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Here, we examined the effects of the BMP signaling pathway inhibitor LDN-193189 on the pluripotency of porcine embryonic stem cells (ESCs) in the absence of feeder cells using molecular and transcriptomic techniques. Additionally, the effects of some extracellular matrix components on porcine ESC pluripotency were evaluated to develop an optimized and sustainable feeder-free culture system for porcine ESCs. Feeder cells were found to play an important role in supporting the pluripotency of porcine ESCs by blocking trophoblast and mesodermal differentiation through the inhibition of the BMP pathway. Additionally, treatment with LDN-193189, an inhibitor of the BMP pathway, maintained the pluripotency and homogeneity of porcine ESCs for an extended period in the absence of feeder cells by stimulating the secretion of chemokines and suppressing differentiation, based on transcriptome analysis. Conclusively, these results suggest that LDN-193189 could be a suitable replacement for feeder cells in the maintenance of porcine ESC pluripotency during culture. Additionally, these findings contribute to the understanding of pluripotency gene networks and comparative embryogenesis. [Display omitted] • Inhibiting the SMAD signaling supports the stemness of pig ESCs without feeder cells. • LDN-193189 enhances the pig pluripotency by suppressing mesodermal differentiation. • These conditions enhance the stemness of pig ESCs, even in FBS-contained conditions. • The culture conditions ensure consistency and reproducibility of pig ESC cultures. [ABSTRACT FROM AUTHOR]

The BMP signaling pathway plays a crucial role in regulating early embryonic development and tissue homeostasis. SMAD6 encodes a negative regulator of BMP, and rare variants of SMAD6 are recurrently found in individuals with birth defects. However, we observed that a subset of rare pathogenic variants of SMAD6 consistently exhibited positive regulatory effects instead of the initial negative effects on the BMP signaling pathway. We sought to determine whether these SMAD6 variants have common pathogenic mechanisms. Here, we showed that pathogenic SMAD6 variants accompanying this functional reversal exhibit similar increases in deamidation. Mechanistically, increased deamidation of SMAD6 variants promotes the accumulation of the BMP receptor BMPR1A and the formation of new complexes, both of which lead to BMP signaling pathway activation. Specifically, two residues, N262 and N404, in SMAD6 were identified as the crucial sites of deamidation, which was catalyzed primarily by glutamine-fructose-6-phosphate transaminase 2 (GFPT2). Additionally, treatment of cells harboring SMAD6 variants with a deamidase inhibitor restored the inhibitory effect of SMAD6 on the BMP signaling pathway. Conversely, when wild-type SMAD6 was manually simulated to mimic the deamidated state, the reversed function of activating BMP signaling was reproduced. Taken together, these findings show that deamidation of SMAD6 plays a crucial role in the functional reversal of BMP signaling activity, which can be induced by a subset of various SMAD6 variants. Our study reveals a common pathogenic mechanism shared by these variants and provides a potential strategy for preventing birth defects through deamidation regulation, which might prevent the off-target effects of gene editing. [ABSTRACT FROM AUTHOR]

Elastic cartilage constitutes a major component of the external ear, which functions to guide sound to the middle and inner ears. Defects in auricle development cause congenital microtia, which affects hearing and appearance in patients. Mutations in several genes have been implicated in microtia development, yet, the pathogenesis of this disorder remains incompletely understood. Here, we show that Prrx1 genetically marks auricular chondrocytes in adult mice. Interestingly, BMP-Smad1/5/9 signaling in chondrocytes is increasingly activated from the proximal to distal segments of the ear, which is associated with a decrease in chondrocyte regenerative activity. Ablation of Bmpr1a in auricular chondrocytes led to chondrocyte atrophy and microtia development at the distal part. Transcriptome analysis revealed that Bmpr1a deficiency caused a switch from the chondrogenic program to the osteogenic program, accompanied by enhanced protein kinase A activation, likely through increased expression of Adcy5/8. Inhibition of PKA blocked chondrocyte-to-osteoblast transformation and microtia development. Moreover, analysis of single-cell RNA-seq of human microtia samples uncovered enriched gene expression in the PKA pathway and chondrocyte-to-osteoblast transformation process. These findings suggest that auricle cartilage is actively maintained by BMP signaling, which maintains chondrocyte identity by suppressing osteogenic differentiation.

A 54‐year‐old man with a history of unimelic, post‐traumatic multifocal heterotopic ossification (HO) and normal genetic analysis of ACVR1 and GNAS had variants of unknown significance (VUS) in PDLIM‐7 (PDZ and LIM Domain Protein 7), the gene encoding LMP‐1 (LIM Mineralization Protein‐1), an intracellular protein involved in the bone morphogenetic protein (BMP) pathway signaling and ossification. In order to determine if the LMP‐1 variants were plausibly responsible for the phenotype observed, a series of in vitro experiments were conducted. C2C12 cells were co‐transfected with a BMP‐responsive reporter as well as the LMP‐1 wildtype (wt) construct or the LMP‐1T161I or the LMP‐1D181G constructs (herein designated as LMP‐161 or LMP‐181) corresponding to the coding variants detected in the patient. A significantly increased BMP‐reporter activity was observed in LMP‐161 or LMP‐181 transfected cells compared to the wt cells. The LMP‐181 variant exhibited BMP‐reporter activity with a four‐fold increase over the LMP‐1 wt protein. Similarly, mouse pre‐osteoblastic MC3T3 cells transfected with the patient's LMP‐1 variants expressed higher levels of osteoblast markers both at mRNA and protein levels and preferentially mineralized when stimulated with recombinant BMP‐2 compared to control cells. Presently, there are no pathogenic variants of LMP‐1 known to induce HO in humans. Our findings suggest that the germline variants in LMP‐1 detected in our patient are plausibly related to his multifocal HO (LMP1‐related multifocal HO). Further observations will be required to firmly establish this gene–disease relationship. [ABSTRACT FROM AUTHOR]

