ATM对DNA损伤后细胞反应信号传导通路的调控作用

物理、化学及生物体自身的因素均可以导致染色体DNA的损伤，引起DNA单链或双链的断裂，受损的DNA能够继续复制并畸变或激活致癌基因。机体多通过激活细胞周期检测点、DNA修复蛋白以及保持与调控端粒功能等方式，或修复损伤的DNA，或使受损细胞凋亡，从而保持生物体DNA的稳定。本文重点讨论放射线辐射造成DNA损伤后，共济失调－毛细血管扩张症突变基因（Ataxia－telangiectasiamutatedgene，ATM基因）的产物－ATM编码蛋白（ATM）在细胞反应信号传导通路的中枢调控作用。１共济失调－毛细血管扩张症与ATM１．１共济失调－毛细血…     

乳腺癌中ATM基因的作用机制与突变种系

目前针对共济失调毛细血管扩张症的致病基因ATM基因的研究不断增多,该基因位于人类染色体的11q22～23,主要参与DNA损伤识别和修复,参与多个复杂的细胞周期关卡G1/S,S和G2/M,通过相应的信号转导途径,介导特定的分子间相互作用激活或抑制相应的细胞因子,起到调节细胞周期的作用。自从Swift等第一次报道了ATM杂合子患乳腺癌的风险增加后,作为乳腺癌危险因子ATM基因受到国内外学者的关注。本文将对ATM基因的作用机制以及乳腺癌中存在的ATM突变种系简要概述。     

共济失调性毛细血管扩张症一例

共济失调性毛细血管扩张症(ataxia telangiectasia)、又称Louis-Bar syndrome,是以机体免疫功能缺陷、眼与皮肤毛细血管扩张和进行性小脑共济失调为特征的一种遗传性疾病。本病国内较少见,现将我科发现一例报告如下。     

细胞周期检测点激酶1/2在相关肿瘤诊断治疗中的研究进展

细胞周期检测点激酶(Checkpoint kinase)的主要作用是在细胞DNA损伤后,通过及时阻滞细胞周期有效介导受损DNA的检测与修复,进而维持细胞基因组的稳定。当前细胞周期检测点激酶主要包括细胞周期检测点激酶1(Checkpoint kinase 1,Chk1)和细胞周期检测点激酶2(Checkpoint kinase 2,Chk2),二者均为蛋白激酶,具有相似的化学结构和重叠的底物谱,但在不同肿瘤的组织细胞中有不同的表达。本文就Chk1和Chk2的基本结构、生理功能及相应分子生物学机制,分析总结Chk1和Chk2作为肿瘤细胞的重要靶点与人类相关肿瘤诊断治疗的关系,旨在为今后相关肿瘤的临床治疗提供新思路。     

共济失调毛细血管扩张突变基因与恶性肿瘤关系研究进展

共济失调毛细血管扩张突变基因(ATM)是一种丝氨酸/苏氨酸蛋白激酶,作为DNA损伤反应的主要调控因子之一,调控p53、乳腺癌相关蛋白1、检查点激酶2等多种蛋白,在细胞对DNA损伤的反应和氧化应激中起着至关重要的作用,其也决定了基因组的稳定性。ATM突变或缺失的患者表现为恶性肿瘤易感性和放疗敏感性,同时恶性肿瘤的侵袭能力及预后也与ATM相关。通过抑制ATM的表达可显著提高癌细胞对放化疗的敏感性,有望作为恶性肿瘤治疗的新靶点。     

ATM启动子甲基化与肿瘤相关性的研究进展

毛细血管扩张-共济失调症突变基因(ATM)是重要的抑癌基因和DNA修复基因,其参与细胞DNA损伤的识别、修复,细胞周期及凋亡调控等。DNA甲基化是表观遗传修饰的主要方式之一,ATM启动子异常甲基化可引起ATM表观沉默,导致细胞的肿瘤易感性及放射敏感性增加,研究ATM启动子甲基化可为精准放射治疗及放射治疗增敏提供新的思路。该文主要对ATM启动子甲基化与肿瘤的相关性研究进行综述,为深入研究ATM启动子甲基化对肿瘤早期诊断、肿瘤放射治疗增敏的作用提供依据。     

