Mesenteric lymph nodes: a critical site for the up-regulatory effect of hUC-MSCs on Treg cells by producing TGF-ß1 in colitis treatment.

BACKGROUND: Mesenchymal stem cells (MSCs) demonstrate a wide range of therapeutic capabilities in the treatment of inflammatory bowel disease (IBD). The intraperitoneal injection of MSCs has exhibited superior therapeutic efficacy on IBD than intravenous injection. Nevertheless, the precise in vivo distribution of MSCs and their biological consequences following intraperitoneal injection remain inadequately understood. Additional studies are required to explore the correlation between MSCs distribution and their biological effects. METHODS: First, the distribution of human umbilical cord MSCs (hUC-MSCs) and the numbers of Treg and Th17 cells in mesenteric lymph nodes (MLNs) were analyzed after intraperitoneal injection of hUC-MSCs. Subsequently, the investigation focused on the levels of transforming growth factor beta1 (TGF-ß1), a key cytokine to the biology of both Treg and Th17 cells, in tissues of mice with colitis, particularly in MLNs. The study also delved into the impact of hUC-MSCs therapy on Treg cell counts in MLNs, as well as the consequence of TGFB1 knockdown hUC-MSCs on the differentiation of Treg cells and the treatment of IBD. RESULTS: The therapeutic effectiveness of intraperitoneally administered hUC-MSCs in the treatment of colitis was found to be significant, which was closely related to their quick migration to MLNs and secretion of TGF-ß1. The abundance of hUC-MSCs in MLNs of colitis mice is much higher than that in other organs even the inflamed sites of colon. Intraperitoneal injection of hUC-MSCs led to a significant increase in the number of Treg cells and a decrease in Th17 cells especially in MLNs. Furthermore, the concentration of TGF-ß1, the key cytokine for Treg differentiation, were also found to be significantly elevated in MLNs after hUC-MSCs treatment. Knockdown of TGFB1 in hUC-MSCs resulted in a noticeable reduction of Treg cells in MLNs and the eventually failure of hUC-MSCs therapy in colitis. CONCLUSIONS: MLNs may be a critical site for the regulatory effect of hUC-MSCs on Treg/Th17 cells and the therapeutic effect on colitis. TGF-ß1 derived from hUC-MSCs promotes local Treg differentiation in MLNs. This study will provide new ideas for the development of MSC-based therapeutic strategies in IBD patients.

Hsa-miR-134-5p predicts cardiovascular risk in circulating mononuclear cells and improves angiogenic action of senescent endothelial progenitor cells.

This research explores the role of microRNA in senescence of human endothelial progenitor cells (EPCs) induced by replication. Hsa-miR-134-5p was found up-regulated in senescent EPCs where overexpression improved angiogenic activity. Hsa-miR-134-5p, which targeted transforming growth factor ß-activated kinase 1-binding protein 1 (TAB1) gene, down-regulated TAB1 protein, and inhibited phosphorylation of p38 mitogen-activated protein kinase (p38) in hsa-miR-134-5p-overexpressed senescent EPCs. Treatment with siRNA specific to TAB1 (TAB1si) down-regulated TAB1 protein and subsequently inhibited p38 activation in senescent EPCs. Treatment with TAB1si and p38 inhibitor, respectively, showed angiogenic improvement. In parallel, transforming growth factor Beta 1 (TGF-ß1) was down-regulated in hsa-miR-134-5p-overexpressed senescent EPCs and addition of TGF-ß1 suppressed the angiogenic improvement. Analysis of peripheral blood mononuclear cells (PBMCs) disclosed expression levels of hsa-miR-134-5p altered in adult life, reaching a peak before 65 years, and then falling in advanced age. Calculation of the Framingham risk score showed the score inversely correlates with the hsa-miR-134-5p expression level. In summary, hsa-miR-134-5p is involved in the regulation of senescence-related change of angiogenic activity via TAB1-p38 signalling and via TGF-ß1 reduction. Hsa-miR-134-5p has a potential cellular rejuvenation effect in human senescent EPCs. Detection of human PBMC-derived hsa-miR-134-5p predicts cardiovascular risk.

Rhizoma coptidis can inhibit the excessive proliferation, inflammation, and transformation of lung fibroblasts into myofibroblasts.