BMP-8a is a member of bone morphogenetic proteins (BMPs) and plays a regulatory role in human growth and development as a transcription regulator. This review aims to summarize the current research on the impact and mechanism of BMP-8a in female and male reproduction, formation and eruption of teeth, bone and cartilage development, tissue differentiation, disease occurrence, progression and prognosis. The phrases "BMP-8a," "BMPs," "regulator," "mechanism," "osteoblast," "cartilage," "cancer," "disease," and "inflammation" were searched in the PubMed database. The abstracts were evaluated, and a series of original publications and reviews were examined. According to the search, BMP-8a affects the development of the uterus by inhibiting luteinization and plays an important role in late spermatogenesis. It is highly expressed in osteogenesis and differentially expressed in chondrogenesis. Furthermore, BMP-8a has a significant impact on the occurrence, development and prognosis of various diseases. BMP-8a regulates important factors and pathways, such as SMAD2/3 and SMAD1/5/8, to promote or inhibit the developmental processes of human reproductive organs. BMP-8a is also a member of the BMP family of proteins that regulates chondrogenesis and osteogenesis. In addition to its osteoinductive capabilities, BMP-8a is involved in the progression of diverse cancers. [ABSTRACT FROM AUTHOR]

Graphene quantum dots (GQDs) have been broadly applied in biomedicine in recent years, and their environmental exposure and toxicological impacts have raised increasing concerns. The nanosafety assessment on the nervous system is one of the most important aspects, and potential effects of GQDs on neurodevelopment and the underlying mechanism are still elusive. In this study, the neural developmental toxicities of OH-GQDs and NH2-GQDs were investigated using the mouse embryonic stem cells (mESCs). The results revealed that OH-GQDs significantly inhibited the ectoderm development, and reduced the neural precursor formation and neurogenesis during the neural differentiation of the mESCs. The exploration on the mechanism uncovered that the increased enrichment of H3K27me3 at the promoter region of the Smad6 gene was involved in histone modification-activated BMP signal pathway, which consequently influenced its regulatory effects on neural differentiation. Additionally, OH-GQDs elicited a stronger effect on inducing the imbalance of histone modification, and resulted in higher latency of neural differentiation disturbance than did NH2-GQDs, suggesting surface functionalization-specific effects of GQDs on neurodevelopmental toxicity. This study would provide new insights in not only the adverse effects of GQDs on neurodevelopment, but also the influence from the chemical modification of GQDs on their bioactivities.

Objective To investigate the effect of adiponectin (ApN) on osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs) and to explore its possible mechanism. Methods BMSCs were cultured in vitro. ApN over-expression and interfering plasmids were constructed and transfected into BMSCs. The calcification deposition of cells in each group was observed with Alizarin red staining. The osteogenic differentiation ability of cells in each group was observed with alkaline phosphatase (ALP) staining. The expression of ApN receptors, and bone morphogenetic protein (BMP) signaling pathway and osteogenic related genes were detected using qRT-PCR. Results Compared to those in the control group, calcification deposition and ALP positive expression increased, the mRNA expression levels of AdipoRl, AdipoR2, BMP2, RUNX2, Smadl, and Smad5 increased significantly in the over-expression group (P < 0. 05). Calcification deposition and ALP positive expression decreased, and the mRNA expression levels of AdipoRl, AdipoR2, BMP2, RUNX2, Smadl, and Smad5 decreased significantly in the interference group (P

[Display omitted] • Conversion of titanium sulfate to flowery titanium phosphate bioceramics. • Bioactive 3D platforms with controlled ion release, enhance cellular interaction. • Rapid osteogenic differentiation and mineralization via BMP-2 expression. • The platform supports ECM remodeling and bone tissue regeneration in vivo. Regenerative bone implants have been designed to promote new bone formation, however, the search for a suitable implant remains challenging despite the use of various biomaterials (e.g., bone cement, hydroxyapatite, allograft). In this study, we developed a highly bioactive flower-like titanium phosphate (Ti(HPO 4) 2) bioceramic and poly(ε-caprolactone) 3D composite with a hierarchical structure and interconnected pores. The newly synthesized Ti(HPO 4) 2 bioceramic, exhibiting an amorphous surface resembling native bone, high porosity, and enhanced ion release properties, was incorporated into 3D scaffolds using an optimized pneumatic 3D printing process. These scaffolds demonstrated their potential for bone regeneration by promoting early osteogenic differentiation and rapid mineralization with human bone marrow mesenchymal stem cells (hBM-MSCs). Moreover, it was revealed their structural stability and tissue regeneration effects after implantation in mouse calvaria. The developed 3D-printed PCL/Ti(HPO 4) 2 scaffold exhibited enhanced performance in bone regeneration and angiogenesis effects, activated by the bone morphogenetic protein (BMP) signaling pathway. Therefore, this study demonstrates an advanced platform with excellent physicochemical and biological properties for bone regeneration. [ABSTRACT FROM AUTHOR]

