To study the influence of Rhizoma Typhonii extracts in glioma SHG-44 cells and to explore the effects of Rhizoma Typhonii extracts on proliferation and apoptosis of　glioma cells.Methods Glioma SHG-44 cells were cultured and divided into control and 8,40,200,and 1 000  μg•L^-1 Rhizoma Typhonii extracts groups,and the influence of Rhizoma Typhonii extracts in proliferation of SHG-44 cells was measured by MTT assay.The morphology of apoptosis was observed with inverted microscope.The cell cycle and apoptotic rate were analyzed by flow cytometry (FCM).The expressions of Bcl-2 and Bax protein were detected by immunohistochemistry.Results Compared with control group,the poliferation of SHG-44 cells was inhibited after treated with Rhizoma Typhonii extracts at concentrations of 8 and 200 μg•L^-1 for 30 min and 3 h and the proliferation activities were decreased(P＜0.05);in 1 000 μg•L^-1 group　after treated for  30 min,1 h and 3 h,the proliferation activities were  significantly decreased (P＜0.05).The proliferation inhibition of differentconcentrations of Rhizoma Typhonii extracts was time- and dose-dependent.The apoptotic bodies were found under inverted microscope in different Rhizoma Typhonii extracts groups.The FCM results showed that part of cells were arrested at G2/M phase,and the apoptotic rates were increased with the  increasing of drug concentrations.Cell immunohistochemisty showed that compared with control group the expression of Bcl-2 protein in 200 μg•L^-1 Rhizoma Typhonii extracts groups was decreased(P<0.01) and the expression of Bax protein  was increased (P<0.01).Conclusion Rhizoma Typhonii extracts can inhibit the proliferation and induce apoptosis by down-regulating the Bcl-2 protein expression and up-regulating the Bax protein expression in SHG-44 cells.

Abstract:Objective To observe the influence of urantide on the injury of thoracic aorta in rats with atherosclerosis(AS) and to investigate its mechanism of prevention and treatment of AS.Methods The AS rat model was induced by feeding high fat diet and arterial intimal injury with intraperitoneal injection of vitamin D3,and the rats were randomly divided into normal control group,AS model group,positive medicine group and urantide group;biochemistry was used to detect the  levels of serum Ca ²⁺ ,triglycerides (TG),total cholesterol (TC),high-density lipoprotein (HDL) and low density lipoprotein (LDL);hematoxylin-eosin staining was performed to observe the morphogical changes of rat thoracic aorta; RT-PCR method was used to measure the mRNA expressions of urotensin Ⅱ(UⅡ) and G protein couple receptor 14(GPR14).Results The levels of serum Ca²⁺,TG,TC,HDL and LDL  of the rats in AS model group were significantly higher than those in normal control group（P<0.01）;compared with AS model group,the expression levels of the serum indicators were gradually decreased with the prolongation of time,and reached or approached the serum levels in  positive medicine group（P<0.01）.The rats in AS model group had the typical AS changes in  thoracic aorta;compared with normal control group,the expression levels of UⅡ and  GPR14 mRNA in AS model group were significantly increased（P<0.01）;compared with AS model group,the expression levels of UⅡ and GPR14 mRNA of the rats in urantide group were decreased（P<0.01） with the prolongation of time.Conclusion Urantide has significantly protective effect on the injury of thoracic aorta in AS rats.
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Abstract:Objective
To investigate the expressions of Notch signal molecules and their downstream target genes in human NK/,NK,T and B lymphoma cells and to expound the effect and significance of Notch signal molecules in NK cell tumor.Methods The expression levels of Notch signal  in YTS,SNK-6,Raji and Jurkat cells were detected  by Real-time PCR.Results The expression levels of Notch 1,3,4 signal molecules in YTS and SNK-6 cells were significantly higher than those in Raji and Jurkat cells（P<0.01）,but the expression levels of Notch 1,3,4 signal molecules in SNK-6 cells were lower than that in YTS cells(P<0.01).The expression levels of  Notch 2  in YTS and SNK-6 cells were significantly higher than that in Jurkat cells（P<0.01）,but there was  no  statistically significant difference compared  with  Raji cells （P>0.05）.The expression level of Hes1 in YTS cells was significantly higher than those in Raji and Jurkat cells（P<0.05 or P<0.01）,but the expression level in SNK-6 cells was lower than those in Raji and Jurkat cells（P<0.01）.Conclusion Notch signal molecules  express abnormally in   NK cell tumor,and they are significantly different with B and T lymphoma cells.

Abstract:Objective
To investigate the effect of rhein lysinate (RHL) on the proliferation and apoptosis of human lung cancer cell line H460 and the role of human epidermal growth factor receptor-2 (HER2) signal pathway in apoptosis,and to provide theoretical foundation for clinical research on RHL against lung cancer. Methods The H460 cells in which HER2 was over-expressed in logarithm growth phase were selected and randomly divided into blank control group and different concentrations (25,50,75,100,125,and 150 μmol•L^-1) of RHL groups. MTT assay was used to detect the proliferation abilities of H460 cells in various groups. The H460 cells were randomly divided into blank control group and 50,100,and 150 μmol•L^-1 RHL groups; flow cytometry was used to analyze apoptosis in various groups after 48 h. The expression levels of  apoptosis-related proteins and   HER2 signal pathway proteins were detected after 48 h by Western blotting method. Results  After 48 h cell culture,the cell viability rates in RHL groups were lower than that in blank control group (P<0.05),whereas the apoptotic rates in RHL groups were higher than that in blank control group (P<0.05). The inhibition of cell proliferation of RHL presented a dose-dependent manner in H460 cells,and so the apoptosis induced by RHL. The expressions of cleaved poly ADP-ribose polymerase (PARP) and caspase-3/7 in RHL groups were higher than those in blank control group (P<0.05) after 48 h cell culture in H460 cells,whereas the expressions of phosphorylation of HER2 and AKT and NF-κB were lower than those in control group (P<0.05).Conclusion RHL can induce apoptosis by inhibiting HER2/AKT/NF-κB signal pathway in H460 cells.

