大肠癌组织中Fas配体表达与肿瘤浸润淋巴细胞凋亡的关系

目的　探讨大肠癌组织Fas配体 (FasL)表达与肿瘤浸润淋巴细胞 (TIL)凋亡的关系。方法　应用免疫组织化学染色和原位杂交方法 ,检测 3 0例大肠癌组织连续切片的FasL蛋白和mRNA的表达。应用免疫组织化学染色和原位末端杂交标记法 (TUNEL)检测 3 0例大肠癌组织连续切片的FasL表达阴性区和阳性区中TIL细胞总量和正处于凋亡状态的TIL细胞数。TUNEL法检测 3 0例大肠癌不同FasL表达区肿瘤细胞的凋亡情况。结果　①大肠癌组织FasL呈高表达 ,3 0例大肠癌组织中均有FasL表达 ,且每张切片中均有 75%以上的组织表达FasL。②FasLmRNA的表达部位与FasL蛋白的表达部位相对应。③不论是在同一组织切片不同部位或两组织切片间相比 ,FasL表达程度和范围都不均匀。同一组织切片中 ,FasL阳性表达区TIL细胞数比FasL阴性表达区减少 ,后者是前者的 2 .88倍。④同一组织切片中 ,FasL阳性表达区正在凋亡的TIL细胞数比FasL阴性区增多 ,前者是后者的 2 .13倍。⑤FasL表达阴性区肿瘤细胞的凋亡率 (81.2 % )比FasL阳性区 (3 7.4% )高 (P <0 .0 1)。结论　大肠癌细胞可通过表达FasL ,诱导TIL发生凋亡 ,反击机体免疫系统     

p73和p51基因在结直肠癌中的表达和意义

背景：p53基因是一种肿瘤抑制基因，其家族成员p73和p51在结构上与p53具有高度同源性，影响细胞转录和凋亡的功能与p53相似。目的：研究p73和p51基因在结直肠癌中的表达及其与细胞凋亡和肿瘤临床病理特征的关系，探讨两者在结直肠癌发生、发展中的可能作用。方法：以逆转录聚合酶链反应（RT—PCR）检测60例结直肠癌组织和相应癌旁组织中p73、p51mRNA表达，以原位末端标记（TUNEL）法检测细胞凋亡。结果：结直肠癌组织p73、p51AmRNA表达阳性率显著高于相应癌旁组织（p73：71．7％对5．0％，P〈0．01：p51A：46．7％对11．7％，P〈0．01）：p51B mRNA在结直肠癌组织与相应癌旁组织中的相对表达量无明显差异（0．7318±0．3628对0．6836±0．3516，P〉0．05）。p73、p51A mRNA表达阳性者肿瘤细胞凋亡指数分别显著低于p73、p51A mRNA表达阴性者（p73：3．2％±2．5％对5．5％±2．8％．P=0．003；p51A：2．6％±2．3％对4．9％±2．7％，P=0．001）。p73mRNA表达与结直肠癌的分化程度、TNM分期和淋巴结转移相关（P〈0．05），p51A mRNA表达仅与淋巴结转移相关（P〈0．05）。结论：结直肠癌中p73、p51A基因表达上调，两者可能通过抑制肿瘤细胞凋亡而参与了结直肠癌的发生、发展。p73过表达可能与结直肠癌预后不良有关。     

Fas、FasL蛋白在非小细胞肺癌组织的表达及其生物学意义

目的探讨Fas和FasL蛋白在非小细胞肺癌（NSCLC）的表达及其生物学意义。方法应用免疫组化法对45例NSCLC和25例癌旁正常肺组织中Fas和FasL蛋白表达进行检测。结果①与受检正常肺组织Fas表达相比，NSCLC Fas表达明显下调（P〈0．05）；Fas表达与NSCLC组织类型及分化程度有关（P〈0．05，P〈0．01）。②FasL表达与NSCLC组织类型、分化程度、临床分期及淋巴结转移有关（P〈0．01，P〈0．05）。NSCLC Fas与FasL表达无相关关系，NSCLC瘤团或癌巢周边区与中央区FasL表达差异显著。结论Fas和FasL异常表达在NSCLC发生、发展及转移过程中起重要作用。     

