Expert consensus on the molecular diagnosis of early-stage pancreatic cancer （2023 edition） / 临床肝胆病杂志

Pancreatic cancer is a relatively common tumor of the digestive system， with difficulties in early-stage diagnosis and an extremely high degree of malignancy. Molecular diagnostic technology based on tumor biomarkers， combined with the existing gold standard in clinical practice， is of great clinical significance to achieve early accurate identification， timely treatment and intervention， and reduction in mortality. Previous studies have shown that miRNAs show high specificity in terms of types and expression levels in different pathological stages of pancreatic cancer and can thus be used in monitoring the development and progression of pancreatic cancer. Since a single miRNA has a limited diagnostic potential， the combination of different miRNAs may effectively improve the diagnostic efficiency of early-stage pancreas carcinogenesis. Based on related research advances in recent years， this consensus document aims to fill the gap in molecular diagnostic technology in the guidelines for the clinical diagnosis and treatment of pancreatic cancer and provide expert guidance and recommendations.

Research progress on non-coding small RNA in oral submucosal fibrosis / 口腔疾病防治

@#Oral submucous fibrosis (OSF) is one of the most common precancerous lesions of the oral mucosa, and its pathogenesis has not been fully elucidated. Small noncoding RNAs (SncRNAs), a class of RNA molecules that do not code for proteins, have been widely reported to be involved in the regulation of a variety of human diseases. An increasing number of studies have shown that a variety of SncRNAs play important roles in the pathogenesis of OSF. Current studies have shown that microRNAs (miRNAs) are involved in OSF disease progression by regulating the expression of related transcription factors and genes or epithelial mesenchymal transformation to regulate the activation of fibroblasts (FBs). Long noncoding RNAs (lncRNAs) that transform growth factor-β/suppressor of mothers against decapentaplegic (TGF-β/Smad) signaling pathways or interact with miRNAs are involved in the development of OSF. Circular RNAs (circRNAs) play a role in OSF by interacting with miRNAs. tRNA-derived small RNAs (tsRNAs) are involved in the progression of various fibrotic diseases, but their specific mechanism of action in OSF still needs to be further explored. In the future, it is still necessary to focus on the targets of SncRNAs mediating OSF progression and explore their function and molecular mechanism in OSF to provide new ideas for the diagnosis and treatment of OSF.

Progressive Changes of Colitis Into Colorectal Cancer Associated With Changes in Neutrophil Lymphocyte Ratio, Serum, Micrornas and Infiltrate Neutrophils

@#Introduction: Understand the progression of colorectal cancer from the beginning until the advance stages is difficult and challenging. However, this could be overcome with a good animal model. Methods: In this study, a modified approach had been used to develop colorectal cancer model. The model was developed and monitored from colitis formation until the late stage of colorectal cancer. The changes of neutrophil lymphocyte ratio (NLR), serum microRNAs and infiltrate neutrophil in different stages of colorectal cancer were assessed in this study. Results: Results showed that the progression of the disease is correlated with NLR as early as the formation of colitis (r=0.121, p<0.026). Meanwhile, the size of the tumor at each stage is also associated with the NLR value (r=0.185, p<0.0012). In the serum microRNAs study, it was found microRNAs expression in blood serum change in different stages of colorectal cancer. In the early stage of colitis formation, miR223 (> 3 fold expression, p < 0.0025) were abundantly found in the blood serum. Meanwhile in others stage mild (miRNA345 > 2.5 fold, p<0.0011), moderate (miRNA347 & miR512 > 3 fold, p<0.002) and severe (miR31 & miR145 > 2 fold, p<0.0001) microRNAs were also found expressed differently. The quantities of infiltrate neutrophil were varied in different stages of the disease. Conclusion: This study provides an insight into the immunity and molecular level of colorectal cancer and it allows a progressive monitoring on the changes in the molecular, cellular and histological level.

A molecular approach to triple-negative breast cancer: targeting the Notch signaling pathway

ABSTRACT Introduction Triple-negative breast cancer is an aggressive subtype of breast cancer characterized by the absence of estrogen receptor, progesterone receptor, and human epidermal growth factor receptor 2 expression. This phenotype renders triple-negative breast cancer cells refractory to conventional therapies, resulting in poor clinical outcomes and an urgent need for novel therapeutic approaches. Recent studies have implicated dysregulation of the Notch receptor signaling pathway in the development and progression of triple-negative breast cancer. Objective This study aimed to conduct a comprehensive literature review to identify potential therapeutic targets of the Notch pathway. Our analysis focused on the upstream and downstream components of this pathway to identify potential therapeutic targets. Results Modulating the Notch signaling pathway may represent a promising therapeutic strategy to treat triple-negative breast cancer. Several potential therapeutic targets within this pathway are in the early stages of development, including upstream (such as Notch ligands) and downstream (including specific molecules involved in triple-negative breast cancer growth). These targets represent potential avenues for therapeutic intervention in triple-negative breast cancer. Comments Additional research specifically addressing issues related to toxicity and improving drug delivery methods is critical for the successful translation of these potential therapeutic targets into effective treatments for patients with triple-negative breast cancer.

