Expression Of Bone Morphogenetic Protein Research Articles

Effects of constitutively active IKKβ on cardiac development

NF-κB is a major transcription factor regulating cell survival, organ development and inflammation, but its role in cardiac development has been inadequately explored. To examine this function, we generated mice in which IKKβ, an essential kinase for NF-κB activation, was constitutively activated in embryonic cardiomyocytes. For this purpose, we used smooth muscle-22α (SM22α)-Cre mice, which are frequently used for gene recombination in embryonic cardiomyocytes. Embryonic hearts of SM22αCre-CA (constitutively active) IKKβflox/flox mice revealed remarkably thin, spongy and hypoplastic myocardium. In exploring the mechanism, we found that the expression of bone morphogenetic protein 10 (BMP10) and T-box transcription factor 20 (Tbx20), major regulators of cardiac development, was significantly downregulated and upregulated, respectively, in the SM22αCre-CAIKKβflox/flox mice. We also generated NK2 homeobox 5 (Nkx2.5) Cre-CAIKKβflox/wt mice since Nkx2.5 is also expressed in embryonic cardiomyocytes and confirmed that the changes in these genes were also observed. These results implicated that the activation of NF-κB affects cardiac development.

MO425: Rosuvastatin Activates Hox13-Usag-1 Pathway and Prevents Renal Fibrosis

Abstract BACKGROUND AND AIMS Statins, 3-hydroxy-3-methylglutaryl-coenzyme A reductase inhibitors, are primarily cholesterol-lowering drugs that have become standard of care in the primary and secondary prevention of cardiovascular diseases. Apart from lipid-lowering, statins may act beneficially through anti-fibrotic mechanisms to protect the diseased kidney. This study evaluated anti-fibrotic effects of rosuvastatin (RSV) in a chronic kidney fibrosis model and against the TGF-ß1 stimulated Madin-Darby canine kidney (MDCK) cells in vitro. METHOD Mice subjected to unilateral ischemic reperfusion injury with contralateral nephrectomy (uIRIx) were treated with vehicle or RSV (10 mg/kg, by oral gavage) daily for 4 weeks and kidneys were analyzed for markers of fibrosis, bone morphogenetic protein-7 (BMP-7), uterine sensitization-associated gene-1 (USAG-1), and SMAD signaling. Control and homeobox protein Hox-A13 (HOXA13) knocked down MDCK cells were stimulated with TGF- ß1 (5 ng/ml) and then treated with RSV. RESULTS Kidneys from uIRIx mice showed increased expression of α-SMA, Collagen 1 and decrease in BMP-7 (20.35 ± 2.37 versus 1.00 ± 0.27, P < 0.05; 8.43 ± 1.55 versus 1.00 ± 0.35, P < 0.05; 0.75 ± 0.06 versus 1.00 ± 0.15, P < 0.05, respectively). In contrast, expression of USAG-1, a BMP-7 antagonist, was markedly increased in fibrotic kidney (6.60 ± 1.11 versus 1.00 ± 0.02, P < 0.05). Interestingly, RSV treatment not only attenuated expression of USAG-1 (0.72 ± 0.08 versus 1.36 ± 0.14, P < 0.05) but also showed a tendency to activate expression of HOXA13 (0.36 ± 0.14 versus 0.51 ± 0.15, P > 0.05) and improved other markers of fibrosis. Moreover, RSV treatment significantly reduced phosphorylated Smad3 (3.94 ± 0.81 versus 7.17 ± 1.50, P < 0.05) and increased phosphorylation levels of Smad 1/5/9 (0.67 ± 0.10 versus 0.30 ± 0.09, P < 0.05) that is associated with BMP-7 signaling in the fibrotic kidney. MDCK cells stimulated with TGF-β1 in vitro showed increased expression of α-SMA, fibronectin, vimentin, collagen 1, USAG-1, and phosphorylation of Smad3 as well as decreased expression of phosphorylated Smad 1/5/9. RSV treatment significantly reversed these changes as well as increased level of transcriptional factor HOXA13 (0.89 ± 0.12 versus 0.33 ± 0.08, P < 0.05), which negatively regulates USAG-1, without changes in BMP-7 expression. In addition, effect of RSV treatment on USAG-1 expression was significantly decreased in HOXA13 gene knocked down MDCK cells (1.01 ± 0.19 versus 2.04 ± 0.38, P < 0.05; 1.66 ± 0.13 versus 2.04 ± 0.38, P > 0.05; TGF- ß1 + RSV versus TGF- ß1 and TGF- ß1 + RSV + siRNA vs TGF- ß1, respectively). These in vitro data suggest that RSV enhanced anti-fibrotic pathway by downregulating a dominant BMP-7 antagonist USAG-1 via HOXA13 upregulation and not by enhancing BMP-7 production. CONCLUSION The present results demonstrate that RSV inhibits the progression of kidney fibrosis in part by upregulating BMP-7-mediated signaling via HOXA13 expression and down regulation of USAG-1.

