MBNL1­AS1 attenuates tumor cell proliferation by regulating the miR­29c­3p/BVES signal in colorectal cancer.

Dysregulation of long non­coding RNAs (lncRNAs) is involved in the development of colorectal cancer (CRC). In the present study, the identification of muscle blind like splicing regulator 1 antisense RNA 1 (MBNL1­AS1) lncRNA was reported. Firstly, Cell Counting Kit­8, EdU and colony formation assays were uesed to explore the role of MBNL1­AS1 in regulating the proliferation of CRC cells. According to TCGA database, it was found that MBNL1­AS1 was correlated with microRNA (miR)­29c­3p and blood vessel epicardial substance (BVES) expression in CRC cells. Then, the regulation among MBNL1­AS1, miR­29C­3P and BVES was detected by dual luciferase reporter assay and the function of MBNL1­AS1/miR­29C­3P/BVES axis was explored by rescue assay. The results demonstrated that MBNL1­AS1 expression was decreased in CRC and was associated with the size of tumors derived from patients with CRC. Functionally, the upregulation of MBNL1­AS1 suppressed CRC cell proliferation in vitro and inhibited tumor growth in vivo, while knockdown of MBNL1­AS1 expression caused the opposite effects. MBNL1­AS1 expression correlated with BVES expression in CRC tissues and MBNL1­AS1 enhanced the stability of BVES mRNA by functioning as a competing endogenous RNA to sponge miR­29c­3p; the latter directly targeted MBNL1­AS1 and BVES mRNA 3'UTR. Collectively, the results indicated that MBNL1­AS1 suppressed CRC cell proliferation by regulating miR­29c­3p/BVES signaling, suggesting that the MBNL1­AS1/miR­29c­3p/BVES axis may be a potential therapeutic target for CRC.

Tumor-suppressor p53 specifically binds to miR-29c-3p and reduces ADAM12 expression in hepatocellular carcinoma.

BACKGROUND: Hepatocellular carcinoma (HCC) is an extremely aggressive malignant tumor associated with high migratory and invasive potential. The present study intends to explore regulatory mechanism of p53/microRNA (miR)-29c-3p/A disintegrin and metalloproteinase 12 (ADAM12) axis in HCC based on clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) technology. METHODS: Putative miR-29c-3p binding sites on ADAM12 3'UTR were verified by a luciferase assay. The binding affinity of p53 to miR-29c-3p was assessed based on CRISPR/Cas9 technology to construct a p53 knockout (p53-/-) HCCLM3 cell line. Furthermore, the effect of p53/miR-29c-3p/ADAM12 was assessed on maligant phenotypes in vitro and tumor formation and metastasis in nude mice. RESULTS: ADAM12 was highly expressed but miR-29c-3p was poorly expressed in HCC. miR-29c-3p inhibited migratory and invasive abilities of HCC cells by targeting ADAM12 expression. p53 was found to target and upregulate miR-29c-3p, thus downregulating ADAM12 and conferring inhibitory effect on HCC cell activities. Moreover, ADAM12 knockout or p53 overexpression reduced HCC tumor formation and metastasis, which were reversed by further silencing of miR-29c-3p. CONCLUSION: The identification of the p53/miR-29c-3p/ADAM12 axis in migration and invasion of HCC may potentially further our understanding of mechanisms underpinning HCC, and also bear translational value as novel molecular targets.

MicroRNA-29c-3p participates in insulin function to modulate polycystic ovary syndrome via targeting Forkhead box O 3.

MicroRNAs (miRNAs) are gene expression regulators and changes in miRNA levels are associated with diabetes, insulin resistance, and inflammation, the latter two of which are characteristic of polycystic ovary syndrome (PCOS). The purpose of this study was to explore the specific mechanism in which miR-29 c-3p participated in insulin function to regulate PCOS by targeting Forkhead box O 3 (Foxo3). Peripheral blood from PCOS patients and healthy volunteers were first collected, and the expression levels of miR-29 c-3p and Foxo3 were detected by reverse transcription quantitative polymerase chain reaction or Western blot. Then human granular tumor cell line (KGN) was treated with insulin, and transfected with plasmid vectors interfering with miR-29 c-3p or Foxo3 expression. Cell proliferation was detected by Cell counting kit-8 and plate cloning, and cell apoptosis was tested by flow cytometry. In addition, PCOS rat model was established. PCOS rats were injected with plasmids vectors interfering with miR-29 c-3p or Foxo3 expression, respectively. Pathological changes in ovarian tissues of rats in each group were observed by hematoxylin-eosin staining, and serum sex hormones and glucose metabolism-related indicators were detected. Finally, via bioinformatics website, luciferase digestion report assay was detected the targeting relationship between miR-29 c-3p and Foxo3. The experimental results showed that miR-29 c-3p was down-regulated in PCOS, but Foxo3 was up-regulated. Up-regulated miR-29 c-3p or down-regulated Foxo3 promoted KGN cell proliferation, inhibited apoptosis in vitro, restored PCOS rat sex hormone levels and improved glucose metabolism in vivo. These results suggest that miR-29 c-3p is involved in insulin function to improve PCOS by targeting Foxo3.

