【Abstract】Objective To explore the relation between the expression of telomerase and DNA ploidy with biliarypancreatic system cancer, so as to find a better way to diagnose and distinguish jaundice between malignance and benign disease.Methods Endoscopic retrograde cholangiopancreatography (ERCP) were performed before operation in patients with obstructive jaundice. The bile and pancreatice juice were collected before ERCP. Biopsy specimens from part of patients were obtained during ERCP. All cancer specimens were possessed once again during operation and were assessed by the activity of telomerase and DNA ploidy. Results ① Telomerase positive rate 〔87.50%(56/64)〕 of tissue specimens in malignant obstructive jaundice were higher than that in benign obstructive jaundice 〔3.33%(2/60)〕，P=0.000. ② Telomerase positive rate〔71.88%(46/64)〕of Bile and pancreatice juice in malignant obstructive jaundice were higher than that in benign obstructive jaundice 〔3.33%(2/60)〕， P=0.000, tissue specimens obtained by endoscopy with malignant obstructive jaundice had detectable telomerase activity, positive rate was 83.33%（20/24）. ③ The rate of DNA heteroploid with malignant obstructive jaundice was 62.50%（40/64）, that of diploid can be seen in all patients with benign obstructive jaundice, the difference was statistically significant (P=0.000). ④ The rate of telomerase positive and DNA heteroploid in high differentiation tumor were significantly lower than in middlelow differentiation tumor （P=0.028，P=0.001）.Conclusion Applying the duodenoscope we collected the bile and pancreatic fluid before operation and obtain biopsy specimens whose telomerase activity and DNA ploid were detected. This is simple, safe, quick method which can identify the malignant and benign obstructive jaundice.
【摘要】 目的 探讨宫颈上皮内瘤变（CIN）和宫颈鳞癌（SCC）组织中人类染色体端粒酶基因（hTERC）的表达和临床意义。 方法 收集2007年10月-2009年6月经病理学证实的116例宫颈脱落细胞标本，其中LSIL（CINⅠ）30例、HSIL（CINⅡ/Ⅲ）37例、SCC 16例、宫颈炎33例，用荧光原位杂交（FISH）方法检测脱落细胞hTERC基因。 结果 在宫颈炎、LSIL、HSIL和SCC组中hTERC基因的表达率分别是6.1%、16.7%、51.4%和93.8%，其中，HSIL、SCC组与宫颈炎组比较，hTERC基因阳性率差异有统计学意义（Plt;0.05），LSIL组与HSIL组比较、LSIL组与SCC组比较、HSIL组与SCC组比较，差异有统计学意义（Plt;0.05），且随着病变程度增加，hTERC基因表达率增加。 结论 hTERC基因在细胞学LSIL、HSIL和SCC中表达异常，且随病变程度增加阳性表达率也增加，可作为宫颈癌癌前病变进展的生物遗传学监测指标，并有望成为宫颈癌早期筛查方法之一。【Abstract】 Objective To explore the clinical significance and expression of the human telomerase gene (hTERC) in the cervical intraepithelial neoplasia (CIN) and squamous carcinoma of the cervix (SCC). Methods According to histological biopsy from October 2007 to June 2009, 116 pap smears were divided into LSIL (n=30), HSIL (n=37), SCC (n=16), and cervicitis (n=33) groups. Fluorescence in situ hybridization (FISH) was used to detect the expression of hTERC. Results Positive expression rate of hTERC was 6.1% in cervicitis group, 16.7% in LSIL group, 51.4% in HSIL group, and 93.8% in SCC group, respectively. Compared to cervicitis group, the expression of hTERC in HSIL and SCC groups was significantly higher (Plt;0.05). Among LSIL, HSIL, and SCC groups, there were significant differencec in hTERC expression between every two groups (Plt;0.05). From LSIL to SCC, the expression of hTERC increased obviously. Conclusion Abnormal expression of hTERC exists in LSIL, HSIL, and SCC patients, which significantly increases during malignant development. It may be a biogenetics monitor index of cervical precancerosis and will be a screening marker for cervical cancer.
OBJECTIVE: To construct a plasmid which has a reporter gene for exploring the role of human telomerase reverse transcriptase(hTRT) in in-vitro cell cultivation. METHODS: hTRT was cut by restricted enzyme from plasmid pGRN145 and inserted to plasmid pEGFP-C1 (enhanced green fluorescent protein). RESULTS: Restricted enzyme analysis and DNA sequencing showed that the sequence of the pEGFP -hTRT transgenic plasmid was correct. CONCLUSION: The recombinant vector pEGFP-hTRT has been successfully constructed, and it can be used as a transgenic plasmid in generating immortalized cell lines.