Although germ cell fate is believed to be determined by signaling factors from differentiated somatic cells, the molecular mechanism behind this process remains obscure. In this study, premature meiosis in male germ cells was observed during the embryonic stage by conditional activation of β‐catenin in Sertoli cells. Somatic and germ cell transcriptome results indicated that the BMP signaling pathway was enriched after β‐catenin activation. In addition, we observed a decreased DNA methylation within a reduction of DNMT3A in germ cells of β‐catenin activated testes and reversed increase after inhibiting BMP signaling pathway with LDN‐193189. We also found that Dazl expression was increased in β‐catenin activated testes and decreased after LDN treatment. Taken together, this study demonstrates that male germ cells entered meiosis prematurely during the embryonic stage after β‐catenin activated in Sertoli cells. BMP signaling pathway involved in germ cell meiosis initiation by mediating DNA methylation to induce meiotic genes expression. [ABSTRACT FROM AUTHOR]

Abstract Background Amelogenesis imperfecta (AI) is a type of hereditary diseases that manifest defects in the formation or mineralization of enamel. Recently, it is reported that inactivation of FAM20C, a well-known Golgi casein kinase, caused AI. However, the mechanism of it is still unknown. The aim of this study was to explore the molecular mechanism of AI, which caused by ablation of FAM20C. Results In the Sox2-Cre;Fam20C fl/fl (cKO) mouse, we found abnormal differentiation of ameloblasts, improper formation and mineralization of enamel, and downregulation of both mRNA and protein level of enamel matrix proteins, including amelogenin (AMEL), ameloblastin (AMBN) and enamelin (ENAM). The levels of BMP2, BMP4 and BMP7, the ligands of BMP signaling pathway, and phosphorylation of Smad1/5/8, the key regulators of BMP signaling pathway, were all decreased in the enamel matrix and the ameloblast of the cKO mice, respectively. The expression of cyclin-dependent kinase inhibitor (P21), muscle segment homeobox genes 2 (Msx2), which are the target genes of the BMP signaling pathway, and laminin 3, the downstream factor of Msx2, were all significantly decreased in the ameloblasts of the cKO mice compared to the control mice. Conclusion the results of our study suggest that ablation of FAM20C leads to AI through inhibiting the Smad dependent BMP signaling pathway in the process of amelogenesis.

Pulmonary hypertension (PH) is an extremely serious cardiopulmonary disease, finally leading to progressive right ventricular failure and death. Our previous studies have nominated HLQ2g, a pyrazolo[3,4-b] pyridine derivative stimulating soluble guanylate cyclase (sGC), as a new candidate for the treatment of PH, but the specific mechanism is still not clear. The PH model induced by hypoxia was established in rats. Right ventricular systolic pressure (RVSP) was assessed by jugular vein catheterization. RV weight was the index to evaluate RV hypertrophy. The protein levels of cGMP-dependent protein kinase type I (cGKI), bone morphogenetic protein receptor 2 (BMPR2), phosphorylated Smad1/5/8 (p-Smad1/5/8), and inhibitor of differention 1 (Id1) in pulmonary artery and human pulmonary artery smooth muscle cells (HPASMCs) were determined by western blotting. Cell proliferation and migration were evaluated. In the whole experiment, the first clinically available sGC stimulator Riociguat was used as the reference. In hypoxic PH rat model, elevated RVSP and RV hypertrophy were significantly reduced by HLQ2g treatment. Both Riociguat and HLQ2g attenuated vascular remodeling accompanied with up-regulated cGKI expression and BMP signaling pathway, which was characterized by elevated expression of BMPR2, p-Smad1/5/8, and Id1 in HPH rats. In addition, HLQ2g inhibited proliferation and migration of HPASMCs induced by hypoxia and platelet-derived growth factor (PDGF), restored BMPR2 signaling, which was recalled by Rp-8-Br-PET-cGMPS, the inhibitor of cGKI. In summary, the novel pyrazolo[3,4-b] pyridine derivative HLQ2g can alleviate HPH progression by up-regulating cGKI protein and BMP signaling pathway. [ABSTRACT FROM AUTHOR]

Abstract Background This study was conducted with the aim of exploring the effect of the BMP signaling pathway on osteoblastic differentiation in rat bone marrow mesenchymal stem cells (rBMSCs) in rats with osteoporosis (OP). Methods The bilateral ovaries of female SD rats were resected for the establishment of a rat OP model. The osteoblastic differentiation of isolated rBMSCs was identified through osteogenic induction. Adipogenetic induction and flow cytometry (FCM) were used to detect adipogenic differentiation and the expression of rBMSC surface markers. The rBMSCs were grouped into the blank group, NC group, si-BMP2 group, and oe-BMP2 group. The expression levels of key factors and osteogenesis-related factors were determined by Western blot and quantitative real-time polymerase chain reaction (qRT-PCR). The formation of calcified nodules was observed by alizarin red staining. ALP activity was measured by alkaline phosphatase staining. Results The rats with OP had greater weight but decreased bone mineral density (BMD) than normal rats (all P

