TRAF2 decrease promotes the TGF-ß-mTORC1 signal in MAFLD-HCC through enhancing AXIN1-mediated Smad7 degradation.

According to recent research, metabolic-associated fatty liver disease (MAFLD) has emerged as an important underlying etiology of hepatocellular carcinoma (HCC). However, the molecular mechanism of MAFLD-HCC is still unclear. Tumor necrosis factor receptor-associated factor 2 (TRAF2) is the key molecule to mediate the signal of inflammatory NF-κB pathway. This study aims to investigate the potential dysregulation of TRAF2 and its biological function in MAFLD-HCC. Huh7 TRAF2-/- demonstrated increased tumor formation ability compared to huh7 TRAF2+/+ when stimulated with transforming growth factor-ß (TGF-ß). The decisive role of TGF-ß in the development of MAFLD-HCC was confirmed through the specific depletion of TGF-ß receptor II gene in the hepatocytes (Tgfbr2ΔHep) of mice. In TRAF2-/- cells treated with TGF-ß, both the glycolysis rate and lipid synthesis were enhanced. We proved the signal of the mechanistic target of rapamycin complex 1 (mTORC1) could be activated in the presence of TGF-ß, and was enhanced in TRAF2-/- cells. The coimmunoprecipitation (co-IP) experiments revealed that TRAF2 fortified the Smurf2-mediated ubiquitination degradation of AXIN1. Hence, TRAF2 depletion resulted in increased Smad7 degradation induced by AXIN1, thus promoting the TGF-ß signal. We also discovered that PLX-4720 could bind with AXIN1 and restrained the tumor proliferation of TRAF2-/- in mice fed with high-fat diet (HFD). Our findings indicate that TRAF2 plays a significant role in the pathogenesis of MAFLD-HCC. The reduction of TRAF2 expression leads to the enhancement of the TGF-ß-mTORC1 pathway by facilitating AXIN1-mediated Smad7 degradation.

Circular RNA circWNK1 inhibits the progression of gastric cancer via regulating the miR-21-3p/SMAD7 axis.

Gastric cancer (GC) is a highly aggressive malignancy with limited treatment options for advanced-stage patients. Recent studies have highlighted the role of circular RNA (circRNA) as a novel regulator of cancer progression in various malignancies. However, the underlying mechanisms by which circRNA contributes to the development and progression of GC remain poorly understood. In this study, we utilized microarrays and real-time quantitative polymerase chain reaction (qRT-PCR) to identify and validate a downregulated circRNA, hsa_circ_0003251 (referred to as circWNK1), in paired GC and normal tissues. Through a series of in vitro and in vivo gain-of-function and loss-of-function assays, we demonstrated that circWNK1 exerts inhibitory effects on the proliferation, migration, invasion, and epithelial-mesenchymal transition (EMT) of GC cells. Additionally, we discovered that circWNK1 acts as a competitive endogenous RNA (ceRNA) for SMAD7 by sequestering miR-21-3p. Our findings were supported by comprehensive biological information analysis, as well as RNA pull-down, luciferase reporter gene, and western blot assays. Notably, the downregulation of circWNK1 in GC cells resulted in reduced SMAD7 expression, subsequently activating the TGF-ß signaling pathway. Collectively, our study reveals that circWNK1 functions as a tumor suppressor in GC by regulating the miR-21-3p/SMAD7-mediated TGF-ß signaling pathway. Furthermore, circWNK1 holds promise as a potential biomarker for the diagnosis and treatment of GC.

TGF-ß1 signaling and Smad7 control T-cell responses in health and immune-mediated disorders.

Transforming growth factor (TGF)-ß1, a member of the TGF-ß superfamily, is produced by many immune and nonimmune cells and has pleiotropic effects on both innate and adaptive immunity, especially in the control of T-cell differentiation and function. Consistently, loss of TGF-ß1 function is associated with exacerbated T-cell-dependent inflammatory responses that culminate in pathological processes in allergic and immune-mediated diseases. In this review, we highlight the roles of TGF-ß1 in immunity, focusing mainly on its ability to promote differentiation of regulatory T cells, T helper (Th)-17, and Th9 cells, thus contributing to amplifying or restricting T-cell responses in health and human diseases (e.g., inflammatory bowel diseases, type 1 diabetes, asthma, and MS). In addition, we discuss the involvement of Smad7, an inhibitor of TGF-ß1 signaling, in immune-mediated disorders (e.g., psoriasis, rheumatoid arthritis, MS, and inflammatory bowel diseases), as well as the discordant results of clinical trials with mongersen, an oral pharmaceutical compound containing a Smad7 antisense oligonucleotide, in patients with Crohn's disease. Further work is needed to ascertain the reasons for such a discrepancy as well as to identify better candidates for treatment with Smad7 inhibitors.

