lncRNA XIST-miR137-ATG5调节细胞自噬功能在肠癌细胞5-氟胞嘧啶耐药性中的作用

摘要 目的：本文旨在研究长链非编码RNA XIST-miR137-ATG5的相互作用,同时探讨其调节细胞自噬功能与肠癌细胞5-氟胞嘧啶敏感性的关系。方法：实时聚合酶链反应(real time PCR)检测XIST与miR-137在肠癌细胞中的表达;采用脂质体转染法将si-XIST,miR-137转染入肠癌SW480及HCT116细胞中。采用CCK-8检测瞬时转染si-XIST对肠癌细胞增殖及5-FU敏感性的影响;并利用双荧光素酶报告实验检测miR-137与XIST, miR-137与ATG5相互关系。Western blot方法检测XIST- miR137- ATG5对细胞自噬的影响。结果：与正常结肠细胞FHC比较, XIST在结肠癌细胞系明显高表达,miR-137在结肠癌细胞系明显低表达。与阴性对照组比较,转染si-XIST后,SW480及HCT116细胞增殖能力明显受到抑制,对F-5U的敏感性增强,且抑制自噬蛋白Beclin-1及LC3II/LC3 I的表达。miR-137可与XIST,ATG5 3''UTR结合,抑制XIST和ATG5的表达及功能。在结肠癌SW480细胞中共转染miR-137 inhibitor或过表达ATG5可逆转XIST沉默引起的5-FU耐药,同时可逆转因XIST沉默引起的自噬蛋白表达的抑制。结论：LncRNA XIST或可通过调控mir137-ATG促进结直肠癌细胞SW480自噬从而提高其对5-FU的耐药,针对其这一机制,可为将来针对结肠癌的靶向治疗提供一定的实验基础。     

萝卜硫素诱导RASD1的表达促进宫颈癌HELA细胞凋亡及周期阻滞的机制研究

目的:探讨萝卜硫素靶向RASD1,对宫颈癌细胞(HELA)凋亡及周期的影响机制。方法:不同浓度的萝卜硫素(SFN)作用于HELA细胞48h,采用CCK-8法检测SFN对HELA细胞增殖的影响;采用RT-PCR法检测SFN对RASD1基因mRNA表达水平的影响;采用Western印迹法检测SFN对RASD1蛋白表达水平的影响;采用流式细胞技术检测加药以及过表达RASD1对HELA细胞凋亡及周期的影响,采用Western blot检测HELA中cleaved-caspase3和cleaved-parp相关凋亡蛋白以及p21和cdc2周期相关蛋白的影响。结果:SFN抑制宫颈癌细胞增殖,且加药组中RASD1的mRNA和蛋白表达水平明显高于对照组,差异有统计学意义(P0.05)。流式细胞凋亡分析结果提示:加SFN或是过表达RASD1细胞组凋亡率明显高于对照组,Western blot结果表明加SFN或过表达RASD1后细胞cleaved-caspase3以及cleaved-parp蛋白水平升高,差异具有统计学意义(P0.05);ModFit LT软件分析结果表明加SFN或过表达RASD1细胞S期显著低于对照组,G2/M期细胞增多,同时Western blot结果示p21蛋白的表达水平增高,而cdc2蛋白表达降低,其差异有统计学意义(P0.05)。结论:在宫颈癌HELA细胞中RASD1作为SFN的作用靶点,诱导细胞凋亡和细胞周期阻滞。     

p16基因导入致人乳腺癌MCF-7细胞端区缩短及细胞周期阻滞

为进一步探讨 p1 6基因在抗肿瘤及细胞衰老中的作用 ,以脂质体介导的方法 ,将重组的含全长 p1 6c DNA的逆转录病毒载体导入人乳腺癌 MCF- 7细胞 ,获得稳定整合有效表达 .检测其对MCF- 7细胞的端区长度、细胞形态、增殖特性及细胞周期的影响 .结果显示 :导入 p1 6c DNA后的MCF- 7细胞端区长度明显缩短、增殖减慢 ,细胞周期阻滞于 G1期 .由此推测 ,野生型 p1 6基因可能通过诱导端区缩短效应及抑制细胞增殖从而抑制肿瘤和启动细胞衰老 .     