BACKGROUND: Pulmonary fibrosis (PF) is a chronic, progressive, and irreversible heterogeneous disease of lung interstitial tissue. To combat progression of PF, new drugs are required to be developed. Rhizoma coptidis (COP), one of the main alkaloids of Coptis chinensis, is a traditional herbal medicine used to treat various inflammatory diseases. OBJECTIVE: To investigate the possible effects of Coptisine (Cop) on the growth, inflammation, as well as FMT of TNF-ß1-induced HFL1 cells and uncover the mechanism. MATERIAL AND METHODS: Human fetal lung fibroblast 1 (HFL1) was induced using 6ng/mL TGF-ß1 as a model of pulmonary fibrosis. CCK-8, Brdu, and transwell assays indicated the effects on cell growth as well as motility. qPCR and the corresponding kits indicted the effects on cell inflammation. Immunoblot showed the effects on FMT and further confirmed the mechanism. RESULTS: Coptisine inhibits excessive growth as well as motility of TNF-ß1-induced HFL1 cells. It further inhibits inflammation and ROS levels in TNF-ß1-induced HFL1 cells. Coptisine inhibits the FMT process of TNF-ß1-induced HFL1 cells. Mechanically, coptisine promotes the Nrf2/HO-1 pathway. CONCLUSION: Coptisine can inhibit the excessive growth, inflammation as well as FMT of lung fibroblasts into myofibroblasts. It could serve as a promising drug of PF.

FOXK2 facilitates the airway remodeling during chronic asthma by promoting glycolysis in a SIRT2-dependent manner.

Asthma is a chronic pulmonary disease with the worldwide prevalence. The structural alterations of airway walls, termed as "airway remodeling", are documented as the core contributor to the airway dysfunction during chronic asthma. Forkhead box transcription factor FOXK2 is a critical regulator of glycolysis, a metabolic reprogramming pathway linked to pulmonary fibrosis. However, the role of FOXK2 in asthma waits further explored. In this study, the chronic asthmatic mice were induced via ovalbumin (OVA) sensitization and repetitive OVA challenge. FOXK2 was upregulated in the lungs of OVA mice and downregulated after adenovirus-mediated FOXK2 silencing. The lung inflammation, peribronchial collagen deposition, and glycolysis in OVA mice were obviously attenuated after FOXK2 knockdown. Besides, the expressions of FOXK2 and SIRT2 in human bronchial epithelial cells (BEAS-2B) were increasingly upregulated upon TGF-ß1 stimulation and downregulated after FOXK2 knockdown. Moreover, the functional loss of FOXK2 remarkably suppressed TGF-ß1-induced epithelial-mesenchymal transition (EMT) and glycolysis in BEAS-2B cells, as manifested by the altered expressions of EMT markers and glycolysis enzymes. The glycolysis inhibitor 2-deoxy-d-glucose (2-DG) inhibited the EMT in TGF-ß1-induced cells, making glycolysis a driver of EMT. The binding of FOXK2 to SIRT2 was validated, and SIRT2 overexpression blocked the FOXK2 knockdown-mediated inhibition of EMT and glycolysis in TGF-ß1-treated cells, which suggests that FOXK2 regulates EMT and glycolysis in TGF-ß1-treated cells in a SIRT2-dependnet manner. Collectively, this study highlights the protective effect of FOXK2 knockdown on airway remodeling during chronic asthma.

More Velcro for the TGF-ß1 straitjacket: A new antibody straps latent TGF-ß1 to the matrix.

In this issue of Science Signaling, Jackson et al. present a new antibody strategy to-quite literally-strap transforming growth factor-ß1 (TGF-ß1) to latent complexes in the extracellular matrix. The antibody has no effect on latent TGF-ß1 presented on the surface of immune cells and thus allows targeting of the detrimental effects of TGF-ß1 in fibrosis without affecting its beneficial immune-suppressing activities.

An antibody that inhibits TGF-ß1 release from latent extracellular matrix complexes attenuates the progression of renal fibrosis.

Inhibitors of the transforming growth factor-ß (TGF-ß) pathway are potentially promising antifibrotic therapies, but nonselective simultaneous inhibition of all three TGF-ß homologs has safety liabilities. TGF-ß1 is noncovalently bound to a latency-associated peptide that is, in turn, covalently bound to different presenting molecules within large latent complexes. The latent TGF-ß-binding proteins (LTBPs) present TGF-ß1 in the extracellular matrix, and TGF-ß1 is presented on immune cells by two transmembrane proteins, glycoprotein A repetitions predominant (GARP) and leucine-rich repeat protein 33 (LRRC33). Here, we describe LTBP-49247, an antibody that selectively bound to and inhibited the activation of TGF-ß1 presented by LTBPs but did not bind to TGF-ß1 presented by GARP or LRRC33. Structural studies demonstrated that LTBP-49247 recognized an epitope on LTBP-presented TGF-ß1 that is not accessible on GARP- or LRRC33-presented TGF-ß1, explaining the antibody's selectivity for LTBP-complexed TGF-ß1. In two rodent models of kidney fibrosis of different etiologies, LTBP-49247 attenuated fibrotic progression, indicating the central role of LTBP-presented TGF-ß1 in renal fibrosis. In mice, LTBP-49247 did not have the toxic effects associated with less selective TGF-ß inhibitors. These results establish the feasibility of selectively targeting LTBP-bound TGF-ß1 as an approach for treating fibrosis.

