pSTAT3 activation of Foxl2 initiates the female pathway underlying temperature-dependent sex determination.

Ovarian development was traditionally recognized as a "default" sexual outcome and therefore received much less scientific attention than testis development. In turtles with temperature-dependent sex determination (TSD), how the female pathway is initiated to induce ovary development remains unknown. In this study, we have found that phosphorylation of the signal transducer and activator of transcription 3 (pSTAT3) and Foxl2 exhibit temperature-dependent sexually dimorphic patterns and tempo-spatial coexpression in early embryos of the red-eared slider turtle (Trachemys scripta elegans). Inhibition of pSTAT3 at a female-producing temperature of 31 °C induces 64.7% female-to-male sex reversal, whereas activation of pSTAT3 at a male-producing temperature of 26 °C triggers 75.6% male-to-female sex reversal. In addition, pSTAT3 directly binds to the locus of the female sex-determining gene Foxl2 and promotes Foxl2 transcription. Overexpression or knockdown of Foxl2 can rescue the sex reversal induced by inhibition or activation of pSTAT3. This study has established a direct genetic link between warm temperature-induced STAT3 phosphorylation and female pathway initiation in a TSD system, highlighting the critical role of pSTAT3 in the cross talk between female and male pathways.

Downregulation of CIAPIN1 regulates the proliferation, migration and glycolysis of breast cancer cells via inhibition of STAT3 pathway.

Cytokine-induced apoptosis inhibitor 1 (CIAPIN1) is a protein that regulates apoptosis and programmed cell death. This research aims to evaluate its potential role in inhibiting breast cancer cell proliferation, migration, and glycolysis and uncover its underlying molecular mechanism. We collected breast cancer tissue samples from eight patients between January 2019 and June 2023 in our Hospital to analyse CIAPIN1 expression. We transfected human breast cancer cell lines (MCF7, MDA-MB-231, MDA-MB-453, and MDA-MB-468) with siRNA of CIAPIN1. Finally, we determined protein expression using RT-qPCR and Western blotting. CIAPIN1 expression was elevated in both breast cancer tissue and serum. Overexpression of CIAPIN1 detected in the breast cancer cell lines MCF7 and MDA-MB-468. In addition, CIAPIN1 overexpression increased cell proliferation and migration rate. CIAPIN1 downregulation suppressed cell proliferation while elevated cellular apoptosis, reactive oxygen species (ROS) production and oxidative stress in breast cancer cells. Moreover, CIAPIN1 inhibition remarkably suppressed pyruvate, lactate and adenosine triphosphate (ATP) production and reduced the pyruvate kinase M2 (PKM2) protein expression and phosphorylation of signal transducer and activator of transcription 3 (STAT3) in breast cancer cells. Downregulation of CIAPIN1 suppresses cell proliferation, migration and glycolysis capacity in breast cancer cells by inhibiting the STAT3/PKM2 pathway.

Linoleyl acetate and mandenol alleviate HUA-induced ED via NLRP3 inflammasome and JAK2/STAT3 signalling conduction in rats.

Hyperuricemia (HUA) is characterized by elevated blood uric acid levels, which can increase the risk of erectile dysfunction (ED). Clinical studies have demonstrated satisfactory efficacy of a traditional Chinese medicine formula QYHT decoction in improving ED. Furthermore, the main monomeric components of this formula, linoleyl acetate and mandenol, demonstrate promise in the treatment of ED. This study established an ED rat model induced by HUA and the animals were administered with linoleyl acetate and mandenol. HE and TUNEL were performed to detect tissue changes, ELISA to measure the levels of serum testosterone (T), MDA, NO, CRP, and TNF-α and qPCR and WB to assess the expression levels of NLRP3, ASC, Caspase-1, JAK2, and STAT3 in whole blood. The findings showed that linoleyl acetate and mandenol improved kidney tissue morphology, reduced cell apoptosis in penile tissue, significantly increased T and NO levels, while substantially decreasing levels of MDA, CRP, and TNF-α. Meanwhile, the expression of NLRP3, ASC, and Caspase-1 mRNAs and proteins was markedly reduced, and the phosphorylation of JAK2 and STAT3 was inhibited. These findings were further validated through faecal microbiota transplantation results. Taken together, linoleyl acetate and mandenol could inhibit NLRP3 inflammasome activation, reduce inflammatory and oxidative stress responses, suppress the activity of JAK-STAT signalling pathway, ultimately providing a potential treatment for HUA-induced ED.

