微小RNA在宫颈癌上皮-间充质转化调控中的作用及机制

在许多发展中国家,宫颈癌是女性癌症相关发病和死亡的主要原因。本文阐明了微小RNA(miRNA)在上皮-间充质转化(EMT)中的作用机制,并发现miRNA通过调控EMT在宫颈癌的转移侵袭和肿瘤耐药中发挥重要作用。miR-92a-3p、miR-21、miR-138等可以直接靶向EMT促进宫颈癌转移侵袭;miR-98-5p、miR-204-5p、miR-340等可以通过抑制EMT来抑制宫颈癌转移侵袭;miR-101-3p、miR-106b、miR-4677-3p等可通过内源性竞争机制影响EMT进而影响宫颈癌发生和进展。在宫颈癌肿瘤耐药性中,miR-25-3p可通过靶向Sema4C,将EMT逆转为间质表型上皮样转化,增强宫颈癌耐药细胞对顺铂的敏感性。靶向miRNA可能是一种新型治疗宫颈癌的方法。     

内皮素3对恶性黑素瘤A375细胞上皮基质转化的影响

目的 研究内皮素-3（ET-3）对人恶性黑素瘤（MM）A375细胞上皮基质转化（EMT）的影响。方法 体外培养A375细胞,分别设立3组：空白对照组、100 nmol/L ET-3组、100 nmol/L ET-3和100 μmol/L BQ788（内皮素受体B阻断剂）组。采用Transwell小室检测细胞转移,细胞爬片技术检测细胞形态变化,实时PCR和Western印迹检测上皮基质转化相关分子上皮细胞钙黏蛋白、波形蛋白及转录因子（Twist、 Slug）表达情况,使用方差分析及Scheffe法对结果进行分析。结果 各干预条件中,与空白对照组比较,ET-3可以促进A375细胞的转移,BQ788可阻断该效应（3组穿膜细胞数分别为4.200 ± 0.837、9.400 ± 0.548、3.400 ± 0.894,F = 88.44,P < 0.01）;ET-3 可以促进A375细胞由上皮型向成纤维细胞样形态转变,促进A375上皮细胞钙黏蛋白表达下调（3组分别为0.330 ± 0.002、0.280 ± 0.007、0.420 ± 0.008,F = 329.98,P < 0.01）,波形蛋白表达上调（0.830 ± 0.014、1.160 ± 0.003、0.750 ± 0.030,F = 262.94,P < 0.01）,而BQ788可阻断这种效应。ET-3可以促进上皮基质转化相关转录因子Slug mRNA（F = 376.94,P < 0.01）及Twist mRNA（F = 215.62,P < 0.01）及其蛋白水平（FSlug = 288.87,P < 0.01;FTwist = 156.96,P < 0.05）上的表达上调。结论 ET-3/ETRB通过上调波形蛋白,下调上皮细胞钙黏蛋白的表达,并上调转录因子（Twist、 Slug）的表达,促进黑素瘤A375细胞上皮基质转化。     

EMT在瘢痕疙瘩中的作用研究进展

上皮间质转化(epithelial-to-mesenchymal transition,EMT)是上皮细胞极性丧失且发展为具有迁移和侵袭特性的间充质细胞的动态过程.EMT已被证实与伤口愈合、器官纤维化及癌症发展有关.瘢痕疙瘩的研究都专注于瘢痕疙瘩成纤维细胞,近几年来广大学者们开始注重瘢痕疙瘩角化上皮细胞侵袭周围正常皮肤...     

上皮-间质转化在皮肤肿瘤中的研究进展

上皮-间质转化(epithelial-mesenchymal transition,EMT)是细胞由上皮状态向间质状态转变,并使其获得迁移和侵袭性行为的生物学过程.皮肤肿瘤是起源于皮肤组织的一组肿瘤性疾病,其中以鳞状细胞癌(squamous cell carcinoma,SCC)、黑素瘤(malignant melan...     

