Abstract:Objective:To examine the regulatory effect of corticotrophin-releasing hormone (CRH) on glutamate-mediated current (IGLU) in cultured hippocampal neurons and to study the related mechanism. Methods: Immunofluorescence analysis was used to investigate whether the cultured hippocampal neurons express CRH receptors, and the whole-cell patch-clamp technique was used to examine the direct modulation of CRH on IGLU and the possible intracellular signal pathway. Results: Two minutes’ exposure to CRH obviously depressed IGLU in the neurons in a dose-dependent manner. CRH receptor antagonist α-helical CRH or CRH receptor type 1 (CRHR1) antagonist antalarmin completely blocked CRH-induced depression of IGLU; whereas, CRH receptor type 2 (CRHR2) antagonist astressin-2B failed to block the effects of CRH. Application of the PKC inhibitor G6976 totally blocked the CRH-induced decrease of IGLU. Conclusion: CRH can inhibit IGLU in primary cultured hippocampal neurons, which is mediated by CRHR1 and may involve the PKC signal pathway.

Abstract:Objective:To investigate and compare the influences of Ghrelin and growth hormone releasing peptide 6 (GHRP6) on the contractility of stomach smooth muscle in guinea pigs, and to study the related mechanism.Methods: The myenteric plexuses of gastric fundus and antrium in guinea pigs were stimulated with electrical field stimulation (EFS) to observe the influence of Ghrelin and GHRP6 on the contractility of stomach smooth muscle. The influences of NnitroLarginine (LNNA) and LArginine (LAA) on the effect of Ghrelin and GHRP6 were studied to disclose the mechanism of the effects of Ghrelin and GHRP6. Results: The circular muscle tissues of the gastric fundus generated onrelaxations and offcontractions when stimulating myenteric plexuses with 116 Hz electrical field; the onresponses induced relaxation could be reduced by LNNA and the offcontractions induced contraction could be blocked by atropine and guanethidine. In fundic strips, ghrelin and GHRP6 could decrease the onresponse induced relaxation and increase offresponse induced contraction of the muscle, with the effect of Ghrelin obviously stronger than that of GHRP6.LNNA could increase the effects of Ghrelin and GHRP6induced muscle contraction, and LAA could decrease their effects. In the antral strips, electrical field stimulation of myenteric plexuses led to disappearance of relaxation wave, only leaving offcontractions. Both ghrelin and GHRP6 could increase that contraction.Conclusion: Both ghrelin and GHRP6 can promote the contractility of stomach smooth muscle in guinea pigs through stimulating myenteric plexuses of gastric fundus and antrium, which might be related to the NO pathway.

Abstract:Objective:To observe the immunoreactive changes of P2X3 receptor and examine P2X3 receptor mRNA level in the dorsal root ganglia (DRG) in a rat model of bone cancer pain. Methods: Sixty female Wistar rats, weighing 150-200 g, were randomly allocated to three groups (n=20 each): group CP (cancer pain) received intra-tibial injection of Syngeneic Walker 256 mammary gland carcinoma cells (2×105 ), group S received intra-tibial injection of equal volume 0.9% sodium chloride, and group C received blank control. Mechanical allodynia and thermal hyperalgesia tests were taken on 4, 7, 10, 14, 17, and 21 days after inoculation. On day 14 and 21, ten rats of each group were chosen for either immunohistochemistry or real-time RT-PCR to examine the changes of P2X3 receptor and mRNA. Results: The rats in group CP began to display hyperalgesia on day 10 and the pain became most obvious between day 14 and 21. Immunohistochemistry showed that the proportion of DRG neurons positive of P2X3 receptor in group CP was significantly increased on 14 and 21 d(P<0.05), and RT-PCR showed that the expression of P2X3 receptor mRNA was also significantly increased. Conclusion: These results suggest that our rat model of bone cancer pain has hyperalgesia, which might be related to the increase of P2X3 receptor expression.