Microvesicles-delivering Smad7 have advantages over microvesicles in suppressing fibroblast differentiation in a model of Peyronie's disease.

BACKGROUND: This study compared the differences of microvesicles (MVs) and microvesicles-delivering Smad7 (Smad7-MVs) on macrophage M1 polarization and fibroblast differentiation in a model of Peyronie's disease (PD). METHODS: Overexpression of Smad7 in rat BMSCs was obtained by pCMV5-Smad7 transfection. MVs were collected from rat BMSCs using ultracentrifugation. In cells, 100 µg/mL of MVs or Smad7-MVs were used to treat the 100 ng/mL of lipopolysaccharide (LPS)-induced RAW264.7 cells or 10 ng/mL of recombinant transforming growth factor-ß1 (TGF-ß1)-induced fibroblasts. The pro-inflammatory cytokines and markers of M1 macrophages were measured in RAW264.7 cells, and the migration and markers of fibroblast differentiation were measured in fibroblasts. In rats, 50 µg of MVs or Smad7-MVs were used to treat the TGF-ß1-induced animals. The pathology of tunica albuginea (TA), the markers of M1 macrophages and fibroblast differentiation in the TA were measured. RESULTS: The MVs or Smad7-MVs treatment suppressed the LPS-induced macrophage M1 polarization and TGF-ß1-induced fibroblast differentiation. Moreover, the Smad7-MVs treatment decreased the fibroblast differentiation compared with the MVs treatment. In the TGF-ß1-induced TA of rats, MVs or Smad7-MVs treatment ameliorated the TA fibrosis by suppressing the macrophage M1 polarization and fibroblast differentiation. There was no significance on the M1-polarized macrophages between the MVs treatment and the Smad7-MVs treatment. Meanwhile, the Smad7-MVs treatment had an edge in terms of suppressing the fibroblast differentiation in the TGF-ß1-induced PD model compared with the MVs treatment. CONCLUSIONS: This study demonstrated that Smad7-MVs treatment had advantages over MVs treatment in suppressing of fibroblast differentiation in a model of PD.

Mir-195-5p targets Smad7 regulation of the Wnt/ß-catenin pathway to promote osteogenic differentiation of vascular smooth muscle cells.

In this study, we sought to investigate the mechanisms of action of miR-195-5p in the osteogenic differentiation of vascular smooth muscle cells (VSMCs), and thereby provide novel insights and a reference for the targeted therapy of arterial media calcification. VSMC differentiation was induced using sodium ß-glycerophosphate, and we investigated the effects of transfecting cells with miR-195-5p mimics, vectors overexpressing Smad7, and the Wnt/ß-catenin pathway inhibitor (KYA1797K) on VSMC differentiation by determining cell viability and apoptosis, and the mRNA and protein expression of factors associated with osteogenic differentiation and the Wnt/ß-catenin pathway. The results revealed that miR-195-5p mimics enhanced the osteogenic differentiation of VSMCs induced by ß-glycerophosphate, whereas the overexpression of Smad7 reversed this phenomenon. In addition, KYA1797K was found to promote the effects of Smad7 overexpression. In conclusion, by targeting, Smad7, miR-195-5p promotes the Wnt/ß-catenin pathway. and thus the osteogenic differentiation of VSMCs. These findings will provide a reference for elucidating the mechanisms whereby miR-195-5p regulates osteogenic differentiation.

Expression of SMADs in orthotopic human endometrium, ovarian endometriosis, and endometriotic lesions in a murine model.

