人胆管癌CD24+ CD44+ EpCAMhigh细胞亚群的分选及其肿瘤干细胞样特性的鉴定

目的 检测及分选人胆管癌中的CD24+ CD44+ EpCAMhigh细胞亚群,探讨其是否具有肿瘤干细胞样生物学特性.方法 流式细胞术检测6例人胆管癌中CD24、CD44、EpCAM的表达率;取2例人胆管癌新鲜标本种植到NOD/SCID鼠皮下,建立荷人胆管癌小鼠模型.流式细胞术分选CD24+ CD44+ EpCAMhigh亚群细胞,NOD/SCID鼠移植瘤试验鉴定其成瘤和分化能力.结果 6例人胆管癌组织标本和2例移植瘤中,CD24+ CD44+ EpCAMhigh在人胆管癌中表达率为0.58%～2.43%(平均值0.94%).运用NOD/SCID鼠移植瘤模型,分选得到CD24+ CD44+ EpCAMhigh亚群细胞.NOD/SCID鼠移植瘤试验,1×103个CD24+ CD44+ EpCAMhigh细胞能成瘤(3/8),而CD24- CD44- EpCAMlow/-细胞在5×104才能成瘤(1/8).CD24+ CD44+ EpCAMhigh胆管癌细胞NOD/SCID鼠移植瘤的组织类型及标记物的表达率和原代肿瘤相似.结论 人胆管癌中含有CD24+ CD44+ EpCAMhigh细胞亚群,具有强成瘤能力、自我更新和分化的能力,可能是胆管癌干细胞.     

小干扰RNA对肾癌细胞Ki67基因表达及其增殖的抑制作用

目的探讨小干扰RNA(siRNA)对人肾癌细胞增殖基因Ki67表达及其增殖、凋亡的影响。方法将Ki67 siRNA(100 nmol/L)转染人肾癌786-0细胞。采用RT-PCR、免疫印迹、免疫组化技术检测Ki67 mRNA及蛋白表达,MTT法检测细胞增殖,免疫组化TUNEL法检测细胞凋亡。结果Ki67 siRNA处理组786-0细胞Ki67 mRNA表达(37．6±1．9)%、Ki67蛋白表达(46．4±0．9)%、免疫组化Ki67表达吸光度(A)值52．5±2．3,阴性siRNA对照组分别为(97．3±0．9)%、(95．3±0．9)%、114．5±4．9,2组比较差异均有统计学意义(P＜0．01)。Ki67 siRNA处理组细胞增殖抑制率(63．6±1．6)%、凋亡细胞阳性率(41．7±0．6)%,阴性siRNA对照组分别为(2．8±0．2)%、(10．3±1．4)%,2组比较差异有统计学意义(P＜0．01)。结论肿瘤增殖基因Ki67 siRNA能抑制人肾癌786-0细胞Ki67基因表达,进而抑制其增殖,促进其凋亡,有望成为肾癌基因治疗的有效工具。     

利用人源化小鼠研究组织工程材料的体内免疫原性

目的建立并利用人源化小鼠模型,研究组织工程材料的体内免疫原性。方法利用NOD／SCID小鼠（非肥胖型糖尿病鼠与重症联合免疫缺陷鼠反交鼠模型）．通过腹腔注射人外周血单个核细胞（PBMC）（100×10^6）建立人源化小鼠,在人源化小鼠皮下植入异体人皮肤（阳性对照）、人体脂肪干细胞与脱钙骨构建的组织工程骨（实验组）、单纯脱钙骨材料（阴性对照）。在植入后1周、2周、3周、4周,取小鼠尾静脉血,流式细胞仪检测人CD4^＋和CD8^＋淋巴细胞比例。4周后,取小鼠的移植局部皮肤进行HE染色分析,同时检测脾细胞人CD4^＋和CD8^＋淋巴细胞比例。结果NOD／SCID小鼠腹腔注射人PBMC4周内,在外周血和脾中均可测到人CD4^＋和CD8^＋淋巴细胞。病理分析结果显示：在人源化小鼠皮下植入的组织工程材料及成体干细胞构建的组织工程骨组未见炎性细胞浸润;而单独植入人皮肤组可见明显的炎性细胞浸润。结论人源化小鼠可作为组织工程材料体内免疫原性的研究的良好模型。     