Microvesicles from quiescent and TGF-ß1 stimulated hepatic stellate cells: Divergent impact on hepatic vascular injury.

BACKGROUND: This study evaluated the effect of microvesicles(MVs) from quiescent and TGF-ß1 stimulated hepatic stellate cells (HSC-MVs, TGF-ß1HSC-MVs) on H2O2-induced human umbilical vein endothelial cells (HUVECs) injury and CCl4-induced rat hepatic vascular injury. METHODS: HUVECs were exposed to hydrogen peroxide (H2O2) to establish a model for vascular endothelial cell injury. HSC-MVs or TGF-ß1HSC-MVs were co-cultured with H2O2-treated HUVECs, respectively. Indicators including cell survival rate, apoptosis rate, oxidative stress, migration, invasion, and angiogenesis were measured. Simultaneously, the expression of proteins such as PI3K, AKT, MEK1+MEK2, ERK1+ERK2, VEGF, eNOS, and CXCR4 was assessed, along with activated caspase-3. SD rats were intraperitoneally injected with CCl4 twice a week for 10 weeks to induce liver injury models. HSC-MVs or TGF-ß1HSC-MVs were injected into the tail vein of rats. Liver and hepatic vascular damage were also detected. RESULTS: In H2O2-treated HUVECs, HSC-MVs increased cell viability, reduced cytotoxicity and apoptosis, improved oxidative stress, migration, and angiogenesis, and upregulated protein expression of PI3K, AKT, MEK1/2, ERK1/2, VEGF, eNOS, and CXCR4. Conversely, TGF-ß1HSC-MVs exhibited opposite effects. CCl4- induced rat hepatic injury model, HSC-MVs reduced the release of ALT and AST, hepatic inflammation, fatty deformation, and liver fibrosis. HSC-MVs also downregulated the protein expression of CD31 and CD34. Conversely, TGF-ß1HSC-MVs demonstrated opposite effects. CONCLUSION: HSC-MVs demonstrated a protective effect on H2O2-treated HUVECs and CCl4-induced rat hepatic injury, while TGF-ß1HSC-MVs had an aggravating effect. The effects of MVs involve PI3K/AKT/VEGF, CXCR4, and MEK/ERK/eNOS pathways.

TGF-ß1-Induced LINC01094 promotes epithelial-mesenchymal transition in hepatocellular carcinoma through the miR-122-5p/TGFBR2-SAMD2-SMAD3 Axis.

Hepatocellular carcinoma (HCC) is a common malignancy with a poor prognosis. It has been proven that long non-coding RNAs (lncRNAs) play an essential role in regulating HCC progression. However, the involvement of LINC01094 in regulating epithelial-mesenchymal transition (EMT) in HCC remains unclear. LINC01094 expression in HCC patients was retrieved from the Cancer Genome Atlas database. Overexpressing and downregulating LINC01094 were conducted to investigate its biological functions using Hep3B, SNU-387, and HuH-7 cells. Western blotting and morphological observation were performed to study the EMT in HCC cells. Transwell assay was adopted to determine the migration and invasion of HCC cells. The underlying mechanism of competitive endogenous RNAs (ceRNAs) was investigated using bioinformatics analysis, quantitative reverse-transcription polymerase chain reaction, and rescue experiments. Elevated LINC01094 expression was observed in HCC and associated with a poor prognosis. Knockdown of LINC01094 expression in SNU-387 and HuH-7 cells could inhibit migration, invasion, and EMT markers. Overexpression of LINC01094 indicated that LINC01094 promoted EMT via the TGF-ß/SMAD signaling pathway. The bioinformatics analysis revealed that miR-122-5p was a target of LINC01094. The miRWalk database analysis showed that TGFBR2, SMAD2, and SMAD3 were downstream targets of miR-122-5p. Mechanically, LINC01094 acted as a ceRNA that facilitated HCC metastasis by sponging miR-122-5p to regulate the expression of TGFBR2, SMAD2, and SMAD3. Further, TGF-ß1 could enhance the expression of LINC01094, forming a positive feedback loop. TGF-ß1-induced LINC01094 expression promotes HCC cell migration and invasion by targeting the miR-122-5p/TGFBR2-SMAD2-SMAD3 axis. LINC01094 may be a potential prognostic biomarker and therapeutic target for HCC metastasis.