Abstract:Objective:To investigate the influence of gender difference on the activation of platelets in septic rats.Methods: Totally 20 female and 20 male SD rats were randomly divided into four groups: male sham-operated group, female sham-operated group, female septic group, and male septic group, with each group containing 10 rats．Sepsis model in the last two groups was produced by cecal ligation and puncture (CLP); rats in the sham-operated group received no CLP．The rats were sacrificed 24 hours after surgery; the blood samples were collected for blood routine test; and the serum β-TG levels were detected by ELISA assay. Serum estrogen (E2) levels were detected by radioimmunoassay.Results: The blood platelet counts were decreased in both septic rats of both genders; the count in female rats was significantly higher than in the male rats (P<0.01). The serum β-TG levels were increased in septic rats of both genders; the level in the female rats was significantly lower than that in the male rats (P<0.01). The blood platelets count in female or male septic rats was significantly positive correlation with the level of serum estrogen and the β-TG expression in female or male septic rats was significantly negative correlation with the level of serum estrogen(P<0.05).Conclusion: There is a gender difference in the activation of platelets in septic rats, and the endogenous estrogen may have protective effect on the platelets of female septic rats.

Abstract:Objective:To observe the expression of CD133 in the clear cell renal cell carcinoma (RCC) and to investigate the related drug resistance.Methods: Flow cytometry and immunohistochemical method were used to examine the expression of CD133 in metastatic RCC cell line RCC05-TXJ, low metastatic RCC cell line RCC05-ZYJ and two clinical non-metastatic RCC primary cultures isolated from a male and a female patient. The four cell lines were treated with IFN-α and 5-FU and the viability of cells were examined by MTT assay before and after treatment.Results: Flow cytometry showed that RCC05-TXJ and RCC05-ZYJ expressed CD133. The in situ carcinomas of male and female patients hardly expressed CD133. Immunohistochemistry examination showed that the local membrane of RCC05-TXJ and RCC05-ZYJ cells expressed CD133. RCC cells of female and male patients hardly expressed CD133. RCC05-TXJ and RCC05-ZYJ cells exposed to IFN-α or 5-FU showed a rebound of survival 24 h after withdrawal of drugs. The survival rates of RCC cells of male and female patients kept at a low level after withdrawal of drugs.Conclusion: There are a small number of CD133+ RCC cells in RCC patients, with a property similar to tumor stem cells, which might be one of the important factors influencing the immunotherapy and chemotherapy of tumors.

Abstract:Objective:To observe the effects of 5-aza-2′-deoxycitydine (5-aza-CdR) on the proliferation and transcription of tumor suppressor gene GSTP1 and RASSF1A in prostate cancer cell line PC3. Methods: The status of 5′CpG island methylation of RASSF1A and GSTP1 genes in PC3 was analyzed by methylation specific PCR (MSP) before treatment with 5-aza-CdR. RASSF1A and GSTP1 mRNA were quantified by real-time PCR during the demethylation process by 5-aza-CdR. MTT assay and flow cytometry were used to examine the proliferative activity of PC3 cells before and after 5-aza-CdR treatment. Results: The 5′CpG island methylation of RASSF1A and GSTP1 genes were detected in human prostate cancer cell line PC3. Compared with control group, RASSF1A and GSTP1 mRNA expression had no significant change 24 h after culture with 5-aza-CdR; their expression was up-regulated 48 h after cultured with 5-aza-CdR, with significant difference found between 5 μmol/L and 10 μmol/L 5-aza-CdR groups. Compared with control group, the expression of RASSF1A and GSTP1 mRNA was significantly increased 72 h after cultured with all concentrations of 5-aza-CdR. MTT assay and cell cycle examination indicated that exposure to 5-aza-CdR for 24 h and 48 h resulted in no obvious growth inhibition and cell cycle change; exposure to 5-aza-CdR for 72 h induced significant growth inhibition (P<0.05) and cell cycle change (P<0.05); and cells were arrested at G0/G1 phase. Conclusion: The 5′CpG island methylation of RASSF1A and GSTP1 genes is probably responsible for RASSF1A and GSTP1 silencing in PC3 cells. 5-aza-CdR can inhibit the proliferation of PC3 cells, disturb the cell cycle, and elevate transcription of GSTP1 and RASSF1A.