Association between microRNA expression and risk of male idiopathic infertility in Iraq

SUMMARY OBJECTIVE: The World Health Organization defines infertility as the inability to get pregnant after 12 months of unprotected sexual activity. This study was conducted to estimate the levels of gene expression for two mature miRNAs (i.e., miR-122 and miR-34c-5p) to evaluate susceptibility to male infertility. METHODS: This study included 50 male patients with idiopathic infertility who were admitted to hospital from the period November 2021 to May 2022 and another group consisting of 50 apparently healthy individuals used as controls. RESULTS: miR-122 level was significantly highest in azoospermia and followed by oligospermia, 39.22 (31.88) versus 37.34 (20.45), respectively. In addition, there was a very significant difference in miR-34c-5p levels between the study groups (p<0.05). CONCLUSION: Two miRNAs, namely, miR-34c-5p and miR-122, can be used as predictive and diagnostic biomarkers for infertility.

3D-Structural Characterization of MicroRNA Expressed in Leprosy / Caracterização Estrutural em 3D de MicroRNAs Expressos na Hanseníase

Introduction: Hansen's disease, or leprosy is caused by Mycobacterium leprae (M. leprae), is a major public health problem in developing countries, and affecting the skin and peripheral nerves. However, M. leprae can also affect bone tissue, mucous membranes, liver, eyes, and testicles, producing a variety of clinical phenotypes. MicroRNAs (miRNAs) have been expressed in the various clinical forms of leprosy and could potentially be used for its diagnosis. Objective: in silico design of the molecular structure of miRNAs expressed in leprosy. Methodology: we performed a nucleotide sequence search of 17 miRNAs expressed in leprosy, designing in silico the molecular structure of the following miRNAs: miRNA-26a, miRNA-27a, miRNA-27b, miRNA-29c, miRNA-34c, miRNA-92a-1, miRNA- 99a-2, miRNA-101-1, miRNA-101-2, miRNA-125b-1, miRNA-196b, miRNA-425-5p, miRNA-452, miRNA-455, miRNA-502, miRNA-539, and miRNA-660. We extracted the nucleotides were from the GenBank of National Center for Biotechnology Information genetic sequence database. We aligned the extracted sequences with the RNA Folding Form, and the three-dimensional molecular structure design was performed with the RNAComposer. Results: we demonstrate the nucleotide sequences, and molecular structure projection of miRNAs expressed in leprosy, and produces a tutorial on the molecular model of the 17 miRNAs expressed in leprosy through in silico projection processing of their molecular structures. Conclusion: we demonstrate in silico design of selected molecular structures of 17 miRNAs expressed in leprosy through computational biology.

Introdução: a doença de Hansen, ou hanseníase é causada pelo Mycobacterium leprae (M. leprae), é um grande problema de saúde pública nos países em desenvolvimento e afeta, a pele e os nervos periféricos. Entretanto, o M. leprae também pode comprometer o tecido ósseo, membranas mucosas, fígado, olhos e testículos, produzindo uma variedade de fenótipos clínicos. MicroRNAs (miRNAs) têm sido expressos nas várias formas clínicas da hanseníase e podem ser potencialmente utilizados para seu diagnóstico. Objetivo: objetivou-se com esse experimento modelar computacionalmente a estrutura molecular dos miRNAs expressos na hanseníase. Metodologia: realizou-se como metodologia uma pesquisa das sequências nucleotídicas de 17 miRNAs expressos na hanseníase, desenhando em modelo computacional a estrutura molecular dos seguintes miRNAs: miRNA-26a, miRNA-27a, miRNA-27b, miRNA- 29c, miRNA-34c, miRNA-92a-1, miRNA-99a-2, miRNA-101-1, miRNA-101-2, miRNA-125b-1, miRNA-196b, miRNA-425-5p, miRNA-452, miRNA-455, miRNA-502, miRNA-539, e miRNA-660. Extraiu-se os nucleotídeos do banco de dados do GenBank of National Center for Biotechnology Information . Alinhou-se as sequências extraídas com o RNA Folding Form, e o projeto da estrutura molecular tridimensional foi realizado com o RNAComposer. Resultados: demonstrou-se como resultados as sequências dos nucleotídeos e a projeção da estrutura molecular dos miRNAs expressos na hanseníase, e produzimos um tutorial sobre o modelo molecular dos 17 miRNAs expressos em hanseníase através do processamento de suas estruturas moleculares em projeção computacional. Conclusão: foi demonstrado computacionalmente o projeto de estruturas moleculares selecionadas de 17 miRNAs expressos em hanseníase através da biologia computacional.

miR-27a/b Level is Associated with ABCA1 Expression and is a Potential PBMC-Based Biomarker for Coronary Artery Disease

Abstract Background: Coronary artery disease (CAD) is the most common form of cardiac disease with high morbidity and mortality rates. Objectives: In this study, we evaluated the expression of miR-27a and miR-27b as biomarkers in peripheral blood mononuclear cells (PBMCs) of patients with CAD and investigated its correlation with cholesterol-efflux transporter, ATP-binding cassette transporter A1 (ABCA1). Method: This study was performed on 54 men with CAD and 51 healthy, sex- and age-matched control participants. The expression of miR-27a/b and ABCA1 genes in PBMCs were measured by quantitative real-time polymerase chain reaction (qRT-PCR). The protein expression of ABCA1 was assessed by Western blotting. Concurrently, the specificity and sensitivity of miR-27a/b was evaluated through receiver operating characteristic (ROC) curve. The significance level adopted in the statistical analysis was 5%. Results: We found that miR-27a and miR-27b expression were significantly increased, while both mRNA and protein expression of ABCA1 were markedly reduced in the PBMCs of CAD patients in comparison to non-CAD controls. miR-27a/27b expression was also shown to be inversely correlated with ABCA1. ROC analysis showed that the miR-27a had an area under the ROC curve (AUC) of about 92.6 (sensitivity 83.3٪ and specificity 86.6٪) and miR-27b had an AUC of about 93.0 (sensitivity 86.6٪ and specificity 80.0 (%, suggesting the diagnostic potential of miR-27a/b in CAD patients. Conclusions: Our data suggested a possible role of miR-27a/b in CAD pathogenesis. Additionally, we proposed that miR-27a/b expression in PBMCs may have potential clinical implications in the diagnosis of CAD patients, but further validations in large cohorts are required.

