TCT检测、阴道镜及荧光原位杂交在子宫颈病变诊断中的作用

目的比较液基细胞学检测(TCT法)、阴道镜检测及荧光原位杂交(FISH)方法对宫颈癌及癌前病变的诊断作用,并探讨其单独及联合检测对子宫颈病变诊断的临床意义。方法收集2007年11月至2009年2月苏州大学附属第三医院妇科115例宫颈脱落细胞标本,其中CIN患者54例、宫颈癌患者25例、正常对照组36例,上述标本均接受TCT液基细胞学检测及荧光原位杂交hTERC基因检测,其中CIN及宫颈癌患者接受阴道镜检查。结果 CINI、CINIⅠ/Ⅲ和宫颈癌患者行液基细胞学检查,宫颈癌诊断的敏感性、特异性及准确度分别为52.0%、100%和84.8%。阴道镜检查的敏感性、特异性及准确度分别为76.0%、94.4%和88.6%。FISH检测的敏感度、特异度及准确度分别为92.0%、72.2%和78.5%。在正常人、CINI、CINIⅠ/Ⅲ和宫颈癌患者宫颈脱落细胞hTERC基因扩增阳性表达率分别是2.8%、6.7%、35.9%和92.0%,组间差异具有统计学意义(P0.05)。结论荧光原位杂交hTERC基因检测宫颈癌具有较高的敏感性,能与TCT法和阴道镜检测较高的特异性形成互补;hTERC基因扩增阳性表达率与组织学分级具有紧密的相关性,可作为宫颈病变组织损伤程度及预后的一个指标。     

乳腺癌浸润淋巴细胞的细胞毒活性研究

目的 研究乳腺癌的肿瘤浸润淋巴细胞(TIL)细胞毒活性。方法 采用生物技术提取乳腺癌的肿瘤浸润淋巴细胞，经过一定时间诱导培养，在7天、14天、21天、28天和35天分别检测TIL对K562细胞的细胞毒活性。结果 TIL对K562细胞的细胞毒活性4～5周时细胞毒活性最强。结论 临床可在乳腺癌TIL诱导培养4～5周时进行临床应用，可取得最佳疗效。     

未转移结直肠癌患者术后转移风险预测列线图的构建

目的构建结直肠(colorectal cancer,CRC)患者预后列线图预测模型,并验证其准确性。方法回顾性分析就诊于苏州大学第三附属医院的438例CRC患者的临床病理资料。建立COX单因素、多因素回归模型确定预后的独立危险因素。用R软件建立列线图,绘制术后3年、5年无病生存率(disease free survival,DFS)校准图线并与实际观察情况比较。用Bootstrap法进行内部验证,计算一致性指数(C-index)评估模型准确性。分析时间依赖的ROC曲线比较其与第7版美国癌症联合委员会(American Joint Committee On Cancer,AJCC)分期系统(TNM系统)在预测术后的3年、5年的DFS的敏感性和特异性。结果438例CRC患者中233例患者出现转移,其中肝脏转移105例,肺转移57例。COX回归模型分析示肿瘤分化程度,肿瘤TNM分期,癌胚抗原(CEA)水平,糖类抗原19-9(CA19-9)水平,中性粒细胞与淋巴细胞之比(Neutrophil lymphocyte radio,NLR),抑癌基因P53是患者预后的独立影响因素。列线图用于预测生存的C指数为0.678。校正曲线表明预测的3年、5年DFS与实际观察情况高度符合。时间依赖的ROC曲线结果表明,相较于传统AJCC-TNM分期,列线图预后模型在预测术后3年和5年的DFS具有更高的敏感性和特异性。结论列线图可较准确预测个体CRC患者的预后,利于临床工作者对其随访或及时开展对患者有益治疗。     

