槲皮素通过miR-101/EZH2轴介导的EMT途径改善NSCLC吉西他滨耐药的机制研究 #br#

目的 从微小RNA-101（miR-101）/Zeste基因同源蛋白2（EZH2）轴的角度探究槲皮素逆转非小细胞肺癌 （NSCLC）上皮间质转分化（EMT）及吉西他滨（Gb）耐药性的分子机制。方法 用不同浓度槲皮素培养A549及A549/ Gb，CCK-8法筛选适宜的槲皮素浓度进行后续试验；取A549细胞分为空白组、A549+EMT诱导剂组、A549+槲皮素 组、槲皮素+EMT诱导剂组；A549细胞和A549/Gb分为A549组、A549/Gb组、A549/Gb+槲皮素组、A549/Gb+EMT诱导 剂组、miR-101-inhibitor组、槲皮素+miR-101-inhibitor组、miR-NC组；实时荧光定量PCR（qPCR）检测各组miR-101 表达水平；Western blot法检测EZH2、转化生长因子（TGF）-β1、果蝇母亲DPP同源物4（SMAD4）、p-SMAD4和EMT标 志蛋白E钙黏蛋白（E-cadherin）、N钙黏蛋白（N-cadherin）、波形蛋白（Vimentin）表达；CCK-8法检测各组细胞光密度 （OD）值，并计算半数抑制浓度（IC50）及耐药指数（IR）。结果 槲皮素对A549及A549/Gb的IC50分别为64、128 μmol/L。 EMT诱导剂增强A549细胞EMT特性（N-cadherin及Vimentin表达升高，E-cadherin表达降低）后，A549细胞miR-101 表达降低，EZH2、TGF-β1、p-SMAD4/SMAD4蛋白表达水平，IC50及IR升高（P＜0.05）；槲皮素可抑制A549细胞EMT 特性、降低IC50及IR，并上调miR-101，抑制EZH2、TGF-β1、p-SMAD4/SMAD4蛋白表达（P＜0.05）；EMT诱导剂可逆 转槲皮素的上述作用（P＜0.05）。与 A549 细胞相比，A549/Gb 细胞的 EMT 特性增强，IC50、IR、EZH2、TGF-β1、pSMAD4/SMAD4表达均升高（P＜0.05）；抑制miR-101或EMT 诱导剂处理，均可进一步增强A549/Gb 细胞的EMT 特 性，升高IC50、IR，上调EZH2、TGF-β1、p-SMAD4/SMAD4蛋白表达（P＜0.05）；槲皮素可抑制A549/Gb细胞的EMT进 程、降低其IC50、IR及EZH2、TGF-β1、p-SMAD4/SMAD4蛋白表达（P＜0.05）；抑制miR-101表达可逆转槲皮素的上述 作用（P＜0.05）。结论 槲皮素可通过上调miR-101，抑制EZH2表达，进而抑制EMT进程，降低NSCLC细胞耐药性。     

LINC00691负调控miR-512-5p对非小细胞肺癌细胞增殖、凋亡、迁移和侵袭的影响

目的 探讨LINC00691对非小细胞肺癌（NSCLC）细胞恶性生物学行为的影响。方法 RT-qPCR检测30例NSCLC患者癌组织和对应癌旁组织中LINC00691和miR-512-5p的表达。双荧光素酶报告基因实验验证LINC00691和miR-512-5p的靶向关系。将A549细胞分为si-NC组、si-LINC00691组、miR-NC组、miR-512-5p组、si-LINC00691+anti-miR-NC组和si-LINC00691+anti-miR-512-5p组,MTT和克隆形成实验检测细胞增殖情况,流式细胞术、Transwell检测细胞凋亡、迁移和侵袭,Western blotting检测细胞中Ki67、Cleaved-caspase3、Ecadherin和N-cadherin蛋白表达。结果 LINC00691在NSCLC癌组织中表达水平上升（P<0.05）,而miR-512-5p表达水平下降（P<0.05）。LINC00691在A549细胞中负调控miR-512-5p。沉默LINC00691表达或过表达miR-512-5p可降低A549细胞的存活率、克...     

