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Nature Communications May 2024ARID1B haploinsufficiency in humans causes Coffin-Siris syndrome, associated with developmental delay, facial dysmorphism, and intellectual disability. The role of...
ARID1B haploinsufficiency in humans causes Coffin-Siris syndrome, associated with developmental delay, facial dysmorphism, and intellectual disability. The role of ARID1B has been widely studied in neuronal development, but whether it also regulates stem cells remains unknown. Here, we employ scRNA-seq and scATAC-seq to dissect the regulatory functions and mechanisms of ARID1B within mesenchymal stem cells (MSCs) using the mouse incisor model. We reveal that loss of Arid1b in the GLI1+ MSC lineage disturbs MSCs' quiescence and leads to their proliferation due to the ectopic activation of non-canonical Activin signaling via p-ERK. Furthermore, loss of Arid1b upregulates Bcl11b, which encodes a BAF complex subunit that modulates non-canonical Activin signaling by directly regulating the expression of activin A subunit, Inhba. Reduction of Bcl11b or non-canonical Activin signaling restores the MSC population in Arid1b mutant mice. Notably, we have identified that ARID1B suppresses Bcl11b expression via specific binding to its third intron, unveiling the direct inter-regulatory interactions among BAF subunits in MSCs. Our results demonstrate the vital role of ARID1B as an epigenetic modifier in maintaining MSC homeostasis and reveal its intricate mechanistic regulatory network in vivo, providing novel insights into the linkage between chromatin remodeling and stem cell fate determination.
Topics: Animals; Mesenchymal Stem Cells; Signal Transduction; Mice; DNA-Binding Proteins; Repressor Proteins; Transcription Factors; Cell Proliferation; Activins; Tumor Suppressor Proteins; Humans; Zinc Finger Protein GLI1
PubMed: 38816354
DOI: 10.1038/s41467-024-48285-2 -
Science Translational Medicine May 2024Fibrodysplasia ossificans progressiva (FOP) is a rare genetic disease driven by gain-of-function variants in activin receptor-like kinase 2 (ALK2), the most common...
Fibrodysplasia ossificans progressiva (FOP) is a rare genetic disease driven by gain-of-function variants in activin receptor-like kinase 2 (ALK2), the most common variant being ALK2. In FOP, ALK2 variants display increased and dysregulated signaling through the bone morphogenetic protein (BMP) pathway resulting in progressive and permanent replacement of skeletal muscle and connective tissues with heterotopic bone, ultimately leading to severe debilitation and premature death. Here, we describe the discovery of BLU-782 (IPN60130), a small-molecule ALK2 inhibitor developed for the treatment of FOP. A small-molecule library was screened in a biochemical ALK2 binding assay to identify potent ALK2 binding compounds. Iterative rounds of structure-guided drug design were used to optimize compounds for ALK2 binding, ALK2 selectivity, and other desirable pharmacokinetic properties. BLU-782 preferentially bound to ALK2 with high affinity, inhibiting signaling from ALK2 and other rare FOP variants in cells in vitro without affecting signaling of closely related homologs ALK1, ALK3, and ALK6. In vivo efficacy of BLU-782 was demonstrated using a conditional knock-in ALK2 mouse model, where prophylactic oral dosing reduced edema and prevented cartilage and heterotopic ossification (HO) in both muscle and bone injury models. BLU-782 treatment preserved the normal muscle-healing response in ALK2 mice. Delayed dosing revealed a short 2-day window after injury when BLU-782 treatment prevented HO in ALK2 mice, but dosing delays of 4 days or longer abrogated HO prevention. Together, these data suggest that BLU-782 may be a candidate for prevention of HO in FOP.
Topics: Animals; Myositis Ossificans; Ossification, Heterotopic; Disease Models, Animal; Mice; Humans; Activin Receptors, Type II; Activin Receptors, Type I; Signal Transduction
PubMed: 38809966
DOI: 10.1126/scitranslmed.abp8334 -
Journal of Biomedical Research May 2024The main pathogenic factor leading to cardiac remodeling and heart failure is myocardial fibrosis. Recent research indicates that microRNAs are essential for the...
