端粒及端粒酶的研究进展

端粒是染色体末端独特的蛋白质-DNA结构,在保护染色体的完整性和维持细胞的复制能力方面起着重要的作用.端粒酶则是由RNA和蛋白质亚基组成的、能够延长端粒的一种特殊反转录酶.端粒长度和端粒酶活性的变化与细胞衰老和癌变密切相关.端粒结合蛋白可能通过调节端粒酶的活性来调节端粒长度,进而控制细胞的衰老、永生化和癌变.研制端粒酶的专一性抑制剂在肿瘤治疗方面有着广阔的前景.     

细胞衰老与癌症治疗

细胞衰老是生物界普遍存在的现象。肿瘤细胞是一类摆脱细胞周期束缚,突破Hayflick界限,能够无限增殖而不衰老的细胞。癌症是一种与细胞衰老密切相关的疾病。从进化的角度来看,衰老对于生物体是有益的,可以导致细胞不可逆的周期阻滞,被认为是一种自主的肿瘤抑制机制。在恶性增殖的癌细胞中,在胞外及胞内多种刺激下,如端粒缩短、DNA损伤、氧化应激以及化疗药物的处理等,都会出现细胞周期阻滞,生长迟缓等细胞衰老现象。诱导肿瘤细胞衰老也被认为是一种治疗癌症的有效手段。衰老细胞可以向胞外分泌数十种因子,维持细胞自身衰老表型,并影响周围细胞的生长,这种特性被称为衰老相关的分泌表型(SASP)。本文详细综述细胞衰老的形态学特征与分子标记物及检测方法,细胞衰老的信号调控通路（p53-p21,p16-pRB和PTEN-p27）,以及细胞衰老与恶性肿瘤发生发展的关系等。尤其是应激压力诱导下的细胞衰老在癌症治疗中的潜在作用,并进一步讨论当下流行的促衰老癌症治疗的靶点与药物以及存在的问题,以期为今后的研究提供新思路和新方向。     

细胞衰老与肿瘤发生

细胞衰老（cell senescence）是指细胞在信号转导作用下不可逆地脱离细胞周期并丧失增殖能力后进入的一种相对稳定的状态。细胞衰老有增殖衰老与早熟衰老两种形式：增殖衰老由端粒缩短激发的信号转导激发,与TP53/CDKN1a（p21^WAF-1/Cip1）/pRB/E2F信号通路密切相关;早熟衰老由细胞内在或外在急慢性应激信号引发,与TP53/CDKN1a（p21^WAF-1/Cip1）/pRB/E2F或CDKN2a（p16^ink4A）/pRB/E2F信号通路相关。目前研究已经证实早熟衰老是细胞在癌变过程中的天然屏障,是继DNA修复、细胞凋亡后的第三大细胞内在抗癌机制,在机体防止肿瘤形成中起重要作用。     

细胞衰老与癌症治疗

细胞衰老是生物界普遍存在的现象。肿瘤细胞是一类摆脱细胞周期束缚,突破Hayflick界限,能够无限增殖而不衰老的细胞。癌症是一种与细胞衰老密切相关的疾病。从进化的角度来看,衰老对于生物体是有益的,可以导致细胞不可逆的周期阻滞,被认为是一种自主的肿瘤抑制机制。在恶性增殖的癌细胞中,在胞外及胞内多种刺激下,如端粒缩短、DNA损伤、氧化应激以及化疗药物的处理等,都会出现细胞周期阻滞,生长迟缓等细胞衰老现象。诱导肿瘤细胞衰老也被认为是一种治疗癌症的有效手段。衰老细胞可以向胞外分泌数十种因子,维持细胞自身衰老表型,并影响周围细胞的生长,这种特性被称为衰老相关的分泌表型(SASP)。本文详细综述细胞衰老的形态学特征与分子标记物及检测方法,细胞衰老的信号调控通路（p53-p21,p16-pRB和PTEN-p27）,以及细胞衰老与恶性肿瘤发生发展的关系等。尤其是应激压力诱导下的细胞衰老在癌症治疗中的潜在作用,并进一步讨论当下流行的促衰老癌症治疗的靶点与药物以及存在的问题,以期为今后的研究提供新思路和新方向。     

miRNAs与衰老相关信号通路的研究进展

miRNAs是一类负调控基因表达的内源性非编码小分子RNA,在细胞衰老过程中发挥重要作用. 细胞衰老是指可增殖细胞在各种应激下出现细胞周期阻滞,并且丧失增殖能力,进入一种不可逆的、相对稳定的状态. p53、p21、p16、SIRT1、胰岛素/IGF-1及mTOR等蛋白是衰老相关信号通路中的重要分子,参与细胞衰老过程. 研究表明,miRNAs可以通过调控这些衰老相关蛋白所在的信号通路,促进或延缓细胞衰老. 本文综述细胞衰老相关的miRNAs,以及它们对衰老相关信号通路的影响,为深化认识衰老和衰老相关疾病的分子机制奠定基础.     

