RESUMO
Integrin αvß8 binds with exquisite specificity to latent transforming growth factor-ß (L-TGF-ß). This binding is essential for activating L-TGF-ß presented by a variety of cell types. Inhibiting αvß8-mediated TGF-ß activation blocks immunosuppressive regulatory T cell differentiation, which is a potential therapeutic strategy in cancer. Using cryo-electron microscopy, structure-guided mutagenesis, and cell-based assays, we reveal the binding interactions between the entire αvß8 ectodomain and its intact natural ligand, L-TGF-ß, as well as two different inhibitory antibody fragments to understand the structural underpinnings of αvß8 binding specificity and TGF-ß activation. Our studies reveal a mechanism of TGF-ß activation where mature TGF-ß signals within the confines of L-TGF-ß and the release and diffusion of TGF-ß are not required. The structural details of this mechanism provide a rational basis for therapeutic strategies to inhibit αvß8-mediated L-TGF-ß activation.
Assuntos
Microscopia Crioeletrônica/métodos , Integrinas/química , Integrinas/metabolismo , Proteínas de Ligação a TGF-beta Latente/química , Proteínas de Ligação a TGF-beta Latente/metabolismo , Fator de Crescimento Transformador beta1/química , Fator de Crescimento Transformador beta1/metabolismo , Animais , Anticorpos/imunologia , Sítios de Ligação , Brônquios/citologia , Células CHO , Cricetulus , Feminino , Humanos , Fragmentos Fab das Imunoglobulinas/imunologia , Integrinas/imunologia , Ativação Linfocitária , Masculino , Vison , Ligação Proteica , Conformação Proteica em alfa-Hélice , Domínios e Motivos de Interação entre Proteínas , Linfócitos T Reguladores/imunologiaRESUMO
The last few decades have seen striking changes in the field of food allergy. The prevalence of the disease has risen dramatically in many parts of the globe, and management of the condition has undergone major revision. While delayed introduction of common allergenic foods during infancy was advised for many years, the learning early about peanut allergy (LEAP) trial and other studies led to a major shift in infant feeding practices, with deliberate early introduction of these foods now recommended. Additionally, the Food and Drug Administration approved the first treatment for food allergy in 2020-a peanut oral immunotherapy (OIT) product that likely represents just the beginning of new immunotherapy-based and other treatments for food allergy. Our knowledge of the environmental and genetic factors contributing to the pathogenesis of food allergy has also undergone transformational advances. Here, we will discuss our efforts to improve the clinical care of patients with food allergy and our understanding of the immunological mechanisms contributing to this common disease.
RESUMO
TGFß signaling and the DNA damage response (DDR) are two cellular toolboxes with a strong impact on cancer biology. While TGFß as a pleiotropic cytokine affects essentially all hallmarks of cancer, the multifunctional DDR mostly orchestrates cell cycle progression, DNA repair, chromatin remodeling and cell death. One oncogenic effect of TGFß is the partial activation of epithelial-to-mesenchymal transition (EMT), conferring invasiveness, cellular plasticity and resistance to various noxae. Several reports show that both individual networks as well as their interface affect chemo-/radiotherapies. However, the underlying mechanisms remain poorly resolved. EMT often correlates with TGFß-induced slowing of proliferation, yet numerous studies demonstrate that particularly the co-activated EMT transcription factors counteract anti-proliferative signaling in a partially non-redundant manner. Collectively, evidence piled up over decades underscore a multifaceted, reciprocal inter-connection of TGFß signaling / EMT with the DDR / cell cycle progression, which we will discuss here. Altogether, we conclude that full cell cycle arrest is barely compatible with the propagation of oncogenic EMT traits and further propose that 'EMT-linked DDR plasticity' is a crucial, yet intricate facet of malignancy, decisively affecting metastasis formation and therapy resistance.