The main pathogenic factor leading to cardiac remodeling and heart failure is myocardial fibrosis. Recent research indicates that microRNAs are essential for the progress of cardiac fibrosis. Myocardial fibrosis is considered to be alleviated through the bone morphogenetic protein and activin membrane-bound inhibitor (BAMBI), which does this by blocking the transforming growth factor β1 (TGF-β1) signaling pathway. Here, this study sought to elucidate the post-transcriptional regulation of miR-19a-3p on BAMBI and its role in TGF-β1-induced cardiac fibroblast activation. Transverse aortic constriction (TAC) caused both myocardial interstitial and perivascular collagen deposition. RT-PCR showed that miR-19a-3p was upregulated in the myocardial tissue of cardiac fibrosis, and TGF-β1 induced an increase of miR-19a-3p expression in cardiac fibroblasts. The dual-luciferase reporter test and qRT-PCR confirmed that miR-19a-3p directly combined with BAMBI mRNA 3'UTR, thus reduced BAMBI expression, which diminished the capability of BAMBI to inhibit TGF-β1. Furthermore, miR-19a-3p mimic increased the activation of TGF-β1/SMAD2/3 pathway signaling, which supported cardiac fibroblast activation, which blocked by overexpression of BAMBI. These findings imply that miR-19a-3p enhances the activation of TGF-β1/SMAD2/3 by inhibiting BAMBI, further boosting the activation of cardiac fibroblasts, and may thus offer a novel strategy to tackling myocardial fibrosis.
PubMed: 38807415
DOI: 10.7555/JBR.37.20230313 -
Kidney Diseases (Basel, Switzerland) Jun 2024The increasing prevalence of kidney diseases has become a significant public health issue, with a global prevalence exceeding 10%. In order to accurately identify... (Review)
Review
BACKGROUND
The increasing prevalence of kidney diseases has become a significant public health issue, with a global prevalence exceeding 10%. In order to accurately identify biochemical changes and treatment outcomes associated with kidney diseases, novel methods targeting specific genes have been discovered. Among these genes, leucine-rich α-2 glycoprotein 1 (LRG1) has been identified to function as a multifunctional pathogenic signaling molecule in multiple diseases, including kidney diseases. This study aims to provide a comprehensive overview of the current evidence regarding the roles of LRG1 in different types of kidney diseases.
SUMMARY
Based on a comprehensive review, it was found that LRG1 was upregulated in the urine, serum, or renal tissues of patients or experimental animal models with multiple kidney diseases, such as diabetic nephropathy, kidney injury, IgA nephropathy, chronic kidney diseases, clear cell renal cell carcinoma, end-stage renal disease, canine leishmaniosis-induced kidney disease, kidney fibrosis, and aristolochic acid nephropathy. Mechanistically, the role of LRG1 in kidney diseases is believed to be detrimental, potentially through its regulation of various genes and signaling cascades, i.e., fibronectin 1, GPR56, vascular endothelial growth factor (VEGF), VEGFR-2, death receptor 5, GDF15, HIF-1α, SPP1, activin receptor-like kinase 1-Smad1/5/8, NLRP3-IL-1b, and transforming growth factor β pathway.
KEY MESSAGES
Further research is needed to fully comprehend the molecular mechanisms by which LRG1 contributes to the pathogenesis and pathophysiology of kidney diseases. It is anticipated that targeted treatments focusing on LRG1 will be utilized in clinical trials and implemented in clinical practice in the future.
PubMed: 38799248
DOI: 10.1159/000538443 -
Vascular Pharmacology Jun 2024Bone morphogenetic protein-9 (BMP9) is critical for bone morphogenetic protein receptor type-2 (BMPR2) signalling in pulmonary vascular endothelial cells. Furthermore,...