The Role of MicroRNAs in the Pathogenesis of Chagas Disease

Abstract Chagas disease (CD) is caused by the protozoan Trypanosoma cruzi and affects about six to seven million individuals worldwide. The distribution of cases is concentrated mainly throughout Latin America, especially in rural areas. This study aims to evaluate microRNAs (miRNAs) as indicators in CD diagnosis for possible contributions to its management. This is a literature review study, carried out in the PubMed, SciELO, Bireme Library, NCBI, Science Direct, and Embase databases, through which a total of 12 articles were included for qualitative analysis. The discussion of this review was based on the thematic axes regarding the modulation of T. cruzi in the immune system and the expression of miRNAs, their production and action, the modulation mechanism of host gene expression, how they act as biomarkers, the importance of miRNAs in the diagnosis of CD, and how their regulation occurs in Chronic Chagas Cardiomyopathy (CCC). Moreover, T. cruzi infection is associated with the downregulation of several miRNAs, which directly related to the findings of hypertrophy and fibrosis. When quantified, these could be used as consistent indicators for CD to support the diagnosis of patients with CD complications, as well as a possible therapeutic target. However, the need for clinical studies that evaluate the usefulness of this biomarker in humans is emphasized, considering that in the present study, only experimental in vitro studies were evaluated, reflecting a lack of studies with practical applicability.

MicroARN circulantes y su potencialidad como futuros biomarcadores óseos / Circulating microRNAs and their potential use as future bone biomarkers / MicroRNAs circulantes e a sua potencialidade como futuros biomarcadores ósseos

Resumen El remodelamiento óseo es ejercido por la actividad de osteoblastos y osteoclastos y puede evaluarse por marcadores bioquímicos de formación y resorción ósea. Sin embargo, el nivel de los marcadores óseos está sometido a una enorme cantidad de variables y, además, carece o presenta escaso valor pronóstico. Los microARN (miARN) fueron recientemente estudiados como una alternativa potencial para ser utilizados como nuevos marcadores óseos. Los miARN son pequeñas moléculas de ARN no codificantes (15-25 nucleótidos) que, a través de la inhibición o degradación de ARN mensajeros, modifican una serie de funciones biológicas. Los miARN específicos de hueso ejercen funciones reguladoras sobre factores transcripcionales involucrados en la osteoblastogénesis y osteoclastogénesis, modificando el remodelamiento óseo. La mayoría de los miARN permanecen dentro de la célula, pero algunos son liberados a la circulación donde pueden ser dosados. Los miARN circulantes presentan gran estabilidad en fluidos biológicos, lo que los hace potenciales candidatos a ser utilizados como nuevos biomarcadores óseos. Cambios en el patrón normal de miARN circulantes específicos de hueso reflejarán modificaciones en el metabolismo óseo y señalan el posible inicio o progresión de enfermedades óseas, como la osteoporosis. Si bien es promisorio, el uso en la práctica clínica de los miARN específicos circulantes como nuevos biomarcadores óseos, ello implica primeramente cumplir con una serie de requisitos que permitan estandarizar las condiciones preanalíticas, analíticas y posanalíticas de estas moléculas. La presente revisión brinda información reciente sobre los estudios clínicos tendientes a determinar el posible uso de los miARN circulantes como nuevos biomarcadores óseos, ya que cuentan con elevada sensibilidad y especificidad diagnósticas, valor predictivo positivo y valor predictivo negativo.

Abstract Osteoblasts and osteoclasts activity determines the level of the bone remodelling process which can be assessed by biochemical markers of bone formation and resorption. However, bone marker levels are subject to a series of variables resand, furthermore, they lack or have little prognostic values. MicroRNAs (miRNAs) were recently studied as a potential alternative to be used as new bone biomarkers. The miRNAs are endogenous small noncoding RNA molecules (15-25 nucleotides) that regulate many biological functions by inhibiting or degrading specific messenger RNAs. Bone-specific miRNAs exert regulatory functions on key transcriptional factors involved in osteoblastogenesis and osteoclastogenesis, modifying the bone remodelling process. Most miRNAs remain within the cell, but some of them are released into circulation. Circulating miRNAs show great stability in biological fluids, which makes them excellent candidates to be used as new bone biomarkers. Modifications in the normal pattern of bone-specific circulating miRNA might reflect changes in bone metabolism, signalling the possible onset or progression of bone diseases, such as osteoporosis. Although promising, the use of specific circulating miRNAs as new bone biomarkers in clinical practice implies fulfilling a series of requirements that lead to standardising the pre-analytical, analytical and post-analytical conditions of these molecules. The present review gives an overview on the clinical studies related to the possible use of specific circulating miRNAs as new bone biomarkers.