Activin A promotes the development of endometriotic lesions in a murine model of endometriosis, and the immunohistochemical localization of phosphorylated suppressor of mothers against decapentaplegic homolog 2/3 (pSMAD2/3) complex in endometriotic lesions has been reported. Activin may therefore be involved in the development and proliferation of endometriotic cells via the SMAD signaling pathway. However, few detailed reports exist on SMAD7 expression in endometriosis. The purpose of this study was to investigate the expression of pSMAD2/3 or pSMAD3 and SMAD7 in the orthotopic human endometrium, ovarian endometriosis, and endometriotic lesions in a murine model and the effect of activin A on pSMAD2/3 and SMAD7 expression. We established an endometriosis murine model via the intraperitoneal administration of endometrial tissue and blood from donor mice. Activin A was intraperitoneally administered to the activin group. We immunohistochemically evaluated orthotopic endometria, ovarian endometriotic tissues, and endometriotic lesions in the murine model followed by western blotting. We found that pSMAD3 and SMAD7 were expressed in ovarian endometriosis and orthotopic endometria from patients with and without endometriosis. In the murine model, endometriotic lesions expressed pSMAD2/3 and SMAD7 in the activin and control groups, and higher SMAD7 expression was found in the activin group. To the best of our knowledge, this study is the first to show that SMAD7 expression is upregulated in endometriosis. In conclusion, these results suggest that activin A activates the SMAD signaling pathway and promotes the development of endometriotic lesions, thus identifying SMAD7 as a potential therapeutic target for endometriosis.

LncRNA-Smad7 mediates cross-talk between Nodal/TGF-ß and BMP signaling to regulate cell fate determination of pluripotent and multipotent cells.

Transforming growth factor ß (TGF-ß) superfamily proteins are potent regulators of cellular development and differentiation. Nodal/Activin/TGF-ß and BMP ligands are both present in the intra- and extracellular milieu during early development, and cross-talk between these two branches of developmental signaling is currently the subject of intense research focus. Here, we show that the Nodal induced lncRNA-Smad7 regulates cell fate determination via repression of BMP signaling in mouse embryonic stem cells (mESCs). Depletion of lncRNA-Smad7 dramatically impairs cardiomyocyte differentiation in mESCs. Moreover, lncRNA-Smad7 represses Bmp2 expression through binding with the Bmp2 promoter region via (CA)12-repeats that forms an R-loop. Importantly, Bmp2 knockdown rescues defects in cardiomyocyte differentiation induced by lncRNA-Smad7 knockdown. Hence, lncRNA-Smad7 antagonizes BMP signaling in mESCs, and similarly regulates cell fate determination between osteocyte and myocyte formation in C2C12 mouse myoblasts. Moreover, lncRNA-Smad7 associates with hnRNPK in mESCs and hnRNPK binds at the Bmp2 promoter, potentially contributing to Bmp2 expression repression. The antagonistic effects between Nodal/TGF-ß and BMP signaling via lncRNA-Smad7 described in this work provides a framework for understanding cell fate determination in early development.

MicroRNA-30e-3p reduces coronary microembolism-induced cardiomyocyte pyroptosis and inflammation by sequestering HDAC2 from the SMAD7 promoter.

This study investigates the mechanism by which microRNA (miR)-30e-3p reduces coronary microembolism (CME)-induced cardiomyocyte pyroptosis and inflammation. Cardiac function tests, histological staining, and transmission electron microscopy were performed on CME-model rats injected with adeno-associated viral vectors. Cardiomyocytes were transfected 24 h before a cellular model of pyroptosis was established via treatment with 1 µg/mL lipopolysaccharide (LPS) for 4 h and 5 mM ATP for 30 min. Pyroptosis, inflammation, and Wnt/ß-catenin signaling in cardiomyocytes were detected. Dual-luciferase reporter assays and/or RNA pull-down assays were performed to verify the binding of miR-30e-3p to HDAC2 mRNA or HDAC2 to the SMAD7 promoter. Chromatin immunoprecipitation was used to assess the level of H3K27 acetylation at the SMAD7 promoter. miR-30e-3p and SMAD7 expression levels were downregulated and HDAC2 expression was upregulated with CME. The overexpression of miR-30e-3p restored cardiac functions in CME-model rats and reduced serum cTnI, IL-18, and IL-1ß levels, microinfarcts, inflammatory cell infiltration, apoptosis, collagen content, and GSDMD-N, cleaved caspase-1, and NLRP3 expression in the myocardium, but these effects were reversed by SMAD7 knockdown. The overexpression of miR-30e-3p or knockdown of HDAC2 reduced LDH, IL-18, and IL-1ß secretion, propidium iodide intake, and GSDMD-N, NLRP3, cleaved caspase-1, Wnt3a, Wnt5a, and ß-catenin expression in the cardiomyocyte model. miR-30e-3p inhibited the expression of HDAC2 by binding HDAC2 mRNA. HDAC2 repressed the expression of SMAD7 by catalyzing H3K27 deacetylation at the SMAD7 promoter. miR-30e-3p, by binding HDAC2 to promote SMAD7 expression, reduces CME-induced cardiomyocyte pyroptosis and inflammation.