Pulmonary hypertension (PH) is an extremely serious cardiopulmonary disease, finally leading to progressive right ventricular failure and death. Our previous studies have nominated HLQ2g, a pyrazolo[3,4-b] pyridine derivative stimulating soluble guanylate cyclase (sGC), as a new candidate for the treatment of PH, but the specific mechanism is still not clear. The PH model induced by hypoxia was established in rats. Right ventricular systolic pressure (RVSP) was assessed by jugular vein catheterization. RV weight was the index to evaluate RV hypertrophy. The protein levels of cGMP-dependent protein kinase type I (cGKI), bone morphogenetic protein receptor 2 (BMPR2), phosphorylated Smad1/5/8 (p-Smad1/5/8), and inhibitor of differention 1 (Id1) in pulmonary artery and human pulmonary artery smooth muscle cells (HPASMCs) were determined by western blotting. Cell proliferation and migration were evaluated. In the whole experiment, the first clinically available sGC stimulator Riociguat was used as the reference. In hypoxic PH rat model, elevated RVSP and RV hypertrophy were significantly reduced by HLQ2g treatment. Both Riociguat and HLQ2g attenuated vascular remodeling accompanied with up-regulated cGKI expression and BMP signaling pathway, which was characterized by elevated expression of BMPR2, p-Smad1/5/8, and Id1 in HPH rats. In addition, HLQ2g inhibited proliferation and migration of HPASMCs induced by hypoxia and platelet-derived growth factor (PDGF), restored BMPR2 signaling, which was recalled by Rp-8-Br-PET-cGMPS, the inhibitor of cGKI. In summary, the novel pyrazolo[3,4-b] pyridine derivative HLQ2g can alleviate HPH progression by up-regulating cGKI protein and BMP signaling pathway.

Elastic cartilage constitutes a major component of the external ear, which functions to guide sound to the middle and inner ears. Defects in auricle development cause congenital microtia, which affects hearing and appearance in patients. Mutations in several genes have been implicated in microtia development, yet, the pathogenesis of this disorder remains incompletely understood. Here, we show that Prrx1 genetically marks auricular chondrocytes in adult mice. Interestingly, BMP-Smad1/5/9 signaling in chondrocytes is increasingly activated from the proximal to distal segments of the ear, which is associated with a decrease in chondrocyte regenerative activity. Ablation of Bmpr1a in auricular chondrocytes led to chondrocyte atrophy and microtia development at the distal part. Transcriptome analysis revealed that Bmpr1a deficiency caused a switch from the chondrogenic program to the osteogenic program, accompanied by enhanced protein kinase A activation, likely through increased expression of Adcy5/8 . Inhibition of PKA blocked chondrocyte-to-osteoblast transformation and microtia development. Moreover, analysis of single-cell RNA-seq of human microtia samples uncovered enriched gene expression in the PKA pathway and chondrocyte-to-osteoblast transformation process. These findings suggest that auricle cartilage is actively maintained by BMP signaling, which maintains chondrocyte identity by suppressing osteogenic differentiation., Competing Interests: RY, HC, HY, ZZ, HL, BL No competing interests declared, YM Reviewing editor, eLife, (© 2023, Yang, Chu et al.)

BACKGROUND: Noggin is an inhibitory molecule of bone morphogenetic protein (BMP) signaling, which can bind to BMP2/4 to form a complex, thereby blocking BMP signaling and affecting normal development. Some studies have suggested that dermal sheath cells are a kind of dermal stem cells that can self-renew for a long time, and participate in the formation of dermal papilla and dermal sheath during hair follicle regeneration. During the development of hair follicles, Noggin is involved in the induction of hair follicle regeneration, and its loss leads to a decrease in hair follicle number and slows hair follicle growth. However, little is known about the Noggin in the study of hair follicle dermal sheaths. OBJECTIVE: To study the biological function of Noggin protein in the dermal sheath of hair follicles. METHODS: The dermal sheath specific α-SMA-Cre ERT2 tool mice were used to specifically overexpress Noggin protein in the dermal sheath. At 8 and 9 days after birth, αSMA-CreER; pMES-Noggin mice in the experimental group and αSMA-CreER mice in the control group were injected with 4-hydroxytamoxifen. At 21, 23, and 28 days after birth, mouse skin tissues were obtained. Hematoxylin-eosin staining was used to observe hair follicle growth and subcutaneous fat layer thickness, and immunohistochemical staining was used to analyze cell phenotype. RESULTS AND CONCLUSION: Based on the findings of hematoxylin-eosin staining, we found that the growth of hair follicle was not affected, but the growth and development of subcutaneous adipose tissue of hair follicles were seriously impacted, presenting with the subcutaneous fat layer becoming thinned. The immunohistochemical results indicated that the decrease of BMP signaling may result in the thinning of the hair follicle fat layer. This discovery expands the understanding of dermal sheath cells and Noggin protein, and provides an important basis for more accurate understanding of hair follicle regeneration and tissue growth mechanisms. [ABSTRACT FROM AUTHOR]