Hypermethylation of the FOXP3 gene regulates Tregs immunodysregulation in chronic idiopathic thrombocytopenic purpura.

BACKGROUND: Chronic idiopathic thrombocytopenic purpura (ITP) is an autoimmune disease characterized by a breakdown of immune tolerance; in ITP, the body's immune system mistakenly attacks and destroys platelets. This study aims to investigate the role and underlying mechanisms of FOXP3 in chronic ITP. METHODS: Flow cytometry was used to detect the proportion of CD4+CD25+FOXP3+ regulatory T cells (Tregs) in CD4+CD25+ T lymphocytes from 20 patients with chronic ITP (CITP), 20 acute ITP (AITP) controls, and 20 healthy individuals.CD4+CD25+ Treg cells were isolated from peripheral blood of patients with CITP using magnetic beads and then treated with phosphate-buffered saline solution or decitabine (a methylation inhibitor) for 48 h. The levels of interleukin-2 (IL-2), IL-10, and transforming growth factor-beta1 (TGF-ß1) in the plasma and CD4+CD25+ Treg cells were assessed by Enzyme-linked-immunosorbent serologic assay and quantitative real-time polymerase chain reaction (qRT-PCR). FOXP3 level was measured by qRT-PCR and Western blot analysis. Methylation-specific PCR (MS-PCR) was adopted to detect the status of FOXP3 methylation. RESULTS: The number of Treg cells and the contents of IL-2, IL-10, and TGF-ß1 decreased in patients with CITP, compared to the AITP control group and normal group. FOXP3 expression was reduced and FOXP3 methylation increased in patients with CITP, compared to the AITP control group and normal group. Hypermethylation of FOXP3 promoter led to decrease in FOXP3 level in Treg cells. Inhibition of FOXP3 promoter hypermethylation promoted the secretion of IL-2, IL-10, and TGF-ß1 in Treg cells. CONCLUSION: The number of Treg cells in CITP patients decreased, and the hypermethylation of FOXP3 promoter led to reduction of its expression in Treg cells, thus affecting the immune functioning of Treg cells.

Fibrogenesis in Human Mucosa and Muscularis Precision-Cut Intestinal Slices.

In Crohn's Disease (CD), intestinal fibrosis is a prevalent yet unresolved complication arising from chronic and transmural inflammation. The histological assessment of CD intestines shows changes in tissue morphology in all the layers, including the mucosa and muscularis. This study aimed to determine the differences in fibrogenesis between mucosa and muscularis. Human precision-cut intestinal slices (hPCIS) were prepared from human intestine mucosa and muscularis and treated with TGF-ß1 and/or PDGF-BB for 72 h. Gene and protein expression and matrix metalloproteinase (MMP) activity were determined. The basal gene expression of various fibrosis markers was higher in muscularis compared to mucosa hPCIS. During incubation, Pro-Collagen-1A1 secretion increased in muscularis but not in mucosa hPCIS. MMP gene expression increased during incubation in mucosa and muscularis hPCIS, except for MMP9, MMP12, and MMP13 in muscularis hPCIS. Incubation with TGF-ß1 caused increased COL1A1 expression in the mucosa but not in muscularis hPCIS. In muscularis hPCIS, TGF-ß1 treatment caused a decrease in MMP1 and CTSK expression, while MMP13 was increased. In the presence of TGF-ß1, protease inhibitor expression was stable, except for SERPINE1, which was increased in muscularis hPCIS. We conclude that fibrogenesis is more pronounced in muscularis hPCIS compared to mucosa hPCIS, especially when stimulated with TGF-ß1.

Cancer-Derived Small Extracellular Vesicles Affect Vascular Endothelial Cells and Promote Adhesiveness of Pancreatic Cancer Cells.

