Receptor mimicking TGF-β1 binding peptide for targeting TGF-β1 signaling.
Ontology highlight
ABSTRACT:
SUBMITTER: Belair DG
PROVIDER: S-EPMC9254699 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Background:TGF-? isoforms play crucial roles in diverse cellular processes. Therefore, targeting and inhibiting TGF-? signaling pathway provides a potential therapeutic opportunity. TGF-? isoforms bind and bring the receptors (T?RII and T?RI) together to form a signaling complex in an ordered manner. Objectives:Herein, an antagonistic variant of TGF-? (AnT?) has been designed and prepared to inhibit the formation of signaling complex and consequently its signaling pathway. This TGF-? homodimeric variant contains intact T?RII binding sites and blocked T?RI binding sites by substituting three peptide segments. So, AnT? could only bind to T?RII, but prevent binding and recruitment of T?RI to form a signaling complex. Materials and Methods:A reliable model of AnT? was built and re?ned using molecular dynamics (MD) simulation, followed by investigating the interactions of AnT? with the receptors using in silico docking studies. After expression of disulfide-linked AnT? in a SHuffle strain and purification of the protein using affinity chromatography, its biological activity was evaluated using Mink lung epithelial cells (Mvl Lu). Results:No meaningful significant changes in AnT? structure were observed when compared with the native protein. Based on the docking analysis, AnT? binds to T?RII similar to TGF-? and its binding to T?RI was diminished considerably which was consistent with our design purpose. Cell-based bioassay indicated that AnT? could modulate TGF-?-induced cell growth inhibition. Conclusions:Our analysis suggests that the antagonistic potency of AnT? can be used as an anti-TGF? signaling factor in the future perspectives.
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Project description:Hydroxysafflor yellow A (HSYA) is an active ingredient of Carthamus tinctorius L.. This study aimed to evaluate the effects of HSYA on transforming growth factor-β1 (TGF-β1)-induced changes in proliferation, migration, differentiation, and extracellular matrix accumulation and degradation in human fetal lung fibroblasts (MRC-5), to explore the mechanisms whereby HSYA may alleviate pulmonary fibrosis. MRC-5 cells were incubated with various doses of HSYA and/or the TGF-β receptor type I kinase inhibitor SB431542 and then stimulated with TGF-β1. Cell proliferation was measured by 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfo-phenyl)-2H-tetrazolium inner salt assay. Cell migration was detected by wound-healing assay. Protein levels of α-smooth muscle actin (α-SMA), collagen I α 1 (COL1A1), and fibronectin (FN) were measured by immunofluorescence. Protein levels of matrix metalloproteinase-2, tissue inhibitor of matrix metalloproteinase-1, tissue inhibitor of matrix metalloproteinase-2, TGF-β type II receptor (TβRII), and TGF-β type I receptor were detected by western blotting. TβRII knockdown with siRNA interfered with the inhibitory effect of HSYA on α-SMA, COL1A1, and FN expression, and TGF-β1-induced Sma and Mad protein (Smad), and extracellular signal-regulated kinase/mitogen-activated protein kinase signaling pathway activation. The antagonistic effect of HSYA on the binding of fluorescein isothiocyanate-TGF-β1 to MRC-5 cell cytoplasmic receptors was measured by flow cytometry. HSYA significantly suppressed TGF-β1-induced cell proliferation and migration. HSYA could antagonize the binding of FITC-TGF-β1 to MRC-5 cell cytoplasmic receptors. Also HSYA inhibited TGF-β1-activated cell expression of α-SMA, COL1A1, and FN and phosphorylation level of Smad2, Smad3, and ERK by targeting TβRII in MRC-5 cells. These findings suggest that TβRII might be the target responsible for the inhibitory effects of HSYA on TGF-β1-induced pathological changes in pulmonary fibrosis.
