Mir-520a-3p Affect The Biological Behavior Of PTC By Regulating JAK/STAT Signaling Pathways

Thyroid cancer(THCA)is the most common malignant tumor of endocrine system,whose incidence ranks first in the head and neck malignant tumors,accounting for about 94.5% of all endocrine tumor and 2.6% of all malignant tumor.Papillary thyroid carcinoma(PTC)is the most common type of thyroid cancer,accounting for about 90% of all differentiated thyroid cancer.PTC is popular among teenagers and women,the majority of which were with cervical lymph node metastasis.Accumulating studies show that micro RNAs(miRNAs)are involved in the occurrence and development of PTC.Micro RNA-520a-3p(miR-520a-3p)is a member of the family of miR-520,which is a relatively conservative family located on human chromosome 19.Some studies found that miR-520 a played an important role in tumor suppression,including breast cancer,esophageal squamous carcinoma,colon cancer and myeloid leukemia.However,its clinical significance in the PTC and the underlying molecular mechanism is still unclear.Therefore,to study the invasion and metastasis process of PTC and the molecular mechanism of cancer cell metastasis are the key problems to be solved in clinical work,which is also the research direction of this study.This study is set to investigate roles of miR-520a-3p in PTC development and related mechanisms.Tumor tissues and normal thyroid tissues were collected from 217 cases of PTC.Histopathological changes were observed by hematoxylin & eosin(HE)staining.The expression of miR-520a-3p in PTC cells and tissues was detected by real-time fluorescence quantitative PCR,and the relationship between the expression of miR-520a-3p and the clinicopathological characteristics of PTC patients was analyzed.The results showed that the relative expression of miR-520a-3p in PTC cells and tissues was downregulated,and it was related to lymph node metastasis,invasion and TNM stage.To further explore the specific mechanism of miR-520a-3p,In this study,the bioinformatics website(micro RNA.org)was used to predict the target molecule of miR-520a-3p,and it was found that the 3'-UTR of the JAK1 gene contains a base sequence that highly matches miR-520a-3p,and through dual luciferase gene reporter experiment confirmed that miR-520a-3p can specifically bind to JAK1 3'-UTR and down-regulate JAK1 gene expression after transcription.Subsequently the positive rate of JAK1 protein was detected by immunohistochemistry.The results showed that JAK1 protein expression in normal tissues was significantly lower than that in PTC tissues,and it was significantly related to tumor differentiation,lymph node metastasis,invasion and TNM stage.At the same time,the results of q RT-PCR and Western blot showed that the expression levels of JAK1,JAK2 and STAT3 were significantly upregulated in PTC,showing that JAK/STAT pathway was activated in PTC tissues.Next,normal and PTC cells were assigned into 7 groups,including Normal,Blank,NC,miR-520a-3p mimic,miR-520a-3p inhibitor,siRNA-JAK1 and miR-520a-3p inhibitor + siRNA-JAK1 groups.Then,q RT-q PCR and Western blot were used to detect expression of JAK/STAT pathway related genes(JAK2 and STAT3)in all transfected cells.And the results showed that miR-520a-3p inhibited the activation of JAK/STAT pathway by negatively regulating JAK1.In order to further clarify the effect of miR-520a-3p on the biological behavior of PTC cells and its related molecular mechanism,MTT method was used to detect cell growth,flow cytometry to detect cell cycle distribution and apoptosis,scratch test to detect cell migration,transwell to detect cell invasion,and q RT-PCR and Western blot to detect the relative expression of E-cadherin and Vimentin.We found it significantly decreased the invasion of PTC cells and partially reverses epithelial mesenchymal transition.It was found that the ability of cell invasion,migration and epithelial-mesenchymal transformation in PTC tissues decreased significantly,and apoptosis increased,suggesting that miR-520a-3p can inhibit the growth,migration,invasion,epithelial-mesenchymal transformation and promote apoptosis of PTC cells by negatively regulating JAK1 expression.The research in this paper shows that miR-520a-3p has the ability to inhibit the invasion and migration of PTC cells,can significantly reduce the malignancy of cell tumors,and by targeting down-regulation of JAK1,the activation of JAK/STAT signaling pathway is inhibited,thereby preventing PTC invasion and metastasis and epithelial mesenchymal transformation.This study reveals the principle of miR-520a-3p in the malignant biological behavior of PTC,and provides a theoretical basis for the early diagnosis,intervention treatment and prognosis of PTC.Moreover,it is of great significance to further improve the cure rate and survival rate of patients,and to find new targets for the treatment of PTC.