FASN对Erlotinib耐药细胞株生长的影响及其机制

目的：探讨脂肪酸合酶（fatty acid synthase,FASN）在非小细胞肺癌（non smallcell carcinoma,NSCLC）中,对厄洛替尼（Erlotinib）耐药细胞株生长的影响和其可能的机制。方法： 应用实时定量PCR技术以及Western blot技术分别检测HCC827-EP细胞株与HCC827-ER细胞株中的FASN mRNA和蛋白水平变化。利用小干扰RNA技术阻抑FASN表达后,以SRB法检测干扰前后对HCC827-EP/ER细胞株生长作用的影响。以及阻抑FASN表达后,用实时定量PCR技术和Western blot检测叉头蛋白O1（forkhead box protein O1 ,FoxO1）水平的变化。用活性形式的FoxO1腺病毒感染细胞后,以SRB法检测细胞的存活率。结果： Erlotinib耐药细胞株的FASN表达水平高于亲代细胞株;阻抑FASN的表达,抑制了耐药细胞株的生长;下调FASN的表达,FoxO1 mRNA及蛋白水平均有所升高。上调FoxO1的表达,抑制了耐药细胞株的生长。结论：FASN的高表达发生于Erlotinib耐药细胞株中,下调FASN表达可以抑制耐药细胞株的生长,其可能机制是抑制FASN表达上调了FoxO1,从而抑制了细胞的生长。     

HKB99, an allosteric inhibitor of phosphoglycerate mutase 1, suppresses invasive pseudopodia formation and upregulates plasminogen activator inhibitor-2 in erlotinib-resistant non-small cell lung cancer cells

Acquired resistance to epidermal growth factor receptor tyrosine kinase inhibitors(EGFR-TKIs),such as erlotinib,remains a major challenge in the targeted therapy of non-small cell lung cancer(NSCLC).HKB99 is a novel allosteric inhibitor of phosphoglycerate mutase 1(PGAM1)that preferentially suppresses cell proliferation and induces more apoptosis in acquired erlotinib-resistant HCC827ER cells compared with its parental HCC827 cells.In this study we identified the molecular biomarkers for HKB99 response in erlotinib-resistant HCC827ER cells.We showed that HCC827ER cells displayed enhanced invasive pseudopodia structures as well as downregulated plasminogen activator inhibitor-2(PAI-2).Meanwhile,PAI-2 knockdown by siPAI-2 candidates decreased the sensitivity of HCC827 parental cells to erlotinib.Moreover,HKB99(5μM)preferentially inhibited the invasive pseudopodia formation and increased the level of PAI-2 in HCC827ER cells.Collectively,this study provides new insight into the role of PAI-2 in regulating the sensitivity of erlotinib resistant NSCLC cells to PGAM1 inhibitor.Furthermore,PAI-2 level might be considered as a potential biomarker for predicting the efficacy of the PGAM1 allosteric inhibitor on the erlotinib resistant NSCLC cells.     

LncRNA ITGB2-AS1靶向miR-338-3p对肺癌细胞多西他赛耐药性的影响

目的 研究lncRNA ITGB2-AS1对肺癌细胞增殖、凋亡及多西他赛耐药性的影响及潜在的分子机制.方法 用不同浓度(6.25μg/L、12.5μg/L、25μg/L、50μg/L、100μg/L、200μg/L)多西他赛(DTX)处理肺癌A549和多西他赛耐药细胞A549/DTX细胞,CCK8法测定DTX对A549...     