Stat3-mediated Atg7 expression regulates anti-tumor immunity in mouse melanoma.

Epigenetic modifications to DNA and chromatin control oncogenic and tumor-suppressive mechanisms in melanoma. Ezh2, the catalytic component of the Polycomb Repressive Complex 2 (PRC2), which mediates methylation of lysine 27 on histone 3 (H3K27me3), can regulate both melanoma initiation and progression. We previously found that mutant Ezh2Y641F interacts with the immune regulator Stat3 and together they affect anti-tumor immunity. However, given the numerous downstream targets and pathways affected by Ezh2, many mechanisms that determine its oncogenic activity remain largely unexplored. Using genetically engineered mouse models, we further investigated the role of pathways downstream of Ezh2 in melanoma carcinogenesis and identified significant enrichment in several autophagy signatures, along with increased expression of autophagy regulators, such as Atg7. In this study, we investigated the effect of Atg7 on melanoma growth and tumor immunity within the context of a wild-type or Ezh2Y641F epigenetic state. We found that the Atg7 locus is controlled by multiple Ezh2 and Stat3 binding sites, Atg7 expression is dependent on Stat3 expression, and that deletion of Atg7 slows down melanoma cell growth in vivo, but not in vitro. Atg7 deletion also results in increased CD8 + T cells in Ezh2Y641F melanomas and reduced myelosuppressive cell infiltration in the tumor microenvironment, particularly in Ezh2WT melanomas, suggesting a strong immune system contribution in the role of Atg7 in melanoma progression. These findings highlight the complex interplay between genetic mutations, epigenetic regulators, and autophagy in shaping tumor immunity in melanoma.

METTL14-mediated lncRNA-FAS-AS1 promotes osteoarthritis progression by up-regulating ADAM8.

BACKGROUND: Osteoarthritis (OA) is a prevalent degenerative disease. We explored the role and regulatory mechanisms of lncRNA-FAS-AS1 in OA progression. METHODS: We exposed human immortalized chondrocytes to IL-1ß for 24 h to induce an OA cell model. The target molecule levels were assessed using western blot and quantitative real-time PCR (RT-qPCR). Cell viability and apoptosis were measured using CCK-8 and flow cytometry. The m6A modification of FAS-AS1 was determined using MeRIP. We examined the binding relationships between FAS-AS1, Fragile X mental retardation 1 (FMR1), and A disintegrin and metalloproteinase 8 (ADAM8) using RIP and RNA pull-down. The OA animal model was established by separating the medial collateral ligament and medial meniscus. Safranin-O staining and Mankin's scale were employed to evaluate pathological changes within the cartilage. RESULTS: FAS-AS1, METTL14, and ADAM8 were upregulated, and the JAK/STAT3 signaling pathway was activated in OA mice and IL-1ß-induced chondrocytes. FAS-AS1 knockdown inhibited extracellular matrix degradation in IL-1ß-induced chondrocytes; however, ADAM8 overexpression reversed this effect. FAS-AS1 maintained the stability of ADAM8 mRNA by recruiting FMR1. METTL14 knockdown repressed FAS-AS1 expression in an m6A-dependent manner. FAS-AS1 overexpression reversed the inhibitory effects of METTL14 knockdown on JAK/STAT3 signaling and cartilage damage in the OA model both in vitro and in vivo. CONCLUSION: METTL14-mediated FAS-AS1 promotes OA progression through the FMR1/ADAM8/JAK/STAT3 axis.

IL-6 signaling accelerates iron overload by upregulating DMT1 in endothelial cells to promote aortic dissection.

