Explore the vast range of titles available.

Acidosis, a common characteristic of the tumor microenvironment, is associated with alterations in metabolic preferences of cancer cells and progression of the disease. Here we identify the TGF-β2 isoform at the interface between these observations. We document that acidic pH promotes autocrine TGF-β2 signaling, which in turn favors the formation of lipid droplets (LD) that represent energy stores readily available to support anoikis resistance and cancer cell invasiveness. We find that, in cancer cells of various origins, acidosis-induced TGF-β2 activation promotes both partial epithelial-to-mesenchymal transition (EMT) and fatty acid metabolism, the latter supporting Smad2 acetylation. We show that upon TGF-β2 stimulation, PKC-zeta-mediated translocation of CD36 facilitates the uptake of fatty acids that are either stored as triglycerides in LD through DGAT1 or oxidized to generate ATP to fulfill immediate cellular needs. We also address how, by preventing fatty acid mobilization from LD, distant metastatic spreading may be inhibited. The tumour microenvironment is known to have an acidic pH but how this influences cancer cell phenotype is unclear. Here, the authors show that tumour cells upregulate TGF-β2 under acidosis, which leads to the increased formation of lipid droplets allowing for invasiveness and metastases.

Fibrosis is a chronic process involving development and progression of multiple diseases in various organs and is responsible for almost half of all known deaths. Epithelial-mesenchymal transition (EMT) is the vital process in organ fibrosis. Lens is an elegant biological tool to investigate the fibrosis process because of its unique biological properties. Using gain- and loss-of-function assays, and different lens fibrosis models, here we demonstrated that microRNA (miR)-26a and miR-26b, members of the miR-26 family have key roles in EMT and fibrosis. They can significantly inhibit proliferation, migration, EMT of lens epithelial cells and lens fibrosis in vitro and in vivo. Interestingly, we revealed that the mechanisms of anti-EMT effects of miR-26a and -26b are via directly targeting Jagged-1 and suppressing Jagged-1/Notch signaling. Furthermore, we provided in vitro and in vivo evidence that Jagged-1/Notch signaling is activated in TGFβ2-stimulated EMT, and blockade of Notch signaling can reverse lens epithelial cells (LECs) EMT and lens fibrosis. Given the general involvement of EMT in most fibrotic diseases, cancer metastasis and recurrence, miR-26 family and Notch pathway may have therapeutic uses in treating fibrotic diseases and cancers.

Background Posterior capsular opacification (PCO) is the main reason affecting the long-term postoperative result of cataract patient, and it is well accepted that fibrotic PCO is driven by transforming growth factor beta (TGFβ) signaling. Ferroptosis, closely related to various ocular diseases, but has not been explored in PCO. Methods RNA sequencing (RNA-seq) was performed on both TGF-β2 treated and untreated primary lens epithelial cells (pLECs). Differentially expressed genes (DEGs) associated with ferroptosis were analyzed using Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) to investigate their biological function. Additionally, protein-to-protein interactions among selected ferroptosis-related genes by PPI network and the top 10 genes with the highest score (MCC algorithm) were selected as the hub genes. The top 20 genes with significant fold change values were validated using quantitative real-time polymerase chain reaction (qRT-PCR). Results Our analysis revealed 1253 DEGs between TGF-β2 treated and untreated pLECs, uncovering 38 ferroptosis-related genes between two groups. Among these 38 ferroptosis-related genes,the most prominent GO enrichment analysis process involved in the response to oxidative stress (BPs), apical part of cell (CCs),antioxidant activity (MFs). KEGG were mainly concentrated in fluid shear stress and atherosclerosis, IL-17 and TNF signaling pathways, and validation of top 20 genes with significant fold change value were consistent with RNA-seq. Conclusions Our RNA-Seq data identified 38 ferroptosis-related genes in TGF-β2 treated and untreated pLECs, which is the first observation of ferroptosis related genes in primary human lens epithelial cells under TGF-β2 stimulation. Graphical Abstract Our study initially observed ferroptosis related genes in primary human lens epithelial cells stimulated by TGF-β2. These findings may improve the understanding of the molecular mechanisms of PCO and provide a new direction for exploring the potential mechanisms of PCO