OBJECTIVES: Pancreatic cancer (PC) is one of the most aggressive malignancies due to the high rate of metastasis. The mechanisms underlying metastasis need to be elucidated. Small extracellular vesicles (sEVs) mediate cell-to-cell communication, and cancer-derived sEVs contribute to the formation of premetastatic niches. The present study examined changes in adhesiveness by the internalization of PC-derived sEVs into vascular endothelial cells, and investigated the molecular mechanisms underlying metastasis. MATERIALS AND METHODS: Pancreatic cancer-derived sEVs were internalized into vascular endothelial cells, and changes in adhesiveness were evaluated. We evaluated the effects of sEVs on the formation of liver metastasis in vivo. We also assessed molecular changes in vascular endothelial cells by the internalization of PC-derived sEVs. RESULTS: The internalization of PC-derived sEVs into vascular endothelial cells promoted the adhesiveness of vascular endothelial cells and PC cells. Pancreatic cancer-derived sEVs contained high levels of transforming growth factor ß1 mRNA and acted as its transporter. Once PC-derived sEVs were internalized into vascular endothelial cells, the expression of fibronectin 1 increased on the cell surface, and the adhesiveness of vascular endothelial cells was enhanced. CONCLUSIONS: We investigated association between PC-derived sEVs and adhesiveness. Regulation of PC-derived sEVs has potential as a therapeutic modality to suppress the metastasis of PC.

A Dynamic Cellular Model as an Emerging Platform to Reproduce the Complexity of Human Vascular Calcification In Vitro.

Vascular calcification (VC) is a cardiovascular disease characterized by calcium salt deposition in vascular smooth muscle cells (VSMCs). Standard in vitro models used in VC investigations are based on VSMC monocultures under static conditions. Although these platforms are easy to use, the absence of interactions between different cell types and dynamic conditions makes these models insufficient to study key aspects of vascular pathophysiology. The present study aimed to develop a dynamic endothelial cell-VSMC co-culture that better mimics the in vivo vascular microenvironment. A double-flow bioreactor supported cellular interactions and reproduced the blood flow dynamic. VSMC calcification was stimulated with a DMEM high glucose calcification medium supplemented with 1.9 mM NaH2PO4/Na2HPO4 (1:1) for 7 days. Calcification, cell viability, inflammatory mediators, and molecular markers (SIRT-1, TGFß1) related to VSMC differentiation were evaluated. Our dynamic model was able to reproduce VSMC calcification and inflammation and evidenced differences in the modulation of effectors involved in the VSMC calcified phenotype compared with standard monocultures, highlighting the importance of the microenvironment in controlling cell behavior. Hence, our platform represents an advanced system to investigate the pathophysiologic mechanisms underlying VC, providing information not available with the standard cell monoculture.

TGFß1 Regulates Cellular Composition of In Vitro Cardiac Perivascular Niche Based on Cardiospheres.

The cardiac perivascular niche is a cellular microenvironment of a blood vessel. The principles of niche regulation are still poorly understood. We studied the effect of TGFß1 on cells forming the cardiac perivascular niche using 3D cell culture (cardiospheres). Cardiospheres contained progenitor (c-Kit), endothelial (CD31), and mural (αSMA) cells, basement membrane proteins (laminin) and extracellular matrix proteins (collagen I, fibronectin). TGFß1 treatment decreased the length of CD31+ microvasculature, VE cadherin protein level, and proportion of NG2+ cells, and increased proportion of αSMA+ cells and transgelin/SM22α protein level. We supposed that this effect is related to the stabilizing function of TGFß1 on vascular cells: decreased endothelial cell proliferation, as shown for HUVEC, and activation of mural cell differentiation.

[Role and mechanism of epithelial-mesenchymal transition in a rat model of bronchopulmonary dysplasia induced by hyperoxia exposure]. / 上皮-间å è´¨è½¬åŒ–åœ¨é«˜æ°§è¯±å¯¼å¤§é¼ æ”¯æ°”ç®¡è‚ºå‘è‚²ä¸è‰¯æ¨¡åž‹ä¸­çš„ä½œç”¨åŠæœºåˆ¶ç ”ç©¶.

