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.

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.

SMAD7 expression in CAR-T cells improves persistence and safety for solid tumors.

Despite the tremendous progress of chimeric antigen receptor T (CAR-T) cell therapy in hematological malignancies, their application in solid tumors has been limited largely due to T-cell exhaustion in the tumor microenvironment (TME) and systemic toxicity caused by excessive cytokine release. As a key regulator of the immunosuppressive TME, TGF-ß promotes cytokine synthesis via the NF-κB pathway. Here, we coexpressed SMAD7, a suppressor of TGF-ß signaling, with a HER2-targeted CAR in engineered T cells. These novel CAR-T cells displayed high cytolytic efficacy and were resistant to TGF-ß-triggered exhaustion, which enabled sustained tumoricidal capacity after continuous antigen exposure. Moreover, SMAD7 substantially reduced the production of inflammatory cytokines by antigen-primed CAR-T cells. Mechanistically, SMAD7 downregulated TGF-ß receptor I and abrogated the interplay between the TGF-ß and NF-κB pathways in CAR-T cells. As a result, these CAR-T cells persistently inhibited tumor growth and promoted the survival of tumor-challenged mice regardless of the hostile tumor microenvironment caused by a high concentration of TGF-ß. SMAD7 coexpression also enhanced CAR-T-cell infiltration and persistent activation in patient-derived tumor organoids. Therefore, our study demonstrated the feasibility of SMAD7 coexpression as a novel approach to improve the efficacy and safety of CAR-T-cell therapy for solid tumors.

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.

SMAD7 gene polymorphisms and their influence on patients with colorectal cancer.

Colorectal cancer (CRC) is a prevalent malignant tumor, and its pathogenesis is still not fully understood. Studies have shown that SMAD7 gene polymorphisms can affect CRC susceptibility, but the results have been inconsistent and require additional confirmation. Our study aimed to evaluate the effect of SMAD7 variants on the risk of CRC in the Chinese Han population. A total of five single nucleotide polymorphisms (SNPs) in SMAD7 were genotyped among 696 CRC patients and 696 healthy participants using the MassARRAY iPLEX platform. SNPs were evaluated for their associations with CRC using logistic regression analysis under multiple genetic models. The false-positive report probability (FPRP) analysis was used to validate the positive findings. Our study indicated that rs11874392 showed an increased association with CRC risk (odds ratio, 1.31; 95% confidence interval, 1.04-1.67; p = 0.024). Stratified analysis showed that rs11874392 might increase the risk of CRC in females (OR = 1.70, p = 0.028), individuals with smoking (OR = 1.87, p = 0.026), and drinking (OR = 1.38, p = 0.027). The rs11874392 was found to be related to an elevated risk of rectal cancer (OR = 1.73, p = 0.003), but not with colon cancer. FPRP analysis demonstrated that all of these associations were statistically significant (FPRP <0.2). Additionally, rs11874392 was the strongest predictive model for CRC. This study provides evidence that the SMAD7 rs11874392 is related to an increased susceptibility to CRC.

[Silencing of SMAD family member 3 promotes M2 polarization of macrophages and the expression of SMAD7 in rheumatoid arthritis].

Objective To investigate the effect of SMAD family member 3(SMAD3) silenced by small interfering RNA (siRNA) on macrophage polarization and transforming growth factor ß1 (TGF-ß1)/ SMAD family signaling pathway in rheumatoid arthritis (RA). Methods RA macrophages co-cultured with rheumatoid arthritis fibroblast-like synoviocytes (RA-FLS) were used as a cell model. TGF-ß1 was used to stimulate macrophages, and SMAD3-specific siRNA (si-SMAD3) and negative control siRNA (si-NC) were transfected into human RA macrophages co-cultured in TranswellTM chamber. The expression of SMAD3 mRNA was detected by real-time fluorescence quantitative PCR, and the expression of TGF-ß1, SMAD3 and SMAD7 protein was detected by Western blot analysis. The contents of TGF-ß1 and IL-23 in cell culture supernatant were determined by ELISA. Cell proliferation was detected by CCK-8 assay. TranswellTM chamber was used to measure cell migration. Results Compared with the model group and the si-NC group, the expression of TGF-ß1, SMAD3 mRNA and protein in RA macrophages decreased significantly after silencing SMAD3. In addition, the secretion of IL-23 decreased significantly, and the cell proliferation activity and cell migration were inhibited, with high expression of SMAD7. Conclusion Knockdown of SMAD3 can promote M2 polarization and SMAD7 expression in RA macrophages.