Correlation between the expressions of serum microRNA-29c-3p, microRNA-378a-3p and serum inflammatory factors, myocardial injury indexes and echocardiographic indexes in sepsis patients with myocardial injury and the value of prognosis evaluation / 中国医师进修杂志

Objective:To investigate the correlation between the expressions of microRNA (miR)-29c-3p, miR-378a-3p and serum inflammatory factors, myocardial injury indexes and echocardiographic indexes in sepsis patients with myocardial injury, and analyze the value of prognosis evaluation.Methods:Prospective research methods were used. Two hundred and eighty-six patients with sepsis in Handan Central Hospital from February 2019 to October 2021 were selected. The serum levels of miR-29c-3p and miR-378a-3p were detected by polymerase chain reaction, and serum Troponin I (TnI), creatine kinase isoenzyme MB (CK-MB), myoglobin (Mb), B-type brain natriuretic peptide (BNP), tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), interleukin-1β (IL-1β) and procalcitonin (PCT) were detected. The left ventricular end diastolic diameter (LVEDD), left ventricular end systolic diameter (LVESD), left ventricular ejection fraction (LVEF), stroke output (SV), cardiac output (CO) and the ratio of early diastolic maximum mitral valve velocity to atrial systolic maximum velocity (E/A) were measured by echocardiography. Serum TnI≥0.15 μg/L was defined as myocardial injury, the survival status within 28 d of hospitalization in patients with myocardial injury was recorded, and the acute physiology and chronic health Ⅱ evaluation (APACHE Ⅱ) and sequential organ failure assessment (SOFA) were evaluated. Pearson method was used for correlation analysis; Multivariate Logistic regression was used to analyze the independent risk factors of death within 28 d of hospitalization in sepsis patients with myocardial injury; the receiver operating characteristic (ROC) curve was drawn to analyze the efficacy of miR-29c-3p and miR-378a-3p in predicting the death within 28 d of hospitalization in septic patents with myocardial injury patients.Results:Among 286 patients with sepsis, 131 had myocardial injury (myocardial injury group), and 155 had no myocardial injury (non-myocardial injury group). The miR-29c-3p, Mb, CK-MB, BNP, TnI, TNF-α, IL-6, IL-1β, PCT, CRP, LVEDD and LVESD in myocardial injury group were significantly higher than those in non-myocardial injury group: 5.02 ± 1.69 vs. 2.01 ± 0.57, (102.35 ± 23.56) μg/L vs. (32.15 ± 9.12) μg/L, (25.01 ± 6.09) U/L vs. (13.02 ± 4.16) U/L, (905.23 ± 135.49) ng/L vs. (92.31 ± 26.35) ng/L, (0.23 ± 0.05) μg/L vs. (0.12 ± 0.02) μg/L, (13.41 ± 3.71) μg/L vs. (3.26 ± 0.95) μg/L, (9.02 ± 2.46) ng/L vs. (4.18 ± 1.03) ng/L, (71.45 ± 15.29) ng/L vs. (30.02 ± 6.39) ng/L, (1.05 ± 0.21) μg/L vs. (0.72 ± 0.13) μg/L, (21.35 ± 6.13) mg/L vs. (16.23 ± 4.57) mg/L, (37.45 ± 3.39) mm vs. (34.01 ± 2.15) mm and (50.12 ± 3.49) mm vs. (44.17 ± 3.02) mm, the miR-378a-3p, LVEF, SV, CO and E/A were significantly lower than those in non-myocardial injury group: 1.67 ± 0.36 vs. 3.02 ± 0.79, (46.32 ± 3.26)% vs. (56.24 ± 4.98)%, (48.21 ± 2.81) ml vs. (56.02 ± 3.49) ml, (3.11 ± 0.29) L/min vs. (4.15 ± 0.31) L/min and 0.98 ± 0.21 vs. 1.19 ± 0.32, and there were statistical differences ( P<0.01). Pearson correlation analysis result showed that miR-29c-3p was positive correlation with TNF-α, IL-6, IL-1β, PCT, CRP, Mb, CK-MB, BNP, TnI, LVEDD and LVESD in sepsis patients with myocardial injury ( P<0.01 or <0.05), and it was negative correlation with LVEF, SV, CO and E/A ( P<0.01); miR-378a-3p was negative correlation with TNF-α, IL-6, IL-1β, PCT, CRP, Mb, CK-MB, BNP, TnI, LVEDD and LVESD ( P<0.01 or <0.05), and it was positive correlation with LVEF, SV, CO and E/A P<0.01). Among 131 sepsis patients with myocardial injury, 55 patients died within 28 d of hospitalization (death subgroup), and 76 patients survived (survival subgroup). The mechanical ventilation rate, continuous renal replacement therapy rate, APACHE Ⅱ, SOFA, miR-29c-3p, Mb, CK-MB, TnI, BNP, TNF-α, IL-6, IL-1β, PCT, CRP, LVEDD and LVESD in death subgroup were significantly higher than those in survival subgroup, the miR-378a-3p, LVEF, SV, CO and E/A were significantly lower than those in survival subgroup, and there were statistical differences ( P<0.05 or <0.01). Multivariate Logistic regression analysis result showed that APACHE Ⅱ, TnI, miR-29c-3p were independent risk factors for death within 28 d of hospitalization in sepsis patients with myocardial injury ( OR = 1.203, 2.451 and 1.394; 95% CI 1.085 to 1.334, 1.498 to 4.008 and 1.141 to 1.702; P<0.01); and the miR-378a-3p an independent protective factor for death within 28 d of hospitalization in sepsis patients with myocardial injury ( OR = 0.367, 95% CI 0.217 to 0.622, P<0.01). ROC curve analysis result show that the best cut-off values of miR-29c-3p and miR-378a-3p for predicting the death within 28 d of hospitalization in sepsis patients with myocardial injury were 5.00 and 1.65; the area under the curve of miR-29c-3p combined with miR-378a-3p for predicting the death within 28 d of hospitalization in septic patients with myocardial injury was significantly larger than that of the two separate applications (0.890 vs. 0.695 and 0.732), with a sensitivity of 87.0%, a specificity of 87.3%, and an accuracy of 86.8%. Conclusions:The expression of serum miR-29c-3p is increased and the expression of miR-378a-3p is decreased in sepsis patients with myocardial injury. The expressions of miR-29c-3p and miR-378a-3p are related to the degree of myocardial injury, echocardiogram indexes, the levels of inflammatory factors and prognosis. The miR-29c-3p and miR-378a-3p can be used as potential prognostic indexes for sepsis patients with myocardial injury.

