels  of BLA of mice in SGFE low dosage,middle dosage and high dosage  groups were significantly lower than that in control group(P<0.05),and the BLA levels showed  a decreasing trend with the  increase of SGFE  in SGFE  low,middle  and high dosage  groups.The levels of BUN  of mice in SGFE middle  and high dosage groups were significantly lower than that in control group (P<0.05),but there was no significantdifference between SGFE  low dosage group and control group.The levels of liver glycogen and muscle glycogen of mice in SGFE low,middle  and high dosage  groups were significantly higher than that in control group(P<0.05),and they showed a increasing trend with the increase of SGFE  dosagein   SGFE low,middle and high dosage groups. Conclusion SGFE  has significant anti-fatigue effect on the  mice with exercise fatigue.

Objective To construct the  gene expression vector  of  enhanced green fluorescent protein （eGFP）regulated by human CD133 gene promoter and to observe the expression ability of the human CD133 gene promoter in laryngocarcinoma Hep-2 cell line,and to provide experimental basis for the target therapy of tumor stem cells(CSCs) regulated by  human CD133 gene promoter.  Methods The CD133 gene promoter was cloned from the genomic DNAs of human peripheral blood by PCR method and  cloned into lentivirus pWPXLd-eGFP by using gene recombination technology to construct eGFP gene expression vector induced by human CD133 gene promoter. The recombined vector was identified by PCR and enzyme digestion. The recombined vector was transfected into Hep-2 cells induced by liposome. The cells were divided into  blank control group (non-transfected with plasmid),positive control  group (transfected with pWPXLd),and experiment  group(transfected with pWPXLd-eGFP-of CD133);and the expression of eGFP gene in Hep-2 cells was observed under fluorescence microscope. Results  The specific 1 810 bp CD133 gene promoter  fragment was obtained successfully by PCR method. The eGFP  gene expression vector regulated by  human CD133 promoter was constructed successfully by PCR and enzyme digestion method.The  eGFP expressions in Hep-2 cells in  positive control group and experiment group were found,but weren’t in blank control group under fluorescence microscope. Conclusion The eGFP  gene expression vector regulated by  human CD133 gene promoter can regulate the expression of eGFP gene in laryngocarcinoma Hep-2 cells.

Abstract:Objective To investigate the therapeutic  effect of gink gobioba extract (GBE) on diabetic cardiomyopathy (DCM) induced by streptozotocin (SZT) and isoprenaline (ISO) in rats. Methods 20 male Wistar rats were randomly divided into normal control group(n=5),model group(n=5),positive control group(n=5),and treatment group (n=5).The rats in normal control group were given nothing and the other rats were given STZ and ISO to construct DCM rat models.The rats in model group were given saline and the rats in positive control group and treatment groupwere given benazepril and GBE,respectively.The body weight,blood sugar level,serum AST and LDH levels,expression level of TGF-β1 protein,and morphology of myocardium of rats  in various groups were observed. Results 1 and 2  weeks after treatment,the body weight and blood sugar level of the  rats  in model group were lower than those in  normal control group（P<0.05）. The body weights and blood sugar levels of the rats in positive control and treatment group were lower than those in  normal control group（P<0.05）. The levels of AST,LDH,and TGF-β1 in model group were higher than those in   normal group(P<0.05）;and the levels of AST,LDH,and TGF-β1 in positive control and treatment group were higher than those in model group(P<0.05).The myocardial  cells of the  rats in normal control group had no swelling.The cells in model group showed clear boundaries and more scattered necrosis,and there were lots of fibroblasts  and some collagen fibers in focal necrosis.The myocardial cells of the  rats intreatment group had smaller focal necrosis and lower fibrosis.Conclusion GBE can alleviate myocardial injury induced by STZ plus ISO of rats.

Abstract:Objective To investigate the protective effects of ginsenoside Re  on human embryonic fibroblast RSa cells damaged by ultraviolet C(UVC),and to discuss the anti-UVC me
chanism of ginsenoside Re.Methods The RSa cells were irradiated with different dosages of UVC,and the RSa cells were treated with  50 mg?L-1 ginsenosides   before and after  irradiation(UVC+Re group),at the same time,  UVC model group and UVC+Rg1 group were established. MTT assay was used to analyze the  survival rate of cells;apoptosis and cell cycle were measured by flow cytometry (FCM) method,and the activity of superoxide dismutase (SOD) was determined by enzyme biochemical methods.The survival rate of cells after treated with SOD inhibitor was detected by colony formation assay. Results The results of MTT assay showed that the survival rate of  the cells in  UVC+ Re group and UVC+ Rg1 group were significantly increased   compared with UVC model group(P<0.01 or P<0.05). The FCM results showed that  different dosages of UVC could induce early apoptosis of the RSa cells. The apoptotic  rate of cells in UVC+Re group was significantly decreased compared  with UVC model group. The cell cycle detection results showed that the  percentage of the RSa cells at G2 phase was significantly increased after UVC irradiation（P<0.01）,and the percentage  of the Rsa cells at G2 phase  in UVC+ Re group was significantly decreased   compared with UVC model group（P<0.01),and the ratio of the RSa cells at G2 phase in UVC+Re group was significantly decreased.Compared with UVC model group,the activity of SOD in UVC+ Re group was incresed （P<0.01）. Conclusion Ginsenoside Re has anti-UVC efficiency and can enhance the survival rate of RSa cells,and inhibit early   apoptosis and increase the SOD activity.