Pulmonary arterial hypertension (PAH) is a malignant disease with high mortality and closely involves the bone morphogenetic protein (BMP) pathway. Mutations in BMPR2 caused proliferation of pulmonary artery smooth muscle cells (PASMCs) leading to PAH. Isorhamnetin, one of the main naturally occurring flavonoids extracted from Hippophae rhamnoides L, shows antiinflammatory and anti-proliferative properties. Nevertheless, the effects of isorhamnetin on PAH remain unclear. This study aimed to investigate whether isorhamnetin has protective effects against PAH and explore possible mechanisms. An in vivo model of PAH induced by monocrotaline (MCT) was employed, and sildenafil and isorhamnetin were orally administered for 21 consecutive days. An in vitro model induced by TNF-α was employed, and cell proliferation of HPASMCs was detected. Results indicated that isorhamnetin significantly improved hemodynamic, histopathological, and echocardiographic changes in MCT-induced PAH in rats. In vitro, isorhamnetin suppressed TNF-α-induced HPASMCs proliferation. Furthermore, isorhamnetin improved protein expression of BMPR2 and suppressed protein expression of TNF-α and IL-6 in rat lungs. Isorhamnetin improved protein expression of BMPR2 and p-smad1/5 and mRNA expression of Id1 and Id3 in HPASMCs. Isorhamnetin ameliorated MCT-induced PAH in rats and inhibited TNF-α-induced HPASMCs proliferation by a mechanism likely involving the regulation of the BMP signaling pathway. [ABSTRACT FROM AUTHOR]

Background: This study was conducted with the aim of exploring the effect of the BMP signaling pathway on osteoblastic differentiation in rat bone marrow mesenchymal stem cells (rBMSCs) in rats with osteoporosis (OP). Methods: The bilateral ovaries of female SD rats were resected for the establishment of a rat OP model. The osteoblastic differentiation of isolated rBMSCs was identified through osteogenic induction. Adipogenetic induction and flow cytometry (FCM) were used to detect adipogenic differentiation and the expression of rBMSC surface markers. The rBMSCs were grouped into the blank group, NC group, si-BMP2 group, and oe-BMP2 group. The expression levels of key factors and osteogenesis-related factors were determined by Western blot and quantitative real-time polymerase chain reaction (qRT-PCR). The formation of calcified nodules was observed by alizarin red staining. ALP activity was measured by alkaline phosphatase staining. Results: The rats with OP had greater weight but decreased bone mineral density (BMD) than normal rats (all P < 0.01). The rBMSCs from rats with OP were capable of osteoblastic differentiation and adipogenic differentiation and showed high expression of CD44 (91.3 ± 2.9%) and CD105 (94.8 ± 2.1%). Compared with the blank group, the oe-BMP2 group had elevated BMP-2 and Smad1 levels and an increase in calcified nodules and ALP-positive staining areas (all P < 0.05). Moreover, the expression levels of Runx2, OC, and OPN in the oe-BMP2 group were relatively higher than those in the blank group (all P < 0.05). The findings in the si-BMP2 group were opposite to those in the oe-BMP2 group. Conclusion: BMP signaling pathways activated by BMP-2 can promote the osteoblastic differentiation of rBMSCs from rats with OP. [ABSTRACT FROM AUTHOR]

Hypoxia‐induced excessive pulmonary artery smooth muscle cell (PASMC) proliferation plays an important role in the pathology of pulmonary arterial hypertension (PAH). Berberine (BBR) is reported as an effective antiproliferative properties applied in clinical. However, the effect of BBR on PAH remains unclear. In the present study, we elucidated the protective effects of BBR against abnormal PASMC proliferation and vascular remodeling in chronic hypoxia‐induced hearts. Furthermore, the potential mechanisms of BBR were investigated. For this purpose, C57/BL6 mice were exposed to chronic hypoxia for 4 weeks to mimic severe PAH. Hemodynamic and pulmonary pathomorphology data showed that chronic hypoxia significantly increased the right ventricular systolic pressure (RVSP), the right ventricle/left ventricle plus septum RV/(LV + S) weight ratio, and the median width of pulmonary arterioles. BBR attenuated the elevations in RVSP and RV/(LV + S) and mitigated pulmonary vascular structure remodeling. BBR also suppressed the hypoxia‐induced increases in the expression of proliferating cell nuclear antigen (PCNA) and of α‐smooth muscle actin. Furthermore, administration of BBR significantly increased the expression of bone morphogenetic protein type II receptor (BMPR‐II) and its downstream molecules P‐smad1/5 and decreased the expression of transforming growth factor‐β (TGF‐β) and its downstream molecules P‐smad2/3. Moreover, peroxisome proliferator‐activated receptor γ expression was significantly decreased in the hypoxia group, and this decrease was reversed by BBR treatment. Our study demonstrated that the protective effect of BBR against hypoxia‐induced PAH in a mouse model may be achieved through altered BMPR‐II and TGF‐β signaling. [ABSTRACT FROM AUTHOR]