CDCA7 promotes TGF-ß-induced epithelial-mesenchymal transition via transcriptionally regulating Smad4/Smad7 in ESCC.

Cell division cycle associated 7 (CDCA7) is a copy number amplification gene that contributes to the metastasis and invasion of tumors, including esophageal squamous cell carcinoma (ESCC). This present study aimed at clarifying whether high expression of CDCA7 promotes the metastasis and invasion of ESCC cell lines and exploring the underlying mechanisms implicated in epithelial-mesenchymal transition (EMT) of ESCC. The role of CDCA7 in the regulation of ESCC metastasis and invasion was evaluated using ESCC cell lines. Expression of EMT-related markers including E-cadherin, N-cadherin, Vimentin, Snail, and Slug, transforming growth factor ß (TGF-ß) signaling pathway including Smad2/3, p-Smad2/3, Smad4, and Smad7 were detected in CDCA7 knockdown and overexpressed cell lines. Dual-luciferase reporter assay and rescue assay were used to explore the underlying mechanisms that CDCA7 contributed to the metastasis and invasion of ESCC. High CDCA7 expression significantly promoted the metastasis and invasion of ESCC cell lines both in vivo and in vitro. Additionally, the expression of CDCA7 positively correlated with the expression of N-cadherin, Vimentin, Snail, Slug, TGF-ß signaling pathway and negatively correlated with the expression of E-cadherin. Furthermore, CDCA7 transcriptionally regulated the expression of Smad4 and Smad7. Knockdown of CDCA7 inhibited the TGF-ß signaling pathway and therefore inhibited EMT. Our data indicated that CDCA7 was heavily involved in EMT by regulating the expression of Smad4 and Smad7 in TGF-ß signaling pathway. CDCA7 might be a new therapeutic target in the suppression of metastasis and invasion of ESCC.

The role of endogenous Smad7 in regulating macrophage phenotype following myocardial infarction.

Smad7 restrains TGF-ß responses, and has been suggested to exert both pro- and anti-inflammatory actions that may involve effects on macrophages. Myocardial infarction triggers a macrophage-driven inflammatory response that not only plays a central role in cardiac repair, but also contributes to adverse remodeling and fibrosis. We hypothesized that macrophage Smad7 expression may regulate inflammation and fibrosis in the infarcted heart through suppression of TGF-ß responses, or via TGF-independent actions. In a mouse model of myocardial infarction, infiltration with Smad7+ macrophages peaked 7 days after coronary occlusion. Myeloid cell-specific Smad7 loss in mice had no effects on homeostatic functions and did not affect baseline macrophage gene expression. RNA-seq predicted that Smad7 may promote TREM1-mediated inflammation in infarct macrophages. However, these alterations in the transcriptional profile of macrophages were associated with a modest and transient reduction in infarct myofibroblast infiltration, and did not affect dysfunction, chamber dilation, scar remodeling, collagen deposition, and macrophage recruitment. In vitro, RNA-seq and PCR arrays showed that TGF-ß has profound effects on macrophage profile, attenuating pro-inflammatory cytokine/chemokine expression, modulating synthesis of matrix remodeling genes, inducing genes associated with sphingosine-1 phosphate activation and integrin signaling, and inhibiting cholesterol biosynthesis genes. However, Smad7 loss did not significantly affect TGF-ß-mediated macrophage responses, modulating synthesis of only a small fraction of TGF-ß-induced genes, including Itga5, Olfml3, and Fabp7. Our findings suggest a limited role for macrophage Smad7 in regulation of post-infarction inflammation and repair, and demonstrate that the anti-inflammatory effects of TGF-ß in macrophages are not restrained by endogenous Smad7 induction.

Hydronidone ameliorates liver fibrosis by inhibiting activation of hepatic stellate cells via Smad7-mediated degradation of TGFßRI.