AIMS
Bone morphogenetic protein-9 (BMP9) is critical for bone morphogenetic protein receptor type-2 (BMPR2) signalling in pulmonary vascular endothelial cells. Furthermore, human genetics studies support the central role of disrupted BMPR2 mediated BMP9 signalling in vascular endothelial cells in the initiation of pulmonary arterial hypertension (PAH). In addition, loss-of-function mutations in BMP9 have been identified in PAH patients. BMP9 is considered to play an important role in vascular homeostasis and quiescence.
METHODS AND RESULTS
We identified a novel BMP9 target as the class-3 semaphorin, SEMA3G. Although originally identified as playing a role in neuronal development, class-3 semaphorins may have important roles in endothelial function. Here we show that BMP9 transcriptional regulation of SEMA3G occurs via ALK1 and the canonical Smad pathway, requiring both Smad1 and Smad5. Knockdown studies demonstrated redundancy between type-2 receptors in that BMPR2 and ACTR2A were compensatory. Increased SEMA3G expression by BMP9 was found to be regulated by the transcription factor, SOX17. Moreover, we observed that SEMA3G regulates VEGF signalling by inhibiting VEGFR2 phosphorylation and that VEGF, in contrast to BMP9, negatively regulated SEMA3G transcription. Functional endothelial cell assays of VEGF-mediated migration and network formation revealed that BMP9 inhibition of VEGF was abrogated by SEMA3G knockdown. Conversely, treatment with recombinant SEMA3G partially mimicked the inhibitory action of BMP9 in these assays.
CONCLUSIONS
This study provides further evidence for the anti-angiogenic role of BMP9 in microvascular endothelial cells and these functions are mediated at least in part via SOX17 and SEMA3G induction.
Topics: Humans; Cell Movement; Semaphorins; Growth Differentiation Factor 2; Endothelial Cells; Signal Transduction; Vascular Endothelial Growth Factor A; Smad5 Protein; Activin Receptors, Type I; Bone Morphogenetic Protein Receptors, Type II; Smad1 Protein; Lung; Neovascularization, Physiologic; Cells, Cultured
PubMed: 38795838
DOI: 10.1016/j.vph.2024.107381 -
The Canadian Journal of Cardiology Mar 2024Preeclampsia remains a major cause of maternal and fetal adverse outcomes in pregnancy; however, accurate and universally acceptable predictive tools remain elusive. We...
BACKGROUND
Preeclampsia remains a major cause of maternal and fetal adverse outcomes in pregnancy; however, accurate and universally acceptable predictive tools remain elusive. We investigated whether a panel of biomarkers could improve risk prediction for preeclampsia when measured at various pregnancy time points.
METHODS
In this prospective cohort study, 192 women with first-trimester high-risk singleton pregnancies were consecutively recruited from tertiary obstetrics clinics in Montréal, Canada. Clinical information (height, pre-pregnancy weight, personal and family medical history, medication use) was collected at baseline. Blood pressure was measured and blood samples collected at each trimester to quantify soluble Fms-like tyrosine kinase 1 (sFlt-1), placental growth factor (PlGF), pregnancy-associated plasma protein A2 (PAPP-A2), PAPP-A, activin A, inhibin A, follistatin, and glycosylated fibronectin. A random-effects hierarchic logistic regression model was used to relate change in biomarker levels to incidence of preeclampsia.
RESULTS
When added to a clinical model composed of maternal age, pre-pregnancy body mass index, race, and mean arterial pressure, a positive third-trimester result for both PAPP-A2 and activin A had a better positive predictive value than the sFlt-1:PlGF ratio added to the clinical model (91.67% [95% confidence interval (CI) 78.57%-100%] vs 66.67% [57.14%-100%]), while maintaining a comparable high negative predictive value (97.69% [95% CI 95.34%-100%] vs 96.00% [92.19%-99.21%]).
CONCLUSIONS
Whereas the third-trimester sFlt-1:PlGF ratio can predict short-term absence of preeclampsia, PAPP-A2 and activin A had both high positive and negative predictive values and therefore could serve as biomarkers to predict the occurrence (and absence) of preeclampsia; these findings will be validated in future studies.
