AIM To study the expression of telomerase activity in malignant esophageal neoplasms and normal humanesophageal epithelia.METHODS Telomerase activity was assayed by the telomere repeat amplification protocol (TRAP)method. All the neoplasms and epithelia of esophagus were confirmed by routine pathological diagnosis.RESULTS Telomerase activity was assayed in 18 normal esophageal epithelial tissues and in 35 malignantneoplasms of esophagus, including 27 cases of esophageal carcinoma and 8 cases of cardiac carcinoma.Telomerase activity was detected in most of malignant neoplasms of esophagus (91.4%, 32/35) and in allthe normal esophageal epithelial tissues except one (18/19).CONCLUSION The results suggest that in addition to contributing to proliferation of immortal blast cellsand neoplastic cells, telomerase activity may also play a similar role in regeneration of normal epithelia ofhuman esophagus. The potential use of telomerase activity as a diagnostic marker in human esophagealneoplasm might not be suitable.

AIM To investigate the diagnostic significance of cytology and telomerase activity in the exfoliated cells ofcardia obtained from endoscopic brushing in the cardiac cancer.METHODS The techniques of the qualitative TRAP-silver staining and quantitative TRAP-PCR-ELISAwere employed to detect telomerase activity in the exfoliated cells of cardia obtained from endoscopicbrushing in 72 cases with cardial lesions, cytological diagnosis was made at the same time.RESULTS Telomerase activity with cardiac cancer group (1.521 ± 0. 192) was significantly higher than thatwith cardialitis group (0.065± 0.014). Positive rate of telomerase activity detected in cardiac cancer group(88.89%) was significantly higher than that with cardialitis group (11.11%), the former was significantlyhiger than cytological examination (77.78%). The diagnostic rate of cardiac cancer reached 93.33% iftelomerase activity and cytology were examined at the same time.CONCLUSION Cytology and telomerase activity in the exfoliated cardiac cells may be an effective andsensitive methods in the diagnosis of cardiac cancer. This research can be a basis for the mass screening ofcardiac cancer.

hTERC基因与子宫颈上皮内瘤变相关性的研究进展

子宫颈癌是妇女常见的恶性肿瘤之一,其发病的主要原因是人乳头状瘤病毒(HPV)感染,约90％以上的宫颈癌患者伴有高危型HPV感染.且其发生发展的主要特点是由癌前病变即宫颈上皮内瘤变(CIN)逐步发展形成.所以,宫颈癌的筛查多采用高危型HPV检测和宫颈脱落细胞学检查.但两者均不能达到较高的敏感性及特异性.因此,探索更为有效、方便、敏感性及特异性更高的方法成为当务之急.宫颈细胞由非典型性异常向宫颈癌转变过程中几乎都伴有3号染色体长臂的扩增,其中涉及到的重要基因可能是人类染色体末端酶基因(telomerase human gene,hTERC),其有望成为非典型细胞癌变的基因[1].一、hTERC基因端粒酶是一种核糖核蛋白酶复合物,可以增加染色体末端的端粒重复序列.人类端粒酶RNA (hTERC)基因位于3号染色体26区,hTERC基因及其产物参与维持染色体的长度和稳定性,端粒酶的上调常常与恶性肿瘤的发生相关[1].细胞克隆进展伴随TERC基因拷贝数的增加,导致了端粒酶长度随正常细胞的增殖不断缩短,使癌细胞的增殖超过了正常增殖上限,进而导致了恶性肿瘤的发生[2].

Objective. To investigate the expression of telomerase gene hTRT mRNA in HeLa cells and to obtain hTRT pro-tein for futher study. Methods. The gene for encoding hTRT catalytic domain was cloned based on RT-PCR amplification from HeLa cells and sequenced. The cloned hTRTcDNA was in-frame inserted into His-tag fusion expression vector pEK318. The His-tag hTRT fusion proteins were purified by Ni-NTA chromatography and stained by westerm blotting. Results. An approximately 620bp fragment was generated and cloned into pBluescript SK + between Sail and BamHI sites. DNA sequencing showed the isolated fragment was consistem to those reported. SDS-PAGE present that a 17kDa protein was expressed stably in E. coli JM109 harboring pEKTRTM4 containing 6 × His-tag and hTRT 150aa, and the expression level of the protein was about 26% of the total bacterial proteins, while the expression of pEKTRT containing 6 × His-tag and hTRT 243aa was only detectable as 27 kDa band in western blotting. Both of fu-sion proteins were purified by Ni-NTA chromatography and showed single band( ＞ 95% purifity) in Coomassie Bril-liant staining. Westem-blotting confirmed that two proteins could be recognized by the Ni-NTA AP conjugate. Conclusions. The hTRT catalytic domain was highly conserved. The expressed hTRT protein contained recogniz-able His-tag, telomerase-specific and strong antigenic epitops, which may be convenient for further investigation.

放射性皮肤溃疡中端粒酶逆转录晦表达与癌变及难愈合的机制

AIM: To study the expression of the catalytic subunit of telomerase, telomerase revere transcriptase(TRT) and explore the possible relationship between the TRT and cancer txanaformation or poor healing in radiation-induced chronic human skin ulcer. METHODS: Rabbit antibody to human TRT and SP immunohistochemical method were used to detect TRT expression in 24 cases of formalin-fixed, paraffin-embeded chronic human skin ulcer tissues induced by radiation, 5 cases of normal skin, 2 of burnt skin, and 8 of carcinoma. RESULTS: The TRT was detected positive in 14 of 24 (58.3%) chronic radiation ulcers, of which the stxongly positive was 10 of 24 (41.7%) and the weakly positive 4 of 24 (16. 7% ); in 0 of 5 normal and 0 of 2 burnt skins; and in 8 of 8 (100%) carcinomas. The expression of TRT was observed almost always strongly positive in the cytoplasm and nucleus of squamous epithelial cells of epidermis but negatively in the endoepithelial cells of capillaries and small blood vessels, or weakly in the cytoplasm of smooth myocytes of media and fibroblssts, of dermis. Chronic inflammtory cells, such an plasma cells and lymphocytes also showed weakly positive for TRT. CONCLUSION: The strong TRT expression in the epidermis could be involved in the cancer transformation from chronic radiation ulcer to scuamous carcinoma, whereas the negative or weak TRT expression in the capillaries, small blood vessels and fibroblasts of dermis might be responsible for the poor healing of chronic ulcers induced by radiation, caused by sclerosis of small blood vessels and lack of granulation tissue consisting of capillaries and fibroblasts.

目的:研究端粒酶反义抑制剂对肿瘤细胞的影响.方法:以端粒酶RNA模板区为靶点,序列为TAGGGTTAGACAA的硫代磷酸修饰的反义寡核苷酸(PS-ASON)及13个寡核苷酸的随机引物与肾癌细胞株RCZ细胞共同孵育,计数1～43 d细胞数,计算细胞倍增时间,通过MTT法测定ASON对细胞生长的抑制率.结果:ASON实验组与随机引物对照组比较能明显减少细胞增殖数目(P<0.01)、增加抑制率(P<0.01)及延长倍增时间(P<0.005);并且其抑制效果显示出剂量的依赖性(P<0.05),具有序列选择特异性.结论:以端粒酶RNA模板区为靶点的反义寡核苷酸抑制肾癌RCZ细胞增殖,可能对肿瘤治疗有潜在的意义.