Resumo A remodelação óssea é exercida pela atividade dos osteoblastos e osteoclastos e pode ser avaliada para a medição dos marcadores bioquímicos de formação e reabsorção óssea. No entanto, o nível dos marcadores ósseos está sujeito a grande quantidade de variáveis e, além disso, carece ou tem pouco valor prognóstico. Os microARN (miARN) foram estudados recentemente como uma alternativa potencial para serem utilizados como novos marcadores ósseos. Os MicroRNAs (miRNAs) são pequenas moléculas de RNA não codificantes (15-25 nucleotídeos) que, através da inibição ou degradação de RNA mensageiros modificam uma série de funções biológicas. Os miRNAs específicos de osso exercem funções reguladoras sobre fatores transcricionais envolvidos na osteoblastogênese e na osteoclastogênese, modificando o processo de remodelação óssea. A maioria dos miRNAs permanece dentro da célula, mas de RNA mensageiros alguns são liberados na circulação, onde podem ser determinados bioquimicamente. Os miRNAs circulantes apresentam grande estabilidade em fluidos biológicos, o que os torna excelentes candidatos para serem usados como novos biomarcadores ósseos. Mudanças no padrão normal de miRNA circulantes específicos do osso mostrarão mudanças no metabolismo ósseo, sinalizando o possível início ou progressão de doenças ósseas, como osteoporose. Embora promissor, o uso de miRNAs específicas circulantes na prática clínica como novos biomarcadores ósseos, implica primeiramente, atender uma série de requisitos que permitem padronizar as condições pré-analíticas, analíticas e pós-analíticas dessas moléculas. A presente revisão fornece informações recentes sobre estudos clínicos destinados a determinar o possível uso dos miRNAs circulantes como novos biomarcadores ósseos, visto que contam com elevada sensibilidade e especificidade diagnósticas, valor preditivo positivo e valor preditivo negativo.

Mechanism of miRNA-related Signaling Pathway Against Diabetic Nephropathy and Intervention of Traditional Chinese Medicine： A Review / 中国实验方剂学杂志

Diabetic nephropathy （DN）， as one of the common chronic microvascular complications of diabetes， has become the main cause of renal failure in end-stage renal disease in China， increasing the risks of renal dialysis and kidney transplantation in diabetes patients. It is the leading cause of death in people with diabetes. The latest research on DN has focused on the gene level. microRNAs （miRNAs） are a family of endogenous accessible short-chain non-coding RNA molecules. By acting on a particular target， they activate or inhibit its mediated signaling pathways and related molecules， playing an important role in the occurrence and development of DN. They have become microeconomic factors for the prevention and treatment of DN. Traditional Chinese medicine （TCM） has a long history in the diagnosis and treatment of DN and has unique advantages such as significant curative effect and few side effects. A large number of studies have proved that TCM can target miRNA to affect multiple signaling pathways， participate in the regulation of inflammatory response， pyroptosis， mesenchymal transdifferentiation， and other pathological changes， and delay the further development of DN. Therefore， this study discusses the biogenesis mechanism of miRNA and its action mechanism in disease-related signaling pathways based on TCM diagnosis and treatment approaches from the perspective of miRNA， and summarizes the effect of TCM targeting miRNA on the disease-related signaling pathways and on DN. Thus， this study is expected to provide a theoretical reference for exploring the progress of TCM intervention in DN from the perspective of genes.

Molecular mechanism of microRNAs regulating sorafenib resistance in hepatocellular carcinoma / 中国药理学通报

Hepatocellular carcinoma (HCC) is one of the most deadly malignancies in the world, with strong invasiveness, low cure rate, high metastasis rate and poor prognosis. Sorafenib is the most important and effective first-line drug for the treatment of advanced hepatocellular carcinoma, but its clinical efficacy is severely limited by primary and acquired drug resistance. Mi-crornas ( micrornas) are small non-coding Rnas that play a key regulatory role in the occurrence and development of hepatocellular carcinoma and the progression of sorafenib resistance. This paper summarizes the role of micrornas in the initiation and development of sorafenib resistance in hepatocellular carcinoma, in order to further understand the mechanism of sorafenib anti-hep-atocellular carcinoma, and to provide valuable theoretical basis for clinical targeted therapy and prognosis improvement in hepatocellular carcinoma.

From viral hepatitis to hepatocellular carcinoma: The role of exosomal microRNAs / 临床肝胆病杂志

Exosomes are nano-sized phospholipid bilayer vesicles containing abundant and complex biomolecules, such as DNA, mRNAs, microRNAs (miRNAs), lipids, and proteins. Exosomes can be secreted and ingested by most types of cells to transfer information through intercellular transport. After uptake by recipient cells, exosomes release bioactive substances to regulate the biological processes of recipient cells, such as promoting tumor growth and metastasis. Changes of exosomes and their contents are associated with a variety of diseases. In recent years, the role of exosomal miRNAs in the development and progression of hepatocellular carcinoma (HCC) caused by viral hepatitis has attracted wide attention, and exosomal miRNAs from different sources play different roles in this process. This article briefly reviews the research on the role of exosomal miRNAs in the development and progression of viral hepatitis-related HCC and proposes that exosomal miRNAs may be the targets for immunotherapy for HCC microenvironment.

miR-142-3p alleviates house dust mite induced airway inflammation among children / 中国学校卫生