Developmental Expression of Bone Morphogenetic Protein 2 (BMP2)-Induced Genes in Fetal Mouse Ovaries [A335

INTRODUCTION: Granulosa cells (GCs) and oocytes assemble to form the primordial follicle (PF) reserve. Reduced PF reserve results in consequential infertility and menopause-related disorders. Ovarian transcriptome analysis was done to understand the mechanism of bone morphogenetic proteins (BMPs) or estradiol-17β (E2)induced PF formation. METHODS: RNA from fetal (E16-E19) and postnatal (P1 and P2) mouse ovaries and E16 ovaries cultured with or without BMP2 or E2 for 24 hours were analyzed by RNA sequencing and bioinformatics. In vivo and in vitro expression of selected candidate genes and spatio-temporal expression of corresponding proteins were examined using real-time quantitative reverse transcription polymerase chain reaction (RT-qPCR) and immunofluorescence localization, respectively. RESULTS: Remarkable increases in Gata2, Gata4, Egr2 and Hepsin mRNA were evident on E18 (one day prior to PF formation). Protein immunosignal was present in somatic cells (SC) but the intensity was higher in GCs on E18 through P1. BMP2 or E2 upregulated gene expression in vivo and in vitro; however, the effect of E2 was less for Gata4 and Egr2. BMP2 significantly upregulated all proteins in GCs, whereas E2 upregulated GATA2 and HEPSIN proteins in SCs as well. CONCLUSION: In summary, Gata2, Gata4, Egr2 and Hepsin mRNA levels increase around PF formation. Correspondingly, intensities of immunosignal increase in differentiating GCs. BMP2 or E2 may induce differentiation of ovarian SCs into GCs by cell-specific activation of Gata2, Gata4 and/or Egr2. Hepsin, a serine proteinase, may participate in ovary remodeling. We conclude that BMP2-induced GATA2, GATA4, EGR2 and Hepsin may regulate SC to GC transition, leading to the formation of PFs.

Bone morphogenetic protein 4 alleviates nonalcoholic steatohepatitis by inhibiting hepatic ferroptosis

Nonalcoholic steatohepatitis (NASH) is a state of simple steatosis that progresses to inflammation and liver injury accompanied by ferroptosis. Bone morphogenetic protein 4 (BMP4) plays an important role in adipogenesis and differentiation, as well as in hepatic steatosis and iron regulation. However, the direct impact of BMP4 on NASH remains unclear. In this study, our aim was to investigate the effect of BMP4 on NASH and its underlying mechanism. We first explored BMP4 expression in vivo in mice and patients and in vitro in HepG2 and LO2 cell lines, and then, determined whether ferroptosis occurs in NASH. Further overexpression or inhibition of BMP4 was induced to observe the effect of BMP4 on liver ferroptosis in NASH. BMP4 expression was upregulated in patients and mice with nonalcoholic fatty liver disease, and free fatty acid (FFA)-induced HepG2 and LO2 cell lines. We observed ferroptosis in high-fat diet and high-fructose diet-fed mice and FFA-induced HepG2 and LO2 cell lines. BMP4 overexpressing plasmid was constructed and the HepG2 and LO2 cells were transfected with lentivirus (oe-BMP4), or treated with exogenously added recombinant human BMP4 or BMP antagonist noggin. BMP4 suppressed the markers of hepatic steatosis, liver inflammation, and liver injury. Upregulated BMP4 expression in HepG2 and LO2 cells reduced reactive oxygen species and malondialdehyde content and relieved ferroptosis. Mechanistically, BMP4 overexpression in hepatocytes upregulated the mRNA and protein levels of glutathione peroxidase 4 (GPX4), a central regulator of ferroptosis, while exogenous inhibition of BMP4 by noggin decreased their levels. Immunoprecipitation assays demonstrated a physical interaction between BMP4 and GPX4 in HepG2 and LO2 cells, and confocal imaging confirmed colocalization of BMP4 and GPX4. Consistently, BMP4 overexpression plays an important role in NASH by increasing GPX4 expression, therefore decreasing hepatic ferroptosis. This study proposes BMP4 as a therapeutic target for preventing steatohepatitis.