转化生长因子β对A431细胞上皮向间质转换相关分子表达的影响

目的 研究转化生长因子β（TGF-β）对人表皮鳞状细胞癌A431细胞上皮向间质转换（EMT）相关分子表达的影响。 方法 用含7.5 μg/L TGF-β处理A431细胞72 h后观察细胞形态,采用实时定量PCR和Western印迹分别在mRNA水平和蛋白水平检测EMT相关基因E钙黏着蛋白、N钙黏着蛋白、波形蛋白、β联蛋白表达水平的变化。未经TGF-β处理的A431细胞为对照组。 结果 TGF-β处理后,A431细胞独立分散,呈梭形。实时定量PCR结果显示,E钙黏着蛋白表达量为未经TGF-β处理组的31.7%,N钙黏着蛋白、波形蛋白、β联蛋白表达量分别为未经处理组的2.475、11.340、2.615倍;Western印迹结果,N钙黏着蛋白、波形蛋白、β联蛋白表达量上升,E钙黏着蛋白表达量下降。 结论 TGF-β可诱导A431细胞产生EMT现象,TGF-β表达量上升,可能参与表皮鳞状细胞癌发生侵袭、转移。     

全反式维A酸对恶性黑素瘤A375细胞上皮-间质转化相关分子表达的影响

【摘要】 目的 研究全反式维A酸（ATRA）对人恶性黑素瘤A375细胞中上皮-间质转化（EMT）相关分子表达的影响。方法 用含10 μmol/L ATRA的DMEM培养基和DMEM培养基分别处理A375细胞24和48 h作为ATRA-1组和ATRA-2组、对照1组和对照2组,采用实时定量PCR法检测EMT相关基因上皮钙黏着蛋白、神经钙黏着蛋白、波形蛋白、β联蛋白mRNA的表达。Western印迹法检测ATRA-1组、ATRA-2组、对照1组中上述蛋白的相对表达量,直接免疫荧光法检测上皮钙黏着蛋白和波形蛋白的荧光强度。统计分析采用两因素方差分析、单因素方差分析及LSD-t检验。结果 ATRA-1组和ATRA-2组分别与对照1组和对照2组相比,上皮钙黏着蛋白mRNA的表达均显著增加（F = 13.148、31.529,P < 0.05）,而神经钙黏着蛋白、波形蛋白、β联蛋白mRNA的表达均显著降低（均P < 0.05）;ATRA-2组上皮钙黏着蛋白mRNA的表达显著高于ATRA-1组（F = 13.148,P < 0.05）,而其他3种蛋白mRNA的表达显著低于ATRA-1组（均P < 0.05）;对照1组与对照2组上述蛋白的mRNA表达差异无统计学意义（均P > 0.05）。Western印迹法显示,与对照1组相比,ATRA-1组和ATRA-2组上皮钙黏着蛋白表达上调,而神经钙黏着蛋白、波形蛋白、β联蛋白表达均下调（均P < 0.05）;与ATRA-1组相比,ATRA-2组上皮钙黏着蛋白表达上调（P < 0.05）,神经钙黏着蛋白、波形蛋白、β联蛋白表达下调（均P < 0.05）。直接免疫荧光显示,ATRA-1组、ATRA-2组上皮钙黏着蛋白的荧光强度（6.23 ± 0.08、10.37 ± 0.13）显著高于对照1组（2.37 ± 0.14,均P < 0.05）,而波形蛋白的荧光强度（15.17 ± 0.18、10.29 ± 0.03）显著低于对照1组（50.16 ± 0.26,均P < 0.05）,抑制上皮向间质转化。结论 ATRA可上调A375细胞中上皮钙黏着蛋白的表达,降低神经钙黏着蛋白、波形蛋白、β联蛋白的表达,可能抑制A375细胞发生EMT现象。     