Sestrin2在miR-19诱导的非小细胞肺癌细胞上皮-间质转化中的表达及六神丸干预研究

目的 探讨Sestrin2在六神丸调控miR-19诱导的非小细胞肺癌细胞上皮-间质转化(epithelial-mesenchymal transition,EMT)中的作用.方法 实验分为对照组、miR-19过表达组、六神丸组.对照组为常规血清培养的A549细胞;miR-19过表达组为常规血清培养的过表达miR-19 A549细胞;六神丸组予以六神丸含药血清干预过表达miR-19 A549细胞.48 h后分别以CCK-8检测A549细胞增殖,Transwell小室检测A549细胞迁移能力,实时荧光定量PCR(quantitative Real-time PCR,qRT-PCR)检测miR-19表达,蛋白印迹检测Sestrin2、PTEN以及EMT标记分子E-cadherin、Vimentin的表达.结果 细胞计数试剂盒-8(cell counting kit-8,CCK-8)检测结果显示,六神丸组A549细胞增殖速度较对照组、miR-19过表达组明显减慢;Transwell小室实验显示,六神丸组A549细胞迁移数量较miR-19过表达组、对照组明显减少;qRT-PCR检测结果显示,六神丸组A549细胞中miR-19表达较miR 19过表达组、对照组明显降低;Western blot检测结果显示,与对照组、miR-19过表达组相比,六神丸组A549细胞中Sestrin2、E-cadherin、PTEN表达明显上调,Vimentin表达明显下调.结论 六神丸能够抑制miR-19诱发的人肺腺癌细胞株A549发生EMT,与激活Sestrin2信号通路有关.     

MiR-142-3p enhances chemosensitivity of breast cancer cells and inhibits autophagy by targeting HMGB1

MiR-142-3p has been reported to act as a tumor suppressor in breast cancer. However, the regulatory effect of miR-142-3p on drug resistance of breast cancer cells and its underlying mechanism remain unknown. Here, we found that miR-142-3p was significantly downregulated in the doxorubicin (DOX)-resistant MCF-7 cell line (MCF-7/DOX). MiR-142-3p overexpression increased DOX sensitivity and enhanced DOX-induced apoptosis in breast cancer cells. High-mobility group box 1 (HMGB1) is a direct functional target of miR-142-3p in breast cancer cells and miR-142-3p negatively regulated HMGB1 expression. Moreover, overexpression of HMGB1 dramatically reversed the promotion of apoptosis and inhibition of autophagy mediated by miR-142-3p up-regulation. In conclusion, miR-142-3p overexpression may inhibit autophagy and promote the drug sensitivity of breast cancer cells to DOX by targeting HMGB1. The miR-142-3p/HMGB1 axis might be a novel target to regulate the drug resistance of breast cancer patients.     

miR-142-3p靶向HMGB1逆转乳腺癌细胞MCF-7对阿霉素的耐药性

目的化疗耐药是乳腺癌复发转移、治疗效果不理想的主要因素。本文探讨miR-142-3p通过靶向高迁移率族蛋白1(high-mobility group box 1,HMGB1)提高乳腺癌化疗敏感性的分子机制。方法实时荧光定量PCR检测人乳腺癌MCF-7细胞及阿霉素耐药株MCF-7/DOX细胞中的miR-142-3p水平;MTT法检测阿霉素(doxorubicin,DOX)处理后各组的增殖情况、流式细胞术检测凋亡率、Western blot检测HMGB1和自噬有关蛋白的表达水平;双荧光素酶报告实验评价miR-142-3p对HMGB1的靶向调控作用。结果阿霉素耐药株MCF-7/DOX细胞中的miR-142-3p水平明显下调。miR142-3p的过度表达增强了乳腺癌细胞对阿霉素的敏感性和提高阿霉素诱导的凋亡比率。HMGB1是乳腺癌细胞中miR-142-3p的直接功能靶点,HMGB1的过表达可以明显解除由miR-142-3p上调所产生的细胞凋亡和自噬抑制。结论miR-142-3p的过表达可能通过抑制自噬靶向HMGB1,增强乳腺癌细胞对阿霉素的化学敏感性。miR-142-3p/HMGB1为提高乳腺癌对药物的敏感性提供了新的靶点,具有一定价值。     