Assuntos
Neoplasias , Fator de Crescimento Transformador beta , Humanos , Fator de Crescimento Transformador beta/metabolismo , Transdução de Sinais , Fatores de Transcrição , Neoplasias/genética , Ciclo Celular/genética , Transição Epitelial-Mesenquimal/genéticaRESUMO
RECK is a candidate tumor suppressor gene isolated as a gene that induces flat reversion in a cell line transformed by the KRAS oncogene. Since RECK knockout mice die in utero, they are not suitable for studying the effects of RECK on tumor formation. In this study, we found an increased incidence of spontaneous pulmonary adenomas in mice with reduced RECK expression (RECK-Hypo mice). To evaluate the effects of RECK expressed by either tumor cells or host cells on tumor growth, we established a tumorigenic cell line (MKER) from the kidney of a C57BL/6 mouse and performed syngeneic transplantation experiments. Our results indicate that when RECK expression is low in host cells, transplanted MKER cells grow faster and kill the animal more rapidly. Since RECK is required for the formation of proper fibrillin fibers that serve as a tissue reservoir for precursors of TGFß-family cytokines, we assessed the levels of TGFß1 in the peripheral blood. We found a significant increase in TGFß1 in RECK-Hypo mice compared to wild-type mice. We also found that the proportion of FOXP3-positive regulatory T (Treg) cells among splenocytes was higher in RECK-Hypo mice compared to the control mice. Furthermore, the number of FOXP3-positive cells in spontaneous hematopoietic neoplasms in the lungs as well as tumors that formed after MKER transplantation was significantly higher in RECK-Hypo mice compared to the control mice. These findings indicate that RECK-mediated tumor suppression involves a non-cell-autonomous mechanism and that possible roles of TGFß1 and Treg cells in such a mechanism warrant further study.
RESUMO
Importin 8, encoded by IPO8, is a ubiquitously expressed member of the importin-ß protein family that translocates cargo molecules such as proteins, RNAs, and ribonucleoprotein complexes into the nucleus in a RanGTP-dependent manner. Current knowledge of the cargoes of importin 8 is limited, but TGF-ß signaling components such as SMAD1-4 have been suggested to be among them. Here, we report that bi-allelic loss-of-function variants in IPO8 cause a syndromic form of thoracic aortic aneurysm (TAA) with clinical overlap with Loeys-Dietz and Shprintzen-Goldberg syndromes. Seven individuals from six unrelated families showed a consistent phenotype with early-onset TAA, motor developmental delay, connective tissue findings, and craniofacial dysmorphic features. A C57BL/6N Ipo8 knockout mouse model recapitulates TAA development from 8-12 weeks onward in both sexes but most prominently shows ascending aorta dilatation with a propensity for dissection in males. Compliance assays suggest augmented passive stiffness of the ascending aorta in male Ipo8-/- mice throughout life. Immunohistological investigation of mutant aortic walls reveals elastic fiber disorganization and fragmentation along with a signature of increased TGF-ß signaling, as evidenced by nuclear pSmad2 accumulation. RT-qPCR assays of the aortic wall in male Ipo8-/- mice demonstrate decreased Smad6/7 and increased Mmp2 and Ccn2 (Ctgf) expression, reinforcing a role for dysregulation of the TGF-ß signaling pathway in TAA development. Because importin 8 is the most downstream TGF-ß-related effector implicated in TAA pathogenesis so far, it offers opportunities for future mechanistic studies and represents a candidate drug target for TAA.
Assuntos
Aneurisma da Aorta Torácica/etiologia , Mutação com Perda de Função , Perda de Heterozigosidade , Fenótipo , beta Carioferinas/genética , Adulto , Animais , Aneurisma da Aorta Torácica/metabolismo , Aneurisma da Aorta Torácica/patologia , Criança , Pré-Escolar , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Linhagem , Transdução de Sinais , Síndrome , Fator de Crescimento Transformador beta/genética , Fator de Crescimento Transformador beta/metabolismo , Adulto Jovem , beta Carioferinas/metabolismoRESUMO
The immortalized human renal proximal tubular epithelial cell line HK-2 is most commonly used to study renal cell physiology and human kidney diseases with tubulointerstitial fibrosis such as diabetic nephropathy, obstructive uropathy or allograft fibrosis. Epithelial-to-mesenchymal transition (EMT) is the main pathological process of tubulointerstitial fibrosis in vitro. Transforming growth factor-beta (TGF-ß) is a key inducer of EMT. Several pro-fibrotic gene expression differences have been observed in a TGF-ß-induced EMT model of HK-2 cells. However, growth conditions and medium formulations might greatly impact these differences. We investigated gene and protein expression of HK-2 cells cultured in six medium formulations. TGF-ß1 increased the expression of ACTA2, TGFB1, COL4A1, EGR2, VIM and CTGF genes while reducing PPARG in all medium formulations. Interestingly, TGF-ß1 treatment either increased or decreased EGR1, FN, IL6 and C3 gene expression, depending on medium formulations. The cell morphology was slightly affected, but immunoblots revealed TGFB1 and vimentin protein overexpression in all media. However, fibronectin expression as well as the nuclear translocation of EGR1 was medium dependent. In conclusion, our study demonstrates that, using the HK-2 in vitro model of EMT, the meticulous selection of appropriate cell culture medium formulation is essential to achieve reliable scientific results.