BACKGROUND AND PURPOSE: Liver fibrosis is a wound-healing reaction that eventually leads to cirrhosis. Hydronidone is a new pyridine derivative with the potential to treat liver fibrosis. In this study, we explored the antifibrotic effects of hydronidone and its potential mode of action. METHODS: The anti-hepatic fibrosis effects of hydronidone were studied in carbon tetrachloride (CCl4 )- and 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC)- induced animal liver fibrosis. The antifibrotic mechanisms of hydronidone were investigated in hepatic stellate cells (HSCs). The antifibrotic effect of hydronidone was further tested after Smad7 knockdown in HSCs in mouse models of fibrosis. RESULTS: In animal models, hydronidone attenuated liver damage and collagen accumulation, and reduced the expression of fibrosis-related genes. Hydronidone decreased the expression of fibrotic genes in HSCs. Impressively, hydronidone significantly upregulated Smad7 expression and promoted the degradation of transforming growth factor ß receptor I (TGFßRI) in HSCs and thus inhibited the TGFß-Smad signalling pathway. Specific knockdown of Smad7 in HSCs in vivo blocked the antifibrotic effect of hydronidone. CONCLUSION: Hydronidone ameliorates liver fibrosis by inhibiting HSCs activation via Smad7-mediated TGFßRI degradation. Hydronidone is a potential drug candidate for the treatment of liver fibrosis.

HIF-1α-induced upregulated miR-322 forms a feedback loop by targeting Smurf2 and Smad7 to activate Smad3/ß-catenin/HIF-1α, thereby improving myocardial ischemia-reperfusion injury.

Myocardial ischemia/reperfusion injury (MIRI) is a major cause of heart failure after myocardial infarction. It has been reported that miR-322 is involved in MIRI progression, while the molecular mechanism of miR-322 in regulating MIRI progression needs to be further probed. MIRI cell model was established by oxygen-glucose deprivation/reoxygenation (OGD/R). Cell viability was assessed using MTS assay. Flow cytometry and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling staining were employed to analyze cell apoptosis. In addition, the interactions between miR-322, Smad7/Smurf2, hypoxia-inducible factor alpha (HIF-1α), and ß-catenin were verified by dual-luciferase reporter gene assay. Our results displayed that miR-322 was significantly downregulated in OGD/R-treated H9c2 cells, and its overexpression resulted in increased cell viability and reduced the apoptosis. Smurf2 and Smad7 were identified as the direct targets of miR-322. Smad7 knockdown or Smurf2 knockdown increased OGD/R-treated H9c2 cell viability and suppressed the apoptosis. Meanwhile, miR-322 mimics abolished the mitigating effect of Smad7 or Smurf2 overexpression on MIRI. In addition, the Smad3/ß-catenin pathway was identified as the downstream pathway of Smurf2/Smad7. Moreover, it was found that HIF-1α interacted with the miR-322 promoter, and ß-catenin interacted with the HIF-1α promoter to form a loop. HIF-1α-induced upregulated miR-322 activated the Smad3/ß-catenin pathway by targeting Smurf2 and Smad7 to improve MIRI; meanwhile, ß-catenin/HIF-1α formed a positive feedback loop to continuously improve MIRI.

Transcription factor Sp2 promotes TGFB-mediated interstitial cell osteogenic differentiation in bicuspid aortic valves through a SMAD-dependent pathway.

Calcification of the bicuspid aortic valve (BAV) involves differential expression of various RNA genes, which is achieved through complex regulatory networks that are controlled in part by transcription factors and microRNAs. We previously found that miR-195-5p regulates the osteogenic differentiation of valvular interstitial cells (VICs) by targeting the TGF-ß pathway. However, the transcriptional regulation of miR-195-5p in calcified BAV patients is not yet clear. In this study, stenotic aortic valve tissues from patients with BAVs and tricuspid aortic valves (TAVs) were collected. Candidate transcription factors of miR-195-5p were predicted by bioinformatics analysis and tested in diseased valves and in male porcine VICs. SP2 gene expression and the corresponding protein levels in BAV were significantly lower than those in TAV, and a low SP2 expression level environment in VICs resulted in remarkable increases in RNA expression levels of RUNX2, BMP2, collagen 1, MMP2, and MMP9 and the corresponding proteins. ChIP assays revealed that SP2 directly bound to the transcription promoter region of miR-195-5p. Cotransfection of SP2 shRNA and a miR-195-5p mimic in porcine VICs demonstrated that SP2 repressed SMAD7 expression via miR-195-5p, while knockdown of SP2 increased the mRNA expression of SMAD7 and the corresponding protein and attenuated Smad 2/3 expression. Immunofluorescence staining of diseased valves confirmed that the functional proteins of osteogenesis differentiation, including RUNX2, BMP2, collagen 1, and osteocalcin, were overexpressed in BAVs. In Conclusion, the transcription factor Sp2 is expressed at low levels in VICs from BAV patients, which has a negative impact on miR-195-5p expression by binding its promoter region and partially promotes calcification through a SMAD-dependent pathway.