Fuchs endothelial corneal dystrophy (FECD) is a progressive corneal disease characterized by corneal endothelial cell (CEC) loss and guttae formation. Elevated levels of Transforming Growth Factor-Beta 1 and 2 (TGF-β1/-β2) have been reported in the aqueous humor (AH) of FECD patients and have been implicated with abnormal extracellular matrix (ECM) production, endothelial-to-mesenchymal transition (EndoMT), the unfolded protein response, and cell death. However, how TGF-β signaling affects cell migration in FECD remains to be elucidated. In this study, we found that TGF-β2 levels were significantly elevated in the AH of FECD patients compared to controls. We performed bulk RNA sequencing on FECD CECs treated with TGF-β1 or TGF-β2 and identified the epithelial-to-mesenchymal (EMT) pathway as one of the top dysregulated pathways. We found that TGF-β1 and TGF-β2 increased EMT markers, filamentous-actin (F-actin) expression and produced more EMT-like phenotype in FECD and control CECs. We also observed that TGF-β1 and TGF-β2 significantly increased FECD CEC migration speed as detected by scratch assay and individual cell tracking and promoted individual cellular migration behavior. This study provides novel insight into FECD pathogenesis and how increased TGF-β signaling promotes EndoMT and alters cellular migration in FECD CECs.

To establish appropriate ex vivo models for a glaucomatous trabecular meshwork (TM), two-dimensional (2D) and three-dimensional (3D) cultures of human trabecular meshwork cells (HTM) were prepared in the presence of 250 nM dexamethasone (DEX) or 5 ng/mL TGFβ2, and characterized by the following analyses; transendothelial electrical resistance (TEER) measurements, FITC dextran permeability, scanning electron microscopy and the expression of the extracellular matrix (ECM) including collagen (COL)1, 4 and 6, and fibronectin (FN), α-smooth muscle actin (α-SMA), tissue inhibitor of metalloproteinase (TIMP)1–4, and matrix metalloproteinase (MMP)2, 9 and 14. DEX and TGFβ2 both caused a significant increase or decrease in the TEER values and FITC dextran permeability. During the 3D spheroid culture, DEX or TGFβ2 induced a mild and significant down-sizing and an increase in stiffness, respectively. TGFβ2 induced a significant up-regulation of COL1 and 4, FN, α-SMA, and MMP 2 and 14 (2D) or COL1 and 6, and TIMP2 and 3 (3D), and DEX induced a significant up-regulation of FN (3D) and TIMP4 (2D and 3D). The findings presented herein indicate that DEX or TGFβ2 resulted in mild and severe down-sized and stiff 3D HTM spheroids, respectively, thus making them viable in vitro HTM models for steroid-induced and primary open angle glaucoma.

The hypoxia associated with the transforming growth factor-β2 (TGF-β2)-induced epithelial mesenchymal transition (EMT) of human retinal pigment epithelium (HRPE) cells is well recognized as the essential underlying mechanism responsible for the development of proliferative retinal diseases. In vitro, three-dimensional (3D) models associated with spontaneous O2 gradients can be used to recapitulate the pathological levels of hypoxia to study the effect of hypoxia on the TGF-β2-induced EMT of HRPE cells in detail, we used two-dimensional-(2D) and 3D-cultured HRPE cells. TGF-β2 and hypoxia significantly and synergistically increased the barrier function of the 2D HRPE monolayers, as evidenced by TEER measurements, the downsizing and stiffening of the 3D HRPE spheroids and the mRNA expression of most of the ECM proteins. A real-time metabolic analysis indicated that TGF-β2 caused a decrease in the maximal capacity of mitochondrial oxidative phosphorylation in the 2D HRPE cells, whereas, in the case of 3D HRPE spheroids, TGF-β2 increased proton leakage. The findings reported herein indicate that the TGF-β2-induced EMT of both the 2D and 3D cultured HRPE cells were greatly modified by hypoxia, but during these EMT processes, the metabolic plasticity was different between 2D and 3D HRPE cells, suggesting that the mechanisms responsible for the EMT of the HRPE cells may be variable during their spatial spreading.