Multiple domains of the mouse p19ARF tumor suppressor are involved in p53-independent apoptosis

The ARF (p19ARF for the mouse ARF consisting of 169 amino acids and p14ARF for the human ARF consisting of 132 amino acids) genes upregulate p53 activities to induce cell cycle arrest and sensitize cells to apoptosis by inhibiting Mdm2 activity. p53-independent apoptosis also is induced by ectopic expression of p19ARF. We constructed various deletion mutants of p19ARF with a cre/loxP-regulated adenoviral vector to determine the regions of p19ARF which are responsible for p53-independent apoptosis. Ectopic expression of the C-terminal region (named C40) of p19ARF whose primary sequence is unique to the rodent ARF induced prominent apoptosis in p53-deficient mouse embryo fibroblasts. Relatively low-grade but significant apoptosis also was induced in p53-deficient mouse embryo fibroblasts by ectopic expression of p19ARF1-129, a p19ARF deletion mutant deficient in the C40 region. In contrast, ectopic expression of the wild-type p14ARF did not induce significant apoptosis in human cells. Taken together, we concluded that p53-independent apoptosis was mediated through multiple regions of the mouse ARF including C40, and the ability of the ARF gene to mediate p53-independent apoptosis has been not well conserved during mammalian evolution.     

Cell Cycle Arrest of Proliferating Neuronal Cells by Serum Deprivation Can Result in Either Apoptosis or Differentiation

Abstract: Apoptotic cell death plays a critical role in the development of the nervous system. The death of mature nondividing neurons that fail to receive appropriate input from the target field has been extensively studied. However, the mechanisms mediating the extensive cell death occurring in areas of the developing brain where proliferating neuroblasts differentiate into mature nondividing neurons have not been analyzed. We show here that the cell cycle arrest of a proliferating cell of neuronal origin by removal of serum results in either apoptotic cell death or differentiation to a mature nondividing neuronal cell. The proportion of cells undergoing death or differentiation is influenced in opposite directions by treatment of the cells with cyclic AMP and retinoic acid. This suggests that following the withdrawal of signals stimulating neuroblast cell division, neuronal cells either can cease to suppress a constitutive suicide pathway and hence die by apoptosis or, alternatively, can differentiate into a mature neuronal cell. Regulation of the balance between apoptosis and neuronal differentiation could therefore play a critical role in controlling the numbers of mature neurons that form.     

Deoxyelephantopin from Elephantopus scaber L. induces cell-cycle arrest and apoptosis in the human nasopharyngeal cancer CNE cells

Deoxyelephantopin (ESD), a naturally occurring sesquiterpene lactone present in the Chinese medicinal herb, Elephantopus scaber L. exerted anticancer effects on various cultured cancer cells. However, the cellular mechanisms by which it controls the development of the cancer cells are unavailable, particularly the human nasopharyngeal cancer CNE cells. In this study, we found that ESD inhibited the CNE cell proliferation. Cell cycle arrest in S and G2/M phases was also found. Western blotting analysis showed that modulation of cell cycle regulatory proteins was responsible for the ESD-induced cell cycle arrest. Besides, ESD also triggered apoptosis in CNE cells. Dysfunction in mitochondria was found to be associated with the ESD-induced apoptosis as evidenced by the loss of mitochondrial membrane potential (ΔΨm), the translocation of cytochrome c, and the regulation of Bcl-2 family proteins. Despite the Western blotting analysis showed that both intrinsic and extrinsic apoptotic pathways (cleavage of caspases-3, -7, -8, -9, and -10) were triggered in the ESD-induced apoptosis, additional analysis also showed that the induction of apoptosis could be achieved by the caspase-independent manner. Besides, Akt, ERK and JNK pathways were found to involve in ESD-induced cell death. Overall, our findings provided the first evidence that ESD induced cell cycle arrest, and apoptosis in CNE cells. ESD could be a potential chemotherapeutic agent in the treatment of nasopharyngeal cancer (NPC).     