Objective To explore the expressions of poly ADP-ribose polymerase(PARP-1),cysteinyl aspartate specific proteinase(Caspase-3),survivin and Bax in cervical squamous cell carcinoma tissues and their relationships with the  clinicopathogical characteristics of cervical carcinoma,and to provide experimental basis for early diagnosis and  treatment for cervical carcinoma.Methods Immunohistochemical method was used to detect the expression rates of PARP-1,Caspase-3,survivin and Bax in specimens of 60 cases of cervical squamous cell carcinoma,40 cases of cervical carcinoma in situ and 20 cases of  normal cervical epithelium (NCE) adjacent to carcinoma.The relationships between their expression levels and clinical stages,pathological types and lymph node metastasis or not were analyzed.Results The positive expression rates of PARP-1 in cervical squamous cell carcinoma,cervical carcinoma in situ and NCE tissue were 68.33%,62.50%,and 25.00%,respectively;the positive expression rates of Caspase-3 in cervical squamous cell carcinoma,cervical carcinoma in situ and NCE tissue were 43.33%,47.50%,and 70.00%, respectively;the positive expression rates of survivin  were 95.00%,92.50%,and 5.00%,respectively;the positive expression rates of Bax  were 31.67%,32.50%,and 75.00%,respectively.The positive expression rates of PARP-1 and survivin    in  squamous cell carcinoma and carcinoma in situ groups were lower than those in NCE group (P<0.001);the positive expression rates of Caspase-3 and Bax  in  squamous cell carcinoma and carcinoma in situ groups were higher than those in NCE group (P<0.01).The  positive expression rates  of  PARP-1,survivin and Bax had relationships  with the clinical stage,histological type,and lymph node metastasis(P<0.01);but the positive expression rate of  Caspase-3 didn’t have relationship with those indexes mentioned above(P>0.01).The  positive expression rates of PARP-1 and Caspase-3 in cervical squamous cell carcinoma tissue were negatively correlated （r=－0.71,P=0.043）;the positive  expression rates of  survivin and Bax in cervical squamous cell carcinoma tissue  were negatively correlated（r=－0.63,P=0.038）.Conclusion The up-regulation  of the expression levels  of PARP-1 and survivin and the down-regulation of the levels of Caspase-3 and Bax expression are likely to play an important role in the invasion and metastasis of cervical squamous cell carcinoma.

Objective To study the relationship between the level of B-type natriuretic peptide(BNP)  and liver function of rats with  acute decompensated heart failure   and to explore the values of the changes of liver function in evaluation of curative effect and prognosis of the patients with heart failure.Methods 60 male SD rats were chosen and  intraperitoneally injected with isoproterenol for establishment of acute decompensated heart failure models.20 rats were randomly selected from the survival  rats after  successful modeling as heart failure group.And 20 normal rats were selected as control group.All rats were underwent heart echocardiography check.The left ventricular end-diastolie inner diameter（LVEDD）,the left ventricular end-systolic diameter (LVESD),and the left ventricular ejection fraction(LVEF) were caculated.The levels of  serum BNP,aspartate aminotransferase(AST),γ-glutamyl (γ-GT) were detected.  Results Compared with control group,the LVEF level of the  rats in heart failure group was decreased (P<0.01),the   BNP level  was  increased(P<0.01),and the AST and γ-GT levels were significantly increased(P<0.01). When LVEF and BNP were regarded  as independent variables,and  AST and γ-GT were regarded as the dependent variables,the linear regression analysis was performed;the γ-GT and AST levels were  elevated with the decreasing of LVEF level,they had a negative correlation (λ =－0.765,λ=－0.789) (P<0.01);the AST and γ-GT levels were elevated with the increasing of BNP level,and they had a positive correlation (λ=0.793,λ= 0.648)(P<0.01). Conclusion The heart function and liver function of the rats with  acute decompensated heart failure  are abnormale;the LVEF level is declined,and the  BNP,AST and γ-GT levels are increased;the  γ-GT and AST levels have  negative correlation with LVEF,and the AST and γ-GT levels have   positive correlation with BNP.It indicates that the changes of liver function can be used to evaluate the curative effect and prognosis of the patients with heart failure.

Abstract:Objective To investigate the hippocamal neuron injury   in pup rats caused by lead exposure  from pregnancy to juvenile stage and the mechanism related to zinc balance. Methods Twenty-five female Wistar rats were randomly divided into control group (n=10) and lead exposure group (n=15).The pup rats of each group were subdivided into three subgroups depending on different observation periods from pregnancy to postnatal 7,14,and 21 d (P7,P14 and P21 subgroups),respectively; each subgroup was composed of 8 pup rats.The female rats in  lead exposure  group were bred with 0.2% lead acetate in drinking water from pregnancy to P7,P14,and P21 of the pup rats,respectively,while the mothers in control group drank water consistently.The  blood and brain lead contents were detected on P7,P14,and  P21 of the pup rats in each subgroup.Timm’s staining was used to observe the mossy fiber (MF) in hippocamspus,and immunohistochemistry assay was performed to detect the expression levels of  zinc transporter 1 (ZnT-1).Results The blood and brain lead contents  in lead exposure subgroups were significantly higher than those in  corresponding control subgroups(P<0.01).There were mass of degenerated cells  in CA2 and CA3 regions of  hippocampus in pup rats after lead exposure,and the nerve cell arrangement was unconsolidated,and the number of cell layers was decreased.The Timm’s staining results showed that MF  sprouting presented on P21 of the pups in control group,and the absorbtion(A) value was 0.135 0±0.024 3.The A valueof P21 in lead exposure  group was 0.096 7±0.024 2,which was significantly lower than that in  control group (P<0.01).The expression of ZnT-1 was significantly decreased on  P7 of the pups in lead exposure  group compared with  control  group (P<0.01).There was no significant difference of ZnT-1 expression on P14 and P21 between lead exposure and control group.Conclusion There may exist neuronal zinc imbalance in pup rats after low-level lead exposure  from pregnancy to juvenile stage,and it may cause the decrease of the intracellular Zn2+level in hippocampal neurons after lead exposure.