RNA sequence analysis identified bone morphogenetic protein-2 (BMP2) as a biomarker underlying form deprivation myopia

PurposeForm deprivation myopia (FDM) is an urgent public issue characterized by pathological changes, but the underlying mechanism remained unclear. The aim was to investigate bone morphogenetic proteins (BMPs) utilizing the pathogenesis of FDM. Material and methodsGene expression omnibus (GEO) database was used to analyze one mRNA profile (GSE89325) of FDM. Sixteen retina samples (8 FDM and 8 controls) were randomly divided into seven groups for differential gene expression analysis in R. software. The gene pathway and protein-protein interaction (PPI) analysis were performed by the DAVID and STRING databases. Cytoscape was used to draw the PPI network. The gene ontology (GO) enrichment and Kyoto encyclopedia of genes and Genomes (KEGG) analysis were determined to achieve gene annotation and visualization. ResultsA total of 18420 differentially expressed genes (DEGs) were identified associated with FDM. The only non-significant gene (BEND6) was separately analyzed between two groups. Thirteen hub genes were discovered, ACVR1, ACVR2A, ACVR2B, RGMB, BMPR2, BMPR1A, BMP2, BMPR1B, CHRD, PTH, PTH1R, PTHLH, and WNT9A. The expression alteration in FDM were mainly enriched in cytokine-cytokine, and neuroactive ligand receptor interaction pathways. BMP2 was the key gene in myopia progression. ConclusionsOf clinical perspective, our findings reveal that expression of BMP2 as an underlying mechanism of FDM, providing an insight for therapeutic interventions.

Lysine (K)-specific demethylase 5C regulates the incidence of severe preeclampsia by adjusting the expression of bone morphogenetic protein-7

ABSTRACT This study aimed to investigate the roles of the lysine (K)-specific demethylase 5C (KDM5C)–bone morphogenetic protein-7 (BMP-7) signaling pathway in the pathogenesis of severe preeclampsia (sPE). A total of 180 pregnant patients were enrolled in the study and classified into three groups: an early-onset sPE group (EOsPE) (n = 60), a late-onset sPE group (LOsPE) (n = 60), and a control group (normal pregnancy; n = 60). The messenger RNA (mRNA) and protein expression levels of bone morphogenetic protein receptor II (BMPRII), BMP-7, and KDM5C were detected in placenta samples from the two sPE groups, and their sites were evaluated using immunohistochemistry (IHC). The sPE groups showed an increased KDM5C mRNA expression, and the EOsPE group showed a decreased BMP-7 and BMPRII mRNA expression compared with the LOsPE group. However, contradictory results were discovered in terms of protein expression. Immunostaining of KDM5C, BMP-7, and BMPRII was observed in villous trophoblast and extravillous trophoblast cells. Compared with the control group, the staining intensity of KDM5C in the placental tissue trophoblast cell nucleus and vascular endothelial cells of the sPE groups was weaker, while that of BMP-7 and BMPRII was stronger, and the staining intensity was more subjective in the LOsPE group. Consistent findings were obtained by IHC and Western blot analysis. KDM5C nuclear-cytoplasmic translocation may regulate sPE through BMP-7 and its receptors. The KDM5C–BMP-7 signaling pathway may also lead to less invasion and increased apoptosis of the trophoblast cells, which is involved in the pathogenesis of sPE.