Abstract:Objective
To examine the regulatory mode of neuropeptide Y (NPY) in the central nucleus of amygdala(CeA) on feeding of fasted rats，and to explore the possible mechanism. Methods 34 male SD  rats were chosen,and 10 rats of them were divided into feeding group(n=5) and fasted 24 h group(n=5),and the rat brain was obtained. The expressions of NPY and neuropeptide Y 1 receptor(Y1R)in CeA were detected by immunohistochemical staining method. The other 24 rats were implanted cannula in CeA. The postoperative 12 rats were randomly divided into 2 groups,and the rats in one group were injected with NPY and the rats in another group were injected with saline in CeA(control 1 group),after 1,2 and 4 h,the intake of food was detected. The other postoperative 12 rats were randomly divided into 2 groups,and the rats in one group were injected with Y1R antagonist and the rats in another group were injected with saline in CeA(control 2 group),after 1,2 and 4 h,the intake of food was detected. Results  Compared with feeding group,the NPY expression in CeA of the rats in fasted 24 h group was significantly increased (P<0.05),and the Y1R expression in CeA of the rats in fasted 24 h group was significantly decreased (P<0.05). Compared with control 1 group,the intake of food of the rats   injected with NPY in CeA within 1 h was significantly increased(P<0.05);compared with control 2 group,the intake of food of the rats  injected with Y1R antagonist in CeA within 1 h was significantly decreased(P<0.05). Conclusion NPY in CeA has promotion effect on the feeding of fasted rats,and the mechanism may be regulated by Y1 receptor.
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Abstract:Objective
To observe the change of neuronal autophagy in hippocampus of depression model rats,and to investigate the reason of the abnormal hippocampal structure of the depression rats. Methods Twenty male SD rats were randomly divided into control group and model group,10 rats in each group. The depression rat model was produced by giving the  chronic unpredictable mild stress (CUMS). Electron microscope was used to detect the ultrastructure of neurons and the changes of autophagy. The expression levels of LC-3 and Beclin-1 in hippocampus tissue of rats  were detected by　Western blotting method. Results  Many autophagsome vacuoles were seen obviously in hippocampal neurons of the rats in model group under electron microscope. The  ratio of LC-3Ⅱ to LC-3Ⅰ　in control group was 0.99±0.11,while in model goup was 3.13±0.21;the expression levels of Beclin-1 in control and model groups were 0.79±0.09 and 2.15±0.28,respectively. The ratio of LC-3Ⅱ to LC-3Ⅰ and the expression level of Beclin-1 in model group were higher than those in control group (P<0.05). Conclusion Atuophagy is increased in hippocampal neurons of depression model rats,which indicats that autophagy may be duced to the abnormality of hippocampus volume.
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Abstract:Objective
To observe the inhibitory effect of andrographolide (Andro) on cell proliferation of  glioblastoma U87 cells,and to explore its regulatory mechanism.Methods The U87 cells at logarithmic growth phase were collected and divided into dimethy1 sulfoxide(DMSO) control group,4H-Andro control group and Andro group.The U87 cells were treated with DMSO,4H-Andro,and Andro with  different concentrations (0,5,10,15,20,25 μmol•L^-1).The cell viability was detected by MTT assay,and the mean inhibitory concentraion (IC50) was caculated, and Western blotting method was used to detect the expression intensity  of NF-κB protein  in U87 cells.Results The U87 cells were treated with  different concentrations of Andro,and the  IC50 of Andro was 8 μmol•L^-1.Compared with DMSO and 4H-Andro control groups,24 h after treatment of 8 μmol•L^-1 Andro,the proliferation activity of U87 cells was inhibited (P<0.01) and the inhibitory rate of proliferation was 50%;and the cell proliferation was obviously decreased after treated with Andro for 96 h (P<0.01) in U87 cells,and the inhibitory rate of proliferation was 65%.The expression intensity of NF-κB(pp65) protein was  down-regulated by 8 μmol•L^-1 Andro for 48 h compared with DMSO control  group　and 4H-Andro control group(P<0.01).Conclusion Andro, as an anti-inflammatory drug in clinic,plays a new function of suppressing glioblastoma cell proliferation  through inhibiting the expression intensity  of NF-κB protein.
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Abstract:Objective
To investigate the changes of each bone layer volume and mechanical properties of osteoarthritis(OA) rabbit femur and to clarify the correlation between the each bone layer volume and mechanical properties. Methods 0.3 mL•kg^-1 3% papain were injected into the joint space of right knee of 10 rabbits on the 1st,5th and 12th days,respectively,to establish OA models (model group); the left legs of the rabbits were used as normal control group. Four times of CT scanning were performed 3 d  before modeling and 7,27,37 d after modeling,then the CT data was imported into MIMICS software,the femur model was extracted in the MIMICS software and the total volume of each femur bone layer was acquired. After CT scanning for 4  times,the left and right rabbit femurs were taken as specimen. The tree-point bending test was performed and the load-deflection curves were obtained. The elastic load and the elastic deflection were contrasted and the maximum bending stress was calculated. Results Compared with normal control group,the deflection and cross-sectional moment of inertia of the right leg of the rabbits in model group had on significant changes in the rabbit bone growth process (P> 0.05); but the growth rate and maximum flexural strength and maximum bending normal stress and cortical bone growth rate and compact bone growth rate were greatly decreased ( P<0.05),and the soft bone reduce rate was　basically consistent( P<0.05). Conclusion The increasing rates of each bone layer and the mechanical properties of the  femur in the rabbits with OA are significantly decreased and the mechanical properties have correlation with each bone layer.
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Abstract:Objective
To investigate the influence of Meconopsis racemosa Maxim.of a traditional Tibetan medicine in the biological activity of human leukemia cell line K562 cells in vitro and  to clarify its related mechanism.Methods The K562 cells were treated with different concentrations (30,60,90,120,150 mg?L-1) of Meconopsis racemosa Maxim.alcohol extract in vitro,meanwhile blank control group was set up.MTT assay was used to detect the growth inhibitory rates of K562 cells treated with Meconopsis racemosa Maxim.alcohol extract.The  morphological changes of K562 cells were observed by inverted microscope.DNA injury was evaluated by single cell gel electrophoresis assay (SCGE).The changes of cell cycle were detected using flow cytometry.Results The inhibitory rates of K562 cells proliferation in Meconopsis racemosa Maxim.alcohol extract groups were significantly higher than those in blank control group  after 24,48 and 72 h(P＜0.01) in dose-dependent and time-dependent manner.The numbers of K562 cells was reduced,and the cells shapes were irregular with cuntour fuzzy under inverted  microscope.The SCGE results showed that nucleus DNA fragmentation and TailDNA% were increased after treated with 60,90,and 120 mg•L^-1 Meconopsis racemosa Maxim.alcohol extract,there were significant differences compared with blank control group(P＜0.05,P＜0.01). The cell cycle detection results showed that in 60,90 and 120 mg•L^-1 Meconopsis racemosa Maxim. alcohol extract groups,the percentages of K562 cells at G0/G1 phase and S  phase were  decreased,the percentages of K562 cells at G2/M phase were increased(1.88%±0.30%,5.46%±0.57%,19.8%±1.03%),they were significantly higher than that in blank control group(1.10%±0.12%)（P＜0.05）,which suggested a G2/M cell cycle arresting in Meconopsis racemosa Maxim.alcohol extract groups.Conclusion Meconopsis racemosa Maxim.alcohol extract could obviously inhibit the proliferation of K562 cells,and the mechanism may be related to inducing DNA injury of K562 cells and arresting cell cycle in G2/ M phase.

Abstract:Objective
To clone and express the nitrite reductase gene  (nir)  from Alcaligenes xylosoxidans and detect the enzyme activity,and to explore the  degration characteristics of the nitrite reductase(NiR) on nitrite.Methods The DNA from Alcaligenes xylosoxidans was purified.The primers were designed according to nir sequences in GenBank.The nir gene was amplified by PCR and cloned into expression vector pET-28a.The recombinant plasmid pET-28a-nir was constructed.After the identification of restriction digestion and sequencing,the positive recombinant plasmid was transferred into E.coli BL21(DE3).The recombinant protein was obtained by induction.The cell fermentation was broken with ultrasonic.The crude enzyme was extracted and the enzyme activity was calculated.Results A 1 083 bp DNA fragment was obtained by PCR.The target gene was induced by IPTG with the final concentration of 0.6 mmol?L-1 at 30℃,and the induction time was 5 h.A kind of recombinant protein with the molecular weight of 37 000 was obtained.Under the condition of pH 6.5 and reaction temperature 35℃,the enzyme activity was 51.74 U•mL^-1.Conclusion Nitrite reductase gene is successfully cloned and expressed in E.coli　BL21(DE3).The expression product has enzyme activity.
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Abstract:Objective To study the differences in efficacy of N- Acety- L- cysteine (NAC) and ipratropium bromide alone or co-administered in rats with chronic obstructive pulmonary disease (COPD),and to clarify the mechanism of co-administration.Methods 30 healthy male Wistar rats were randomly divided into blank control group(n=6),model group(n=6),NAC group(n=6),ipratropium bromide group(n=6) and combination of NAC and ipratropium bromide group(n=6).The rat COPD model was established by intratracheal instillation of lipopolysaccharide ( LPS) for 3 times and exposing to cigarette smoke for 3 months.1 week after modeling,on the eighth day the drug NAC or ipratropium bromide were used for 11 weeks alone or in combination.HE staining was used to detect the morphological changes of lung tissue;ELISA was used to determine the changes of serum interleukin-6(IL-6),nitric oxide (NO),tumor necrosis factor-α(TNF-α) expression;blood-gas analyzer was used to measure the changes of arterial blood oxygen indicators.Results The morphological examination results of lung tissue showed that compared with model group,the lung airway remodeling in drug-treated groups were mitigated,especially in combined treatment group.In expression of inflammatory factors of IL-6,NO,TNF-α,the serum concentration of IL-6,NO,TNF-α of model group was respectively high,compared with blank control group,there is statistical significance（P<0.01）.The serum concentration of IL-6,NO,TNF-α of NAC group and ipratropium bromide group was low,compared with model group,there is statistical significance（P< 0.05）.The results of blood gas analysis showed that the value of PaO2 in model group was decreased,compared with blank control group,there was statistical significance（P<0.01);the value of PaO2 was increased(P<0.01).Compared with model group,the values of PaO2 of the rats in NAC group,ipratropium bromide group and combination group were significantly increased(P<0.05 or P<0.01)，and the values of PaCO2 were decreased significantly（P<0.05 or P<0.01).Compared with blank control group,the serum IL-6,NO and TNF-α levels of the rats in model group were significantly increased(P<0.01);compared with model group,the serum IL-6,NO and TNF-α levels of the rats in NAC group,ipratropium bromide group and combination group were significantly decreased(P<0.05).Conclusion Combination of ipratropium bromide and NAC can prevent lung function of COPD rats from decreasing,and control the progression of early COPD.

