中药活性成分抑制肿瘤端粒酶活性的研究进展

端粒酶的活性与肿瘤的发生发展密切相关,因而抑制端粒酶活性研究必将成为肿瘤治疗的热点。本文综述了多糖类、生物碱类、皂苷类、黄酮类及其他中药活性成分抑制端粒酶活性的研究现状,并对其研究和应用前景作简要分析。     

哺乳动物早期胚胎端粒酶活性的研究进展

端粒位于真核染色体末端，是稳定染色体末端的重要元件。端粒酶（TER）是一种特殊的细胞核蛋白（RNP）反转录酶（RT），其核心酶包括蛋白亚基和RNA元件。在DNA复制过程中的端粒丢失可以被有活性的端粒酶补偿回来。哺乳动物端粒酶在发育中受调控，端粒的重编程可能是由于早期胚胎不同时期的端粒酶活性而造成的，因此，研究胚胎发育早期端粒和端粒酶重编程是非常重要的。本文对端粒和端粒酶的结构和功能，及其与哺乳动物早期胚胎发育的关系进行了综述．并在此基础上展望了端粒和端粒酶在克隆动物胚胎发育上的基础作用。     

端粒和端粒酶与肿瘤关系的研究进展

端粒是末端染色体 ,维持染色体的稳定。端粒酶是染色体末端转移酶 ,是合成端粒DNA的酶 ,对端粒的结构起着稳定的作用。端粒、端粒酶的研究始于 2 0世纪初期 ,但直到近几年科研人员才发现他们与肿瘤有着密切的关系。在人类端粒酶的研究过程中发现端粒酶在约 85 %以上的恶性肿瘤中呈阳性 ,而动物肿瘤与端粒酶的研究还是空白。文章主要介绍端粒、端粒酶的结构功能及它们与肿瘤关系的研究进展 ,并讨论世界上首例蛋鸡 J亚群禽白血病的自然发病的病例的端粒酶 TERT表达。作者曾用辣根过氧化物酶标记的链酶卵白素进行免疫组化检测 ,结果在病鸡的心、肝、脾、肺、肾、气管、十二指肠、睾丸、骨髓、脑、胸腺、法氏囊等组织中发现 TERT表达呈阳性 ,说明在蛋鸡 J亚群禽白血病发生时端粒酶活性升高 ,提示端粒酶参与了该病的发生发展机制 ,并且该结果与人类肿瘤端粒酶的活性的研究结果相一致     

小鼠卵母细胞及体细胞核移植重构早期端粒酶活性的测定

采用端粒酶重复序列扩增(telomeric repeat amplification protocol,TRAP)-银染法,对小鼠卵母细胞、体内受精胚胎及重构胚的各阶段端粒酶活性进行测定,对PCR产物的电泳条带进行灰度值分析,计算端粒酶总产品总量。结果表明:(1)体内受精胚胎及核移植重构胚各阶段的端粒酶活性随胚胎分裂呈上升趋势,囊胚期最高(P0.05);(2)体内受精胚胎的端粒酶活性高于体细胞核移植重构胚端粒酶活性。同时进行囊胚计数并测定单卵裂球相对端粒酶活性,结果表明:单卵裂球的端粒酶活性呈下降趋势,囊胚期端粒酶活性最低(P0.05)。以上结果说明小鼠卵母细胞及体细胞核移植重构胚端粒酶活性变化与细胞发育水平及细胞发育全能性有关。     

端粒酶活性检测方法的改进及dNTP浓度对端粒酶活性的影响

利用重新设计的引物和反应步骤 ,对检测端粒酶活性的 TRAP反应进行了改进。与原有方法相比 ,新方法在PCR扩增过程中不改变端粒酶产物的长度和比例 ,能更准确地反映端粒酶反应产物的性质。利用这个方法 ,考察了d NTP浓度对端粒酶的影响 ,发现不同的脱氧核苷酸对端粒酶活性的影响有差异。     

中药抑制肿瘤细胞端粒酶活性的研究进展

肿瘤严重威胁着人类健康,近年来癌症患者不断增多且死亡率逐年上升。中药以其安全性高、价格低、毒副作用小等优点成为抗肿瘤的首选药物。研究证实,肿瘤细胞与端粒酶之间存在相互激发的关系,肿瘤细胞缺乏端粒酶调节机制,一旦端粒酶活性被激活,肿瘤细胞将会有无限增殖的能力。因此,抑制肿瘤细胞端粒酶活性可作为治疗肿瘤的方法之一。文章将从中药的几种有效成分及复方中药对肿瘤细胞端粒酶活性的抑制作用作一综述,探讨中药抑制肿瘤细胞端粒酶活性的研究进展及其应用前景。     

端粒和端粒酶的研究进展

端粒和端粒酶是现代生物学研究的热点,端粒封闭了染色体的末端并维持了染色体的稳定性,端粒的缺失会引起染色体融合并导致细胞的衰老及死亡.端粒酶的活化可延长染色体末端DNA,维持基因组的稳定.并且端粒酶活性的异常表达又会引起细胞永生化或转化成癌细胞.因此端粒和端粒酶的结构和功能的研究对于治疗肿瘤和控制细胞寿命有着极其重要的意义.文章综述了端粒和端粒酶的结构和功能,及其与细胞老化的关系,并在此基础之上展望了端粒酶在抑制肿瘤、抗衰老等方面的应用.     

