Hepatitis C virus in human B lymphocytes transformed by Epstein-Barr virus in vitro by in situ reverse transcriptase-polymerase chain reaction

AIM: To study persistence and replication of hepatitis C virus (HCV) in patients' peripheral blood mononuclear cells (PBMC) cultured in vitro. METHODS: Epstein Barr virus (EBV) was used to transform the hepatitis C virus from a HCV positive patient to permanent lymphoblastoid cell lines (LCL). Positive and negative HCV RNA strands of the cultured cells and growth media were detected by reverse transcriptase-polymerase chain reaction (RT-PCR) each month. Core and NS5 proteins of HCV were further tested using immunohistochemical SP method and in situ RT-PCR. RESULTS: HCV RNA positive strands were consistently detected the cultured cells for one year. The negative-strand RNA in LCL cells and the positive-strand RNA in supernatants were observed intermittently. Immunohistochemical results medicated expression of HCV NS3 and C proteins in LCL cytoplasm mostly. The positive signal of PCR product was dark blue and mainly localized to the LCL cytoplasm. The RT-PCR signal was eliminated by overnight RNase digestion but not DNase digestion. CONCLUSION: HCV may exist and remain functional in a cultured cell line for a long period.

COMBINED DETECTION OF BREAST CANCER MICROMETASTASES IN THE LYMPH NODES AND BONE MARROW USING REVERSETRANSCRIPTASE CHAIN REACTION AND SOUTHERN HYBRIDIZATION

Objective: The presence of lymph nodes and bone marrow micrometastases of patients with breast carcinoma by immunohistochemistry (IHC) methods has been strongly correlated to early recurrence and shorter overall survival. The aim of this study was to detect micrometastases in matched sample pairs of lymph nodes and the bone marrow of primary breast cancer patients using a more sensitive method, and compare with other clinical parameters. Methods: Cytokeratin 19 (CK-19) gene mRNA expression was detected by reverse transcriptase-polymerase chain reaction (RT-PCR) and Southern blot hybridization. Human breast cancer cell line T47D was mixed with bone marrow cells at different proportions. The positive detection rate was compared among RT-PCR, Southern blotting and IHC methods. Results: Cytokeratin 19 gene was expressed in all 6 positive control samples, while the expression wasn’t seen in 18 negative control samples. CK-19 IHC positive cells were detected at a dilution of one T47D cell in 5×105 bone marrow cells, while the sensitivity detected by PCR and Southern blot hybridization was at 1:5×104 and 1:106, respectively. In the samples from the 35 patients, we found CK-19 positive cells in 2 cases (5.7%) by IHC. CK-19 gene expression signal was detected in 14/35 (40%) by RT-PCR, and 17/35 (48.6%) by southern blotting. Four cases were micrometastases positive both in lymph node and bone marrow (11.4%). There was no correlation between CK-19 detection and other clinical parameters. Conclusion: combined detection of micrometastases in lymph node and bone marrow by RT-PCR and Southern blotting, using CK-19 as a biological marker, is a highly sensitive method for breast cancer.

Detection of SYT-SSX fusion transcripts in paraffin-embedded tissues of synovial sarcoma by reverse transcription-polymerase chain reaction

OBJECTIVE: To assess the feasibility of detecting SYT-SSX fusion transcripts in paraffin-embedded tissues of synovial sarcoma by reverse transcription-polymerase chain reaction (RT-PCR). METHODS: RT-PCR was used to amplify the SYT-SSX fusion transcripts using archival formalin-fixed paraffin-embedded tumor specimens from a series of 37 synovial sarcoma cases. To investigate the specificity of the SYT-SSX fusion transcripts, a variety of non-synovial sarcoma tumors were included in the study as negative controls. The detected messages derived from fusion genes were confirmed by subsequent sequence analysis. RESULTS: SYT-SSX fusion transcripts were detected in 33 of 37 (89.2%) synovial sarcomas. None of the 34 cases of non-synovial sarcoma tumors showed amplified products of SYT-SSX fusion transcripts, although PBGD mRNA was detected in all specimens. Among 33 SYT-SSX-positive synovial sarcomas, 22 tumors had an SYT-SSX 1 fusion transcript, whereas 6 tumors had an SYT-SSX2 fusion transcript. Fusion types can not be distinguished in the remaining 5 cases. There was a significant relationship between SYT-SSX fusion type and histologic subtype. All 10 biphasic synovial sarcomas had the SYT-SSX1 fusion, whereas all tumors with SYT-SSX2 were of monophasic morphology (P