Abstract:Objective To investigate the influence of oridonin combined with melphalan in cell proliferation,apoptosis and cell cycle of myeloma RPMI8226 cells and their synergistic effect,and to clarify the cytotoxin enhanced effect of the combination of the two drugs on myeloma cells. Methods The RPMI-8226 cells were divided into blank  control group,oridonin（10  μmol/L）　group,melphalan（30  μmol/L）group and oridonin(10 μmol/L) + melphalan(30 μmol/L) group.The inhibitory rates of proliferation of RPMI-8226 cells were detected by MTT assay after treated with oridonin,melphalan alone or in  combination  at different time.The drug synergistic effect was expressed by coefficient of drug interaction (CDI).The change of cell cycle was analyzed by flow cytometry,and the apoptotic rate was detected by Annexin-Ⅴ/PI staining.Results Compared with blank control group,the inhibitory rates of proliferation of RPMI-8226 in oridonin,melphalan and oridonin+melphalan group were significantly increased(P<0.01) in a time-dependent manner.Compared with single drug groups,the inhibitory rates in oridonin+melphalan group at different time were significantly increased (P<0.01).The CDI values of two drugs at different time were less than 1,which indicated that oridonin could improve the cytotoxicity of melphalan.The apoptotic rate of combination use of two drugs was 43.06%±1.96％,which was significantly higher than single oridonin (15.01%±0.47％) and melphalan (20.03%±1.30％) (P<0.01).Flow cytometry DNA analysis showed that oridonin,melphalan or combination use could induce the arrest of most RPMI-8226 cells at G0/G1 phase,the ratio of RPMI-8226 cells at  S phase was reduced,there was significant difference between drug groups and blank control group (P<0.01),and there also was significant difference between oridonin+melphalan  group and  single drug  groups (P<0.01).Conclusion Oridonin combined with melphalan can synergisticly inhibit the proliferation of RPMI-8226 myeloma cells by arresting cell cycle and inducing apoptosis.

Abstract:Objective To clone,identify,express and purify the S-adenosyl-L-homocysteine hydrolase (sahH) gene of Veillonella dispar and to provide theoretical basis for  the mechanism of dental caries disease. Methods The DNA  exacted from Veillonella dispar sahH gene were amplified by PCR method,and the recombinant plasmid PET28a-sahH was constructed and transformed into E.coil BL21.The sequencing was performed after PCR identification and restriction enzyme digestion.Then the protein expression was induced by IPTG and the protein was purified.Results The PCR product  was 1 410 bp.The sequencing data was analyzed by BLAST software.The homology of the sequencing result was 99 % compared with the sahH gene of Pseudomonas aeruginosa strain PAO1 which was reported in GenBank.The sequence had been submitted to the NCBI GenBank,and the accession number was KC477409.With SDS-PAGE electrophoresis,the recombinant plasmid PET28a-sahH was expressed and purified in  E.coil JM109,and the molecular weight of 50 000 was noted in sahH protein bands.The highest amount of protein expression was generated 5 h after  induction,The protein concentration was 5.4 g/L after concentrated.The size of protein obtained by Western blotting method was corresponded with target  protein.Conclusion The sahH gene of Veillonella dispar is successfully cloned,and it is proved to have a full reading framework through the analysis on gene sequences and amino acids.And the recombinant vector could  express fusion protein,and the interest protein is obtained by isolation and  purification.

Abstract:Objective To study the localization and expressions of somatostatin(SS) and islet amyloid polypeptide(IAPP) in rabbit gastric antrum mucosa during the postnatal development period,and to explore the relationship between these hormones and digestive function.Methods Sixty rabbits were divided into 6 groups according to their age:5 d,15 d,25 d,35 d,60 d,and 90 d;10 rabbits in each group.The rabbits were excuted and their gastric antrum tissues were taken and embedded in routine paraffin,then the  serial sections were made．Immunohistochemical SABC staining and image analysis were performed to detect the expression and  localization of SS and IAPP immunoreactive positive cells.Results The expressions of SS and IAPP were found in the gastric antrum mucosa of rabbits at different postnatal development periods.In  5 d  and 15 d groups,a few SS positive cells were scattered into the mucosa epithelium and the staining  was weaker.From the 35th  day,the number of SS positive cells was ascended and obviously reached a peak in 60 d group （P<0.05）,and the number of SS positive cells in 90 d group was similar with that in 35 d group.The mean grey value of SS positive cells was decreased from the 35th day and reached the lowest in 60 d group （P<0.05）.The positive IAPP positive cells were predominantly located in the connective tissue of gastric antrum mucosa,occasionally in the gastric mucosal epithelium.The number of IAPP positive cells was gradually increased from the 5th day,especially in 60 d group（P<0.05）. The mean grey value of IAPP positive cells was decreased from the 35th day（P<0.05）.Conclusion The expressions of SS and IAPP in gastric antrum mucosa of rabbits change with the growth,which suggests that both of them should participate in the process of the digestive function by endocrine and paracrine ways．They play an important role in the growth repair and self-protection of the gastric mucosa.

Abstract:Objective To study the changes of Bmi-1 and p16 gene expressions in human fetal bone marrow-derived mesenchymal stem cells(MSCs) with the increasing of passage number,and to clarify  the relationship between Bmi-1,p16 gene expressions and replicative senescence of MSCs. Methods By using the adherence culture method,MSCs were isolated from the femoral bone marrow of three abortuses: 16-week-old,17-week-old and 25-week-old,respectively; and then the MSCs were isolated,cultured and amplified. The expressions of CD13,CD34，CD44，CD45，CD73，CD90，CD105，CD133 and HLA-DR in the fifth generation of MSCs were detected with flow cytometry. After successively subcultured,the fetal bone marrow-derived MSCs were induced to replicative senescence. The proliferation  abilities of MSCs at passage 5(P5),passage 15(P15) and passage 30(P30) were detected with BrdU incorporation assays. The expression levels of Bmi-1 and  p16 gene mRNA in MSCs at P5,P15 and P30 were determined with Real-time PCR. Results The MSCs were isolated from the femoral bone marrow of the three abortuses and then cultured successfully,with positive expressions for CD13,CD44, CD73,CD90,CD105 and almost no expressions for CD34,CD45,CD133 and HLA-DR; With the increasing of passage number,the cell growth rate was  gradually slowed down,and the cells became flat,and the refractivity was decreased. The BrdU incorporation rates of  MSCs between P5 and P15 had no significant difference（P>0.05），but the BrdU incorporation rate of  MSCs at P30 was lower than those of MSCs at P5 and P15（P<0.05）. The expression level of Bmi-1 gene mRNA of MSCs at P5 was higher than those of MSCs at P15 and P30 (P<0.05),and the expression level of Bmi-1 gene mRNA of MSCs at P15 was higher than that of MSCs at P30 (P<0.05). The expression levels of p16 gene mRNA of MSCs between P5 and P15 had no significant difference（P>0.05），but the expression level of p16 gene mRNA of MSCs at P30 was lower than those of MSCs at P5 and P15 (P<0.05). Conclusion MSCs can be successfully isolated from the femoral bone marrow of the human fetus. The replicative senescence of fetal bone marrow-derived MSCs has correlation with the mRNA expression level of Bmi-1,but the signal pathway may not mediate through p16INK4a-Rb pathway .