Aortic dissection (AD), caused by tearing of the intima and avulsion of the aortic media, is a severe threat to patient life and organ function. Iron is closely related to dissection formation and organ injury, but the mechanism of iron ion transport disorder in endothelial cells (ECs) remains unclear. We identified the characteristic EC of dissection with iron overload by single-cell RNA sequencing data. After intersecting iron homeostasis and differentially expressed genes, it was found that hypoxia-inducible factor-1α (HIF-1α) and divalent metal transporter 1 (DMT1) are key genes for iron ion disorder. Subsequently, IL-6R was identified as an essential reason for the JAK-STAT activation, a classical iron regulation pathway, through further intersection and validation. In in vivo and in vitro, both high IL-6 receptor expression and elevated IL-6 levels promote JAK1-STAT3 phosphorylation, leading to increased HIF-1α protein levels. Elevated HIF-1α binds explicitly to the 5'-UTR sequence of the DMT1 gene and transcriptionally promotes DMT1 expression, thereby increasing Fe2+ accumulation and endoplasmic reticulum stress (ERS). Blocking IL-6R and free iron with deferoxamine and tocilizumab significantly prolonged survival and reduced aortic and organ damage in dissection mice. A comparison of perioperative data between AD patients and others revealed that high free iron, IL-6, and ERS levels are characteristics of AD patients and are correlated with prognosis. In conclusion, activated IL-6/JAK1/STAT3 signaling axis up-regulates DMT1 expression by increasing HIF-1α, thereby increasing intracellular Fe2+ accumulation and tissue injury, which suggests a potential therapeutic target for AD.

Ruxolitinib-based senomorphic therapy mitigates cardiomyocyte senescence in septic cardiomyopathy by inhibiting the JAK2/STAT3 signaling pathway.

Background: Cellular senescence has emerged as a pivotal focus in cardiovascular research. This study investigates the previously unrecognized role of cellular senescence in septic cardiomyopathy (SCM) and evaluates senomorphic therapy using ruxolitinib (Rux) as a potential treatment option. Methods: We employed lipopolysaccharide (LPS)-induced neonatal rat cardiomyocytes (NRCMs) and two mouse models-LPS-induced and cecal ligation and puncture (CLP)-induced SCM models-to assess Rux's effects. RNA sequencing, western blotting (WB), quantitative polymerase chain reaction (qPCR), immunofluorescence, immunohistochemistry, senescence-associated ß-galactosidase (SA-ß-gal) assay, and other techniques were utilized to investigate underlying mechanisms. Results: Senescence-associated secretory phenotype (SASP) and cellular senescence markers were markedly elevated in LPS-induced NRCMs and SCM animal models, confirmed by the SA-ß-gal assay. Rux treatment attenuated SASP in vitro and in vivo, alongside downregulation of senescence markers. Moreover, Rux-based senomorphic therapy mitigated mitochondrial-mediated apoptosis, improved cardiac function in SCM mice, restored the balance of antioxidant system, and reduced reactive oxygen species (ROS) levels. Rux treatment restored mitochondrial membrane potential, mitigated mitochondrial morphological damage, and upregulated mitochondrial complex-related gene expression, thereby enhancing mitochondrial function. Additionally, Rux treatment ameliorated SCM-induced mitochondrial dynamic dysfunction and endoplasmic reticulum stress. Mechanistically, Rux inhibited JAK2-STAT3 signaling activation both in vitro and in vivo. Notably, low-dose Rux and ABT263 showed comparable efficacy in mitigating SCM. Conclusions: This study highlighted the potential significance of cellular senescence in SCM pathogenesis and suggested Rux-based senomorphic therapy as a promising therapeutic approach for SCM.

RNF122 promotes glioblastoma growth via the JAK2/STAT3/c-Myc signaling Axis.