Topics: Humans; Female; Pre-Eclampsia; Pregnancy; Pregnancy-Associated Plasma Protein-A; Biomarkers; Activins; Adult; Placenta Growth Factor; Prospective Studies; Vascular Endothelial Growth Factor Receptor-1; Predictive Value of Tests; Pregnancy Trimester, First
PubMed: 38787345
DOI: 10.1016/j.cjca.2023.10.017 -
Molecular Metabolism Dec 2023The liver is a central regulator of energy metabolism exerting its influence both through intrinsic processing of substrates such as glucose and fatty acid as well as by...
OBJECTIVE
The liver is a central regulator of energy metabolism exerting its influence both through intrinsic processing of substrates such as glucose and fatty acid as well as by secreting endocrine factors, known as hepatokines, which influence metabolism in peripheral tissues. Human genome wide association studies indicate that a predicted loss-of-function variant in the Inhibin βE gene (INHBE), encoding the putative hepatokine Activin E, is associated with reduced abdominal fat mass and cardiometabolic disease risk. However, the regulation of hepatic Activin E and the influence of Activin E on adiposity and metabolic disease are not well understood. Here, we examine the relationship between hepatic Activin E and adipose metabolism, testing the hypothesis that Activin E functions as part of a liver-adipose, inter-organ feedback loop to suppress adipose tissue lipolysis in response to elevated serum fatty acids and hepatic fatty acid exposure.
METHODS
The relationship between hepatic Activin E and non-esterified fatty acids (NEFA) released from adipose lipolysis was assessed in vivo using fasted CL 316,243 treated mice and in vitro using Huh7 hepatocytes treated with fatty acids. The influence of Activin E on adipose lipolysis was examined using a combination of Inhbe knockout mice, a mouse model of hepatocyte-specific overexpression of Activin E, and mouse brown adipocytes treated with Activin E enriched media.
RESULTS
Increasing hepatocyte NEFA exposure in vivo by inducing adipose lipolysis through fasting or CL 316,243 treatment increased hepatic Inhbe expression. Similarly, incubation of Huh7 human hepatocytes with fatty acids increased expression of INHBE. Genetic ablation of Inhbe in mice increased fasting circulating NEFA and hepatic triglyceride accumulation. Treatment of mouse brown adipocytes with Activin E conditioned media and overexpression of Activin E in mice suppressed adipose lipolysis and reduced serum FFA levels, respectively. The suppressive effects of Activin E on lipolysis were lost in CRISPR-mediated ALK7 deficient cells and ALK7 kinase deficient mice. Disruption of the Activin E-ALK7 signaling axis in Inhbe KO mice reduced adiposity upon HFD feeding, but caused hepatic steatosis and insulin resistance.
CONCLUSIONS
Taken together, our data suggest that Activin E functions as part of a liver-adipose feedback loop, such that in response to increased serum free fatty acids and elevated hepatic triglyceride, Activin E is released from hepatocytes and signals in adipose through ALK7 to suppress lipolysis, thereby reducing free fatty acid efflux to the liver and preventing excessive hepatic lipid accumulation. We find that disrupting this Activin E-ALK7 inter-organ communication network by ablation of Inhbe in mice increases lipolysis and reduces adiposity, but results in elevated hepatic triglyceride and impaired insulin sensitivity. These results highlight the liver-adipose, Activin E-ALK7 signaling axis as a critical regulator of metabolic homeostasis.
Topics: Animals; Lipolysis; Mice; Liver; Adipose Tissue; Humans; Male; Inhibin-beta Subunits; Fatty Acids; Activins; Mice, Inbred C57BL; Hepatocytes; Fatty Acids, Nonesterified; Mice, Knockout; Adiposity
PubMed: 38787338
DOI: 10.1016/j.molmet.2023.101830 -
Human Reproduction (Oxford, England) May 2024Does the chemokine/chemokine receptor axis, involved in immune cell trafficking, contribute to the pathology of testicular inflammation and how does activin A modulate...