结直肠癌组织间质细胞中CD10蛋白的表达变化及临床意义

目的观察CD10蛋白在结直肠癌组织间质细胞中的表达情况,探讨其在结直肠癌发生、发展中的作用。方法同期手术留取的80份结直肠癌组织及50份癌旁正常结肠黏膜组织,采用快捷免疫组化Max、Vision“法检测两者间质细胞中CD10蛋白表达情况,并分析阳性表达率与结直肠癌临床病理特征的关系。结果CD10蛋白在结直肠癌组织和正常结肠黏膜组织问质细胞中的阳性表达率分别为70．00％（56/80）、0,差异有统计学意义（P〈0．01）;结直肠癌组织间质细胞中CD10蛋白表达与患者性别、年龄、肿瘤部位、临床分期无关（P均〉0．05）,而与肿瘤细胞的分化程度、浸润深度、有无脉管侵犯及淋巴结转移有关（P均〈0．05）。结论CD10蛋白在结直肠癌组织间质细胞中呈高表达,可能与肿瘤的发生、发展及浸润转移有关。     

胎儿生长受限孕妇胎盘Fas、FasL的表达变化

选择孕龄为37-42周的单胎妊娠妇女，根据新生儿出生体质量定为胎儿生长受限（FGR）30例，用免疫组化法检测其胎盘组织中Fas及FasL的表达，并与相同孕龄及新生儿出生体质量正常者（对照组）作比较。结果显示，FGR组Fas的表达明显强于对照组（P〈0．01），FasL的表达则明显弱于对照组（P〈0．01）。认为FGR者胎盘Fas的高表达、FasL的低表达使母-胎易发免疫排斥，致胎盘组织免疫病理损伤，导致FGR。     

肝癌组织M30蛋白表达及其与凋亡的关系

目的探讨原发性肝癌早期凋亡相关蛋白M30的表达及其与凋亡的关系。方法采用免疫组织化学链霉素抗生物素蛋白-过氧化物酶（SP）法分别检测了80例原发性肝癌及相应的癌旁组织、10例正常肝组织M30蛋白的表达情况，采用脱氧核糖核酸末端转移酶介导的缺口末端标记（TUNEL）技术，检测肝癌凋亡指数。结果80例原发性肝癌、相应的癌旁组织、10例正常肝组织M30平均指数分别为9．10±7．05、3．31±3．02、0．10±0．32，肝癌组织M30指数和TNM分期、组织分化程度、生长方式、生存期及复发时间密切相关（P〈0．05）。M30指数与凋亡指数呈显著正相关（P〈0．01）。结论M30有可能成为对肝癌预后进行推测的分子标志之一。     

结直肠癌患者血清MIC-1的表达及意义

目的评价结直肠癌患者血清中巨噬细胞抑制因子1（MIC-1）的表达水平及其临床意义。方法测定152例未经放化疗结直肠癌患者及120例健康对照组的血清MIC-1、癌胚抗原（CEA）及糖链抗原19—9（CA19—9）zk平。结果结直肠癌患者血清MIC-1水平较健康对照组显著升高（P〈0．01），MIC-1水平与Dukes分期、分化程度、浸润深度、淋巴转移、远处转移相关（P〈0．01），与年龄、组织类型无关。MIC-1表达与CEA呈正相关（r=0．514；P〈0．01），联合CEA及CA19-9检测可提高结直肠癌诊断的敏感性和准确率。结论血清MIC-1联合CEA及CA19-9对结直肠癌诊断和鉴别诊断有一定的临床应用价值。     

特发性肺纤维化患者肺泡和支气管上皮细胞凋亡及相关信号表达

目的探讨细胞凋亡和促凋亡信号在特发性肺纤维化（IPF）发生中的意义。方法应用末端原位杂交和免疫组化技术，检测12例IPF的肺活检组织和10例正常对照肺组织肺泡和支气管上皮细胞凋亡、P53、Fas抗原和Fas配体（Fas／FasL）蛋白表达的变化。结果IPF肺组织的肺泡和支气管上皮细胞凋亡率（12／12）明显高于对照组（0／10），IPF组肺泡上皮细胞中Fas、FasL、P53蛋白表达的阳性率（分别为12／12、12／12、11／12）均明显高于对照组（分别为5／10、2／10、0／10）。IPF组支气管上皮细胞中Fas、FasL、P53蛋白表达的阳性率（分别为12／12、12／12、11／12）也均明显高于对照组（分别为6／10、3／10、0／10）。相关分析结果表明，肺泡和支气管上皮细胞Fas／FasL、P53蛋白表达与其凋亡百分率之间均呈显著正相关（r值为0．625—0．839，P均〈0．01），肺泡和支气管上皮细胞Fas／FasL与P53蛋白表达之间也均呈显著正相关（r值为0．571～0．760，P均〈0．01）。结论肺纤维化时肺泡和支气管上皮细胞凋亡率增高，P53、Fas／FasL蛋白表达增强，这些因素在肺纤维化的发生和发展中起重要作用。     