Assuntos
Nefropatias Diabéticas , Fator de Crescimento Transformador beta1 , Humanos , Fator de Crescimento Transformador beta1/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Transição Epitelial-Mesenquimal , Nefropatias Diabéticas/metabolismo , Fibrose , Técnicas de Cultura de Células , Células Epiteliais/metabolismoRESUMO
BACKGROUND: Extracellular vesicles (EVs) and their cargoes, including MicroRNAs (miRNAs) play a crucial role in cell-to-cell communication. We previously demonstrated the upregulation of bta-mir-148b in EVs from oviductal fluid of cyclic cows. This miRNA is linked to the TGF-ß pathway in the cell proliferation. Our aim was to verify whether miR-148b is taken up by embryos through gymnosis, validate its target genes, and investigate the effect of miR-148b supplementation on early embryo development and quality. METHODS: Zygotes were cultured in SOF + 0.3% BSA (Control) or supplemented with: 1 µM miR-148b mimics during: D1-D7 (miR148b) or D1-D4 (miR148b-OV: representing miRNA effect in the oviduct) or D4-D7 (miR148b-UT: representing miRNA effect in the uterus) or 1 µM control mimics was used during: D1-D7 (CMimic). Embryos at ≥ 16-cells and D7 blastocysts (BD7) were collected to examine the mRNA abundance of transcripts linked to the TGF-ß pathway (TGFBR2, SMAD1, SMAD2, SMAD3, SMAD5, BMPR2, RPS6KB1, POU5F1, NANOG), total cell number (TC), trophectoderm (TE), and inner cell mass (ICM) were also evaluated. One-way ANOVA was used for all analyses. RESULTS: We demonstrated that miR-148b can be taken up in both 16-cell embryos and BD7 by gymnosis, and we observed a decrease in SMAD5 mRNA, suggesting it's a potential target of miR-148b. Cleavage and blastocysts rates were not affected in any groups; however, supplementation of miR-148b mimics had a positive effect on TC, TE and ICM, with values of 136.4 ± 1.6, 92.5 ± 0.9, 43.9 ± 1.3 for miR148b and 135.3 ± 1.5, 92.6 ± 1.2, 42.7 ± 0.8, for miR148b-OV group. Furthermore, mRNA transcripts of SMAD1 and SMAD5 were decreased (P ≤ 0.001) in 16-cell embryos and BD7 from miR148b and miR148b-OV groups, while POU5F1 and NANOG were upregulated (P ≤ 0.001) in BD7 and TGFBR2 was only downregulated in 16-cell embryos. pSMAD1/5 levels were higher in the miR148b and miR148b-OV groups. CONCLUSIONS: Our findings suggest that supplementation of bta-miR-148b mimics during the entire culture period (D1 - D7) or from D1 - D4 improves embryo quality and influences the TGF-ß signaling pathway by altering the transcription of genes associated with cellular differentiation and proliferation. This highlights the importance of miR-148b on embryo quality and development.
