Overexpression of epithelial-mesenchymal transition-related markers according to cell dedifferentiation: clinical implications as an independent predictor of poor prognosis in cholangiocarcinoma

Ryu, H.S.,Chung, J.H.,Lee, K.,Shin, E.,Jing, J.,Choe, G.,Kim, H.,Xu, X.,Lee, H.E.,Kim, D.G.,Lee, H.,Jang, J.J. W. B. Saunders Co ; Centrum Philadelphia 2012 Human pathology Vol.43 No.12

Although increased evidence has suggested that epithelial-mesenchymal transition has been implicated in cancer invasion and is associated with poor prognosis, its significance in cholangiocarcinoma remains unclear. We evaluated the levels of expression of epithelial-mesenchymal transition-related genes and proteins in 2 established human cholangiocarcinoma cell lines with different morphological characteristics and performed transwell cell invasion assays. Furthermore, we investigated the association between altered expression of 6 epithelial-mesenchymal transition-related proteins and clinical outcomes in human cholangiocarcinoma patients (n = 119) by immunohistochemistry using a tissue microarray approach. Comparative analysis of protein and messenger RNA expression revealed that the cell line with less differentiation (JCK) showed increased expression of mesenchymal markers and zinc-finger proteins and decreased expression of epithelial markers. The invasion activity of JCK cells was significantly higher than that of cells from OZ cell lines. Tissue microarray analysis revealed that the combined expression pattern of 6 epithelial-mesenchymal transition-related proteins predicted shortened disease-free survival (13.0 versus 22.0 months, P = .033) and overall survival (23.0 versus 63.0 months, P = .003) and was confirmed as an independent unfavorable prognostic factor for survival in multivariate survival analysis (disease-free survival, P = .028 for the 3 epithelial-mesenchymal transition-related markers; overall survival, P = .010 for the 6 epithelial-mesenchymal transition-related markers). In conclusion, our results suggest that altered expression of a number of epithelial-mesenchymal transition-related genes in tumor cells with poor differentiation may explain their increased invasive ability. Our results also suggest that altered expression of a suite of epithelial-mesenchymal transition-related proteins could be used as a tool to predict poor outcomes in human cholangiocarcinoma patients.

P-129 Histone deacetylase 3 induces alveolar epithelial cell mesenchymal transition under hypoxic condition

박정웅,손은숙,정성환,김세희 대한결핵 및 호흡기학회 2017 대한결핵 및 호흡기학회 추계학술대회 초록집 Vol.124 No.-

Background & Aims: Epithelial-mesenchymal transition (EMT), a process by which epithelial cells undergo a phenotypic conversion that gives rise to myofibroblasts, plays a pivotal process in the development of pulmonary fibrosis. From recent studies that hypoxia induces EMT of alveolar epithelial cells, and histone deacetylases (HDACs) are aberrantly overexpressed in basal cells of idiopathic pulmonary fibrosis (IPF) lungs, we aimed to elucidate whether hypoxia promotes mesenchymal transition of alveolar epithelial cell through activating HDACs. Methods: To evaluate the effect of HDACs in alveolar EMT, we performed quantitative real-time PCR, Western blot analysis, immunocytochemistry using siRNA transfection in human and rat alveolar epithelial cells, and immunohistochemistry using a bleomycin-induced fibrosis mouse model. Results: Hypoxia induced expression of alveolar EMT markers in A549 and RLE-6TN cells. Specifically, exposure to hypoxia decreased an epithelial marker, E-cadherin, and reversely increased expression of mesenchymal markers, Fibronectin and α -SMA in both cell lines. Additionally, a HDAC inhibitor, TSA was able to repress hypoxic effect such as loss of E-cadherin and gain of Fibronectin in alveolar epithelial cell. Among HDAC family, inhibition of HDAC3 using siRNA increased E-cadherin, and decreased Fibronectin, Snail, α-SMA in hypoxic conditions, and this process is dependent on Akt pathway. Conclusions: Histone deacetylase 3 induces mesenchymal transition of alveolar epithelial cell via repressing Akt via repressing Akt pathway in hypoxic conditions.