Objective@#To investigate the role of miR-142-3p in alleviation of house dust mite induced allergic airway inflammation among children, so as to provide insights into unraveling the pathogenesis of allergic airway inflammation.@*Methods@#Serum samples were collected from 15 patients with house dust mite induced allergic asthma and 15 healthy children in Jiangnan University Medical Center from September to November 2022, and serum miR-142-3p expression was quantified using a fluorescent quantitative real time PCR (qPCR) assay. The levels of interleukin 6 (IL 6) and tumor necrosis factor α (TNF α) were measured in the cell culture supernatant using enzyme linked immunosorbent assay (ELISA), and the expression of high mobility group box 1 (HMGB1) was detected at transcriptional and translational lvels using qPCR and Western blotting assays. The negative regulation of the HMGB1 gene by miR 142 3p was identified using a dual luciferase gene reporter assay, and the expression of downstream regulatory proteins was determined in human normal lung epithelial cells (BEAS 2B) cells transfected with miR 142 3p using Western blotting. In addition, female C57BL/6 mice at ages of 6-8 weeks were randomly assigned to the phosphate buffer saline (PBS) group, house dust mite sensitized airway inflammation group and house dust mite sensitized airway inflammation + miR 142 3p intervention group. Mouse airway inflammation was evaluated using hematoxylin eosin staining, and the expression of inflammatory cells and inflammatory cytokines were detected in mouse bronchoalveolar lavage fluid (BALF) using Giemsa staining and ELISA.@*Results@#Lower serum miR-142-3p expression was quantified among children with house dust mite induced allergic asthma than among healthy controls (1.33±0.21 vs. 4.74±0.62, t=5.22, P <0.05). Stimulation with dermatophagoides farinae extract (DFE) resulted in a reduction in miR-142-3p expression in BEAS-2B cells (0.82±0.25), while transfection with miR-142-3p mimics resulted in a rise in miR-142-3p expression in BEAS-2B cells (0.55±0.14)( t=3.31, 3.94, P <0.05). Pre treatment with miR-142-3p reduced the expression of IL 6(2.25±0.46)and TNF α(6.58±1.95) ( t=4.86, 3.38, P <0.05) in BEAS 2B cells stimulated with DFE, and treatment with miR-142-3p mimics resulted in a reduction in TLR4 and NF-κB expression in BEAS-2B cells via negative regulation of the HMGB1 expression. In addition, treatment with miR-142-3p was found to alleviate inflammatory cell infiltration in lung tissues of house dust mite sensitized mice, and results in a reduction in interleukin 4 (IL-4)[(107.60±10.43)pg/mL], interleukin 5 (IL 5)[(95.78±13.14)pg/mL] and HMGB1[(2.52±0.87)pg/mL] expression in BALF ( t=10.32, 7.29, 2.90, P <0.05).@*Conclusion@#miR-142-3p alleviates house dust mite induced allergic airway inflammation among children via negative regulation of the HMGB1/TLR4/NF-κB pathway.

Mechanism of microRNA-223-3p inhibiting hepatic stellate cell activation by targeting microtubule-associated protein 1B / 临床肝胆病杂志

ObjectiveTo investigate the effect of microRNA-223-3p （miR-223-3p） on hepatic stellate cell （HSC） activation and its mechanism. MethodsHuman HSC LX2 cells were selected for the study， and LX2 cells were stimulated by TGF-β to establish a model of HSC activation； quantitative real-time PCR was used to measure the change in the expression level of miR-223-3p during HSC activation. After LX2 cells were transfected with miR-223-3p mimic， quantitative real-time PCR， Western blot， and immunofluorescence assay were used to clarify the regulatory effect of miR-223-3p on HSC activation， and dual-luciferase reporter assay was used to verify the association between miR-223-3p and the target gene MAP1B. After LX2 cells were transfected with MAP1B siRNA， Western blot was used to clarify the influence of inhibiting MAP1B expression on HSC activation； after LX2 cells were transfected with miR-223-3p， quantitative real-time PCR and Western blot were used to verify the regulatory effect of miR-223-3p on MAP1B. The independent-samples t test was used for comparison of continuous data between two groups. ResultsHSC in the activated state had a significant reduction in the expression level of miR-223-3p compared with those in the resting state （t=9.12， P<0.001）. Overexpression of miR-223-3p inhibited the mRNA and protein expression levels of the markers for HSC activation alpha-smooth muscle actin and collagen type Ⅰ （mRNA expression： t=8.35 and 12.23， both P<0.01； protein expression： t=16.24 and 20.90， both P<0.001）. The dual-luciferase reporter assay confirmed that MAP1B was a potential target gene of miR-223-3p. Compared with the control group， LX2 cells with miR-223-3p overexpression had significant reductions in the mRNA and protein expression levels of MAP1B （mRNA expression： t=5.95， P<0.01； protein expression： t=11.12， P<0.001）. ConclusionThis study shows that miR-223-3p can inhibit HSC activation by targeting MAP1B.