High BMP4 expression in low/intermediate risk BCP-ALL identifies children with poor outcomes

AbstractPediatric B-cell precursor acute lymphoblastic leukemia (BCP-ALL) outcome has improved in the last decades, but leukemic relapses are still one of the main problems of this disease. Bone morphogenetic protein 4 (BMP4) was investigated as a new candidate biomarker with potential prognostic relevance, and its pathogenic role was assessed in the development of disease. A retrospective study was performed with 115 pediatric patients with BCP-ALL, and BMP4 expression was analyzed by quantitative reverse transcription polymerase chain reaction in leukemic blasts at the time of diagnosis. BMP4 mRNA expression levels in the third (upper) quartile were associated with a higher cumulative incidence of relapse as well as a worse 5-year event-free survival and central nervous system (CNS) involvement. Importantly, this association was also evident among children classified as having a nonhigh risk of relapse. A validation cohort of 236 patients with BCP-ALL supported these data. Furthermore, high BMP4 expression promoted engraftment and rapid disease progression in an NSG mouse xenograft model with CNS involvement. Pharmacological blockade of the canonical BMP signaling pathway significantly decreased CNS infiltration and consistently resulted in amelioration of clinical parameters, including neurological score. Mechanistically, BMP4 favored chemoresistance, enhanced adhesion and migration through brain vascular endothelial cells, and promoted a proinflammatory microenvironment and CNS angiogenesis. These data provide evidence that BMP4 expression levels in leukemic cells could be a useful biomarker to identify children with poor outcomes in the low-/intermediate-risk groups of BCP-ALL and that BMP4 could be a new therapeutic target to blockade leukemic CNS disease.

Protein kinase A is activated during bone morphogenetic protein 2-induced osteogenic differentiation of dental follicle stem cells via endogenous parathyroid hormone-related protein

ObjectiveThe aim of this study was to investigate the mechanisms of how protein kinase A (PKA) is activated during bone morphogenetic protein 2 (BMP2)-induced osteogenic differentiation in dental follicle stem cells. DesignHuman dental follicle stem cells were cultured and treated with a BMP2-containing osteogenic differentiation medium or differentiation medium without BMP2. Specific siRNAs and substances/proteins were used to modulate pathways. PKA activity and activity of alkaline phosphatase were determined. Expression of targets was measured by Western Blots and reverse transcription-quantitative polymerase chain reaction, while protein interactions were investigated by immunoprecipitation. Immunofluorescence staining was used for subcellular target localization. ResultsPKA activity is stimulated after osteogenic induction by BMP2. Differentiation medium without BMP2 strongly induces BMP2 gene expression, which correlates with downstream target expression. Elevation of cAMP levels does not affect alkaline phosphatase activity and PKA does not directly interact with Smad 4. However, PKA activation requires expression of parathyroid hormone-related protein (PTHrP), which is stimulated after BMP2-induced differentiation. Furthermore, neither supplementation with PTHrP nor with the receptor antagonist parathyroid hormone (7−34) affects PKA activity. Thus, endogenous PTHrP expression is required for PKA activation and immunofluorescence staining shows that PTHrP is mainly located in the nucleus of dental follicle stem cells. Beyond, knockdown of PKA stimulates the BMP2 signaling pathway and down-stream expression of PTHrP. ConclusionsBMP2-induced osteogenic differentiation activates PKA in dental follicle stem cells via endogenous expression of PTHrP. Additionally, PKA inhibits BMP2 signaling and expression of PTHrP in a negative feedback loop.

BMP-7 ameliorates partial epithelial-mesenchymal transition by restoring SnoN protein level via Smad1/5 pathway in diabetic kidney disease