Abstract:Objective:To explore whether short hairpin RNA (shRNA) targeting Bcl-2 can enhance the inhibitory effect of methotrexate (MTX) on growth of subcutaneously-transplanted human lymphoma in nude mice. Methods: Recombinant shRNA expression vector targeting the coding region of Bcl-2 mRNA was constructed and preserved in our lab. Human lymphoma Raji cells were injected subcutaneously into 45 nude mice to establish lymphoma models. The polyethylenimine (PEI)/shRNA complex and (or) MTX were injected into tumors. The influence of Bcl-2 shRNA and (or) MTX on tumor growth was observed. The animals were sacrificed 21 days after administration of drugs and the tumors were removed and weighed; the tumor inhibitory rate was calculated. H-E staining was used to observe the pathological morphology of the tumor. The expression of Bcl-2 mRNA in the tumor tissues was examined by RT-PCR. Results: The tumor growth was significantly slower in Bcl-2 shRNA/MTX group than in Bcl-2 shRNA or MTX alone groups (P<0.05). The tumor weight of mice in Bcl-2 shRNA plus MTX group was significantly lower than those in negative shRNA and blank plasmid group (P<0.05). The inhibition rate of tumor growth in Bcl-2 shRNA/MTX was significantly higher than those in the Bcl-2 shRNA or MTX alone groups (P<0.05). H-E staining showed obvious apoptosis and necrosis in Bcl-2 shRNA group and MTX group. RT-PCR result showed that the expression of Bcl-2 mRNA in tumor cell suspension was significantly decreased in Bcl-2 shRNA group (P<0.05), and kept unchanged in the control group. Conclusion: The shRNA targeting Bcl-2 can enhance the inhibitory effect of MTX on the growth of subcutaneously-transplanted human lymphoma in nude mice.

Abstract:Objective:To construct of PSAspecific dendritic cell (DC) vaccine and to observe its in vitro antitumour activity, so as to pave a way for future study. Methods: Bone marrow precursors were isolated and bone marrow derived DCs were prepared. Mature DCs were pulsed by PSA, Lysate of cancer cells, OVA and PBS to yield PSADC, Lys DC, OvaDC, and NonDC, respectively. After primed by antigen, the changes of IL12 p70 and IL1β in the supernatant of dendritic cells were assessed by ELISA. The antigenspecific proliferation and cytotoxicity activity of T cellprimed by PSApulsed DCs were observed and the results were compared with those by Lys, Ova and PBSpulsed DCs. Results: Mature DCs were successfully derived from bone marrow cells with a purity higher than 95%. ELISA assay showed PSADC, LysDC and OvaDC group secreted high levels of IL12 p70 and IL1β than NonDC group (P＜0.05). In addition, PSADCs and LysDCs had significantly stronger ability to stimulate the proliferation of CD4+T cells in 3day classic mixed lymphocyte reaction (MLR) compared with OvaDCs and NonDCs (P＜0.01). Higher levels of IFNγ and IL2 were detected in PSADCs and LysDCs groups compared with the other two groups (P＜0.01), whereas the levels of IL10 and IL4 were lower than the other two groups (P＜0.05). Moreover, PSADCs and Lys DCs enhanced DTH responses of C57BL/6 mice after antigen immunization; the third antigen and control did not show the enhancement effect (P＜0.05). To observe the in vitro antiPSA CTL reactions induced by PSADCs and LysDCs, the LNCaP cell line (PSA specific) was used as syngeneic target and the E.G7 cell line (H2b) was used as Ovaspecific target cells. Compared with OvaDCs and NonDCs, CTL cells induced by PSADCs, LysDCs had significantly enhanced antigenspecific CTL activity to LNCaP cells (P＜0.05).Conclusion: DCbased PSAepitope vaccine can be prepared by pulsing DCs with PSA protein; the prepared vaccine has strong in vitro immune activity and can kill LNCaP cells.

Abstract:Objective:To study the therapeutic effect of alginate-barium sulfate microspheres via transcatheter hepatic arterial infusion in treatment of liver VX2 tumors in rabbits.Methods: A total of 24 rabbits were randomly divided into 3 groups, with 5 rabbits in group A (normal control group); 19 rabbits implanted with liver VX2 tumors were further divided into 2 groups, with 10 rabbits in group B (tumor control group), and 9 in group C (therapy group). Rabbits in group C were catheterized with 3F microcatheter by Seldinger technique for interventional therapy. Spiral CT scanning was performed in group B and group C 14 days after implantation and 14 days after treatment. Liver function tests (TB, ALT, and AST) were performed before and 7 days after treatment. Five rabbits in group B and C were sacrificed to measure the tumor weight and volume; MVD and expression of CD34 and VEGF expression were examined by immunohistochemical technique. Survival periods of the animals were observed and animals were sacrificed 70 days after treatment. Results: Seven days after treatment, the ALT and AST in group B were significantly higher than those in group A and group C (P<0.01). Fourteen days after treatment, the average tumor weights in group C was significantly lower than that in group B (\[2.434±0.992\] g vs \[4.696±1.246\] g, P<0.01); and the tumor volume in group C was also significantly lower than that in group B ( \[2.126±0.929\] cm3 vs \[3.962±1.101\]cm3, P<0.01). Pathological examination showed large necrotic areas in the tumors in group C. CD34 stained necrotic area had no obvious microvessels. The angiogenesis was decreased greatly in the tumor tissues left. Weak VEGF expression was only found in the survived tumor cells. In contrast, group B had abundant cancer cells, large cancerous nests, abundant CD34 positive angiogenesis and strong cytoplasmic staining of VEGF. The survival of rabbits in group C was obviously longer than in group B. Conclusion: Treatment with alginate-barium sulfate microsphere via hepatic artery infusion is safe and feasible for treating liver VX2 tumor. Alginate-barium sulfate microspheres can obviously inhibit tumor growth and have less toxicity to the normal liver tissue.