过表达microRNA-144通过靶向调节泛素样PHD和环指结构域1基因诱导非小细胞肺癌凋亡

目的探讨microRNA-144 (miR-144)对非小细胞肺癌(Non-small-cell lung cancer,NSCLC)细胞增殖以及预后的影响。方法通过癌症基因组图谱(The cancer genome atlas,TCGA)检测NSCLC患者组织中miR-144以及泛素样PHD和环指结构域包含1(Ubiquitin like PHD and ring finger domain 1,UHRF1)的表达预后以及两者表达的相关性。采用CCK-8、AO/EB以及γH2A检测不同表达miR-144对NSCLC细胞活性、细胞凋亡、增殖以及DNA损伤的影响,Western blot检测PCNA、Bax、Bcl-2及UHRF1的表达。荧光素酶报告证明miR-144以及UHRF1的靶向关系。结果在NSCLC患者组织中,miR-144表达水平较低,低表达miR-144 NSCLC患者生存时间明显低于高表达miR-144者。UHRF1在NSCLC患者组织中明显升高,且在不同年龄、性别、TNM分期和种族有显著差异。低表达UHRF1非小细胞肺癌患者生存率明显高于高表达UHRF1的NSCLC患者。过表达miR-144能够明显降低A549细胞活性,诱导A549细胞凋亡,增加γH2ax表达;抑制PCNA以及Bax蛋白表达,上调Bcl-2表达。荧光素酶报告证明UHRF1是miR-144的靶基因。同时,低表达miR-144能够使UHRF1表达增加,过表达miR-144能够抑制UHRF1的表达。结论过表达miR-144通过靶向UHRF1诱导A549细胞凋亡以及DNA损伤,参与NSCLC的病理生理过程,进而影响预后。     

The miRNA‐149‐5p/MyD88 axis is responsible for ursolic acid‐mediated attenuation of the stemness and chemoresistance of non‐small cell lung cancer cells

Although the inhibitory roles of ursolic acid (UA) have been established in various tumors, its effects on the stemness of non‐small cell lung cancer (NSCLC) cells are still unclear. Here, we constructed NSCLC cells with paclitaxel resistance (A549‐PR) and showed that A549‐PR exhibited a remarkably stronger stemness than the parental A549 cells, which is evident by the increase of spheroid formation capacity, stemness marker expression, and ALDH1 activity. Additionally, UA significantly reduced the stemness and paclitaxel resistance of A549‐PR cells. Mechanistic investigations revealed that UA inhibited the miR‐149‐5p/MyD88 signaling, which is responsible for UA‐mediated effects on the stemness of A549‐PR cells. Notably, miR‐149‐5p/MyD88 axis promoted the stemness of A549 cells, while inhibition of this axis attenuated the stemness of A549‐PR cells. Therefore, these results suggest that UA could attenuate the stemness and chemoresistance of NSCLC cells through targeting miR‐149‐5p/MyD88 axis.     

MiR-215-5p inhibits the inflammation injury in septic H9c2 by regulating ILF3 and LRRFIP1

MicroRNAs (miRNAs) are small non-coding RNAs (ncRNAs) playing crucial roles in sepsis-induced diseases, including myocardial inflammation. Nevertheless, the expression pattern and role of miR-215-5p in myocardial inflammation are still un-investigated up to now. The purpose of our study is to further inquire the effect of miR-215-5p on lipopolysaccharide (LPS)-activated inflammation injury in H9c2 cells and the possibly associated mechanisms. First of all, LPS-induced H9c2 cells models were constructed and affirmed via detection of pro-inflammatory factors, the viability and apoptosis. MiR-215-5p was overtly down-regulated in LPS-treated H9c2 cells and miR-215-5p overexpression could suppress the inflammation injury. LRRFIP1 was proved to be the target gene of miR-215-5p and meanwhile, miR-215-5p also targeted ILF3 that experimented to bind to and stabilize LRRFIP1. Final rescue assays confirmed that the overexpression of LRRFIP1 or ILF3 rescued the repressive effect of miR-215-5p up-regulation on the inflammation injury in septic H9c2. Totally, miR-215-5p exerted protective function in the inflammation injury in septic H9c2 via targeting ILF3 and LRRFIP1, suggesting an additional treatment method for sepsis-activated myocardial inflammation.     