Dieckol exerts anticancer activity in human osteosarcoma (MG-63) cells through the inhibition of PI3K/AKT/mTOR signaling pathway

BackgroundOsteosarcoma (OS) is the most common malignant bone cancer with more metastasis and increased occurrence in children and teen-agers and being responsible for more number of morbidity and mortality worldwide.ObjectiveThe current exploration was planned study the in vitro anticancer actions of dieckol against human OS MG-63 cells via PI3K/AKT/mTOR signaling inhibition.MethodologyThe cytotoxicity of dieckol was scrutinized by MTT assay. Effects of dieckol on the ROS accumulation, apoptotic cell death, and MMP level in the MG-63 cells were studied by respective fluorescence staining assays. The levels of proliferative, inflammatory, and apoptotic markers in the dieckol treated MG-63 cells were scrutinized by marker specific kits. The expressions of PI3K, AKT, and mTOR was assayed by RT-PCR.ResultsThe MTT assay revealed that the dieckol dose dependently prevented MG-63 cells viability and the IC50 was found at 15 µM. Dieckol treatment effectively reduced the MMP level and improved the ROS generation and apoptosis in MG-63 cells. Dieckol also regulated the proliferative (cyclin D1), inflammatory (COX-2, IL-6, TNF-α, and NF-κB), and apoptotic (caspase-3, Bax, Bcl-2) markers in the MG-63 cells. The PI3K/AKT/mTOR signaling in the MG-63 cells were effectively inhibited by the dieckol treatment.ConclusionIn conclusion, our findings from this study recommends that the dieckol could be a talented anticancer candidate for the OS management in the future.     

NFBD1/MDC1 participates in the regulation of G2/M transition in mammalian cells

NFBD1/MDC1 is a large nuclear protein involved in the early cellular response to DNA damage. Upon DNA damage, NFBD1 has an ability to facilitate the efficient DNA repair. In the present study, we have found that, in addition to DNA damage response, NFBD1 plays a critical role in the regulation of G2/M transition. Expression study using synchronized HeLa cells demonstrated that, like the mitotic kinase Plk1, NFBD1 expression level is maximal in G2/M-phase of the cell cycle. siRNA-mediated knockdown of NFBD1 resulted in G2/M arrest as well as simultaneous apoptosis in association with a significant increase in the amounts of γH2AX and pro-apoptotic p73. Since a remarkable down-regulation of mitotic phospho-histone H3 was detectable in NFBD1-knocked down cells, it is likely that knocking down of NFBD1 inhibits G2/M transition. Taken together, our present findings suggest that NFBD1 has a pivotal role in the regulation of proper mitotic entry.     

Mdm4 loss in the intestinal epithelium leads to compartmentalized cell death but no tissue abnormalities

Mdm4 is a critical inhibitor of the p53 tumor suppressor. Mdm4 null mice die early during embryogenesis due to increased p53 activity. In this study, we explore the role that Mdm4 plays in the intestinal epithelium by crossing mice carrying the Mdm4 floxed allele to mice with the Villin Cre transgene. Our data show that loss of Mdm4 (Mdm4^intΔ) in this tissue resulted in viable animals with no obvious morphological abnormalities. However, these mutants displayed increased p53 levels and apoptosis exclusively in the proliferative compartment of the intestinal epithelium. This phenotype was completely rescued in a p53 null background. Notably, the observed compartmentalized apoptosis in proliferative intestinal epithelial cells was not due to restricted Mdm4 expression in this region. Thus, in this specific cellular context, p53 is negatively regulated by Mdm4 exclusively in highly proliferative cells.     

Taiwanin A induced cell cycle arrest and p53-dependent apoptosis in human hepatocellular carcinoma HepG2 cells

Taiwanin A, a lignan isolated from Taiwania cryptomerioides Hayata, has previously been reported to have cytotoxicity against human tumor cells, but the mechanisms are unclear. In this study, we examined the molecular mechanism of cell death of human hepatocellular carcinoma HepG2 cells induced by Taiwanin A. Taiwanin A has been found to induce cell cycle arrest at G2/M phase as well as caspase-3-dependent apoptosis within 24 h. We performed both in vitro turbidity assay and immunofluorescence staining of tubulin to show that Taiwanin A can inhibit microtubule assembly. Moreover, the tumor suppressor protein p53 in HepG2 cells was activated by Taiwanin A within 12 h. Inhibition of p53 by either pifithrin-alpha or by short hairpin RNA which blocks p53 expression attenuates Taiwanin A cytotoxicity. Our results demonstrate that Taiwanin A can act as a new class of microtubule damaging agent, arresting cell cycle progression at mitotic phase and inducing apoptosis through the activation of p53.     