水牛卵母细胞和体外受精早期胚胎端粒酶活性测定

为了解水牛卵母细胞和体外受精（IVF）胚胎早期发育过程中端粒酶的活性变化,本研究利用端粒重复序列扩增法（TRAP）进行了水牛未成熟卵母细胞,成熟卵母细胞和2～4细胞,8～16细胞,桑椹胚以及囊胚各阶段的早期胚胎端粒酶活性的测定。依据电泳条带在成像系统下的光密度值,计算端粒酶的相对活性（RTA）。结果发现,未成熟卵母细胞端粒酶活性比成熟卵母细胞高（P〈0.05）,受精后2～4和8～16细胞胚胎端粒酶活性相对较低,桑椹胚端粒酶活性明显升高（P〈0.05）,囊胚阶段达到最高水平。通过对水牛不同发育阶段胚胎细胞数计数及单细胞相对端粒酶活性的分析比较结果显示,卵母细胞的单细胞端粒酶活性最高,囊胚阶段的最低。单细胞端粒酶活性从未成熟卵母细胞到IVF囊胚阶段呈逐渐降低的趋势。这些结果表明,水牛卵母细胞及早期胚胎的端粒酶活性变化与其成熟、发育阻断及全能性的逐步降低有关。     

端粒和端粒酶与衰老关系的研究进展

端粒是存在于线性染色体末端的一段特殊的DNA－蛋白质复合物，由于末端不能复制，正常体细胞随着细胞分裂的进行而逐渐丢失端粒序列，导致细胞老化和死亡。端粒酶是维持端粒长度的一种特殊的DNA聚合酶，在细胞的增殖、衰老及永生中起重要作用。本文就端粒和端粒酶及其与衰老的作一综述。     

Measurement of telomerase activity in dog tumors

Telomeres are specific structures present at the end of liner chromosomes. DNA polymerase can not synthesize the end of liner DNA and, as a result, the telomeres become progressively shortened by successive cell divisions. To overcome the end replication problem, telomerase adds new telomeric sequences to the end of chromosomal DNA. The enzyme activity is undetectable in most normal human adult somatic cells, in which shortening of the telomere is thought to limit the somatic-cell life span. In contrast to normal somatic cells, many human tumors possess telomerase activity. The present study looked at whether telomerase activity might serve as a marker for canine tumors. Telomerase activity was measured using the telomeric repeat amplification protocol assay. Normal dog somatic tissues showed little or no telomerase activity, while normal testis exhibited a high level of telomerase activity. We measured telomerase activity in tumor samples from 45 dogs; 21 mammary gland tumors, 16 tumors developed in the skin and oral cavity, 7 vascular tumors and 1 Sertoli cell tumor. Greater than 95% of the tumor samples contained telomerase activity (3-924 U/2 micrograms protein). The results obtained in this study indicated that telomerase should be a useful diagnostic marker for a variety of dog tumors, and it may serve as a target for antitumor chemotherapy.     

A cohort study of telomere and telomerase biology in cats

OBJECTIVE: To investigate telomere lengths in tissues of domestic shorthair (DSH) cats of various ages, evaluate the relationship between telomere length and age of cats, and investigate telomerase activity in the somatic tissues of cats. SAMPLE POPULATION: Tissues obtained from 2 DSH cats and blood samples obtained from 30 DSH cats. PROCEDURE: DNA isolated from blood cells and somatic tissue samples was subjected to terminal restriction fragment (TRF) analysis to determine mean telomere repeat lengths. Protein samples were subjected to analysis by use of a telomeric repeat-amplification protocol to assess telomerase activity. RESULTS: MeanTRF values of cats ranged from 4.7 to 26.3 kilobase pairs, and there was significant telomeric attrition with increasing age of cat. Telomerase activity was not found in a wide range of normal tissues obtained from 2 cats. CONCLUSIONS AND CLINICAL RELEVANCE: Analysis of these results clearly indicates that telomeres are shorter in older cats, compared with young cats; therefore, telomeres are implicated in the aging process. The analysis of telomerase activity in normal somatic tissues of cats reveals a pattern of expression similar to that found in human tissues. IMPACT FOR HUMAN MEDICINE: Fundamental differences in the biological characteristics of telomeres and telomerase exist between humans and the other most widely studied species (ie, mice). The results reported here reveal similarities in telomere and telomerase biologic characteristics between DSH cats and humans. Hence, as well as developing our understanding of aging in cats, these data may be usefully extrapolated to aging in humans.     