Abstract:Objective To observe the activational effect of   PAX2 transfection on Wnt4 pathway and  the effect of WNT4 gene on PAX2-induced epithelial- mesenchymal transition,and to discuss the mechanism of PAX2 transition.  Methods The pEGFP-PAX2 plasmid was transfected into rat normal renal tubular epithelial cells(NRK52E) with lipofectamine 2000.The cell line NRK52E stable expressing PAX2 was constructed by G418.NRK52E were divided into control group,empty vector group and stable transfection PAX2 group.The expressions of WNT4 and β-catenin were examined by Western blotting and Real-time PCR in various groups.The WNT4 gene siRNA silencing  was stably transfected into WNT4 of  PAX2 cells  and evaluated through fluorescence detection,and the best chain was chosen.After the best WNT4 siRNA chain was transfected into NRK52E,the expressions of β-catenin,E-cadherin and α-SMA were examined by Western blotting and Real time-PCR at 24,48,72, and 96 h after silencing.Results The Western blotting and Real-time PCR results showed that the protein and mRNA expressions of WNT4 and β-catenin  in stable transfection PAX2 group  were significantly increased  compared with control group and empty vector group(P<0.05).The transfection efficiency of negtive siRNA was(47.46±4.48)％.The third WNT4 transfection chain  was the best transfection chain detected by Real-time PCR and Western blotting. The inhibitory rate of WNT4 m RNA was (66.9±3.8)％（P<0.05),and the inhibitory rate of WNT4 protein was （63.7±2.5）％（P<0.05). 24,48,72, and 96 h after WNT4 siRNA transfection,compared with control group,the mRNA and protein expressions of β-catenin were decreased,especially at 72 h(P<0.05);the mRNA and protein expressions of E-cadherin were increased,especially at 72 h(P<0.05);the mRNA and protein expressions of α-SMA were decreased,especially at 72 h(P<0.05).Conclusion PAX2 could activate WNT4/β-catenin pathway.PAX2 may induce epitheial-mesenchymal transition in normal renal tubular epithelial cells through  WNT4/β-catenin pathway.
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Abstract:Objective To establish intrahepatic cholestasis of pregnancy (ICP) models of rats and detect the expression of heme oxygenase-1(HO-1) in liver tissue of pregnant rats with ICP and   brain tissue of  fetal rats,and to explore the  pathogenesis of  ICP.Methods 40 pregnant  rats were randomly divide into experiment group and control group,20 rats in each group. From the 15th day after pregnancy,the  pregnant rats in experiment group were administered with estrogen and progestrone to induce ICP，while  the rats in control group were administered with vegetable oil.At the 21st day after pregnancy the rats were executed and the blood  samples were obtained to detect the biochemical indicators,and the pathologic changes in liver tissue of pregnant rats and brain tissue of  fetal rats were observed by  HE staining; the HO-1 expression in hippocampal CA1 section of  brain tissue of fetal rats was detected by immunohistochemistry  staining and image analysis. Results Compared with control group,the levels of serum alanine aminotransferas
es (ALT), total bile salt (TBA),aspartate amino transaminase(AST) of the pregnant rats in experiment group were significantly increased (P<0.01). The pregnant rats in experiment group showed  granule and vacuolization,natural hepatic foliole structure in liver;and the structural changes such as swelling,vacuolar degeneration,partial cells dissolution,and  disappearance in brain tissue of fetal rats were found;but there was no pathologic changes in control group. The immunohistochemistry staining and image analysis results showed that compared with control group,the number of  HO-1 immunoreaction positive cells in  brain tissue of fetal rats was significantly increased in experiment group,and  the HO-1 expression level in  experiment group was significantly decreased(P<0.05). Conclusion The expression of HO-1 in brain tissue of fetal rats with ICP is decreased,which suggests that HO-1 might be  important to cause fetal anoxia and death.

Abstract:Objective To explore the impact of hepatic stellate cell （HSC）in hepatocellular carcinoma invasion through SDF-1/CXCR4 axis and its mechanism,and to provide basis for research on hepatocellular carcinoma  invasion. Methods The expressions of SDF-1 and CXCR4 in HSC LX02,hepatocellular carcinoma cells(MHCC97,SMMC7721,Hep3B,and HepG2) were examined by Western blotting  method  and RT-PCR method. In addition,Transwell invasion assay was carried out to analyze the influence of HSC LX02 and SDF-1 on invasion of hepatocellular carcinoma cells HepG2 under normal condition or CXCR4 gene silence condition. Then Western blotting method was performed to evaluate the expressions of epithelial marker E-Cadherin protein and mesenchymal marker Vimentin protein. Results Compared with hepatocellular carcinoma cells,SDF-1 was over-expressed in HSC LX02 (P<0.05).CXCR4 highly expressed  in all hepatocellular carcinoma cells. Co-culture with HSC LX02 or treatment with SDF-1 both increased the invasion of hepatocellular carcinoma cells HepG2 (P<0.05),and decreased E-Cadeherin protein expression (P<0.05) and increased Vimentin protein expression (P<0.05). Furthermore,after silence of CXCR4 gene by RNA jamming technique in HepG2 cells,neither co-culture with HSC LX02 nor treatment with SDF-1 had impact in invasion of hepatocellular carcinoma cells(P>0.05),and there were no changes in the E-Cadeherin (P>0.05) and Vimentin (P>0.05) protein expression levels. Conclusion HSC can promote hepatocellular carcinoma cells invasion through chemokine SDF-1/CXCR4 axis,and the mechanism may be realted to epithelial-mesenchymal transition of carcinoma cells.

Abstract:Objective To observe the influence of swimming exercise in learning and memory,the concents of amino acids and nNOS expression in prefrontal cortex (PF) of brain in the aging rats,and to explore the mechanism of swimming exercise to improve learning and memory of the aging rats.Methods Twenty SD aging rats (24 months) were chosen and randomly divided into aging control group(SC) and swimming exercise group(SE),and other ten SD adult rats(5 months) were used as adult control group(control).The rats in control and SC groups kept normal activities and the rats in SE group undertook an 8-week swimming exercise with progressively increasing load.Then an 8-arms radial maze was used to assess the capability of learning and memory of the rats and the contents of glutamic acid (Glu) and gamma aminobutyric acid (GABA) in PF,and the nNOS expression in PF was detected and analyzed by using ABC immunohistochemical technique and semi quantitative method.Results Compared with SC group,the Glu content  in PF of the rats in SE group was significantly increased(P<0.05);the GABA content was also increased,but there was no significant difference (P>0.05);the ratio of Glu/GABA was obviously increased(P<0.05).Compared with SC group,the time of completing 8-arms radial maze  of the rats in SE group was shortened (P<0.05);the reference memory error (RME) and total memory error (TE) were obviously decreased (P<0.05,P<0.01);the working memory error (WME) was decreased,but there was no significant difference(P>0.05).Compared with SC group,the amount and area of nNOS in PF  of the rats in SE group were obviously increased (P<0.05),and the gray degree was enhanced (P<0.05).Conclusion Swimming exercise may enhance the learning and memory  abilities of the aging rats and its mechanism might be that the swimming exercise can increase the content of Glu,effectively adjust the ratio of Glu/GABA and reinforce the expression of nNOS in PF at the same time.

Abstract:Objective To study the effect of Sanshen Weixin Capsula (Sanshen) on cardiac function in rats with heart failure induced by myocardial infarction,and  to provide basis for developing the new drugs to treat heart failure inducedby myocardial infarction. Methods The heart failure model was induced by myocardial infarction in rats with left anterior descending coronary (LAD) occulusion.The Wistar rats with heart failure were divided into four groups after detecting cardiac function by  echocardiography: sham operation group(n=9),heart failure model group(n=8),ramipril group(n=9) andSanshen group(n=9).After 1 week,the rats in sham operation group and model group were treated  with distilled water,and the rats in Ramipril group and Sanshen group were treated with ramipril and Sanshen,respectively.After 5 weeks,the changes of echocardiography and hemodynamics were observed,and the heartorgan index and infarction area were evaluated,and the histomorphogical changes were observed.Results Compared with model group,the left ventricular end-diastolic dimension (LVEDD) and the left ventricular end-systolic dimension (LVESD)in Sanshen group  were decreased significantly(P<0.01)；the left ventricular ejection fraction（LVEF） and the franctional shortening（FS）were increased significantly(P<0.01).Compared with model group,the left ventricular end diastolic pressu(LVEDP) in Sanshen group  was decreased significantly(P<0.05)；the left ventricular systolic pressure（LVSP） and the maximal ventricular pressure rise ratio during systolic period/decrease ratio during diastolic period（±dp/dtmax） were increased significantly (P<0.05 or P<0.01).Compared with model group,the infraction area of left ventricle and heart organ index in Sanshen group were decreased significantly (P<0.05).Compared with model group,the ventricular myocardial cells in Sanshen group arranged in neat rows,the interstitial edema was significantly reduced,the interstitial connective tissue proliferation and inflammatory cell infiltration were decreased,and the myocardial fibers were not significantly enlarged.Conclusion  Sanshen can improve cardiac function in the rats with heart failure induced by myocardial infarction，suggestting that it can be used as a new drug for treatment of heart failure.