LincRNA-P21通过靶向miR-17-3p对三阴性乳腺癌细胞迁移和侵袭能力的影响

目的　探讨LincRNA-P21通过靶向miR-17-3p对三阴性乳腺癌（TNBC）细胞迁移和侵袭作用的影响。方法　qPCR检测30例TNBC患者肿瘤组织、癌旁正常组织以及6种乳腺细胞（乳腺正常细胞MCF-10A,乳腺癌细胞MDA-MB-231、MDA-MB-435、MDA-MB-468、BT549、T47D）LincRNA-P21和miR-17-3p的表达。取MDA-MB-231细胞,单独过表达LincRNA-P21（实验设对照组、pcDNA3.1空白组、pcDNA-LincRNA-P21组）,抑制miR-17-3p表达（实验设对照组、miR-17-3p空白组、miR-17-3p抑制剂组）,过表达LincRNA-P21+抑制miR-17-3p表达（实验设对照组、miR-17-3p抑制剂组和pcDNA-LincRNA-P21+miR-17-3p inhibitor组）。CCK-8法检测MDA-MB-231细胞增殖,集落形成实验检测细胞的集落数量,细胞划痕实验检测细胞的迁移能力,Transwell实验检测细胞侵袭能力。Western blot检测E-cadherin和Vimentin蛋白表达。结果　TNBC患者组织中LincRNA-P21的表达明显低于癌旁组织（0.48±0.03 vs. 1.03±0.06,t=11.714,P＜0.01）,miR-17-3p的表达显著高于癌旁组织（2.93±0.17 vs. 1.02±0.04,t=15.593,P＜0.01）。乳腺癌细胞系中LincRNA-P21表达量明显低于MCF-10A,miR-17-3p表达量高于MCF-10A（P＜0.01）。单独过表达LincRNA-P21或抑制miR-17-3p后,MDA-MB-231细胞增殖能力、集落形成数量、迁移和侵袭能力均明显下降。过表达LincRNA-P21的同时抑制miR-17-3p,MDA-MB-231细胞增殖能力、集落形成数量、迁移和侵袭能力出现进一步下降,且上调E-cadherin的表达,抑制Vimentin的表达（P＜0.05）。结论　LincRNA-P21通过竞争性结合miR-17-3p来抑制MDA-MB-231细胞增殖、集落形成、迁移和侵袭。     

MicroRNA-367-3p overexpression represses the proliferation and invasion of cervical cancer cells through downregulation of SPAG5-mediated Wnt/β-catenin signalling

MicroRNA-367-3p (miR-367-3p) has been previously reported as a cancer-related miRNA that is dysregulated in various cancer types and functions either as an oncogenic or as tumour suppressive miRNA. However, whether miR-367-3p is dysregulated in cervical cancer and, further, whether it contributes to the development and progression of the disease remains unknown. Here, our results demonstrated that miR-367-3p expression was markedly decreased in both cervical cancer tissues and cell lines compared with corresponding controls. In vitro experiments revealed that miR-367-3p overexpression repressed the proliferation and invasion of cervical cancer cells. Notably, sperm-associated antigen 5 (SPAG5) was identified as a target gene of miR-367-3p. Moreover, decreased expression of miR-367-3p was correlated with high expression of SPAG5 in cervical cancer tissue specimens. SPAG5 inhibition or miR-367-3p overexpression significantly downregulated Wnt/β-catenin signalling in cervical cancer cells. However, the antitumour effect mediated by miR-367-3p overexpression was partially reversed by SPAG5 overexpression. Overall, these findings demonstrate that miR-367-3p overexpression restricts the proliferation and invasion of cervical cancer cells through targeting SPAG5 to downregulate Wnt/β-catenin signalling, suggesting a mechanism for the tumour suppressive function of miR-367-3p in cervical cancer. Our study highlights the involvement of miR-367-3p/SPAG5/Wnt/β-catenin signalling axis in regulating the malignant progression of cervical cancer.     