巨噬细胞移动抑制因子在大肠癌中的表达及其与细胞凋亡的关系

目的观察巨噬细胞移动抑制因子在大肠癌中的表达及其与细胞凋亡的关系。方法应用免疫组化技术及DNA原位末端标记TUNEL法分别测定43例大肠癌及10例正常大肠组织中的MIF蛋白表达和细胞凋亡。结果大肠癌组织中的MIF表达率为65．1％（28／43），明显高于正常大肠组织0．0％（0／10），二者之间有显著性差异（P〈0．01）。MIF表达与大肠癌组织学类型及Dukes分期无明显相关（P〉0．05），但与大肠癌分化程度、淋巴结转移呈显著相关（P〈0．05）。MIF蛋白的表达与细胞凋亡指数呈负相关（r=-0．436，P〈0．01）。结论MIF蛋白的异常表达而引起的细胞凋亡抑制或逃避，在大肠癌的发生、发展中起一定的作用。联合检测MIF蛋白和凋亡指数，有助于对肿瘤细胞的分化程度作出正确评价，以指导临床治疗及估计预后。     

结直肠癌组织中AKT2的表达变化及意义

目的观察结直肠癌组织中AKT2蛋白的表达,探讨其在结直肠癌预后判断中的应用价值。方法采用免疫组化SP法检测75例结直肠癌组织和20例癌旁正常组织中的AKT2。结果AKT2阳性表达在正常组织中低于结直肠癌组织（P〈0．01）,并与淋巴结转移、Dukes分期、浸润深度、远处转移有关（P均〈0．05）;AKT2阳性者的生存时间明显短于阴性者（P=0．0007）。多因素分析结果显示,淋巴结转移、Dukes分期、远处转移和AKT2是独立的预后指标（P分别为0．013、0．018、0．001、0．038）。结论结直肠癌组织中AKT2表达上调;AKT2阳性表达与结直肠癌转移、侵袭性有关,可作为评价结直肠癌预后的指标之一。     

Fas receptor counterattack against tumor-infiltrating lymphocytes in vivo as a mechanism of immune escape in gastric carcinoma

We investigated the presence and functional status of surface expression of the Fas receptor (FasR) and its ligand (FasL) in tumor and tumor-infiltrating lymphocytes (TIL) in gastric carcinoma (n=36) from the primary locus, metastatic gastric carcinoma (n=30) from malignant ascites, and benign gastric mucosa (n=30) for the control. The quantitative analysis was based on the percentage of positive cells by a flow cytometry. The results showed that the membrane-bound FasL molecule was constitutively expressed in primary and metastatic gastric carcinomas as well as normal gastric epithelium in nearly all the patients. In particular, metastatic carcinoma proved to aberrantly express the FasL molecule. On the other hand, FasR expression ranged from minimal or absent in primary and metastatic gastric carcinomas, suggesting that the carcinoma might be rendered less sensitive toward FasR-induced killing. Apoptotic tumor cells detected by terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick and labeling (TUNEL) were barely identified in primary and metastatic carcinomas. In the analysis of TIL, the expression of FasR and FasL, and apoptotic TIL could not usually be observed in primary gastric carcinoma. In metastatic carcinoma, however, there was significant overexpression of FasR and FasL in immune TIL associated with a higher frequency of apoptotic cell death detected by TUNEL. The results suggest that metastatic carcinoma expressing FasL, but not FasL⁺ primary carcinoma, might evade the immune attack by apoptotic depletion of activated TIL through the FasR/FasL systems. These results provide the direct and quantitative evidence of FasR counterattacks and/or paracrine fratricides as a mechanism of tumor-immune escape in vivo in human cancer. Received: 29 March 2000 / Accepted: 30 June 2000     