Abstract:Objective To study the inhibitory effect of 3-bromopyruvate(3-BP) on proliferation of colon carcinoma HCT116 cells  and to explore its mechanism,and to provide  evidence for the clinical application of 3-BP.Methods HCT116 cells were cultivated in vitro and divided into control group and different concentrations of 3-BP groups(25,50,75,and 100 mg/L).The hexokinaseⅡ(HKⅡ) protein expression levels in various groups were determined by Western blotting method;the concentrations of glucose-6-phosphatase (G-6-P) were determined by G-6-P a
ssay kit;the proliferation activities of the cells were observed by MTT method;the cell cycle was measured by flow cytometry. Results Compared with control group,the HKⅡ protein level in 25 mg/L 3-BP group did not change significantly（P>0.05）;compared with control group and 25 mg/L 3-BP group,the HKⅡprotein levels in 50,75, and 100 mg/L 3-BP groups were decreased significantly（P<0.01）.Compared with control group,the concentration of G-6-P in culture medium in 25 mg/L 3-BP group did  not change significantly（P>0.05）;compared with control group and 25 mg/L 3-BP group,the concentration of G-6-P in culture medium in 50,75,and 100 mg/L 3-BP groups  were decreased significantly（P<0.01）.The MTT results showed that compared with control group,the survival rate in 25 mg/L 3-BP group did  not change significantly(P>0.05);compared with control group and 25 mg/L 3-BP group,the survival rates
 in 50,75,and 100 mg/L 3-BP groups were decreased markedly(P<0.05).The flow cytometry analysis results showed that compared with control group,the percentages of the cells at G1 phase and S phase  in 25 mg/L 3-BP group did  not  change significantly（P>0.05）;compared with  control group and  25 mg/L 3-BP group,the percentages of the cells at G1 phase  in 50,75 and 100 mg/L 3-BP groups were increased significantly(P<0.01)  and the percentages of the cells at S phase  were decreased(P<0.05).The cell cycle was arrested at G1 stage.Conclusion 3-BP could inhibit the proliferation of  HCT116 cells  significantly,which  may be related to the decreasing of HKⅡ protein expression and  use of glucose.

Abstract:Objective To observe the  changes of expression level  of matrix metalloproteinase-8(MMP-8) in periapical tissues of SD rats and to investigate the relationship between MMP-8 and the development of periapical periodontitis  reinfected by Enterococcus faecalis. Methods The maxillary first molar teeth of 42 SD rats  received lipopolysaccharide(LPS) and  were exposed to oral environment to establish primary infected periapical periodontitis  model. The rats were randomly divided into chronic periapical periodontitis group (no extra treatment)，calcium hydroxide disinfection group (disinfection with calcium hydroxide paste after the root canal preparation),formaldehyde cresol (FC) disinfection group (disinfection with FC cotton ball after the root canal preparation),and periapical periodontitis  reinfected by Enterococcus faecalis group (reinfection group,implantion with  Enterococcus faecalis 2 weeks after FC disinfection),respectively. Six rats were respectively sacrificed at the 6th week in chronic periapical periodontitis group and calcium hydroxide disinfection group,the 2nd week  of FC disinfection group and the 1st,2nd,3rd,and 4th week in reinfection  group. The suppermaxilla were obtained and subjected to immunohistochemical staining in order to test the changes of integral absorbance (IA) values  of MMP-8 in periapical tissue in various groups. Results The expression levels of MMP-8 in FC disinfection group(4.49±1.08)and calcium hydroxide disinfection group(14.84±2.60)were both lower than that in chronic periapical periodontitis group(27.76±2.60)(P<0.01),and the expression level of MMP-8 in  FC disinfection group was lower than that in calcium hydroxide disinfection group(P<0.01). The expression level of MMP-8 at the 1st week in reinfection group(42.08±4.19)was higher than that in chronic periapical periodontitis group,and the MMP-8 expression levels were gradually increased at  the  1st week,2nd week (61.32±5.81),3rd week(79.45±7.40),and 4th week (114.67±10.40) in a time-dependent manner,which were higher than those in the other groups,respectively(P<0.05).Conclusion The expression level of MMP-8 is correlated with the progression of tissue destruction of SD rats with periapical  periodontitis reinfected by Enterococcus faecalis,and the higher the expression of MMP-8 is,the worse of the course expansion is.

Abstract:Objective To explore the mechanism of  targeted inhibition of hTERTp/TK/pGL3 vector on  the telomerase activity and its killing effect on nasopharyngeal cancer stem cells.Methods The nasopharyngeal carcinaoma CD133+ stem cells,CD133－tumor cells, human umbilical vein endothelial cells (ECV cells) (control group) and nasopharyngeal carcinaoma SUNE unsorted cells were transfected by hTERTp/TK/pGL3 vector and its control vectors (CMV/TK/pGL3 and TK/pGL3).The telomerase avtivities of nasopharyngeal carcinaoma CD133+ stem cells and ECV cells were detected by Stretch PCR.The survival rates of CD133+stem cells and ECV cells were detected by  MTT method.Results After culture for 7 d in vitro,the number of nasopharyngeal carcinaoma CD133+ stem cells was increased, the tumorigenicity test of CD133+ stem cells  in vivo was positive.The telomerase activity of nasopharyngeal carcinaoma CD133+ stem cells was decreased after transfected by hTERTp/TK/pGL3 or CMV/TK/pGL3; the telomerase activity of EVC cells was negative.After transfected by TK/pGL3,CMV/TK/pGL3,and hTERTp/TK/pGL3,the  survival rates of CD133+ stem cells were 87.4％±0.4％,20.5％±0.4％,and 27.9％±0.2％,respectively. After transfected by TK/pGL3,CMV/TK/pGL3,and hTERTp/TK/pGL3,the  survival rates of ECV cells were 90.7％±0.1％,18.1％±0.2％,and 86.2％±0.1％,respectively.The efficiency of  hTERTp/TK/pGL3 vector  in  killing CD133+ stem cells was higher than that of  ECV cells（P<0.01）.Conclusion hTERTp/TK/pGL3 vector can targetedly inhibit the telomerase-positive nasopharyngeal carcinaoma CD133+ stem cells by down-regulation of telomerase activity.