实时荧光定量聚合酶链反应检测B7-H4基因表达的方法学构建

目的 建立实时荧光定量聚合酶链反应(PCR)检测协同刺激分子B7-H4基因表达的方法.方法 采用自行设计的检测协同刺激分子B7-H4及内参照基因GAPDH的引物及TaqMan探针,经优化反应体系及反应条件后,检测B7-H4及GAPDH的基因表达.将B7-H4和内参照基因GAPDH的PCR扩增产物经测序鉴定正确并纯化后分别与pMD19-T载体连接,克隆构建含B7-H4及GAPDH基因片段的重组质粒,作为实时荧光定量PCR检测B7-H4及GAPDH基因表达的标准品.将重组质粒标准品进行定量及梯度稀释,建立标准曲线.结果 PCR扩增产物分别经测序证实为B7-H4及GAPDH基因的特异性片段.该法检测B7-H4基因表达的灵敏度为527 copies/ml,线性范围为5.27×102～5.27×107 copies/ml,标准曲线方程为:Y=-3.1395X+41.805,直线回归相关系数r=0.995,批间变异系数范围为2.39%～3.59%,扩增效率108.2%.该法检测GAPDH基因表达的灵敏度为386 copies/ml,线性范围为3.86×102～3.86×107copies/ml,标准曲线方程为:Y=-3.2436X+41.083,直线回归相关系数r=0.999,批间变异系数范围为2.26%～3.86%,扩增效率103.4%.结论　成功构建实时荧光定量PCR检测协同刺激分子B7-H4基因表达的方法,该方法 特异性好、灵敏度高、重复性好.

Abstract：

Objective To establish a real-time polymerase chain reaction (PCR) method to detect costimulatory molecule B7-H4 gene expression. Methods The mRNA levels of costimulatory molecule B7-H4 and the reference gene GAPDH were detected by using real-time PCR using TaqMan technology. The primers and TaqMan probes of B7-H4 and GAPDH were designed. The B7-H4 and internal reference gene GAPDH fragments in pure form from classical PCR amplification were cloned into pMD19-T vector, and recombinant plasmids were used as the standard substances of B7-H4 and GAPDH quantitative detection.Standard curves were established using a serial dilution of quantified plasmids to measure B7-H4 and GAPDH. The reaction systems were optimized, and the sensitivity, specificity and reproducibility were evaluated. Results The amplification products were confirmed as the specific fragments of B7-H4 and GAPDH by DNA sequencing instrument. For B7-H4, the sensitivity was 527 copies/ml. The linear range was from 5.27 × 102 to 5.27 × 107 copies/ml, the standard curve equation was Y = - 3. 1395X +41. 805, the correlation coeffecient was 0. 995, the interassay coefficient of variations was 2. 39% -3.59%, and the amplification efficiency was 108.2%; For GAPDH, the sensitivity was 386 copies/ml. The linear range was from 3. 86 × 102 to 3.86 × 107 copies/ml, the standard curve equation was Y = - 3. 2436X +41. 083, the correlation coeffecient was 0. 999, the interassay coefficient of variations was 2. 26% -3. 86%, and the amplification efficiency was 103.4%. Conclusion The real-time PCR system for quantifying costimulatory molecule B7-H4 mRNA levels has been established successfully with high specificity, sensitivity and good repeatability.     

CD8+T细胞在胃癌组织中的表达及其临床意义

目的 观察胃癌组织中浸润性淋巴细胞CD8+T的表达及临床作用.方法 应用免疫组织化学染色检测102例胃癌患者手术标本中浸润淋巴细胞CD8+T的表达.结果 在胃癌、胃炎及胃息肉组织中均可见浸润淋巴细胞CD8+T的表达,但胃癌组浸润淋巴细胞CD8+T的表达[(29.53±14.71)个/HPF]明显高于慢性胃炎组[(19.54±8.10)个/HPF]和胃息肉组[(14.26±6.83)个/HPF],差异有统计学意义(P＜0.05).CD8+T细胞的表达与胃癌患者的性别、年龄、肿瘤部位、肿瘤大小、淋巴结转移、复发、病理分级和TNM分期无明显相关(P＞0.05);但与组织学类型和浸润深度有关,其中分化型胃癌患者CD8+T细胞的表达[(33.9±18.4)个/HPF]高于低分化型患者的表达[(27.6±12.0)个/HPF],差异有统计学意义(t=2.04,P＜0.05);未侵入肌层组的CD8+T细胞表达[(38.8±13.8)个/HPF]明显高于侵入肌层组[(27.7±14.1)个/HPF],差异有统计学意义(t=2.45,P＜0.05).与CD8+T细胞低表达组比较,CD8+T细胞高表达组胃癌患者中位生存时间延长5个月,但差异无统计学意义(χ2=1.0274,P＞0.05).多因素COX模型分析显示,与浸润淋巴细胞CD8+T低表达组比较,高表达组有降低胃癌死亡风险的趋势(RR=0.82,95%CI=0.41～1.65).结论 浸润性淋巴细胞CD8+T的表达与胃癌的预后有关.