OBJECTIVE: The E3 ubiquitin ligase is well recognized as a significant contributor to glioblastoma (GBM) progression and has promise as a prospective therapeutic target. This study explores the contribution of E3 ubiquitin ligase RNF122 in the GBM progression and the related molecular mechanisms. METHODS: RNF122 expression levels were evaluated using qRT-PCR, WB, and IHC, while functional assays besides animal experiments were used to assess RNF122's effect on GBM progression. We also tested the RNF122 impact on JAK2/STAT3/c-Myc signaling using WB. RESULTS: RNF122 was upregulated in GBM and correlated to the advanced stage and poor clinical outcomes, representing an independent prognostic factor. Based on functional assays, RNF122 promotes GBM growth and cell cycle, which was validated further in subsequent analyses by JAK2/STAT3/c-Myc pathway activation. Moreover, JAK2/STAT3 signaling pathway inhibitor WP1066 can weaken the effect of overexpression RNF122 on promoting GBM progression. CONCLUSION: Our results revealed that RNF122 caused an aggressive phenotype to GBM and was a poor prognosticator; thus, targeting RNF122 may be effectual in GBM treatment.

Unraveling the unphosphorylated STAT3-unphosphorylated NF-κB pathway in loss of function STAT3 Hyper IgE syndrome.

Background: Patients with loss of function signal transducer and activator of transcription 3-related Hyper IgE Syndrome (LOF STAT3 HIES) present with recurrent staphylococcal skin and pulmonary infections along with the elevated serum IgE levels, eczematous rashes, and skeletal and facial abnormalities. Defective STAT3 signaling results in reduced Th17 cells and an impaired IL-17/IL-22 response primarily due to a compromised canonical Janus kinase-signal transducer and activator of transcription (JAK-STAT) pathway that involves STAT3 phosphorylation, dimerization, nuclear translocation, and gene transcription. The non-canonical pathway involving unphosphorylated STAT3 and its role in disease pathogenesis, however, is unexplored in HIES. Objective: This study aims to elucidate the role of unphosphorylated STAT3-unphosphorylated NF-κB (uSTAT3-uNF-κB) activation pathway in LOF STAT3 HIES patients. Methodology: The mRNA expression of downstream molecules of unphosphorylated STAT3-unphosphorylated NF-κB pathway was studied in five LOF STAT3 HIES patients and transfected STAT3 mutants post-IL-6 stimulation. Immunoprecipitation assays were performed to assess the binding of STAT3 and NF-κB to RANTES promoter. Results: A reduced expression of the downstream signaling molecules of the uSTAT3-uNF-κB complex pathway, viz., RANTES, STAT3, IL-6, IL-8, ICAM1, IL-8, ZFP36L2, CSF1, MRAS, and SOCS3, in LOF STAT3 HIES patients as well as the different STAT3 mutant plasmids was observed. Immunoprecipitation studies showed a reduced interaction of STAT3 and NF-κB to RANTES in HIES patients. Conclusion: The reduced expression of downstream signaling molecules, specially RANTES and STAT3, confirmed the impaired uSTAT3-uNF-κB pathway in STAT3 LOF HIES. Decreased levels of RANTES and STAT3 could be a significant component in the disease pathogenesis of Hyper IgE Syndrome.

BioID-based intact cell interactome of the Kv1.3 potassium channel identifies a Kv1.3-STAT3-p53 cellular signaling pathway.

Kv1.3 is a multifunctional potassium channel implicated in multiple pathologies, including cancer. However, how it is involved in disease progression is not fully clear. We interrogated the interactome of Kv1.3 in intact cells using BioID proximity labeling, revealing that Kv1.3 interacts with STAT3- and p53-linked pathways. To prove the relevance of Kv1.3 and of its interactome in the context of tumorigenesis, we generated stable melanoma clones, in which ablation of Kv1.3 remodeled gene expression, reduced proliferation and colony formation, yielded fourfold smaller tumors, and decreased metastasis in vivo in comparison to WT cells. Kv1.3 deletion or pharmacological inhibition of mitochondrial Kv1.3 increased mitochondrial Reactive Oxygen Species release, decreased STAT3 phosphorylation, stabilized the p53 tumor suppressor, promoted metabolic switch, and altered the expression of several BioID-identified Kv1.3-networking proteins in tumor tissues. Collectively, our work revealed the tumor-promoting Kv1.3-interactome landscape, thus opening the way to target Kv1.3 not only as an ion-conducting entity but also as a signaling hub.