CircMACF1 alleviates myocardial fibrosis after acute myocardial infarction by suppressing cardiac fibroblast activation via the miR-16-5p/SMAD7 axis.

Circular RNAs (circRNAs) played a pivotal role in myocardial fibrosis after acute myocardial infarction (AMI). The activation of cardiac fibroblasts (CFs) and accumulation of extracellular matrix are the main characteristics of myocardial fibrosis. In our research, we aimed to elucidate the functional roles of circMACF1 in CF activation after AMI as well as the underlying mechanism. Human CFs were activated by TGF-ß1 treatment. qPCR and western blotting were performed to investigate gene and protein expression. CCK-8 and transwell assays were carried out to measure cell proliferation, and migration. Immunofluorescence was used to investigate α-SMA level. The interaction between miR-16-5p and circMACF1 or SMAD7 was revealed by RIP or dual luciferase reporter gene assays. CircMACF1 and SMAD7 were repressed in AMI patients and CFs treated with TGF-ß1, and miR-16-5p was increased. In addition, circMACF1 was resistant to RNase R and abundantly expressed in the cytoplasm. Overexpression of circMACF1 inhibited cell proliferation and migration and reduced the expression levels of fibrosis-related proteins, including Collagen I, Collagen III, and α-SMA. Furthermore, circMCAF1 could directly bind to miR-16-5p, and SMAD7 was a target gene of miR-16-5p. Knockdown of miR-16-5p suppressed the activation, proliferation, and migration of TGF-ß1-treated CFs, but silencing circMACF1 or SMAD7 partially reversed this phenomenon. CircMACF1 attenuated the TGF-ß1-induced activation, proliferation and migration of CFs via the miR-16-5p/SMAD7 signaling pathway, indicating that circMACF1 might be a new therapeutic target for AMI.

Identification and characterization of colorectal-cancer-associated SNPs on the SMAD7 locus.

PURPOSE: Genome-wide association studies have identified SMAD7 as a colorectal cancer (CRC) susceptibility gene. However, its underlying mechanism has not yet been characterized. This study screened functional SNPs (fSNPs) related to colorectal cancer through Reel-seq and obtained regulatory proteins on functional SNPs. METHODS: The candidate fSNPs on the SMAD7 locus were screened by Reel-seq method. Eight SNPs such as rs8085824 were identified as functional SNPs by luciferase reporter assay and EMSA, SDCP-MS and AIDP-WB revealed that HNRNPK can specifically bind to the rs8085824-C allele. The knockdown of HNRNPK by RNAi proved that HNRNPK could affect cell function by regulating SMAD7. RESULTS: Eight functional SNPs was found on the SMAD7 locus in linkage disequilibrium (LD) with R2 > 0.8, i.e., rs12953717, rs7227023, rs34007497, rs58920878, rs8085824, rs4991143, rs4939826, and rs7227023. We also identified allele-imbalanced binding of HNRNPK to rs8085824, H1-3 to rs12953717, THOC6 to rs7227023, and DDX21 to rs58920878. Further functional analysis revealed that these proteins are the regulatory proteins that modulate the expression of SMAD7 in the human colorectal cancer cell line DLD1. In particular, we discovered that siRNA knockdown of HNRNPK inhibits cell proliferation and cell clonal formation by downregulating SMAD7, as the decreased cell proliferation and cell clonal formation in the siRNA HNRNPK knockdown cells was restored by SMAD7 overexpression. CONCLUSION: Our findings reveal a mechanism which underlies the contribution of the fSNP rs8085824 on the SMD7 locus to CRC susceptibility.

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.

Genetic Risk Score Constructed from Polymorphisms in the VEGFA, TBX5, and SMAD7 Genes Provides Novel Insights into the Molecular Mechanisms of the Tetralogy of Fallot and Ventricular Septal Defect (Case-Control Study from the Pakistani Population).

Congenital heart defects are common and complex birth-defect malformations in developed and developing countries. It is a multifactorial disease that involves the interaction of either gene-gene or gene-environment. This comparative study was the first report on the genotypic-phenotypic correlation in the Pakistani population. The single nucleotide polymorphisms (SNPs) were further tested for association with maternal diabetes mellitus or hypertension. In addition, the cumulative genetic risk score (GRS) for low to moderately-associated SNPs was calculated for each study subject, which can ultimately guide us for better therapeutic options and prevention strategies. According to the predefined selection criteria, 376 subjects were recruited. The multiplex mini-sequencing genotyping technique opted for the cost-effective genotyping of selected loci. The association of variants with the disease was examined using logistic regression analysis. The statistical and graphical analysis was conducted using SPSS, Haploview, SNPStats, and GraphPad Prism. The results for all SNPs analysis suggested a nonsignificant association with overall congenital heart defect risk except rs3809923. However, interestingly on stratified analysis variants, rs3809923 and rs3809922 showed an association only with tetralogy of Fallot. The remaining risk factor analysis for maternal hypertension and diabetes mellitus association with SNPs were nonsignificant. The GRS was the first time constructed for this low to moderately-associated variants. Interestingly, the cumulative GRS was significantly different from the control group revealing the cumulative effect of these polymorphisms panel in patients. In conclusion, the use of GRS in the clinical setting can predict better risk association and patient outcomes.