In the normal eye, most of the aqueous humor drains through the trabecular meshwork (TM) and Schlemm’s canal (SC). The concentration of transforming growth factor beta 2 (TGF-β2) is increased in the aqueous humor of primary open angle glaucoma patients. TGF-β2 increases outflow resistance by affecting the TM and SC, and endothelial–mesenchymal transition (EndMT) of SC cells is involved in these changes. Here, we investigated the effect of a ROCK inhibitor on TGF-β2-induced EndMT in SC cells. The ROCK inhibitor Y-27632 suppressed the TGF-β2-induced increase in the trans-endothelial electrical resistance (TER) and proliferation of SC cells. Y-27632 suppressed the expression of α-SMA, N-cadherin, and Snail, which are upregulated by TGF-β2. Moreover, TGF-β2 decreased mRNA levels of bone morphogenetic protein (BMP) 4 and increased those of the BMP antagonist gremlin (GREM1), but Y-27632 significantly suppressed these changes. Y-27632 also inhibited TGF-β2-induced phosphorylation of p-38 mitogen-activated protein kinase (MAPK). BMP4 and the p-38 MAPK inhibitor SB203580 suppressed the TGF-β2-induced TER elevation in SC cells. Moreover, SB203580 suppressed TGF-β2-induced upregulation of fibronectin, Snail, and GREM1. These results indicate that a ROCK inhibitor inhibited the TGF-β2-induced EndMT in SC cells, implying the involvement of p38 MAPK and BMP4 signaling.

This study aimed to investigate the tyrosine kinase inhibitor Nintedanib and its potential role in reversing epithelial–mesenchymal transition (EMT) induced by transforming growth factor beta 2 (TGF-β2) in retinal pigment epithelial (RPE) cells, along with its therapeutic potential using a mouse model of subretinal fibrosis. We hypothesized that the blockade of angiogenesis promoting and fibrosis inducing signaling using the receptor tyrosine kinase inhibitor Nintedanib (OfevTM) can prevent or reverse EMT both in vitro and in our in vivo model of subretinal fibrosis. Primary human retinal pigment epithelial cells (phRPE) and adult retinal pigment epithelial cell line (ARPE-19) cells were treated with TGF-β210 ng/mL for two days followed by four days of Nintedanib (1 µM) incubation. Epithelial and mesenchymal phenotypes were assessed by morphological examination, quantitative real-time polymerase chain reaction(qPCR) (ZO-1, Acta2, FN, and Vim), and immunocytochemistry (ZO-1, vimentin, fibronectin, and αSMA). Metabolites were measured using luciferase-based assays. Extracellular acidification and oxygen consumption rates were measured using the Seahorse XF system. Metabolic-related genes (GLUT1, HK2, PFKFB3, CS, LDHA, LDHB) were evaluated by qPCR. A model of subretinal fibrosis using the two-stage laser-induced method in C57BL/6J mice assessed Nintedanib’s therapeutic potential. Fibro-vascular lesions were examined 10 days later via fluorescence angiography and immunohistochemistry. Both primary and ARPE-19 RPE stimulated with TGF-β2 upregulated expression of fibronectin, αSMA, and vimentin, and downregulation of ZO-1, consistent with morphological changes (i.e., elongation). Glucose consumption, lactate production, and glycolytic reserve were significantly increased in TGF-β2-treated cells, with upregulation of glycolysis-related genes (GLUT1, HK2, PFKFB3, CS). Nintedanib treatment reversed TGF-β2-induced EMT signatures, down-regulated glycolytic-related genes, and normalized glycolysis. Nintedanib intravitreal injection significantly reduced collagen-1+ fibrotic lesion size and Isolectin B4+ neovascularization and reduced vascular leakage in the two-stage laser-induced model of subretinal fibrosis. Nintedanib can induce Mesenchymal-to-Epithelial Transition (MET) in RPE cells and reduce subretinal fibrosis through metabolic reprogramming. Nintedanib can therefore potentially be repurposed to treat retinal fibrosis.