Abstract:Objective:To investigate the relationship of PPP4R1 gene with the oncogenesis and metastasis of gastric carcinoma by using bioinformatics analysis, and to verify the result by RT-PCR.Methods: IntNetDB was used to search for the neighborhoods of gene PPP4R1 and cliques by CFinder；Chilibot was used to explore the association between the cliques and carcinoma, then the association of PPP4R1 with gastric carcinoma was deduced. Eighteen pairs of primary and metastatic specimens from the same patient and 12 pairs primary carcinoma and the adjacent normal specimens from the same patient were analyzed by RT-PCR for PPP4R1 expression. Results: It was found that the community genes of PPP4R1 included PPP2R5E, MTMR4, PPP3CC, CTDP1, CTDSP1, FLJ22405, PPP1CB, PPP1CC, PPP2R2A, PPP2R5A, PPP2R5C, CTDSP2, and PPP1CA as searched by Cfinder2.0 when k=14. Nine of the genes were associated with carcinoma and 2 associated with carcinoma and metastasis. The result of RT-PCR showed that higher expression of PPP4R1 was found in 15 of the 18 primary gastric cancer specimens compared with the paired metastatic specimens (P<0.01); higher expression was also found in 9 of the 12 adjacent normal specimens compared with the paired cancer specimens (P<0.01). Conclusion: Gene PPP4R1 may be associated with the oncogenesis and metastasis of gastric carcinoma；bioinformatics is an efficient way to investigate function of new genes.

Abstract:Objective:To investigate the distribution of different hepatitis B virus（HBV）genotypes in patients with chronic hepatitis B (CHB) and hepatocellular carcinoma（HCC）, and to analyze the clinical laboratory examination outcomes and pathological characteristics of CHB and HCC by infection with different HBV genotypes.Methods: Totally 89 patients with CHB and 86 patients with HCC were randomly chosen for this study. HBV genotypes were determined by real-time fluorescence quantitative polymerase chain reaction (FQ-PCR) combined with double immunofluorescence staining TaqMan MGB probes. The general information and the laboratory and pathological data of patients were obtained by reviewing of the clinical documentation of patients. Statistical software SPSS10.0 was used to for statistical analyses. A P value less than 0.05 was considered statistically significant.Results: HBV B was dominant in the CHB patients in our group, accounting for 78.65%; the mixed B and C type accounted for 3.37%. HCC patients in our group were dominated by C type (70.93%). There were no other genotypes other than B and C in our group, and there was significant different between their frequency in our group (P<0.001). There were no significant differences in the clinical experimental and pathological parameters in CHB patients with different HBV subtypes.In HCC patients, those with genotype C had higher positive rate of HBV e antigen than those with genotype B（P<0.05）. HCC patients infected with HBV genotype B had larger tumor size (P<0.05). No associations were found between HBV genotypes with TNM stage, vascular invasion, or metastasis.Conclusion: Patients with CHB are dominantly infected with genotype B in our group. HBV genotype C and positive HBV e antigen are risk factors of HCC. Antiviral therapy and promoting e antigen seroconversion may reduce the incidence of HCC. HBV genotype B might be associated with larger tumor size.

