BACKGROUND

This study aims to characterize the function of downregulated MicroRNA miR-205 in renal cell carcinoma (RCC), and show how the downstream zinc finger E-box-binding homeobox 2 (ZEB2) is negatively regulated by miR-205.

METHODS

The expression of miR-205 was detected in RCC and adjacent non-tumor tissues using real-time polymerase chain reaction (PCR). The expression of miR-205 and ZEB2 was detected in RCC cell lines using real-time PCR. The luciferase reporter assay was used to assess ZEB2 as a target of miR-205. Protein levels of ZEB2, E-cadherin, and vimentin were measured by western blot after overexpression of miR-205 in ACHN cells. In vivo functions of miR-205 in ACHN cells were measured by MTT assays, migration and invasion assays, and flow cytometry.

RESULTS

MiR-205 was significantly downregulated in RCC samples and cell lines compared with matched non-tumor tissues and HK-2 cells, respectively. No significant difference was found in miR-205 expression between well differentiated and poorly to moderately differentiated groups or between phase I and phase II-III. ZEB2 was upregulated in RCC cell lines compared with expression in HK-2 cells. Upregulation of miR-205 expression caused the downregulation of ZEB2 and vimentin, and the upregulation of E-cadherin in ACHN cells. miR-205 also inhibited proliferation, migration, and invasion, and induced apoptosis of ACHN cells.

CONCLUSIONS

miR-205 is a candidate tumor suppressor that targets ZEB2 in RCC.

Bladder cancer (BC) is a common urologic tumor characterized by high risk of recurrence and mortality. Capsaicin (CPS), used as an intravesical drug for overactive bladder, was demonstrated to induce cell death in different cancer cells including BC cells.Here we found that treatment of high-grade BC cells with high dose of CPS triggers autophagy. Infact, the CPS treatment alters the redox homeostasis by inducing production of radicals, mitochondrial depolarization, alterations of ADP/ATP ratio and activation of AMPK pathway stimulating the autophagic process in BC cells. The inhibition of autophagy, by using the specific inhibitor bafilomycin A or Beclin 1 knock-down, enhanced the CPS-induced cell death, demonstrating that CPS-induced autophagy acts as a pro-survival process in BC cells. By using PCR arrays and FACS analysis, we found that the CPS-treated BC cells displayed typical mesenchymal features of the epithelial mesenchymal transition (EMT) as elongated shape and over-expression of vimentin, α5 and β1 integrin subunits, integrin-like kinase and the anti-apoptotic Bcl-2 proteins. Moreover, we demonstrated that CPS treatment stimulates upregulation of Dhh/Ptch2/Zeb2 members of the Hedgehog signaling pathway, increases CD24, VEGFA and TIMP1 and decreases CD44 and ALCAM mRNA expression levels. By PTCH2 knock-down we found that the Hedgehog signaling pathway is involved in the CPS-induced autophagy and EMT phenotype.Finally, we also showed that the CPS-resistant EMT-positive BC cells displayed an increased drug-resistance to the cytotoxic effects of mitomycin C, gemcitabine and doxorubicine drugs commonly used in BC therapy.

The purpose of this study was to test the hypothesis that KLF4 promotes corneal epithelial (CE) cell fate by suppressing the epithelial-mesenchymal transition (EMT), using spatiotemporally regulated CE-specific ablation of Klf4 in Klf4Δ/ΔCE (Klf4LoxP/LoxP/Krt12rtTA/rtTA/Tet-O-Cre) mice.

CE-specific ablation of Klf4 was achieved by feeding Klf4Δ/ΔCE mice with doxycycline chow. The wild-type (WT; normal chow-fed littermates) and the Klf4Δ/ΔCE histology was compared by hematoxylin and eosin-stained sections; EMT marker expression was quantified by quantitative PCR, immunoblots, and immunofluorescent staining; and wound healing rate was measured by CE debridement using Algerbrush. KLF4 and EMT markers were quantified in human corneal limbal epithelial (HCLE) cells undergoing TGF-β1-induced EMT by quantitative PCR, immunoblots, and immunofluorescent staining.

