目的 研究C-C亚族趋化因子单核细胞趋化蛋白-4（MCP-4/CCL13）在系统性红斑狼疮（SLE）患者外周血的表达水平，并分析MCP-4的水平与狼疮肾炎的关系，以探讨MCP-4在SLE发病机制中可能起的作用。 方法 选取2007年9月－2010年8月在四川大学华西医院和宜宾市第一人民医院诊断明确的SLE及类风湿关节炎（RA）患者各40例。另收入正常健康对照组（20例），应用酶联免疫吸附试验定量方法测定SLE组、RA患者和正常健康对照者血清中MCP-4的水平，SLE患者根据有无肾脏受累分为非狼疮肾炎组和狼疮肾炎组，其中非狼疮肾炎组20例，狼疮肾炎组20例，并分析SLE组血清MCP-4水平是否与抗核抗体、补体C3、C4等指标及SLE疾病活动指数SLEDAI评分相关性，血清MCP-4水平采用方差分析、LSD-t检验和Spearman相关进行统计分析。 结果 血清MCP-4水平SLE组为（216.32 ± 12.65）pg/mL，RA组为（203.79 ± 18.64）pg/mL，正常健康对照组为（125.13 ± 11.08）pg/mL。SLE组、RA组血清MCP-4水平与正常健康对照组相比均有统计学意义（P＜0.05），SLE组与RA组比较血清MCP-4水平无统计学意义（P＞0.05）；SLE患者中狼疮肾炎组与非狼疮肾炎组比较血清MCP-4水平无统计学意义（P＞0.05）。SLE组血清MCP-4水平与抗核抗体、补体C3、C4等指标及SLEDAI评分无相关性。 结论 MCP-4在SLE组患者血清中表达增高，MCP-4可能参与了SLE的发病过程，可能成为SLE新的血清学有用指标并作为治疗的靶点。
目的:β淀粉样蛋白（β-amyloid precursor protein，β-APP）是已知的参与阿尔茨海默病机制的关键因子。β-APP是否参与难治性癫痫中的病理机制并不清楚。这项研究在于了解β-APP的蛋白在难治性癫痫患者术后颞叶皮质和海马组织中的表达是否异常。方法:免疫荧光法半定量测定难治性癫痫患者术后颞叶皮质和海马组织中的β-APP阳性蛋白的荧光值，并应用统计软件对实验数据进行单因素方差分析。结果:免疫荧光强度值分析结果显示β-APP在耐药性癫痫脑组织中表达较对照组明显增高且有统计学意义。结论:β-APP在难治性癫痫脑组织中异常增高，增高的β-APP可能参与了难治性癫痫的病理机制。
目的:探讨Livin基因在人胆管癌组织及胆管癌细胞系中的表达情况及其与胆管癌发 生发展之间的关系。方法:采用免疫组织化学技术（SP法）检测Livin基因蛋白在45例人胆管 癌标本及及40例癌旁胆管组织、20例正常胆管组织标本中的表达；同时采用RTPCR法及SP 法检测了Livin基因mRNA和蛋白在人胆管癌细胞系QBC939及非肿瘤细胞系HT1080中表达。 结果:Livin在胆管癌组织中表达阳性率为57.8%，而癌旁胆管组织、非癌胆管组织中未能检 测到Livin表达。Livin表达与性别、年龄、肿瘤大小及肿瘤分化程度无关。在有淋巴结转组 中，Livin阳性表达率（70.4%）明显高于无淋巴结转移组（38.9%）。在人胆管癌细胞QBC939 中，Livin mRNA及蛋白均特异性表达，而非肿瘤细胞系HT1080未见Livin表达。结论:Livin 基因在人类胆管癌组织和细胞系中选择性高表达，其可能与胆管癌发生、发展及预后密切相 关。
Objective To explore the pathogenesis of the level of gene and therapeutic target genes associated with intestinal obstruction by analyzing the differential expression gene. Methods The gene expression data that came from public database gene expression omnibus （GEO） which provided adhesion formation’ gene expression data on 1， 3， 7，and 14 days after operation （n=8） and normal intestinal tissues’ gene expression data （n=2） of mouse were collected. The gene function and differential expression of genes were analyzed by using gene ontology （GO） and significance analysis of microarray （SAM）. Results There were a lot of response stimulated up-regulation of gene expression when occurrence of adhesion， and the products of these genes were distributed on cell membrane. The analysis results of gene expression at different time point after operation showed that expression up-regulated of Hmgcs 2 gene occurred on 3-14 days ofter operation and expression up-regulated of Stxbp 5 gene occurred on 14 days ofter operation. Conclusions The adhesion formation may be closely associated with the genes of response to stimulus and the gene product in membrane. The Hmgcs 2 and Stxbp 5 genes may be closely associated with the occurrence of other diseases which induced by adhesion formation.This provides a basis for the discovery of potential therapeutic targets.
Objective To summarize the current advancement of preoperative radiotherapy for rectal cancer. Methods Relevant literatures about current advancement of preoperative radiotherapy for rectal cancer published domesticly and abroad recently were collected and reviewed. Results The lower local recurrence rate and longer disease-free survival time were observed in preoperative radiotherapy， compared with postoperative radiotherapy for rectal cancer. The recurrence rate was higher in short-course radiotherapy， compared with conventionally radiotherapy for stageⅢrectal cancer， but there was no significant difference for stageⅡrectal cancer. The biology molecular such as p53， CEA， Cox-2， EGFR， and VEGF had shown to be radiosensitive. Conclusions The proposal of preoperative radiotherapy for rectal cancer， could be prone to conventionally radiotherapy. There are more screening targets for preoperative radiotherapy in extensive exploration of diverse radiosensitivity. Biology molecular， developed gene expression profiling， and gene chips for rectal cancer may contribute to the individualization treatment.