Abstract:Objective:To investigate the reliability, validity and sensitivity of Chinese scale of clinical neurological deficit of stroke patients (China Stroke Scale, CSS), so as to assess its clinical application value.Methods: A total of 126 consecutive inpatients with acute stroke onset were included in our study and they were scored by CSS and the United States National Institutes of Health Stroke Scale (NIHSS) score separately; the reliability, validity, and sensitivity of CSS were evaluated. Reliability was evaluated by correlation coefficient r and Cronbach’s α coefficient; construct validity was analyzed by factor analysis method of appraisal; criterion validity was analyzed by the correlation coefficient analysis with NIHSS scale as the criterion. Sensitivity in various fields was assessed through standardization of effect (SES).Results: Totally 123 valid questionnaires were collected. CSS showed high intrarater reliability, interrater reliability (0.911-1.000) and good internal consistency, with the Cronbach’s α>0.8. There was concurrent validity between CSS and NIHSS (r=0.86). The prognosis prediction accuracy of CSS was 92.4%, slightly lower than that of NIHSS (94.1%). Logistic regression showed that CSS’s “gaze function” and “facial paralysis” were not included in the prediction equation. The facial paralysis had a SES of 0.38, all others had a SES higher than 0.5. Most fields showed a good sensitivity.Conclusion: CSS shows an acceptable reliability, validity and sensitivity in patients with stroke, but the predicative validity of CSS is inferior to that of NIHSS, which needs be further revised.

Abstract:Objective:To analyze the association of type 2 diabetes mellitus (DM) with the clinical and coronary angiographic features of coronary heart disease (CHD) in patients aged over 70 years old. Methods: A total of 310 elderly patients with coronary angiographconfirmed coronary diseases, who were treated in Changhai Hospital during Apr. 2006 to Jul. 2008, were retrospectively analyzed. The patients were further divided into 2 subgroups according to the presence of DM: DMCHD group (n=155) and nonDMCHD group (n=155). The age, gender, blood pressure, blood lipid, ejection fraction (EF), the angiographic outcomes, etc. were analyzed and compared between the two groups.Results: The incidence of hypertension was significantly higher in the DMCHD group than in the nonDMCHD group (P<0.05). The fasting blood glucose (FBG), triglyceride (TG), low density lipoprotein cholesterol (LDLC), and fibrinogen (FIB) were significantly higher than those in the other group (P<0.01), and the high density lipoprotein cholesterol(HDLC) was lower than that in the other group (P<0.05).The EF value in the DMCHD group was significantly lower than that in the nonDMCHD group (P<0.01). Among the 35 (22%) patients preliminarily diagnosed as having DM, 14 (40%) would be misdiagnosed if the diagnosis depends solely on FBG without oral glucose tolerance test (OGTT). The prevalence of diffusive coronary lesions in the DMCHD group was significantly higher than that in the nonDM group (P<0.01). DMCHD group also had significantly higher coronary stenosis index (P<0.01) and more occlusive vessels than nonDMCHD group. Conclusion: The missed diagnosis rate of DM is high in CHD patients. Compared with nonDMCHD patients, DMCHD patients are at higher risks for coronary disease and have more severe coronary lesions.

Abstract:Objective:To analyze the changes of the complementarity-determining region 3(CDR3) of T-cell receptor (TCR) beta chain variable region (TCR BV) in peripheral blood mononuclear cells (PBMCs) of patients with pemphigus vulgaris (PV), so as to understand the association of proliferated T cells with autoimmune diseases.Methods: Immunoscope spectratyping technique was used to analyze the distribution of TCR β chain CDR3 in 6 normal blood donors and the dominant CDR3 in the PBMCs in 6 PV patients. Results: The spectratypes of TCR BV subfamily CDR3 region was in a Gaussian distribution manner in all the 6 normal blood donors. The 6 PV patients, however, displayed abnormal proliferation, and oligoclonal expansion of the T cells was observed in TCR BV families with different CDR3 sequences. Conclusion: The abnormal spectratypes of TCR BV subfamily CDR3 region in the peripheral blood of PV patients might be related to the pathogenesis of PV.