A water-soluble caveolin-1 peptide inhibits psoriasis-like skin inflammation by suppressing cytokine production and angiogenesis.

The plasma membrane protein caveolin-1 (CAV-1) regulates signaling by inhibiting a wide range of kinases and other enzymes. Our previous study demonstrated that the downregulation of CAV-1 in psoriatic epidermal cells contributes to inflammation by enhancing JAK/STAT signaling, cell proliferation, and chemokine production. Administration of the CAV-1 scaffolding domain (CSD) peptide suppressed imiquimod (IMQ)-induced psoriasis-like dermatitis. To identify an optimal therapeutic peptide derived from CAV-1, we have compared the efficacy of CSD and subregions of CSD that have been modified to make them water soluble. We refer to these modified peptides as sCSD, sA, sB, and sC. In IMQ-induced psoriasis-like dermatitis, while all four peptides showed major beneficial effects, sB caused the most significant improvements of skin phenotype and number of infiltrating cells, comparable or superior to the effects of sCSD. Phosphorylation of STAT3 was also inhibited by sB. Furthermore, sB suppressed angiogenesis both in vivo in the dermis of IMQ-induced psoriasis mice and in vitro by blocking the ability of conditioned media derived from CAV-1-silenced keratinocytes to inhibit tube formation by HUVEC. In conclusion, sB had similar or greater beneficial effects than sCSD not only by cytokine suppression but by angiogenesis inhibition adding to its ability to target psoriatic inflammation.

The expansion of MDSCs induced by exosomal PD-L1 promotes the progression of gastric cancer.

BACKGROUND: Myeloid-derived suppressor cells (MDSCs) are the major factor in gastric cancer (GC) immune evasion. Nevertheless, the molecular process underlying the expansion of MDSCs induced by tumor-derived exosomes (TDEs) remains elusive. METHODS: The levels of exosomal and soluble PD-L1 in ninety GC patients were examined via enzyme-linked immunosorbent assay (ELISA) to determine their prognostic value. To investigate the correlation between exosomal PD-L1 and MDSCs, the percentage of MDSCs in the peripheral blood of 57 GC patients was assessed via flow cytometry. Through ultracentrifugation, the exosomes were separated from the GC cell supernatant and detected via Western blotting, nanoparticle tracking analysis (NTA), and transmission electron microscopy (TEM). The function of exosomal PD-L1 in MDSCs was evaluated via immunofluorescence, Western blotting and flow cytometry in a GC cell-derived xenograft (CDX) model. RESULTS: The overall survival (OS) of GC patients in the high exosomal PD-L1 group was significantly lower than that of patients in the low exosomal PD-L1 group (P = 0.0042); however, there was no significant correlation between soluble PD-L1 and OS in GC patients (P = 0.0501). Furthermore, we found that the expression of exosomal PD-L1 was positively correlated with the proportions of polymorphonuclear MDSCs (PMN-MDSCs, r = 0.4944, P < 0.001) and monocytic MDSCs (M-MDSCs, r = 0.3663, P = 0.005) in GC patients, indicating that exosomal PD-L1 might induce immune suppression by promoting the aggregation of MDSCs. In addition, we found that exosomal PD-L1 might stimulate MDSC proliferation by triggering the IL-6/STAT3 signaling pathway in vitro. The CDX model confirmed that exosomal PD-L1 could stimulate tumor development and MDSC amplification. CONCLUSIONS: Exosomal PD-L1 has the potential to become a prognostic and diagnostic biomarker for GC patients. Mechanistically, MDSCs can be activated by exosomal PD-L1 through IL-6/STAT3 signaling and provide a new strategy against GC through the use of exosomal PD-L1 as a treatment target.

Interleukin 6 and cancer resistance in glioblastoma multiforme.

