BACKGROUND: Resistance to chemotherapy containing cisplatin (DDP) is a main challenge in the treatment of triple-negative breast cancer (TNBC). Forkhead box O4 (FOXO4) is frequently downregulated in DDP-resistant cells. However, it is unclear whether FOXO4 down-regulation is related to DDP resistance. Here, we investigated the relationship between FOXO4 and DDP resistance in TNBC.
METHODS: We established the DDP-resistant cell lines MDA-MB-231/DDP and BT-549/DDP through in vitro selection. CCK-8 and colony formation assays analyzed cell growth. The resistance index was calculated. Cell autophagy was evaluated. Western blotting and qRT-PCR measured protein and gene expression. The binding between FOXO4 and TGF-β1 was determined by the dual-luciferase reporter assay.
RESULTS: FOXO4 expression was significantly lower in MDA-MB-231/DDP and BT-549/DDP cells. FOXO4 overexpression increased the sensitivity of TNBC cells to DDP. The PI3K class Ⅲ and Beclin-1 levels and LC3-II/LC3-I ratio elevated significantly in DDP-resistant cells. Moreover, the autophagic flux was enhanced in DDP-resistant cells. 3-MA enhanced the sensitivity of TNBC cells to DDP by inhibiting autophagy. Overexpression of FOXO4, treatment with 3-MA, and their combination significantly reduced the drug resistance index. FOXO4 directly targeted TGF-β1. Additionally, TGF-β1 knockdown inhibited autophagy and restored the sensitivity of DDP-resistant cells to DDP. Mechanistically, FOXO4 affected TNBC resistance to DDP by regulating autophagy and TGF-β1.
CONCLUSIONS: FOXO4 overexpression, in combination with autophagy inhibitors, can significantly improve the sensitivity of TNBC-resistant cells to DDP. These findings reveal the role and mechanism of FOXO4 in DDP sensitivity and may provide evidence for the development of TNBC therapies.

UNASSIGNED: Pathological fibrosis of the myodural bridge (MDB) affects cerebrospinal fluid circulation. However, no optimal drug treatments are available. We aimed to explore the antifibrotic effect of resveratrol on bleomycin-induced pathological fibrosis of the MDB and its underlying mechanisms.
UNASSIGNED: Genes common to the potential targets of resveratrol were determined using network pharmacology, genes related to muscle and tendon fibrosis were acquired from the GeneCards database, and genes related to MDB development were determined using Venny. These genes were considered potential resveratrol treatment targets in bleomycin-induced pathological fibrosis of the MDB and were annotated using bioinformatics methods. We validated the intersected genes using quantitative real-time polymerase chain reaction (qRT-PCR) and performed molecular docking analysis to calculate the binding activity between the target gene and resveratrol. Hematoxylin and eosin and Masson staining were used to detect the morphological changes in bleomycin-induced fibrosis of the MDB following resveratrol treatment. We used qRT-PCR and immunohistochemistry to evaluate the expression of the sirtuin 3 (SIRT3)/transforming growth factor-β1 (TGF-β1)/Smad pathway and the profibrotic markers α-smooth muscle actin (α-SMA) and Collagen Ⅰ.
UNASSIGNED: Through network pharmacology and bioinformatics analyses, we identified four core intersected genes, and SIRT3 expression was validated using qRT-PCR. Molecular docking analysis revealed that resveratrol had good binding affinity for SIRT3. Resveratrol ameliorated morphological abnormalities in bleomycin-induced pathological fibrosis of the MDB by inhibiting fibroblast activation and excessive collagen fiber deposition. Resveratrol exerted its antifibrotic effect by regulating the SIRT3/TGF-β1/Smad pathway.
UNASSIGNED: Resveratrol has an antifibrotic effect in bleomycin-induced pathological fibrosis of the MDB in vivo and may be considered a novel therapeutic strategy.