The epithelial markers E-cadherin, Krt12, claudin-3, and claudin-4 were down-regulated, whereas the mesenchymal markers vimentin, β-catenin, survivin, and cyclin-D1 and the EMT transcription factors Snail, Slug, Twist1, Twist2, Zeb1, and Zeb2 were up-regulated in the Klf4Δ/ΔCE corneas. The Klf4Δ/ΔCE cells migrated faster, filling 93% of the debrided area within 16 hours compared with 61% in the WT. After 7 days of wounding, the Klf4Δ/ΔCE cells that filled the gap failed to regain epithelial characteristics, as they displayed abnormal stratification; down-regulation of E-cadherin and Krt12; up-regulation of β-catenin, survivin, and cyclin-D1; and a 2.5-fold increase in the number of proliferative Ki67+ cells. WT CE cells at the migrating edge and the HCLE cells undergoing TGF-β1-induced EMT displayed significant down-regulation of KLF4.

Collectively, these results reveal that KLF4 plays an essential role in CE homeostasis by promoting epithelial cell fate and suppressing EMT.

Osteosarcoma is the most common primary bone sarcoma around the world. The poor prognosis and high recurrence rate of osteosarcoma are largely due to the high rate of pulmonary metastasis. H19 has been reported to play a potential role in osteosarcoma progression. However, the exact molecular mechanism of metastasis involving H19 remains unclear. In the present study, we performed gain- and loss-of-function assays and found that H19 promotes migration and invasion in osteosarcoma cells. Furthermore, we showed that H19 promotes metastasis through upregulation of ZEB1 and ZEB2 by competitively binding the miR-200 family. Thus, our findings suggest important roles of H19 in osteosarcoma metastasis and indicate its potential application in osteosarcoma therapy.

Cancer cells acquire essential characteristics for metastatic dissemination through the process of epithelial-to-mesenchymal transition (EMT), which is regulated by gene expression and chromatin remodeling changes. The enhancer of zeste homolog 2 (EZH2), the catalytic subunit of the polycomb repressive complex 2 (PRC2), catalyzes trimethylation of lysine 27 of histone H3 (H3K27me3) to repress gene transcription. Here we report the functional roles of EZH2-catalyzed H3K27me3 during EMT in ovarian cancer (OC) cells. TGF-β-induced EMT in SKOV3 OC cells was associated with decreased levels of EZH2 and H3K27me3 (P<0.05). These effects were delayed (~72 h relative to EMT initiation) and coincided with increased (>15-fold) expression of EMT-associated transcription factors ZEB2 and SNAI2. EZH2 knockdown (using siRNA) or enzymatic inhibition (by GSK126) induced EMT-like changes in OC cells. The EMT regulator ZEB2 was upregulated in cells treated with either approach. Furthermore, TGF-β enhanced expression of ZEB2 in EZH2 siRNA- or GSK126-treated cells (P<0.01), suggesting that H3K27me3 plays a role in TGF-β-stimulated ZEB2 induction. Chromatin immunoprecipitation assays confirmed that TGF-β treatment decreased binding of EZH2 and H3K27me3 to the ZEB2 promoter (P<0.05). In all, these results demonstrate that EZH2, by repressing ZEB2, is required for the maintenance of an epithelial phenotype in OC cells.

Prior study shows that maternal protein-restricted (LP) 16-wk-old offspring have pronounced reduction of nephron number and arterial hypertension associated with unchanged glomerular filtration rate, besides enhanced glomerular area, which may be related to glomerular hyperfiltration/overflow and which accounts for the glomerular filtration barrier breakdown and early glomerulosclerosis. In the current study, LP rats showed heavy proteinuria associated with podocyte simplification and foot process effacement. TGF-β1 glomerular expression was significantly enhanced in LP. Isolated LP glomeruli show a reduced level of miR-200a, miR-141, miR-429 and ZEB2 mRNA and upregulated collagen 1α1/2 mRNA expression. By western blot analyzes of whole kidney tissue, we found significant reduction of both podocin and nephrin and enhanced expression of mesenchymal protein markers such as desmin, collagen type I and fibronectin. From our present knowledge, these are the first data showing renal miRNA modulation in the protein restriction model of fetal programming. The fetal-programmed adult offspring showed pronounced structural glomerular disorders with an accentuated and advanced stage of fibrosis, which led us to state that the glomerular miR-200 family would be downregulated by TGF-β1 action inducing ZEB 2 expression that may subsequently cause glomeruli epithelial-to-mesenchymal transition.