Abstract:Objective:To construct a lentiviral vector(LV) of RNA interference (RNAi) targeting HBs gene and to observe its effect on the replication of HBV and expression of antigens.Methods: The effective sequence of siRNA targeting HBs gene was confirmed in our previous study. The complementary DNA containing both sense and antisense Oligo DNA of the targeting sequence was designed, synthesized and cloned into the lentivirus vector（pGCLM-GFP）. The resulting lentivirus vector containing HBs shRNA was named LVshHBs, and it was confirmed by PCR and sequencing. 293T cells were cotransfected with lentivirus vector pGCLM-GFP,pHelper 1.0 and pHelper 2.0. All virus stocks were produced by calcium phosphate-mediated transfection. The titer of virus was tested according to the expression level of GFP. HepG2.2.15 cells were infected with LVshHBs and the supernatant of the cells was subjected to ELISA, Western blotting analysis and HBV DNA quantitative analysis.Results: PCR and DNA sequencing demonstrated that the LVshHBs-producing HBs shRNA was successfully constructed. The titer of concentrated virus was 5×108-2×109 TU /ml. The inhibitory effect was efficient and the corresponding viral transcript and expression of antigens were decreased after infection. The inhibitory effect was observed 4 days after infection and peaked 9 days after the initial treatment with RNAi. Secreted HBsAg was reduced by >70% in cell culture compared with the negative control, which is also confirmed by Western blotting and real-time PCR. After quantification of HBV DNA, the level of DNA relative to the controls was also significantly reduced after RNAi treatment(P<0.05).Conclusion: The lentivirus RNAi vector of HBs has been successfully constructed. The lentiviral microRNA-based RNAi targeting HBs can specifically mediate the down-regulation of HBs expression, inhibiting HBV replication and antigen expression.

Abstract:Objective:To screen for the transcription factor (TF) closely related to the development of cervical cancer tissue by microarrry gene chip and to testify the results at protein level, so as to assess clinico-pathologic significance of these TFs. Methods: The differential genes in 2 fresh cervical cancer specimens and their adjacent normal tissues and 2 normal cervical tissues were examined by Illumine Human-6 gene chip. The expression of the important TFs selected was verified by RT-PCR, and the major TFs regulating the differential genes were selected. The selected TFs were further investigated in 44 cervical cancer tissues, 22 cervical intraepithelial neoplasia(CIN), and 17 normal cervical tissues using immunohistochemical method. The relationship between the expression of major TFs in cervical cancer and the clinico-pathological characteristics was evaluated. Results: There were 67 upregulated genes and 28 downregulated genes in cervical cancer tissue and CIN when compared to normal tissue, these were testified by RT-PCR. Among these genes,the expression of NF-κB p65 in cervical cancer tissues was up-regulated compared with that in the CIN tissues and normal tissues (P<0.05). Expression of NF-κB p65 was positively correlated with the pathological grade of cervical cancer (P<0．05), and was not associated with FIGO stage (P>0．05). The expression of Foxq1 in cervical cancer was significantly lower than that in CIN and normal tissues (P<0．05), and the expression was not correlated with FIGO stage, pathological grade, or NF-κB p65 expression (P>0.05). Conclusion: NF-κB p65 and Foxq1 are associated with the development of cervical cancer; they may play different roles in the oncogenesis of cervical cancer.

Abstract:The brain structural imaging changes in patients with post-traumatic stress disorders (PTSD) and the related research progress are reviewed. It is elucidated that the volume decrease and the density reductions of gray and white matters in PTSD patients are mainly manifested as the imaging changes of the hippocampus, callus, and anterior cingulate cortex (ACC). The imaging change of callus is characteristic in PTSD patients. The image structure analyses of the white matter and gray matter are the current frontiers and strategic trends of PTSD-related research. Meanwhile, it is pointed out that the relationship between PTSD and brain structure imaging is a problem needs to be urgently solved.

Abstract:Multinolular and intrahepatic recurrent HCC can originate from intrahepatic metastasis and multiple origins, and their colnal origin is closely related to the clinical diagnosis and treatment. To designate suitable therapeutic strategies according to their colnal origin is a new challenge needs to be tackled urgently. This paper reviews recent progress in the clinicopathological features, molecular diagnosis and clinical outcomes of multiple origin HCC.

Abstract:CD99, a cell adhesion molecule, is involved in the cell adhesion, apoptosis and tumor cell migration, infiltration, and invasion, and plays an important role in the development and progression of tumor. CD99 is abnormally expression in some tumors, which has potential value in the differential diagnosis, predication of metastasis capability, treatment and predication of tumors.