Overexpression of SMAD7 improves the function of EGFR-targeted human CAR-T cells against non-small-cell lung cancer.

BACKGROUND AND OBJECTIVE: Recent advancements in immunotherapy led to the development of Chimeric antigen receptor (CAR) T-cell therapy. CAR-T cell therapy in non-small cell lung cancer (NSCLC) is hindered by overexpression of transforming growth factor (TGFß) in the cancer cells that have a negative regulatory role on T-cells activity. This study characterized CAR-T with overexpression of mothers against decapentaplegic homologue 7 (SMAD), a negative regulator of TGFß downstream signalling. METHODS: We have generated three types of CAR-T: epidermal growth factor receptor (EGFR)-CAR-T, EGFR-dominant-negative TGFbeta receptor 2 (DNR)-CAR-T, and EGFR-SMAD7-CAR-T by transducing human T-cells with the lentivirus constructs. We characterized the proliferation, expression of proinflammatory cytokines, activation profile, and lysis capacity in co-cultures with A549 lung carcinoma cells with and without TGFß neutralizing antibodies. We also tested the therapeutic potential of EGFR-SMAD7-CAR-T in the A549 cells tumour-bearing mice model. RESULTS: Both EGFR-DNR-CAR-T and EGFR-SMAD7-CAR-T demonstrated a higher proliferation rate and lysis capacity to A549 than traditional EGFR-CAR-T. Neutralization of TGFß by the antibodies resulted in increased performance of EGFR-CAR-T. In vivo, both EGFR-DNR-CAR-T and EGFR-SMAD7-CAR-T resulted in complete tumour resorption by day 20, whereas conventional CAR-T only has a partial effect. CONCLUSION: We demonstrated the high efficacy and resistance to negative TGFß regulation of EGFR-SMAD7-CAR-T comparable with EGFR-DNR-CAR-T and without the systemic effect of TGFß inhibition.

Exosomal miR-21-5p derived from multiple myeloma cells promote renal epithelial-mesenchymal transition through targeting TGF-ß/SMAD7 signalling pathway.

The prognosis of multiple myeloma (MM) patients combined with renal insufficiency is poor. Renal fibrosis is an important pathological cause for MM patients combined with renal insufficiency. It is reported that epithelial-mesenchymal transition (EMT) of renal proximal tubular epithelial cells is an important mechanism in renal fibrosis. We speculated that EMT might play an important role in the renal insufficiency of MM with unclear mechanism. MM cells derived exosomes could affect the function of targeted cells by delivering microRNAs (miRNAs). Literature has shown that the expression of miR-21 is closely related to EMT. In this research, we found that co-culture of HK-2 cells (human renal proximal tubular epithelial cells) and exosomes derived from MM cells promoted the EMT of HK-2 cells, resulting in the down-regulation of epithelial-related marker (E-cadherin), and up-regulation of stroma-related marker (Vimentin). Meanwhile, the expression of SMAD7, one of the downstream targets in the TGF-ß signalling pathway, was suppressed and the expression of TGF-ß was increased. After transfecting the inhibitor of miR-21 in MM cells, the expression of miR-21 in exosomes secreted by MM cells was significantly decreased, and the co-culture of these treated exosomes and HK-2 cells inhibited the EMT of HK-2 cells. In conclusion, these findings showed that exosomal miR-21 derived from MM cells could promote renal EMT through targeting TGF-ß/SMAD7 signalling pathway.

Current perspectives on inhibitory SMAD7 in health and disease.