OBJECTIVES: To investigate the role and mechanism of epithelial-mesenchymal transition (EMT) in a rat model of bronchopulmonary dysplasia (BPD). METHODS: The experiment consisted of two parts. (1) Forty-eight preterm rats were randomly divided into a normoxia group and a hyperoxia group, with 24 rats in each group. The hyperoxia group was exposed to 85% oxygen to establish a BPD model, while the normoxia group was kept in room air at normal pressure. Lung tissue samples were collected on days 1, 4, 7, and 14 of the experiment. (2) Rat type II alveolar epithelial cells (RLE-6TN) were randomly divided into a normoxia group (cultured in air) and a hyperoxia group (cultured in 95% oxygen), and cell samples were collected 12, 24, and 48 hours after hyperoxia exposure. Hematoxylin-eosin staining was used to observe alveolarization in preterm rat lungs, and immunofluorescence was used to detect the co-localization of surfactant protein C (SPC) and α-smooth muscle actin (α-SMA) in preterm rat lung tissue and RLE-6TN cells. Quantitative real-time polymerase chain reaction and protein immunoblotting were used to detect the expression levels of EMT-related mRNA and proteins in preterm rat lung tissue and RLE-6TN cells. RESULTS: (1) Compared with the normoxia group, the hyperoxia group showed blocked alveolarization and simplified alveolar structure after 7 days of hyperoxia exposure. Co-localization of SPC and α-SMA was observed in lung tissue, with decreased SPC expression and increased α-SMA expression in the hyperoxia group at 7 and 14 days of hyperoxia exposure compared to the normoxia group. In the hyperoxia group, the mRNA and protein levels of TGF-ß1, α-SMA, and N-cadherin were increased, while the mRNA and protein levels of SPC and E-cadherin were decreased at 7 and 14 days of hyperoxia exposure compared to the normoxia group (P<0.05). (2) SPC and α-SMA was observed in RLE-6TN cells, with decreased SPC expression and increased α-SMA expression in the hyperoxia group at 24 and 48 hours of hyperoxia exposure compared to the normoxia group. Compared to the normoxia group, the mRNA and protein levels of SPC and E-cadherin in the hyperoxia group were decreased, while the mRNA and protein levels of TGF-ß1, α-SMA, and E-cadherin in the hyperoxia group increased at 48 hours of hyperoxia exposure (P<0.05). CONCLUSIONS: EMT disrupts the tight connections between alveolar epithelial cells in a preterm rat model of BPD, leading to simplified alveolar structure and abnormal development, and is involved in the development of BPD. Citation:Chinese Journal of Contemporary Pediatrics, 2024, 26(7): 765-773.

Nebulized milk exosomes loaded with siTGF-ß1 ameliorate pulmonary fibrosis by inhibiting EMT pathway and enhancing collagen permeability.

Pulmonary Fibrosis (PF) is a fatal disease in the interstitial lung associated with high mortality, morbidity, and poor prognosis. Transforming growth factor-ß1 (TGF-ß1) is a fibroblast-activating protein that promotes fibrous diseases. Herein, an inhalable system was first developed using milk exosomes (M-Exos) encapsulating siRNA against TGF-ß1 (MsiTGF-ß1), and their therapeutic potential for bleomycin (BLM)-induced PF was investigated. M-siTGF-ß1 was introduced into the lungs of mice with PF through nebulization. The collagen penetration effect and lysosomal escape ability were verified in vitro. Inhaled MsiTGF-ß1 notably alleviated inflammatory infiltration, attenuated extracellular matrix (ECM) deposition, and increased the survival rate of PF mice by 4.7-fold. M-siTGF-ß1 protected lung tissue from BLM toxicity by efficiently delivering specific siRNA to the lungs, leading to TGF-ß1 mRNA silencing and epithelial mesenchymal transition pathway inhibition. Therefore, M-siTGF-ß1 offers a promising avenue for therapeutic intervention in fibrosis-related disorders.

Decorin attenuates hypertrophic scar fibrosis via TGFß/Smad signalling.

The management of hypertrophic scars (HSs), characterized by excessive collagen production, involves various nonsurgical and surgical interventions. However, the absence of a well-defined molecular mechanism governing hypertrophic scarring has led to less-than-ideal results in clinical antifibrotic treatments. Therefore, our study focused on the role of decorin (DCN) and its regulatory role in the TGF-ß/Smad signalling pathway in the development of HSs. In our research, we observed a decrease in DCN expression within hypertrophic scar tissue and its derived cells (HSFc) compared to that in normal tissue. Then, the inhibitory effect of DCN on collagen synthesis was confirmed in Fc and HSFc via the detection of fibrosis markers such as COL-1 and COL-3 after the overexpression and knockdown of DCN. Moreover, functional assessments revealed that DCN suppresses the proliferation, migration and invasion of HSFc. We discovered that DCN significantly inhibits the TGF-ß1/Smad3 pathway by suppressing TGF-ß1 expression, as well as the formation and phosphorylation of Smad3. This finding suggested that DCN regulates the synthesis of collagen-based extracellular matrix and fibrosis through the TGF-ß1/Smad3 pathway.

The flavonoids from the fruits of Psoralea corylifolia and their potential in inhibiting metastasis of human non-small cell lung cancers.