Tubulointerstitial fibrosis (TIF) is involved in the development of diabetic kidney disease (DKD). Transforming growth factor β1 (TGF-β1) is involved in the extensive fibrosis of renal tissue by facilitating the partial epithelial-mesenchymal transition (EMT), increasing the synthesis of extracellular matrix (ECM), inhibiting degradation, inducing apoptosis of renal parenchyma cells, and activating renal interstitial fibroblasts and inflammatory cells. Recent studies indicated that bone morphogenetic protein-7 (BMP-7) upregulated the expression of endogenous SnoN against renal TIF induced by TGF-β1 or hyperglycemia. Nevertheless, the mechanisms underlying the BMP-7-mediated restoration of SnoN protein level remains elusive. The present study demonstrated the increased expression of BMP-7 in diabetic mellitus (DM) mice by hydrodynamic tail vein injection of overexpressed BMP-7 plasmid, which attenuated the effects of DM on kidney in mice. Partial tubular EMT and the accumulation of Collagen-III were resisted in DM mice that received overexpressed BMP-7 plasmid. Similar in vivo results showed that BMP-7 was competent to alleviate NRK-52E cells undergoing partial EMT in a high-glucose milieu. Furthermore, exogenous BMP-7 activated the Smad1/5 pathway to promote gene transcription of SnoN and intervened ubiquitination of SnoN; both effects repaired the SnoN protein level in renal tubular cells and kidney tissues of DM mice. Therefore, these findings suggested that BMP-7 could upregulate SnoN mRNA and protein levels by activating the classical Smad1/5 pathway to refrain from the partial EMT of renal tubular epithelial cells and the deposition of ECM in DKD-induced renal fibrosis.

Fluoride induces hypomethylation of BMP2 and activates osteoblasts through the Wnt/β-catenin signaling pathway

BackgroundSkeletal fluorosis has become a public health issue in recent years as its serious impact on patients’ life expectancy. Bone morphogenetic protein 2 (BMP2) plays a key role in promoting osteogenesis. However, the mechanism of BMP2-Wnt/β-catenin axis in skeletal fluorosis needs further exploration. MethodsThe RT-qPCR and western blot assay were carried out to examine the mRNA and protein levels. Cell viability was measured by MTT assay. A commercial ALP assay kit was used to detect ALP activities. Alizarin Red staining was performed to measure the formation of mineralized nodules. Methylation-specific PCR (MSP) was performed to measure the methylation level of BMP2. ResultsFluoride promoted the expression of osteogenic marker genes (OPN, OCN, OSX and RUNX2) and induced the proliferation and differentiation of MC3T3-E1 cells. Fluoride induced hypomethylation and high expression of BMP2. Furthermore, knockdown of BMP2 reversed the promoting effect of fluoride on osteogenic differentiation of MC3T3-E1. The expression of β-catenin, glycogen synthase kinase 3β (GSK3β), wingless/integrated 3α (Wnt3α), low-density lipoprotein receptor-related protein 5 (LRP5) and dishevelled 1 (Dv1) were increased in osteoblasts treated with fluoride, however, knockdown of BMP2 reversed this phenomenon. Simultaneous knockdown of BMP2 and β-catenin significantly inhibited the differentiation of osteoblasts induced by fluoride. ConclusionFluoride contributed to proliferation and differentiation of osteoblasts through BMP2-Wnt/β-catenin axis, providing a feasible theoretical basis for the treatment of skeletal fluorosis.

Role of Bone Morphogenetic Protein 4 in the Inflammation of the Myocardium and Vascular Tissue of Obese Mice.

Bone morphogenetic protein 4 (BMP4) is a proinflammatory factor. The expression of BMP4 is reduced in the adipose and enhanced in the myocardium and vascular during obesity. It is possibly involved in the process of inflammatory response of the myocardium and vascular. Obesity, often regarded as a risk factor for cardiovascular diseases, is a kind of inflammatory response. This study aimed to investigate the relationship of BMP4 with obesity and cardiovascular disease. Ob/ob mice were used as the experimental group, and C57BL/6 mice were used as the control group. The two groups were further divided into 2 subgroups based on the mice carrying adenovirus-encoding shRNA for BMP4 or Lac Z genes. The messenger RNA and protein levels of BMP4, interleukin-1β, and interleukin-9 were significantly higher in the myocardial tissue and aorta of ob/ob+ Lac Z shRNA than those in the other 3 groups, whereas the levels in the ob/ob+ BMP4 shRNA group were significantly decreased and comparable with those in the control groups. BMP4 is significantly upregulated in the myocardial tissue and aorta of obese mice, and this suggests that BMP4 is an risk factor involved in the local inflammatory response.

Lower urinary tract symptoms in patients with small prostates: Smooth muscle proliferation and calcification might be causative factors.