人肝癌细胞系MHCC97H中侧群细胞初探

目的 从人肝癌细胞系MHCC97H中分选侧群细胞,探索其形态、表型和功能特征.方法 利用流式细胞仪从MHCC97H中分选得到侧群细胞,用相差显微镜和透射电镜观察其形态学特点;Western blotting观察其ABCG2表达;用流式细胞仪评估侧群细胞与干细胞基因CD133、CD117的关系;用克隆形成实验和NOD/SCID接种实验分别观察侧群细胞体外增殖和体内成瘤情况.结果 侧群细胞体积较小,呈圆形或短梭形;侧群细胞的细胞器如线粒体、内质网等明显少于非侧群细胞;流式检测显示侧群细胞中含有CD133、CD117阳性细胞;Western blotting显示侧群细胞和非侧群细胞中均有ABCG2表达,两者无明显差异;平板克隆形成实验显示SP具有较强的克隆形成能力;体内移植实验显示2000个SP细胞即能在体内形成肿瘤.结论 肝癌细胞系MHCC97H中侧群细胞具有肿瘤干细胞样细胞的表型和特性;ABCG2可能不是决定肝癌侧群细胞表型的主要因素.     

人脐血单个核细胞体外扩增后植入NOD/SCID小鼠重建多系造血

目的探讨人脐血单个核细胞（MNC）体外扩增后植入能力的改变，以及对NOD／SCID小鼠造血重建的影响。方法采用不同细胞因子组合对人脐血MNC进行短期无血清培养扩增，比较扩增后的效果及细胞凋亡的变化；将扩增6d后的人脐血MNC植入经半致死剂量照射的NOD／SCID小鼠体内，观察小鼠6周后的存活率和造血重建情况。结果人脐血MNC在干细胞因子（SCF）、酪氨酸激酶受体3配基（FL）、白细胞介素6（IL-6）和白细胞介素3（IL-3）细胞因子共同作用下，培养6～11）d获得最佳扩增效果，同时细胞表面膜联蛋白V（Annxin V）的表达明显减少；移植6周后有55．6％的小鼠存活，存活小鼠的骨髓、脾和胸腺细胞中均能检测出人类CD45、CD34、CD33、CD3和CDl9抗原，外周血提取的DNA可检测出人特异的Cart—Ⅰ基因和Alu基因。结论SCF＋FL＋IL-6＋IL-3细胞因子协同作用能有效扩增人脐血MNC，MNC扩增6d后可成功植入N（）D／SCID小鼠体内，并帮助小鼠重建多系造血。     

microRNA-200c在胰腺癌干细胞中的表达及作用

目的：探讨microRNA-200c（miRNA-200c）在胰腺癌干细胞中的表达及作用。方法：应用流式细胞术在人胰腺癌PANC-1细胞中以CD24+CD44+ESA+为标记物分选胰腺癌干细胞,并通过NOD/SCID小鼠移植瘤实验验证其肿瘤干细胞特性;分别用RFQ-PCR法Transwell试验检测PANC-1细胞、胰腺癌干细胞以及转染miRNA-200c前体片段或阴性对照序列的胰腺癌干细胞的miRNA-200c表达与侵袭能力。结果：PANC-1细胞中分选出CD24+CD44+ESA+细胞（占0.8%）具有肿瘤干细胞特性,其在小鼠皮下移植后的移植瘤体积明显大于同期移植的PANC-1细胞[（1 725.14±261.29）mm3 vs.（479.65± 99.67）mm3,P<0.05];胰腺癌干细胞中miRNA-200c的表达量明显低于PANC-1细胞（0.15±0.01 vs. 1.00±0.09,P<0.05）,平均穿膜细胞数明显多于PANC-1细胞[（321±7.62）个vs.（70±16.47）个,P<0.05],但转染miRNA-200c前体片段后,胰腺癌干细胞中miRNA-200c表达量明显升高,平均穿膜细胞数明显减少（P<0.05）。结论：胰腺癌干细胞中miRNA-200c的表达降低,miRNA-200c具有抑制胰腺癌干细胞生长及侵袭力的作用。

     