Mechanism of counterattack of colorectal cancer cell by Fas/Fas ligand system

AIM: To determine the role of Fas/Fas ligand (FasL) in the immune escape of colon cancer cells. METHODS: Immunohistochemistry was used to observe the expression of Fas and FasL in the tissues of colon cancer patients. In situ hybridization was used to detect the localization of FasL mRNA expression in cancer tissues. Terminal deoxynucleotide transferase-mediated dUTP nick end labeling (TUNEL) assay and CD45 staining were performed to detect the apoptosis of tumor-infiltrating lymphocytes (TILs). Co-culture assays of colon cancer cells (SW480) and Jurkat cells (Fas-sensitive cells) were performed to observe the counterattack of colon cancer cells to lymphocytes. RESULTS: Of 53 cases of colon carcinomas, 23 cases (43.4%) expressed Fas which was significantly lower as compared to the normal colonic mucosa (73.3%, P<0.01), and 45 cases (84.9%) of colon carcinomas expressed FasL, whereas only two cases (3.75%) in normal mucosa expressed FasL. FasL expression in the colon cancer cells was found to be associated with increased cell death of TILs. The apoptotic rate of TIL in the FasL-positive staining regions of tumor cells was significantly higher than that in the FasL-negative staining region (54.84±2.79% vs 25.73±1.98%, P<0.01). The co-culture of SW480 cells and Jurkat cells confirmed the function of FasL on the SW480 cells. The apoptotic rates of Jurkat cells were found to be related with the amount of SW480 cells. CONCLUSION: Colon cancer cells can escape the immune surveillance and killing via decreasing Fas expression, and can counterattack the immune system via increasing FasL expression. Fas/FasL can serve as potential targets for effective antitumor therapy.     

Action and mechanism of Fas and Fas ligand in immune escape of gallbladder carcinoma

AIM: To study the role of Fas and Fas ligand (FasL) in biological behaviors of gallbladder carcinoma, and their correlated action and mechanism in tumor escape. METHODS: Streptavidin-biotin-peroxidase immunohistochemistry technique was used to study the expression of Fas and FasL protein in 26 gallbladder carcinoma tissues, 18 gallbladder adenoma tissues, 3 gallbladder dysplasia tissues and 20 chronic cholecystitis tissues. Apoptosis of the infiltrating lymphocytes in these tissues was studied by terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick-end labeling (TUNEL) method. Expression of both proteins and apoptosis of the tumor infiltrating lymphocytes in cancer tissues of primary foci was compared with clinicopathological features of gallbladder carcinoma. RESULTS: The positive rates of Fas were not significantly different among carcinoma, adenoma, dysplasia and chronic cholecystitis. The positive rate of FasL in carcinoma was significantly higher than that in chronic cholecystitis (X^2= 4.89, P&lt;0.05). The apoptotic index (AI) in carcinoma was significantly higher than that in adenoma (t‘= 4.19, P=&lt;0.01) and chronic cholecystitis (t‘= 8.06, P=&lt;0.01). The AI was significantly lower in well-differentiated carcinoma and Nevin Ⅰ-Ⅲ carcinoma than that in poorly-differentiated carcinoma (t‘= 2.63, P=&lt;0.05) and Nevin Ⅳ-Ⅴ carcinoma (t‘= 3.33, P&lt;0.01). The confidence interval (CI) of infiltrating lymphooltes in adenoma, chronic cholecystitis, well-differentiated carcinoma and Nevin Ⅰ-Ⅲ carcinoma was very significantly lower than that in carcinoma (t‘ = 6.99, P&lt;0.01), adenoma (t‘ = 3.66, P&lt;0.01), poorly-differentiated carcinoma (t‘ = 5.31, P&lt;0.01) and Nevin Ⅳ-Ⅴ carcinoma (t‘ = 3.76, P&lt;0.01), respectively. The CI of apoptosis of infiltrating lymphocytes in well-differentiated carcinoma was significantly lower than that in poorly-differentiated carcinoma (t = 2.52, P&lt;0.05), and was not significantly lower in Nevin Ⅰ-Ⅲ carcinoma than in Nevin Ⅳ-Ⅴ carcinoma (t = 1.42, P&gt;0.05). Apoptosis of infiltrating lymphocytes was not discovered in adenoma and chronic cholecystitis. CONCLUSION: FasL expressed in gallbladder carcinoma cells permits tumor cells to escape from immune surveillance of organism by inducing apoptosis in infiltrating lymphocytes of carcinoma tissues. Up-regulation of FasL expression plays an important role in invasive depth, histological classification and metastasis of gallbladder carcinoma.     