Transforming growth factor ß (TGF-ß) family members play an extensive role in cellular communication that orchestrates both early development and adult tissue homeostasis. Aberrant TGF-ß family signaling is associated with a pathological outcome in numerous diseases, and in-depth understanding of molecular and cellular processes could result in therapeutic benefit for patients. Canonical TGF-ß signaling is mediated by receptor-regulated SMADs (R-SMADs), a single co-mediator SMAD (Co-SMAD), and inhibitory SMADs (I-SMADs). SMAD7, one of the I-SMADs, is an essential negative regulator of the pleiotropic TGF-ß and bone morphogenetic protein (BMP) signaling pathways. In a negative feedback loop, SMAD7 inhibits TGF-ß signaling by providing competition for TGF-ß type-1 receptor (TßRI), blocking phosphorylation and activation of SMAD2. Moreover, SMAD7 recruits E3 ubiquitin SMURF ligases to the type I receptor to promote ubiquitin-mediated proteasomal degradation. In addition to its role in TGF-ß and BMP signaling, SMAD7 is regulated by and implicated in a variety of other signaling pathways and functions as a mediator of crosstalk. This review is focused on SMAD7, its function in TGF-ß and BMP signaling, and its role as a downstream integrator and crosstalk mediator. This crucial signaling molecule is tightly regulated by various mechanisms. We provide an overview of the ways by which SMAD7 is regulated, including noncoding RNAs (ncRNAs) and post-translational modifications (PTMs). Finally, we discuss its role in diseases, such as cancer, fibrosis, and inflammatory bowel disease (IBD).

Long noncoding RNA NEAT1 promotes cardiac fibrosis in heart failure through increased recruitment of EZH2 to the Smad7 promoter region.

Cardiac fibrosis, a well-known major pathological process that ultimately leads to heart failure, has attracted increasing attention and focus in recent years. A large amount of research indicates that long noncoding RNAs (lncRNAs) play an important role in cardiac fibrosis, but little is known about the specific function and mechanism of the lncRNA NEAT1 in the progression of cardiac fibrosis to heart failure. In the present study, we have demonstrated that the lncRNA NEAT1 is upregulated in patients with heart failure. Similarly, the expression of Neat1 was also increased in the left ventricular tissue of transverse aortic constriction (TAC) surgery mice and cardiac fibroblasts treated with TGF-ß1. Further, gain-of-function and loss-of-function experiments showed that silencing of Neat1 attenuated cardiac fibrosis, while overexpression of Neat1 with adenovirus significantly aggravated the in vitro progression of fibrosis. With regard to the underlying mechanism, our experiments showed that Neat1 recruited EZH2 to the promoter region of Smad7 through physical binding of EZH2 to the promoter region, as a result of which Smad7 expression was inhibited and the progression of cardiac fibrosis was ultimately exacerbated. We found that the introduction of shNeat1 carried by adeno-associated virus-9 significantly ameliorated cardiac fibrosis and dysfunction caused by TAC surgery in mice. Overall, our study findings demonstrate that the lncRNA Neat1 accelerates the progression of cardiac fibrosis and dysfunction by recruiting EZH2 to suppress Smad7 expression. Thus, NEAT1 may serve as a target for the treatment of cardiac fibrosis.

Overexpression of Mothers Against Decapentaplegic Homolog 7 Activates the Yes-Associated Protein/NOTCH Cascade and Promotes Liver Carcinogenesis in Mice and Humans.

BACKGROUND AND AIMS: Mothers against decapentaplegic homolog (SMAD) 7 is an antagonist of TGF-ß signaling. In the present investigation, we sought to determine the relevance of SMAD7 in liver carcinogenesis using in vitro and in vivo approaches. APPROACH AND RESULTS: We found that SMAD7 is up-regulated in a subset of human HCC samples with poor prognosis. Gene set enrichment analysis revealed that SMAD7 expression correlates with activated yes-associated protein (YAP)/NOTCH pathway and cholangiocellular signature genes in HCCs. These findings were substantiated in human HCC cell lines. In vivo, overexpression of Smad7 alone was unable to initiate HCC development, but it significantly accelerated c-Myc/myeloid cell leukemia 1 (MCL1)-induced mouse HCC formation. Consistent with human HCC data, c-Myc/MCL1/Smad7 liver tumors exhibited an increased cholangiocellular gene expression along with Yap/Notch activation and epithelial-mesenchymal transition (EMT). Intriguingly, blocking of the Notch signaling did not affect c-Myc/MCL1/Smad7-induced hepatocarcinogenesis while preventing cholangiocellular signature expression and EMT, whereas ablation of Yap abolished c-Myc/MCL1/Smad7-driven HCC formation. In mice overexpressing a myristoylated/activated form of AKT, coexpression of SMAD7 accelerated carcinogenesis and switched the phenotype from HCC to intrahepatic cholangiocarcinoma (iCCA) lesions. In human iCCA, SMAD7 expression was robustly up-regulated, especially in the most aggressive tumors, and directly correlated with the levels of YAP/NOTCH targets as well as cholangiocellular and EMT markers. CONCLUSIONS: The present data indicate that SMAD7 contributes to liver carcinogenesis by activating the YAP/NOTCH signaling cascade and inducing a cholangiocellular and EMT signature.