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Project description:BackgroundHoneysuckle is a time-honored herb with anticancer activity in traditional Chinese medicine. Recently, accumulating reports are suggesting that the microRNAs in this medicinal plant not only play a physiological role in their original system, but also can be transmitted to another species as potential therapeutic components. In the numerous bioactive investigations, the anti-tumor effects of these microRNAs in the magical herb are rarely studied, especially the special miR2911, a honeysuckle-encoded atypical microRNA, with high stability during the boiling process and unique biological activity to target TGF-β1 mRNA.MethodsLuciferase assay was conducted to test the ability of miR2911 to target TGF-β1 mRNA. ELISA was performed to determine the expression level of TGF-β1 of mouse colorectal adenocarcinoma CT26 cells when treated with miR2911 and tumor tissue in Sidt1+/+ and Sidt1-/- mice. qRT-PCR was performed to examine the level of expression of miR2911. Tumor-bearing wild and nude mice were employed to evaluate the anti-tumor effect of honeysuckle and miR2911 in vivo. Tumor tissue necrosis was observed by H&E staining. Besides, the infiltration of T lymphocytes across solid tumors was tested by immunostaining staining.ResultsOur results showed that honeysuckle slowed the development of colon cancer down. Further research showed that miR2911 could bind strongly to TGF-β1 mRNA and down-regulate the expression of TGF-β1 and had a high stability under boiling and acid condition. Moreover, SIDT1 mediated dietary miR2911 inter-species absorption. And we found that miR2911 had a similar anticancer effect as honeysuckle. Mechanistically, miR2911 reversed the tumor-promoting effect of TGF-β1 by an increase of T lymphocytes infiltration, resulting in slowing the colon cancer process in immunocompetent mice. Consistent with this inference, the anti-tumor effect of miR2911 was revealed to be abolished in T cell immune deficiency mice.ConclusionTaken together, honeysuckle-derived miR2911 showed an anti-tumor effect in colon cancer through targeting TGF-β1 mRNA. The down-regulation of TGF-β1 promoted T lymphocytes infiltration, and accordingly impeded the colon tumor development.
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Project description:Loss of Klotho, an anti-aging protein, plays a critical role in the pathogenesis of chronic kidney diseases. As Klotho is a large transmembrane protein, it is challenging to harness it as a therapeutic remedy. Here we report the discovery of a Klotho-derived peptide 1 (KP1) protecting kidneys by targeting TGF-β signaling. By screening a series of peptides derived from human Klotho protein, we identified KP1 that repressed fibroblast activation by binding to TGF-β receptor 2 (TβR2) and disrupting the TGF-β/TβR2 engagement. As such, KP1 blocked TGF-β-induced activation of Smad2/3 and mitogen-activated protein kinases. In mouse models of renal fibrosis, intravenous injection of KP1 resulted in its preferential accumulation in injured kidneys. KP1 preserved kidney function, repressed TGF-β signaling, ameliorated renal fibrosis and restored endogenous Klotho expression. Together, our findings suggest that KP1 recapitulates the anti-fibrotic action of Klotho and offers a potential remedy in the fight against fibrotic kidney diseases.
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Project description:BackgroundPeritoneal fibrosis (PF) is the main complication of peritoneal dialysis (PD) and the most common cause of cessation from PD. There is still no effective therapeutic approach to reserve PF. We aimed to investigate the role of miR-132-3p and underlying potential mechanisms in PF.MethodsA total of 18 Sprague-Dawley (SD) rats were divided randomly into three groups (n = 6): (i)Control group (ii)PF group (iii)PF+Losartan group; Rats in the PF group and PF+Losartan group received daily intraperitoneal injections of 3 mg/kg chlorhexidine for 14 days, and rats in the PF+Losartan group simultaneously received daily intraperitoneal injections of 2 mg/kg losartan for 14 days. The control group was injected with saline in the same volume. Met-5A cells were treated for 24h with TGF-β1 dissolved in recombinant buffered saline at a concentration of 10 ng/ml, meanwhile, PBS solution as a negative control. The human peritoneal solution was collected for the detection of miR-132-3p.ResultsIn vivo, SD rats were infused with chlorhexidine to establish PF model, and we found that miR-132-3p significantly decreased and the expressions of transforming growth factor-β1 (TGF-β1), and Smad2/3 were up-regulated in PF. In vitro, miR-132-3p mimics suppressed TGF-β1/Smad2/3 activity, whereas miR-132-3p inhibition activated the pathway. In human peritoneal solution, we found that the expression of miR-132-3p decreased in a time-dependent model and its effect became more pronounced with longer PD duration.ConclusionMiR-132-3p ameliorated PF by suppressing TGF-β1/Smad2/3 activity, suggesting that miR-132-3p represented a potential therapeutic approach for PF.
| S-EPMC11008817 | biostudies-literature