角化性皮肤病

20110564毛囊角化病一家系中ATP2A2基因新的剪接突变/孙忠辉(温州医学院),李明,张国龙∥中国麻风皮肤病杂志.-2010,26(11).-762~764为了检测毛囊角化病一家系中ATP2A2基因新的剪接突变,提取家系中2例患者和2名正常人外周血DNA,采用PCR对ATP2A2基因进行扩增,并对其产物进行测序,同时设立正常对照。结果发现患者ATP2A2基因的第13号内含子第1761+2位碱基由T转化为C。认为该家系发病可能是由ATP2A2基因发生剪接突变所致。图3参8(赵恩兵)20     

角朊细胞基因表达的转录调控

角朊细胞的增殖和分化是一个受到精细调节的过程，并伴随着一系列形态学和生化改变，最终形成角质细胞，这就必然涉及到许多结构基因的同时活化与灭活，即基因表达的调控，而转录水平的调控尤为重要。现已发现许多转录因子如ＡＰＩ、ＡＰ２、Ｓｐ１、ＰＯＵ结构域及Ｃ／ＥＢＰ等可调节角朊细胞基因的表达。     

上皮样血管瘤1例

报告1例发生于头部的上皮样血管瘤。患者男，40岁。左侧头皮多发性丘疹、结节7年组织病理检查示：病变由许多增生的大小不等的血管组成，血管内皮细胞较肥大，呈立方形或上皮样突向腔内形成鞋钉状，血管间散在灶性炎性细胞浸润，以淋巴细胞为主，可见少量嗜酸粒细胞，病理诊断为上皮样血管瘤。     

多发性家族性毛发上皮瘤致病基因的确定

目的 对多发性家族性毛发上皮瘤一家系进行基因定位及候选基因突变检测。方法 共用18对覆盖9p21和16q12-q13的微卫星标记对一个多发性家族性毛发上皮瘤家系进行局部基因组扫描,并用Linkage软件进行两点参数连锁分析,最后PCR扩增CYLD1基因的17个编码外显子及其邻近剪接子并进行双向直接测序。结果 ①两点参数连锁分析在常染色体显性遗传模式下,外显率为99.9%、基因频率0.00001时在D16S3068位点处得出LOD值=3.31(θ=0.00),排除与9号染色体连锁;②突变分析在CYLD1基因第18号外显子出现连续的4个碱基缺失,即c.2355-2358delCAGA。结论 多发性家族性毛发上皮瘤存在着遗传异质性,本家系的致病基因位于16q12-q13,而不在9p21。     

SMAD inhibition attenuates epithelial to mesenchymal transition by primary keratinocytes in vitro

Epithelial to mesenchymal transition (EMT) is a process whereby epithelial cells undergo transition to a mesenchymal phenotype and contribute directly to fibrotic disease. Recent studies support a role for EMT in cutaneous fibrotic diseases including scleroderma and hypertrophic scarring, although there is limited data on the cytokines and signalling mechanisms regulating cutaneous EMT. We investigated the ability of TGF‐β and TNF‐α, both overexpressed in cutaneous scleroderma and central mediators of EMT in other epithelial cell types, to induce EMT in primary keratinocytes and studied the signalling mechanisms regulating this process. TGF‐β induced EMT in normal human epidermal keratinocytes (NHEK cells), and this process was enhanced by TNF‐α. EMT was characterised by changes in morphology, proteome (down‐regulation of E‐cadherin and Zo‐1 and up‐regulation of vimentin and fibronectin), MMP secretion and COL1α1 mRNA expression. TGF‐β and TNF‐α in combination activated SMAD and p38 signalling in NHEK cells. P38 inhibition with SB203580 partially attenuated EMT, whereas SMAD inhibition using SB431542 significantly inhibited EMT and also reversed established EMT. These data highlight the retained plasticity of adult keratinocytes and support further studies of EMT in clinically relevant in vivo models of cutaneous fibrosis and investigation of SMAD inhibition as a potential therapeutic intervention.     