Histone deacetylase 3 induces alveolar epithelial cell mesenchymal transition via Akt pathway under hypoxic condition

박정웅,손은숙,정성환,김세희 대한결핵 및 호흡기학회 2018 대한결핵 및 호흡기학회 추계학술대회 초록집 Vol.126 No.-

Background and Aims: Epithelial-mesenchymal transition (EMT), a process by which epithelial cells undergo a phenotypic conversion to myofibroblasts, plays a pivotal role of developing chronic lung diseases such as idiopathic pulmonary fibrosis (IPF) and chronic obstructive pulmonary disease (COPD). Recent studies showed that hypoxia induces EMT of alveolar epithelial cells and histone deacetylases (HDACs) are aberrantly overexpressed in basal cells of IPF lungs. Furthering on these studies, we aimed to elucidate whether hypoxia promotes mesenchymal transition of alveolar epithelial cell through activating HDACs. Methods: To evaluate the effect of HDAC3, we performed quantitative real-time PCR, Western blot analysis, immunocytochemistry using siRNA transfection and HDAC3 lentivirus in human and rat alveolar epithelial cells under hypoxic conditions. Results: Exposure to hypoxia decreased an epithelial marker, E-cadherin, and reversely increased the expression of mesenchymal markers, Fibronectin and α-SMA in A549 and RLE-6TN cells. Among HDAC family, inhibition of HDAC3 using siRNA increased E-cadherin, and decreased Fibronectin, Snail, α-SMA, ZEB1, N-cadherin, Vimentin, and β-catenin in hypoxic conditions. Additionally, overexpression of HDAC3 using lentivirus repressed E-cadherin, and reversely induced mesenchymal markers under hypoxia. This process which HDAC3 promotes alveolar epithelial-mesenchymal transition was dependent on AKT pathway. Conclusions: Histone deacetylase 3 induces mesenchymal transition of alveolar epithelial cell via AKT pathway in hypoxic conditions.

Early Growth Response 1 Induces Epithelial-to-mesenchymal Transition via Snail

Hyun Min Jeon(전현민),Su Yeon Lee(이수연),Min Kyung Ju(주민경),Hye Gyeong Park(박혜경),Ho Sung Kang(강호성) 한국생명과학회 2013 생명과학회지 Vol.23 No.8

Epithelial-to-mesenchymal transition (EMT)는 embryogenesis에서 중요한 역할을 하며 tumor metastasis, invasion에도 관여함으로써 tumor progression 및 aggressiveness에 기여한다. EMT는 EMT hallmark인 epithelial E-cadherin의 발현 감소와 mesenchymal-like cell morphology를 획득함으로써 epithelial cell polarity를 잃어버리는 특징을 가지고 있다. O₂-, H₂O₂, OH-와 같은 활성산소가 EMT를 유도하는 것으로 알려져 있다. Snail이 E-cadherin의 발현을 억제함으로써 ROS에 의한 EMT에 관여하는 것으로 알려져 있으나, 그 기작은 완전히 밝혀져 있지 않다. 본 연구에서는, noninvasive breast tumor cell line인 MCF-7 세포에 Egr-1을 과발현시킨 후 그 영향을 조사하였다. Egr-1이 과발현되면, MCF-7 세포는 epithelial cell polarity를 잃고 spindle-shaped로 변화되므로, Egr-1이 EMT를 유도할 가능성이 대두되었다. 또한 Snail이 Egr-1에 의한 EMT에 관여함을 확인하였다. 나아가, 본 연구진은 Egr-1-Snail axis가 ROS에 의해 활성화 되고, ROS에 의한 EMT에서 중요한 역할을 함을 발견하였다. The epithelial-to-mesenchymal transition (EMT) plays an essential role in embryogenesis and is involved in tumor metastasis and invasion; it significantly contributes to tumor progression and aggressiveness. The EMT is characterized by a loss of epithelial cell polarity as a result of the reduced expression of epithelial E-cadherin, a hallmark of the EMT, and the acquisition of mesenchymal-like cell morphology. Reactive oxygen species (ROS) such as O₂ -, H₂O₂, and OH- have been demonstrated to induce the EMT; although Snail is involved in ROS-induced EMT by transcriptionally repressing E-cadherin, its mechanism is not fully understood. In this study, we examined the effects of early growth response 1 (Egr-1) overexpression in noninvasive breast tumor cell line MCF-7 cells. Upon Egr-1 overexpression, MCF-7 cells lost epithelial cell polarity and became more spindle-shaped, indicating that Egr-1 may induce EMT. We found that Snail is implicated in Egr-1 induced EMT. We further demonstrate that the Egr-1-Snail axis is activated by ROS and plays a critical role(s) in ROS-induced EMT.