Learning to discriminate between environmental visual stimuli is essential to make right decisions and guide appropriate behaviors. Moreover, impairments in visual discrimination learning are observed in several neuropsychiatric disorders. Visual discrimination learning requires perception and memory processing, in which the hippocampus critically involved. To understand the molecular mechanisms underpinning hippocampus function in visual discrimination learning, we examined the hippocampal gene expression profiles of Sprague-Dawley rats with different cognitive performance (high cognition group vs. low cognition group) in the modified visual discrimination learning task, using high-throughput RNA sequencing technology. Compared with the low cognition group, bioinformatics analysis indicated that 319 genes were differentially expressed in the high cognition group with statistical significance, of which 253 genes were down-regulated and 66 genes were up-regulated. The functional enrichment analysis showed that protein translation and energy metabolism were up-regulated pathways, while transforming growth factor beta receptor signaling pathway, bone morphogenetic protein signaling pathway, apoptosis, inflammation response, transport, and glycosaminoglycan metabolism were down-regulated pathways, which were related to good cognitive performance in the visual discrimination learning task. Taken together, our finding reveals the differential gene expression and enrichment biological pathways related to cognitive performance differences in visual discrimination learning of rats, which provides us direct insight into the molecular mechanisms of hippocampus function in visual discrimination learning and may contribute to developing potential treatment strategies for neuropsychiatric disorders accompanied with cognitive impairments. • Compared with pigmented rats, the image parameters for SD rats were optimized in visual discrimination paradigm. • Hippocampal transcriptome was associated with different cognitive performance in visual discrimination learning task. • Energy metabolism pathway and TGF-beta signaling pathway, etc., may contribute to cognitive performance differences. [ABSTRACT FROM AUTHOR]

OBJECTIVE: To study the effect of micro-ribonucleic acid (miR)-20b on osteocyte apoptosis in rats with steroid-induced necrosis of the femoral head (SNFH) and to analyze whether the bone morphogenetic protein (BMP) signaling pathway is involved in the regulation. MATERIALS AND METHODS: A total of 36 Sprague-Dawley rats were randomly divided into control group (n=12), model group (n=12) and intervention group (n=12). The rat model of SNFH was established in the model and intervention groups, while the rats in the intervention group were intraperitoneally injected with the bone morphogenetic protein (BMP) signaling pathway inhibitor. After modeling, the femoral head in each group was taken, and the morphology of osteocytes was observed via hematoxylin-eosin (HE) staining. The apoptosis level of femoral head cells was detected via terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) staining. The miR-20b expression level in femoral head cells in each group was detected via quantitative Polymerase Chain Reaction (qPCR). The expression levels of inflammatory factors in femoral head cells in each group were detected via enzyme-linked immunosorbent assay (ELISA). The expression levels of apoptotic proteins and BMP signaling pathway-related proteins in femoral head cells in each group were detected via Western blotting. RESULTS: Compared with those in the control group, the bone trabecula was sparse, the number of osteocytes significantly declined and the number of apoptotic osteocytes markedly increased (p<0.01); the expression level of miR-20b in bone tissues remarkably increased (p<0.01), the content of interleukin-1β (IL-1β), IL-6 and tumor necrosis factor-α (TNF-α) in bone tissues increased (p<0.01), the content of IL-10 significantly declined (p<0.01), the expression level of cleaved caspase-3 protein in bone tissues markedly increased (p<0.01), the Bcl-2/ Bax expression level evidently declined (p<0.01) and the expression levels of anaplastic lymphoma kinase3 (ALK3), GATA4 and NKX2.5 in bone tissues remarkably increased (p<0.01) in the model group. Compared with those in the model group, the necrosis of bone tissues significantly decreased, the apoptosis level of osteocytes remarkably declined (p<0.01), the content of IL-1β, IL-6 and TNF-α in bone tissues markedly decreased (p<0.01), the content of IL-10 increased (p<0.01), the expression level of cleaved caspase-3 protein in bone tissues significantly declined (p<0.01), the B-cell lymphoma 2/BCL2-Associated X (Bcl-2/Bax) expression level markedly increased (p<0.01) and the expression levels of ALK3, GATA4 and NKX2.5 in bone tissues significantly decreased (p<0.01) in the intervention group. CONCLUSIONS: SNFH will significantly increase the expression level of miR-20b in bone tissues, thereby activating the BMP signaling pathway, promoting the release of inflammatory factors and leading to osteocyte apoptosis. Inhibiting the BMP signaling pathway can effectively reduce the osteocyte apoptosis level. [ABSTRACT FROM AUTHOR]

Glaucoma is the leading cause of irreversible blindness worldwide. Although the etiology of glaucoma is incompletely understood, it is known that the extracellular matrix (ECM) of the trabecular meshwork, oxidative stress, TGFβ signaling pathways, and apoptosis are important components of glaucoma pathogenesis. These components appear to be interrelated, but knowledge on their interactions remains incomplete. Relevant to this gap in knowledge, LTBP2 , glaucoma causing gene, may also be related to the mentioned components of glaucoma pathogenesis because of its putative roles in TGFβ signaling and ECM functions. This background prompted us to further query interactions among some molecules and pathways thought to be important in glaucoma etiology, with emphasis on oxidative stress and LTBP2. To this end, effects of LTBP2 siRNA knockdown, oxidative stress induction, TGFβ2 and gremlin exposures on canonical TGFβ and BMP signaling pathways, expression of ECM related genes, and apoptosis were assayed in primary human trabecular meshwork cell cultures. We found that oxidative stress induction and LTBP2 knockdown both affected all the processes queried, and that their affects paralleled one another. We suggest that effects of both oxidative stress and LTBP2 knockdown on the ECM and apoptosis may be mediated by TGFβ and BMP signaling pathway activation. [ABSTRACT FROM AUTHOR]