Assuntos
Vesículas Extracelulares , MicroRNAs , Humanos , Feminino , Bovinos , Animais , Fator de Crescimento Transformador beta/metabolismo , Receptor do Fator de Crescimento Transformador beta Tipo II/metabolismo , MicroRNAs/genética , Oviductos/metabolismo , Vesículas Extracelulares/metabolismo , RNA Mensageiro/genéticaRESUMO
BACKGROUND: Osteoporosis is a genetic disease caused by the imbalance between osteoblast-led bone formation and osteoclast-induced bone resorption. However, further gene-related pathogenesis remains to be elucidated. METHODS: The aberrant expressed genes in osteoporosis was identified by analyzing the microarray profile GSE100609. Serum samples of patients with osteoporosis and normal group were collected, and the mRNA expression of candidate genes was detected by quantitative real-time polymerase chain reaction (qRT-PCR). The mouse cranial osteoblast MC3T3-E1 cells were treated with dexamethasone (DEX) to mimic osteoporosis in vitro. Alizarin Red staining and alkaline phosphatase (ALP) staining methods were combined to measure matrix mineralization deposition of MC3T3-E1 cells. Meanwhile, the expression of osteogenesis related genes including alkaline phosphatase (ALP), osteocalcin (OCN), osteopontin (OPN), Osterix, and bone morphogenetic protein 2 (BMP2) were evaluated by qRT-PCR and western blotting methods. Then the effects of candidate genes on regulating impede bone loss caused by ovariectomy (OVX) in mice were studied. RESULTS: Cyclin A1 (CCNA1) was found to be significantly upregulated in serum of osteoporosis patients and the osteoporosis model cells, which was in line with the bioinformatic analysis. The osteogenic differentiation ability of MC3T3-E1 cells was inhibited by DEX treatment, which was manifested by decreased Alizarin Red staining intensity, ALP staining intensity, and expression levels of ALP, OCN, OPN, Osterix, and BMP2. The effects of CCNA1 inhibition on regulating osteogenesis were opposite to that of DEX. Then, Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis demonstrated that genes negatively associated with CCNA1 were enriched in the TGF-beta signaling pathway. Inhibitor of TGF-beta signaling pathway partly reversed osteogenesis induced by suppressed CCNA1. Furthermore, suppressed CCNA1 relieved bone mass of OVX mice in vivo. CONCLUSION: Downregulation of CCNA1 could activate TGF-beta signaling pathway and promote bone formation, thus playing a role in treatment of osteoporosis.
Assuntos
Antraquinonas , Osteoporose , Fator de Crescimento Transformador beta , Animais , Feminino , Humanos , Camundongos , Fosfatase Alcalina/metabolismo , Diferenciação Celular , Ciclina A1/metabolismo , Osteoblastos/metabolismo , Osteogênese , Osteoporose/induzido quimicamente , Fator de Crescimento Transformador beta/metabolismo , Fatores de Crescimento Transformadores/efeitos adversos , Fatores de Crescimento Transformadores/metabolismoRESUMO
A role for substance P has been proposed in musculoskeletal fibrosis, with effects mediated through transforming growth factor beta (TGFß). We examined the in vitro effects of substance P on proliferation, collagen secretion, and collagen deposition in rat primary dermal fibroblasts cultured in medium containing 10% fetal bovine serum, with or without TGFß. In six-day cultures, substance P increased cell proliferation at concentrations from 0.0002 to 100 nM. TGFß increased proliferation at concentrations from 0.0002 to 2 pg/mL, although higher concentrations inhibited proliferation. Substance P treatment alone at concentrations of 100, 0.2, and 0.00002 nM did not increase collagen deposition per cell, yet when combined with TGFß (5 ng/mL), increased collagen deposition compared to TGFß treatment alone. Substance P treatment (100 nM) also increased smooth muscle actin (SMA) expression at 72 h of culture at a level similar to 5 ng/mL of TGFß; only TGFß increased SMA at 48 h of culture. Thus, substance P may play a role in potentiating matrix deposition in vivo when combined with TGFß, although this potentiation may be dependent on the concentration of each factor. Treatments targeting substance P may be a viable strategy for treating fibrosis where both substance P and TGFß play roles.
Assuntos
Substância P , Fator de Crescimento Transformador beta , Ratos , Animais , Fator de Crescimento Transformador beta/metabolismo , Substância P/farmacologia , Substância P/metabolismo , Células Cultivadas , Fibroblastos/metabolismo , Colágeno/metabolismo , Fibrose , Fator de Crescimento Transformador beta1/metabolismoRESUMO
Alveolar rhabdomyosarcoma (ARMS), an invasive subtype of rhabdomyosarcoma (RMS), is associated with chromosomal translocation events resulting in one of two oncogenic fusion genes, PAX3-FOXO1 or PAX7-FOXO1. ARMS patients exhibit an overexpression of the pleiotropic cytokine transforming growth factor beta (TGF-ß). This overexpression of TGF-ß1 causes an increased expression of a downstream transcription factor called SNAIL, which promotes epithelial to mesenchymal transition (EMT). Overexpression of TGF-ß also inhibits myogenic differentiation, making ARMS patients highly resistant to chemotherapy. In this review, we first describe different types of RMS and then focus on ARMS and the impact of TGF-ß in this tumor type. We next highlight current chemotherapy strategies, including a combination of the FDA-approved drugs vincristine, actinomycin D, and cyclophosphamide (VAC); cabozantinib; bortezomib; vinorelbine; AZD 1775; and cisplatin. Lastly, we discuss chemotherapy agents that target the differentiation of tumor cells in ARMS, which include all-trans retinoic acid (ATRA) and 5-Azacytidine. Improving our understanding of the role of signaling pathways, such as TGF-ß1, in the development of ARMS tumor cells differentiation will help inform more tailored drug administration in the future.