Knockdown of long noncoding RNA colorectal neoplasia differentially expressed inhibits hepatocellular carcinoma progression by mediating the expression of nuclear autoantigenic sperm protein.

Long noncoding RNAs (lncRNAs) play critical roles in the tumorigenesis and progression of hepatocellular carcinoma (HCC). As the most common malignant cancer type in humans, HCC poses a great threat to human health. However, the function of lncRNA colorectal neoplasia differentially expressed (CRNDE) in HCC has not been extensively studied. The chief aim of the present study was to reveal the potential role of CRNDE in HCC. Expression levels of CRNDE in HCC tissues and cell lines were detected by reverse transcription­quantitative (RT­q) PCR, and Cell Counting kit 8, wound­healing and Transwell assays were used to evaluate the influences of CRNDE on in vitro cellular proliferation, migration and invasiveness, respectively. The interaction between CRNDE and microRNA (miR)­29c­3p was determined by dual­luciferase reporter assay, and rescue experiments were conducted to evaluate the interactive relationships between CRNDE and miR­29c­3p or nuclear autoantigenic sperm protein (NASP). CRNDE was found to be upregulated in HCC tissues and cells, and to be positively associated with the poor prognosis of patients with HCC. Furthermore, CRNDE­knockdown suppressed cell proliferation, migration and invasion abilities. Bioinformatics and RT­qPCR analysis indicated miR­29c­3p as a potential target of CRNDE. In line with previous reports, as a tumor suppressor, downregulated expression of miR­29c­3p was observed in HCC. In addition, the present study revealed that miR­29c­3p directly targeted NASP. NASP expression was markedly elevated following transfection with an miR­29c­3p inhibitor, while knocking down CRNDE inhibited NASP expression. Moreover, the effects of CRNDE and NASP on HCC cells were reversed by miR­29c­3p. Collectively, the results of the present study revealed that CRNDE was upregulated and exerted an oncogenic role in HCC by targeting miR­29c­3p, and that the upregulation of CRNDE also promoted NASP expression. These findings indicate a novel role for CRNDE in the progression of HCC.