Smad7 in the hippocampus contributes to memory impairment in aged mice after anesthesia and surgery.

BACKGROUND: Postoperative cognitive dysfunction (POCD) is a common neurological complication following anesthesia and surgery. Increasing evidence has demonstrated that neuroinflammation caused by systemic inflammatory responses during the perioperative period is a key factor in the occurrence of POCD. In addition, SMAD family member 7 (Smad7) has been confirmed to play vital roles in the pathogenesis and treatment of inflammatory diseases, such as inflammatory bowel disease. However, whether Smad7 participates in the regulatory process of neuroinflammation and apoptosis in the development of POCD is still unknown. METHODS: In this study, a POCD mouse model was constructed by unilateral nephrectomy under anesthesia, and cognitive function was assessed using the fear conditioning test and open field test. The expression of Smad7 at the mRNA and protein levels in the hippocampus 3 days after surgery was examined by qRT-PCR, western blot and immunofluorescence assays. Furthermore, to identify whether the elevation of Smad7 in the hippocampus after unilateral nephrectomy contributes to cognitive impairment, the expression of Smad7 in the hippocampal CA1 region was downregulated by crossing Smad7fl/fl conditional mutant mice and CaMKIIα-Cre line T29-1 transgenic mice or stereotaxic injection of shRNA-Smad7. Inflammation and apoptosis in the hippocampus were assessed by measuring the mRNA levels of typical inflammatory cytokines, including TNF-α, IL-1ß, IL-6, CCL2, CXCL1, and CXCL2, and the protein levels of apoptotic proteins, including Bax and Bcl2. In addition, apoptosis in the hippocampus postoperation was investigated by a terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining assay. Finally, western blotting was used to explore how Smad7 mediates inflammation and apoptosis postoperation. RESULTS: The results unequivocally revealed that elevated Smad7 in the hippocampal CA1 region significantly inhibited TGF-ß signal transduction by blocking Smad2/3 phosphorylation, which enhanced neuroinflammation and apoptosis in the hippocampus and further led to learning and memory impairment after surgery. CONCLUSIONS: Our results revealed that Smad7 contributes to cognitive impairment after surgery by enhancing neuroinflammation and apoptosis in the hippocampus and might serve as a promising therapeutic target for the treatment of memory impairment after anesthesia surgery.

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.

YES-associated protein-regulated Smad7 worsen epithelial barrier injury of chronic sinusitis with nasal polyps.

BACKGROUND: Chronic rhinosinusitis with nasal polyps (CRSwNP) was potentially due to the epithelial barrier injury. YES-associated protein (YAP) is a multifunctional transcriptional factor and plays versatile roles in the regulation and maintenance of epithelial barrier in different organs and tissues. The purpose of this study is to elucidate possible effect and mechanism of YAP on the epithelial barrier of CRSwNP. METHODS: Patients were divided into CRSwNP group (n = 12) and control group (n = 9). The location of YAP, PDZ-binding transcriptional co-activator (TAZ), and Smad7 were estimated by immunohistochemistry and immunofluorescence. Meanwhile, the expression of YAP, TAZ, Zona occludens-1 (ZO-1), E-cadherin, and transforming growth factor-beta1 (TGF-ß1) were detected by Western blot. After primary human nasal epithelial cells were treated with YAP inhibitor, the expression level of YAP, TAZ, ZO-1, E-cadherin, TGF-ß1, and Smad7 were measured by Western blot. RESULTS: Compared with the control group, the protein levels of YAP, TAZ, and Smad7 were significantly upregulated, while TGF-ß1, ZO-1, and E-cadherin were downregulated in CRSwNP. YAP and Smad7 demonstrated lower levels, while the expression of ZO-1, E-cadherin, and TGF-ß1 rose slightly after YAP inhibitor treatment in primary nasal epithelial cells. CONCLUSIONS: Higher level of YAP may lead to CRSwNP epithelial barrier injury via the TGF-ß1 signaling pathway, and the inhibition of YAP can partially reverse epithelial barrier function.