The current study is designed to evaluate and compare the histological changes in the surgical samples of prostate taken from patients undergoing transurethral resection of prostate (TURP) for benign prostate hyperplasia (BPH) with different sizes. Prostate surgical tissue samples were obtained from BPH patients undergoing TURP after taking informed consent. Ultrasound measure of prostatic weight and prostate-specific antigen (PSA) levels were obtained from the patients along with other clinical and demographic details. Tissue samples were fixed, processed, sectioned and stained with hematoxylin and eosin and Masson's trichrome to look for histological features, specifically smooth muscle proliferation. Immunohistochemical expression of bone morphogenetic protein (BMP)-2 was recorded to assess the calcification potential. Fifty-nine surgical samples were obtained from the patients of age range 50-90 years and body mass index (BMI) 15.6-33.3kg/m2 . The range of ultrasound measures of prostate weight was 20-137 g with PSA ranged 1.03-93.3ng/mL. Patients with small-sized prostate had significant severe smooth muscle proliferation (P < .001). Prostate size/weight had significant positive association with BMI (P < 0.001, r=0.543) and negative association with BMP-2 (P < 0.001, r=-0.654). Samples with severe smooth muscle proliferation were with increased BMP-2 expression (P < .001) and higher levels of PSA levels (P=0.004). BMP-2 expression revealed positive significant association with PSA (P < .001, r=0.432). From this study we conclude that BPH patients with small-sized glands and high PSA levels have increased smooth muscle proliferation and calcification potential causing the symptoms of lower urinary tract symptoms in these patients.

Pinctada Maxima Pearl Shells as a Promising Bone Graft Material in the World of Dentistry

BACKGROUND: Pinctada maxima pearl shell contains inorganic and organic materials that have a bone-like basic structure that facilitates bone remodeling. AIM: This study aimed to describe the effectiveness of Pinctada maxima pearl shells as bone graft material in the world of dentistry using an animal model. METHODS: Research uses Pinctada maxima pearl shell that was processed into hydroxyapatite Pinctada maxima (HPM) powder. Chemical and surface characteristics of HPM were performed with X-ray fluorescence (XRF), Fourier transform infra-red (FTIR), and X-ray diffraction (XRD), respectively. Thirty male guinea pigs were randomly assigned into three groups: Negative control (NC), positive control, and HPM. After 14–21 days of observation, guinea pigs were sacrificed. Bone formation was seen through immunohistochemical examination of osteoprotegerin (OPG) and bone morphogenetic protein (BMP2) expression. Data were analyzed through Shapiro wills and analysis of variance with a significance level of 5%. RESULTS: There was a high expression of OPG and BMP2 on days 14–21 in the HPM group when compared to the NC group with a significance level of 5%. CONCLUSION: HPM powder can be used as a promising bone graft material in the world of dentistry.

Effect of polycaprolactone scaffolds containing different weights of graphene on healing in large osteochondral defect model

Now it is possible to combine the different biomaterial properties of graphene and 3 D printing scaffolds produced by tissue engineering for cartilage repair. In the study graphene-containing (1, 3, 5, 10 wt%), porous and oriented poly-ε-caprolactone-based scaffolds were prepared by robocasting method to use in the regeneration of large osteochondral defects. The scaffolds were implanted into the full-thickness osteochondral defect in a rabbit model to evaluate the regeneration of the defect in vivo. For this purpose, twenty female New Zealand white rabbits were used and they were euthanized at 4 and 8 weeks of implantation. The reparative osteochondral tissues were harvested from rabbit distal femurs and then processed for gross appearance assessment, radiographic imaging, histopathological, histochemical and immunohistochemical examinations. Results revealed that graphene-containing graft materials caused significant amelioration at the defect areas. Graphene-containing graft materials improved the fibrous, chondroid and osseous tissue regeneration compared to the control group. The expressions of bone morphogenetic protein-2 (BMP-2), collagen-1 (col-1), vascular endothelial growth factor (VEGF) and alkaline phosphatase (ALP) expressions were more prominent in graphene-containing PCL implanted groups (p < .001). Picrosirrius red method was used for to evaluate connective and muscle tissues. Results also revealed that the ameliorative effect of graphene increased by the elevation in concentration. The most prominent healing was observed in 10 wt% graphene-containing PCL based composite scaffold implanted group. This study results showed that graphene-containing PCL scaffolds enhanced the healing significantly in large osteochondral defect areas compared to the control groups.

Uremic Toxin Lanthionine Induces Endothelial Cell Mineralization In Vitro.