胃癌CD133阳性细胞亚群肿瘤起始细胞样特性的检测

目的 探讨人胃癌细胞CD133＋亚群的分选及其特性的鉴定.方法 对50例胃癌原发灶和癌旁胃黏膜组织标本行免疫组织化学染色及Western blot检测CD133蛋白的表达.采用流式细胞仪检测不同分化胃癌细胞系CD133所占百分比,免疫磁珠法分选CD133＋细胞亚群,悬浮培养并观察其生长特性,并检测CD133阳性细胞裸鼠皮下接种致瘤能力.单细胞克隆观察单个CD133＋细胞生长特性,半定量反转录聚合酶链反应法鉴定相关干细胞标记的表达.结果 CD133蛋白多定位于胃癌原发灶黏膜及黏膜下层的肿瘤细胞膜表面,其相对表达量高于癌旁胃黏膜组织（P＜0.05）.不同分化程度的人胃癌细胞系KATO-Ⅲ、SGC-7901、AGS及MKN-45中CD 133＋亚群所占相对百分比为（28±2）％、（17±2）％、（6±2）％及（4±2）％.人胃癌细胞系KATO-Ⅲ分选后阳性组中CD133＋亚群比例分别为（91±3）％;培养1周后,达到（95±2）％,并不断增殖形成细胞球.而且其增殖能力强于阴性细胞[群体倍增时间分别为（21±3）h和（40±8）h,P＜0.05].CD133阳性组和未分选组细胞裸鼠皮下接种时成瘤率分别为100％和80％;而CD133阴性组不成瘤,CD133阳性组移植瘤平均体积及重量均大于未分选组（P＜0.05,P＜0.05）.单克隆形成实验示单个CD133＋细胞可形成新的细胞克隆.半定量RT-PCR检测示其表达干细胞标记物Oct-4、Nanog、Sox-2、Musashi-1及EGFR.结论 体外可成功分离、纯化和扩增人胃癌细胞CD133＋亚群,其具有自我更新、增殖能力及较强的致瘤能力,并表达部分干细胞相关基因.     

肿瘤标志物和Ki67在结直肠癌中的表达及预后因素分析

目的探讨术前血清肿瘤标志物与术后Ki67联合检测对结直肠癌病人预后评估的价值。方法收集2012年1月至2014年6月在武汉大学人民医院胃肠外科就诊的结直肠癌病人的临床病理资料和随访资料。癌胚抗原(CEA)>5μg/L定义为阳性,癌抗原(CA)19-9>35 kU/L定义为阳性。Ki67 LI大于其对应截断值定义为高表达(阳性)。绘制受试者工作特征(ROC)曲线寻找Ki67 LI的截断值[曲线下面积(AUC)最大时];Cox回归分析寻找影响结直肠癌病人预后的独立危险因素。结果研究共纳入311例结直肠癌病人,截至2019年6月1日,中位随访67个月(3~89个月),随访率94.2%,病死率32.5%。单因素生存分析提示,CEA阳性、CA19-9阳性,Ki67高表达与结直肠癌病人预后不良相关(均P<0.01)。联合血清肿瘤标志物和Ki67的生存分析结果提示,均阴性时病人预后最好,单阳性时次之,均阳性时最差(均P<0.01)。Cox回归结果提示:CEA[HR=6.461,95%CI(3.693,11.304),P<0.01]、CA19-9[HR=2.046,95%CI(1.214,3.449),P=0.007]、TNM[HR=2.104,95%CI(1.597,5.242),P=0.028]、CEA+Ki67[HR=9.992,95%CI(5.337,18.706),P<0.01]、CA19-9+Ki67[HR=5.345,95%CI(2.282,12.521),P<0.01]是影响预后的独立危险因素。结论联合检测术前血清肿瘤标志物和术后Ki67指标对评估结直肠癌病人预后具有重要价值。     