p38丝裂原活化蛋白激酶在肺部疾病中的研究进展

 p38丝裂原活化蛋白激酶(p38 mitogen-activated protein kinase,p38 MAPK)是体内重要的信号转导通路,能够调控细胞的增殖、分化、死亡、炎症因子分泌等过程,大量研究已证实p38 MAPK信号通路在肺部疾病中的炎症反应、细胞增殖与凋亡、上皮间质转化、肿瘤侵袭等多个方面中起重要作用。目前已有p38 MAPK抑制剂在慢性阻塞性肺疾病中的临床研究,故p38 MAPK有望成为治疗肺部疾病的靶点。本文就p38 MAPK在肺部疾病中的相关研究作一综述。     

p38MAPK信号通路与前列腺炎疼痛关系研究进展

丝裂原活化蛋白激酶（mitogen activated protein kinase,MAPK）是细胞内的一类丝氨酸/苏氨酸蛋白激酶,它存在于大多数细胞内,是真核细胞转导细胞外信号到细胞内引起细胞反应的一类重要信号系统。p38丝裂原活化蛋白激酶（p38 mitogen activated protein kinase,p38MAPK）信号通路为     

p38丝裂原活化蛋白激酶在雄性小鼠生殖细胞株中的表达

目的 检测p38 丝裂原活化蛋白激酶(p38 mitogen-activated protein kinase,p38 MAPK)在雄性小鼠生殖细胞株TM4、GC-1spg、GC-2spd(ts)中的表达,为深入研究p38 MAPK基因在雄性小鼠精子发生中的作用机制奠定理论基础.方法 应用逆转录-多聚合酶链反应(RT-PCR)检测雄性小鼠生殖细胞株TM4、GC-1spg、GC-2spd(ts)中p38 MAPK基因的表达.结果 p38 MAPK基因在TM4、GC-1spg、GC-2spd(ts)细胞株中均为阳性表达.结论 p38 MAPK基因的表达在雄性小鼠精子发生中可能发挥重要作用.     

p38丝裂原活化蛋白激酶与骨代谢

丝裂原活化蛋白激酶(mitogen-activated protein kinases,MAPK)是机体广泛表达的丝氨酸/酪氨酸激酶,在哺乳动物细胞多种信号转导通路中起重要作用。p38MAPK信号通路是MAPK通路的一个重要分支,在细胞增殖、分化、凋亡和细胞周期调控等多种生理和病理过程中发挥重要作用。近年来,有关p38MAPK信号通路在与骨代谢相关的破骨细胞、成骨细胞、软骨细胞生长、代谢及功能方面的研究倍受关注。本文就 p38MAPK与骨代谢相关研究进展进行综述,旨在探讨p38MAPK在骨代谢相关疾病中的作用机制。     

Multiple defects of cell cycle checkpoints in U937-ASPI3K, an U937 cell mutant stably expressing anti-sense ATM gene cDNA

Ataxia--telangiectasia(AT)isanautosomalre-cessivemultisystemdisordercausedbytheheterozy-gousorhomozygousmutationofATMgene(ATrnutant).Atthecellu1arlevel,itischaracterizedbyhypersensitivitytoradiationorradiation--mimiccytcrtoxicagentsandinefficientcelIcycIecheckpointacti-vation[]'2J.ATMgeneencodesa350kdnuc1earphosphoproteinthatcontainsatitsCOOHterminusaphosphatidylinositol3kinase(Pl3K)catalyticdo-main.ATMproteinfunctionsincontrollingcellcyclecheckpointsinrespondingtoDNAdamageandpro-tect…     

PI3K-like kinases restrain Pim gene expression in endothelial cells

Pim kinases contribute to tumor formation and development of lymphoma,which shows enhanced DNA replication,DNA recombination and repair.Endothelial cells (ECs) express all the three members of Pim kinase gene family.We hypothesized that DNA repair gene would regulate Pim ex-pression in ECs.Human umbilical vein endothelial cells (HUVECs) were isolated and maintained in M199 culture medium.The cellular distribution of Pim-3 in ECs was determined by immunofluorescent staining.The siRNA fragments were synthesized and transfected by using Lipofectamine LTX.The total cellular RNA was extracted from the cells by using Trizol reagent.cDNAs were quantified by semi-quantity PCR.The effects of LY294002 and wortmannin on RNA stability in ECs were also ex-amined.Our data showed that LY294002 and wortmannin,phosphatidylinositol 3-kinase (PI3K) and PI3K-like kinase inhibitors,increased Pim mRNA expression in ECs without altering the mRNA stabil-ity.RNA interference (RNAi) targeting DNA-dependent protein kinase catalytic subunit (DNA-PKcs) and ataxia telangiectasia mutated (ATM) increased mRNA expression of Pim-3 and Pim-1,respectively.Silencing of Akt decreased Pim-1 instead of Pm-2 and Pim-3 gene expression in ECs.But etoposide,a nucleoside analogue,which could activate DNA-PKcs and ATM,increased Pim expression in ECs.Our study indicates that the expression of Pim kinases is physiologically related to DNA-PKcs and ATM in ECs.     