NANOG is a key pluripotency factor in embryonic stem cells that is frequently expressed in squamous cell carcinomas (SCCs). However, a direct link between NANOG and SCCs remains to be established. Here, we show that inducible overexpression of NANOG in mouse skin epithelia favours the malignant conversion of skin papillomas induced by chemical carcinogenesis, leading to increased SCC formation. Gene expression analyses in pre-malignant skin indicate that NANOG induces genes associated to epithelial-mesenchymal transition (EMT). Some of these genes are directly activated by NANOG, including EMT-associated genes Zeb1, Zeb2, Twist1, Prrx1 and miR-21. Finally, endogenous NANOG binds to the promoters of theses genes in human SCC cells and, moreover, NANOG induces EMT features in primary keratinocytes. These results provide in vivo evidence for the oncogenic role of NANOG in squamous cell carcinomas.

MicroRNAs from the miR-200 family are commonly associated with the inhibition of the metastatic potential of cancer cells, following inhibition of ZEB transcription factors expression and epithelial-to-mesenchymal transition. However, previous studies performed in pancreatic adenocarcinoma revealed a more complex picture challenging this canonical model. To gain better insights into the role of miR-200 family members in this disease, we analyzed the expression of miR-200a, miR-200b, miR-200c, miR-141, miR-429, and miR-205, and ZEB1, ZEB2, and CDH1 in pancreatic tumors and matching normal adjacent parenchyma and patient-derived xenografts. We found that miR-200a, miR-429, and miR-205 are frequently overexpressed in pancreatic tumors, whereas CDH1 is downregulated, and ZEB1 and ZEB2 levels remain unchanged. Furthermore, we measured a positive correlation between miR-200 family members and CDH1 expression, and a negative correlation between ZEB1 and miR-200c, miR-141, and miR-205 expression, respectively. Interestingly, we identified significant changes in expression of epithelial-to-mesenchymal transition regulators and miR-200 members in patient-derived xenografts. Lastly, functional studies revealed that miR-141 and miR-429 inhibit the tumorigenic potential of pancreatic cancer cells. Taken together, this comprehensive analysis strongly suggests that miRNAs from the miR-200 family, and in particular miR-429, may act as a tumor suppressor gene in pancreatic cancer.

Clinically, serum level of folate has been negatively correlated to the stage and progression of liver cancer. Nevertheless, the functional consequence of folate deficiency (FD) in malignancy has not been fully investigated. Human hepatocellular carcinoma (HCC) cells (as study model) and other cancer types such as lung and glioma were cultured under folate deficient (FD) and folate complete (FD) conditions. Molecular characterization including intracellular ROS/RNS (reactive oxygen/nitrogen species), viability, colony formation, cancer stem-like cell (CSC) phenotype analyses were performed. In vivo tumorigenesis under FD and FC conditions were also examined. FD induced a significant increase in ROS and RNS, suppressing proliferative ability but inducing metastatic potential. Mesenchymal markers such as Snail, ZEB2, and Vimentin were significantly up-regulated while E-cadherin down-regulated. Importantly, CSC markers such as Oct4, β-catenin, CD133 were induced while PRRX1 decreased under FD condition. Furthermore, FD-conditioned HCC cells showed a decreased miR-22 level, leading to the increased expression of its target genes including HDAC4, ZEB2 and Oct4. Finally, xenograft mouse model demonstrated that FD diet promoted tumorigenesis and metastasis as compared to their FC counterparts. Our data provides rationales for the consideration of folate supplement as a metastasis preventive measure.

Mowat-Wilson syndrome (MWS) is a rare intellectual disability/multiple congenital anomalies syndrome caused by heterozygous mutation of the ZEB2 gene. It is generally underestimated because its rarity and phenotypic variability sometimes make it difficult to recognize. Here, we aimed to better delineate the phenotype, natural history, and genotype-phenotype correlations of MWS.

In a collaborative study, we analyzed clinical data for 87 patients with molecularly confirmed diagnosis. We described the prevalence of all clinical aspects, including attainment of neurodevelopmental milestones, and compared the data with the various types of underlying ZEB2 pathogenic variations.

All anthropometric, somatic, and behavioral features reported here outline a variable but highly consistent phenotype. By presenting the most comprehensive evaluation of MWS to date, we define its clinical evolution occurring with age and derive suggestions for patient management. Furthermore, we observe that its severity correlates with the kind of ZEB2 variation involved, ranging from ZEB2 locus deletions, associated with severe phenotypes, to rare nonmissense intragenic mutations predicted to preserve some ZEB2 protein functionality, accompanying milder clinical presentations.