Neuroendocrine prostate cancer (NEPC) is a highly aggressive and treatment‐resistant subtype of castration‐resistant prostate cancer (CRPC) that often emerges during progression under androgen‐receptor (AR) pathway inhibition. While lineage plasticity in cancer cells has been recognized as a key mechanism of resistance, the role of the tumor microenvironment in driving this transition remains unclear. Among its cellular components, vascular endothelial cells can undergo endothelial‐mesenchymal transition (EndoMT), a phenotypic shift associated with tumor progression and fibrosis. Here, we investigated whether EndoMT contributes to NEPC development. Human umbilical vein endothelial cells (HUVEC) were induced to undergo EndoMT using IL‐1β and TGF‐β2, and are hereafter referred to as EndoMTed HUVEC. EndoMTed HUVEC promoted neuroendocrine features and functional changes in LNCaP cells. Transcriptome analysis revealed marked upregulation of granulocyte‐macrophage colony‐stimulating factor (GM‐CSF) in EndoMTed HUVEC. Neutralization of GM‐CSF signaling using mavrilimumab, a monoclonal antibody targeting the GM‐CSF receptor alpha (CSF2RA), and siRNA‐mediated CSF2RA knockdown both suppressed the neuroendocrine phenotype and STAT3 signaling of LNCaP cells. Conversely, GM‐CSF stimulation alone reproduced these changes. Enzalutamide‐treated LNCaP cells secreted IL‐1β and TGF‐β2, which in turn triggered EndoMT, suggesting a reciprocal loop. These findings indicate that anti‐androgen therapy may inadvertently promote NEPC through a paracrine loop involving tumor‐derived cytokines and endothelial GM‐CSF secretion, highlighting EndoMT as a microenvironmental driver of treatment resistance. Endothelial‐mesenchymal transition (EndoMT) contributes to neuroendocrine differentiation (NED) in prostate cancer. Using in vitro co‐culture models, we show that EndoMTed endothelial cells promote NED in prostate cancer cells via GM‐CSF signaling. These findings reveal a previously unrecognized paracrine mechanism by which the tumor microenvironment drives therapy‐induced NED.

This study aimed to investigate the effects of substratum stiffness on the sensitivity of human conjunctival fibroblasts to transforming growth factor (TGF)-β, and to explore the molecular mechanism of action. Human conjunctival fibroblasts were cultured on collagen-coated plastic or silicone plates. The stiffness of the silicone plates was 0.2 or 64 kPa. Cells were treated by 2.5 ng/mL TGF-β2 with or without fibroblast growth factor (FGF)-2 (0-100 ng/mL) for 24 h or 48 h. The protein expression levels were determined by Western blot analysis. Cell proliferation was assessed using the WST-8 assay. FGF-2 suppressed the TGF-β-induced expression of α-smooth muscle actin (SMA) and collagen type I (Col I), but not fibronectin (FN). Both FGF-2 and TGF-β2 increased cell proliferation without an additive effect. The induction of α-SMA by TGF-β2 was decreased on the soft substratum, without any change in the expression level or subcellular location of Yes-associated protein/transcriptional coactivator with PDZ-binding motif (YAP/TAZ). FGF-2 suppressed TGF-β-induced α-SMA expression even on the soft substratum. FGF-2 treatment and a soft substratum suppressed TGF-β-induced transdifferentiation of conjunctival fibroblasts into myofibroblasts. FGF-2 attenuated the TGF-β-induced expression of α-SMA, even on a soft substratum.