microRNA-21促进人肺腺癌细胞A549对顺铂耐药性的研究

目的 研究microRNA-21(miR-21)对人肺腺癌细胞A549对于顺铂耐药性的影响及分子机制.方法 MTT法检测A549和顺铂耐药株A549/CDDP对于顺铂的敏感性,以及在A549细胞中过表达miR-21和在A549/CDDP细胞中抑制miR-21表达后对顺铂敏感性的影响;RT-PCR方法检测A549和A549/CDDP中miR-21表达情况;Western blotting检测PI3K/AKT,核因子-κB (NF-κB)信号通路的激活情况以及在过表达miR-21和抑制miR-21表达后细胞中AKT信号通路的活性变化情况.结果 相对于A549细胞,A549/CDDP对顺铂的敏感性有明显降低,而细胞中miR-21表达则明显增多.Western blotting结果表明:与A549细胞相比,A549/CDDP细胞PI3K/AKT通路被激活,NF-κB通路没有明显变化.过表达miR-21可以激活PI3K/AKT信号通路并提高A549细胞对顺铂的耐药性,相反地,抑制miR-21表达抑制了A549/CDDP细胞中的AKT通路的活化情况并降低了A549/CDDP细胞对顺铂的耐药性.结论 miR-21可以促进肺癌细胞A549对顺铂的耐药性,其机制与PI3K/AKT信号通路的激活有关.     

MicroRNA expression is differentially altered by xenobiotic drugs in different human cell lines

Several noncoding microRNAs (miR or miRNA) have been shown to regulate the expression of drug-metabolizing enzymes and transporters. Xenobiotic drug-induced changes in enzyme and transporter expression may be associated with the alteration of miRNA expression. Therefore, this study investigated the impact of 19 xenobiotic drugs (e.g. dexamethasone, vinblastine, bilobalide and cocaine) on the expression of ten miRNAs (miR-18a, -27a, -27b, -124a, -148a, -324-3p, -328, -451, -519c and -1291) in MCF-7, Caco-2, SH-SY5Y and BE(2)-M17 cell systems. The data revealed that miRNAs were differentially expressed in human cell lines and the change in miRNA expression was dependent on the drug, as well as the type of cells investigated. Notably, treatment with bilobalide led to a 10-fold increase of miR-27a and a 2-fold decrease of miR-148a in Caco-2 cells, but no change of miR-27a and a 2-fold increase of miR-148a in MCF-7 cells. Neuronal miR-124a was generally down-regulated by psychoactive drugs (e.g. cocaine, methadone and fluoxetine) in BE(2)-M17 and SH-SY5Y cells. Dexamethasone and vinblastine, inducers of drug-metabolizing enzymes and transporters, suppressed the expression of miR-27b, -148a and -451 that down-regulate the enzymes and transporters. These findings should provide increased understanding of the altered gene expression underlying drug disposition, multidrug resistance, drug-drug interactions and neuroplasticity.     

OTU deubiquitinase 5 inhibits the progression of non‐small cell lung cancer via regulating p53 and PDCD5

Non‐small cell lung cancer (NSCLC) has the highest morbidity and mortality worldwide. OTU deubiquitinase 5 (OTUD5), a deubiquitinating enzyme, can enhance the stability of p53 and programmed cell death 5 (PDCD5), a protein related to the apoptosis, by deubiquitination. This study aimed to explore the biological function and underlying mechanism of OTUD5 in NSCLC. Western blot and qRT‐PCR were used to detect the expression of OTUD5 protein and mRNA in NSCLC tissues and cells, respectively. RNAi was adopted to construct an OTUD5 low‐expression model while the plasmids overexpressing p53 and PDCD5 were used to establish the overexpression models, respectively. CCK‐8 assay, transwell assay, and apoptosis assay were carried out to analyze the changes in the proliferation, migration, and chemoresistance of A549 and HCC827 cells. The mechanism of OTUD5 in NSCLC was studied by Western blot. Down‐regulated OTUD5 in NSCLC tissues was significantly correlated to a poor prognosis. The knockdown of OTUD5 inactivated p53 and PDCD5, promoting the proliferation and metastasis of NSCLC cells while inhibiting their apoptosis. OTUD5 knockdown also enhanced the resistance of NSCLC cells to doxorubicin and cisplatin. OTUD5 acted as a tumor suppressor in NSCLC by regulating the p53 and PDCD5 pathways.     