布地奈德通过干预线粒体钙单转运蛋白影响哮喘大鼠气道上皮细胞自噬和屏障功能的机制

摘要 目的：探究布地奈德通过干预线粒体钙单转运蛋白影响哮喘大鼠气道上皮细胞自噬和屏障功能的机制。方法：30只雄性SD大鼠作为研究对象,并根据实验目的分为3组：对照组（正常大鼠,生理盐水干预,n=10）,哮喘组（通过OVA诱导大鼠哮喘模型,n=10）,布地奈德组（气雾剂布地奈德用于治疗过敏性哮喘的大鼠,n=10）。通过钙测定试剂盒和蛋白印迹分析大鼠气道上皮细胞中Ca²⁺的吸收和MCU蛋白表达;TEER和TRITC 荧光分析检测大鼠气道上皮中的屏障功能;免疫组化分析分气道上皮细胞屏障功能相关因子ZO-1、E-cadherin的蛋白表达;ELISAF分析BALF上清液中炎性因子IL-4、IL-5和IL-13的水平;二氢乙锭衍生物和蛋白印迹分析BALF中ROS含量和caspase-3活性。结果：哮喘组较对照组Ca²⁺浓度降低,MCU蛋白表达升高（P<0.05）,布地奈德组较哮喘组Ca²⁺浓度升高,MCU蛋白表达降低（P<0.05）。哮喘组较对照组TEER降低,TRITC升高（P<0.05）,布地奈德组较哮喘组TEER升高,TRITC降低（P<0.05）。哮喘组较对照组ZO-1、E-cadherin的蛋白表达降低（P<0.05）,布地奈德组较哮喘组ZO-1、E-cadherin的蛋白表达升高（P<0.05）。哮喘组较对照组IL-4、IL-5和IL-13的水平升高（P<0.05）,布地奈德组较哮喘组IL-4、IL-5和IL-13的水平降低（P<0.05）。对照组组支气管和肺泡结构未见异常,与对照组相比哮喘组大鼠表现出肺泡间隔增厚,可见的肺毛细血管水肿,以及肺毛细血管和肺泡间隙中的大量炎性细胞浸润（P<0.05）,与哮喘组相比,布地奈德组显著减轻肺部病变的严重程度（P<0.05）。哮喘组较对照组LC3B II/I、ATG5、Beclin-1 和LC3II 的表达升高（P<0.05）,布地奈德组较哮喘组LC3B II/I、ATG5、Beclin-1 和LC3II 的表达降低（P<0.05）。哮喘组较对照组ROS含量和caspase-3活性升高（P<0.05）,布地奈德组较哮喘组ROS含量和caspase-3活性降低（P<0.05）。结论：布地奈德通过调节MCU表达介导气道上皮细胞的屏障完整性和自噬水平,缓解哮喘气道炎症,改善哮喘症状。     

分泌蛋白YKL-40增强肿瘤细胞的上皮间质样转化

目的:分泌糖蛋白YKL-40在多种晚期肿瘤病人的血液中显著升高,提示YKL-40蛋白的血浓度是肿瘤恶变的生物标志物。本课题研究YKL-40重组蛋白和过表达YKL-40肿瘤细胞对肿瘤细胞的上皮间质样转化的作用。方法:构建YKL-40过表达的纤维状乳腺癌细胞系MDA-MB-231和非纤维状结肠癌细胞系HCT-116,观察细胞形态学变化,收集细胞和细胞培养液用于Western Blot(WB)检测YKL-40和上皮间质转化标记蛋白Vimentin和N-cadherin。观察重组蛋白YKL-40对原代MDA-MB-231细胞在无血清条件下的细胞存活影响;另外,用细胞存活试剂盒检测YKL-40过表达HCT-116细胞在无血清的培养液中细胞存活情况。最后,用细胞侵袭试验检测YKL-40过表达MDA-MB-231细胞的侵袭力,并用WB和Zymography来测定细胞分泌MMP9蛋白的表达和酶活性。结果:YKL-40过表达增强MDA-MB-231细胞的形态向上皮间质样转化,并显著提高Vimentin、N-cadherin蛋白的表达,但对HCT-116细胞无法诱导上皮间质样转化。在无血清培养基培养条件下,YKL-40可以增强两种细胞的存活能力,并且YKL-40过表达的MDA-MB-231细胞增强了细胞的侵袭能力,促进了MMP9蛋白表达和蛋白活性。结论:YKL-40可以增加肿瘤细胞的存活力,增强纤维状细胞向上皮间质样转化;并且,YKL-40增加MMP9蛋白表达和活性,增强细胞侵袭力。YKL-40是间质样肿瘤细胞EMT的增强子,此发现为抑制肿瘤恶变提供新靶点。     