Transcription Factor DLX5 Promotes Hair Follicle Stem Cell Differentiation by Regulating the c-MYC/microRNA-29c-3p/NSD1 Axis.

INTRODUCTION: Adult stem cell function has been one of the most intensively explored areas of biological and biomedical research, with hair follicle stem cells serving as one of the best model systems. This study explored the role of the transcription factor DLX5 in regulating hair follicle stem cell (HFSC) differentiation. METHODS: HFSCs were isolated, characterized, and assessed for their expression of DLX5, c-MYC, NSD1, and miR-29c-3p using RT-qPCR, Western blot analysis, or immunofluorescence. Next, the ability of HFSCs to proliferate as well as differentiate into either sebaceous gland cells or epidermal cells was determined. The binding of DLX5 to the c-MYC promoter region, the binding of c-MYC to the miR-29c-3p promoter region, and the binding of miR-29c-3p to the 3'-UTR of NSD1 mRNA were verified by luciferase activity assay and ChIP experiments. RESULTS: DLX5 was highly expressed in differentiated HFSCs. DLX5 transcriptionally activated c-MYC expression to induce HFSC differentiation. c-MYC was able to bind the miR-29c-3p promoter and thus suppressed its expression. Without miR-29c-3p mediated suppression, NSD1 was then able to promote HFSC differentiation. These in vitro experiments suggested that DLX5 could promote HFSC differentiation via the regulation of the c-MYC/miR-29c-3p/NSD1 axis. DISCUSSION: This study demonstrates that DLX5 promotes HFSC differentiation by modulating the c-MYC/miR-29c-3p/NSD1 axis and identifies a new mechanism regulating HFSC differentiation.

miR-29c-3p regulates TET2 expression and inhibits autophagy process in Parkinson's disease models.

Autophagy in dopamine (DA) neurons is concerned to be associated with Parkinson's disease (PD), but the detailed mechanism remains unknown. Herein, we aimed to investigate the function of microRNA (miR)-29c-3p in autophagy in PD models. Intraperitoneal injection of MPTP (20 mg/kg) was given to C57BL/6 mice to establish PD mouse model. SH-SY5Y cells were treated with MPP+ (1 mmol/L) to establish in vitro PD model. The results indicated that in the substantia nigra pars compacta (SNpc) DA neurons of PD mice, autophagy was activated accompanied by down-regulated miR-29c-3p and up-regulated ten-eleven translocation 2 (TET2) expression. Up-regulation of miR-29c-3p inhibited TET2 expression and SNpc (including DA neurons) autophagy in PD mice. In vitro PD model confirmed that MPP+ treatment markedly down-regulated miR-29c-3p expression and up-regulated TET2 expression in SH-SY5Y cells in a dose/time-dependent manner. Moreover, miR-29c-3p up-regulation also inhibited autophagy and TET2 expression in vitro. Additionally, TET2 was proved to be targeted and down-regulated by miR-29c-3p. TET2 knockdown inhibited MPP+ -induced autophagy, whereas TET2 over-expression reversed the effects of miR-29c-3p over-expression on SH-SY5Y cell autophagy. Overall, miR-29c-3p over-expression inhibits autophagy in PD models, which may be mediated by TET2. Our finding may provide new insights for regulating autophagy to improve PD progression.

Mesenchymal stem cells-derived extracellular vesicles ameliorate Alzheimer's disease in rat models via the microRNA-29c-3p/BACE1 axis and the Wnt/ß-catenin pathway.