B7 molecule mRNA expression in colorectal carcinoma

AIM: To observe the status of tumor-associated B_7 molecule mRNA expression in human colorectal cancer tissue by in situ hybridization. METHODS: The mRNA expression patterns of cancerassociated B_(7-1),B_7H_1,B_7H_2,ICOS in 22 specimens of human colorectal cancer tissue were monitored by in situ hybridization (ISH) with digoxin-iabeled oligonucieotide probes. RESULTS: B_(7-1)B_7H_1,B_7H_2,ICOS mRNA were detected in both cancer cells and tumor infiltrating lymphocytes (TIL). The mRNA expression level of these molecules in tumor cells was higher than that in TIL (0.76±0.54-1.62±0.82 vs 0.38±0.19-0.65±0.33, P<0.001). There was no relationship between expression level of tested B_7 family molecules and patients'sex, age, differentiation status of cancer and regional lymph node metastasis. CONCLUSION: Th2 cytokine predominant in tumor microenvironment might be related to the expression of B_7H_1,B_7H_2 co-signal molecules in tumor cells and TIL.     

Suppression of FasL expression in tumor cells and preventing TNF-induced apoptosis was better for immune cells survival

OBJECTIVE: To elucidate if Fas/FasL signal pathway participates in the immune escape of tumor cells, and if contemporarily preventing Fas/FasL and TNF-induced apoptosis is better for immune cells survival than just blocking Fas/FasL-induced apoptotic signal. METHODS: Suppression of FasL expression in mouse H22 hepatocellular cancer cells by siRNA technique. Wild-type Ad5 14.7K gene was amplified by PCR and transduced into Jurkat T cells. Detecting apoptosis of target Jurkat cells by Flow Cytometry. Detection of TNF-alpha in the culture supernatant of H22 cells by ELISA. FasL and 14.7K gene expression in stably transfected or transduced clones were determined by western blotting. RESULTS: FasL expression in H22 cells was down-regulated following stable transfection with a plasmid encoding antisense FasL cDNA. Down-regulation of FasL expression in H22 cells had no effect on tumor growth in vitro. There was an apparent decrease in the number of apoptotic Jurkat T cells following coculture with transfected H22 cells, relative to coculture with FasL-expressing untransfected cells. Compared with untransduced Jurkat cells, apoptotic rates in 14.7K transduced Jurkat cells were significantly reduced in three different E/T ratios (P < 0.01), respectively. CONCLUSIONS: Fas/FasL signal pathway participated in the immune escape of tumor cells by inducing immune cells apoptosis. Reducing the expression of FasL in tumor cells can decrease the apoptotic rate of immune cells. Further blocking of apoptotic signal pathway of immune cells by preventing TNF-induced apoptosis can increase the survival of immune cells.     

凋亡相关基因caspase-9,bax及bcl-2在大肠癌中的表达及其意义

目的探讨凋亡相关基因easpase-9,bax和bel-2在大肠癌中的表达及其在大肠癌发生、发展中的可能作用及相互关系。方法应用免疫组化S-P法检测20例正常大肠黏膜、48例大肠腺瘤及56例大肠癌中的caspase-9、bax和bcl-2蛋白的表达。用TUNEL 法检测细胞凋亡。结果正常大肠黏膜、大肠腺瘤和大肠癌中caspase-9的阳性表达率分别为5．00％、33．33％、64．29％,其表达率在三者间差异有显著性(P<0．01)。bax在三种组织中的表达率分别为5．00％、35．42％、62．50％,三者间差异有显著性(P<0．01)。bcl-2 在三者中的阳性表达率分别为15．00％、87．50％、60．71％,三者间差异亦有显著性(P<0．01)。caspase-9,bax和bcl-2的表达与大肠癌的分化程度有关(P<0．01),与Dukes分期无关(P>05)。正常大肠黏膜、腺瘤和大肠癌中细胞凋亡指数差异有显著性(P<0．01),细胞凋亡指数与肿瘤的分化程度有关(P<0．01),与Dukes分期无关(P>0．05)。caspase-9、bax和bcl-2的表达与细胞凋亡指数有密切联系 (P<0．01)。结论肿瘤早期阶段的细胞凋亡异常,可能是大肠癌的发病原因之一。bcl-2和bax通过调节caspase-9参与大肠癌的发生。