MEK AND p38 MAPK-DEPENDANT PATHWAYS ARE INVOLOVED IN THE POSITIVE EFFECT OF INTERLEUKIN-6 ON HUMAN GROWTH HORMONE GENE EXPRESSION IN RAT MtT/S SOMATOTROPH CELLS

Objective To investigate the effect of interleukin-6（IL-6） on the human growth hormone （hGH） gene expression in a rat somatotropic pituitary cell line MtT/S. Methods The plasmids containing various lengths of hGH gene 5＇-promoter fragments were constructed. Stably transfected MtT/S cells were created by cotransfecting the above plasmids and pcDNA3.1 （＋） with DMRIE-C transfection reagent. After the administration of these cells with IL-6 and/or various inhibitors of signaling transduction path-ways, the luciferase activities in MtT/S cells lysis were assayed to demonstrate the effects of IL-6 on hGH gene promoter activity and possibly involved mechanism. Results The 10^3U/mL IL-6 stimulated GH secretion and synthesis, and promoted the 5＇-promoter activity of GH gene in stably transfected MtT/SGL cells with the action of 1.69 times above the control. Among inhibitors of signaling transduction pathways, mitogen-activated protein kinase kinase （MAPKK/MEK） inhibitor PD98059 （40μmol/L） and p38 mitogen-activated protein kinase （MAPK） inhibitor SB203580 （5μmol/L） completely blocked the stimulatory effect of IL-6. Western blot analysis further confirmed the activation of phosphorylated MEK and p38 MAPK in MtT/SGL cells. Neither over-expression of Pit-1 nor inhibition of Pit-1 expression affected IL-6 induction of hGH promoter activity. A series of deletion constructs of hGH promoter were created to identify the DNA sequence that mediated the effect of IL-6. The results showed that the stimulatory effect of IL-6 was abolished following deletion of the - 196 to - 132 bp fragment. Conclusions IL-6 promotes GH secretion and synthesis by rat MtT/S somatotroph cells. The stimulatory effect of IL-6 on hGH gene promoter appears to require the activation of MEK and p38 MAPK, and a fragment of promoter se- quence that spans the - 196 to - 132 bp of the gene, but may be unlinked with Pit-1 protein.     

Caffeine Suppresses Apoptosis of Bladder Cancer RT4 Cells in Response to Ionizing Radiation by Inhibiting Ataxia Telangiectasia Mutated-Chk2-p53 Axis

Background:

Caffeine suppresses ataxia telangiectasia and Rad3 related and ataxia telangiectasia mutated (ATM) activities; ATM is the major kinase for DNA damage detection. This study aimed to investigate the effects of caffeine on DNA damage responses in cells from the bladder cancer cell line RT4 those were exposed to ionizing radiation (IR).

Methods:

Immunofluorescent staining was performed to investigate changes in the proteins involved in DNA damage responses with or without caffeine. A mouse xenograft model was used to study the effects of caffeine on the DNA damage responses. Western blotting was used to investigate the effects of caffeine pretreatment on the ATM-Chk2-p53-Puma axis, while real-time polymerase chain reaction (RT-PCR) assessed changes in messenger RNA levels of p53 and downstream targets responding to IR. Finally, terminal deoxynucleotidyl transferase-dUTP nick end labeling assay. Western blotting and colony formation assay were used to measure the effects of caffeine on radiation-related apoptosis. All of the data were analyzed with a two-tailed Student''s t-test.

Results:

Immunofluorescent staining showed that caffeine pretreatment profoundly suppressed the formation of γH2AXand p53-binding protein 1 foci in RT4 cells in response to irradiation. Cellular and animal experiments suggested that this suppression was mediated by suppression of the ATM-Chk2-p53-Puma DNA damage-signaling axis. RT-PCR indicated caffeine also attenuated transactivation of p53 and p53-inducible genes. The colony formation assay revealed that caffeine displayed radioprotective effects on RT4 cells in response to low-dose radiation compared to the radiosensitization effects on T24 cells.

Conclusion:

Caffeine may inhibit IR-related apoptosis of bladder cancer RT4 cells by suppressing activation of the ATM-Chk2-p53-Puma axis.     