Currently, Alzheimer's disease (AD) cannot be treated effectively. Mesenchymal stem cells (MSCs)-derived extracellular vesicles (EVs) (MSC-EVs) exhibit therapeutic effects on many diseases. This study investigated the mechanism of bone marrow MSC-EVs (BM-MSC-EVs) in a rat model of AD. The cognitive function, amyloid-ß (Aß) plaques, Aß deposition areas and levels of Aß1-42, Aß decomposition-related factors (NEP and IDE), and inflammatory cytokines in BM-MSC-EVs-treated AD rats were measured. The effect of BM-MSC-EVs was studied in AD neuron model. microRNA (miR)-29c-3p and BACE1 expression, as well as levels of Wnt/ß-catenin pathway-related factors in AD and EVs-treated AD models were detected. miR-29c-3p relationship with BACE1 was predicted and confirmed. miR-29c-3p and BACE1 were interfered to verify the mechanism of EVs in AD. The Wnt/ß-catenin pathway inhibitor DKK1 was further added to EVs-treated AD neurons. BM-MSC-EVs showed therapeutic effects on AD rats and neurons. BM-MSC-EVs carried miR-29c-3p into AD neurons. miR-29c-3p targeted BACE1. Silencing miR-29c-3p in BM-MSCs reduced BM-MSC-EV therapeutic effect on AD, which was reversed after BACE1 knockdown. miR-29c-3p targeted BACE1 and activated the Wnt/ß-catenin pathway, and the Wnt/ß-catenin pathway inhibition impaired EV therapeutic effects on AD. We highlighted that BM-MSC-EVs delivered miR-29c-3p to neurons to inhibit BACE1 expression and activate the Wnt/ß-catenin pathway, thereby playing a therapeutic role in AD. This study may provide a novel perspective for elucidating the mechanism of MSCs in the treatment of AD.

MiRNA-29c-3p Promotes Intestinal Inflammation via Targeting Leukemia Inhibitory Factor in Ulcerative Colitis.

BACKGROUND: Dysregulation of micro-RNAs (miRNAs) is profoundly linked to inflammatory bowel diseases (IBD), but little is known about the specific biological functions of miRNAs in IBD. This study sought to elucidate the effect and the underlying target of miR-29c-3p in ulcerative colitis (UC). METHODS: The levels of miR-29c-3p and leukemia inhibitory factor (LIF) were measured in inflamed lesions of UC patients and dextran sulfate sodium (DSS)-induced colitis mice by quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting. MiR-29c-3p was predicted to target LIF by bioinformatics software, which was verified via luciferase reporter assay and transfection of miR-29c-3p mimics or inhibitor. The role of miR-29c-3p/LIF axis in intestinal inflammation was explored in experimental colitis mice and Caco-2 cells. RESULTS: MiR-29c-3p was markedly downregulated while LIF was upregulated in colon tissues of UC patients and DSS-challenged colitis mice as well as in primary intestinal epithelial cells (IECs) and LPS-treated Caco-2 cells. MiR-29c-3p inhibited LIF expression at the transcriptional level via binding to LIF 3'-untranslated region (UTR) in Caco-2 cells. Targeting miR-29c-3p/LIF axis regulated inflammatory cytokines production, cell proliferation and apoptosis. Overexpression of miR-29c-3p aggravated mice experimental colitis via suppressing LIF. CONCLUSION: Our findings demonstrate that the upregulation of miR-29c-3p promotes gut inflammation and the expression of pro-inflammatory mediators via suppressing LIF, thereby modulating the pathogenesis of UC.

MicroRNA­29c­3p acts as a tumor suppressor gene and inhibits tumor progression in hepatocellular carcinoma by targeting TRIM31.

Aberrant expression of microRNAs (miRNAs) has been widely reported in many malignant tumors, and dysregulated miRNAs play an important role in the malignant progression of tumors. It has been reported that miR­29c­3p expression is dysregulated in tumors and promotes the development of tumors, especially in hepatocellular carcinoma (HCC). However, the specific mechanism of miR­29c­3p in HCC is not clear. The present study demonstrated that miR­29c­3p was expressed at low levels in HCC patients and cell lines and that its decreased expression was closely related to poor prognosis of HCC patients. Overexpression of miR­29c­3p could significantly inhibit the proliferation and migration of HCC cells in vitro and suppress the HCC tumor growth in vivo. The luciferase reporter assay demonstrated that miR­29c­3p directly bound to tripartite motif containing 31 (TRIM31) and suppressed TRIM31 expression. Finally, upregulation of TRIM31 could partially abolish the tumor suppressing roles of miR­29c­3p in HCC. Overall, miR­29c­3p, as a tumor suppressor gene, was revealed to inhibit the malignant progression of HCC by reducing the expression of TRIM31 and may be used as a potential therapeutic target for the precise treatment of HCC.