DETECTION OF BREAST CANCER MICROMETASTASES IN BONE MARROW USING REVERSE-TRANSCRIPTASE CHAINREACTION AND SOUTHERN HYBRIDIZATION

Objective: The aim of this study was to detect micrometastases in bone marrow of primary breast cancer patients, and compare with other clinical parameters. Methods: Cytokeratin 19 (CK-19) gene mRNA expression was detected by reverse transcriptase-polymerase chain reaction (RT-PCR) and Southern blot hybridization. Human breast cancer cell line T47D was mixed with bone marrow cells in different proportions. The positive detection rate was compared among RT-PCR, Southern blotting and immunohistochemistry (IHC) methods. Results: Cytokeratin 19 gene was expressed in all 6 positive control samples while the expression was not seen in 8 negative control samples. In all 54 patients 14 cases were CK-19 positive (25.9%) by RT-PCR, another positive signal was obtained in 5/54 (9.3%) of bone marrow samples by Southern blotting. The total positive cases are 19/54 (35.2%). CK-19 IHC+ cells were detected at a dilution of one T47D cell in 5×104 bone marrow cells, while the sensitivity detected by PCR and Southern blot hybridization was at 1∶5×105 and 1∶1×106, respectively. This demonstrates that RT-PCR and Southern blotting was at least 20 times more sensitive than the IHC method. The micrometastases positive rate of the larger tumor size group (>5.0 cm) was significantly (P<0.05) greater than that of the smaller tumor size group (0–2.0 cm). Conclusion: detection of micrometastases in bone marrow by RT-PCR and Southern blotting, using CK-19 as a biological marker, is highly sensitive and it is a method to be used for anticipating the prognosis of breast cancer patients.

Epidemiological and Subtype Characterization of Influenza Viruses Infection in Children in Shenzhen, China during Three Consecutive Seasons (January 2016-December 2018)

Background: Children with seasonal influenza infection cause a significant burden of disease each year in the pediatric clinic. Influenza A and B viruses are the major types responsible for illness. A better understanding of the periodicity facilitates the prevention and control of influenza in children. Objective: This study aims to analyze the epidemiological patterns and subtype characterization of influenza viruses among children in Shenzhen, China. Methods: Influenza samples were collected by nasopharyngeal swabs from influenza like illness patients in Shenzhen Children’s Hospital from January 2016 to December 2018. The positive cases and influenza subtypes were determined by gold labeled antigen detection and reverse transcriptase polymerase chain reaction. The influenza periodicity and age, subtype distribution as well as the association between climate parameters and different influenza subtypes were analyzed by SPSS 22.0. Results: The influenza positive rate during 2016-2018 was 21.0%, with a highest positive rate in the year 2018. The positive rate varied by month, season, and year describing a sequence of peaks presenting primarily in all year including spring, summer and winter. The characteristics of influenza peak were different in each year, with a spring peak in 2016 and a summer plus a winter-spring peaks in 2017 and 2018. In addition, influenza B exhibited a winter-spring seasonal pattern while influenza A displayed a more variable seasonality, highlighting influenza B rather than influenza A which had a negative association with climate parameters. Influenza-positive cases were older than influenza-negative cases (P P Conclusion: Influenza activity in children from Shenzhen typically displays both winter-spring and summer peaks. Influenza A epidemic occurred separately or co-circulated with influenza B, with a winter-spring pattern for influenza B and a much more variable seasonality for influenza A. Influenza B had a negative association with climate parameters. In addition, hospitalization with influenza often occurs in younger individuals infected with influenza A.