Action mechanism of microRNA-484 involved in myocardial fibrosis in hypertrophic cardiomyopathy / 中国胸心血管外科临床杂志

@#Objective     To search for the key microRNAs (miRNAs) involved in myocardial fibrosis in hypertrophic cardiomyopathy, and to further explore the mechanisms involved in the regulation of myocardial fibrosis. Methods    Forty-two patients with hypertrophic cardiomyopathy diagnosed and treated surgically in West China Hospital of Sichuan University from January 2014 to June 2018 were selected, including 29 males and 13 females, with a median age of 46 (15-69) years. In the myocardial tissue of patients with hypertrophic cardiomyopathy with different degrees of fibrosis, miRNAs with significantly different expression were screened and further verified at the cellular level. By regulating the expression of the target miRNAs, the expressions of fibrosis-related proteins and target genes were detected respectively. Finally, the target-binding relationship was verified by dual-luciferase reporter gene detection. Results    miR-484 was up-regulated in severely fibrotic myocardial tissue and activated cardiac fibroblasts. After cardiac fibroblasts were activated by TGF-β1, the expression of miR-484 was significantly up-regulated, the expression of fibrosis-related proteins (CollagenⅠ, α-SMA) increased, and the expression of the target gene HIPK1 decreased (P<0.05). After inhibiting the expression of miR-484 by transfection of miR-484 antagomir, the expression of fibrosis-related proteins decreased, while expression of HIPK1 was up-regulated (P<0.05). The detection of dual luciferase reporter gene showed that the luciferase activity of the transfected WT-miRNA-484 mimics group was lower than that of the control group (P<0.05). Conclusion    miR-484 is a pro-fibrotic miRNA that participates in the process of myocardial fibrosis by negatively regulating the expression of HIPK1. It can be used as a regulatory target to provide a therapeutic strategy for myocardial fibrosis.

miRNA-3653-3p reduces the proliferation and invasion ability of endometrial cancer cells by inhibiting PLAC8-Wnt-β-catenin signaling pathway / 肿瘤研究与临床

Objective:To investigate the effect of miRNA-3653-3p (miR-3653-3p) on the proliferation and invasion ability of endometrial cancer cells and its related mechanisms.Methods:The data of 356 endometrial cancer patients were downloaded from the OncoLnc database (http://www.oncolnc.org, updated version 2020), and the Kaplan-Meier method was used to analyze the relationship between the expression level of miR-3653-3p and the overall survival of endometrial cancer patients. The miRGator database (https://bio.tools/mirgator_v2.0, updated version 2019) was used to predict the target gene binding to miR-3653-3p. Human endometrial cancer cell lines AN3CA, HEC-1A, HEC-1B, Ishikawa and human normal endometrial epithelial cell line ESC were selected, and the relative expression level of miR-3653-3p was detected by using quantitative real-time fluorescent polymerase chain reaction (qRT-PCR). The cell line with the lowest expression of miR-3653-3p was selected as the research object, which was divided into the negative control group and miR-3653-3p group, and transfected with the control empty vector plasmid and miR-3653-3p overexpression plasmid. CCK-8 method was used to detect the proliferation ability of cells, Transwell method was used to detect the invasion ability of cells, and qRT-PCR and Western blot were used to detect the expression of miR-3653-3p target gene. The effect of miR-3653-3p on the related protein expression of Wnt- β-catenin signaling pathway was detected by using Western blot.Results:Data analysis in the OncoLnc database showed that compared with endometrial cancer patients with low miR-3653-3p expression, patients with high miR-3653-3p expression had better overall survival ( P < 0.01). Compared with human normal endometrial epithelial ESC, the expression levels of miR-3653-3p in endometrial cancer cell lines AN3CA, HEC-1A, HEC-1B, and Ishikawa were all decreased (all P < 0.05), and the relative expression level of miR-3653-3p was the lowest in HEC-1A cells, and HEC-1A cells were selected for subsequent experiments. The result of CCK-8 showed that compared with the negative control group, the ability of HEC-1A cells in the miR-3653-3p group decreased on the 2nd, 3rd, 4th, and 5th days (all P < 0.05). The result of the Transwell chamber invasion test showed that the number of HEC-1A cell invasion after culturing for 26 h in the negative control group and the miR-3653-3p group was (80±11) and (21±4), respectively, and the difference was statistically significant ( t = 5.18, P < 0.01); compared with the negative control group, the number of cell invasion in the miR-3653-3p group decreased. The miRGator database was used to predict that the target gene of miR-3653-3p might be placenta-specific protein 8 (PLAC8). The relative expression levels of PLAC8 mRNA in HEC-1A cells in the negative control group and miR-3653-3p group were (6.26±0.83) and (0.97±0.31), respectively, and the difference was statistically significant ( t = 6.00, P < 0.01); the relative expression level of PLAC8 mRNA in the miR-3653-3p group was lower than that in the negative control group. Compared with the negative control group, the PLAC8 protein of HEC-1A cells decreased, and the expression of Wnt-β-catenin signaling pathway related proteins β-catenin, transforming growth factor β (TGF-β), GSK-3β, and Rac1 decreased in the miR-3653-3p group. Conclusions:miR-3653-3p may inhibit the proliferation and invasion of endometrial cancer cells by regulating the PLAC8-Wnt-β-catenin signaling pathway.

Clinical application value and prospect of peripheral blood circulating microRNA in hepatocellular carcinoma / 肿瘤研究与临床

Early diagnosis, effective treatment and monitoring of recurrence and metastasis of hepatocellular carcinoma have always been difficult problems for clinicians. MicroRNA (miRNA) plays an important role in hepatocellular carcinoma cells' proliferation, apoptosis, metabolism and other processes, and can be released into body fluids such as blood, urine and saliva. The peripheral blood miRNA in hepatocellular carcinoma can be used as biomarkers for the diagnosis, efficacy assessment, recurrence and metastasis monitoring and prognosis judgment, and may even become therapeutic targets for hepatocellular carcinoma. This article summarizes the research progress of circulating miRNA in peripheral blood as markers for diagnosis and treatment monitoring of hepatocellular carcinoma.