Nineteen flavonoids were isolated from the fruits of Psoralea corylifolia L., including a novel flavanol (3) and three novel isoflavones (12-14). Their chemical structures were unequivocally determined through comprehensive spectral data analysis. The anti-proliferative effect of the isolated flavonoids was assessed in vitro using the MTT assay. Molecular docking and ELISA were employed to determine the inhibitory effects of the active compounds on ALK5. Isobavachalcone was found to inhibit TGF-ß1 induced EMT in A549 cells by Wound healing assay and Transwell chamber assay. Immunofluorescence assay and Western blot assay showed that IBC could inhibit cytoskeleton rearrangement, reduce the phosphorylation of ALK5, ERK, and Smad, down-regulate Snail expression, and up-regulate E-cadherin expression in TGF-ß1 induced A549 cells, thereby exerting the potential inhibitory effects on epithelial-mesenchymal transition (EMT) process in A549 cells. The findings presented herein establish a fundamental basis for investigating the anti-proliferative and anti-metastatic properties of psoralen flavonoids in human non-small cell lung cancer.

TGF-ß1 mediates hypoxia-preconditioned olfactory mucosa mesenchymal stem cells improved neural functional recovery in Parkinson's disease models and patients.

BACKGROUND: Parkinson's disease (PD) is a neurodegenerative disorder characterized by the degeneration of dopaminergic neurons in the substantia nigra (SN). Activation of the neuroinflammatory response has a pivotal role in PD. Mesenchymal stem cells (MSCs) have emerged as a promising therapeutic approach for various nerve injuries, but there are limited reports on their use in PD and the underlying mechanisms remain unclear. METHODS: We investigated the effects of clinical-grade hypoxia-preconditioned olfactory mucosa (hOM)-MSCs on neural functional recovery in both PD models and patients, as well as the preventive effects on mouse models of PD. To assess improvement in neuroinflammatory response and neural functional recovery induced by hOM-MSCs exposure, we employed single-cell RNA sequencing (scRNA-seq), assay for transposase accessible chromatin with high-throughput sequencing (ATAC-seq) combined with full-length transcriptome isoform-sequencing (ISO-seq), and functional assay. Furthermore, we present the findings from an initial cohort of patients enrolled in a phase I first-in-human clinical trial evaluating the safety and efficacy of intraspinal transplantation of hOM-MSC transplantation into severe PD patients. RESULTS: A functional assay identified that transforming growth factor-ß1 (TGF-ß1), secreted from hOM-MSCs, played a critical role in modulating mitochondrial function recovery in dopaminergic neurons. This effect was achieved through improving microglia immune regulation and autophagy homeostasis in the SN, which are closely associated with neuroinflammatory responses. Mechanistically, exposure to hOM-MSCs led to an improvement in neuroinflammation and neural function recovery partially mediated by TGF-ß1 via activation of the anaplastic lymphoma kinase/phosphatidylinositol-3-kinase/protein kinase B (ALK/PI3K/Akt) signaling pathway in microglia located in the SN of PD patients. Furthermore, intraspinal transplantation of hOM-MSCs improved the recovery of neurologic function and regulated the neuroinflammatory response without any adverse reactions observed in patients with PD. CONCLUSIONS: These findings provide compelling evidence for the involvement of TGF-ß1 in mediating the beneficial effects of hOM-MSCs on neural functional recovery in PD. Treatment and prevention of hOM-MSCs could be a promising and effective neuroprotective strategy for PD. Additionally, TGF-ß1 may be used alone or combined with hOM-MSCs therapy for treating PD.

[Effects of Luhong Yixin Granules on myocardial fibrosis in heart failure rats based on TGF-ß1/Smads signaling pathway].