Abstract:Objective To investigate the regulation of puerarin on the expression levels of endorphin and dopamine(DA) of   cortex,cerebellum,hippocampus,and striatum of the rats with  acute alcoholism and to provide  basis for clinical treatment of alcoholism and the exploitation of traditional Chinese herb pueraria . Methods 24 male SD rats were randomly divided into control group,alcoholism group and puerarin group,and there were 8 rats in each group. The expression levels of β-endorphin(β-EP),dynorphin（DnyA） and Leu-enkephalin（L-EK） in cortex,cerebellum,hippocampus and striatum were detected by radioimmunoassay,and the expression levels of DA,dihydroxy-phenyl acetic acid（DOPAC） and homovanillic acid(HVA) of the rats in various groups were detected by high performance liquid chromatography(HPLC) method. The ratio of (DOPAC+HVA)/DA was calculated.Results Compared with control group,the exrpession levels  of β-EP,DnyA and L-EK in  cortex of the rats in alcoholism group and puerarin group were decreased (P<0.01). The expression levels  of DA,DOPAC and HVA in various  brain regions were increased (P<0.01).Compared with alcoholism group,and the expression levels of β-EP,DnyA and L-EK in rat cortex in puerarin group were increased (P<0.05,P<0.01),and the expression levels of DA(P<0.01),DOPAC(P<0.01) and HVA in cortex and all the indexes mentioned above in the cerebellum,hippocampus and striatum were decreased(P<0.05,P<0.01),and the ratios of（DOPAC+HVA）/DA in cortex,cerebellum and hippocampus were increased,but it was decreased in striatum(P>0.05).Conclusion Puerarin has protective effect on rat brain damage induced by acute alcoholism and the mechanism may be related to the regulation  of puerarin on endorphin and DA on the rat brain.

Abstract:Objective To observe the effects of α-Fodrin siRNA on the  expressions of IFN-γ and IL-4 of Sjogren’s syndrome (SS)  model non-obese diabetic (NOD) mice and to discuss its possibility  on treatment of NOD. Methods 12 female NOD mice were randomly divided into control group (n=3),empty vector group (n=3),α-Fodrin siRNA1 group (n=3),and α-Fodrin siRNA2 group (n=3). The vectors expressing α-Fodrin and  siRNA  1 and  α-Fodrin siRNA2 were injected by tail vein into NOD mice in α-Fodrin siRNA1 group and α-Fodrin siRNA2 group,while the  NOD mice in control group and empty vector group were injected with the same dose of PBS and pGFP-V-RS empty vector,respectively. The water drinking amount of each mouse was detected in each week.The expression levels of IFN-γ and IL-4 in serum of the mice in various groups were detected by ELISA method on the 1st,3rd,5th,and 7th day after injection,and the expression levels of α-Fodrin mRNA in lung tissue of NOD mice were detected by RT-PCR method and the expression level of α-Fodrin protein was detected by immunohistochemistry method.Results There were no significant differences of water drinking amounts of the mice between four groups(P>0.05).The levels of IL-4 were higher and IFN-γ/IL-4 ratios were lower in siRNA1 group and siRNA2 group than control group and empty vector group on the 1st and 3rd day after injection (P<0.05);the levels of IL-4 and IFN-γ/IL-4 in control group and empty vector group had no  significant differences on the 1st and 3rd day after injection;so did α-Fodrin siRNA1 group and α-Fodrin siRNA2 group. There were no significant differences  of the levels of IL-4 and the ratios of IFN-γ/IL-4 between various groups on the 5th and the 7th day. The expression levels of α-Fodrin mRNA and protein were lower in siRNA1 group and siRNA2 group than control group and empty vector group on the 7th day after injection (P<0.05);the expression levels of α-Fodrin protein were lower in siRNA1 group and siRNA2 than control group and empty vector group on the 7th day after injection (P<0.05);the expression levels of α-Fodrin mRNA and protein in control group and empty vector group had no significant differences;the expression levels of α-Fodrin mRNA and protein in siRNA1 group and siRNA2 group had  no   significant difference on the 7th day after injection. Conclusion α-Fodrin siRNA can elevate the expression of IL-4 and suppress the expression levels of α-fodrin mRNA and protein,at the same time,it can decrease the ratios of IFN-γ/IL-4;α-Fodrin siRNA could reduce the levels of inflammatory cytokines of NOD mice and delay  the development of SS.

Abstract:Objective To study the expressions of P33ING1b and KAI1/CD82in gastric cardia carcinoma(GCA) tissue and their roles in the occurrence and  malignancy progress of　GCA. Methods  20 cases  of adult normal cardia mucosa tissue (normal group) and 51 cases  of GCA tissue (GCA group) were selected.Immunohistochemistry method was used to detect the positive expression rates  of P33INGlb and KAI1/CD82 in the tissue  in  two groups. The data  combined with the clinicopathological features was analyzed. Results KAI1/CD82 protein mainly expressed in the cytoplasm of GCA cells and cardia mucosal cells. The positive expression rate of KAI1/CD82 protein in normal gastric cardia tissue was 85.0%,and the positive expression rate of KAI1/CD82 protein in GCA tissue was 65.7%.The expression level of KAI1/CD82 protein in GCA tissue with lymph node metastasis was lower than that in GCA tissue  without lymph node metastasis(P<0.05).  The expression level of KAI1/CD82 protein in GCA tissue  with  serosa  invasion was lower than that in GCA tissue without serosa invasion (P<0.05). The loss or reduction of KAI1/CD82 protein expression had relationships with clinical grade of tumor(P<0.05).  P33INGlb protein mainly expressed in the cytoplasm and nucleolus of GCA cells and cardia mucosal cells. The expression level of P33INGlb protein in GCA tissue was lower than that in normal　gastric cardia　mucosa tissue (P<0.05). The loss or reduction of P33INGlb protein expression had relationships with clinical grade of tumor (P<0.05). Conclusion The expression of　KAI1/CD82 protein in GCA tissue has relationship with clinical grade,the depth of  tumor invasion,and lymth metastasis.The expression rate of P33INGlb protein in GCA tissue has relationship with  the clinical grade of tumor. The expressions of KAI1/CD82 protein and P33INGlb protein can be used as indicators to judge the malignant biological behavior of human GCA at molecular level.