Knowledge of the phenotypic spectrum of MWS and its correlation with the genotype will improve its detection rate and the prediction of its features, thus improving patient care.

Epithelial-to-mesenchymal plasticity (EMP) has been linked to metastasis, stemness, and drug resistance. In prostate cancer, EMP has been associated with both suppression and activation of the androgen receptor (AR) signaling. Here we investigated the effect of the potent AR antagonist enzalutamide on EMP in multiple preclinical models of prostate cancer and patient tissues. Enzalutamide treatment significantly enhanced the expression of EMP drivers (ZEB1, ZEB2, Snail, Twist, and FOXC2) and mesenchymal markers (N-cadherin, fibronectin, and vimentin) in prostate cancer cells, enhanced prostate cancer cell migration, and induced prostate cancer transformation to a spindle, fibroblast-like morphology. Enzalutamide-induced EMP required concomitant suppression of AR signaling and activation of the EMP-promoting transcription factor Snail, as evidenced by both knockdown and overexpression studies. Supporting these findings, AR signaling and Snail expression were inversely correlated in C4-2 xenografts, patient-derived castration-resistant metastases, and clinical samples. For the first time, we elucidate a mechanism explaining the inverse relationship between AR and Snail. Specifically, we found that AR directly repressed SNAI1 gene expression by binding to specific AR-responsive elements within the SNAI1 promoter. Collectively, our findings demonstrate that de-repression of Snail and induction of EMP is an adaptive response to enzalutamide with implications for therapy resistance. Cancer Res; 77(11); 3101-12. ©2017 AACR.

The epithelial-mesenchymal transition (EMT), a key process in the tumor metastatic cascade, is characterized by the loss of cell-cell junctions and cell polarity, as well as the acquisition of migratory and invasive properties. LEF-1 is a member of the lymphoid enhancer-binding factor/T-cell factor (LEF/TCF) family of DNA-binding transcription factors, which interact with nuclear β-catenin and act as central transcriptional mediators of Wnt signaling. To investigate the role of LEF-1 in EMT, we generated stable LEF-1 transfectants using MDCK cells. The transfectants had a spindle-shaped mesenchymal morphology, and enhanced migration and invasiveness relative to control cells. These EMT changes were accompanied by the downregulation of an epithelial marker protein, E-cadherin, and the upregulation of mesenchymal marker proteins, vimentin and N-cadherin. Consistent with these observations, the mRNA levels of Slug, ZEB1, and ZEB2-EMT-related transcription factors-increased significantly. Although the N-terminally deleted mutant LEF-1 cannot interact with β-catenin, it retained the ability to induce EMT. Consistent with these observations, neither the expression of a dominant negative β-catenin/engrailed chimera, nor the expression of a cytoplasmic domain of E-cadherin that sequesters β-catenin from binding to LEF/TCF, reversed LEF-1-induced EMT. Together, these data indicated that the nuclear function of β-catenin was not necessary for the induction of Slug, ZEB1, and ZEB2 expression leading to EMT.

Bladder cancer (BCa) threatens human health due to the high occurrence and mortality. Nowadays, more and more researchers focused on the molecular mechanisms and biological functions of microRNAs (miRNAs) in human cancers. This study aims to study the biological role of miR-454-3p and miR-374b-5p in bladder cancer. The expression levels of miR-454-3p and miR-374b-5p were detected in BCa tissues and cell lines by qRT-PCR analysis. Kaplan-Meier analysis revealed that the expression levels of miR-454-3p and miR-374b-5p were positively correlated with the overall survival rate of BCa patients. Gain-of-function assays were conducted to demonstrate the inhibitory effects of miR-454-3p and miR-374b-5p on the invasion, migration and epithelial-mesenchymal transition (EMT) of BCa cells. Mechanically, ZEB2 was found to be a target of both miR-454-3p and miR-374b-5p. Rescue assays revealed that ZEB2 reversed the inhibitory effects of miR-454-3p and miR-374b-5p on the invasion and migration of BCa cell lines. In summary, miR-454-3p and miR-374b-5p negatively regulated invasion and migration of BCa cell lines via targeting ZEB2.