核素显像在131 I治疗甲状腺两叶不均等Graves病中的应用价值

目的 探讨核素甲状腺显像在¹³¹I治疗甲状腺两叶不均等Graves病病人中的应用,为临床Graves病的¹³¹I个性化治疗提供依据。方法 选择2010年1月至2015年12月间,在安徽医科大学第二附属医院核医学科首次接受¹³¹I治疗的Graves病病人194例。¹³¹I治疗前均行甲状腺核素显像,勾画甲状腺两叶ROI(region of interest),分别记录两叶放射性计数,根据两叶放射性计数比值差值绝对值分为两组:A组为比值差值≥20%,共40例;B组为差值<5%,共154例。参照¹³¹I治疗格雷夫斯甲亢指南(2013版)又分为:治愈组(完全缓解和甲减)和未愈组(部分缓解、无效和复发)。结果 (1)A、B组病人治愈率分别为75.00%、89.61%,两组治愈率间比较差异有统计学意义(χ²=5.84,P<0.05);(2)40例Graves病病人中甲状腺两叶长轴比值、短轴比值及面积比值的差值与放射性计数比值的差值均无相关性(均P>0.05);(3)A、B两组病人的治疗剂量分别为(474.00±310.80)MBq、(393.68±306.73) MBq,差异无统计学意义(均P>0.05);而A组中未愈组和治愈组与B组未愈组和治愈组比较均差异有统计学意义(均P<0.05)。结论 (1)¹³¹I治疗甲状腺两叶大小不均等Graves病的效果明确,但较均等增大病人的重复治疗机率增大,可适当增加¹³¹I治疗计划量;(2)甲状腺的长轴、短轴及面积等指标不能替代放射性计数来评估Graves病病人甲状腺两叶的不均等性。     

miR-208a促进非小细胞肺癌A549细胞增殖、侵袭和迁移的功能和机制研究

目的　探讨miR-208a对非小细胞肺癌A549细胞增殖、侵袭和迁移的影响及其机制。方法　（1）分别用miR-208a mimic、NC mimic、miR-208a inhibitor和NC inhibitor转染A549细胞。采用荧光定量PCR（qPCR）检测A549细胞中miR-208a过表达和干扰效率;采用CCK-8实验、Transwell实验和划痕实验检测miR-208a对A549细胞增殖、侵袭和迁移的影响。利用生物信息学网站PicTar、miRanda、Targetscan和miRDB预测与miR-208a结合的下游靶基因;构建蛋白酪氨酸磷酸酶受体G（PTPRG）的3′UTR野生型和突变型双荧光素报告酶基因载体,和miR-208a共转染到HEK-293T细胞后观察荧光素酶活性变化。（2）取对数生长期的A549细胞,设置阴性对照组（si-NC组）、PTPRG敲低组（si-PTPRG组）和PTPRG敲低组＋miR-208a干扰组（si-PTPRG＋miR-208a inhibitor组）,采用CCK-8、Transwell实验和划痕实验检测PTPRG对A549增殖、侵袭和迁移的影响,qPCR和Western blot检测3组细胞PTPRG的mRNA和蛋白表达变化。结果　（1）miR-208a mimic组miR-208a基因表达水平、细胞增殖、细胞侵袭和细胞迁移率均高于NC mimic组（P＜0.05）;miR-208a inhibitor组的miR-208a基因表达水平、细胞增殖、细胞侵袭和细胞迁移率均低于NC inhibitor组（P＜0.05）。生物信息学分析显示PTPRG是miR-208a的靶基因,双荧光素报告酶实验结果显示miR-208a过表达能够降低PTPRG 3′UTR野生型的荧光素酶活性,而敲低miR-208a能够增加PTPRG 3′UTR野生型的荧光素酶活性,miR-208a能够与PTPRG的3′UTR序列结合。（2）与si-NC组相比,si-PTPRG组细胞增殖、细胞侵袭和细胞迁移能力明显增高,PTPRG基因和蛋白表达水平下降（P＜0.05）;而与si-PTPRG组相比,si-PTPRG＋miR-208a inhibitor组细胞增殖、细胞侵袭和细胞迁移能力下降,PTPRG基因和蛋白表达水平升高（P＜0.05）。结论　miR-208a可能通过靶向调控PTPRG来促进A549细胞的增殖、侵袭和迁移。