Urolithin A induces cell cycle arrest and apoptosis by inhibiting Bcl-2, increasing p53-p21 proteins and reactive oxygen species production in colorectal cancer cells

Colorectal cancer (CRC) is the second most common gastrointestinal cancer globally. Prevention of tumor cell proliferation and metastasis is vital for prolonging patient survival. Polyphenols provide a wide range of health benefits and prevention from cancer. In the gut, urolithins are the major metabolites of polyphenols. The objective of our study was to elucidate the molecular mechanism of the anticancer effect of urolithin A (UA) on colorectal cancer cells. UA was found to inhibit the cell proliferation of CRC cell lines in a dose-dependent and time-dependent manner in HT29, SW480, and SW620 cells. Exposure to UA resulted in cell cycle arrest in a dose-dependent manner along with alteration in the expression of cell cycle–related protein. Treatment of CRC cell lines with UA resulted in the induction of apoptosis. Treatment of HT29, SW480, and SW620 with UA resulted in increased expression of the pro-apoptotic proteins, p53 and p21. Similarly, UA treatment inhibited the anti-apoptotic protein expression of Bcl-2. Moreover, exposure of UA induced cytochrome c release and caspase activation. Furthermore, UA was found to generate reactive oxygen species (ROS) production in CRC cells. These findings indicate that UA possesses anticancer potential and may be used therapeutically for the treatment of CRC.Supplementary InformationThe online version contains supplementary material available at 10.1007/s12192-020-01189-8.     

1950 MHz射频电磁场对SRA01/04细胞周期和凋亡的影响

目的：研究显示射频电磁场与白内障的发生关系密切,为了评价晶状体上皮细胞在射频电磁场诱导的白内障发生中的作 用,本实验探讨了1950 MHz射频电磁场暴露对人眼晶状体上皮细胞株（SRA01/04）细胞周期与凋亡的影响。方法：将处于对数生 长期的SRA01/04 细胞暴露或假暴露于频率为1950 MHz,比吸收率（SAR）为2.79 W/kg 的射频电磁场中,每天暴露1 h,每周暴露 5 天,连续暴露4 周。暴露结束后立即收集细胞,显微镜下观察细胞形态变化,噻唑蓝（MTT）法检测细胞存活力,流式细胞仪 （FCM）检测细胞周期与凋亡。结果：与假辐照组相比,暴露组细胞形态未见明显变化;细胞存活力、细胞周期分布及细胞凋亡率亦 无显著改变（P>0.05）。结论：1950 MHz射频电磁场暴露4 周对SRA01/04 细胞的形态、活力、周期以及凋亡均无明显影响,提示在 本实验条件下1950 MHz 射频电磁场不会诱发白内障的发生。     

survivin在非小细胞肺癌中的表达及其与bc1-2、p63表达的关系

目的:探讨survivin在非小细胞肺癌组织(non small cell lung cancer,NSCLC)中的表达,及其与bc 1 2、p63蛋白表达的相关性。方法:应用二步法免疫组织化法,检测survivin、bc 1-2、p63蛋白在60例NSCLC组织和20例正常肺组织中的表达。结果:肺癌组织中的survivin蛋白阳性率(56．67%)明显高于正常肺组织15%),有显著性差异;(p＜0．05)Ⅲ期surviving蛋白阳性表达率72．73%(16/22)明显高于Ⅰ Ⅱ期survivin47．37%(18/38)。有显著差异;(p＜0．05)survivin蛋白表达与患者年龄、病理类型、组织分化程度,淋巴结转移情况无关(P＞0．05)NSCLC组织bc 1-2蛋白表达阳性、阴性组中,survivin蛋白阳性表达率分别为66．67% (18/27)和48．48%(16/33),两者比较,差异有显著性(P＜0．05);p63蛋白表达阳性、阴性组中,survivin蛋白阳性表达率分别为53．33%(16/30)和60%(18/30),两者比较,差异有显著性(P＜0．05)survivin,蛋白与bc 1-2蛋白的表达呈正相关。survivin蛋白与p63蛋白的表达呈正相关。结论:survivin在NSCLC组织中表达上调,通过抑制细胞凋亡,在NSCLC的发生和发展中起到重要作用。survivin,bc 1-2与p63它们分别在肺癌发生发展过程中不同途径上抑制肺癌细胞的凋亡,对肺癌早期诊断有一定的意义。对3种蛋白进行联合检测,更有利于肺癌的早期诊断和判断肺癌的分化程度、临床分期、淋巴结是否转移及病人的预后。survivin与bc1-2及survivin与p63可能起协同作用,并可能会成为NSCLC基因治疗的新靶点。     