The ACVR1 gene encodes a type I receptor of bone morphogenetic proteins (BMPs). Activating mutations in ACVR1 are responsible for fibrodysplasia ossificans progressiva (FOP), a rare disease characterized by congenital toe malformation and progressive heterotopic endochondral ossification leading to severe and cumulative disability. Until now, no therapy has been available to prevent soft-tissue swelling (flare-ups) that trigger the ossification process. With the aim of finding a new therapeutic strategy for FOP, we developed a high-throughput screening (HTS) assay to identify inhibitors of ACVR1 gene expression among drugs already approved for the therapy of other diseases. The screening, based on an ACVR1 promoter assay, was followed by an in vitro and in vivo test to validate and characterize candidate molecules. Among compounds that modulate the ACVR1 promoter activity, we selected the one showing the highest inhibitory effect, dipyridamole, a drug that is currently used as a platelet anti-aggregant. The inhibitory effect was detectable on ACVR1 gene expression, on the whole Smad-dependent BMP signaling pathway, and on chondrogenic and osteogenic differentiation processes by in vitro cellular assays. Moreover, dipyridamole reduced the process of heterotopic bone formation in vivo. Our drug repositioning strategy has led to the identification of dipyridamole as a possible therapeutic tool for the treatment of FOP. Furthermore, our study has also defined a pipeline of assays that will be useful for the evaluation of other pharmacological inhibitors of heterotopic ossification.

The isoflavone calycosin-7-O-β-d-glucopyranoside (CG) is a principal constituent of Astragalus membranaceus (AR) and has been reported to inhibit osteoclast development in vitro and bone loss in vivo. The aim of this study was to investigate the osteogenic effects of CG and its underlying mechanism in ST2 cells. The results show that exposure of cells to CG in osteogenic differentiation medium increases ALP activity, osteocalcin (Ocal) mRNA expression and the osteoblastic mineralization process. Mechanistically, CG treatment increased the expression of bone morphogenetic protein 2 (BMP-2), p-Smad 1/5/8, β-catenin and Runx2, all of which are regulators of the BMP- or wingless-type MMTV integration site family (WNT)/β-catenin-signaling pathways. Moreover, the osteogenic effects of CG were inhibited by Noggin and DKK-1 which are classical inhibitors of the BMP and WNT/β-catenin-signaling pathways, respectively. Taken together, the results indicate that CG promotes the osteoblastic differentiation of ST2 cells through regulating the BMP/WNT signaling pathways. On this basis, CG may be a useful lead compound for improving the treatment of bone-decreasing diseases and enhancing bone regeneration.

During endochondral ossification, cartilage template is eventually replaced by bone. This process involves several well characterized, stereotypic, molecular and cellular changes in the cartilage primordia. These steps involve transition from resting to proliferative and then pre-hypertrophic to finally hypertrophic cartilage. BMP signaling is necessary and sufficient for osteogenesis. However, the specific step(s) of endochondral ossification in which BMP signaling plays an essential role is not yet known. In this study we have identified Prdx1, a known scavenger of ROS, to be expressed in pre-hypertrophic chondrocytes in a BMP signaling-dependent manner. We demonstrate that BMP signaling inhibition increases ROS levels in osteogenic cells. Further, Prdx1 regulates osteogenesis in vivo by helping maintenance of Ihh expressing pre-hypertrophic cells, in turn regulating these cells’ transition into hypertrophy. Therefore, our data suggests that one of the key roles of BMP signaling in endochondral ossification is to maintain pre-hypertrophic state. [ABSTRACT FROM AUTHOR]

The eye is a complex organ with precise function. In early embryonic development, eye is composed of several parts which are from the different germ layers through a series of induction and interaction. Multiple signaling pathways and their associated regulatory factors play important roles in the development of eyes. This review focuses on the relationship between eye development and BMP signaling pathway. The activation of BMP signal can induce the regeneration of the lens and the process of cornea to lens trans-differentiation (CLT). The activation of BMP signal can also maintain the function of the ciliary body, promote the development of retina, affect scleral remodeling and the development of lacrimal gland. Many ocular diseases are closely related to the dysregulation of BMP signaling pathway. Therefore, the regulation of BMP signal pathway can serve as a potential target for therapies of ocular diseases. [ABSTRACT FROM AUTHOR]

Background Abusive alcohol utilization of pregnant woman may cause congenital heart disease ( CHD) of fetus, where alcohol ignites histone H3 hyperacetylation leading to abnormal development of heart morphogenesis and associated genes. Knowledge about the regularized upstream genes is little, but bone morphogenetic protein ( BMP) signaling may actively and prominently take part in alteration in acetylation of histone H3. The supreme objective of this study was to unearth the involvement of BMP signaling pathway in alcohol-driven hyperacetylation of histone H3 in cardiomyoblast cells. Methods Cardiomyoblast cells (H9c2 cells) were addicted with alcohol (100 mM) for 24 hours. Dorsomorphin (5 μM) was used for the inhibition of BMP signaling pathway. We detected the phosphorylation activity of SMAD1/5/8, mRNA expression, histone acetyltransferases ( HAT)/histone deacetylase ( HDAC) activity, and acetylation of histone H3. Results Following alcohol exposure, phosphorylation of SMAD1/5/8 and HAT activities was increased to a significant extent, while histone H3 acetylation and expression of heart development-related genes were also increased. The said phenomenon influenced by alcohol was reverted upon dorsomorphin treatment to the cells without effecting HDAC activity. Conclusions The data clearly identified that BMP-mediated histone H3 acetylation of heart development-related genes might be one of the possible cellular mechanisms to control alcohol-induced expression of heart development-related genes. Dorsomorphin, on the other hand, may modulate alcohol-induced hyperacetylation of histone H3 through BMP targeting, which could be a potential way to block CHD. [ABSTRACT FROM AUTHOR]