Hepatocellular carcinoma (HCC) is the most common type of cancer and the rapid tumor growth, drug resistance and metastasis are the major problems for HCC therapy. MicroRNAs (miRNAs) involve in various cell biological processes in HCC. ZEB2 plays crucial roles in HCC progression. ZEB2 is regulated by some identified miRNAs, but there needs to find new miRNAs regulating ZEB2 expression for better understanding the molecular mechanism of HCC. In the present study, ZEB2 was identified as a direct target of miR-211-5p, which was a potential oncogene in cancer. We found that miR-211-5p levels in HCC tissues were lower than the compared normal tissues. ZEB2 expression was higher in HCC tissues and was negatively related to miR-211-5p levels. Overexpression of miR-211-5p in human HCC cell lines (HepG2 and 7721) caused the delay of cell proliferation, apoptosis and drug sensitivity. Summarily, our study demonstrates that miR-211-5p may play a suppressing role in HCC by inhibiting ZEB2 expression.

Metastasis is the most common cause of mortality in patients with gastric cancer. Epithelial-to-mesenchymal transition (EMT), which may be stimulated by insulin-like growth factor-I (IGF-I) is involved in the metastasis of numerous tumors; however, the molecular mechanism by which IGF-I may induce tumor cell EMT remains to be elucidated in gastric cancer. The present study aimed to investigate the induction of EMT in BGC-823 gastric cancer cells. It was identified that IGF-I induced EMT by upregulating the levels of ZEB2 transcription factor, and this was dependent on the phosphoinositide 3-kinase (PI3K)/Akt signaling pathway in these cells. In addition, glycogen synthase kinase 3β (GSK-3β), an intracellular downstream effector of PI3K/Akt, sustained the epithelial phenotype by repressing ZEB2 expression and the subsequent inhibition of EMT induced by IGF-I, suggesting the involvement of a potential PI3K/Akt-GSK-3β-ZEB2 signaling pathway in IGF-I-induced EMT in gastric cancer BGC-823 cells. Overall, the results of the present study suggest that IGF-I induced EMT by the activation of a PI3K/Akt-GSK-3β-ZEB2 signaling pathway in gastric cancer BGC-823 cells. Therefore, this study may provide more useful information regarding the mechanism of gastric cancer metastasis.

Aging imposes a barrier to somatic cell reprogramming through poorly understood mechanisms. Here, we report that fibroblasts from old mice express higher levels of Zeb2, a transcription factor that activates epithelial-to-mesenchymal transition. Synthesis of Zeb2 protein is controlled by a natural antisense transcript named Zeb2-NAT. We show that transfection of adult fibroblasts with specific LNA Gapmers induces a robust downregulation of Zeb2-NAT transcripts and Zeb2 protein and enhances the reprogramming of old fibroblasts into pluripotent cells. We further demonstrate that Zeb2-NAT expression is precociously activated by differentiation stimuli in embryonic stem (ES) cells. By knocking down Zeb2-NAT, we were able to maintain ES cells challenged with commitment signals in the ground state of pluripotency. In conclusion, our study identifies a long noncoding RNA that is overlapping and antisense to the Zeb2 locus as a target for rejuvenation strategies.

Cervical carcinoma (CC) is the second most common cancer in females. In order to improve current anti-metastasis strategies for CC, it is important to improve our understanding of the mechanisms involved in epithelial-to-mesenchymal transition (EMT). This study aimed to elucidate the potential role of a novel long non-coding RNA (lncRNA)-CTS and the mechanisms underlying EMT in CC. The expression levels of lncRNA-CTS and miR-505 were detected using quantitative reverse transcriptase polymerase chain reaction in CC specimens and cells (HeLa, SiHa, Ca-Ski, C-33A, and HT-3). Further experiments including wound scratch and transwell invasion assays, Western blotting, immunofluorescence, and luciferase assays were used to investigate the function of lncRNA-CTS/miR-505/ZEB2 in vitro. In addition, a tumor xenograft model was used to assess the effect of lncRNA-CTS in vivo. The expression levels of lncRNA-CTS and miR-505 were correlated with the metastasis-associated clinicopathological features of CC patients. Moreover, lncRNA-CTS was associated with a poor prognosis in CC patients. In vitro and in vivo experiments, along with gain- and loss-of-function studies, showed that lncRNA-CTS enhanced cell migration, invasion, and the transforming growth factor (TGF)-β1-induced-EMT process. Data also showed that lncRNA-CTS could function as a competing endogenous RNA for miR-505 in CC cells. Further investigations disclosed that ZEB2 was demonstrated as a downstream target of miR-505, and subsequently exerted its metastatic effects via the lncRNA-CTS/miR-505/ZEB2 axis in CC cells. Finally, lncRNA-CTS activated the SMAD/TGF pathway via miR-505 in CC cells. Collectively, our results demonstrate the importance of the lncRNA-CTS/miR-505/ZEB2 axis in CC. LncRNA-CTS can predispose CC patients to metastases and may represent a promising therapeutic target for CC.