Despite unprecedented survival in patients with glioblastoma (GB), the aggressive primary brain cancer remains largely incurable and its mechanisms of treatment resistance have gained particular attention. The cytokine interleukin 6 (IL-6) and its receptor weave through the hallmarks of malignant gliomas and may represent a key vulnerability to GB. Known for activating the STAT3 pathway in autocrine fashion, IL-6 is amplified in GB and has been recognized as a negative biomarker for GB prognosis, rendering it a putative target of novel GB therapies. While it has been recognized as a biologically active component of GB for three decades only with concurrent advances in understanding of complementary immunotherapy has the concept of targeting IL-6 for a human clinical trial gained scientific footing.

The role of IL-22 in cancer.

Interleukin-22, discovered in the year of 2000, is a pleiotropic Th17 cytokine from the IL-10 family of cytokines. IL-22 signals through the type 2 cytokine receptor complex IL-22R and predominantly activates STAT3. This pathway leads to the transcription of several different types of genes, giving IL-22 context-specific functions ranging from inducing antimicrobial peptide expression to target cell proliferation. In recent years, it has been shown that IL-22 is involved in the pathogenesis of neoplasia in some cancers through its pro-proliferative and anti-apoptotic effects. This review highlights studies with recent discoveries and conclusions drawn on IL-22 and its involvement and function in various cancers. Such a study may be helpful to better understand the role of IL-22 in cancer so that new treatment could be developed targeting IL-22.

Maggot extract accelerates skin wound healing of diabetic rats via enhancing STAT3 signaling.

BACKGROUND: Diabetic skin wound is a complex problem due to the disruption of normal repairing program and lack of effective remedy. Lucilia sericata larvae (maggot) is a folk method to treat chronic skin wound, while its therapeutic effects on that caused by diabetic remains unknown. OBJECTIVE: This study aims to investigate the therapeutic effects of maggot extract (M.E.) on diabetic skin wound and its molecular mechanism by establishing the skin wound model of diabetic Sprague Dawley (SD) rats. METHODS: Diabetic model was established by injecting intraperitoneally streptozotocin in SD rats under specific pathogen-free (SPF) conditions. The rat fasting blood glucose values were ≧16.7 mmol/L 72 hours after intraperitoneal streptozotocin (60mg/kg body weight) injection. The rats were divided into five groups (n = 10/group): normal group: normal SD rats without any treatment, diabetic blank group: the diabetic rats without any treatment, Vaseline group: the diabetic rats dressed with Vaseline, recombinant human epidermal-growth-factor (rhEGF) group: the diabetic rats dressed with a mixture of Vaseline and 200 µg/g rhEGF, M.E. group: the diabetic rats dressed with a mixture of Vaseline and 150 µg/ml maggot extract. The round open wounds (1.0 cm in diameter) down to the muscle fascia were made on both sides of rat dorsa by removing the skin layer (epidermis and dermis) and were daily photographed for calculating their healing rates. Hematoxylin-eosin (HE) and Masson's trichrome staining were performed on skin wound sections to analyze re-epithelialization and granulation tissue formation. Immunohistochemical (IHC), immunofluorescent (IF) stainings and Western blotting were conducted to analyze the statuses of STAT3 signaling. RESULTS: The average wound healing rates on the 14th day were 91.7% in the normal, 79.6% in M.E., 71% in rhEGF, 55.1% in vaseline and 43.3% in the diabetes blank group. Morphological staining showed more active granulation tissue formation, re-epithelialization and neovascularization in M.E.-group than those in the blank and the vaseline-treated groups. Decreased p-STAT3 nuclear tranlocation and down-regulated Bcl-2, CyclinD1 and vascular endothelial growth factor (VEGF) expression were evidenced in the diabetic rats, which could be improved by rhEGF and especially M.E. CONCLUSION: Maggot extract would be an alternative and/or adjuvant candidate for the better management of diabetic skin wounds because of its activity in enhancing STAT3 activation.

JAK inhibitors to treat STAT3 gain-of-function: a single-center report and literature review.