Abstract:Objective:To assess the value of central venous catheter in treatment of refractory urinary retention in children. Methods: Seven boys with acute urinary retention, who underwent failed routine drainage from Aug. 2007 to Jun. 2008, received paracentric suprapubic cystostomy via disposable central venous catheter. The patients were followed up for 2-11 months and the outcomes and complications were observed. Results: The catheterization processes were successful at the first try, with the mean operation time being (3.2±0.4) min,ranging 2-5 min. The drainage outcomes were satisfactory and there were no complications.Conclusion: Suprapubic puncture and drainage with central venous catheter is safe and effective for children with urinary retention and is worth popularizing.

Abstract:目的:表达纯化可卡因-苯丙胺调节转录肽（CART）及其突变体，观察其对大鼠进食行为的影响。方法:PCR扩增CART活性片段CART55-102 cDNA，构建表达CART55-102原核表达载体，并对其68位和94位半胱氨酸残基突变，构建表达CART55-102（Cys68Ser）和CART55-102（Cys94Ser）原核表达菌株，经IPTG诱导表达后，亲和层析纯化3种小肽；行为学技术检测原核表达小肽对大鼠进食行为的影响。结果:获得的重组CART55-102及其突变体的纯度在95%以上，产物得率约75%；CART55-102可显著抑制饥饿大鼠的进食行为，而68和94位突变体对饥饿大鼠的进食行为无明显影响。结论:原核表达系统可以成功制备高纯度CART活性小肽和突变体，68位和94位突变导致CART抑制进食的作用消失。

Abstract:目的:构建含鼠疱疹病毒侵入介质（herpesvirus entry mediator,HVEM）胞外区和IgG2a Fc段重组表达质粒，真核表达HVEM-Ig融合蛋白，鉴定、纯化，并检测其生物学活性。方法:通过RT-PCR从BALB/c小鼠脾细胞总RNA中扩增HVEM胞外区片段，WT-1杂交瘤细胞总RNA中扩增鼠IgG2a Fc片段，克隆入真核分泌型表达载体pSecTag2A，转染中国仓鼠卵巢细胞（CHO），表达产物用rProtein A凝胶进行亲和层析纯化，获得目的蛋白，并进行SDS-PAGE和Western印迹分析。在单向混合淋巴细胞反应中加入不同浓度的HVEM-Ig融合蛋白作为处理组，同时以加入IgG蛋白的作为阴性对照和未处理的混合淋巴细胞作为空白对照，分别采用\[3H\]-TdR掺入法和ELISA法观察淋巴细胞增殖程度和细胞因子分泌情况。结果:成功构建重组质粒pSecTag2A-HVEM-Ig，表达及纯化HVEM-Ig融合蛋白，并对蛋白的理化性质进行了鉴定。\[3H\]-TdR掺入法和ELISA法结果显示：与对照组相比，该融合蛋白呈剂量依赖性抑制淋巴细胞增殖和IFN-γ、TNF-α、IL-2等细胞因子的分泌(P<0.05或P<0.01)。结论:成功构建了HVEM-Ig融合蛋白真核表达体系，融合蛋白具有预期的抑制淋巴细胞活性及细胞因子分泌的功能，为后续研究该分子在自身免疫和移植免疫反应中的机制奠定了基础。

Abstract:目的:通过比较帕金森病与特发性震颤、头痛患者脑脊液蛋白质谱，筛选和鉴定与帕金森病密切相关的疾病特异性蛋白（disease-specific proteins, DSPs），寻找帕金森病的生物学标志物。方法:以固相pH梯度等电聚焦(IPG-IEF)为第一向，垂直平板十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)为第二向进行脑脊液蛋白质分离；用图像分析软件PDQuest8.0 分析电泳图谱，寻找有意义的差异蛋白点；运用MALDI-TOF-MS质谱鉴定，并对其中差异显著蛋白采用酶联免疫吸附实验（ELISA）进行鉴定。结果:帕金森病与特发性震颤患者及头痛对照组患者脑脊液蛋白质谱的比较，发现3个有意义的差异蛋白质点，其中载脂蛋白E (apolipoprotein E,Apo E)差异显著。ELISA鉴定结果表明，与特发性震颤\[(3.59±1.07) mg/L\]、头痛患者\[(3.67±0.71) mg/L\]相比，PD组患者脑脊液Apo E水平\[(2.47±1.27) mg/L\]明显下调，差异有统计学意义（P＜0.05)。结论:帕金森病患者与特发性震颤患者、头痛对照组患者脑脊液蛋白表达存在差异，这些差异蛋白可能是帕金森病的DSPs，并有可能成为帕金森病诊断的分子标志物。