从分子水平看c-Myc在细胞衰老中的作用

细胞衰老是指细胞生长永久阻滞于细胞周期的G1期,出现形态、生化及表观遗传的变化特性.细胞衰老由端粒缩短、DNA损伤、缺氧或癌基因失调等因素引起,它是抵抗肿瘤发生的主要壁垒.原癌基因c-myc编码转录因子,可调控很多基因,进而影响细胞周期演进、衰老、凋亡、代谢等生物学过程.c-Myc蛋白与细胞衰老密切相关,它可影响hTERT、p16、p53、Bmi-1和p27等衰老相关基因转录.c-Myc不仅可抑制复制性衰老,也能抑制癌基因诱发的衰老.c-Myc抑制ras诱导的细胞衰老取决于CDK2.c-Myc失活不仅能够诱导非恶性细胞(如人成纤维细胞)衰老,而且在许多肿瘤细胞中也可诱导衰老.然而,与ras基因类似,在特定条件下,c-Myc也可诱导细胞衰老,并可促进维氏综合症(Werner syndrome,WRN)缺失细胞的衰老.     

综述: 从分子水平看c-Myc在细胞衰老中的作用

细胞衰老是指细胞生长永久阻滞于细胞周期的G1期,出现形态、生化及表观遗传的变化特性.细胞衰老由端粒缩短、DNA损伤、缺氧或癌基因失调等因素引起,它是抵抗肿瘤发生的主要壁垒.原癌基因c-myc编码转录因子,可调控很多基因,进而影响细胞周期演进、衰老、凋亡、代谢等生物学过程.c-Myc蛋白与细胞衰老密切相关,它可影响hTERT、p16、p53、Bmi-1和p27等衰老相关基因转录.c-Myc不仅可抑制复制性衰老,也能抑制癌基因诱发的衰老.c-Myc抑制ras诱导的细胞衰老取决于CDK2.c-Myc失活不仅能够诱导非恶性细胞(如人成纤维细胞)衰老,而且在许多肿瘤细胞中也可诱导衰老.然而,与ras基因类似,在特定条件下,c-Myc也可诱导细胞衰老,并可促进维氏综合症(Werner syndrome,WRN)缺失细胞的衰老.     

综述: 衰老及相关疾病细胞分子机制研究进展

人类及其他生物随时间推移逐渐发生细胞功能丧失,即细胞衰老.这个过程如突显在某个组织器官,则可引起这个组织和器官的衰老性疾病.然而,最近的研究表明,哺乳动物在出生之前胚胎发育的生理条件下,即已经出现细胞和组织的复制性衰老现象.机制研究显示多种分子从细胞(核)内外引起生理性和应激性细胞复制性衰老.而自然界中某些生物随时间推移生命力增强、并不发生衰老.这些现象的分子机制,还有如发生在脑及代谢性疾病中的非复制性细胞衰老等,都还是个谜.本文就近期衰老的机制、细胞衰老的类型以及某些衰老相关疾病的分子基础的最新研究进展做一个扼要综述.论文包含以下几个部分:a.细胞衰老的定义、分类和机制;b.生理性衰老:发育中程序化衰老;c.内环境稳态与组织器官衰老;d.一型细胞复制性衰老及相关疾病:端粒长度与预测衰老及肿瘤预后、特发性肺纤维化、高血压;e.二型非复制性细胞衰老及相关疾病:帕金森病、糖尿病;f.衰老与长寿的物种多样性.     

衰老及相关疾病细胞分子机制研究进展

人类及其他生物随时间推移逐渐发生细胞功能丧失,即细胞衰老.这个过程如突显在某个组织器官,则可引起这个组织和器官的衰老性疾病.然而,最近的研究表明,哺乳动物在出生之前胚胎发育的生理条件下,即已经出现细胞和组织的复制性衰老现象.机制研究显示多种分子从细胞(核)内外引起生理性和应激性细胞复制性衰老.而自然界中某些生物随时间推移生命力增强、并不发生衰老.这些现象的分子机制,还有如发生在脑及代谢性疾病中的非复制性细胞衰老等,都还是个谜.本文就近期衰老的机制、细胞衰老的类型以及某些衰老相关疾病的分子基础的最新研究进展做一个扼要综述.论文包含以下几个部分:a.细胞衰老的定义、分类和机制;b.生理性衰老:发育中程序化衰老;c.内环境稳态与组织器官衰老;d.一型细胞复制性衰老及相关疾病:端粒长度与预测衰老及肿瘤预后、特发性肺纤维化、高血压;e.二型非复制性细胞衰老及相关疾病:帕金森病、糖尿病;f.衰老与长寿的物种多样性.     