Epithelial-mesenchymal Transition and Its Role in the Pathogenesis of Colorectal Cancer

Zhu, Qing-Chao,Gao, Ren-Yuan,Wu, Wen,Qin, Huan-Long Asian Pacific Journal of Cancer Prevention 2013 Asian Pacific journal of cancer prevention Vol.14 No.5

Epithelial-to-mesenchymal transition (EMT) is a collection of events that allows the conversion of adherent epithelial cells, tightly bound to each other within an organized tissue, into independent fibroblastic cells possessing migratory properties and the ability to invade the extracellular matrix. EMT contributes to the complex architecture of the embryo by permitting the progression of embryogenesis from a simple single-cell layer epithelium to a complex three-dimensional organism composed of both epithelial and mesenchymal cells. However, in most tissues EMT is a developmentally restricted process and fully differentiated epithelia typically maintain their epithelial phenotype. Recently, elements of EMT, specially the loss of epithelial markers and the gain of mesenchymal markers, have been observed in pathological states, including epithelial cancers. Increasing evidence has confirmed its presence in human colon during colorectal carcinogenesis. In general, chronic inflammation is considered to be one of the causes of many human cancers including colorectal cancer(CRC). Accordingly, epidemiologic and clinical studies indicate that patients affected by ulcerative colitis and Crohn's disease, the two major forms of inflammatory bowel disease, have an increased risk of developing CRC. A large body of evidence supports roles for the SMAD/STAT3 signaling pathway, the NF-kB pathway, the Ras-mitogenactivated protein kinase/Snail/Slug and microRNAs in the development of colorectal cancers via epithelial-tomesenchymal transition. Thus, EMT appears to be closely involved in the pathogenesis of colorectal cancer, and analysis refered to it can yield novel targets for therapy.

Basic, HCC basic : PE-107 ; EGR and WT1 control the differentiation of BM-derived mesenchymal stem cells into hepatic with mesenchymal to epithelial transition

( Hye Lim Kim ),( Jung Hoon Cha ),( Na Ri Park ),( Won Hee Hur ),( Si Hyun Bae ),( Jong Young Choi ),( Seung Kew Yoon ) 대한간학회 2012 춘·추계 학술대회 (KASL) Vol.2012 No.-

Background: Epithelial-to-mesenchymal transition drives mammary epithelial cells to de-differentiate into mammary stem cells which are mesenchymal-like. MET and the reverse process, epithelial mesenchymal transition (EMT), both occur in normal tissue, including gastrulating and regenerating tissue. MET in hepatic stem cells is important to multiple processes, including hepatic differentiation, liver development and regeneration. In this study, We investigated regulator of EMT/MET process in the hepatic differentiation. Methods: Based on the 2 step protocol, bone marrow-derived mesenchymal stem cells were cultured for hepatic differentiation. Pas staining and Urea production test were performed to verify the functionality of hepatic differentiation. To search for novel proteins in MSC before and after differentiation, protein/ membrane array was performed. EMT/ MET markers were identified during the hepatic differentiation of MSC by Western blot. Results: During differentiation proceeds, from day 1 to day 28, stem cells change in shape from spindle to polygon with larger in size and tighter intercellular connections. Intracellular glycogen and urea production was significantly increased from day 1 to day 28. The mRNA expression of liver-specific genes was up-regulated. In order to study key regulators in hepatic differentiation, we performed protein/membrane array. Novel factors of EGR1, EGR2 and WT1 were significantly in differentiated MSCs increased during the culture compared to undifferentiated MSCs. The induction of MET markers, E-cadherin and CK18 and the reduction of EMT markers, vimentin and N-cadherin, were exhibited during hepatic differentiation. Conclusions: We identified the novel factors of EGR and WT1 which were expected to play a role in hepatic differentiation. In the future, the acceleration of differentiation by novel factors and EMT/MET process should be studied further.