Abstract:Objective To detect the expressions of SHIP mRNA and Caspase-3 mRNA in bone marrow tissue of  the patients with   myelodysplastic syndrome （MDS） and to discuss the relationship between SHIP mRNA and Caspase-3 mRNA expressions and the occurrence and development of MDS.Methods 20 low-risk MDS cases(low-risk group)，12 Int-1 MDS cases(Int-1 group),13 Int-2 MDS cases(Int-2 group),and 17 high-risk MDS cases(high-risk group) were selected according to International Prognostic Scoring Syterm(IPSS).30 healthy volunteers were regared as control group.The expression levels of SHIP mRNA and Caspase-3 mRNA in bone marrow mononuclear  cells (BMNC) in various groups were detected by RT-PCR method,and the relationship between them was analyzed. Results The positive rates of SHIP mRNA were 100% and 100% and its expression levels were 4.467±2.899 and 4.529±1.975 in low-risk group and Int-1 group,respectively,while they were 100% and 4.623±2.979 in control group,so there were no significant differences between low-risk group,Int-1 group and control group (P>0.05).The positive rates of SHIP mRNA were 90% and 88% and the expression levels were 2.124±1.880 and 2.574±1.807 in Int-2 group and high-risk group,respectively,so there were significantly differences compared with control group (P<0.05).The Pearson correlation analysis results showed that the positive correlations were found between the level of SHIP mRNA and the level of Caspase-3 mRNA in MDS patients(r=0.714，P>0.05).The expression level of SHIP mRNA was decreased with the increasing of premitive cells(F=3.7，P<0.05).There were no significant differences of SHIP mRNA expression levels of patients between different genders(t=0.940，P>0.05) and different ages(t=1.417，P>0.05).Conclusion The expression level of SHIP  is lower or the SHIP doesn’t  express in  BMNC of the high-risk MDS patients, and  its expression level   can be used as an indicator to judge the prognosis of MDS.
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Abstract:Objective To observe the effect of paired box gene 2 (PAX2) on the surface markers of renal tubular epithelial cells and induction of PAX2 on epithelial-mesenchymal transition (EMT) in vitro,and to provide basis for treatment of renal fibrosis. Methods The  rat normal renal tubular epithelial cell line NRK52E were divided into  control group,empty vector group(transfected with pGC-FU empty vector),and transfection group(transfected  with pGC-FU-GFP-PAX2 vector). The morphology of cells 72 h after transfection was evaluated by inverted phase contrast microscope. The expression levels of PAX2 protein and mRNA were evaluated by Western blotting and Real-time PCR methods.72 h after transfection,the expressions of E-cadherin and α-smooth muscle actin (α-SMA)  and their mRNA expressions were detected by  immunohistochemistry,Western blotting and Real-time PCR methods. Results 72 h after  tranfection of pGC-FU-GFP-PAX2,the fluorescence intensity   in transfection group was stronger than that in control group,and the shape of renal tubular epithelial cells  elongated.The  Western blotting results showed that the expression level of  PAX2 protein in transfection group was increased  compared with control and empty vector groups(P<0.05);the Real-time PCR results showed that  the expression level of PAX2 mRNA  in transfection group was increased  compared with control  group and empty vector group(P<0.05).The dyeing  of E-cadherin got weaker and the dyeing of α-SMA got stronger  in transfection group compared with control and  empty vector groups. The Western blotting results showed that the expression level of E-cadherin in transfection group was decreased and α-SMA was increased  compared with control and empty vector groups (P<0.05).The Real-time PCR results showed that the expression level of E-cadherin mRNA in transfection group was decreased and the expression level of α-SMA mRNA was increased in transfection group  compared with control and empty vector groups (P<0.05). Conclusion PAX2 transfection can reduce E-cadherin expression and increase α-SMA 　expression. PAX2 may  induce renal tubular mesenchymal transition in vitro.

Abstract:Objective To investigate the association between single nucleotide polymorphism (SNP) of Fok Ⅰ in vitamin D receptor (VDR) gene and different degrees of histological prostatitis(HP),and to discuss the role of genetic factors in  the onset of HP.Methods  169 patients with benign prostatic hyperplasia (BPH) complicated with  HP were selected.The mild(79 cases),moderate(52 cases) and  severe(38 cases) HP patients were distinguished by pathological  examination.Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) was used to determine the  SNP of Fok Ⅰ of patients.The distribution of genotypic and allelic frequencies of the patients in various groups was analyzed.Results The  distribution of genotypic frequency of Fok Ⅰ site in  mild and moderate  HP patients didn’t deviate from  Hardy-Weinberg equilibrium(P>0.05).The  distribution of genotypic frequency of Fok Ⅰ gene in  severe HP patients deviated from  Hardy-Weinberg equilibrium(P<0.05).The  distribution of FF,Ff,ff genotypes  in  mild and moderate HP patients were 32%（25/79）,46%（36/79）;22%（18/79）,23%（12/52）;40%（21/52），37%（19/52）, respectively;there was no significant difference between two groups(P>0.05).The  allelic frequencies of F and f were 54%（86/158），46%（72/158） in  mild HP patients  and 43%（45/104），57%（59/104） in moderate  HP patients, respectively;there was no significant difference between two groups (P>0.05).Conclusion VDR gene FokⅠ SNP may not associate with the  occurrence of different degrees of HP.