Vascular calcification (VC) is a pathological event caused by the unusual deposition of minerals in the vascular system, representing the leading cause of cardiovascular mortality in chronic kidney disease (CKD). In CKD, the deregulation of calcium and phosphate metabolism, along with the effect of several uremic toxins, act as key processes conveying altered mineralization. In this work, we tested the ability of lanthionine, a novel uremic toxin, to promote calcification in human endothelial cell cultures (Ea.hy926). We evaluated the effects of lanthionine, at a concentration similar to that actually detected in CKD patients, alone and under pro-calcifying culture conditions using calcium and phosphate. In pro-calcific culture conditions, lanthionine increased both the intracellular and extracellular calcium content and induced the expression of Bone Morphogenetic Protein 2 (BMP2) and RUNX Family Transcription Factor 2 (RUNX2). Lanthionine treatment, in pro-calcifying conditions, raised levels of tissue-nonspecific alkaline phosphatase (ALPL), whose expression also overlapped with Dickkopf WNT Signaling Pathway Inhibitor 1 (DKK1) gene expression, suggesting a possible role of the latter gene in the activation of ALPL. In addition, treatment with lanthionine alone or in combination with calcium and phosphate reduced Inorganic Pyrophosphate Transport Regulator (ANKH) gene expression, a protective factor toward the mineralizing process. Moreover, lanthionine in a pro-calcifying condition induced the activation of ERK1/2, which is not associated with an increase in DKK1 protein levels. Our data underscored a link between mineral disease and the alterations of sulfur amino acid metabolisms at a cell and molecular level. These results set the basis for the understanding of the link between uremic toxins and mineral-bone disorder during CKD progression.

The potential for induction of the expression of bone morphogenetic protein-7 and alkaline phosphatase in postextraction tooth sockets using a combination of mangosteen peel extract and DFDBBX

The potential for induction of the expression of bone morphogenetic protein-7 and alkaline phosphatase in postextraction tooth sockets using a combination of mangosteen peel extract and DFDBBX

Efficacy of Bushen Culuan decoction on ovarian follicle and follicular granulosa cells in mice with premature ovarian insufficiency induced by tripterygium wilfordii polyglycoside.

To investigate the protective efficacy of Bushen Culuan decoction (BCD) on ovarian follicle and follicular granulosa cells in mice with premature ovarian insufficiency (POI) induced by tripterygium wilfordii polyglycoside, and to study the potential mechanism underlying the action. Eighty female Balb/c mice were randomly divided into 4 groups (n = 20 each): blank group, model group, Bushen Culuan decoction intervening group (BCD group) and estradiol valerate intervening group (EV group). In the first 14 model establishing d, mice in model group, BCD group and EV group were under Tripterygium wilfordii polyglycoside (TWP) gavage to establish POI models. In the 14-day therapeutic stage, mice in BCD group were taken BCD 18.35 mg·kg-1d-1, mice in EV group were taken EV solution 0.15 mg·kg-1d-1, while mice in blank group and model group were taken normal saline. When the mice accomplished therapy, whole blood was collected for serum hormone including follicle stimulating hormone (FSH), luteal hormone (LH), estradiol (E2), antimullerian hormone (AMH) levels and vascular endothelial growth factor (VEGF), bone morphogenetic protein-7 (BMP-7) measurement. Ovarian tissues were harvested for morphologic observation, follicle counting, ovarian follicular graulosa cell apoptosis test and testing BMP-7 and caspase-3 expressions. The body weights of the mice kept growing stably in the process expect in TWP intervening stage. Compared with model group, BCD group had significantly higher ovarian index, serum E2, AMH, VEGF, BMP-7 levels and significantly lower FSH level (P < 0.05). Meanwhile the VEGF level in BCD group was higher than in EV group (P < 0.05). Compared with model group, the histopathological damage and GCs apoptosis were mitigated; developing follicle counting, BMP-7 expression were up-regulated, and caspase-3 expression was downregulated in BCD groups (P < 0.05). BCD treatment could attenuate pathological process in POI ovaries, suppress GC apoptosis, probably through promoting BMP-7 expression and following inhibiting caspase-3 activation.