Abstract：

Objective To study the expression and clinical significance of infiltrating lymphocytes CD8 + T in gastric carcinoma tissues.Methods The expression of infiltrating lymphocytes CD8 + T in 102surgically resected specimens of gastric cancer tissues was detected by using immunohistochemistry.Results CD8+ T cells were expressed both in gastric carcinoma,gastritis and gastric polyp tissues,but the number of CD8 + T cells in gastric carcinoma tissues [(29.53 ± 14.71 )/high powered field (HPF)]wassignificantly greater than in gastritis tissues [( 19.54 ±8.10)/HPF]and polyp tissues [( 14.26 ±6.83)/HPF](P ＜0.05).No correlation was found between CD8 + T lymphocyte count and gender,age,tumor location,tumor size,lymph node metastasis,tumor relapse,pathological grade or TNM stage ( P ＞ 0.05 ).However,there was a statistically significant correlation between the number of CD8 + T cells and the histological type,invasive depth.The CD8 + T cell count was obviously higher in the differentiated group [(33.9 ± 18.4)/HPF]than in the undifferentiated group [(27.6 ± 12.0)/HPF]( t = 2.04,P＜0.05 ).And the number of CD8 + T cells was greater in the non-invasive group [(38.8 ± 13.8)/HPF]than in the invasive group [(27.7 ± 14.1 )/HPF]( t = 2.45,P＜0.05 ).The median survival time of gastric cancer patients with high expression of CD8 + T cells was 5 months longer than in those with low expression,but there was no significant difference.Multi-factor COX model analysis demonstrated that the risk of death in gastric cancer patients was lower in the CD8 + T cells high expression group than in the low expression group ( RR = 0.82,95%,CI = 0.41-1.65 ).Conclusion Detection of infiltrating lymphocytes CD8 + T expression in gastric carcinoma tissues will be beneficial to the judgment of the prognosis.     

恶性肿瘤合并抗利尿激素分泌异常综合征13例临床分析

目的 分析合并抗利尿激素分泌异常综合征(SIADH)的恶性肿瘤患者的临床及实验室检查特点.方法 回顾性分析13例对ADH患者的原发肿瘤、临床特点、诊断及治疗情况.13例SIADH患者,基础疾病均为恶性肿瘤.患者无特殊临床症状,低钠血症的相关临床症状主要为倦怠、乏力、纳差、恶心、淡漠和头昏.结果 经限制液体摄入和补充氯化钠后,12例患者血钠水平恢复正常,5例小细胞肺癌化疗有效.结论 并发SIADH的肿瘤患者预后较差,应尽早诊断,必须限制液体入量和有效控制肿瘤.     

创建全科医师规范化培训示范基地的经验与体会

常州市第一人民医院创建于1918年,自2008年4月30日成为江苏省首批全科医师规范化培训基地以后,医院领导非常重视,一直致力于培训基地建设,不断扩大和改善教学条件,在全科医师规范化培训方面积累了近5年的教学经验,并在2011年获得国家发展和改革委员会批准立项"全科医生临床     

ATP和COA活化癌性胸水TIL的增殖力与杀伤活性的研究

目的探讨ATP与COA活化癌性胸水肿瘤浸润淋巴细胞(TIL)的增殖力与杀伤活性.方法应用ATP和COA作为癌性胸水TIL的培养液,在7天、14天、21天、28天分别计算其增殖力,测定其杀伤细胞活性.结果单一的完全培养液TIL增殖量仅为(51.37±18.56)×108/ml,加入ATP和COA的完全培养液在14天时TIL增殖量即可达(105.31±24.76)×108/ml,扩增能力增加一倍以上.加入ATP和COA所培养的TIL其杀伤细胞活性明显增强,在14天百分比达到58.12±12.35,21天百分比时可达78.50±11.93.同时可提前一周达到临床治疗胸腔积液输注所需的TIL数量.结论培养液中加入ATP和COA可以活化癌性胸水ITL的增殖力与杀伤活性,有一定的临床实用价值.