STUDY QUESTION
Does the chemokine/chemokine receptor axis, involved in immune cell trafficking, contribute to the pathology of testicular inflammation and how does activin A modulate this network?
SUMMARY ANSWER
Testicular chemokines and their receptors (especially those essential for trafficking of monocytes) are elevated in orchitis, and activin A modulates the expression of the chemokine/chemokine receptor network to promote monocyte/macrophage and T cell infiltration into the testes, causing extensive tissue damage.
WHAT IS KNOWN ALREADY
The levels of CC motif chemokine receptor (CCR)2 and its ligand CC motif chemokine ligand (CCL)2 are increased in experimental autoimmune orchitis (EAO) compared with healthy testes, and mice deficient in CCR2 are protected from EAO-induced tissue damage. Activin A induces CCR2 expression in macrophages, promoting their migration. Moreover, there is a positive correlation between testicular activin A concentration and the severity of autoimmune orchitis. Inhibition of activin A activity by overexpression of follistatin (FST) reduces EAO-induced testicular damage.
STUDY DESIGN, SIZE, DURATION
EAO was induced in 10-12-week-old male C57BL/6J (wild-type; WT) and B6.129P2-Ccr2tm1Mae/tm1Mae (Ccr2-/-) mice (n = 6). Adjuvant (n = 6) and untreated (n = 6) age-matched control mice were also included. Testes were collected at 50 days after the first immunization with testicular homogenate in complete Freund's adjuvant. In another experimental setup, WT mice were injected with a non-replicative recombinant adeno-associated viral vector carrying a FST315-expressing gene cassette (rAAV-FST315; n = 7-9) or an empty control vector (n = 5) 30 days prior to EAO induction. Appropriate adjuvant (n = 4-5) and untreated (n = 4-6) controls were also examined. Furthermore, human testicular biopsies exhibiting focal leukocytic infiltration and impaired spermatogenesis (n = 17) were investigated. Biopsies showing intact spermatogenesis were included as controls (n = 9). Bone-marrow-derived macrophages (BMDMs) generated from WT mice were treated with activin A (50 ng/ml) for 6 days. Activin-A-treated or untreated BMDMs were then co-cultured with purified mouse splenic T cells for two days to assess chemokine and cytokine production.
PARTICIPANTS/MATERIALS, SETTING, METHODS
Quantitative real-time PCR (qRT-PCR) was used to analyze the expression of chemokines in total testicular RNA collected from mice. Immunofluorescence staining was used to detect activin A, F4/80, and CD3 expression in mouse testes. The expression of chemokine/chemokine-receptor-encoding genes was examined in human testicular biopsies by qRT-PCR. Correlations between chemokine expression levels and either the immune cell infiltration density or the mean spermatogenesis score were analyzed. Immunofluorescence staining was used to evaluate the expression of CD68 and CCR2 in human testicular biopsies. RNA isolated from murine BMDMs was used to characterize these cells in terms of their chemokine/chemokine receptor expression levels. Conditioned media from co-cultures of BMDMs and T cells were collected to determine chemokine levels and the production of pro-inflammatory cytokines tumor necrosis factor (TNF) and interferon (IFN)-γ by T cells.