Luminal A型和HER-2阳性乳腺癌的肿瘤干细胞生物学行为比较

目的:比较人Luminal A型和HER-2阳性乳腺癌的肿瘤干细胞的生物学行为差异。方法:用Luminal A型和HER-2阳性乳腺癌组织培分离并培养原代乳腺癌细胞,获取富含肿瘤干细胞的微球体(MS);用胰酶将MS消化成单细胞(微球体细胞,MSDC),比较两类乳腺癌MSDC的体外增殖和自我更新能力、ALDH1+表型肿瘤干细胞含量,以及移植NOD/SCID小鼠后的成瘤情况。结果:Luminal A型和HER-2阳性乳腺癌组织分离的原代乳腺癌细胞均能培养出MS,但相对于Luminal A型乳腺癌,HER-2阳性乳腺癌来源的MS形成时间短,生成的MS体积大;HER-2阳性乳腺癌来源的MSDC无论在无血清或添加血清的培养基中的增殖与再生能力均明显强于Luminal A型乳腺癌来源的MSDC,且前者含ALDH1表型干细胞比例明显高于后者(均P0.05);HER-2阳性乳腺癌的MSDC在NOD/SCID小鼠体内成瘤能力也强于Luminal A型乳腺癌来源的MSDC。结论:Luminal A型和HER-2阳性乳腺癌分离出来肿瘤干细胞生物学行为存在明显的差异,两者可能有不同肿瘤干细胞起源。     

人膀胱癌组织中肿瘤干细胞样细胞的分离培养及PIWIL2表达

目的从人膀胱移行细胞癌（BTCC）组织中分离培养肿瘤干细胞样细胞（CSLC）,鉴定其生物学特性,并研究PIWIL2在CSLC中的表达和意义。方法收集12例不同临床分期BTCC患者组织标本,通过酶消化和原代培养相结合等方法处理,采用无血清悬浮培养法分离培养获得含CSLC的悬浮细胞球,流式细胞仪检测细胞表面分子标志CD133和CD44的表达,免疫磁珠分选系统分离CD133＋CD44＋细胞;采用半定量逆转录聚合酶链反应检测PIWIL2mRNA在CSLC中的表达;裸鼠皮下接种CS-LC,观察其成瘤能力。结果成功地从人膀胱癌组织分离培养获得可悬浮生长、稳定传代的CSLC,CSLC高表达CD133和CD44,裸鼠皮下接种1×104、1×105个CSLC均可全部成瘤,PIWIL2mRNA在CSLC的表达显著高于正常膀胱组织细胞。结论采用无血清悬浮培养法成功从人膀胱癌组织中分离获得CSLC,具有肿瘤干细胞特性,能高度表达PIWIL2,为靶向肿瘤干细胞的治疗提供了实验依据。     

Is a proliferation index of cancer cells a reliable prognostic factor after hepatectomy in patients with colorectal liver metastases?

BACKGROUND: In spite of many reports focusing on prognostic factors after hepatectomy in patients with colorectal liver metastases, few studies have investigated pathological factors, eg, fibrous pseudocapsulation, growth pattern at the tumor margin, and proliferation activity of cancer cells, other than histological type and surgical margin. The aim of the present study was to investigate whether absence of pseudocapsulation, infiltrative growth pattern of metastases, and higher proliferation of cancer cells shown by Ki-67 immunohistochemical reactivity were associated with poorer survival after hepatectomy among patients with colorectal liver metastases. METHODS: Between 1988 and 1998, 221 patients underwent hepatic resection of colorectal metastases with curative intent in our institution. Pathology analyses were focused on pseudocapsulation of liver metastases, growth pattern at the tumor edge, and Ki-67 labelling index (Ki-67 LI) of cancer cell nuclei. Univariate analyses of survival and of disease-free survival were performed for several clinicopathological factors, and multivariate analyses of survival and disease-free survival were also performed. RESULTS: The univariate survival analyses showed that pseudocapsulation, growth pattern, and Ki-67 LI were significant prognostic factors, besides synchronous versus metachronous occurrence of metastases, carcinoembryonic antigen level before hepatectomy, and number of metastases. A multivariate analysis showed that Ki-67 labeling index was the most reliable prognostic factor of survival. In addition, Ki-67 LI and microscopic growth pattern were multivariately predictive factors of disease-free survival. CONCLUSIONS: This large single-institution study showed that investigation of cancer cell proliferation and pathologic characteristics of the tumor margin are major prognostic factors.     