HCV : PE-107 ; EGR and WT1 control the differentiation of BM-derived mesenchymal stem cells into hepatic with mesenchymal to epithelial transition

( Hye-lim Kim ),( Jung Hoon Cha ),( Na Ri Park ),( Won Hee Hur ),( Si Hyun Bae ),( Jong Young Choi ),( Seung Kew Yoon ) 대한간학회 2012 춘·추계 학술대회 (KASL) Vol.2012 No.1

Background: Epithelial-to-mesenchymal transition (EMT) drives mammary epithelial cells to de-differentiate into mammary stem cells which are mesenchymal-like. MET and the reverse process, epithelial mesenchymal transition (EMT), both occur in normal tissue, including gastrulating and regenerating tissue. MET in hepatic stem cells is important to multiple processes, including hepatic differentiation, liver development and regeneration. In this study, We investigated regulator of EMT/MET process in the hepatic differentiation. Methods: Based on the 2 step protocol, bone marrow-derived mesenchymal stem cells were cultured for hepatic differentiation. Pas staining and Urea production test were performed to verify the functionality of hepatic differentiation. To search for novel proteins in MSC before and after differentiation, protein/ membrane array was performed. EMT/ MET markers were identified during the hepatic differentiation of MSC by Western blot. Results: During differentiation proceeds, from day 1 to day 28, stem cells change in shape from spindle to polygon with larger in size and tighter intercellular connections. Intracellular glycogen and urea production was significantly increased from day 1 to day 28. The mRNA expression of liver-specific genes was up-regulated. In order to study key regulators in hepatic differentiation, we performed protein/membrane array. Novel factors of EGR1, EGR2 and WT1 were significantly in differentiated MSCs increased during the culture compared to undifferentiated MSCs. The induction of MET markers, E-cadherin and CK18 and the reduction of EMT markers, vimentin and N-cadherin, were exhibited during hepatic differentiation. Conclusions: We identified the novel factors of EGR and WT1 which were expected to play a role in hepatic differentiation. In the future, the acceleration of differentiation by novel factors and EMT/MET process should be studied further.

Basic, Research : EGR1 Control the Differentiation of BM-derived Mesenchymal Stem Cells into Functional Hepatocyte through Mesenchymal-Epithelial Transition

( Hye Lim Kim ),( Si Hyun Bae ),( Jung Hoon Cha ),( Na Ri Park ),( Jong Young Choi ),( Seung Kew Yoon ) 대한간학회 2013 춘·추계 학술대회 (KASL) Vol.2013 No.1

Background: Mesenchymal-Epithelial transition (MET) in hepatic stem cells is important to multiple processes, including hepatogenic differentiation and liver development, regeneration. Early growth response (EGR) 1, transcription factor of early growth response gene family, is induced in the responses to a number of growth and differentiation factors. The aim of this study is to investigate that EGR1 acts as a key regulator of EMT/MET in the process of hepatogenic differentiation from bone marrow-derived mesenchymal stem cells (BM-MSCs). Methods: Pas staining and Urea production test were performed to verify the functionality of hepatogenic differentiation. To search for novel proteins in BM-MSCs before and after differentiation, protein/DNA array was performed. Immunohistochemistry were identified the enhanced expression of EGR1 and MET markers, accompanied by decreased expression of EMT markers. To determine the effect of EGR1, BMMSCs were infected by lenti-shEGR1. To determine the role of EGR1 in BM-MSCs which were infected by lenti-shEGR1, MET/EMT markers during the hepatogenic differentiation were identified by immunoblotting, immuno- fluorescence. Results: During differentiation proceeds, from day 1 to day 28, stem cells change in shape from spindle to polygon with larger size and tighter intercellular connections. Intracellular glycogen and urea production was significantly increased from day 1 to day 28. The mRNA expression of liver-specific genes was upregulated. In order to study key regulator in hepatogenic differentiation, we performed protein/DNA array. EGR1 in differentiated BM-MSCs were significantly increased during the culture compared to undifferentiated BM-MSCs. In contrast, downregulation of EGR1 by lenti-shRNA reduced hepatogenic differentiation and increased the expression of EMT related genes in associaton with the decreased expression of MET related gene. Conclusions: In this study, we identified novel factors in the process of hepatogenic differentiation through inducing the MET process. Our study suggests that EGR1 were expected to the “transcription factor therapy” as novel strategy in future clinical treatment.