To investigate the effects of Luhong Yixin Granules on myocardial fibrosis in rats with heart failure and its possible mechanism, a total of 60 male Wistar rats were randomly divided into the control group, model group, and low-, medium-and high-dose Luhong Yixin Granules groups, with 12 rats in each group. Except for those in the control group, rats in the other groups were induced by intraperitoneal injection of doxorubicin(DOX) into a rat model. After the Luhong Yixin Granules were dissolved in the same amount of normal saline, they were given by gavage at low, medium and high doses(2.8, 5.6, 11.2 g·kg~(-1)·d~(-1)), and the control group and the model group were given the same amount of normal saline by gavage for 40 days. After the end of dosing, echocardiography was used to measure left ventricular ejection fraction(LVEF) and left ventricular fractional shortening(LVFS). Rat body weight(BW) and heart weight(HW) were calculated as HW/BW. Enzyme-linked immunosorbent assay was used to measure the levels of interleukin-6(IL-6), interleukin-17(IL-17), tumor necrosis factor-α(TNF-α), transforming growth factor-ß1(TGF-ß1), growth stimulation expressed gene 2 protein(ST2), N-terminal pro-B-type natriuretic peptide(NT-proBNP), galectin-3(Gal-3) and creatine kinase isoenzyme(CK-MB) in serum. Hematoxylin-eosin(HE) staining and Masson staining were used to observe the pathological morphology of myocardial tissue. Western blot and quantitative real-time polymerase chain reaction were used to detect the protein and mRNA expression levels of IL-6, IL-17, TNF-α, TGF-ß1, Smad3, Smad7, α-smooth muscle actin(α-SMA), and collagen Ⅰ(COL-Ⅰ), respectively. RESULTS:: showed that compared with those in the control group, LVEF, LVFS, and HW/BW in the model group were decreased(P<0.05), and the levels of IL-6, IL-17, TNF-α, TGF-ß1, ST2, NT-proBNP, Gal-3, and CK-MB were increased(P<0.05). HE staining showed inflammatory changes in myocardial tissue; Masson staining showed decreases in the cross-sectional area and ventricular cavity area of the heart, and myocardial fibrosis of varying degrees(P<0.05). The protein and mRNA expression of IL-6, IL-17, TNF-α, TGF-ß1, Smad3, α-SMA, and COL-Ⅰ were increased(P<0.05), and the protein and mRNA expression of Smad7 protein was decreased(P<0.01). Compared with those in the model group, LVEF, LVFS and HW/BW of the low-, medium-and high-dose Luhong Yixin Granules groups were increased(P<0.05), and the levels of IL-6, IL-17, TNF-α, TGF-ß1, ST2, NT-proBNP, Gal-3 and CK-MB were decreased(P<0.05). HE staining showed gradually reduced inflammatory changes of myocardial tissue, and Masson staining showed increased cross-sectional area and ventricular cavity area of the heart and decreased area of myocardial fibrosis(P<0.05). The protein and mRNA expression levels of IL-6, IL-17, TNF-α, TGF-ß1, Smad3, α-SMA, and COL-Ⅰ were decreased(P<0.05), while the protein and mRNA expression levels of Smad7 were increased(P<0.05). Luhong Yixin Granules may be of great value in the treatment of heart failure by regulating the TGF-ß1/Smads signaling pathway, inhibiting the expression of inflammation-related proteins, reducing the deposition of extracellular matrix, and alleviating myocardial fibrosis.

[Modified Fangji Huangqi Decoction alleviates renal interstitial fibrosis by inhibiting TGF-ß1/Smad/Snail signaling pathway during epithelial-mesenchymal transition].

This study investigated the effects of modified Fangji Huangqi Decoction on the expression of proteins related to epithelial-mesenchymal transition(EMT) in a mouse model of unilateral ureteral obstruction( UUO) and in a rat renal tubular epithelial cell(NRK-52E) model of fibrosis induced by transforming growth factor ß1(TGF-ß1). It aims to decipher the molecular mechanism by which modified Fangji Huangqi Decoction alleviates renal interstitial fibrosis. C57/BL mice were subjected to UUO.After the surgery, the mice were treated with 0. 5-fold and 2-fold concentrations of modified Fangji Huangqi Decoction and fosinopril sodium(positive control) for 7 days. The interstitial collagen deposition in the kidney was assessed by Masson staining. Western blot and RT-qPCR were employed to determine the expression levels of TGF-ß1, phosphorylated Smad2/3(p-Smad2/3), Smad2/3, Snail,epithelial cadherin(E-cadherin), alpha smooth muscle actin(α-SMA), and vimentin. The NRK-52E cell model induced by TGF-ß1was treated with the serum samples collected from SD rats treated with different concentrations of modified Fangji Huangqi Decoction.The CCK-8 assay was employed to examine the effects of the serum samples on NRK-52E cell proliferation. The cell morphology in different groups was observed under a microscope. Furthermore, the modeled cells were treated with the serum containing 1-fold decoction. Western blot and RT-qPCR were then employed to measure the expression levels of p-Smad2/3, Smad2/3, Snail,E-cadherin, α-SMA, and vimentin in the cells. Under the same conditions, sh RNA was used to silence the Snail gene, and measurements were repeated before and after treatment with the serum containing 1-fold decoction. The results indicated that modified Fangji Huangqi Decoction alleviated the fibrotic injury in the mouse model of UUO and the fibrosis in the NRK-52E cell model. The treatment with the decoction down-regulated the protein and m RNA levels of EMT-related indicators including p-Smad2/3, α-SMA,Snail, and vimentin, while it up-regulated the expression of E-cadherin. After sh RNA silencing of the Snail gene, the protein and m RNA levels of E-cadherin, α-SMA, and vimentin showed no significant differences before and after treatment with the serum containing the decoction. The results suggest that modified Fangji Huangqi Decoction may alleviate renal interstitial fibrosis by inhibiting the TGF-ß1/Smad/Snail signaling pathway and regulating the EMT process.