Objective To study the effect of knockdown of signal transducer and activator of transcription 3 (STAT3) expression by short hairpin RNA (shRNA) on proliferation, apoptosis and invasion of human gastric cancer cell line MKN-45 in vitro . Methods Specific shRNA plasmids to STAT3 were constructed, and then transfected into MKN-45 cells by lipofectamine methods. Cells were divided into three groups: control group, psiRNA-H1 transfected group as negative group and psiRNA-H1/STAT3 transfected group. Semi-quantitative RT-PCR and Western blotting were used to detect the expression of STAT3 mRNA and protein, respectively. Proliferation and apoptosis of the transfected cells were observed by methyl thiazolyl tetrazolium (MTT) method and flow cytometry (FCM), respectively. The invasion of the transfected MKN-45 cells was measured by Boyden chamber. Results Compared with the negative control cells, semi-quantitative RT-PCR and Western blotting showed that the expressions of STAT3 mRNA and protein were down-regulated in the psiRNA-H1/STAT3 transfected group ( P ＜ 0.05) . The subcloned recombinant plasmid expressing shRNA effectively inhibited MKN-45 cell growth and proliferation while empty plasmid had no such specific effect. Cell apoptosis rate increased significantly in psiRNA-H1/STAT3 transfected group （ P ＜ 0.01）, and the invasion of MKN-45 cells was efficiently inhabited in psiRNA-H1/STAT3 transfected group as compared with control group and psiRNA-H1 transfected group( P ＜ 0.01).Conclusion Recombinant plasmid psiRNA-H1/STAT3 shRNA significantly inhibits the proliferation and invasion of MKN-45 cells and promotes their apoptosis.
Objective To establish hepatocellular carcinoma (HCC) cell lines which olig-expressed IGF1R gene stably． Methods An eukaryotic expressing vector pSUPER-IGF1R-siRNA that could block IGF1R expressing was transferred into hepatocellular carcinoma cell lines SMMC7721 and Hep3B with Lipofectamine 2000 reagents． After transferred, cells were selected with G418 to obtain positive clones. The expressions of IGF1R, cyclin D1 and cyclin B1 were detected by RT-PCR and Western-blot． Cell growth curve were painted. Results Two cell lines clones were screened olig-expressing IGF1R gene stably． The experimental cell lines grew more slowly than control cell lines and the expression of cyclin D1 decreased (P＜0.05). Conclusion The HCC cell lines for olig-expressing IGF1R gene stably are established successfully．The plasmid pSUPER-IGF1R-siRNA can inhibit the growth of SMMC7721 and Hep3B cell lines, and the expression of cyclin D1.
Objective To construct a regulatable plasmid containing single chain fusion gene of murine interleukin-12 (mIL-12) which was regulated with mifepristone (RU486) and explore its expression in vitro. Methods The p40 and p35 subunit sequence of mIL-20 were respectively obtained from the plasmid GCp35Ep40PN by polymerase chain reaction (PCR) and they were cloned into pCA14 plasmid after introducing a linker by overlap PCR. The single chain mIL-12 gene was comfirmed by sequencing and subcloned into pRS-17 vector which contains RU486 regulator cassette. The positive clone named pRS-RUmIL-12 was identified by restriction endonuclease digestion and PCR. Lipofectamine 2000 was used to transfect the pRS-RUmIL-12 to HEK293 cells followed by manufacturer’s recommendations. The protein concentration of mIL-12 induced with RU486 in supernatant of the transfected HEK293 cells was measured by ELISA. Results The sequence of single chain mIL-12 what we obtained was the same as the expected result. The results of restriction endonuclease digestion and PCR showed that the RU486-inducible regulatory vector (pRS-RUmIL-12) was successfully constructed. No significant mIL-12 protein concentration in supernatant of HEK293 cells activation was measured without the inducer RU486, whereas higher concentration of the mIL-12 protein was observed in the presence of RU486. The relationship of concentration of the mIL-12 protein and RU486 was positive correlated under definite range. Conclusion A regulatable eukaryotic expression plasmid of mIL-12 single chain fusion gene was constructed, which could be used in the further research of gene regulation and gene therapy.
Objective To construct the responsive plasmid PTRE-HIF-1αof Tet-on gene expression system and examine its expression. Methods RT-nested PCR was performed on the total RNA extracted from hypoxia HepG2 cells to obtain the cDNA of HIF-1α, which was inserted into the responsive plasmid PTRE2hyg. DNA sequencing was performed after the recombinant of responsive plasmid PTRE-HIF-1α was identified by endonuclease digestion. This recombinant vector was transfected into HepG2Tet-on cells by means of liposome and its expression was examined by RT-PCR and Western blot under the control of deoxycycline. Results The amplified products were confirmed as the cDNA of HIF-1α by DNA sequencing. The responsive plasmid PTRE-HIF-1α verified by edonuclease digestion, was capable of expression in HepG2Tet-on cells and could be controlled by deoxycycline. Conclusion The responsive plasmid PTRE-HIF-1α of Tet-on expression system is constructed successfully, and it can express under the regulation of deoxycycline in the HepG2Tet-on cells.