Objective: The signal transducer and activator of transcription 3 (STAT3) gain-of-function (GOF) syndrome (STAT3-GOF) is an inborn error of immunity (IEI) characterized by diverse manifestations of immune dysregulation that necessitate systemic immunomodulatory treatment. The blockade of the interleukin-6 receptor and/or the inhibition of the Janus kinases has been commonly employed to treat diverse STAT3-GOF-associated manifestations. However, evidence on long-term treatment outcome, especially in the case of adult patients, is scarce. Methods: Clinical data, including laboratory findings and medical imaging, were collected from all seven patients, diagnosed with STAT3-GOF, who have been treated at the Hannover University School, focusing on those who received a Janus kinase (JAK) inhibitor (JAKi). Previously published cases of STAT3-GOF patients who received a JAKi were evaluated, focusing on reported treatment efficacy with respect to diverse STAT3-GOF-associated manifestations of immune dysregulation and safety. Results: Five out of seven patients diagnosed with STAT3-GOF were treated with a JAKi, each for a different indication. Including these patients, outcomes of JAKi treatment have been reported for a total of 41 patients. Treatment with a JAKi led to improvement of diverse autoimmune, inflammatory, or lymphoproliferative manifestations of STAT3-GOF and a therapeutic benefit could be documented for all except two patients. Considering all reported manifestations of immune dysregulation in each patient, complete remission was achieved in 10/41 (24.4%) treated patients. Conclusions: JAKi treatment improved diverse manifestations of immune dysregulation in the majority of STAT3-GOF patients, representing a promising therapeutic approach. Long-term follow-up data are needed to evaluate possible risks of prolonged treatment with a JAKi.

GelMA loaded with platelet lysate promotes skin regeneration and angiogenesis in pressure ulcers by activating STAT3.

Pressure ulcers (PU) are caused by persistent long-term pressure, which compromises the integrity of the epidermis, dermis, and subcutaneous adipose tissue layer by layer, making it difficult to heal. Platelet products such as platelet lysate (PL) can promote tissue regeneration by secreting numerous growth factors based on clinical studies on skin wound healing. However, the components of PL are difficult to retain in wounds. Gelatin methacrylate (GelMA) is a photopolymerizable hydrogel that has lately emerged as a promising material for tissue engineering and regenerative medicine. The PL liquid was extracted, flow cytometrically detected for CD41a markers, and evenly dispersed in the GelMA hydrogel to produce a surplus growth factor hydrogel system (PL@GM). The microstructure of the hydrogel system was observed under a scanning electron microscope, and its sustained release efficiency and biological safety were tested in vitro. Cell viability and migration of human dermal fibroblasts, and tube formation assays of human umbilical vein endothelial cells were applied to evaluate the ability of PL to promote wound healing and regeneration in vitro. Real-time polymerase chain reaction (PCR) and western blot analyses were performed to elucidate the skin regeneration mechanism of PL. We verified PL's therapeutic effectiveness and histological analysis on the PU model. PL promoted cell viability, migration, wound healing and angiogenesis in vitro. Real-time PCR and western blot indicated PL suppressed inflammation and promoted collagen I synthesis by activating STAT3. PL@GM hydrogel system demonstrated optimal biocompatibility and favorable effects on essential cells for wound healing. PL@GM also significantly stimulated PU healing, skin regeneration, and the formation of subcutaneous collagen and blood vessels. PL@GM could accelerate PU healing by promoting fibroblasts to migrate and secrete collagen and endothelial cells to vascularize. PL@GM promises to be an effective and convenient treatment modality for PU, like chronic wound treatment.

Unveiling novel insights into human IL-6 - IL-6R interaction sites through 3D computer-guided docking and systematic site mutagenesis.