SEQUENCE ANALYSIS OF THE NS5 REGION OF GBVC/HGV AND DETECTION OF THE VIRUS BY REVERSE TRANSCRIPTASE PCR

GBV C/HGV is a newly identified virus associated with human hepatitis In this study, the nucleotide sequences of the partial NS5 gene of GBV C/HGV derived from sera of 8 Chinese patients were determined The nucleotide homology among the 8 isolates were 92% on average On the basis of sequence analysis, two sets of oligonucleotide primers derived from highly conserved region of GBV C/HGV NS5 gene were designed to establish both sensitive and specific nested PCR for detection of GBV C/HGV RNA 253 Chinese patients were examined for the virus RNA GBV C/HGV RNA positive rates in patients infected with HBV, HCV and patients with chronic non B,non C hepatitis were 18 4%, 19 8% and 8 9% respectively This result suggested that HBV,HCV and GBV C/HGV shared the same transmission risk factors 8 patients with GBV C/HGV and HCV coinfection were retrospectively observed for the response to interferon Coinfection with GBV/HGV did not negatively influence the responsiveness of HCV, and GBV C/HGV was sensitive to interferon to a certain degree

TT virus and hepatitis G virus infections in Korean blood donors and patients with chronic liver disease

AIM:To determine the prevalences of TTV and HGV infections among blood donors and patients with chronic liver disease in Korea,to investigate the association of TTV and HGV infections with blood transfusion,and to assess the correlation between TTV and HGV viremia and hepatic damage. METHODS:A total of 391 serum samples were examined in this study.Samples were obtained from healthy blood donors(n=110),hepatitis B surface antigen(HBsAg)-positive donors(n=112),anti-hepatitis C virus(anti-HCV)-positive donors(n=69),patients with type B chronic liver disease (n=81),and patients with type C chronic liver disease(n=19). Trv DNA was detected using the hemi-nested PCR.HGV RNA was tested using RT-PCR.A history of blood transfusion and serum levels of alanine aminotransferase(ALT)and aspartate aminotransferase(AST)were also determined. RESULTS:TTV DNA was detected in 8.2%of healthy blood donors,16.1%of HBsAg-positive donors,20.3%of anti- HCV-positive donors,21.0%of patients with type B chronic liver disease,and 21.1%of patients with type C chronic liver disease.HGV RNA was detected in 1.8%of healthy blood donors,1.8%of HBsAg-positive donors,17.4%of anti-HCV-positive donors,13.6%of patients with type B chronic liver disease,and 10.5%of patients with type C chronic liver disease.The prevalence of TTV and HGV infections in HBV- or HCV-positive donors and patients was significantly higher than in healthy blood donors(P<0.05), except for the detection rate of HGV in HBsAg-positive donors which was the same as for healthy donors.There was a history of transfusion in 66.7%of TTV DNA-positive patients and 76.9%of HGV RNA-positive patients(P<0.05).No significant increase in serum ALT and AST was detected in the TTV or HGV-positive donors and patients. CONCLUSION:TTV and HGV infections are more frequently found in donors and patients infected with HBV or HCV than in healthy blood donors.However,there is no significant association between TTV or HGV infections and liver injury.

Comparison of gene expression profiles between primary tumor and metastatic lesions in gastric cancer patients using laser microdissection and cDNA microarray

AIM. To study the differential gene expression profiles of target cells in primary gastric cancer and its metastatic lymph nodes using laser microdissection （LMD） in combination with cDNA microarray. METHODS： Normal gastric tissue samples from 30 healthy individuals, 36 cancer tissue samples from primary gastric carcinoma and lymph node metastasis tissue samples from 58 patients during gastric cancer resection were obtained using LMD in combination with cDNA microarray independently. After P27-based amplification, aRNA from 36 of 58 patients （group 1） with lymph node metastasis and metastatic tissue specimens from the remaining 22 patients （group 2） were applied to cDNA microarray. Semiquantitative reverse transcriptase polymerase chain reaction （RT-PCR） and imrnunohistochemical assay verified the results of microarray in group 2 and further identified genes differentially expressed in the progression of gastric cancer. RESULTS： The expression of 10 genes was up-regulated while the expression of 15 genes was down-regulated in 22 gastric carcinoma samples compared with that of genes in the normal controls. The results were confirmed at the level of mRNA and protein, and suggested that four genes （OPCML, RNASE1, YES1 and ACK1） could play a key role in the tumorigenesis and metastasis of gastric cancer. The expression pattern of 3 genes （OPCML, RNASE1 and YES1） was similar to tumor suppressor genes. For example, the expression level of these genes was the highest in normal gastric epithelium, which was decreased in primary carcinoma, and further decreased in metastatic lymph nodes. On the contrary, the expression pattern of gene ACK1 was similar to that of oncogene. Four genes were further identified as differentially expressed genes in the majority of the cases in the progression of gastric cancer. CONCLUSION： LMD in combination with cDNA microaro ray provides a unique support foe the identification of early expression profiles of differential genes and the expression pattern of 3 genes （OPCML, RNASE1 and YES1） associated with the progression of gastric cancer. Further study is needed to reveal the molecular mechanism of lymph node metastasis in patients with gastric cancer.