Ell3 is a testis-specific RNA polymerase II elongation factor whose cellular function is not clear. The present study shows that Ell3 is activated during the differentiation of mouse embryonic stem cells (mESCs). Furthermore, Ell3 plays a critical role in stimulating lineage differentiation of mESCs by promoting epithelial-mesenchymal transition (EMT) and suppressing apoptosis. Mouse ESCs engineered to stably express Ell3 were rapidly differentiated compared with control cells either under spontaneous differentiation or neural lineage-specific differentiation conditions. Gene expression profile and quantitative RT-PCR analysis showed that the expression of EMT markers, such as Zeb1 and Zeb2, two major genes that regulate EMT, was upregulated in Ell3-overexpressing mESCs. Remarkably, knockdown of Zeb1 attenuated the enhanced differentiation capacity of Ell3-overexpressing mESCs, which indicates that Ell3 plays a role in the induction of mESC differentiation by inducing EMT. In contrast to Ell3-overexpressing mESCs, Ell3-knock down mESCs could not differentiate under differentiation conditions and, instead, underwent caspase-dependent apoptosis. In addition, apoptosis of differentiating Ell3-knock out mESCs was associated with enhanced expression of p53. The present results suggest that Ell3 promotes the differentiation of mESCs by activating the expression of EMT-related genes and by suppressing p53 expression.

Plexiform and/or dermal neurofibromas are nerve sheath tumors of the peripheral nervous system that are usually present in individuals with neurofibromatosis type 1 (NF1). Neurofibromas arise from Schwann cells with biallelic inactivation of NF1, the gene that encodes neurofibromin. This protein is responsible for regulation of the Ras-mediated pathway, which has been shown to play a crucial role in epithelial-to-mesenchymal transition (EMT). EMT is a biological process that occurs during embryogenesis and wound healing and is involved in pathological processes such as organ fibrosis and cancer metastasis. However, the relationship between neurofibromin and EMT has not been elucidated. We investigated whether the EMT-related signaling pathway was upregulated in NF1-associated neurofibromas and Schwann cells by assessing the expression levels of the EMT-related transcription factors Snail, Slug, Twist, ZEB1 and ZEB2. Immunohistochemical studies and quantitative reverse transcription polymerase chain reaction revealed an increase in the expression levels of EMT-related transcription factors in neurofibroma specimens and NF1-derived Schwann cells (sNF96.2). In addition, the silencing of NF1 by siRNA induced the expression of EMT-related transcription factors in normal human Schwann cells and in epithelial-like breast cancer cells. Our findings suggest that the loss of neurofibromin activated the EMT-related signaling pathway and that the excessive mesenchymal reaction may play a key role in the development of NF1-associated neurofibromas.

Patients with thyroid cancers refractory to radioiodine (RAI) treatment show a limited response to various therapeutic options and a low survival rate. The recent use of multikinase inhibitors has also met limited success. An alternative approach relies on drugs that induce cell differentiation, as the ensuing increased expression of the cotransporter for sodium and iodine (NIS) may partially restore sensitivity to radioiodine. The inhibition of the ERK1/2 pathway has shown some efficacy in this context. Aggressive thyroid tumors overexpress the isoform-A of the insulin receptor (IR-A) and its ligand IGF-2; this IGF-2/IR-A loop is associated with de-differentiation and stem-like phenotype, resembling RAI-refractory tumors. Importantly, IR-A has been shown to be positively modulated by the non-integrin collagen receptor DDR1 in human breast cancer. Using undifferentiated human thyroid cancer cells, we now evaluated the effects of DDR1 on IGF-2/IR-A loop and on markers of cell differentiation and stemness. DDR1 silencing or downregulation caused significant reduction of IR-A and IGF-2 expression, and concomitant increased levels of differentiation markers (NIS, Tg, TSH, TPO). Conversely, markers of epithelial-to-mesenchymal transition (Vimentin, Snail-2, Zeb1, Zeb2 and N-Cadherin) and stemness (OCT-4, SOX-2, ABCG2 and Nanog) decreased. These effects were collagen independent. In contrast, overexpression of either DDR1 or its kinase-inactive variant K618A DDR1-induced changes suggestive of less differentiated and stem-like phenotype. Collagen stimulation was uneffective. In conclusion, in poorly differentiated thyroid cancer, DDR1 silencing or downregulation blocks the IGF-2/IR-A autocrine loop and induces cellular differentiation. These results may open novel therapeutic approaches for thyroid cancer.