The cytokine interleukin-6 (IL-6) plays a crucial role in autoimmune and inflammatory diseases. Understanding the precise mechanism of IL-6 interaction at the amino acid level is essential to develop IL-6-inhibiting compounds. In this study, we employed computer-guided drug design tools to predict the key residues that are involved in the interaction between IL-6 and its receptor IL-6R. Subsequently, we generated IL-6 mutants and evaluated their binding affinity to IL-6R and the IL-6R - gp130 complex, as well as monitoring their biological activities. Our findings revealed that the R167A mutant exhibited increased affinity for IL-6R, leading to enhanced binding to IL-6R - gp130 complex and subsequently elevated intracellular phosphorylation of STAT3 in effector cells. On the other hand, although E171A reduced its affinity for IL-6R, it displayed stronger binding to the IL-6R - gp130 complex, thereby enhancing its biological activity. Furthermore, we identified the importance of R178 and R181 for the precise recognition of IL-6 by IL-6R. Mutants R181A/V failed to bind to IL-6R, while maintaining an affinity for the IL-6 - gp130 complex. Additionally, deletion of the D helix resulted in complete loss of IL-6 binding affinity for IL-6R. Overall, this study provides valuable insights into the binding mechanism of IL-6 and establishes a solid foundation for future design of novel IL-6 inhibitors.

Deferoxamine preconditioning of canine stem cell derived extracellular vesicles alleviates inflammation in an EAE mouse model through STAT3 regulation.

Extracellular vesicles (EVs) from mesenchymal stem cells (MSCs), specifically those preconditioned with deferoxamine (DFO) in canine adipose tissue-derived MSCs (cAT-MSCs), were explored for treating autoimmune diseases. This study assessed the effects of DFO-preconditioned EVs (EVDFO) in an experimental autoimmune encephalomyelitis (EAE) mouse model. cAT-MSCs were treated with DFO for 48 h, after which EVs were isolated. EAE mice received intranasal EV or EVDFO treatments and were euthanized following histopathologic analysis; RNA and protein expression levels were measured. Histologically, EV and EVDFO groups showed a significant reduction in inflammatory cell infiltration and demyelination. Immunofluorescence revealed increased CD206 and Foxp3 expression, indicating elevated M2 macrophages and regulatory T (Treg) cells, particularly in the EVDFO group. Treg cells also notably increased in the spleen of EVDFO -treated mice. STAT3 and pSTAT3 proteins were upregulated in the EAE groups compared to the naïve group. However, following EV treatment, STAT3 expression decreased compared to the EAE group, whereas pSTAT3 expression was similar in both the EV and EAE groups. In conclusion, EVDFO treatment resulted in reduced STAT3 expression, suggesting its role in T cell regulation and the potential of EVDFO in modulating the STAT3 pathway for reducing inflammation more effectively than non-preconditioned EVs.

Lactucin reverses liver fibrosis by inhibiting TGF-ß1/STAT3 signaling pathway and regulating short-chain fatty acids metabolism.

TGF-ß1 activation of hepatic stellate cells (HSCs), transcriptional activator 3 (Stat3) activation and short chain fatty acids (SCFAs), metabolite of intestinal bacteria, is closely associated with hepatic fibrosis. Previous studies have shown that Lactucin has significant anti-inflammatory and hepatoprotective effects; however, the mechanism of Lactucin's role in liver fibrosis associated with SCFAs remains unknown. This study was intended to investigate whether effect of Lactucin on liver fibrosis was mediated by TGF-ß1/Stat3 and SCFAs. We found that Lactucin induced apoptosis in HSC-T6 cells, and inhibition of nuclear translocation of Stat3 and p-Stat3. And Smad3 and TGF-ß1 protein expression was significantly inhibited, while TLR4 and Smad7 protein expression was significantly enhanced. For in vivo experiments, we demonstrated that Lactucin alleviated liver fibrosis in mice, as evidenced by a reduction in inflammatory factors, collagen deposition, liver injury and fibrosis-related factors expression, especially the expression of Smad3 and TGF-ß1 proteins was significantly suppressed and Smad7 protein expression was significantly increased in the liver. In addition, the levels of acetic acid, butyric acid and valeric acid in the intestine of Lactucin-treated mice were significantly higher than those in the intestine of liver fibrosis mice. In conclusion, based on the results of in vivo and in vitro experiments, preventive mechanism of Lactucin against liver fibrosis in mice may be to improve the enterohepatic circulation by regulating the metabolites of intestinal microorganisms, acetic acid and butyric acid, and to further regulate the Stat3 and TGF-ß1 signaling pathway through the "gut-liver axis" to combat liver fibrosis.