Cytopathological evaluations combined RNA and protein analyses on defined cell regions using single frozen tissue block

The co-existence of multiple cell components in tissue samples is the main obstacle for precise molecular evaluation on defined cell types. Based on morphological examination, we developed an efficient approach for paralleled RNA and protein isolations from an identical histological region in frozen tissue section.The RNA and protein samples prepared were sufficient for RT-PCR and Western blot analyses, and the results obtained were well coincident each other as well as with the corresponding parameters revealed from immunohistochemical examinations. By this way, the sampling problem caused by cell-cross contamination can be largely avoided, committing the experimental data more specific to a defined cell type. These novel methods thus allow us to use single tissue block for a comprehensive study by integration of conventional cytological evaluations with nucleic acid and protein analyses.

Interleukin-1 beta up-regulates tissue inhibitor of matrix metalloproteinase-1 mRNA and phosphorylation of c-jun N-terminal kinase and p38 in hepatic stellate cells

AIM： To study the relationship between interleukin-lbeta （IL-1β） up-regulating tissue inhibitor of matrix metalloproteinase-1 （TIMMP-1） mRNA expression and phosphorylation of both c-jun N-terminal kinase （INK） and p38 in rat heffatic stellate cells （HSC）. METHODS： RT-PCR was performed to measure the expression of TIMMP-1 mRNA in rat HSC. Western blot was performed to measure IL-1β-induced JNK and p38 activities in rat HSC. RESULTS： TIMMP-1 mRNA expression （1.191± 0.079） was much higher after treatment with IL-1β （10 ng/mL） for 24 h than in control group （0.545±0.091） （P〈0.01）. IL-1β activated INK and p38 in a time-dependent manner. After stimulation with IL-1β for 0, 5, 15, 30, 60 and 120 min, the INK activity was 0.982±0.299, 1.501±0.720, 2.133±0.882, 3.360±0.452, 2.181±0.789, and 1.385 ± 0.368, respectively. There was a significant difference in JNK activity at 15 min （P〈 0.01）, 30 min （P〈 0.01） and 60 min （P〈0.01） in comparison to that at 0 min. The p38 activity was 1.061±0.310, 2.050±0.863, 2.380±0.573, 2.973±0.953, 2.421±0.793, and 1.755 ± 0.433 at the 6 time points （0, 5, 15, 30, 60 and 120 min） respectively. There was a significant difference in p38 activity at 5 min （P〈0.05）, 15 min （P〈0.01）, 30 min （P〈0.01） and 60 min （P〈0.01） compared to that at 0 min. TIMMP-1 mRNA expression trended to decrease in 3 groups pretreated with different concentrations of SP600125 （10 μmol/L, 1.022±0.113; 20 μmol/L, 0.869±0.070; 40 μmol/L, 0.666±0.123）. Their decreases were all significant （P〈0.05, P〈0.01, P〈0.01） in comparison to control group （without SP600125 treatment, 1.163±0.107）. In the other 3 groups pretreated with different concentrations of SB203580 （10 μmol/L, 1.507±0.099; 20 μmol/L, 1.698±0.107; 40 μmol/L, 1.857±0.054）, the expression of TIMMP-1 mRNA increased. Their levels were higher than those in the control group （without SB203580 treatment, 1.027 ± 0.061） with a significant statistical significance （P〈 0.01）. CONCLUSION： IL-1β has a direct action on hepatic fibrosis by up-regulating TIMMP-1 mRNA expression in ratessionin in rate HSC.JNK and p38 mitogen-activated protein kinases （MAPKs） are involved in IL-1β-induced TIMMP-1 gene expression, and play a distinct role in this process, indicating that p38 and .INK pathways cooperatively mediate TIMP-1 mRNA expression in rat HSC.