腺病毒介导的p33ING1b过表达显著促进衰老细胞的DNA损伤修复

p33^ING1b是一个较晚发现的肿瘤抑制基因ING1的主要表达形式,自从被成功克隆以后得到了广泛的研究,已有的研究表明,p33^ING1b参与了细胞的生长抑制、凋亡、染色质重塑、DNA损伤修复、肿瘤抑制和细胞衰老等。但是它在细胞衰老过程中的作用特别是对衰老细胞DNA损伤修复的影响还没有被地阐明,在本研究中,我们首先用2BS细胞构建了细胞衰老模型,通过RT-PCR和Western blot技术证实p33^ING1b在衰老细胞中的表达水平是下调的,然后通过构建和包装包含p33^ING1b基因的腺病毒,将p33^ING1b导入年轻和衰老细胞中并使其过表达,用HCR（host cell reactivation）方法检测年轻细胞和衰老细胞DNA损伤修复能力。我们的实验首次表明,相对于年轻细胞,p33^ING1b的过表达使衰老细胞的DNA的损伤修复能力显著增加,这说明p33^ING1b在衰老细胞中的表达下调与衰老细胞DNA损伤修复能力的下降有关,也进一步证实了p33^ING1b在细胞衰老过程中起着十分重要的作用。     

DNA double strand break responses and chromatin alterations within the aging cell

Cellular senescence is a state of permanent replicative arrest that allows cells to stay viable and metabolically active but resistant to apoptotic and mitogenic stimuli. Specific, validated markers can identify senescent cells, including senescence-associated β galactosidase activity, chromatin alterations, cell morphology changes, activated p16- and p53-dependent signaling and permanent cell cycle arrest. Senescence is a natural consequence of DNA replication-associated telomere erosion, but can also be induced prematurely by telomere-independent events such as failure to repair DNA double strand breaks. Here, we review the molecular pathways of senescence onset, focussing on the changes in chromatin organization that are associated with cellular senescence, particularly senescence-associated heterochromatin foci formation. We also discuss the altered dynamics of the DNA double strand break response within the context of aging cells. Appreciating how, mechanistically, cellular senescence is induced, and how changes to chromatin organization and DNA repair contributes to this, is fundamental to our understanding of the normal and premature human aging processes associated with loss of organ and tissue function in humans.     

Senescence-associated heterochromatin foci are dispensable for cellular senescence,occur in a cell type- and insult-dependent manner and follow expression of p16ink4a

Cellular senescence, an irreversible proliferation arrest evoked by stresses such as oncogene activation, telomere dysfunction, or diverse genotoxic insults, has been implicated in tumor suppression and aging. Primary human fibroblasts undergoing oncogene-induced or replicative senescence are known to form senescence-associated heterochromatin foci (SAHF), nuclear DNA domains stained densely by DAPI and enriched for histone modifications including lysine9-trimethylated histone H3. While cellular senescence occurs also in premalignant human lesions, it is unclear how universal is SAHF formation among various cell types, under diverse stresses, and whether SAHF occur in vivo. Here, we report that human primary fibroblasts (BJ and MRC-5) and primary keratinocytes undergoing replicative senescence, or premature senescence induced by oncogenic H-Ras, diverse chemotherapeutics and bacterial cytolethal distending toxin, show differential capacity to form SAHF. Whereas all tested cell types formed SAHF in response to activated H-Ras, only MRC-5, but not BJ fibroblasts or keratinocytes, formed SAHF under senescence induced by etoposide, doxorubicin, hydroxyurea, bacterial intoxication or telomere attrition. In addition, DAPI-defined SAHF were detected on paraffin sections of Ras-transformed cultured fibroblasts, but not human lesions at various stages of tumorigenesis. Overall, our results indicate that unlike the widely present DNA damage response marker γH2AX, SAHF is not a common feature of cellular senescence. Whereas SAHF formation is shared by diverse cultured cell types under oncogenic stress, SAHF are cell-type-restricted under genotoxin-induced and replicative senescence. Furthermore, while the DNA/DAPI-defined SAHF formation in cultured cells parallels enhanced expression of p16^ink4a, such ‘prototypic’ SAHF are not observed in tissues, including premalignant lesions, irrespective of enhanced p16^ink4a and other features of cellular senescence.     