Objective To investigate the effects of curcumin on the expression  of connective tissue growth factor(CTGF) in kidney and kidney excretion of the rats with   diabetic nephropathy(DN) rats,and to clarify the protection mechanism of curcumin on the kidney of DN rats.Methods 40male SD rats were  divided into normal control group(n=12) and DN model group(n=28). The diabetic model was induced by intraperitoneal injection of streptozotocin(STZ).24 DN model rats were randomly divided into model group(n=12) and  curcumin treatment group(n=12).The blood sugar,body weight,kidney weight,kidney weight/ body weight,24 h urinary albumin excretion rate(UAER),serum creatinine (Scr),bloodnitrogen（BUN）of the rats were detected 8 weeks after treatment;the pathological changes of the rat kidney tissues was observed by HE staining,and the expression of CTGF protein was determined by immunohistochemistry.Results Compared with model group,the general satus of the rats in curcumin group was significantly improved;the fasting blood glucose was significantly decreased(P<0.01),the body weight of the rats was increased (P<0.01)，and the kidney weight,kidney weight / body weight,24 h UAER,Scr and BUN were significantly reduced(P<0.01);the staining degree of the kidnry tissue was significantly decreased,the expression of CTGF protein was significantly alleviated in curcumin group(P<0.01);glomerular hypertrophy,mesangial cell proliferation and glomerular lesions were significantly alleviated.Conclusion Curcumin can obviously improve the kidney function,and its mechanism may be correlated with inhibition of CTGF expression in kindey.

Objective To investigate the influence of restoration of oncogenic microRNA-21 (miR-21) in CD44 mRNA and   protein expressions,one of the most common cancer stem cell (CSC) markers,in human gastric cancer Kato Ⅲ cells,and to  provide a new theoretical foundation for research on gastric cancer.Methods Human
gastric cancer Kato Ⅲ cells were cultured and divided into blank control group,Kato Ⅲ/miR-21 group and Kato Ⅲ/control group.Hsa-miR-21 mimics and  hsa-miR-21 control were transfected into Kato Ⅲ cells in Kato Ⅲ/miR-21 group and Kato Ⅲ/control group.Then the expressions of CD44 mRNA and protein in the abovecells were detected by  RT-PCR and Western blotting method.Results The relative expression levels of miR-21 in Kato Ⅲ/miR-21 cells was 1.000 00±0.107 41,which was significantly increased compared with Kato Ⅲ/control cells (0.109 91±0.013 99),there was significant difference( P<0.01). The mRNA level of CD44 in Kato /miR-21 cells was 2.342 23±0.223 47,which was significantly  increased compared with Kato Ⅲ/control cells (0.674 58±0.064 53),there was significant difference(P<0.01).The protein level of CD44 in Kato Ⅲ/miR-21 cells was 0.176 048±0.009 900,which was also significantly increased as compared with Kato Ⅲ/control cells (0.045 929±0.007 396),there was significant difference (P<0.01).Conclusion Restoration of miR-21 expression level can induce the over-xpressions of CD44 mRNA and protein in gastric cancer Kato Ⅲ cells,and may induce the transformation of gastric cancer CSCs.

Objective To investigate the targeting effect of recombinant human tumor necrosis factor(rhTNF)combined with cyclophosphamide (CTX) on glioma and to clarify its mechanism.Methods C6 glioma rat models were established and divided into  rhTNF group(n=10),CTX group(n=10) and rhTNF-Dex-CTX group(n=10).The immunofluorescence method was used to observe tumor necrosis factor receptor 1(TNFR1) expression in glioma tissue; radioligand receptor binding analysis  was performed to determine the rhTNF concentration in brain tissue and glioma tissue; ELISA assay was used to detect the CTX levels in brain tissue and glioma tissue; the influence of rhTNF, CTX and rhTNF-Dex-CTX in  survival time of glioma rats was observed.Results The TNFR1 expression in glioma cells was significantly higher than that in tumor vascular endothelial cells.The amount of rhTNF combined with TNFR1 in glioma tissue was significantly higher than that in normal brain tissue (P<0.01).The CTX levels in the brain tissue and glioma tissue of the rats in CTX group were lower; compared with CTX group,the CTX level  in glioma tissue of the rats in rhTNF-Dex-CTX group was significantly increased;compared with CTX and rhTNF groups,the survival time of the  glioma rats in rhTNF-Dex-CTX group was significantly prolonged(P<0.01).Conclusion rhTNF-DexCTX may increase targeting effect of CTX on glioma by combining with TNFR1.

Objective To construct the eukaryotic expression vector of Hath1 gene and to transfect neuroblastoma cells (SH-SY5Y),and to observe its expression in SH-SY5Y cells.Methods DNA was extracted from human whole blood，the Hath1 gene was cloned and the green fluorescent protein(GFP) gfp gene was amplified at the same time; the primers GFP-R and Hath1-F was used to amplify fusion gene gfp-Hath1,and gfp-hath1 was cloned into pMD18-T and digested with XhoⅠ and EcoRⅠ. The recombinant expression plasmid pCDNA3.1 (+) :: gfp-Hath1 was constructed and   transfected into SH-SY5Y cells.Results The gfp-Hath1 fusion gene with 2 023 bp was successfully amplified.The eukaryotic expression plasmid pCDNA3.1 (+) :: gfp-Hath1 was constructed successfully after identified with double enzyme digestion.The GFP was found in SH-SY5Y cells  under immunofluorescence  microscope.Conclusion The Hath1 gene is expressed in SH-SY5Y cells successfully, which lays  a foundation for transfection cochlea experiment and exploring new methods to treat deafness.

Objective To detect  the expression of key factors of toll-like receptor 4 (TLR4) signaling pathway in pancreas and serum of the patients with chronic pancreatitis(CP), and to explore the significance of TLR4 signaling pathway in the occurrence and development of CP. Methods Pancreas tissues from 24 patients with CP and 6 controls with normal pancreatic histology were  collected to detect TLR4 mRNA by in situ hybridization. The MyD88 and TNF-ɑ were detected by immunohistochemistry. Computer iag nalysis was performed to measure integrated optimal density (IA) of TLR4 mRNA, MyD88 and  TNF-ɑ in pancreas sections. The fasting vein blood samples were collected in 84 patients with CP and 30 normal individuals. The serum soluble TLR4 (sTLR4), TNF-ɑ,  IL-6 and IL-12 levels were measured by ELISA. The lipopolysaccharide (LPS) level in plasma was determined by spectrophotometry. The relationships between the plasma PLS level and different cytokines were analyzed. Results In situ 　hybridization analysis showed that the expression levels of TLR4 mRNA, MyD88 and TNF-ɑ in the pancreas tissue of the CP patients were significantly higher than those in  normal control group(P<0.01),and they were 4.2-fold, 6.1-fold and 10.2-fold of the normal controls. The blood LPS, sTLR4, TNF-ɑ, IL-6, and IL-12 levels in the CP patients were significantly increased compared with normal control group(P<0.05 or P<0.01). Bivariate correlation analysis showed that there were significantly positive correlations between the level of plasma LPS and the levels of serum TNF-α, IL-6 and IL-12 in the patients with endotoxemic CP (r=0.859,P<0.01; r=0.688, P<0.01; r=0.539, P<0.05). Conclusion LPS-TLR4 signaling pathway and their response gene products in pancreas tissue and peripheral blood of the   CP patients are up-regulated，and it plays an important role in acute attack stage and inflammation progression of CP.