Exosomal microRNA-16-5p from macrophage exacerbates atherosclerosis via modulating mothers against decapentaplegic homolog 7.

OBJECTIVE: Studies have probed the function of microRNA (miR)-16-5p in the progression of atherosclerosis (AS), while the regulatory function of exosomal miR-16-5p from macrophage on AS remains largely unknown. This study commits to exploring the efficiency of exosomal miR-16-5p from macrophage on AS via modulating mothers against decapentaplegic homolog 7 (SMAD7). METHODS: Macrophages were cultured and transfected with miR-16-5p antagomir, then, the exosomes from macrophages were extracted. The AS mouse model was established, and miR-16-5p or SMAD7 expression in AS mice was detected. Thereafter, the effects of macrophage-derived exosomes, miR-16-5p or SMAD7 on serum inflammatory response, oxidative stress response, pathological changes and apoptosis in AS mice were observed by immunohistochemical and biochemical analysis. Finally, the binding relation between miR-16-5p and SMAD7 was examined. RESULTS: MiR-16-5p was elevated while SMAD7 was depleted in AS mice. Macrophage-derived exosomes aggravated AS progression via facilitating inflammatory response and oxidative stress, exacerbating pathological changes and increasing cell apoptosis in AS mice; while downregulation of miR-16-5p reversed the exacerbation of AS progression by macrophage-derived exosomes in AS mice. MiR-16-5p targeted SMAD7, and the down-regulated SMAD7 reversed the impacts of depleted miR-16-5p on AS progression. CONCLUSION: Exosomal miR-16-5p from macrophages aggravates AS progression via downregulating SMAD7 expression. This study provides novel therapeutic targets for AS treatment from the animal level.

Vascular smooth muscle cell-specific miRNA-214 knockout inhibits angiotensin II-induced hypertension through upregulation of Smad7.

Vascular remodeling is a prominent trait during the development of hypertension, attributable to the phenotypic transition of vascular smooth muscle cells (VSMCs). Increasing studies demonstrate that microRNA plays an important role in this process. Here, we surprisingly found that smooth muscle cell-specific miR-214 knockout (miR-214 cKO) significantly alleviates angiotensin II (Ang II)-induced hypertension, which has the same effect as that of miR-214 global knockout mice in response to Ang II stimulation. Under the treatment of Ang II, miR-214 cKO mice exhibit substantially reduced systolic blood pressure. The vascular medial thickness and area in miR-214 cKO blood vessels were obviously reduced, the expression of collagen I and proinflammatory factors were also inhibited. VSMC-specific deletion of miR-214 blunts the response of blood vessels to the stimulation of endothelium-dependent and -independent vasorelaxation and phenylephrine and 5-HT induced vasocontraction. In vitro, Ang II-induced VSMC proliferation, migration, contraction, hypertrophy, and stiffness were all repressed with miR-214 KO in VSMC. To further explore the mechanism of miR-214 in the regulation of the VSMC function, it is very interesting to find that the TGF-ß signaling pathway is mostly enriched in miR-214 KO VSMC. Smad7, the potent negative regulator of the TGF-ß/Smad pathway, is identified to be the target of miR-214 in VSMC. By which, miR-214 KO sharply enhances Smad7 levels and decreases the phosphorylation of Smad3, and accordingly alleviates the downstream gene expression. Further, Ang II-induced hypertension and vascular dysfunction were reversed by antagomir-214. These results indicate that miR-214 in VSMC established a crosstalk between Ang II-induced AT1R signaling and TGF-ß induced TßRI /Smad signaling, by which it exerts a pivotal role in vascular remodeling and hypertension and imply that miR-214 has the potential as a therapeutic target for the treatment of hypertension.