Assuntos
Rabdomiossarcoma Alveolar , Rabdomiossarcoma , Humanos , Rabdomiossarcoma Alveolar/genética , Rabdomiossarcoma Alveolar/metabolismo , Rabdomiossarcoma Alveolar/patologia , Fator de Crescimento Transformador beta , Fator de Crescimento Transformador beta1 , Fatores de Transcrição Box Pareados/genética , Transição Epitelial-Mesenquimal , Rabdomiossarcoma/genética , Proteínas de Fusão Oncogênica/genéticaRESUMO
Cnidarians are fascinating creatures at the base of metazoan evolution possessing an almost unlimited regeneration capacity that has attracted the interest of researchers, from Abraham Trembley's discovery of regeneration to the present. They share a simple body plan and a high morphogenetic plasticity that has led to a broad spectrum of life cycles. With molecular genomics it became unequivocally clear that Cnidaria are the sister group of the Bilateria and how similar their molecular toolkit is to that of more complex animals. This has renewed interest in these simple animals, which have had an important role in the establishment of fundamental concepts for developmental biologists from the beginning. This review focuses on our current understanding of signaling centers (organizers) and morphogenetic gradients in cnidarians and how they relate to the emergence of the bilaterian body axes. The data are largely based on the cnidarian models Hydra and Nematostella and are supported by new studies on forms with a complete cnidarian life cycle, such as the medusozoans Aurelia and Clytia. Molecular studies on cnidarian development have revealed the existence of an ancient Wnt signaling center at the site of gastrulation, which was instrumental for the formation of a primary body axis and can be traced back to the common ancestor of bilaterian and non-bilaterian animals. New molecular data also suggest that the molecular vectors for the dorso-ventral and left-right body axis in bilaterians, Bmp and Nodal signaling, respectively, were already present but had different fates in the two clades. The close link of developmental processes in bilaterians and cnidarians but also their distinct differences make cnidarians an indispensable model for tackling fundamental questions in developmental biology from patterning, regeneration and other recent molecular approaches to theoretical concepts.
Assuntos
Padronização Corporal , Anêmonas-do-Mar , Animais , Padronização Corporal/genética , Biologia do Desenvolvimento , Evolução Molecular , Via de Sinalização Wnt/genéticaRESUMO
KIN17 DNA and RNA binding protein (Kin17) is involved in the regulation of tumorigenesis of diverse human cancers. However, its role in the cancer progression and metastasis in hepatocellular carcinoma (HCC) remains largely unknown. Bioinformatics and immunohistochemistry staining were used to investigate the expression pattern of KIN17 and its prognostic value in HCC patients. The transwell, wound-healing assay was employed to determine the effects of KIN17 on migration and invasion of HCC cells in vitro. The tail veins model was employed to determine the effects of KIN17 on lung metastasis in vivo. The biological mechanisms involved in cell migration and invasion regulated by KIN17 were determined with Western blot analysis method. KIN17 expression was significantly increased in HCC tissues compared with adjacent normal tissues, with particularly higher in portal vein tumor thrombus and intrahepatic metastasis tissues. Patients with higher KIN17 expression experienced poor overall and disease free survival. KIN17 knockdown in HuH7 and HepG2 cells significantly reduced cell migration and invasion abilities, whereas its overexpression promoted migration and invasion in MHCC-97L and HepG2 cells in vitro and in vivo. In HuH7 and HepG2 cells, KIN17 knockdown inhibited the TGF-ß/Smad2 pathway. In contrast, KIN17 overexpression stimulated TGF-ß/Smad2 pathway in MHCC-97L and HepG2 cells, along with the genes involved in the epithelial-mesenchymal transition. These findings suggest that KIN17 promotes migration and invasion in HCC cells by stimulating the TGF-ß/Smad2 pathway. KIN17 could be a promising prognostic biomarker, as well as a potential therapeutic target in HCC.
Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , Humanos , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Movimento Celular/genética , Transição Epitelial-Mesenquimal/genética , Regulação Neoplásica da Expressão Gênica , Neoplasias Hepáticas/patologia , Proteína Smad2/genética , Proteína Smad2/metabolismo , Fator de Crescimento Transformador beta/metabolismoRESUMO
BACKGROUND: Limosilactobacillusmucosae (LM) exerts anti-inflammatory and health-promoting effects. However, its role in the modulation of gut serotonin or 5-hydroxytryptamine (5-HT) metabolism and 5-HT receptors (HTRs) in inflammation requires further investigation. OBJECTIVES: We compared LM with Lactobacillus amylovorus (LA) for the regulation of 5-HT, HTRs, inflammatory mediators, and their correlations in the colon of mice with experimental colitis. METHODS: Male C57BL/6 mice were randomly assigned to 6 groups: control (Con), LM, LA, dextran sodium sulfate (DSS), and DSS with pre-administration of LM (+LM) or LA (+LA). After 7 d of DSS treatment, mice were killed to analyze the expression of inflammatory mediators, HTRs, and concentrations of 5-HT and microbial metabolites in the colon. RESULTS: LM was more effective than LA in alleviating DSS-induced colonic inflammation. Compared with mice in the DSS group, mice receiving DSS + LM or DSS + LA treatment had lower (P < 0.05) colonic mRNA expression of proinflammatory cytokines. DSS + LM treatment had lower mRNA expression of Il1b, Tnfa, and Ccl3, an abundance of p-STAT3, and greater expression of Tgfb2 and Htr4 in the colon (P < 0.05). The expression of inflammatory mediators (including Tgfb-1) was positively correlated (P < 0.05) with 5-HT and Htr2a and negatively correlated (P < 0.05) with Htr4. However, the expression of Tgfb-2 showed reversed correlations with the 5-HT and HTRs described above. Patterns for these correlations were different for LM and LA. Mice receiving the DSS + LM treatment had greater (P < 0.05) concentrations of acetate and valerate and lower (P < 0.05) concentrations of indole-3-acetic acid in the cecal and colonic contents. CONCLUSIONS: LM showed greater efficacy than LA in alleviating DSS-induced colonic inflammation. The coordinated regulation of transforming growth factor-ß subtypes and serotonin receptors in the colon may be one of the most important mechanisms underlying the probiotic effects of lactobacilli in gut inflammation.
Assuntos
Colite , Serotonina , Masculino , Animais , Camundongos , Serotonina/metabolismo , Lactobacillus acidophilus/metabolismo , Regulação para Cima , Camundongos Endogâmicos C57BL , Colite/induzido quimicamente , Colite/prevenção & controle , Colite/metabolismo , Colo/metabolismo , Inflamação/metabolismo , RNA Mensageiro/metabolismo , Fatores de Crescimento Transformadores/efeitos adversos , Fatores de Crescimento Transformadores/metabolismo , Sulfato de Dextrana/toxicidade , Modelos Animais de DoençasRESUMO
Clopidogrel, a P2Y12 inhibitor, is a novel anti-fibrosis agent for chronic kidney disease (CKD), but its mechanisms remain unclear, which we investigated by silencing P2Y12 or treating unilateral ureteral obstruction (UUO) in LysM-Cre/Rosa Tomato mice with clopidogrel in vivo and in vitro. We found that P2Y12 was significantly increased and correlated with progressive renal fibrosis in CKD patients and UUO mice. Phenotypically, up to 82% of P2Y12-expressing cells within the fibrosing kidney were of macrophage origin, identified by co-expressing CD68/F4/80 antigens or a macrophage-lineage-tracing marker Tomato. Unexpectedly, more than 90% of P2Y12-expressing macrophages were undergoing macrophage-to-myofibroblast transition (MMT) by co-expressing alpha smooth muscle actin (α-SMA), which was also confirmed by single-cell RNA sequencing. Functionally, clopidogrel improved the decline rate of the estimated glomerular filtration rate (eGFR) in patients with CKD and significantly inhibited renal fibrosis in UUO mice. Mechanistically, P2Y12 expression was induced by transforming growth factor ß1 (TGF-ß1) and promoted MMT via the Smad3-dependent mechanism. Thus, silencing or pharmacological inhibition of P2Y12 was capable of inhibiting TGF-ß/Smad3-mediated MMT and progressive renal fibrosis in vivo and in vitro. In conclusion, P2Y12 is highly expressed by macrophages in fibrosing kidneys and mediates renal fibrosis by promoting MMT via TGF-ß/Smad3 signaling. Thus, P2Y12 inhibitor maybe a novel and effective anti-fibrosis agent for CKD.