[Display omitted] • OH-GQDs inhibited the ectoderm development of the mESCs. • Exposure to OH-GQDs reduced the neural precursor formation and neurogenesis. • OH-GQDs selectively affected BMP pathway by disturbing histone modification. • OH-GQDs elicited higher hazards on neurodevelopment than did NH 2 -GQDs. Graphene quantum dots (GQDs) have been broadly applied in biomedicine in recent years, and their environmental exposure and toxicological impacts have raised increasing concerns. The nanosafety assessment on the nervous system is one of the most important aspects, and potential effects of GQDs on neurodevelopment and the underlying mechanism are still elusive. In this study, the neural developmental toxicities of OH-GQDs and NH 2 -GQDs were investigated using the mouse embryonic stem cells (mESCs). The results revealed that OH-GQDs significantly inhibited the ectoderm development, and reduced the neural precursor formation and neurogenesis during the neural differentiation of the mESCs. The exploration on the mechanism uncovered that the increased enrichment of H3K27me3 at the promoter region of the Smad6 gene was involved in histone modification-activated BMP signal pathway, which consequently influenced its regulatory effects on neural differentiation. Additionally, OH-GQDs elicited a stronger effect on inducing the imbalance of histone modification, and resulted in higher latency of neural differentiation disturbance than did NH 2 -GQDs, suggesting surface functionalization-specific effects of GQDs on neurodevelopmental toxicity. This study would provide new insights in not only the adverse effects of GQDs on neurodevelopment, but also the influence from the chemical modification of GQDs on their bioactivities. [ABSTRACT FROM AUTHOR]

Ethnopharmacological significance Icariin (I), ferulic acid (F) and timosaponin B II (T) derived respectively from the leaf of Epimedium brevicornu Maxim (EBM, Berberidaceae), rhizome of Anemarrhena asphodeloides Bunge (AAB, Liliaceae) and root of Angelica sinensis (Oliv.) Diels (ASD, Umbelliferae) are included in several traditional Chinese medicine (TCM) formulas for the treatment of osteoporosis. In addition, the medicinal materials and chemical constituents in many traditional Chinese formulas have been shown to have potential synergistic, additive and antagonistic effects. Aim of study To explore the action mechanism and interactions between I, T and F as bone anabolic ingredients on osteoblasts, and fully understand their action mechanism and rationality of the formula design. Materials and methods An osteoporotic model was established in bilaterally ovariectomized mice. Bone mineral density (BMD), bone mineral content (BMC) and serum biochemical parameters including alkaline phosphatase (ALP), tartrate resistant acid phosphatase (TRAP), osteoprotegerin (OPG) and deoxypyridinoline cross-links (DPD) were measured to evaluate the effects of I, T or F alone and their combinations on osteoporotic mice. UMR-106 osteoblastic cells and primary osteoblasts in neonatal rat calvarias were used to evaluate the osteogenesis effect. The immunohistochemical method and Western-blot analysis were used to detect the expression of critical proteins in the process of proliferation and differentiation of osteoblasts. Results IFT combinations enhanced the therapeutic effect without increasing the adverse effects on osteoporotic mice, synergistically increased the osteoblast proliferation, ALP activity and mineralized nodule formation, and promoted the expression of bone matrix by regulating BMP and Wnt/β-catenin signaling pathways in osteoblasts. Conclusion IFT combinations reinforced the therapeutic effect on osteoporosis by modulating multi-signaling pathways and action targets. [ABSTRACT FROM AUTHOR]

Background: Mollusca is the second largest phylum in nature. The shell of molluscs is a remarkable example of a natural composite biomaterial. Biomineralization and how it affects mollusks is a popular research topic. The BMP-2 signaling pathway plays a canonical role in biomineralization. SMAD4 is an intracellular transmitter in the BMP signaling pathway in mammals, and some genomic data show SMAD4's involvment in BMP signaling in invertbrates, but whether SMAD4 plays a conservative role in pearl oyster, Pinctada fucata, still need to be tested. Results: In this study, we identified a SMAD4 gene (hereafter designated PfSMAD4) in pearl oyster Pinctada fucata. Bioinformatics analysis of PfSMAD4 showed high identity with its orthologs. PfSMAD4 was located in the cytoplasm in immunofluorescence assays and analyses of PfSMAD4 mRNA in tissues and developmental stages showed high expression in ovaries and D-shaped larvae. An RNA interference experiment, performed by PfSMAD4 double-stranded RNA (dsRNA) injection, demonstrated inhibition not only of nacre growth but also organic sheet formation with a decrease in PfSMAD4 expression. A knockdown experiment using PfBMP2 dsRNA showed decreased PfBMP2 and PfSMAD4 mRNA and irregular crystallization of the nacreous layer using scanning electron microscopy. In co-transfection experiments, PfBMP2-transactivated reporter constructs contained PfSMAD4 promoter sequences. Conclusions: Our results suggest that PfSMAD4 plays a role in biomineralization and can transduce BMP signals in P. fucata. Our data provides important clues about the molecular mechanisms that regulate biomineralization in pearl oyster. [ABSTRACT FROM AUTHOR]