Epilysin (MMP-28)--structure, expression and potential functions

Epilysin (MMP-28) is the newest member of the matrix metalloproteinase (MMP) family of extracellular proteases. Together the MMPs can degrade almost all components of the extracellular matrix (ECM). MMPs also regulate cell behaviour by releasing growth factors and biologically active peptides from the ECM by modulating cell surface receptors and adhesion molecules and by regulating the activity of mediators of the inflammatory pathways. Epilysin differs from most other MMPs as it is expressed in a number of normal tissues, suggestive of functions in tissue homeostasis. The epilysin homologue in Xenopus laevis (XMMP-28) is expressed in neural tissues, where it cleaves the neural cell adhesion molecule. Enhanced expression of epilysin has been observed in basal keratinocytes during wound healing and in different forms of cancer. There are, however, also reports on the downregulation of epilysin in malignant cells. The roles of epilysin in cancer seem to vary based on tumor type and stage of the disease. Importantly, epilysin can induce stable epithelial to mesenchymal transition (EMT) when overexpressed in epithelial lung carcinoma cells. Transforming growth factor beta (TGF-beta) is a crucial mediator of this process, which was characterized by the loss of E-cadherin and increased cell migration and invasion. Current results suggest a plausible interaction between epilysin and TGF-beta also under physiological circumstances, where epilysin activity may not induce EMT but, instead, trigger less permanent changes in TGF-beta signalling and cell motility.     

Matrix metalloproteinases contribute to kidney fibrosis in chronic kidney diseases

Matrix metalloproteinases (MMPs) are members of the neutral proteinase family. They were previously thought to be anti-fibrotic because of their ability to degrade and remodel of extracellular matrix. However, recent studies have shown that MMPs are implicated in initiation and progression of kidney fibrosis through tubular cell epithelial–mesenchymal transition (EMT) as well as activation of resident fibroblasts, endothelial-mesenchymal transition (EndoMT) and pericyte-myofibroblast transdifferentiation. Interstitial macrophage infiltration has also been shown to correlate with the severity of kidney fibrosis in various chronic kidney diseases. MMPs secreted by macrophages, especially MMP-9, has been shown by us to be profibrotic by induction of tubular cells EMT. EMT is mainly induced by transforming growth factor-β (TGF-β). However, MMP-9 was found by us and others to be up-regulated by TGF-β1 in kidney tubular epithelial cells and secreted by activated macrophages, resulting in EMT and ultimately kidney fibrosis. Therefore, MMP-9 may serve as a potential therapeutic target to prevent kidney fibrosis in chronic kidney disease. This review, by a particular focus on EMT, seeks to provide a comprehensive understanding of MMPs, especially MMP-9, in kidney fibrosis.     

Twist1 in tumor cells and α‐smooth muscle actin in stromal cells are possible biomarkers for metastatic giant basal cell carcinoma

We previously reported a case of giant basal cell carcinoma (BCC) in a 75‐year‐old Japanese man, who subsequently developed a pulmonary metastasis. With regard to the pathogenesis of metastasis of BCC, recently, it has been reported that high levels of expression of Twist1 and N‐cadherin in primary and metastatic tumor cells, suggesting that Twist1 expression and an epithelial–mesenchymal transition (EMT) of tumor cells are important for the promotion of tumor invasion and subsequent metastasis. In this report, we identified the expressions of Twist1 in tumor cells and α‐smooth muscle actin (α‐SMA) in stromal cells in the primary and metastatic sites of giant BCC. These results suggest that Twist1‐induced EMT of tumor cells might have been associated with distant organ metastasis in our case, and the presence of α‐SMA‐positive myofibroblasts surrounding a BCC nest can be one of hallmarks of the aggressiveness of BCC.     

Human papillomavirus 16 E6/E7‐immortalized human gingival keratinocytes with epithelial mesenchymal transition acquire increased expression of cIAP‐1, Bclx and p27Kip1

Abstract: Epithelial mesenchymal transition (EMT) has been implicated in neoplastic invasion and metastasis. We have previously generated six cell lines from human gingival mucosal keratinocytes immortalized by E6/E7 of human papillomavirus type 16. Ldk and NuB1 lines represented EMT phenotype and other four lines represented cobblestone non‐EMT phenotype. These cell lines were utilized as model cells to determine whether inhibitors of apoptosis proteins and cell‐cycle regulators were molecular players during EMT. EMT cells exhibited increased expression of vimentin, vascular endothelial growth factor (VEGF) receptor1 and the ability to form tubules on Matrigel as well as to grow anchorage independently. We found that EMT cells expressed significantly elevated levels of cIAP‐1, Bclx and p27^kip higher than non‐EMT cells. These proteins could therefore be used as intrinsic indicators of EMT of human gingival keratinocytes.     