IL-22RA2 Is a SMAD7 Target Mediating the Alleviation of Dermatitis and Psoriatic Phenotypes in Mice.

Long-term management of inflammatory skin diseases is challenging because of side effects from repeated use of systemic treatments or topical corticosteroids. This study sought to identify the mechanisms and developmental therapeutics for these diseases using genetic models and pharmacological approaches. We found that mice overexpressing SMAD7 in keratinocytes but not mice overexpressing the N-terminal domain of SMAD7 (i.e., N-SMAD7) were resistant to imiquimod-induced T helper 1/17- and T helper 2-type inflammation. We generated a Tat-PYC-SMAD7 (truncated SMAD7 protein encompassing C-terminal SMAD7 and PY motif fused with cell-penetrating Tat peptide). Topically applied Tat-PYC-SMAD7 to inflamed skin entered cells upon contact and attenuated imiquimod-, 2,4-dinitrofluorobenzene-, and tape-stripping-induced inflammation. RNA-sequencing analyses of mouse skin exposed to these insults showed that in addition to inhibiting TGFß/NF-κB, SMAD7 blunted IL-22/signal transducer and activator of transcription 3 activation and associated pathogenesis, which is due to SMAD7 transcriptionally upregulating IL-22 antagonist IL-22RA2. Mechanistically, SMAD7 facilitated nuclear translocation and DNA binding of C/EBPß to IL22RA2 promoter for IL22RA2 transactivation. Consistent with the observations in mice mentioned earlier, transcript levels of IL22RA2 were increased in human atopic dermatitis and psoriasis lesions with clinical remission. Our study identified the anti-inflammation functional domain of SMAD7 and suggests the mechanism and feasibility for developing SMAD7-based biologics as a topical therapy for skin inflammatory disorders.

miR-181c targets Parkin and SMAD7 in human cardiac fibroblasts: Validation of differential microRNA expression in patients with diabetes and heart failure with preserved ejection fraction.

BACKGROUND: Cardiac fibrosis represents a key element in the pathophysiology of heart failure with preserved ejection fraction (HFpEF), a condition highly prevalent amongst geriatric patients, especially if diabetic. The microRNA 181c (miR-181c) has been shown to be associated with the response to exercise training in HFpEF patients and has been also linked to diabetic cardiovascular complications. However, the underlying mechanisms have not been fully elucidated. OBJECTIVE: To measure circulating miR-181c in elderly patients with HFpEF and diabetes mellitus (DM) and identify gene targets pathophysiologically relevant in HFpEF. METHODS: We quantified circulating miR-181c in frail older adults with a confirmed diagnosis of HFpEF and DM, and, as control, we enrolled age-matched subjects without HFpEF and without DM. We validated in human cardiac fibroblasts the molecular mechanisms linking miR-181c to a pro-fibrotic response. RESULTS: 51 frail patients were included :34 patients with DM and HFpEF and 17 age-matched controls. We observed that miR-181c was significantly upregulated (p < 0.0001) in HFpEF patients vs controls. We confirmed in vitro that miR-181c is targeting PRKN and SMAD7. CONCLUSIONS: We demonstrate that miR-181c levels are significantly increased in frail elderly adults with DM and HFpEF and that miR-181c targets PRKN and SMAD7 in human cardiac fibroblasts.

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.

Genetic analysis and allele-specific expression of SMAD7 3'UTR variants in human colorectal cancer reveal a novel somatic variant exhibiting allelic imbalance.

BACKGROUND: Considering the impact of SMAD7 deregulation in colorectal cancer (CRC) progression and the significance of single nucleotide variant (SNV)-mediated disruptions of microRNA (miRNA)-dependent regulation for cancer susceptibility, our study aimed to analyze genetic variation in the SMAD7 3' untranslated region ( 3'UTR) in CRC, measure differences in allelic mRNA expression, and evaluate its interference with miRNA-mediated post-transcriptional regulation. PATIENTS AND METHODS: This study included 80 patients with different CRC stages and six human colon cancer cell lines of various histological origins. SMAD7 3'UTR was analyzed by direct sequencing, followed by the relative quantification of differential allelic expression of detected variants by allele-specific qRT-PCR. In silico tools were employed for predictions of regulatory consequences of detected variants. RESULTS: A total of four different SNVs in one cell line and nine patients were found, among which were a novel somatic point variant and three already known germline variants (rs16950113, rs1050799536, and rs1043778717). All evaluated SNVs exhibited variable extents of allelic imbalance in expression. In silico analysis predicted significant effects of SNVs on miRNA binding efficiency, with each SNV disrupting existing and creating new target sites for one or more miRNAs. CONCLUSION: Imbalance observed in the expression of SNV alleles altering miRNA binding suggests that all investigated SNVs are potential contributing factors impacting SMAD7 expression regulation in CRC that further studies should investigate.