Objective To investigate the risk factors for mortality in elderly patients with femoral neck fractures during  one year period and to clarify the re
lationships between the risk factors and postoperative mortality.Methods  196 (64 male,132 female)  patients  with consecutive isolated nonpathologic h
ip fractures were chosen.All patients were more than 65 years old.The patients were treated with hemiarthroplasty.The clinical data of patients was collected.The relationship between age,gender,American Society of Anaesthesiologists (ASA) rating of operative risk,the time  from injury to surgery,
blood albumin level,  haemoglobin level, total lymphocyte count and mortality during 1 year after operation was investigated.
Results In total,13 patients died during one year, among them 1 patient in hospital ,and 12 patients in follow-up period;the survival rates were 99.5% after 3 months,96.9% after 6 months,and 94.8% after 1 year;the ASA scores  were Ⅲ-Ⅳ grades in 12 patients including 8 patients with ASA Ⅲ grade and 4 patients with ASA Ⅳ grade;there were 11 patients with preoperative blood albumin level<35 g?L-1,and 12 patients with 
  preoperative lymphocyte count<1 500 mL-1.The comparisons of blood album in level,haemoglobin level,ASA score,and lymphocyte count  between the death patients and the survival patients had statistical differences(P<0.05).There was no significant difference of ages
 between the death patients and the survival patients(P>0.05).There were no significant differences of constituent ratio between the patients with the time  from injury to surgery less than 5 d and the patients with the time from injury to surgery more than 5 d(P>0.05).
Conclusion A lower lymphocyte count,more than two comorbidities,blood albumin level<35 g/L^-1,haemoglobin level  <10 g/L^-1,and ASA Ⅲ-Ⅳ grade are significant factors in assessing the one-year mortality in elderly patients with femoral neck fractures;age,gender and the time  from injury to  surgery are not significant factors in assessing the one-year mortality in elderly patients with femoral neck fractures.

Objective To investigate the association between aldosterone synthase CYP11B2 (-344C/T)
gene polymorphism and ischemic stroke (IS) by Meta-analysis and to provide evide
nce for the treatment of IS.Methods Case-control studies on the association between CYP11B2 gene polymorphism and IS
 from  1990 to 2012 were collected from China national knowledge internet,Chinese biological medicine disk,Vip fulltext database,Wanfang fulltext database,PubMed and Cochrane Collaboration Database.Meta-analysis was carried out after strict screening.Heterogeneity test，sensitivity analysis and publication bias assessment were also performed.Results A total of 6 studies were included.In IS group,the pooled OR ( 95% CI ) of TT ／(TT + CC) genotype was 1.51( 1.06,2.16,P<0.05). In large artery atherosclerotic stroke LAA group,the pooled OR (95% CI ) of
TT/(TT + CC) genotype was 1.85(1.11,3.08,P<0.05).In multiple lacunar stroke (MLI) group,the pooled OR
(95% CI) of TT/ (TT + CC) genotype was 2.46(1.21,4.99,P<0.05).Conclusion The TT genotype of CYP11B2 gene is associated with IS,a
nd TT genotype is also associated with LAA and MLI,which indicats that CYP11B2 gene
is possiblely the candidate gene of IS.

Objective To explore the correlation between  single nucleotide
polymorphism (SNP) of four sites on  the Toll-like receptor  8 (TLR8) gene and
tuberculosis in Han population from Northeast china,and to analyze the differences of genotypic frequency
distribution between tuberculosis and other lung diseases patients. Methods
A case-control study for 368 cases of tuberculosis patients (case group) and 355 other lung disease patients (control group) in Han population from Northeast China was established.The ligase specific
detection technology was carried out to detect the genotypes of   rs3764880,rs3761624,rs3788935, and rs3764879 sites on   TLR8 gene in two groups.The
 distribution  of  allelic frequencies (GF)  of the patients in two groups were compa
red  and the odds ratio (OR)  and the distribution of genotypic frequencies of the  patients with different gender  were analyzed.Results The　 allelic frequency distribution of   rs3764880,rs3761624,rs3788935,and rs3764879 sites on   TLR8 gene between case group and control group was not statistically
 significant(P>0.05);the  distribution of allelic frequency of four sites on  TLR8 gene   of patients with different gender 
between  case group and control group was not statistically significant(P>0.05).Conclusion The distributions of SNP at  rs3764880,rs3761624,rs3788935,and rs3764879 sites on   TLR8 gene are same  between tuberculosis and other lung disease patients in H
an population from Northeast China,which indicates that there is no association between SNP of four sites on TLR8 gene and the occurrence of tuberculosis.

Objective To observe the effects of dexmedetomidine on levels of tumor necrosis factor-α(TNF-α)，interleukin-6 (IL-6)，serum creatinine(Scr) and blood urea nitrogen (BUN)  in the patients undergoing radical nephrectomy during perioperative period，and to probe the influences of dexmedetomidine on immune function and renal function.Methods Sixty patients with kidney cancer undergoing radical nephrectomy were randomly devided into dexmedetomidine group(n=30) and control group(n=30).The blood samples were taken immediately before induction of anesthesia(T0)，1 h after the beginning of  operation(T1)，at the end of  operation(T2)，1 d after operation(T3) and 3 d after operation(T4) for determining the levels the of TNF-α,IL-6,Scr and BUN.
Results Compared with T0 time point，the levels of serum TNF-α and IL-6 of the patients in dexmedetomidine group had no sianificant differences at
all time points（P>0.05）；the levels of serum TNF-α and IL-6 of the patients in control group were significantly increased at T1-T3 time points (P<0.05).Compared with control group，the levels of serum TNF-α and IL-6 of the patients in dexmedetomidine group were significantly decreased at T1-T3 time points (P<0.05).There were no significant differences of  Scr or BUN  levels between two groups during perioperative period(P>0.05).
Conclusion Continuously intravenous infusion of dexmedetomidine may effectively decrease the levels of serum TNF-α and IL-6 in patients undergoing radical nephrectomy during perioperative period and inhibit the perioperative stress response，and doesn’t have  influence on renal function.