Assuntos
Nefropatias , Insuficiência Renal Crônica , Obstrução Ureteral , Animais , Clopidogrel/metabolismo , Clopidogrel/farmacologia , Clopidogrel/uso terapêutico , Fibrose , Rim , Nefropatias/etiologia , Nefropatias/genética , Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Miofibroblastos/metabolismo , Insuficiência Renal Crônica/metabolismo , Insuficiência Renal Crônica/patologia , Transdução de Sinais , Proteína Smad3/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Fator de Crescimento Transformador beta1/metabolismo , Obstrução Ureteral/complicações , Obstrução Ureteral/tratamento farmacológico , Obstrução Ureteral/genéticaRESUMO
Dupuytren disease is a benign, progressive fibroproliferative disorder of the hands. To date, only one pharmacotherapy (clostridial collagenase) has been approved for use in Dupuytren disease. There is a great need for additional nonsurgical methods that can be used to either avoid the risks of invasive treatments or help minimize recurrence rates following treatment. A number of nonsurgical modalities have been discussed in the past and continue to appear in discussions among hand surgeons, despite highly variable and often poor or no long-term clinical data. This article reviews many of the pharmacotherapies discussed in the treatment of Dupuytren disease and novel therapies used in inflammation and fibrosis that offer potential treatment options.
Assuntos
Contratura de Dupuytren , Humanos , Contratura de Dupuytren/cirurgia , Colagenase Microbiana/uso terapêutico , Resultado do Tratamento , Injeções Intralesionais , Clostridium histolyticumRESUMO
Fibrodysplasia ossificans progressiva (FOP) is a catastrophic, ultra-rare disease of heterotopic ossification caused by genetic defects in the ACVR1 gene. The mutant ACVR1 receptor, when triggered by an inflammatory process, leads to heterotopic ossification of the muscles and ligaments. Activin A has been discovered as the main osteogenic ligand of the FOP ACVR1 receptor. However, the source of Activin A itself and the trigger of its production in FOP individuals have remained elusive. We used primary dermal fibroblasts from five FOP patients to investigate Activin A production and how this is influenced by inflammatory cytokines in FOP. FOP fibroblasts showed elevated Activin A production compared to healthy controls, both in standard culture and osteogenic transdifferentiation conditions. We discovered TGFß1 to be an FOP-specific stimulant of Activin A, shown by the upregulation of the INHBA gene and protein expression. Activin A and TGFß1 were both induced by BMP4 in FOP and control fibroblasts. Treatment with TNFα and IL6 produced negligible levels of Activin A and TGFß1 in both cell groups. We present for the first time TGFß1 as a triggering factor of Activin A production in FOP. As TGFß1 can promote the induction of the main driver of FOP, TGFß1 could also be considered a possible therapeutic target in FOP treatment.
Assuntos
Miosite Ossificante , Ossificação Heterotópica , Humanos , Miosite Ossificante/genética , Miosite Ossificante/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Transdução de Sinais/genética , Ossificação Heterotópica/genética , Fibroblastos/metabolismo , Receptores de Ativinas Tipo I/genética , Receptores de Ativinas Tipo I/metabolismo , MutaçãoRESUMO
Up to 50% of systemic lupus erythematosus (SLE) patients world-wide develop lupus nephritis (LN). In low to middle income countries and in particular in sub-Saharan Africa, where SLE is prevalent with a more aggressive course, LN and end stage renal disease is a major cause of mortality. While developed countries have the funding to invest in SLE and LN research, patients of African descent are often underrepresented in clinical trials. Thus, the complex influence of ethnicity and genetic background on outcome of LN and SLE as a whole, is not fully understood. Several pathophysiological mechanisms including major role players driving LN have been identified. A large body of literature suggest that prevention of fibrosis-which contributes to chronicity of LN-may significantly improve long-term prognosis. Bone morphogenetic protein-7 (BMP-7) was first identified as a therapeutic option in this context decades ago and evidence of its benefit in various conditions, including LN, is ever-increasing. Despite these facts, BMP-7 is not being implemented as therapy in the context of renal disease. With this review, we briefly summarise current understanding of LN pathology and discuss the evidence in support of therapeutic potential of BMP-7 in this context. Lastly, we address the obstacles that need to be overcome, before BMP-7 may become available as LN treatment.