Effects of miRNA-30a-5p and metadherin on proliferation, invasion and migration abilities of breast cancer cells / 肿瘤研究与临床

Objective:To investigate the effects of miRNA-30a-5p (miR-30a-5p) and metadherin (MTDH) on the proliferation, invasion and migration abilities of human breast cancer cells in vitro.Methods:The expression of MTDH in cancer and paracancerous tissues of 112 breast cancer patients in the database and miR-30a-5p in cancer and paracancerous tissues of 103 breast cancer patients in the database were analyzed using data from The Cancer Genome Atlas (TCGA) database. Pearson correlation analysis was used to analyze the correlation between miR-30a-5p and MTDH in 1 222 breast cancer patients in the database; the data were updated to August 2022. Breast cancer MDA-MB-231 cells were divided into negative control group (transfected with negative control sequence), miR-30a-5p overexpression group (transfected with miR-30a-5p mimics), siMTDH group [transfected with small interfering RNA against MTDH (siMTDH)], siMTDH+miR-30a-5p overexpression group (transfected with both siMTDH and miR-30a-5p mimics); cell proliferation ability was detected by methyl thiazol tetrazolium (MTT) assay, cell migration ability was detected by cell scratch assay, cell invasion ability was detected by Transwell assay. The relative expressions of miR-30a-5p, MTDH, matrix metalloproteinase (MMP)-2, MMP-9, vimentin and β-catenin mRNA in cells were detected by quantitative real-time fluorescence polymerase chain reaction (qRT-PCR), and the expressions of MTDH, N-cadherin (N-cad), β-catenin, Snail and MMP-9 proteins were detected by Western blotting.Results:In the TCGA database, MTDH expression level was higher and miR-30a-5p expression level was lower in breast cancer tissues compared with paracancerous tissues, and the differences were statistically significant (both P < 0.001). There was a negative correlation between MTDH and miR-30a-5p expressions in 1 222 patients with breast cancer ( r=-0.134, P < 0.001). Compared with the negative control group, the cell proliferation ability was reduced in both siMTDH group and miR-30a-5p overexpression group at 24, 48 and 72 h (all P < 0.001). The cell scratch healing rate in miR-30a-5p overexpression group and siMTDH group was lower than that in negative control group [(61.6±1.6)%, (54.7±5.9)% vs. (80.3±3.0)%] (both P < 0.05). Compared with the negative control group, The number of migrated cells in miR-30a-5p overexpression group and siMTDH group was less than that in negative control group (881±50, 725±63 vs. 1 172±66) (both P < 0.05). Compared with the negative control group, the relative expressions of MMP-2, MMP-9, vimentin and β-catenin mRNA were all down-regulated in MDA-MB-231 cells of miR-30a-5p overexpression group and siMTDH group (all P < 0.05). Compared with the negative control group, the relative expressions of N-cad, β-catenin, Snail and MMP-9 proteins were down-regulated in MDA-MB-231 cells of miR-30a-5p overexpression group and siMTDH group (all P < 0.05). There was no statistical difference in the number of migrated MDA-MB-231 cells between siMTDH+miR-30a-5p overexpression group and siMTDH group (476±5 vs. 389±46, t = 3.37, P = 0.078). There was no statistical difference in the number of migrated cells between siMTDH+miR-30a-5p overexpression group and miR-30a-5p overexpression group (476±5 vs. 477±22, t = 0.02, P = 0.983). Conclusions:The expression of miR-30a-5p is negatively correlated with the expression of MTDH in breast cancer tissues, and either overexpression of miR-30a-5p or silence of MTDH in breast cancer MDA-MB-231 cells in vitro can inhibit cell proliferation, invasion and migration, but MTDH may not be a target gene of miR-30a-5p.

Identification and study of regulatory axis that miRNA-224-5p targets NEDD4L gene in clear cell renal cell carcinoma / 肿瘤研究与临床