A panel of collagen genes are associated with prognosis of patients with gastric cancer and regulated by microRNA-29c-3p: an integrated bioinformatics analysis and experimental validation.

Background: The systematic expression characteristics and functions of collagen genes in gastric cancer (GC) have not been reported. Through public data integration, combined with bioinformatics analysis, we identified a panel of collagen genes overexpressed in GC. The functions of these genes were analyzed and validated in a GC-related cohort. microRNAs that may potentially target such genes were investigated in vitro. Methods: Four GC-related datasets retrieved from the Gene Expression Omnibus (GEO) were used to extract differentially expressed genes (DEGs) in GC. Functional annotation was performed to identify the potential roles of the identified DEGs. The association of candidate genes involved in the prognosis of GC patients (n=876) was determined using data provided by the Kaplan-Meier-plotter database, The Cancer Genome Atlas Stomach Adenocarcinoma (TCGA-STAD) repository, and a GC-related dataset (GSE15459). The expression characteristics of candidate genes and their associations with clinical parameters were validated in our in-house cohort (n=58). MicroRNAs able to target the identified candidate genes were predicted and confirmed using qRT-PCR, Western blotting, and dual-luciferase reporter assays in vitro. Results: After the integration of four GEO datasets, 76 DEGs were identified. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway analysis indicated that these DEGs were significantly enriched in ECM-related functions and pathways. A group of collagen genes was significantly upregulated in the GC tissues and constituted a protein-protein interaction network as important nodes. Some of these collagen genes were closely associated with poor prognosis in GC patients. Overexpression of COL1A1 and COL4A1 was confirmed in our in-house cohort, and this was related to prognosis and certain clinicopathological parameters. We found that microRNA-29c-3p could directly target COL1A1 and COL4A1 in BGC-823 cells. Conclusions: Collagen genes identified in this study were associated with patient prognosis in GC and may represent diagnostic markers or potential therapeutic targets. Aberrant expression of such candidate genes may be induced by microRNA-29c-3p.

The Smad3-miR-29b/miR-29c axis mediates the protective effect of macrophage migration inhibitory factor against cardiac fibrosis.

Although macrophage migration inhibitory factor (MIF) is known to have antioxidant property, the role of MIF in cardiac fibrosis has not been well understood. We found that MIF was markedly increased in angiotension II (Ang-II)-infused mouse myocardium. Myocardial function was impaired and cardiac fibrosis was aggravated in Mif-knockout (Mif-KO) mice. Functionally, overexpression of MIF and MIF protein could inhibit the expression of fibrosis-associated collagen (Col) 1a1, COL3A1 and α-SMA, and Smad3 activation in mouse cardiac fibroblasts (CFs). Consistently, MIF deficiency could exacerbate the expression of COL1A1, COL3A1 and α-SMA, and Smad3 activation in Ang-II-treated CFs. Interestingly, microRNA-29b-3p (miR-29b-3p) and microRNA-29c-3p (miR-29c-3p) were down-regulated in the myocardium of Ang-II-infused Mif-KO mice but upregulated in CFs with MIF overexpression or by treatment with MIF protein. MiR-29b-3p and miR-29c-3p could suppress the expression of COL1A1, COL3A1 and α-SMA in CFs through targeting the pro-fibrosis genes of transforming growth factor beta-2 (Tgfb2) and matrix metallopeptidase 2 (Mmp2). We further demonstrated that Mif inhibited reactive oxygen species (ROS) generation and Smad3 activation, and rescued the decrease of miR-29b-3p and miR-29c-3p in Ang-II-treated CFs. Smad3 inhibitors, SIS3 and Naringenin, and Smad3 siRNA could reverse the decrease of miR-29b-3p and miR-29c-3p in Ang-II-treated CFs. Taken together, our data demonstrated that the Smad3-miR-29b/miR-29c axis mediates the inhibitory effect of macrophage migration inhibitory factor on cardiac fibrosis.