Abstract:目的:观察栀子浸膏对兔膝骨关节炎模型中关节软骨病理改变及关节液、血清中IL-1β表达的影响。方法:24只大白兔随机分为给药组、模型组和对照组（n=8）。给药组、模型组行Hulth法复制骨关节炎模型，术后第5周，给药组以栀子浸膏600 mg/kg灌胃，模型组与对照组给予等量生理盐水灌胃，给药3周后处死动物，分别对软骨肉眼下行Pelletier评分及光镜下行Mankin评分，取股骨内侧髁关节软骨组织块行电镜、H-E染色观察软骨细胞形态及检测关节液与血清中IL-1β的含量。结果:栀子浸膏能延缓骨关节炎关节软骨病理损伤，给药组关节液与血清中IL-1β的含量明显低于模型组（P<0.05或P<0.01）。结论:栀子浸膏可能通过降低骨关节炎模型关节液与血清炎症因子IL-1β的表达发挥保护关节软骨的作用，从而延缓骨关节炎病程的进展。

Abstract:目的:评价普卢利沙星片治疗呼吸系统和泌尿系统急性细菌性感染的安全性和有效性。方法:采用多中心、随机、双盲双模拟、阳性药平行对照方法。试验组用普卢利沙星片和左氧氟沙星模拟片各2片，2次/d，疗程7～14 d；对照组用左氧氟沙星片和普卢利沙星模拟片各2片，2次/d，疗程7～14 d。结果:本研究共入组265例，可进行疗效评价260例，试验组(普卢利沙星)131例，对照组(左氧氟沙星)129例。试验组和对照组的总有效率分别为89.47%和90.91%，细菌清除率分别为94.06%（95/101）和95.96%（95/99），两组比较差异均无统计学意义（P<0.05）。试验组和对照组不良反应发生率分别为5.26%（7/133）和3.79%（5/132），主要表现为轻中度的恶心、腹泻、头痛、皮疹、ALT升高、BUN升高、Cr升高等，两组比较差异无统计学意义（P<0.05）。结论:普卢利沙星片治疗呼吸系统和泌尿系统急性细菌感染，临床疗效明显，安全性较好。

Abstract:目的:探讨CD44V6在眼附属器淋巴瘤中的表达及意义。方法:采用免疫组化S-P法检测63例眼附属器淋巴瘤中CD44V6的表达情况。其中弥漫性大B细胞淋巴瘤（DLBCL）22例，首次诊断为DLBCL 12例，原发性黏膜相关淋巴组织（MALT）淋巴瘤复发演变为DLBCL 10例；MALT淋巴瘤41例，26例为初发病例，15例为复发病例；另选10例眼附属器淋巴组织反应性增生病例作对照。结果:CD44V6阳性者细胞膜呈棕色。CD44V6在DLBCL的表达率和阳性程度均比MALT淋巴瘤高，差异有统计学意义（P<0.05）；CD44V6在淋巴瘤中的表达率和阳性程度比淋巴组织反应性增生高，差异有统计学意义（P<0.05），但在DLBCL和MALT淋巴瘤各自组内的表达率和阳性程度均无有统计学差异（P>0.05）。CD44V6表达与患者的年龄、性别均无关。结论:在眼附属器淋巴组织异常增生疾病中，CD44V6的表达随病变程度加重而增强，提示其在眼附属器MALT淋巴瘤的发生和发展中起重要作用。

Abstract:目的:以氟尿嘧啶纳米粒(5-Fu-NP)对聚甲基丙烯酸甲酯(PMMA)人工晶状体进行表面修饰,并测定其体外性质。方法:将壳聚糖、聚丙烯酸与氟尿嘧啶通过滴入分散法制备5-Fu-CS-NP。利用载能离子束技术对PMMA人工晶状体进行5-Fu-NP表面修饰。考察人工晶状体表面纳米粒粒径、含药量以及体外释药特性。结果:表面修饰人工晶状体氟离子束剂量为5×1013 ~5×1015ions/cm2,表面分布有大小不等的5-Fu-NP球形颗粒,多数小颗粒粒径在100 nm以下,少数大颗粒粒径主要集中在100~400 nm,含药量为19.55～35.94 μg/枚。5-Fu-NP表面修饰人工晶状体能在4 d内持续释放药物。结论:5-Fu-NP表面修饰人工晶状体缓释作用较好,有望成为可预防后发性白内障的新型人工晶状体。

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