Objective:To screen key genes of renal clear cell carcinoma based on bioinformatics methods, identify possible microRNA (miRNA)-mRNA action axis, and explore the expression of related genes in clear cell renal cell carcinoma tissues and cells.Methods:Gene expression profiles of GSE40435 and GSE71302 datasets were obtained from the Gene Expression Omnibus (GEO) database. TCGA-KIRC datasets were obtained from The Cancer Genome Atlas (TCGA) database. R software was used to identify the differentially expressed mRNA and miRNA, and the functional enrichment analysis was performed. STRING database and Cytoscape software were used to perform the protein interaction analysis. The prognosis-related differentially expressed miRNA was evaluated by the Oncomir database. The potential targeted genes regulated by miRNA were determined by using TargetScan and miRDB targeted gene prediction tools. The tissue samples and clinicopathological features of 34 patients with clear cell renal cell carcinoma in the First Hospital of Shanxi Medical University from June to December 2021 were collected, and normal renal cell line 293T and clear cell renal cell carcinoma cell line 786O were selected. The real-time fluorescence quantitative polymerase chain reaction (qRT-PCR), was used to detect the relative expression of genes; Western blotting and immunohistochemical staining were used to detect the expression levels of the targeted proteins. The dual luciferase reporter gene assay was carried out to verify the targeting relationship between genes.Results:A total of 1 351 differentially expressed mRNA and 50 differentially expressed miRNA were screened and identified. The result of functional enrichment analysis suggested that the fatty acid metabolism pathway and xenobiotic metabolism pathway were suppressed in clear cell renal cell carcinoma, while the apoptosis and immune response pathways were activated. Protein interaction analysis suggested that the signal transduction and protein ubiquitination pathways might play a key role in clear cell renal cell carcinoma. The screening results showed that miRNA-224-5p (miR-224-5p) was most closely associated with clear cell renal cell carcinoma progression and was highly expressed in tumor tissues, and its prognosis-related target gene was NEDD4L. The relative expression of NEDD4L mRNA in clear cell renal cell carcinoma tissues and paraneoplastic tissues were 0.138±0.103 and 1.000±0.026 ( t = 46.23, P < 0.05), and the relative expression of miR-224-5p was 1.000±0.043 and 0.129±0.108 ( t = 45.28, P < 0.05). The differences of NEDD4L mRNA and miR-224-5p expressions in different grades and stages of clear cell renal cell carcinoma tissues were statistically significant (all P < 0.05). The expression of NEDD4L protein was decreased in clear cell renal cell carcinoma. The relative expression of NEDD4L gene in 293T and 786O cells were 1.000±0.125 and 0.210±0.044 ( t = 17.52, P < 0.05); the relative expressions of miR-224-5p gene were 0.209±0.049 and 1.000±0.234 ( t = 10.61, P < 0.05). The relative expressions of NEDD4L mRNA in miRNA mimic group and negative control group were 0.236±0.062 and 1.000±0.024, and the difference was statistically significant ( t = 43.56, P < 0.05). NEDD4L protein expression was reduced in the miRNA mimic group. Dual luciferase reporter gene assay suggested that NEDD4L was a direct target gene of miR-224-5p. Conclusions:In clear cell renal cell carcinoma, miR-224-5p targets and regulates NEDD4L expression, and this mechanism may be related to carcinogenesis and progression of clear cell renal cell carcinoma.

Effects of long non-coding RNA RP11-1212A22.4 on the cell viability and invasive ability of ovarian cancer cell lines through targeting miRNA-483-5p / 肿瘤研究与临床

Objective:To investigate the effects of long non-coding RNA (lncRNA) RP11-1212A22.4 on the cell viability and invasive ability of esophageal cancer cell lines by targeting miRNA-483-5p (miR-483-5p).Methods:The expression of RP11-1212A22.4 in esophageal cancer tissues was analyzed by using GEPIA online database. Real-time quantitative polymerase chain reaction (qRT-PCR) was used to detect the expression of RP11-1212A22.4 in human esophageal cancer cell lines EC9706, KYSE30, TE-13, Eca109 and normal esophageal epithelial cell line HET-1A. The lowest expression level of EC9706 cell line in RP11-1212A22.4 was divided into RP11-1212A22.4 group (transfected with pcDNA-RP11-1212A22.4 plasmid) and the control group (transfected with pcDNA-NC plasmid). The cell viability of EC9706 cell was analyzed by using methyl thiazolyl tetrazolium (MTT) method, and the invasion ability of EC9706 cell was detected by using Transwell assay. The targeting relationship between RP11-1212A22.4 and miR-483-5p was verified by using StarBase database prediction and dual luciferase reporter assay. The relative expression level of miR-483-5p of EC9706 cell in two groups was detected by using qRT-PCR. Western blot was used to detect the expressions of cyclin-dependent kinase 6 (CDK6), matrix metalloproteinase 2 (MMP-2), cyclin-dependent kinase 4 (CDK4), and matrix metalloproteinase 9 (MMP-9) proteins in two groups.Results:In GEPIA online database, compared with adjacent tissues, the relative expression level of RP11-1212A22.4 in esophageal cancer tissues was decreased, and the difference was statistically significant ( P < 0.001). The relative expression levels of RP11-1212A22.4 in esophageal cancer cell lines EC9706, KYSE30, TE-13, Eca109 and normal esophageal mucosal epithelial cell line HET-1A were 0.11±0.08, 0.32±0.09, 0.72±0.09, 0.59±0.13 and 0.97±0.12, and the difference was statistically significant ( F = 40.42, P < 0.001). The relative expression levels of RP11-1212A22.4 in EC9706 cells of RP11-1212A22.4 group and the control group were 11.9±2.4 and 1.0±0.3, respectively, and the difference was statistically significant ( t = 8.89, P < 0.001). Compared with the control group, the cell viability of EC9706 cell in RP11-1212A22.4 group was decreased (all P < 0.05). The number of invasive cells in RP11-1212A22.4 group was lower than that in the control group (48±12 vs. 106±22, t = 4.63, P < 0.001). StarBase database prediction and dual luciferase reporter assay both showed that RP11-1212A22.4 targeted miR-483-5p. The relative expression level of miR-483-5p in RP11-1212A22.4 group was lower than that in the control group (0.24±0.11 vs. 1.02±0.23, t = 5.98, P = 0.001). Compared with the control group, the expressions of CDK6, MMP-2, CDK4 and MMP-9 proteins in the RP11-1212A22.4 group were decreased. Conclusions:RP11-1212A22.4 is lowly expressed in esophageal cancer tissues and cell lines, and it inhibits the cell viability and invasive ability of esophageal cancer cells by targeting miR-483-5p.