Previous research has revealed that platelets promote tumor metastasis by binding to circulating tumor cells (CTCs). However, the role of platelets in epithelial-mesenchymal transition (EMT) of cancer cells at the primary tumor site, the crucial initial step of tumor metastasis, remains to be elucidated. Here, we found that platelet releasate enhanced EMT and motility of hepatocellular carcinoma (HCC) cells via AMPK/mTOR-induced autophagy. RNA-seq indicated that platelet releasate altered TGF-β signaling pathway of cancer cells. Inhibiting TGFBR or deleting platelet TGF-β1 suppressed AMPK/mTOR pathway activation and autophagy induced by platelet releasate. Compared with Pf4cre-; Tgfb1fl/fl mice, HCC orthotopic models established on Pf4cre+; Tgfb1fl/fl mice showed reduced TGF-β1 in primary tumors, which corresponded with decreased cancer cell EMT, autophagy, migration ability and tumor metastasis. Inhibition of autophagy via Atg5 knockdown in cancer cells negated EMT and metastasis induced by platelet-released TGF-β1. Clinically, higher platelet count correlated with increased TGF-β1, LC3 and N-cad expression in primary tumors of HCC patients, suggesting a link between platelets and HCC progression. Our study indicates that platelets promote cancer cell EMT in the primary tumor and HCC metastasis through TGF-β1-induced HCC cell autophagy via the AMPK/mTOR pathway. These findings offer novel insights into the role of platelets in HCC metastasis and the potential therapeutic targets for HCC metastasis.

Histone methyltransferase KMT2D is one of the most frequently mutated genes in diffuse large B-cell lymphoma (DLBCL) and has been identified as an important pathogenic factor and prognostic marker. However, the biological relevance of KMT2D mutations on tumor microenvironment remains to be determined. KMT2D mutations were assessed by whole-genome/exome sequencing (WGS/WES) in 334 patients and by targeted sequencing in 427 patients with newly diagnosed DLBCL. Among all 761 DLBCL patients, somatic mutations in KMT2D were observed in 143 (18.79%) patients and significantly associated with advanced Ann Arbor stage and MYC expression ≥ 40%, as well as inferior progression-free survival and overall survival. In B-lymphoma cells, the mutation or knockdown of KMT2D inhibited methylation of lysine 4 on histone H3 (H3K4), downregulated FBXW7 expression, activated NOTCH signaling pathway and downstream MYC/TGF-β1, resulting in alterations of tumor-induced regulatory T cell trafficking. In B-lymphoma murine models established with subcutaneous injection of SU-DHL-4 cells, xenografted tumors bearing KMT2D mutation presented lower H3K4 methylation, higher regulatory T cell recruitment, thereby provoking rapid tumor growth compared with wild-type KMT2D via FBXW7-NOTCH-MYC/TGF-β1 axis.