MAIN RESULTS AND THE ROLE OF CHANCE
Induction of EAO in the testes of WT mice increased the expression of chemokine receptors such as Ccr1 (P < 0.001), Ccr2 (P < 0.0001), Ccr3 (P < 0.0001), Ccr5 (P < 0.0001), CXC motif chemokine receptor (Cxcr)3 (P < 0.01), and CX3C motif chemokine receptor (Cx3cr)1 (P < 0.001), as well as that of most of their ligands. Ccr2 deficiency reversed some of the changes associated with EAO by reducing the expression of Ccr1 (P < 0.0001), Ccr3 (P < 0.0001), Ccr5 (P < 0.01), Cxcr3 (P < 0.001), and Cx3cr1 (P < 0.0001). Importantly, the biopsies showing impaired spermatogenesis and concomitant focal leukocytic infiltration exhibited higher expression of CCL2 (P < 0.01), CCR1 (P < 0.05), CCR2 (P < 0.001), and CCR5 (P < 0.001) than control biopsies with no signs of inflammation and intact spermatogenesis. The gene expression of CCR2 and its ligand CCL2 correlated positively with the immune cell infiltration density (P < 0.05) and negatively with the mean spermatogenesis score (P < 0.001). Moreover, CD68+ macrophages expressing CCR2 were present in human testes with leukocytic infiltration with evidence of tubular damage. Treatment of BMDMs, as surrogates for testicular macrophages, with activin A increased their expression of Ccr1, Ccr2, and Ccr5 while reducing their expression of Ccl2, Ccl3, Ccl4, Ccl6, Ccl7 Ccl8, and Ccl12. These findings were validated in vivo, by showing that inhibiting activin A activity by overexpressing FST in EAO mice decreased the expression of Ccr2 (P < 0.05) and Ccr5 (P < 0.001) in the testes. Interestingly, co-culturing activin-A-treated BMDMs and T cells reduced the levels of CCL2 (P < 0.05), CCL3/4 (P < 0.01), and CCL12 (P < 0.05) in the medium and attenuated the production of TNF (P < 0.05) by T cells. The majority of cells secreting activin A in EAO testes were identified as macrophages.
LARGE SCALE DATA
N/A.
LIMITATIONS, REASONS FOR CAUTION
BMDMs were used as surrogates for testicular macrophages. Hence, results obtained from the in vitro experiments might not be fully representative of the situation in the testes in vivo. Moreover, since total RNA was extracted from the testicular tissue to examine chemokine expression, the contributions of individual cell types as producers of specific chemokines may have been overlooked.
WIDER IMPLICATIONS OF THE FINDINGS
Our data indicate that macrophages are implicated in the development and progression of testicular inflammation by expressing CCR2 and activin A, which ultimately remodel the chemokine/chemokine receptor network and recruit other immune cells to the site of inflammation. Consequently, inhibition of CCR2 or activin A could serve as a potential therapeutic strategy for reducing testicular inflammation.
STUDY FUNDING/COMPETING INTEREST(S)
This work was supported by the International Research Training Group in 'Molecular pathogenesis on male reproductive disorders', a collaboration between Justus Liebig University (Giessen) and Monash University (Melbourne) (GRK1871/1-2) funded by the Deutsche Forschungsgemeinschaft and Monash University, a National Health and Medical Research Council of Australia Ideas Grant (1184867), and the Victorian Government's Operational Infrastructure Support Programme. The authors declare no competing financial interests.
PubMed: 38775335
DOI: 10.1093/humrep/deae107 -
Bioorganic & Medicinal Chemistry Letters Aug 2024TGF-β is an immunosuppressive cytokine and plays a key role in progression of cancer by inducing immunosuppression in tumor microenvironment. Therefore, inhibition of...
TGF-β is an immunosuppressive cytokine and plays a key role in progression of cancer by inducing immunosuppression in tumor microenvironment. Therefore, inhibition of TGF-β signaling pathway may provide a potential therapeutic intervention in treating cancers. Herein, we report the discovery of a series of novel thiazole derivatives as potent inhibitors of ALK5, a serine-threonine kinase which is responsible for TGF-β signal transduction. Compound 29b was identified as a potent inhibitor of ALK5 with an IC value of 3.7 nM with an excellent kinase selectivity.
Topics: Thiazoles; Receptor, Transforming Growth Factor-beta Type I; Drug Design; Humans; Protein Kinase Inhibitors; Structure-Activity Relationship; Protein Serine-Threonine Kinases; Receptors, Transforming Growth Factor beta; Molecular Structure; Dose-Response Relationship, Drug
PubMed: 38759932
DOI: 10.1016/j.bmcl.2024.129797