Expressions of ICAM-1 and its mRNA in sera and tissues of patients with hepatocellular carcinoma

INTRODUCTIONThe increased expression of ICAM-1 on a widerange of cells and in the sera of patients withmalignancies, chronic liver diseases andinflammation diseases has been described since thelate 1980s[1-22]. Recently rapid progress in studieson expression of ICAM-1 in patients withhepatocellular carcinoma ( HCC ) have beenachieved, including clinical and experimentalresearches[23-31].

Impact of endoscopically minimal involvement on IL-8 mRNA expression in esophageal mucosa of patients with non-erosive reflux disease

AIM:Little has been known about the pathogenesis of non- erosive reflux disease(NERD).Recent studies have implicated interleukin 8(IL-8)in the development and progression of gastroesophgeal reflux disease(GERD).The purpose of this study was to determine IL-8 RNA expression levels in NERD patients with or without subtle mucosal changes. METHODS:We studied 26 patients with NERD and 13 asymptomatic controls.Biopsy sample was taken from the esophagus 3 cm above the gastroesophageal junction and snap frozen for measurement of IL-8 mRNA levels by real-time quantitative polymerase chain reaction(PCR).We also examined mRNA expression of IL-8 receptors,CXCR-1 and -2 by reverse transcriptase PCR.The patients were endoscopically classified into grade M(mucosal color changes without visible mucosal break)and N(neither minimal involvement nor mucosal break)of the modified Los Angeles classification. RESULTS:The relative IL-8 mRNA expression levels were significantly higher in esophageal mucosa of NERD patients than those in esophageal mucosa of the controls.There was a significant difference in IL-8 mRNA levels between grades M and N.The CXCR-1 and -2 mRNAs were constitutively expressed in esophageal mucosa.CONCLUSION: Our results suggest that high IL-8 levels in esophageal mucosa may be involved in the pathogenesis of NERD through interaction with its receptors. NERD seems to be composed of a heterogeneous population in terms of not only endoscopically minimal involvement but also immune and inflammatory processes.

喉癌前病变及喉癌中Livin表达的意义及其与Caspase-3表达的关系

喉癌是呼吸道上皮常见肿瘤,正常上皮由增生开始发展为恶性肿瘤,一般要经过一个量变到质变的过程,即由正常上皮→不典型增生→原位癌,其中包括一个癌前病变阶段,进年来癌前病变的研究已渐成为癌症研究的重点。喉癌前病变包括：喉白斑病、喉厚皮病、成人喉乳头状瘤、慢性肥厚性喉炎。

Rho GTPases途径在非小细胞肺癌中的表达及其临床意义

目的:探讨Rho GTPases在非小细胞肺癌(non-small cell lung cancer,NSCLC)中的表达及其临床意义。方法:应用RT-PCR和免疫组织化学法检测36例NSCLC组织Rho GTPases信号转导途径中的RhoC、E-钙黏附素、基质金属蛋白酶-2(matrix metalloproteinases-2,MMP-2)和MMP-9的表达。通过绘制Kaplan-Meier生存曲线分析RhoC mRNA的表达与患者预后的关系。结果:RhoC mRNA在NSCLC组织和癌旁正常组织中的表达存在差异(P<0.01),RhoC mRNA的表达与患者的性别、年龄、肿瘤浸润程度、淋巴结转移、肿瘤大小、组织学类型和分化程度无关,而与不同TNM分期有关(P<0.05)。RhoC蛋白与MMP-2蛋白的表达呈正相关(r=0.474,P=0.003)。RhoC mRNA低表达组患者的生存时间长于高表达组患者,但2者之间差异无统计学意义。结论:RhoC mRNA的高表达可能与NSCLC的发生和发展有关,也可能与NSCLC的早期及中期浸润和转移有关。