Objective To evaluate the effectiveness of nasotracheal intubation using lightwand in patients with difficult airways in oral and maxillofacial surger
y,and to provide a new method for difficult airway management.Methods 76 ASAⅠor Ⅱ grade patients with difficult airways who required nasotracheal intubation were randomly divided into using lightwand group (lightwand group) (n=38) and blind intubation group (n=38).The intubation procedure was guided by the light spot at neck in lightwand group while in blind intubation group it was guided by the patients’ breathing sound.The intubation success rate,intubation time,changes of  mean arterial pressure(MAP) and heart rate(HR) and postoperative complications of the patients were compared between two groups.Results In lightwand group, the intubation success rate (89.5%,34/38) was obviously higher than blind intubation group (71.1%,27/38)(P<0.05);the intubation time ［(89.9±26.8) s］ was significantly lower than  blind intubation group ［(134.9±32.8) s］ (P<0.001);
the postoperative incidence rate of  pharyngalgia (14.7%,5/34)  was significantly lower than blind intubation group (40.7%,11/27) (P<
0.05);there were no significant differences of the incidence rates of hoarseness and epistaxis between two groups(P>0.05);the   MAP values of patients  were lower than blind intubation group during intubation period and 1 min after intubation,and there were no significant differences of HR  of patients at different time points between two groups(P>0.05).Conclusion To the  patients with difficult airways,the nasotracheal intubation using lightwand is superior to blind intubation with higher success rate,more stable haemodynamic responses,and less postoperative complications.The technique could be used as a simple and practical approach for nasotracheal intubation in difficult airways.

Objective To investigate the significance of the detection of circumferential resection margin(CMR) after rectal cancer excision for the treatment
and prognosis of rectal cancer,and to provide basis for  improving the survival rate of patients with mid-low rectal cancer.
Methods 78 patients with mid-low rectal cancer were operated following the principles of total mesorectal excision(TME).The specimens were made into large slices stained by HE and the positive rate of circumferential margin invasion (CMI)  of the specimen was detected.Then the patients were followed-up,and the  mortality rates,the local recurrence rates and the postoperative metastasis rates of the CRM positive and CRM negative patients were compared.
ResultsThe patients were all followed-up and the follow-up time was  mean 17  months.The  positive rate of CMI was 25.64% (20/78).
 The   positive rates of T1 and T2 tumor (0 % and 0 %) were lower than that of T3 tumor (29.85% ,20/67,P<0.05).The positive rates of CMI in well differentiated and moderately differentiated tumor were 12.50%(1/8) and 16.36%(9/55),respectively,which were lower than that in poorly differentiated tumor(66.67%,10/15,
P<0.05). The  positive rate of CMI in lymph node N0 group (12.77%，6/47) was lower than those in N1 and N2  groups(40.00%，8/20； 54.55%，6/11,P<0.05).The positive rate of CMI in the specimens with the distance from the lower edge of  tumor to the dentate line≤5 cm  (48.15%,13/27) was higher than that in the specimens with the distance >5 cm (13.73%,7/51,P<0.05). During the 17 months of follow-up,10  cases died,in which there were 6 cases of  CMI.Postoperative local recurrence was found in  4 cases of positive  CMI.Moreover,there were 6 cases of CMI in 10 cases of postoperative distant metastasis.There were significant differences of the  mortality,local recurrence rate and postoperative metastasis rate between CRM positive
 (30.0%，20.0%，and 30.0%) and CRM negative patients (6.9%，0%， and 6.9%)(P<0.05).Conclusion CRM should be routinely detected
 in the patients with  mid-low rectal cancer after operation.The mortality,the local recurrence rate and the postoperative metastasis rate of CMI patients are significantly increased;chemotherapy and radiotherapy should be applied in order to reduce the local recurrence rate and the mortality.

Objective To establish the method of using the atlas morphological indexes for sex determination in Jilin province and to evaluate its effect.
Methods The clinic neck CT images were used to reconstruct the 3D image of atlas.A total of 27 linear measurements on 8
aspects of the atlas were measured and the ratios were calculated.The 14 items were selected.Results Of the total 27 linear measurements,14 were sexually dimorphic（P<0.05）,and the accuracies of sex determination of 27 indexes were 52.0%-89.3%.The highest accuracy was width of vertebral body(
 86.7%).A function with variables predicting sex with 95.8% accuracy was derived by using stepwise method of discriminant function analysis:Y=1.308W－0.409CDF－0.469LTPSD－0.849LUACD+0.478RUACD+0.332RDACD+0.363ATH－0.334PTH－0.236PAL.Conclusion
The method of using atlas traits for sex determination in Jilin province  is practicable.