BACKGROUND: Silicosis is an irreversible fibrotic disease of the lung caused by chronic exposure to silica dust, which manifests as infiltration of inflammatory cells, excessive secretion of pro-inflammatory cytokines, and pulmonary diffuse fibrosis. As the disease progresses, lung function further deteriorates, leading to poorer quality of life of patients. Currently, few effective drugs are available for the treatment of silicosis. Bicyclol (BIC) is a compound widely employed to treat chronic viral hepatitis and drug-induced liver injury. While recent studies have demonstrated anti-fibrosis effects of BIC on multiple organs, including liver, lung, and kidney, its therapeutic benefit against silicosis remains unclear. In this study, we established a rat model of silicosis, with the aim of evaluating the potential therapeutic effects of BIC.
METHODS: We constructed a silicotic rat model and administered BIC after injury. The FlexiVent instrument with a forced oscillation system was used to detect the pulmonary function of rats. HE and Masson staining were used to assess the effect of BIC on silica-induced rats. Macrophages-inflammatory model of RAW264.7 cells, fibroblast-myofibroblast transition (FMT) model of NIH-3T3 cells, and epithelial-mesenchymal transition (EMT) model of TC-1 cells were established in vitro. And the levels of inflammatory mediators and fibrosis-related proteins were evaluated in vivo and in vitro after BIC treatment by Western Blot analysis, RT-PCR, ELISA, and flow cytometry experiments.
RESULTS: BIC significantly improved static compliance of lung and expiratory and inspiratory capacity of silica-induced rats. Moreover, BIC reduced number of inflammatory cells and cytokines as well as collagen deposition in lungs, leading to delayed fibrosis progression in the silicosis rat model. Further exploration of the underlying molecular mechanisms revealed that BIC suppressed the activation, polarization, and apoptosis of RAW264.7 macrophages induced by SiO2. Additionally, BIC inhibited SiO2-mediated secretion of the inflammatory cytokines IL-1β, IL-6, TNF-α, and TGF-β1 in macrophages. BIC inhibited FMT of NIH-3T3 as well as EMT of TC-1 in the in vitro silicosis model, resulting in reduced proliferation and migration capability of NIH-3T3 cells. Further investigation of the cytokines secreted by macrophages revealed suppression of both FMT and EMT by BIC through targeting of TGF-β1. Notably, BIC blocked the activation of JAK2/STAT3 in NIH-3T3 cells required for FMT while preventing both phosphorylation and nuclear translocation of SMAD2/3 in TC-1 cells necessary for the EMT process.
CONCLUSIONS: The collective data suggest that BIC prevents both FMT and EMT processes, in turn, reducing aberrant collagen deposition. Our findings demonstrate for the first time that BIC ameliorates inflammatory cytokine secretion, in particular, TGF-β1, and consequently inhibits FMT and EMT via TGF-β1 canonical and non-canonical pathways, ultimately resulting in reduction of aberrant collagen deposition and slower progression of silicosis, supporting its potential as a novel therapeutic agent.

UNASSIGNED: Hypertrophic scar (HS) is a common fibrotic skin disease that occurs secondary to burns or injuries. The activation of the TGF-β signaling pathway contributes immensely to HS formation. Isorhamnetin (ISO) is a type of flavonoid compound that exerts an antifibrotic effect via TGF-β signaling suppression. However, whether ISO can inhibit HS formation via TGF-β signaling is yet to be elucidated. This study aimed to examine the influence of ISO on HS pathogenesis and TGF-β signaling, especially the downstream molecules and networks of TGF-β signaling that facilitate HS formation.
UNASSIGNED: Hypertrophic scar fibroblasts (HSFBs) were isolated from human HS tissues. The in vitro proliferation, migration, contractile ability, cell cycle, and apoptosis of HSFBs after ISO treatment were determined using cell viability assay, EdU staining, wound healing assay, collagen gel contraction assay, and flow cytometry. The expressions of genes and proteins involved in TGF-β signaling and its downstream molecules in ISO-treated HSFBs were determined using quantitative PCR (qPCR), immunofluorescence, and western blotting. In vivo, a rabbit HS model was established, and the effects of ISO on rabbit HS formation were investigated using histological analysis, immunohistochemical staining, and qPCR.
UNASSIGNED: In vitro studies indicated that ISO treatment suppressed the proliferation, migration, and contractile ability of HSFBs; attenuated the expressions of COL Ⅰ, COL Ⅲ, and α-SMA; and inhibited TGF-β1 signaling-induced activation of HSFBs by decreasing the levels of phosphorylated Smad2/3 and cleaved CREB3L1 in a dose-dependent manner. Furthermore, ISO augmented apoptosis and G2 phase cell cycle arrest of HSFBs by upregulating the expressions of the proapoptotic proteins Bax and cleaved caspase-3 and downregulating the expression of the antiapoptotic protein Bcl-2. In vivo studies revealed that ISO ameliorated HS formation in the rabbit ear by lowering the scar elevation index, attenuating the collagen density, facilitating the regular arrangement of collagen fibers, and downregulating the expressions of TGF-β1, CREB3L1, COL Ⅰ, COL Ⅲ, and α-SMA.
UNASSIGNED: ISO suppressed HS pathogenesis by dampening TGF-β1/Smad and TGF-β1/CREB3L1 signaling pathways, which suggests that it may serve as a candidate inhibitor of TGF-β1 signaling and a promising anti-HS drug with a high therapeutic potential.