Histone Deacetylase 3 Promotes Alveolar Epithelial-mesenchymal Transition and Fibroblast Migration in Hypoxic Conditions

( Jeong-woong Park ),( Se-hee Kim ),( Sung Hwan Jeong ),( Eun Suk Son ),( Young Eun Lee ),( Sung Young Kyung ) 대한결핵 및 호흡기학회 2020 대한결핵 및 호흡기학회 추계학술대회 초록집 Vol.128 No.-

Background/Aims Epithelial-mesenchymal transition (EMT), a process by which epithelial cells undergo a phenotypic conversion that gives rise to myofibroblasts, plays a pivotal role in the development of lung diseases such as pulmonary fibrosis. Recent studies revealed that hypoxia induces EMT of alveolar epithelial cells and histone deacetylases (HDACs) are aberrantly overexpressed in basal cells of idiopathic pulmonary fibrosis (IPF) lungs. Here, we aimed to elucidate whether HDAC3 promotes mesenchymal transition of alveolar epithelial cells and fibroblast migration under hypoxic conditions. Methods To evaluate the effect of HDAC3 in hypoxia-induced alveolar EMT, we performed quantitative real-time PCR, Western blot analysis, and immunocytochemistry. In addition, we also conducted migration and invasion assays using HDAC3 siRNA transfection in human IPF lung fibroblast cells, and immunohistochemistry using lung tissues from the bleomycin-induced mice and fibrosis patients. Results We found that HDAC3 decreased E-cadherin, and increased ZEB1, Snail, β-catenin, fibronectin, α-SMA, N-cadherin, and vimentin during hypoxia. Additionally, HDAC3 induced expression of HIF-1α and miR-224 under hypoxic condition, which it contributed to upregulation of Snail. The process by which HDAC3 promoted alveolar EMT was dependent on the AKT pathway. Furthermore, inhibition of HDAC3 using siRNA suppressed migration and invasion of fibroblast cells in hypoxic conditions. In lung tissues from bleomycin-induced mice and IPF patients, expression of HDAC3, Snail, and CAIX was elevated. Conclusions We suggest that HDAC3 induces mesenchymal transition of alveolar epithelial cells through AKT pathway, and migration and invasion of fibroblast cells in hypoxic conditions, which may contribute to the aggravation of lung fibrosis.

Effects of Hypoxia on Epithelial-to-Mesenchymal Transition in Oral Squamous Cell Carcinoma

김경아,김지영,유미현,박봉수,정진,박혜련 대한구강악안면병리학회 2011 대한구강악안면병리학회지 Vol.35 No.1

Malignant tumor cells outgrow new blood vessel formation and tend to be in hypoxic state. Hypoxic cancer cells adapt to hypoxic conditions by transforming its characteristics. On the other hand, one of the most important features of cancer cells is that carcinoma cells loses its inherent epithelial phenotype and acquires mesenchymal characteristics, called as epithelial-mesenchymal transition(EMT). It has been already well known that EMT contributes to tumor invasion and metastasis. The present study investigated whether hypoxia play a major role in induction of phenotypic changes of oral squamous cell carcinoma(OSCC). Furthermore, the mechanism of EMT in oral squamous cell carcinoma cells by hypoxia has been clarified. To mimic hypoxic condition, cobalt chloride and desferoxamine, well-known hypoxic mimetic agents, were used. This study shows that hypoxia suppresses the expression of E-cadherin(epithelial marker) and increases vimentin and N-cadherin(mesenchymal markers) in OSCC. In addition, α5 integrin protein, which is a receptor for fibronectin and an important molecule for tumor invasion, is prominently induced by hypoxia. Malignant tumor cells outgrow new blood vessel formation and tend to be in hypoxic state. Hypoxic cancer cells adapt to hypoxic conditions by transforming its characteristics. On the other hand, one of the most important features of cancer cells is that carcinoma cells loses its inherent epithelial phenotype and acquires mesenchymal characteristics, called as epithelial-mesenchymal transition(EMT). It has been already well known that EMT contributes to tumor invasion and metastasis. The present study investigated whether hypoxia play a major role in induction of phenotypic changes of oral squamous cell carcinoma(OSCC). Furthermore, the mechanism of EMT in oral squamous cell carcinoma cells by hypoxia has been clarified. To mimic hypoxic condition, cobalt chloride and desferoxamine, well-known hypoxic mimetic agents, were used. This study shows that hypoxia suppresses the expression of E-cadherin(epithelial marker) and increases vimentin and N-cadherin(mesenchymal markers) in OSCC. In addition, α5 integrin protein, which is a receptor for fibronectin and an important molecule for tumor invasion, is prominently induced by hypoxia.