Objective To review the research process of telomerase reverse transcriptase (TERT) in the restoration of neurological diseases. Methods The related l iterature on TERT in the restoration of neurological diseases was extensively reviewed and comprehensively analyzed. Results TERT was the significant component of telomerase and the critical regulator of telomerase activity. It played an important role in the pathomechanism of neurological diseases including tumors,neurodevelopmental deficits, and nerve injury. TERT was becoming a research focus in the reparative therapy of neurological diseases. Conclusion TERT has manifested its great academic significance and appl ication prospects in the reparative therapy of neurological diseases, which deserves a further investigation.
【Abstract】ObjectiveTo investigate the expressions of hTERT mRNA and BRCA1 protein and to analyze the correlation between these two factors in breast cancer. MethodsThe expression of hTERT mRNA was examined by reverse transcription polymerase chain reaction (RT-PCR). The expression of BRCA1 protein was examined by immunohistochemistry. ResultsThe positive rates of hTERT mRNA and BRCA1 protein were 72.1%（31/43） and 34.9%（15/43） in breast cancer tissue, were 5.0%(2/40) and 77.5%(31/40) in paracancerous breast tissue respectively. Significant difference existed between breast cancer tissue and paracancerous breast tissue (P＜0.05). Significant negative correlation existed between the expression of BRCA1 protein and expression of hTERT mRNA （r=-0.995, P＜0.01）. ConclusionThe expression of hTERT mRNA is upregulated in breast cancer, and expression of BRCA1 protein is downregulated in breast cancer. BRCA1 protein expression may be associated with expression of hTERT mRNA in breast cancer, which may be involved in the carcinogenesis of breast cancer.
OBJECTIVE: To prevent the senescence of ’seed cells’ for tissue engineering, the life span of human fibroblasts is extended by reconstitution of telomerase activity, and the osteogenic potential of these fibroblasts are tested. METHODS: The pGRN145 plasmids encoding human telomerase reverse transcriptase (hTERT) were introduced into the normal human primary fibroblasts by electroporation. Telomerase activity was analyzed by TRAP-PCR assay. The beta-galactosidase stain was used to indicate the signs of cell senescence. The hTERT positive fibroblasts were then induced to form bone nodules. The bone nodules were stained by tetracycline and Alizarin Red S. RESULTS: Stable telomerase activity could be detected in the transfected fibroblasts and no signs of cell senescence were found in the fibroblasts cultured for more than 50 doublings. The hTERT positive fibroblasts could form bone nodules when they were cultured in vitro induced by bone morphogentic protein 2 and tumor necrosis factor-alpha. CONCLUSION: The fibroblasts with reconstituted telomerase activity reserve their osteogenic potential.
【摘要】 目的 观察晚期糖基化终产物（advanced glycosylation end prodrcts，AGE）对人结肠癌细胞株SW-480增殖的影响，并探讨其可能机制。 方法 不同浓度AGE干预SW-480细胞，噻唑蓝（MTT）法比较各组细胞活力，流式细胞术观察AGE对SW-480细胞周期的影响，蛋白质印迹法观察AGE对SW-480细胞CyclinD1表达的影响，端粒重复序列扩增法（telomeric repeat amplification protocol，TRAP）银染法观察AGE对SW-480细胞端粒酶活性的影响。MTT测细胞活力的检测设置空白对照组、100 μg/mL小牛血清白蛋白（bovine serum albumin，BSA）组及50、100、500 μg/mL AGE组，其余检测只设置100 μg/mL BSA组和100 μg/mL AGE组。 结果 MTT结果示AGE促进SW-480细胞的增殖，且呈浓度依赖性。100 μg/mL BSA组与100 μg/mL AGE组72 h后的细胞G0/G1期所占百分比分别为56.02%±0.58%、51.93%±1.01%，差异有统计学意义（Plt;0.05）。蛋白质印迹法示100 μg/mL AGE组72 h后CyclinD1的表达较100 μg/mL BSA组增加，差异有统计学意义（Plt;0.05）。TRAP银染法检测示100 μg/mL AGE干预SW-480细胞72 h后可以增加端粒酶活性（Plt;0.05）。 结论 AGE可促进人结肠癌细胞SW-480生长，呈剂量依赖性。其作用机制可能与AGE上调CyclinD1的表达加速G1/S期转换及增加端粒酶活性有关。