Objective To investigate the expressions of TPX2 and Aurora A  in oral squamous cell carcinoma(OSCC)tissue and explore the significances of TPX2 and Aurora A expressions in the  treatment and prognosis of oral squamous cell carcinoma.Methods 61 cases of oral squamous cell carcinoma were chosen as experimental group, other 29cases of adjacent tissue of cancer and 61 cases of normal oral mucosa tissue were chosen as control group.　Immunohistochemical staining  was used to detect  the positive expressions  of TPX2 and Aurora A.The percentages of positive cells were observed under  microscope.The correlation between TPX2 and Aurora A protein expressions was analyzed with Spearman analysis.Results The results of immunohistochemical staining showed that the positive expression rates of TPX2 protein in normal mucosa tissue,adjacent tissue of cancer and cancer tissue were 4.9% (3/ 61),51.7% (15/29),and 86.9% (53/61);and there were statistically significant difference among all groups (P<0.05).The expression of TPX2 protein was closely related to the lymph node metastasis of oral squamous cell carcinoma (P<0.05).The positive expression rates of Aurora A protein in normal mucosa tissue,adjacent tissue of cancer and cancer tissue were  3.3% (2/ 61),44.8% (13/29),and 72.1% (44/61);and there were statistically significant difference among all groups (P<0.05).And with the increasing of Aurora A expression level,the malignant  degrees and possibility of lymph node metastasis in oral squamous cell carcinomas were also increased,which had statistically significant difference among groups (P<0.05).The TPX2 protein expression in oral squamous cell carcinoma tissue had positive correlation with the Aurora A protein expression(r=0.445，P<0.01).Conclusion TPX2 and Aurora A may play important roles in promoting the infiltration and metastasis of mucosal epithelium in oral squamous cell carcinoma,and the relationship between the two genes is collabortion and mutual promotion.

Objective To explore the expressions of nuclear factor(NF-κB ) and tumor necrosis factor(TNF-α) protein in  serum and carcinoma tissue of the pati
ents with cervical cancer,and to clarify their biological significance and the values in early combined diagnosis.Methods The blood samples were collected  and colposcope was performed in 100 patients with high-risk factors of cervical lesion (37 cases of cervicitis,39 cases of cervical intraepithlial neoplasia,24 cases of cervical cancer).The ELISA method was used to detect the contents of NF-κB and TNF-α protein in the serum and cervix  tissue.The expression levels of NF-κB and TNF-α mRNA in cervix tissue were determined with RT-PCR assa.The immunohistochemical method was used to detect the expressions of NF-κB and TNF-α protein in cervix tissue;the relationship with clinical and pathological features was analyzed.Results The contents of NF-κB and TNF-α protein in serum and cervix tissue in the patients with cervical intraepithelial neoplasia and cervical cancer were significantly higher than those in the patients with cervicitis(P<0.01).The contents of NF-κB and TNF-α protein in the patients with cervical cancer were significantly higher than those in the patients with cervical intraepithelial neoplasia(P<0.01).The NF-κB and TNF-α  mRNA expression levels and  protein levels in the cervix tissue of the patients with cervicitis were  significantly lower than those of the patients with cervical neoplasia and cervical cancer(P<0.01),and the expression levelsNF-κB and TNF-α mRNA  and NF-κB and TNF-α protein  in cervical tissue of the patients with cervical intraepithelial neoplasia were significantly lower than those of the patients with cervical cancer(P<0.01).The expressions of mRNA and protein of NF-κB and TNF-α showed positive correlation between  the patients with cervicitis,cervical intraepithelial neoplasia and cervical cancer(r=0.69，P=0.037).Conclusion NF-κB and TNF-α can be independent as markers for early diagnosis of cervical precancerous lesions and cervical cancer,and the positive correlation between them can make the combined detection to improve the early diagnosis rate of cervical cancer.

Objective To study the expression of B7-H4 protein in ovarian epithelial tumor tissue and to explore its association with the clinical pathological characteristics of ovarian epithelial tumor  and the relationship between the expressions of CA125 and B7-H4.Methods The immunohistochemistry method was used to detect the expressions of B7-H4 and CA125 in 70 patients with ovarian epithelial carcinoma including 41 cases of ovarian serous cystadenocarcinoma,11 cases of ovarian mucous cystadenocarcinoma,9 cases of ovarian endometrioid carcinoma,9 cases of ovarian clear cell carcinoma,30 cases of ovarian benign epithelial tumor and 10 cases of normal ovary tissue.Results ① The positive expression rate of B7-H4 in  ovarian epithelial carcinoma(84.3%) was significantly higher than that in ovarian benign epithelial tumor (23.3%,P<0.01),and the B7-H4 expression was negative in normal ovary tissue.② The positive expression rates of B7-H4 in ovarian serous cystadenocarcinoma (100.0%),ovarian endometrioid carcinoma (77.8%) and ovarian clear cell carcinoma (88.9%) were significantly higher than that in ovarian mucus cystadenocarcinoma (27.3%,P<0.05).The positive expression rate of B7 - H4 in advanced epithelial ovarian carcinoma group (Ⅲ period and Ⅳ period) (92.9%) was statistically higher than that in early epithelial ovarian carcinoma group  (Ⅰ period and Ⅱ period) (71.4%,P<0.05).④ The positive expression rate of B7-H4 in low differentiation (G3) ovarian epithelial carcinoma group (100%) was statistically higher than that in high-medium differentiation (G1 and G2) ovarian epithelial carcinoma group (67.6%,P<0.01).⑤The positive expression rates of CA125 in ovarian serous cystadenocarcinoma,ovarian mucinous carcinoma,ovarian endometrioid carcinoma,ovarian clear cell carcinoma tissue,excluding ovarian mucous cystadenocarcinoma,were lower than those of B7-H4,and they were positively correlated (r=0.277,P<0.05).Conclusion The positive expression rate of B7-H4 in  ovarian epithelial carcinoma  tissue is related to the pathological type,clinical stage and pathological grade of ovarian epithelial carcinoma.B7-H4 may be regarded as a new marker used for  diagnosis of ovarian cancer.

Abstract:Objective To observe the  changes of expression levels of  hypoxia-inducible factor 1 alpha(HIF-1α)and nuclear factor kappa B(NF-κB) p65 (RelA) in
mononuclear cells  after ascenting of plateau  and their correlations with the incidence
 rate of acute mountain sickness(AMS),and to explore the role and mechanism of
 HIF-1α  and RelA in pathogenesis of AMS.Methods The soldiers’ venous blood before ascentting of plateau and 48 h after arriving Yushu were collected,and the p
rotein and mRNA in mononuclear cells were extracted.The expressions of HIF-1α and RelA mRNA were determined by RT-PCR.The expression of HIF-1α  protein was
determined by Western blotting method.The correlations between the expressions of HIF-1α  and RelA and pathogenesis of  AMS were analyzed with Spearman anal
ysis.Results Compared with before ascentting,after ascentting of plateau the expression levels of  HIF-1α mRNA,RelA mRNA and
 HIF-1α   protein  were increased significantly（P<0.01）.The expression of HIF-1α mRNA was positively correlated with the  RelA mRNA expression
（r=0.806,P<0.01）.At the same time,the expression  of HIF-1α  protein was positively  correlated with the pathogenesis of AMS（r= 0.875,P<0.01）;and the expression levels of HIF-1α  and RelA had no significant difference between women and men（P>0.05).Conclusion
The expression level of HIF-1α  protein is increased significantly after asentting of plateau,and has significantly positive correlation with the pathogenesis of AMS.The expression levels of HIF-1α  and RelA mRNA are significantly increased 48 h after ascentting of plateau,and they have positive correla
tion,prompting that the existence of cross-talk phenomenon between their signal transduction pathways.

Objective To explore the association of low molecular weight protease
( LMP) gene single nucleotide polymorphisms (SNP) with the susceptibility of pulmonary tuberculosis (PTB) of Li population in Hainan Province,and to clarify the LMP gene polymorphism in Li population,and to provide theoretical basis for research on genomics.Methods
Two polymorphicloci of LMP2/LMP7 (Codon60/Codon145/) in 205 cases of Li population  in Hainan with PTB(case group) and 217 healthy Li population in Hainan(control group)  were detected with PCR-RFLP,and the susceptibilities were   analyzed. Results  The polymorphic loci of the LMP gene had polymorphisms in two groups,and LMP7 were C→A,LMP2 were C→T.The genotypic frequencies of  homozygous  Lys/Lys and  heterozygous  Gln/Lys of LMP7 Codon145 polymorphic loci in case group were significantly higher than those in  control group.The genotypic frequencies of  Gln/Gln and homozygous  Lys/Lys were compared,OR
=3.77 (95%CI:1.60-8.89,P<0.05);compared with  Gln/Lys,OR=2.97 (95%CI:1.80-4.90,P<0.01).The genotypic frequency of LMP2 gene polymorphic loci had no significant difference between two groups (P>0.05).The polymorphism of LMP7 gene  was a predisposing factor  for pulmonary tuberculosis infection
  of Li population in Hainan,and the homozygous and heterozygous forms increased the tuberculosis risk for 3.77 and 2.97 times.
Conclusion For the infection of Mycobacterium tuberculosis,LMP7 gene polymorphism is still an important predisposing risk factors.