Assuntos
Falência Renal Crônica , Lúpus Eritematoso Sistêmico , Nefrite Lúpica , Humanos , Nefrite Lúpica/tratamento farmacológico , Nefrite Lúpica/patologia , Proteína Morfogenética Óssea 7/uso terapêutico , PrognósticoRESUMO
BACKGROUND: The Transforming Growth Factor ß (TGFß) family is a group of related proteins that signal through a type I and type II receptors. Betaglycan, also known as the type III receptor (Tgfbr3), is a coreceptor for various ligands of the TGFß family that participates in heart, liver and kidney development as revealed by the tgfbr3-null mouse, as well as in angiogenesis as revealed by Tgfbr3 downregulation in morphant zebrafish. RESULTS: Here, we present CRISPR/Cas9-derived zebrafish Tgfbr3-null mutants, which exhibited unaltered embryonic angiogenesis and developed into fertile adults. One reproducible phenotype displayed by these Tgfbr3-null mutants is delayed chordacentra mineralization, which nonetheless does not result in vertebral abnormalities in the adult fishes. We also report that the canonical TGFß signaling pathway is needed for proper chordacentra mineralization and that Tgfbr3 absence decreases this signal in the notochordal cells responsible for this process. CONCLUSION: Betaglycan's "ligand presentation" function contributes to the optimal TGFß signaling required for zebrafish chordacentra mineralization.
Assuntos
Receptores de Fatores de Crescimento Transformadores beta , Peixe-Zebra , Animais , Camundongos , Proteoglicanas/genética , Receptores de Fatores de Crescimento Transformadores beta/genética , Receptores de Fatores de Crescimento Transformadores beta/metabolismo , Transdução de Sinais/genética , Fator de Crescimento Transformador beta/genética , Fator de Crescimento Transformador beta/metabolismo , Peixe-Zebra/genética , Peixe-Zebra/metabolismoRESUMO
Transforming growth factor (TGF)-ß uses several intracellular signaling pathways besides canonical ALK5-Smad2/3 signaling to regulate a diverse array of cellular functions. Several of these so-called non-canonical (non-Smad2/3) pathways have been implicated in the pathogenesis of fibrosis and may therefore represent targets for therapeutic intervention. This review summarizes our current knowledge on the mechanisms of non-canonical TGF-ß signaling in fibrosis, the potential molecular targets and the use of agonists/antagonists for therapeutic intervention.
Assuntos
Fibrose/metabolismo , Transdução de Sinais , Fator de Crescimento Transformador beta/metabolismo , Animais , HumanosRESUMO
Systemic sclerosis (SSc) is a highly challenging chronic condition that is dominated by the pathogenetic triad of vascular damage, immune dysregulation/autoimmunity and fibrosis in multiple organs. A hallmark of SSc is the remarkable degree of molecular and phenotypic disease heterogeneity, which surpasses that of other complex rheumatic diseases. Disease trajectories in SSc are unpredictable and variable from patient to patient. Disease-modifying therapies for SSc are lacking, long-term morbidity is considerable and mortality remains unacceptably high. Currently-used empirical approaches to disease modification have modest and variable clinical efficacy and impact on survival, are expensive and frequently associated with unfavorable side effects, and none can be considered curative. However, research during the past several years is yielding significant advances with therapeutic potential. In particular, the application of unbiased omics-based discovery technologies to large and well-characterized SSc patient cohorts, coupled with hypothesis-testing experimental research using a variety of model systems is revealing new insights into SSc that allow formulation of a more nuanced appreciation of disease heterogeneity, and a deepening understanding of pathogenesis. Indeed, we are now presented with numerous novel and rationally-based strategies for targeted SSc therapy, several of which are currently, or expected to be shortly, undergoing clinical evaluation. In this review, we discuss promising novel therapeutic targets and rationally-based approaches to disease modification that have the potential to improve long-term outcomes in SSc.