原发性肝细胞癌中P16/CDKN2基因表达的研究

应用原位杂交技术检测了２８ 例原发性肝细胞癌（ＨＣＣ）中Ｐ１６／ＣＤＫＮ２ 基因ｍ ＲＮＡ的表达, 并用免疫组织化学法检测了Ｐ１６ 蛋白及细胞周期蛋白依赖性激酶４ （ｃｙｃｌｉｎ－ｄｅｐｅｎｄｅｎｔｋｉｎａｓｅ ４, ＣＤＫ４） 蛋白的表达。结果显示： ７例ＨＣＣ的Ｐ１６基因ｍ ＲＮＡ为阴性, １３ 例为异质性减弱, ８ 例为保留表达。ｍ ＲＮＡ阳性的ＨＣＣ中, 均可见到Ｐ１６ 蛋白表达。Ｐ１６／ＣＤＫＮ２ 基因表达与ＨＣＣ的分级、分化及转移无关。２８ 例ＨＣＣ中共有６ 例出现ＣＤＫ４ 蛋白阳性,这６ 例ＨＣＣ均有不同程度的Ｐ１６ 蛋白表达, 提示ＣＤＫ４ 蛋白表达可能是灭活Ｐ１６ 蛋白的又一途径。     

丝裂原活化蛋白激酶对佛波酯诱导滋养细胞MMP-9基因表达的影响

目的 探讨佛波酯(PMA)诱导细胞滋养层细胞(CTB)MMP-9基因表达的调控机制。方法 用细胞ELISA法测定CTB细胞的蛋白激酶活性变化;用反转录聚合酶链反应检测CTB中MMP-9的基因表达。结果 100 nmol/L PMA能迅速激活CTB中丝裂原活化蛋白激酶(MAPK)家族中细胞外信号调节蛋白激酶(ERK)、c-jun氨基末端激酶(JNK)以及p38 MAPK激酶的活性。100 nmol/L PMA刺激CTB引起MMP-9 mRNA表达显著增加,能被ERK或p38 MAPK的特异性抑制剂所抑制。结论 ERK和p38 MAPK可能是PMA诱导CTB中MMP-9基因表达增加的重要调节物质。     

EFFECT OF INTERLEUKIN-1β ON GROWTH HORMONE GENE EXPRESSION AND ITS POSSIBLE MOLECULAR MECHANISM IN RAT MtT/S SOMATOTROPH CELLS

Objective To elucidate the effect of interleukin-1β （IL- 1β） on human growth hormone （hGH） gene expression in a rat somatotropic pituitary cell line MtT/S.
Methods Stably transfected MtT/S cells were firstly established by transfecting 484-Lucl plasmid which contained hGH gene promoter --484 to ＋30 bp and luciferase reporter gene. The effect of IL-1β on hGH gene expression was determined by assaying the luciferase activities. RT-PCR method was also used to determine whether IL-1 recepor mRNA was expressed in MtT/S cells.
Results The 10^3 U/mL IL-1β stimulated secretion and synthesis of GH, and promoted the 5＇-promoter activity of GH gene in stably transfected MtT/SGL cells with the action of 1.38 times above the control. Among inhibitors of signaling transduction pathways, mitogen-activated protein kinase kinase （MAPKK/MEK） inhibitor PD98059 （40 μmol/L） and p38 mitogen-activated protein kinase （MAPK） inhibitor SB203580 （5 μmol/L） completely blocked the stimulatory effect of IL-1μ, and phosphatidylinositol-3-kinase （PI3-K） inhibitor LY294002 partly abolished the effect of IL-1μ. Western blot analysis further confirmed the activation of phosphorylated MEK and p38 MAPK in MtT/SGL cells. Neither over-expression of Pit- 1 nor inhibition of Pit- 1 expression affected induction of hGH promoter activity by IL-1μ. A series of deletion constructs of hGH promoter were created to identify the DNA sequence that mediated the effect of IL-1β, and results showed that the stimulatory effect of IL-1β was abolished following deletion of the --196 to -- 132 bp fragment.
Conclusions IL-1β promotes GH secretion and synthesis in rat MtT/S somatotroph cells. The stimulatory effect of IL-1β on hGH gene promoter appears to require the activation of MEK, p38 MAPK, PI3-K, and a fragment of promoter sequence that spans the -196 to -132 bp of the gene, but it may be unlinked with Pit-1 protein.