Abstract:Objective  To investigate the correlation between cranial magnetic resonance imaging (MRI) findings and autoantibody levels  in patients with neuropsychiatric systemic lupus erythematosus (NPSLE),and to elucidate the role of MRI findings in predicting the prognosis of NPSLE.
Methods In total,36 well-documented cases of NPSLE diagnosed definitely were selected.All the patients were divided into survival group (n=27) and
 dead group (n=9).Anti-nuclear antibodies and anti-dsDNA antibodies in serum were detected by indirect immunofluorescence and colloidal gold spot penetration method, respectively.Immunoblotting method was used to detect anti-Sm,anti-SSA,anti-SSB,anti U1-RNP, and anti-ribosomal P protein antibodies.Anti-cardiolipin antibody (ACL) was detected by ELISA method.The correlation between MRI findings of cerebral lesions and  autoantibodies,and prognosis was analyzed. Results  Cranial MRI scans on admission were abnormal in 32 patients (88.9%),among which 21 cases showed diffuse manifestations,10 cases showed focal lesion in brain and 1 case showed brain atrophy.The diffuse lesion on MRI showed multiple spotty or patchy normal intensity signal on T1-weighted image (T1WI) and high-intensity signal changes on T2-weighted image (T2WI).The focal lesion showed single spotty or patchy normal intensity signal on T1WI and high-intensity signal changes on T2WI.Thelesion sites included basal ganglia,subcortical white matter,anterior and posterior horn of lateral ventricle,semiovale center,cerebral cortex,brainstem and cerebellum.Of 9 patients in dead group,6 cases presented with focal lesion and the percentage of focal lesion cases in dead group was significantly higher than that in survival group (P<0.01).The percentages of lesion number in brainstem and basal ganglia in dead group were 11.5% and 26.9%, respectively,which were significantly higher than those in survival group (P<0.05 or P<0.01).The positive rate of ACL in cases with focal lesion was higher than that in cases with diffuse lesion (P<0.05).
Conclusion  NPSLE patients with focal lesion,brainstem and basal ganglia involvement on cranial MR images show an unfavorable outcome.
ACL is associated with focal lesion. There is a certain correlation between MRI findings and autoantibodies in patients with NPSLE.MRI findings may help to  predict the prognosis of NPSLE to some extent.

Abstract:Objective To analyze the effect factors of   depression occurrence in  patients with end-stage renal disease,and to elucida
te the  impact of depression on life quality of patients with end-stage renal disease. Methods 186  patients with end-stage renal disease were
 divided into depression and non-depression patients according to BECK depression questionary.Then the Kidney Disease Quality of Life Short For
m（KDQOL-SF）of patients were measured and the results were analyzed. Results In all 186 patients,138 (74.2%) were depressed,including 26  cases of marginal depression,72 cases of   moderate depression,37 cases of  severe  depression and 3  cases of  critical depression.The incid
ence of depression was not correlated with gender or marital status (P>0.05),but it was correlated with age,occupational status,educational level,economic status,duration of dialysis,and type of medicare (P<0.05).There were significant differences in symptom/problems,effects of kidney disease on daily life(EKD),work status(WS),cognitive function(CF),quality of social interaction(QSI),sexual function(SexF) and sleep between  depr
ession and non-depression patients (P<0.05),while there were no significant differences in burden of kindey disease(BKD),social support(SOS),dialysis staff encouragement(DSE) or patient satisfaction(PS) between two kinds of patients(P>0.05).Depression was correlated with pain,genera
l health preperceptions(GH),emotion well-being(EUB),role linitations caused by emotional health problems(RE),social function(SocF),energy /fatigue,and changes in health(GH) and general health scales (P<0.05),but depression  wasn’t correlated with   physical functioning(DF) or role limitations caused by physical health problems (RP)(P>0.05).  Conclusion 14 dimensionalities of survival quality (symptom/ problems,EKD,WS,CF,QSI,SexF,sleep,pain,GH
,EWB,RE,SocF,energy/fatigue,change in health and general health scales) are positively  correlated with depression occurrence in  patients with end-stage renal disease.

Abstract:Objective To construct the prokaryotic expression vector of  nonstructural protein 1(NS1) of Tick-borne encephalitis virus(TBEV) and to o
btain  high purity expression of NS1 and to provide  experimental basis for preparation of polyclonal antibody against NS1 and study on its function.
Methods The PCR primers were designed according to the target gene,and the prokaryotic expression vector pEASYTM-E1-NS1 was constructed using gene recombination method.The recombinant plasmids pEASYTM-E1-NS1 were transferred into BL21 cells,and the expressed fusi
on proteins were purified by nickel ion affinity purification and  the concentration of protein was analyzed by Bradford method after indu
ction by Isopropy1β-D-1-Thiogalactopyranoside(IPTG).The expression level of fusion protein was detected by SDS-PAGE
and Western blotting methods.Results The prokaryotic expression vector pEASYTM-E1-NS1 carrying  TBEV NS1  expressed fusion protein in the form of
inclusion body after induction;a kind of highly purified protein was obtained through affinity purification. The SDS-PAGE and Western blotting results showed that a kind of specific protein with a relative molecular mass of about 40 000 was found.Conclusion The prokaryotic expression vector pEASYTM-E1-NS1 is constructed successfully,and the fusion protein of  TBEV NS1 with  high purity  is acquired.

Abstract:Objective To establish conventional PCR and fluorescence quantitative PCR(FQ-PCR) methods for detection of NDM1 gene and to compare the detection results of the two methods.Methods Three pairs of primers that specifically recognized the NDM1 gene were respectively designed,and their reaction systems and parameters were confirmed.Under the optimized conditions,the conventional PCR  and FQ-PCR methods were evaluated for the specificity,sensitivity and detection effects on simulative clinical samples.Results  Conventional PCR and FQ-PCR methods were successfully established.The results of specificity analysis showed that conventional PCR and FQ-PCR  both presented  negative resuts  for seven familiar pathogenic bacteria.The threshold of FQ-PCR was 104 mL-1 and that of conventional PCR was 10⁶ mL^-1.The detection results of s
imulative clinical samples indicated that they had accordant results.Conclusion Both conventional PCR and FQ-PCR  methods possess good specificity,and the sensitivity of FQ-PCR is 100 times higher than conventional PCR.