CyclinD1和Ki-67在茉莉酸甲酯抑制人肝癌细胞HepG2增殖作用中的表达变化

目的:探讨CyclinD1、Ki-67蛋白在茉莉酸甲酯(MeJA)抑制人肝癌细胞HepG2细胞增殖过程中的表达变化。方法:免疫细胞化学检测HepG2细胞CyclinD1、Ki-67蛋白的表达。结果:MeJA作用HepG2细胞48 h后,CyclinD1、Ki-67蛋白表达水平明显下降(P<0.01)。结论:MeJA通过下调Cy-clinD1、Ki-67蛋白表达,使细胞周期相关蛋白的表达发生改变,减少处于增殖期的细胞数目,从而发挥抗肝癌作用。     

The relationship between tumour proliferative activity, the systemic inflammatory response and survival in patients undergoing curative resection for colorectal cancer

Background  The aim of the present study was to examine the relationship between Ki-67, C-reactive protein and cancer-specific survival in patients undergoing resection for colorectal cancer.
Method  One hundred and forty-seven patients undergoing potentially curative resection for colorectal cancer had preoperative C-reactive protein concentrations and tumour Ki-67 labelling index measured.
Results  On univariate analysis, age ( P  < 0.001), Dukes stage ( P  < 0.001), C-reactive protein ( P  < 0.001) and expression of Ki-67 (< 0.01) were associated with poorer cancer-specific survival. Ki-67 labelling index and C-reactive protein were correlated ( r _s = 0.172, P  = 0.037). On multivariate analysis, age (HR 1.96, 95% CI 1.26–3.04, P  = 0.003), Dukes stage (HR 4.38, 95% CI 2.11–9.09, P  < 0.001) and C-reactive protein (HR 4.09, 95% CI 2.04–8.24, P  < 0.001) retained significance.
Conclusion  Increased tumour proliferation is associated with a systemic inflammatory response and poor cancer-specific survival in patients undergoing potentially curative surgery for colorectal cancer.     

结直肠癌中F框蛋白2的表达与Ki-67、血管内皮生长因子及预后的关系

目的探讨结直肠癌中F-box蛋白家族成员F框蛋白2（FBXO2）的表达与肿瘤增殖指标（Ki-67）、血管内皮生长因子（VEGF）的表达及预后的关系。 方法采用免疫组织化学MaxVision两步法检测105例结直肠癌组织标本中FBXO2、Ki-67及VEGF的表达情况,统计分析其与临床病理特征及预后的相关性。 结果Ki-67、VEGF和FBXO2阳性表达率分别为53.3%（56/105）、51.4%（54/105）、44.7%（47/105）。Ki-67与结直肠癌的原发灶大小密切相关（P=0.017）,VEGF与结直肠癌临床病理特征无明显相关性,FBXO2与结直肠癌患者的远处转移、AJCC临床分期密切相关（P=0.045、0.027）。Pearson相关分析提示,FBXO2与Ki-67、VEGF的表达呈显著相关（r=0.305、0.223,P=0.002、0.022）,Ki-67与VEGF的表达无相关性（r=0.160,P=0.102）。多因素Cox比例危险模型分析提示,高表达的FBXO2是结直肠癌患者不良预后的独立影响因素（HR=2.183,95%CI=1.075~4.434,P=0.031）。生存分析显示,结直肠癌组织中Ki-67、VEGF及FBXO2高表达的患者生存时间明显低于低表达者,差异有统计学意义（P=0.019、0.036、<0.001）。 结论FBXO2在结直肠癌的发生、发展中起到促癌的作用,是结直肠癌不良预后的独立影响因素,可能机制是通过泛素蛋白酶体途径参与调节结直肠癌的肿瘤增殖活性及新生血管形成。     

Prognostic significance of p27Kip1 and Ki-67 expression in carcinoma of the renal pelvis and ureter