BTG2 antagonizes Pin1 in response to mitogens and telomere disruption during replicative senescence

Cellular senescence limits the replicative capacity of normal cells and acts as an intrinsic barrier that protects against the development of cancer. Telomere shortening–induced replicative senescence is dependent on the ATM‐p53‐p21 pathway but additional genes likely contribute to senescence. Here, we show that the p53‐responsive gene BTG2 plays an essential role in replicative senescence. Similar to p53 and p21 depletion, BTG2 depletion in human fibroblasts leads to an extension of cellular lifespan, and ectopic BTG2 induces senescence independently of p53. The anti‐proliferative function of BTG2 during senescence involves its stabilization in response to telomere dysfunction followed by serum‐dependent binding and relocalization of the cell cycle regulator prolyl isomerase Pin1. Pin1 inhibition leads to senescence in late‐passage cells, and ectopic Pin1 expression rescues cells from BTG2‐induced senescence. The neutralization of Pin1 by BTG2 provides a critical mechanism to maintain senescent arrest in the presence of mitogenic signals in normal primary fibroblasts.     

Reversal of human cellular senescence: roles of the p53 and p16 pathways

Telomere erosion and subsequent dysfunction limits the proliferation of normal human cells by a process termed replicative senescence. Replicative senescence is thought to suppress tumorigenesis by establishing an essentially irreversible growth arrest that requires activities of the p53 and pRB tumor suppressor proteins. We show that, depending on expression of the pRB regulator p16, replicative senescence is not necessarily irreversible. We used lentiviruses to express specific viral and cellular proteins in senescent human fibroblasts and mammary epithelial cells. Expression of telomerase did not reverse the senescence arrest. However, cells with low levels of p16 at senescence resumed robust growth upon p53 inactivation, and limited growth upon expression of oncogenic RAS. In contrast, cells with high levels of p16 at senescence failed to proliferate upon p53 inactivation or RAS expression, although they re-entered the cell cycle without growth after pRB inactivation. Our results indicate that the senescence response to telomere dysfunction is reversible and is maintained primarily by p53. However, p16 provides a dominant second barrier to the unlimited growth of human cells.     

Interleukin-6 decreases senescence and increases telomerase activity in malignant human cholangiocytes

BACKGROUND/AIMS: Cellular senescence results in irreversible growth arrest. In malignant cells, senescence is prevented by maintenance of chromosomal length by telomerase activity. Telomerase activity is increased in malignant, but not in normal cholangiocytes. Interleukin-6 (IL-6) is an autocrine promoter of cholangiocarcinoma growth. Our aims were to assess the relationship between IL-6 activated p38 mitogen-activated protein kinase (MAPK) pathways and senescence in malignant cholangiocytes. METHODS: Cell senescence and telomerase activity was assessed in Mz-ChA-1 malignant human cholangiocytes. The effect of inhibitors of p38 MAPK and telomerase activity on cell proliferation was assessed, and the interaction between these inhibitors was quantitated by median effects analysis. RESULTS: Mz-ChA-1 cells rapidly underwent senescence during repeated passaging. IL-6 increased telomerase activity and decreased cellular senescence during repeated passaging. However, basal telomerase activity was increased by inhibition of p38 MAPK. Inhibition of telomerase activity decreased IL-6 induced proliferation and had a synergistic effect with p38 MAPK inhibitors. Thus, IL-6 increases telomerase activity independent of p38 MAPK signaling and maintenance of telomerase activity promotes cholangiocarcinoma growth. CONCLUSION: Enhanced telomerase activity in response to IL-6 stimulation can prevent cellular senescence and thereby contribute to cholangiocarcinoma growth. Inhibition of telomerase activity may therefore be therapeutically useful in biliary tract malignancies.     

细胞衰老与细胞自噬的生物学关联及其意义

细胞衰老是指细胞生理功能的衰减,包括增殖能力下降、细胞周期停滞、对促凋亡应激不敏感、衰老相关基因和蛋白表达增加,伴有形态学衰老改变,渐趋死亡的现象,其至少可分为复制性衰老和应激诱导的衰老。细胞自噬属于细胞＂自食＂现象,是细胞依赖溶酶体的分解代谢过程,能降解受损蛋白、衰老或损伤的细胞器等细胞结构,可被多种应激所触发。细胞自噬的典型特征是形成自噬体并呈递给溶酶体,该过程在蛋白质和细胞器质量控制中起基础作用并维持了细胞能量的稳态。最新研究表明,自噬与细胞衰老密切相关,参与蛋白酶和自噬相关调节的BAG蛋白家族中BAG3/BAG1比值在复制性衰老时增高,且BAG3在细胞衰老时能介导自噬的激活。在Ras诱导的细胞衰老进程中亦可观察到较高的自噬活性。再者,自噬作为生物机体抗衰老的效应因子的遗传学证据已在低等真核生物中发现。还有研究证实,作为人类精液主要组分的亚精胺能够触发组蛋白H3脱乙酰基作用,此改变上调了自噬相关转录物的表达,继而引发自噬活性增强,从而延缓了多种细胞的衰老进程。另有研究显示,在P53/Arf的正常调节下,小鼠的衰老进程得以延缓,而Arf在细胞自噬过程的调节中亦是不可或缺的。总之,自噬活性的改变影响细胞衰老进程并可作为细胞衰老新的效应机理。