Hmga2 is dispensable for cutaneous squamous cell carcinoma

Hmga2 functions as a chromatin‐associated factor during development, but is not expressed in most adult tissues. Expression of Hmga2 in adult tissues has been associated with a variety of human cancers. Numerous studies have implicated Hmga2 in epithelial‐to‐mesenchymal transition (EMT) and cancer progression through gain of function studies, but it is unclear whether Hgma2 is necessary for EMT, tumor formation or tumor progression. We deleted Hmga2 in two mouse models of squamous cell carcinoma and found this gene to be dispensable. In fact, EMT, tumor initiation and progression all appeared to be mostly unaffected by the absence of Hmga2. Tumors lacking the ability to induce Hmga2 proceeded to initiate cutaneous spindle cell and squamous cell carcinomas with all the typical pathological and molecular hallmarks of these cancers.     

Cancer stem-like cells enriched with CD29 and CD44 markers exhibit molecular characteristics with epithelial–mesenchymal transition in squamous cell carcinoma

Increasing evidences have indicated that only a phenotypic subset of cancer cells, termed as the cancer stem cells (CSCs), is capable of initiating tumor growth and provide a reservoir of cells that cause tumor recurrence after therapy. Epithelial–mesenchymal transition (EMT), a cell type change from an epithelial cobblestone phenotype to an elongated fibroblastic phenotype, plays a critical role not only in tumor metastasis but also in tumor recurrence and contributes to drug resistance. Accumulating evidence has shown that cells with an EMT phenotype are rich sources for CSCs, suggesting a biological link between EMT and CSCs; thus study on the link will help understand the cellular and molecular mechanisms of tumor metastasis and drug resistance. CD29 is involved in EMT through cross-talk with cadherins and CD44 has been reported as a successful used marker for CSCs. Here, we try to address whether combination of CD29 and CD44 could be used to identify cancer stem-like cells undergoing EMT in squamous cell carcinoma (SCC) and compare the molecular differences between CD29high/CD44high and CD29low/CD44low cells in SCC. Expression pattern of CD29 and CD44 was analyzed in tissues of skin SCC and cultured A431 cells by immunostaining. Subtype cells of CD29high/CD44high and CD29low/CD44low A431 were sorted by fluorescence-activated cell sorting and proliferating abilities were assayed by cell counting, colony forming and tumorigenicity in NOD/SCID mice. Finally, to probe more deeply into the molecular differences between CD29high/CD44high and CD29low/CD44low A431 cells, gene microarray analysis was applied to compare gene expression profiling. Staining of CD29 and CD44 showed similar heterogeneous expression pattern with positive cells located in the invasion front of SCC tissue as well as in cultured A431 cells. Sorted CD29high/CD44high A431 cells had higher proliferating ability in vitro and in NOD/SCID mice as compared with CD29low/CD44low cells. Gene profiling identified differentiated gene expressions between CD29high/CD44high and CD29low/CD44low A431 cells. These genes are involved in cell cycle, cell malignant transformation, metastasis, drug resistance and EMT, implying that CD29high/CD44high cells have properties of CSCs and EMT. Our present results demonstrated heterogeneous gene expression patterns and different biological behavior in SCC. Combination of CD29 and CD44 can be used as markers to enrich CSCs in human SCC. Moreover, CD29high/CD44high cells exhibit molecular characteristics of EMT, suggesting that CSC-associated pathways were involved in EMT. Studies on correlation of CSCs and the cells undergoing EMT may explain some aspects of tumor progression and drug resistance.