Ischemic stroke is one of the most disabling and fatal diseases around the world. The damaged brain tissues will undergo excessive autophagy, vascular endothelial cells injury, blood-brain barrier (BBB) impairment and neuroinflammation after ischemic stroke. However, there is no unified viewpoint on the underlying mechanism of brain damage. Transforming growth factor-β1 (TGF-β1), as a multi-functional cytokine, plays a crucial role in the intricate pathological processes and helps maintain the physiological homeostasis of brain tissues through various signaling pathways after ischemic stroke. In this review, we summarize the protective role of TGF-β1 in autophagic flux, BBB, vascular remodeling, neuroinflammation and other aspects after ischemic stroke. Based on the review, we believe that TGF-β1 could serve as a key target for treating ischemic stroke.

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-β/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-β1 could enhance the expression of LINC01094, forming a positive feedback loop. TGF-β1-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.

BACKGROUND: Peyronie\'s disease (PD) causes benign plaques or induration in tunica albuginea (TA). Kindlin-2 regulates the TGF-β1/Smad3 pathway, which accelerates kidney fibrosis. The study is aimed mainly to investigate the impact of Kindlin-2 on PD formation and its signaling pathways, notably the TGF-β/Smad pathway in the presence of TGF-β1.
METHODS: In this mouse investigation, adenovirus TGF-β1 was injected into TA to produce PD. The model was successfully induced 45 days later. Western Blot (WB) and immunohistochemistry (IHC) were utilized to measure Kindlin-2 in PD model tissue. WB and immunofluorescence assays were utilized to confirm the impact of TGF-β1 on Kindlin-2 levels in vitro. The interaction among Kindlin-2, TβRI, and Smad3 was detected using immunoprecipitation (IP) experiments. We examined how TGF-β1 affects Smad3 phosphorylation and downstream gene activation process. Finally, Kindlin-2 and the level of tissue fibrosis were examined in PD model.
RESULTS: Kindlin-2 levels were elevated in the TGF-β1-induced PD model, confirming that TGF-β1 can increase Kindlin-2 levels in primary PD cells. Moreover, Kindlin-2 mediates Smad3-TβRI interaction, activates p-Smad3, and enhances TGF-β1 target gene expression. In vivo investigations reveal that Kindlin-2 promotes PD development and tissue fibrosis. The regulatory effects of Kindlin-2 need the presence of TGF-β1. Tissue fibrosis can be reduced by downregulating Kindlin-2.
CONCLUSIONS: Kindlin-2 does not directly activate Smad3 to induce tissue fibrosis. Instead, it exerts its effect through the combined influence of TGF-β1. Inhibiting Kindlin-2 could potentially be a treatment for PD.

Spinal cord injury (SCI) remains a worldwide challenge due to limited treatment strategies. Repetitive trans-spinal magnetic stimulation (rTSMS) is among the most cutting-edge treatments for SCI. However, the mechanism underlying rTSMS on functional recovery is still unclear. In this study, 8-week-old C57BL/6J female mice were used to design SCI models followed by treatment with monotherapy (1 Hz rTSMS or LY364947) or combination therapy (rTSMS + LY364947). Our results showed obvious functional recovery after monotherapies compared to untreated mice. Immunofluorescence results demonstrated that rTSMS and LY364947 modulate the lesion scar by decreasing fibrosis and GFAP and possess the effect on neural protection. In addition, rTSMS suppressed inflammation and the activation of TGFβ1/Smad2/3 signaling pathway, as evidenced by markedly reduced TGF-βRⅠ, Smad2/3, and p-Smad2/3 compared with untreated mice. Overall, it was confirmed that 1 Hz rTSMS promotes SCI recovery by suppressing the TGFβ1/Smad2/3 signaling, revealing a novel pathological mechanism of 1 Hz rTSMS intervention, and may provide potential targets for clinical treatment.