Overexpression of MERTK Receptor Tyrosine Kinase in Epithelial Cancer Cells Drives Efferocytosis in a Gain-of-Function Capacity

MERTK, a member of the TAM (TYRO3, AXL, and MERTK) receptor tyrosine kinases, has complex and diverse roles in cell biology. On the one hand, knock-out of MERTK results in age-dependent autoimmunity characterized by failure of apoptotic cell clearance, while on the other, MERTK overexpression in cancer drives classical oncogene pathways leading to cell transformation. To better understand the interplay between cell transformation and efferocytosis, we stably expressed MERTK in human MCF10A cells, a non-tumorigenic breast epithelial cell line devoid of endogenous MERTK. While stable expression of MERTK in MCF10A resulted in enhanced motility and AKT-mediated chemoprotection, MERTK-10A cells did not form stable colonies in soft agar, or enhance proliferation compared with parental MCF10A cells. Concomitant to chemoresistance, MERTK also stimulated efferocytosis in a gain-of-function capacity. However, unlike AXL, MERTK activation was highly dependent on apoptotic cells, suggesting MERTK may preferentially interface with phosphatidylserine. Consistent with this idea, knockdown of MERTK in breast cancer cells MDA-MB 231 reduced efferocytosis, while transient or stable expression of MERTK stimulated apoptotic cell clearance in all cell lines tested. Moreover, human breast cancer cells with higher endogenous MERTK showed higher levels of efferocytosis that could be blocked by soluble TAM receptors. Finally, through MERTK, apoptotic cells induced PD-L1 expression, an immune checkpoint blockade, suggesting that cancer cells may adopt MERTK-driven efferocytosis as an immune suppression mechanism for their advantage. These data collectively identify MERTK as a significant link between cancer progression and efferocytosis, and a potentially unrealized tumor-promoting event when MERTK is overexpressed in epithelial cells.     

Folate deprivation induces cell cycle arrest at G0/G1 phase and apoptosis in hippocampal neuron cells through down-regulation of IGF-1 signaling pathway

Folate deficiency contributes to impaired adult hippocampal neurogenesis, yet the mechanisms remain unclear. Here we use HT-22 hippocampal neuron cells as model to investigate the effect of folate deprivation (FD) on cell proliferation and apoptosis, and to elucidate the underlying mechanism. FD caused cell cycle arrest at G0/G1 phase and increased the rate of apoptosis, which was associated with disrupted expression of folate transport and methyl transfer genes. FOLR1 and SLC46A1 were (P < 0.01) down-regulated, while SLC19A1 was up-regulated (P < 0.01) in FD group. FD cells exhibited significantly (P < 0.05) higher protein content of BHMT, MAT2b and DNMT3a, as well as increased SAM/SAH concentrations and global DNA hypermethylation. The expression of the total and all the 3 classes of IGF-1 mRNA variants was significantly (P < 0.01) down-regulated and IGF-1 concentration was decreased (P < 0.05) in the culture media. IGF-1 signaling pathway was also compromised with diminished activation (P < 0.05) of STAT3, AKT and mTOR. CpG hypermethylation was detected in the promoter regions of IGF-1 and FOLR1 genes, while higher SLC19A1 mRNA corresponded to hypomethylation of its promoter. IGF-1 supplementation in FD media significantly abolished FD-induced decrease in cell viability. However, IGF-1 had limited effect in rescuing the cell phenotype when added 24 h after FD. Taken together, down-regulation of IGF-1 expression and signaling is involved in FD-induced cell cycle arrest and apoptosis in HT-22 hippocampal neuron cells, which is associated with an abnormal activation of methyl transfer pathway and hypermethylation of IGF-1 gene promoter.