OBJECTIVE: To determine the significance of p27(Kip1) (p27) for tumour behaviour and prognosis of patients with transitional cell carcinoma (TCC) of the renal pelvis and ureter. PATIENTS AND METHODS: Using immunohistochemical staining, the relationship was evaluated between p27 protein level (low < 50%, high > 50%) and the Ki-67 labelling index (low < 30%, high > 30%) and clinicopathological features of 37 consecutive Japanese patients with TCC of the renal pelvis and ureter. RESULTS: Low levels of p27 correlated with higher tumour stage (P < 0.05) and positive lymph node metastases (P < 0.05). There was no significant association between p27 staining and the grade and tumour proliferation as assessed by the Ki-67 index. A high Ki-67 index correlated with higher grade and stage (P < 0.05). Kaplan-Meier plots of survival rate in patients with low or high p27 staining showed that low levels correlated with a shorter disease-free and overall survival (P < 0.001 and P < 0.01, respectively). Similarly, patients with a high Ki-67 index had lower disease-free and overall survival than those with a low Ki-67 index (P < 0.01 and P < 0.05, respectively). The Cox proportional hazards model showed that a low level of p27 was an independent predictor of a shorter disease-free (P < 0.01) and overall survival (P < 0.05) on univariate analysis, but not of overall survival on multivariate analysis. A high Ki-67 index was an independent prognostic marker for shorter disease-free survival on univariate and multivariate analysis (P < 0.01) and for overall survival on multivariate analysis (P < 0.05). In those with a high Ki-67 index, increased p27 staining was associated with a better prognosis than decreased staining for disease-free and overall survival (log-rank test, P < 0. 01 and P < 0.05, respectively). CONCLUSIONS: The finding that a low level of p27 is associated with tumour invasion and unfavourable prognosis indicates that p27 may be a useful prognostic marker for survival in upper urinary tract cancer.     

Prognostic value of the expression of Ki-67, CD44 and vascular endothelial growth factor, and microvessel invasion, in renal cell carcinoma

OBJECTIVE: To determine if use of cell proliferation, cell adhesion, level of angiogenesis-related factors and presence of microscopic vascular invasion (MVI) could better predict the biological behaviour of renal cell carcinoma (RCC), which has a widely variable clinical outcome despite the use of conventional prognostic factors (staging and grading). MATERIALS AND METHODS: The expression of Ki-67, CD44H and vascular endothelial growth factor (VEGF) were assessed immunohistochemically in formalin-fixed, paraffin-embedded tissues from 48 RCCs, using a Ki-67 labelling index (LI), CD44 LI and level of VEGF expression, respectively. In addition all the pathological slides were reviewed retrospectively for the presence and absence of MVI. The prognostic value of all the variables assessed was then evaluated, and correlated with the usual prognostic variables and cancer-specific survival. RESULTS: Univariate analysis of cancer-specific survival showed that tumour stage (P < 0.001), tumour size (P = 0.005), metastasis, MVI, Ki-67 LI, CD44H LI and VEGF expression (all P < 0.001) were predictors of tumour-related death. There was a statistical correlation between CD44H LI and each of Ki-67 LI (r = 0.61), expression level of VEGF (r = 0.72) and presence of MVI (r = 0.71). Independent predictors of cancer-specific survival in a multivariate analysis were: in all patients with RCC, the MVI (P = 0.003) and VEGF expression (P = 0.01); in those with no metastases, MVI (P = 0.01); in patients with no MVI, VEGF (P = 0.04); and in patients with MVI, Ki-67 LI (P = 0.003). No independent predictor was identified in patient with metastases. CONCLUSION: This study suggests that cell proliferation, cell adhesion, the level of VEGF expression and the presence of MVI represent a complex tumour-host interaction that may favour the progression of RCC. Cell proliferation, CD44H and VEGF expression appear to be powerful markers for identifying patients with an adverse prognosis.     

术前辅助放疗对直肠癌细胞增殖和凋亡的影响

目的通过对直肠癌术前放射治疗的对比研究,观察放疗效果与直肠癌细胞增殖和凋亡关系。方法将80例pTNMⅡ期和Ⅲ期直肠癌患者分为2组,每组40例。对照组直接手术治疗,放疗组术前行40Gy放射治疗,放疗结束后1～2周手术。应用免疫组织化学法检测Ki-67表达,用TUNEL法检测细胞凋亡。结果放疗组术前有14例肿瘤显著缩小,18例部分缩小,8例轻微缩小;显效率35.0%,总有效率80.0%。放疗前癌组织的自发凋亡与Ki-67标记指数也有显著的正相关性(r=0.563,P=0.027);放疗后癌组织的Ki-67表达显著低于放疗前,也显著低于对照组(P=0.017和P=0.011);癌旁2cm正常黏膜Ki-67表达较对照组显著降低。放疗后癌细胞的凋亡指数显著高于放疗前,也显著高于对照组(P=0.013和P=0.018)。结论放疗主要通过抑制肿瘤细胞增殖和促进癌细胞凋亡产生治疗作用。Ki-67表达的高低可作为放疗的筛选指标。     