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.

Association of several loci of SMAD7 with colorectal cancer: A meta-analysis based on case-control studies.

BACKGROUND: Sma-and mad-related protein 7 (SMAD7) can affect tumor progression by closing transforming growth factor-beta intracellular signaling channels. Despite the extensive research on the correlation between SMAD7 polymorphisms and colorectal cancer (CRC), the conclusions of studies are still contradictory. We conducted a study focusing on the association of SMAD7 polymorphisms rs4939827, rs4464148, and rs12953717 with CRC. METHODS: We searched through 5 databases for articles and used odd ratios (ORs) and 95% confidence intervals (CIs) to discuss the correlation of SMAD7 polymorphisms with CRC risk. The heterogeneity will be appraised by subgroup analysis and meta-regression. Contour-enhanced funnel plot, Begg test and Egger test were utilized to estimate publication bias, and the sensitivity analysis illustrates the reliability of the outcomes. We performed False-positive report probability and trial sequential analysis methods to verify results. We also used public databases for bioinformatics analysis. RESULTS: We conclusively included 34 studies totaling 173251 subjects in this study. The minor allele (C) of rs4939827 is a protective factor of CRC (dominant, OR/[95% CI] = 0.89/[0.83-0.97]; recessive, OR/[95% CI] = 0.89/[0.83-0.96]; homozygous, OR/[95% CI] = 0.84/[0.76-0.93]; heterozygous, OR/[95% CI] = 0.91/[0.85-0.97]; additive, OR/[95% CI] = 0.91/[0.87-0.96]). the T allele of rs12953717 (recessive, OR/[95% CI] = 1.22/[1.15-1.28]; homozygous, OR/[95% CI] = 1.25/[1.13-1.38]; additive, OR/[95% CI] = 1.11/[1.05-1.17]) and the C allele of rs4464148 (heterozygous, OR/[95% CI] = 1.13/[1.04-1.24]) can enhance the risk of CRC. CONCLUSION: Rs4939827 (T > C) can decrease the susceptibility to CRC. However, the rs4464148 (T > C) and rs12953717 (C > T) variants were connected with an enhanced risk of CRC.

Deletion of Smad7 Ameliorates Intestinal Inflammation and Contributes to Fibrosis.

BACKGROUND: Patients suffering from inflammatory bowel diseases (IBDs) express increased mucosal levels of transforming growth factor (TGF)-ß compared with non-IBD controls. SMAD7 negatively regulates TGF-ß signaling. An earlier study aiming to target Smad7 showed a lack of clinical benefit. It remains unknown whether inhibition of SMAD7 is beneficial in specific settings of IBD. We evaluated the effect of Smad7 deficiency on inflammation, fibrogenesis, and wound healing. METHODS: For the initiation of fibrosis in Smad7-/- (Smad7Δex-I) CD-1 mice, the dextran sodium sulfate-induced chronic colitis model and the heterotopic transplantation model of fibrosis were used. Wound closure of fibroblasts from Smad7-/- mice was determined using culture inserts and electric cell-substrate impedance sensing in vitro. RESULTS: In dextran sodium sulfate-induced chronic colitis, Smad7 deficiency was associated with ameliorated inflammation, as evidenced by decreased clinical score, histological score, and myeloperoxidase activity. Absence of SMAD7 decreased T-cell accumulation in colonic tissue and tumor necrosis factor (TNF) mRNA expression levels. Smad7-/- mice showed a significant increase in hydroxyproline and collagen content, as well as ColIVa1 mRNA expression. Wild type mice transplanted with terminal ileum from Smad7-/- mice in the heterotopic animal model for intestinal fibrosis showed a significant increase in collagen content and protein expression of α-smooth muscle actin. CONCLUSIONS: Smad7 deficiency is associated with a decrease in intestinal inflammation and an increase in fibrosis. Targeting SMAD7 constitutes a potential new treatment option for IBD; progression of disease-associated fibrosis should be considered.

We evaluated the effect of Smad7 deficiency on inflammation and fibrogenesis. Smad7 deficiency was associated with ameliorated inflammation and increased collagen deposition. When targeting Smad7 as therapeutic strategy in IBD, potential initiation or aggravation of fibrosis should be considered.