The majority of studies on microRNA-200 family members (miR-200s) in human cancers are based on the premise that miR-200s maintain epithelial cell integrity by suppressing epithelial-mesenchymal transition (EMT) through direct inhibition of mesenchymal transcription factors zinc finger E-box-binding homeobox 1/2 (ZEB1/ZEB2) and transforming growth factor-β (TGF-β), a potent inducer of EMT. Hence, downregulation of miR-200 in cancer cells promotes EMT and cancer metastasis. Yet, miR-200s are highly expressed in ovarian cancer, and ovarian cancer metastasizes primarily by dissemination within the pelvic cavity. In this review, we will refocus the epithelial property of ovarian cancer cells and the role of miR-200s in safeguarding this property, as well as the diverse roles of miR-200s in inclusion cyst formation, cancer cell growth, collective movement, angiogenesis, exosome-mediated cell communication, and chemoresponse. Taken together, miR-200s play a significant role in the initiation, progression and metastasis of ovarian cancer and may serve as diagnostic biomarkers and a target in therapeutic development.

Epithelial homeostasis within the epidermis is maintained by means of multiple cell-cell adhesion complexes such as adherens junctions, tight junctions, gap junctions, and desmosomes. These complexes co-operate in the formation and the regulation of the epidermal barrier. Disruption of the epidermal barrier through the deregulation of the above complexes is the cause behind a number of skin disorders such as psoriasis, dermatitis, keratosis, and others. During epithelial-to-mesenchymal transition (EMT), epithelial cells lose their adhesive capacities and gain mesenchymal properties. ZEB transcription factors are key inducers of EMT. In order to gain a better understanding of the functional role of ZEB2 in epidermal homeostasis, we generated a mouse model with conditional overexpression of Zeb2 in the epidermis. Our analysis revealed that Zeb2 expression in the epidermis leads to hyperproliferation due to the combined downregulation of different tight junction proteins compromising the epidermal barrier. Using two epidermis-specific in vivo models and in vitro promoter assays, we identified occludin as a new Zeb2 target gene. Immunohistological analysis performed on human skin biopsies covering various pathogeneses revealed ZEB2 expression in the epidermis of pemphigus vulgaris. Collectively, our data support the notion for a potential role of ZEB2 in intracellular signaling of this disease.

Aberrant expression of microRNA-302a (miR-302a) has been frequently reported in some cancers excluding colorectal cancer (CRC). However, the role of miR-302a in CRC has not been reported. In this paper, we examined the effect of miR-302a overexpression on proliferation and invasion in CRC cells. The mRNA level of miR-302a in CRC cell lines was determined by real-time PCR. The miR-302a mimic was transiently transfected into CRC cells using Lipofectamine™ 2000 reagent. Subsequently, cell proliferation and invasion were assessed by MTT and Transwell assays. Western blot and ELISA assay were used to detect the expressions and secretions of matrix metalloproteinases (MMPs). Moreover, the expressions of epithelial marker, mesenchymal markers and transcription factors were also determined by Western blot. In addition, the effects of miR-302a overexpression on the MAPK and PI3K/Akt signaling pathways were investigated by Western blot. Our results showed that the mRNA level of miR-302a was remarkably decreased in CRC cell lines compared with normal colon epithelium cells. Up-regulation of miR-302a inhibited the proliferation and invasion of CRC cells. The expressions and secretions of MMP-9 and -2 were evidently reduced by increasing miR-302a. Besides, we found a decrease of β-catenin, fibronection, vimentin, Snail, Slug, ZEB1 and ZEB2 expressions and an increase of E-cadherin expression. We also found that miR-302a overexpression might decrease the phosphorylation of Erk1/2 and Akt. Altogether, our results indicated that miR-302a overexpression was shown to inhibit proliferation and invasion of CRC cells by reducing the expressions of related proteins through suppressing the MAPK and PI3K/Akt signaling pathways.