结直肠癌组织中CyclinB1、p34cdc2、PCNA和Ki-67表达的研究

目的探讨结直肠癌发生和分化中CyclinB1、p34cdc2、PCNA和Ki-67表达异常及其临床病理学意义.方法采用免疫组化SABC法检测CyclinB1、p34cdc2、PCNA和Ki-67在16例结直肠癌组织中蛋白的表达.结果CyclinB1和p34cdc2染色定位于细胞胞浆和(或)胞核内.PCNA和Ki-67染色定位于胞核内.在研究病例中有87.5%(14/16)CyclinB1阳性表达,81.2%(13/16)p34cdc2阳性表达,93.7%(15/16)PCNA阳性表达,81.2%(13/16)Ki-67阳性表达.低分化结直肠癌的CyclinB1、p34cdc2、PCNA和Ki-67阳性率明显高于高中分化结直肠癌,CyclinB1、p34cdc2、PCNA和Ki-67的表达具有高度一致性.结论CyclinB1、p34cdc2、PCNA和Ki-67的阳性表达率及表达强度同结直肠癌分化程度有密切关系.     

靶向增殖诱导配体的RNA干扰对人结直肠癌裸鼠移植瘤的影响

目的 利用RNA干扰(RNAi)技术观察增殖诱导配体(APRIL)siRNA(pGC-shAPRIL重组干扰质粒)对裸鼠SW480移植瘤的影响.方法 建立裸鼠人结直肠癌移植瘤模型,待皮下瘤块长到一定体积,全部裸鼠分为APRIL siRNA组、空载体组和磷酸盐缓冲液(PBS)组,分别进行瘤块内注射.逆转录-聚合酶链反应(RT-PCR)检测移植瘤APRIL mRNA水平;瘤块和裸鼠肝肺苏木素-伊红(HE)染色病理检测;免疫组织化学法检测APRIL、核增殖抗原(Ki-67)、基质金属蛋白酶(MMP)-2蛋白表达.结果 APRIL siRNA组,与空载体组和PBS组比较,APRIL mRNA[(1.36±0.03)×10~7、(1.05±0.02)×10~8、(1.04±0.01)×10~8 copy/ml]和蛋白表达得分(2.00±0.40、7.00±0.45、8.00±0.40)明显降低(P<0.05);瘤块体积显著减小[(0.78±0.04)、(1.60±0.07)、(1.66±0.06)cm~3](P<0.05);与之相应Ki-67(2.00±0.45、8.00±0.54、7.00±0.44)和MMP-2(3.00±0.44、7.00±0.45、7.00±0.55)蛋白表达显著减弱(P<0.05).结论 RNAi敲低APRIL基因表达,抑制了裸鼠SW480移植瘤的生长和转移,打靶APRIL基因的RNAi技术有望成为人结直肠癌的治疗途径.     

ALDH1、EpCAM及Ki67 mRNA在大肠癌患者循环肿瘤细胞中的表达及意义

目的探索大肠癌患者循环肿瘤细胞中ALDH1、EpCAM及Ki67mRNA的表达临床价值。方法收集56例健康志愿者和55例大肠癌患者术前外周静脉血,从全血中提取单个核细胞总RNA并反转录成cDNA,用荧光定量PCRTaqman探针法检测外周血样本中ALDH1、EpCAM及Ki67的表达。结果大肠癌组ALDH1、EpCAM及Ki67mRNA表达显著高于对照组（P〈0.05）;ALDH1的表达量与肿瘤浸润程度及淋巴结转移相关,EpCAM的表达量与TNM分期及远处转移相关（P〈0.05）。结论荧光定量PCR检测大肠癌患者循环肿瘤细胞中ALDH1、EpCAM及Ki67mRNA的表达作为一种简便可行的分子生物学方法,能为大肠癌患者的分期、预后及个体化治疗提供有价值的信息。