Objective To explore the protective effect and related mechanism of miR-140-5p on liver fibrosis by interfering with TGF-β/Smad signaling pathway.Methods Liver fibrosis mice models were established by intraperitoneal injection of CCL4. Hematoxylin and eosin (HE) staining was used to detect the structural and morphological changes of the liver. Masson staining was used to detect collagen deposition. Human hepatic stellate cells (HSCs, LX-2) were transfected with miR-140-5p mimic or inhibitor then treated with TGF-β1. The qRT-PCR and Western blotting was used to detect the expression of related molecules. The luciferase reporter assay was used to identify the target of miR-140-5p.Results Our results indicated that miR-140-5p expression was downregulated in fibrotic liver tissues of model mice and LX-2 cells treated with TGF-β1. The overexpression of miR-140-5p decreased the expression of collagen1(COL1) and α-smooth muscle actin(α-SMA), inhibited the phosphorylation of Smad-2/3 (pSmad-2/3) in LX-2 cells. Conversely, the knockdown of miR-140-5p upregulated COL1 and α-SMA expression, increased Smad-2/3 phosphorylation. A dual-luciferase reporter assay showed that TGFβR1 was a target gene of miR-140-5p. The overexpression of miR-140-5p suppressed TGFβR1 expression in LX-2 cells. Additionally, knockdown of TGFβR1 decreased the expression of COL1 and α-SMA. Conversely, the overexpression of TGFβR1 reversed the inhibitory effect of miR-140-5p upregulation on expression of COL1 and α-SMA.Conclusion miR-140-5p bound to TGFβR1 mRNA 3′-untranslated region(3′UTR) and inhibited the expression of TGFβR1, pSmad-2/3, COL1 and α-SMA, thereby exerting a potential therapeutic effect on hepatic fibrosis.
Characterized by high malignancy and limited treatment efficacy, triple-negative breast cancer (TNBC) remains a clinically challenging subtype within breast cancer classifications, marked by rapid progression and high mortality. Abnormal activation of the transforming growth factor-β (TGFβ) pathway signaling, a pathway integral to tumor progression, metastasis, angiogenesis and immune evasion, is a common feature in a broad spectrum of malignancies. Owing to the restricted effectiveness of first-line interventions including surgical resection, cytotoxic agents, and radiation therapy for TNBC, novel agents that modulate TGFβ activity represent a compelling therapeutic avenue. Herein, we reported the identification and preclinical evaluation of YH395A, a novel tetrahydro‐β‐carboline derivative derived from the lead compound YR‐290 with virtual screening from pseudo molecular library generated by generative deep learning method. In vitro studies demonstrated that YH395A dose‐dependently inhibited TNBC cell migration and invasion. In vivo, administration of YH395A not only curtailed metastatic dissemination and prevented the extravasation of breast cancer cells into lung parenchyma in mouse models but also significantly reduced tumor growth in a patient‐derived xenograft (PDX) model. Mechanistic analyses indicated that these antitumor effects are mediated via potent inhibition of TGFβ signaling. These cumulative results demonstrate YH395A’s viability as a novel therapeutic agent for TNBC, while emphasizing the necessity for expanded preclinical validation studies.
Luna Zhang,Qianqian Li,Yufang Ni,Zhaoyang Li,Honglian Wang,Lifeng Meng,Li Wang,Jianchun Li
Background Renal fibrosis represents a common and critical pathological process underlying chronic kidney disease (CKD), invariably leading to progressive renal dysfunction and ultimately end-stage renal disease. Purpose This investigation sought to determine anti-fibrotic efficacy of Deacetyl Asperulosidic Acid Methyl Ester (DA), in ameliorating renal fibrosis, focusing on its interaction with heat shock protein 90 (Hsp90aa1) and its downstream effects on TGF-β/Smad signaling. Study design The anti-fibrotic efficacy of DA was evaluated in vivo using two murine models of CKD (unilateral ureteral obstruction and adenine-induced nephropathy) and in vitro using TGF-β1-stimulated NRK-49F fibroblasts and TCMK-1 tubular cells. Molecular targets were identified and validated through quantitative proteomics, Specific Pupylation as Identity Reporter (SPIDER) assay, and biophysical interaction analyses. Methods Renal function and histological changes were assessed by biochemical analysis, H&E, and Masson's trichrome staining. The expression of fibrotic markers and signaling pathway proteins was determined using IHC, Western blot, and qRT-PCR. The interaction between DA and Hsp90aa1 was verified by Cellular Thermal Shift Assay (CETSA), Drug Affinity Responsive Target Stability (DARTS), and Co-immunoprecipitation (Co-IP). Results Our data demonstrated that DA administration substantially ameliorated renal dysfunction and attenuated fibrotic progression in both animal and cellular models. Quantitative proteomic profiling results revealed a prominent suppression of extracellular matrix components and TGF-β type I receptor (TGFβRI) following DA treatment. Furthermore, biotin-labeled DA binding combined with SPIDER experiments identified Hsp90aa1 as a direct target of DA. Mechanistically, the binding of DA to Hsp90aa1 disrupts the interaction between Hsp90aa1 and TGFβR1, thereby destabilizing TGFβR1 and promoting its degradation. This regulation effectively impedes the transduction of the downstream TGF-β/Smad signaling cascade, ultimately mitigating renal fibrosis. Conclusion Our findings demonstrate that DA is a potent anti-fibrotic agent that targets Hsp90aa1 to modulate TGFBR1 stability and inhibit the TGF-β/Smad pathway. Download: Download high-res image (359KB) Download: Download full-size image
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