Objective To investigate the changes of peripheral blood CD4+CD25+
 regulatory T cells in patients with primary pancreatic cancer before and after
treatment of high intensity focused ultrasound(HIFU),and to elucidate the effects of HIFU treatment on immune fun
ction of the patients with primary pancreatic cancer. Methods
Flow cytometry was used to detect the percentage 
of CD4+CD25+ regulatory T cells in peripheral blood of the patients
in pancreatic carcinoma group(n=36) before and 4 weeks after HIFU treatment and healthy persons in control group(n=28).The KPS scores,the levels of CEA
 and CA-199 of the patients with primary pancreatic cancer before and after treatment were compared.
Results The percentage of CD4+CD25+ regulatory T cells of the patients  before HIFU treatment was significantly higher than that in control group,the difference was statistically significant (P<0.01).The percentage of CD4+CD25+ regulatory T cells of the patients in pancreatic cancer group 4 weeks after HIFU treatment was lower than  before treatment,the difference was statistically significant (P<0.01).The KPS score of 36 patients with primary pancreatic cancer after HIFU treatment was significantly increased compared with before treatment,the difference was statistically significant ( P<0.05 ).The
 levels  of CEA and CA-199 of the patients after HIFU treatment was significantly decreased compared with before treatment,the difference was statistically significant (P<0.05).Conclusion HIFU treatment can improve the cell immune function of the patients with pancreatic cancer.
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Abstract:Objective To detect the periodontal status and the serum levels of h
ypersensitivity C reactive protein (hs-CRP) in the patients with chronic
  periodontitis and essential  hypertension,the patients with single chronic per
iodontitis and healthy subjects,and to explore relationship between chronic periodontitis and essential hypertension.
Methods There were 32 patients with chronic periodontitis and essential hypertension，38 patients with chronic
periodontitis and 34 healthy controls.The clinical periodontal parameters including plaque index（PLI）,probing depth(PD),attachment loss(AL) and bleeding inde
x(BI) were examined，and the serum levels of hs-CRP were tested.Results Compared with healthy group and chronic periodontitis group,the PD,AL and BI in chronic periodontitis and essential hypertension group were significantly increased (P<0.05).The hs-CRP level of  the patients in chronic periodontitis and essential hypertension group was significantly higher than those in chronic periodontitis group and healthy group (P<0.05),and the hs-CRP level of the patients  in
chronic periodontitis group was significantly higher than that in healthy group (P<0.05).Conclusion Hypertension might increase the severity of chronic periodontitis,and  chronic periodontitis may increase the level of serum hs-CRP in patients with hypertension.
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Abstract:Objective To observe the expressions of neuritinrotrophic factor
 Neuritin protein and mRNA in human breast cancer tissue and the relationship between
 its expression and the clinicopathological features,and to explore its roles
 in the carcinogenesis and progression of breast cancer.Methods
74 cases of breast cancer tissue samples and 12 cases of normal breast tissue samples were coll
ected.The expressions of Neuritin protein in samples were detected with immunohistochemical assay and Western blotting method;the expression levels of Neuritin mRNA  in samples were detected by quantitative  RT-PCR.
Results The expression rate of Neuritin  in normal breast tissue was 25%，
and the positive expression rate of Neuritin in breast cancer tissue  was 70.62%,which was higher than that in norma
l breast tissue (P<0.05);Neuritin mainly located in the nucleus and cytoplasm.Meantime,there were positive correlations between the positive expression of Neuritin and lymph node metastasis,TNM stages and the positive rate of ER/PR/HER2.Compared with normal breast tissue,the expression levels of Neuritin protein and mRNA in stage Ⅰ-Ⅱ and Ⅲ-Ⅳ breast cancer  tissue were increased (P<0.05 or P<0.01);compared with stage Ⅰ-Ⅱ breast cancer tissue,the expresson leve
ls of Neuritin protein and mRNA in stage Ⅲ-Ⅳ breast cancer tissue was significantly increased(P<0.05).Conclusion Neuritin may be one
 of the genes to enhance the growth of breast cancer and may be a target for the treatment of breast cancer.

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Abstract:Objective To observe the  changes of haemodynamics during peri-extubation period and analepsia process in patients with general anesthesia for t
otal hip replacement after treated with different doses of dexmedetomidine(DEX)，and to provide theoretical basis for its application in clinic.
Methods 80 ASA Ⅰ-Ⅱ patients undergoing total hip replacement with general anesthesia were randomly divided into control group， low，moderate and high doses of DEX groups(n=20).The patients in various groups were pumped with 0，0.5，1.5 and 2.5 μg? kg-1 DEX 30 min before the end of
 operation，and lasted for 10 min. The systolic blood pressure (SBP)，diastolic blood pressure (DBP) ，heart rate (HR)，myocardial oxygen consumption (MCO) of the patients in various groups before administration(T0)，before extubation(T1)， at extubation(T2)，30 min after extubation(T3) were recorded.The wake-up time(t1)， extubation time(t2)， and Observer’s Assessment of Alterness/Sedation Scores (OAA/S)at 30 min after operation(Ta)，35 min after operation(Tb)，and 40 min after operation(Tc) were also recorded.Results Compared with control group，at T2 and T3，there were no statistical differences of the SBP，DBP，HR and MCO in low dose of DEX  group（P>0.05）;the SBP，DBP，HR and MCO in  moderate and high doses of DEX groups
were descended significantly，there were statistically significant differences(P<0.05);but there was no statistical difference between DEX moderate and high d
ose groups（P>0.05）.Compared with control group，the extubation time(t1) and wake-up time(t2) in high dose of DEX  group were prolonged significantly，and there were statistical differences(P<0.05).Compared with control group， the OAA/S of the patients in  high dose  of DEX group was reduced significantly，and there was statistical difference(P<0.05);but there were no statistical differences among the other three groups（P>0.05）.Conclusion
1.5 μg.kg-1  DEX can stablize the haemodynamics indexes of the patients with general anesthesia during extubation，shorten the time of analepsia and e
xtubation，and sustain OAA/S.
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Abstract:Objective To analyze the results of ⁹⁹Tcm-MIBI imaging and parathyroid hormone (PTH) measurement of the patients withsuspected primary hyperparathyroidism (PHPT) and to evaluate the diagnostic value of ⁹⁹Tcm-MIBI imaging combined with PTH measurement in PHPT.
Methods The results of ⁹⁹Tcm-MIBI imaging and serum PTH measurement in 43 patients with suspected PHPT were retrospectively analyzed.The postoperative pathology was used as the golden standard for PHPT.The sensitivities，specificities，positive predictive values， negative predictive values，accuracies，negative likelihood and positive likelihood of ⁹⁹Tcm-MIBI imaging,PTH measurement,99Tcm-MIBI imaging combined with serum PTH
 measurement were compared and evaluated.Results The sensitivities，specificities，positive predictive values， negative predictive values，negative likelihood,positive likelihood and accuracies of ⁹⁹Tcm-MIBI imaging,PTH measurement and  ⁹⁹Tcm-MIBI imaging combined with serum PTH measurement were 93.75％,84.38％,100%; 45.50％,54.55％,36.36%; 83.33％,84.38％,88.89%; 71.4％,54.55％,100%; 1.72,1.85,1.57; 0.14,0.29,0;81.40％,76.74％,83.72%.
Conclusion 99Tcm-MIBI imaging combined with serum PTH measurement can increase the diagnostic accuracy and decrease the rate of missed diagnosis.
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To study the morphology of first maxillary molar main  mesiobuccal(MB) root canal and to observe the clinical effects of cold lateral condensation(CLC) and continuous wave obturation(CW).Methods 80 first maxillary molars were selected in Changchun area;with 8-10# C pioneer file in MB canal,the digital periapical films in buccolingual direction were used to measure the root canal curvature;CT scanning of root canals was performed to measure and calculate the root canal flatness when the root canals were prepared with Mtwo Ni-Ti instruments.The teeth were divided in groups for different curvature and flatness as QWQB type(n=16),QWRB type(n=26),RWQB type(n=14) and RWRB type(n=24).CLC and CW  technique were used separately,and CT scanning of the root canal was performed to calculate the percentage of gutta-percha filled area (PGFA).Results In CLC group,the PGFA was decreased   as the order: QWQB<RWQB<QWRB<RWRB;there was no significant difference   between various groups（P<0.05）；in CW group,there was no significant difference in PGFA of root canal between four groups（P>0.05）.The PGFA of QWQB,QWRB and RWQB root canals in CLC  group were smaller  than those in CW  group（P<0.05）；there was no significant difference in the PGFA of RWRB root canal between CW group and CLC group (P>0.05).The whole PGFA in CW group was better than that in  CLC group（P<0.05）.Conclusion The effect of CW technique is better than that of CLC  in treatment of first maxillary molar root canal.