Induction of telomerase activity in avian lymphoblastoid cell line transformed by Marek's disease virus, MDCC-MSB1

Telomerase has been studied extensively in human and murine tumors, but little is known about the role of telomerase in the tumor biology of other vertebrate species such as the chicken. We studied the telomerase activity of the lymphoblastoid cell line derived from lymphomas induced by Marek's disease virus (MDCC-MSB1) compared with another avian cell line (PA5) and peripheral blood lymphocytes (PBL) using the telomeric repeat amplification protocol (TRAP) Assay. Telomerase activity in MDCC-MSB1 was 4.5 times greater than in the PA5 cell line and normal avian lymphocytes. These results demonstrate for the first time that telomerase is more intense in one transformed cell line than in normal cells, suggesting a potential role for telomerase in carcinogenesis induced by an avian virus.     

Telomerase Activity in Swamp Buffalo (Bubalus bubalis) Oocytes and Embryos Derived from In Vitro Fertilization, Somatic Cell Nuclear Transfer and Parthenogenetic Activation

The objective of this study was to examine the telomerase activity in swamp buffalo oocytes and pre-implantation stage embryos derived from in vitro fertilization (IVF), somatic cell nuclear transfer (NT) and parthenogenetic activation (PA). Immature and mature oocytes, and embryos at the 2-4 cell, 8-16 cell, morula and blastocyst stages produced by IVF, NT and PA were collected and the telomerase activity was assayed by using a Telomerase PCR ELISA kit. Telomerase activity was detected in all developmental stages evaluated from immature oocytes to blastocyst stage embryos. Telomerase activity was detected in higher amounts in immature as compared with mature oocytes (p < 0.05). Embryos derived from NT showed a profile of telomerase activity similar to that of IVF. In IVF and NT embryos, telomerase activity was low in the 2-4 cell and 8-16 cell stages, but the activity significantly increased (p < 0.05) at the morula stage, reaching its highest level at the blastocyst stage. In PA embryos, low levels of telomerase activity were detected from the 2-4 cell to the morula stage, and the highest level of telomerase activity was found at the blastocyst stage. Telomerase activity in NT blastocysts is higher than that derived from IVF and the activity is highest in PA blastocysts. These results suggest that the successful reprogramming of telomerase activity in buffalo NT embryos follow a pattern similar to that in embryos derived from IVF and PA.     

Inhibition of telomerase in canine cancer cells following telomestatin treatment

Telomere shortening in normal somatic cells has been proposed as a major barrier to unlimited cellular proliferation. Telomerase is an enzyme capable of maintaining telomere length, and thus bypassing this barrier. In human beings, telomerase activity is restricted to cancer cells and cells of stem or germ cell lineages. Dogs represent a potentially useful clinical model for the development of telomerase‐based therapies because telomerase activity is also restricted to cancer cells and stem cells in this species. We examined the ability of telomestatin to inhibit telomerase activity in telomerase‐positive D17 and CMT7 canine cancer cell lines. At a concentration of 2 μM, telomestatin treatment resulted in a decrease in telomerase activity, telomere shortening, growth inhibition and apoptosis in telomerase‐positive cancer cells. These effects were not seen in telomerase‐negative skin fibroblasts or negative controls. These results confirm that telomestatin specifically inhibits telomerase activity in canine cancer cells and strengthens the usefulness of dogs as a model for testing telomerase‐based therapies.     

Telomerase enzyme activity as a diagnostic tool to distinguish effusions of malignant and benign origin

Telomerase enzyme activity is high in populations of cells that are dividing, and is low or undetectable in quiescent cell populations. Activation of telomerase in tissues that normally lack the capacity for self-renewal is strongly correlated with neoplasia. Telomerase activity can be detected in samples containing very small numbers of cells and studies of human patients suggest that measurement of telomerase activity may be useful for the evaluation of samples that can be obtained in a minimally invasive manner. This study compares the presence or absence of telomerase activity with cytologic evaluation of body cavity effusions, to determine if neoplasia is the underlying cause for the effusion in dogs and cats. Detection of telomerase in effusions was no more sensitive than cytologic evaluation for the identification of underlying neoplasia, and was less specific (telomerase assay: sensitivity = 50%, specificity = 83%; cytology: sensitivity = 50%, specificity = 100%). We conclude that although the telomerase assay may constitute a useful adjunctive test for the diagnosis of neoplasia in some dogs and cats with body cavity effusions, the results of this assay are not sufficiently reliable to be used as a sole diagnostic test.