【Abstract】 Objective To observe the effects of advanced glycosylation end products (AGE) on proliferation of SW-480 cells and study the possible mechanism. Methods Various concentrations of AGE were designed to have impact on SW-480 cells. Proliferation of SW-480 cells was assessed by thiazolyl blue tetrazolium bromide (MTT) assay; The impact of AGE on the cell cycle of SW-480 cells was analyzed by flow cytometry (FCM); the influence of AGE on expression of CyclinD1 was checked by Western blotting; and the impact of AGE on telomerase activity was examined by telomeric repeat amplification proctol (TRAP) sliver staining. For the MTT assay, blank control group, 100 μg/mL bovine serum albumin (BSA) group, 50, 100 and 500 μg/mL AGE groups were designed, while for other examinations, there were only 100 μg/mL BSA group and 100 μg/mL AGE group. Results MTT result showed that AGE increased the proliferation of SW-480 cells in a dose-dependent mode. The proportion of the cells at G0/G1 stage of the 100 μg/mL BSA group and the 100 μg/mL AGE experimental group were (56.02±0.58)% and (51.93±1.01)% respectively after 72 hours, with a significant difference (Plt;0.05); western blotting showed that the expression of CyclinD1 in the 100 μg/mL AGE group was significantly higher than that in the 100 μg/mL BSA group after 72 hours; TRAP silver staining demonstrated that telomerase activity increased significantly after treated with 100 μg/mL AGE for 72 hours. Conclusions AGE can promote the growth of SW-480 cells in a dose-dependent mode. Its mechanism is mainly by up-regulating the expression of CyclinD1 to shorten G0/G1 and increasing the telomerase activity significantly.
Objective To evaluated the role of wt-P53 protein in telomerase regulation in keloid fibroblasts(KFBs). Methods The fibroblasts were derived from humankeloid tissue which was proved by pathological diagnosis. KFBs were divided into 2 groups, the transfection group and the untransfection group. wt-p53 gene was transfected into the fibroblasts by adenovirus vectors in the transfection group. The KFBs untransfected with wt-p53 gene served as control (untransfection group). After 48 hours of transfection, the expression of wt-P53 protein was analyzed by both Western blotting and immunofluorescence method, respectively. The telomerase activity was evaluated by TRAP-ELISA after 1-7 days of transfection. Results All the KFBs from 2 groups expressed wt-P53 protein. But the expression level of wt-P53 protein in the transfection group was significantly higher than that in the untransfection group.At the same time of high expression of wt-P53 protein, the telomeraseactivity of KFBs in transfection group was significantly lower than that in theuntransfection group(P＜0.05). Conclusion High level expression of wt-P53 protein can transiently inhibit the telomerase activity of KFBs.
Objective To investigate the variety of telomerase activity in the course of liver cancer development, and the possibility of using telomerase as a marker of HCC. Methods Human liver specimens, comprising 22 HCC and adjacent peritumoral tissues, 12 liver cirrhosistissues, 6 nodulat regenerative hyperplasia (NRH) tissues and 10 normal liver tissues, were examined for telomerase activity by TRAP assay based on PCR. Results Twenty of 22 HCC and 14 of 22 adjacent tissue specimens were positive for telomerase activity with a positive rate of 90.9% and 63.6% respectively. Ten of 12 liver cirrhosis tissues were positive with a positive rate of 83.3%. 5 of 6 NRH were positive with a positive rate of 83.3%. Telomerase activity was negative in 10 normal liver tissues. Conclusion Telomerase may occur in the progress of hepatocarcinogenesis. Telomerase can be used as a tumor marker of HCC.
Objective To introduce telomeres, telomerase and their expression in gastric carcinoma.MethodsThe related literatures were collected and reviewed.Rsults In summary, telomerase activity could be detected in 85%-90% of gastric cancer. Moreover, the patient with telomerase-positive tumors showed poorer prognosis than those with telomerase-negative tumours, indicating that telomerase-positive gastric cancer might have more malignant potential. ConclusionKnowledge of telomerase activity in gastric cancer may be useful in cancer diagnosis, as well as a prognostic indicator of clinical outcome. Future development of drugs aimed at telomerase inhibition may potentially provide a therapy with relatively less side effects.