To restore the extensive posterior defection with all-ceramic onlays made by CEREC3D CAD/CAM system and to explore their instant qualities and short-term assessment.Methods Twenty-four patients with extensive postenor tooth-defection were selected to accept the onlay restoration.All the 24 onlays were scanned and designed in CEREC3D  CAD/CAM system and milled out of feldspathic porcelain blocks,and cemented with composite luting agents.The restorations were recalled and assessed using the modified assessment standard of American Dental Association (ADA) after the operation.The patients were recalled after 3 months and received satisfaction survey.The contents of TNF-α and MMP-8 in gingival crevicular fluid were also checked and compared with  control group.Results None of  the 24 restoration failed during the observation period,the satisfaction rate was 100%.The satisfactory rate of the patients 3 months after operation was 100%.The contents of TNF-α and MMP-8 in gingival crevicular fluid was  close to those in control group,there was no statistical significance (P>0.05).Conclusion The feldspar onlays fabricated with CEREC3D chairside dental CAD/CAM system could gain good result in restoring extensive posterior defection.

Abstract:Objective To study the variety, appearing time and risk factors of side effects after laser photocoagulation for retinopathy of prematurity (ROP) infa
nts and to provide basis for effectively preventing the side effects.Methods 64 ROP infants with stage 3 threshold lesion were selected and divided into side effect group(n=26) and non-side effect group (n=38) according to whether any side effects such as apnea,necrotizing enterocolitis(NEC),sepsis and gastrointestinal hemorrhage accompanied.The WBC,Hgb,PLT,CRP level, screening times, laser points,weight,corrected gestat
ional age and anaesthesia style were compared between two groups. Results 38 (59.4%) infants had no side effect after laser photocoagulation,but 26 (40.6%) infants had side effects including NEC(53.8%), apnea(23.1%),sepsis(15.4%)and gastrointestinal hemorrhage (7.7%);the side effects appeared on the (3.05±1.95) d after  photocoagulation.The WBC,Hgb,PLT,CRP level, laser points and corrected gestationalage  had no significant difference between two groups(P>0.05). However, the screening times in side effect group were more than that in non-side effect group（P<0.05) and the weight was less than in non-side effect group（P<0.05). The rate of general anesthesia in non-side effect group(26.3%) was higher than that in side effect group(15.4%)(P<0.05).
Conclusion Apnea, necrotizing enterocolitis,sepsis and gastrointestinal hemorrhage are the most common side effects  after laser photocoagulation. More screen times and less weight are the risk factors of side effects. General anesthesia could help decrease the side effect of laser photocoagulation.
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Abstract:Objective To explore the most appropriate treatment method for  the  patients with intracranial metastatic tumor，and to provide basis for improving the life quality of the patients and prolonging the living time．Methods 186 cases of intraeranial metastatic tumor in our department form 2000 to 2010 were selected and divided into five groups according to the condition and image data: group Ⅰ (operation + radiotherapy + chemotherapy group) with 81 cases, group Ⅱ (radiotherapy + chemotherapy group) with 66 cases,group Ⅲ (chemotherapy group) with 14 cases, group Ⅳ(radiotherapy group) with 14 cases, Ⅴ group (supportive therapy group) with 11 cases.The nerve compression symptoms and treatment effectiveness of intracranial hypertension and the average survival period of the patients were compared between various groups. The statistics analysis was carried for the patients who were followed up.Results There were significantly differences of nerve compression symptoms and treatment effectiveness of intracranial hypertension and the average survival period between group Ⅰ and group Ⅱ,group Ⅲ,group Ⅴ P<0.05);but there were no significantly differences between group Ⅰ and group Ⅳ(P>0.05).Conclusion Individualized treatment should be used in the patients with intracranial metastatic tumor；operation combined with adiotherapy and chemotherapy is the first choice in the patients with good neurological condition, ollowed by radiotherapy.

Abstract:Objective To compare the clinical application effect and radiation dose of dual axis rotational coronary angiography (XperSwing) and conventional coronary angiography(CCA) technique, and to evaluate the  differences and clinical  values of two kinds of coronary angiography technique.Methods 30 cases induding 20 cases of CCA group and  10 cases of XperSwing group under went coronary angiography were
 collected in clinic; the fluoroscope time, contrast agent usage and the score of image quality of the patients in two groups were analyzed. Using the Chinese anthropomorphic chest phantom, the virtual surgical environment of XperSwing and CCA was simulated. The thermoluminescent dosimeters(TLD) were placed on its major organs,and  the  absorbed doses of different organs were measured and the effective doses were computed. Results Compared with  CCA group, the contrast agent usage was decrea
sed by 27.74%(P<0.05),and the  fluoroscope time and the score of image quality in XperSwing group had no significant difference(P>0.05). The mean radiation dose in XperSwing group was lower than that in CCA group (P<0.05). The effective dose in XperSwing group(4.94 mSv) was lower than that in CCA group(5.57 mSv). Conclusion XperSwing technique can fulfill the image quality requirement  and effectively reduce the contrast agent usage and radiation doseof coronary angiography.
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Objective To clone cholesteryl ester transfer protein (CETP) gene of rabbits and to construct its eukaryotic expression vector,and to study the st
ructure and function of CETP gene,and to lay the foundation for further study on the action mechanism of CETP in atherosclerosis(AS).
Methods Total RNA of rabbit liver tissue  was extracted by  TRIzol.RNA was  reversely transcripted into  cDNA by RT-PCR method.Then the CETP gene was cloned by PCR amplification with LA Taq enzyme and the cDNA was regared as template.The CETP cDNA fragment was
cloned into the pMD19-T vector.The recombinant CETP-pMD19-T was sequenced and alignmented with GenBank sequence.The sec
ondary structure of CETP was analyzed by nnPredict software.Then the CETP cDNA fragment was cloned into the pcDNA3.1 eukaryotic expression vector.And the recombinant plasmid was identified by restriction enzyme and PCR.Results The  sequencing and matching  results showed that  it w
as 98%  corresponded to M27486 gene.The full length of CETP cDNA was 1 659 bp,and the length of coded sequence(CDs zone) was 1
 494 bp,and it encoded 498 amino acids. The secondary structure of CETP was the α-helix structure mainly,its molecular weight was 142 119.4，and pI was 4.91.Conclusion The CETP cDNA is successfully cloned，and the CETP eukaryotic expression vector is constructed and the secondary structure of CETP is forecasted  successfully.

Objective To establish the prokaryotic expression model of IFN-β by constructing the prokaryotic expression vector of recombinant mouse IFN-β and screening the conditions for its inclusion body solubilization,and to provide experimental basis for screening the proper solubilization conditions.Methods DNA was extracted from fresh liver tissue of mice.The prokaryotic expression vector of mouse IFN-β was constructed by traditional methods and was named IFN-β′.Based on the traditional method of solution,the inclusion body dissolving process conditions were improved,such as solution composition pH environment,bacteria and dissolved liquid ratio,and the effect of dissolved temperature  and the dissolution time of inclusion body.Results The mature IFN-β′protein was identical with the predicted results and the induced expression was normal.The changes in pH and DTT  in dissolved liquid had no effects  on inclusion body solubilization.The inclusion body could not be dissolved by washing liquid after liquid washing.Temperature had almost no effect on inclusion body solubilization.When the ratio of dry bacteria to solution was increased from 1 g∶5 mL to 1 g∶20 mL,the IFN-β′protein inc
lusion body could be partially dissolved.Conclusion The recombinant mouse IFN-β prokaryotic expression vector is constructed successfully.The target protein is expressed with the form of inclusion body but not easy to be dissolved,and the  dissolved liquid proportion is the only condition to affect the dissolution of in clusion body protein among the screened inclusion body solubilization conditions.
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