新型冠状病毒Omicron I1566V突变位点MS2噬菌体病毒样颗粒的构建

目的构建新型冠状病毒Omicron I1566V突变位点MS2噬菌体病毒样颗粒。方法选取并合成带有6×His标签的MS2噬菌体的衣壳蛋白(CP)、成熟蛋白(A蛋白)及Omicron I1566V突变基因序列,插入pACYCDuet-1质粒构建重组载体,通过原核系统诱导表达目的蛋白,纯化重组蛋白后利用透射电镜对蛋白质进行物理表征,最后通过反转录聚合酶链反应(RT-PCR)检测病毒样颗粒的热稳定性及耐核酸酶水解能力。结果成功构建包含有6×His标签的CP、A蛋白和Omicron I1566V突变基因序列的重组载体,经限制性内切酶BamHⅠ和KpnⅠ酶切鉴定和测序验证,结果均与预期相符。经诱导并纯化后,通过电镜观察到了大小均匀、直径为23~28 nm的病毒样颗粒,该病毒样颗粒经核酸酶消化后可在37℃条件下稳定储存20 d以上。结论该研究成功利用MS2噬菌体的CP和A蛋白构建了Omicron I1566V突变位点病毒样颗粒,为该突变位点的RT-PCR检测体系提供了可靠的质量保障。

肝内嵌合Sertoli细胞对肝组织中Fas/FasL表达的影响及其意义

目的 :探讨大鼠肝内嵌合Sertoli细胞对肝组织中Fas/FasL表达的影响及其意义 .方法 :将雄性SD大鼠的睾丸Sertoli细胞以不同浓度 (1× 10 5·mL-1、1× 10 6·mL-1,1×10 7·mL-1,1× 10 8·mL-1)和培养 4 8h的上清液 ,经门静脉向每只雌性SD大鼠肝内注入 1mL .2wk后取肝组织 ,应用半定量RT PCR(QRT PCR)的方法检测各组肝组织中Fas/FasLmRNA相对含量 ,与正常肝组织比较 .结果 :正常、上清液组及 1× 10 5·mL-1,1× 10 6·mL-1,1× 10 7·mL-1,1× 10 8·mL-1浓度组肝组织中FasmRNA表达相对量分别为 :0 .77±0 .0 5 ,0 .78± 0 .0 4 ,0 .78± 0 .0 4 ,0 .78± 0 .0 4 ,0 .81± 0 .0 3,0 .80± 0 .0 4 ,FasLmRNA表达相对量分别为 :0 .18± 0 .0 4 ,0 .2 0± 0 .0 4、0 .2 4± 0 .0 4 ,0 .2 4± 0 .0 4 ,0 .4 3± 0 .0 4 ,0 .4 3± 0 .0 4 .各组间FasmRNA表达水平无显著差异 ;FasLmRNA相对含量 ,正常、上清液组与 1× 10 5·mL-1和 1× 10 6·mL-1组比较有显著差异 (P <0 .0 5 ) ,与 1× 10 7·mL-1和 1× 10 8·mL-1组比较相差非常显著 (P <0 .0 1) .结论 :Sertoli细胞经门静脉肝内注入 。

雷公藤红素对内皮细胞E-selectin和ICAM-1及CD31表达的实验研究

目的通过研究雷公藤红素对细胞因子表达的影响探讨其体外抑制血管生成的机制。方法应用RT-PCR检测一定浓度雷公藤红素作用下的内皮细胞株ECV304 E-selectin和ICAM-1的表达,采用流式细胞仪检测不同浓度雷公藤红素作用下内皮细胞株ECV304表面CD31的表达。结果RT-PCR的结果表明,经过雷公藤红素处理后,内皮细胞株ECV304 ICAM-1的表达明显地降低,而E-selectin和CD31并未见表达。结论雷公藤红素在体外可能通过影响内皮细胞粘附分子ICAM-1等的表达,从而抑制血管生成。

Expression pattern of matrix metalloproteinases-13 in a rat model of alcoholic liver fibrosis

BACKGROUND: Interstitial collagenase has been considered as an essential enzyme for collagenolysis in liver fi-brosis, because type Ⅰ and Ⅲ collagens increase predominantly in liver fibrosis. The present study aimed to demonstrate the gene expression of matrix metalloproteinases-13 (MMP-13) in the progressive phases of ethanol induced experimental liver fibrosis in rats. METHODS: Thirty-four Sprague-Dawley rats were randomly divided into two groups. The experimental group (24 rats) was given ethanol (44% , 7 g/kg) every day and the control group (10) was given normal saline. Liver samples were harvested from experimental rats at 4, 12 and 24 weeks respectively. The kinetics of MMP-13 mRNA expression was assayed by semi-quantity reverse transcriptase-poly-merase chain reaction (RT-PCR). RESULTS: In normal rat liver, a faint band for MMP-13 mRNA was observed by RT-PCR (0.24±0.41). The gene expression of MMP-13 was increased in the liver of the rats treated with ethanol for 4 weeks (0. 62 ±0. 54), but it was not considered statistically significant (P >0.05). And the livers from 12-week-treated rats showed a marked mRNA expression(1.65 ±0.47, P <0. 01). Once fibrosis became prominent (24 weeks), a faint band of MMP-13 mRNA was observed (0.39±0.25). CONCLUSION: MMP-13 participates in the degradation of newly-formed matrix in the early phase of rat liver fibrosis induced by ethanol, and it was induced in a distinct time frame.

Quantified gene expression levels for phase Ⅰ/Ⅱ metabolizing enzyme and estrogen receptor levels in benign prostate from cohorts designated as high-risk （UK） versus low-risk （India）for adenocarcinoma at this organ site：a preliminary study

Risk of clinically significant prostate adenocarcinoma （CAP） varies worldwide,although there is a uniform prevalence of latent disease. A hormone-responsive tissue,the prostate possesses the metabolizing capacity to biotransform a variety of environmental procarcinogens or endogenous hormones. Whether such metabolizing capacity or estrogen receptor （ER） status underlies these demographic differences in susceptibility to CaP remains unclear. With appropriate ethical permission,verified-benign tissues were obtained following transurethral resection of the prostate from a high-risk region （n = 12 UK-resident Caucasians） and a typically low-risk region （n = 14 India-resident Asians）. Quantitative gene expression analysis was employed for cytochrome P450 （CYP）1B1,N-acetyltransferase （NAT）1,NAT2,catechol-O-methyl transferase （ COMT）,sulfotransferase （ SULT） 1A1,ERα,ERβ and aromatase （CYP To quantify the presence or absence of CYP1B1,ERα or ERβ,and to identify ther in situ localization,immunohistochemistry was carried out. The two cohorts had reasonably well-matched serum levels of prostate-specific antigen or hormones. Expression levels for the candidate genes investigated were similar.However,clear differences in protein levels for CYP1B1 and ERβ were noted. Staining for CYP1B1 tended to be nuclear-associated in the basal glandular epithelial cells,and in UK-resident Caucasian tissues was present at a higher （P = 0.006） level compared with that from India-resident Asians. In contrast,a higher level of positive ERβ staining was noted in prostates from India-resident Asians. These study findings point to differences in metabolizing capacity and ER status in benign prostate tissues that might modulate susceptibility to the emergence of clinically significant CaP in demographically distinct populations.

小鼠干扰素γ基因克隆、测序及辐射诱导表达质粒pIRESEgr-IFNγ的构建

目的 :克隆小鼠干扰素γ (IFN -γ)编码区 c DNA序列并构建含 Egr- 1启动子的辐射诱导表达质粒p IRESEgr- IFNγ。方法 :利用逆转录多聚酶链反应 (RT- PCR)法 ,以小鼠脾细胞 m RNA为模板 ,扩增获得全长IFNγc DNA,与 p GEMT载体连接做全自动测序 ,并利用基因重组技术构建含 Egr- 1启动子的辐射诱导表达质粒p IRESEgr- IFNγ。结果 :经测序证实获得的小鼠 IFNγ c DNA序列与文献报道完全一致 ,并构建了含 Egr- 1启动子的辐射诱导表达质粒 p IRESEgr- IFNγ。结论 :成功克隆了小鼠 IFNγ的 c DNA序列 ,构建了辐射诱导表达质粒p IRESEgr-