Function of calcium-rich hydroxyapatite nanoparticles on proliferation and differentiation of bone marrow mesenchymal stem cells (BMSCs) in osteoporosis

Out study was aimed to assess the function of calcium-rich hydroxyapatite nanoparticles on proliferation and differentiation of bone marrow mesenchymal stem cells (BMSCs) in osteoporosis. The structure of calciumrich hydroxyapatite nanoparticles was tested with Transmission electron microscopy (TEM) and cell proliferation was detected with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) method. The activity of Caspase3 was detected with ELISA and content of alkaline phosphatase (AP), osteopontin (OPN) and Runx2 was tested with Western Blot assay. The bone marrow mesenchymal stem cells (BMSCs) proliferation was prompted by calcium-rich hydroxyapatite. Results showed that the activity of ALP was increased, while activity of Caspase3 was reduced. The expression levels of OPN and Runx2 were increased while expression of Bone morphogenetic protein 2 (BMP-2) osteogenesis marker was increased. Moreover, the BMSCs could be differentiated effectively onto osteogenic direction, and proliferation and osteogenic differentiation of BMSCs was prompted effectively by calcium-rich hydroxyapatite. These findings could probably provide a brand-new scheme for clinical treatment of OP.

Regulatory role of dihydrotestosterone on BMP-6 receptors in granular cells of sheep antral follicles.

Bone morphogenetic protein-6 (BMP-6) and dihydrotestosterone (DHT) affect steroid synthesis in follicles and regulate cell proliferation in the ovaries of female animals. However, little is known about granular cells (GCs) in sheep. We identified the key BMP-6 receptors, activin receptor-like kinase(ALK-6), and bone morphogenetic protein receptor type 2 (BMPRII) in sheep follicles using immunohistochemistry (IHC) and immunofluorescence (IF). Both ALK-6 and BMPRII were expressed in the GC layer, GC membranes, and cytoplasm. We evaluated ALK-6 and BMPRII expression at the follicular development stage using quantitative real-time PCR and western blotting to detect sheep GCs from large, medium, and small follicles (diameters of≥5, 2-5, and≤2mm, respectively). The mRNA abundance and protein expression of ALK-6 and BMPRII were significantly higher in GCs from large follicles compared to those in GCs from small follicles (P<0.05) and were the lowest in GCs from medium follicles. To assess whether DHT affects ALK-6 and BMPRII expression in sheep GCs, we cultured GCs from large follicles in vitro then incubated them with DHT (10-11, 10-9, 10-7 M). We found that 10-7-M DHT significantly inhibited ALK-6 and BMPRII mRNA and protein (P<0.05). We further explored whether DHT regulates ALK-6 and BMPRII through the nuclear androgen receptor (AR) pathway and found that 10-6-M flutamide, a non-selective androgen inhibitor, partially relieved the inhibitory effect of 10-7-M DHT on ALK-6 and BMPRII expression. Thus, GCs in sheep antral follicles differentially expressed ALK-6 and BMPRII at various stages, indicating that BMP-6 plays different roles to some extent during the development of antral follicles, and that high concentrations of DHT can inhibit the expression of ALK-6 and BMPRII via the androgen receptor pathway in sheep GCs. The present study aimed to determine the expression of the main BMP-6-related main receptors, namely, ALK-6 and BMPRII, during the development of GCs in sheep antral follicles and a potential mechanism of DHT regulation in sheep GCs. Our findings lay a foundation for the further exploration of the effects of ovarian BMP-6 expression on follicular development.

TJ-M2010-5, A self-developed MyD88 inhibitor, attenuates liver fibrosis by inhibiting the NF-κB pathway

Liver fibrosis is the result of most chronic inflammatory liver damage and seriously endangers human health. However, no drugs have been approved to treat this disease. Previous studies showed that the Toll‐like receptors (TLRs)/myeloid differentiation factor-88 (MyD88)/nuclear factor-κB (NF-κB) pathway plays a key role in liver fibrosis. TJ-M2010-5 is a self-developed small molecule MyD88 inhibitor, which has been proven to have a good protective effect in a variety of inflammatory disease models. In the present study, to investigate the anti-fibrotic effect of TJ-M2010-5, mice were injected with carbon tetrachloride (CCl4) in vivo and LX2 cells (a human hepatic stellate cell line) were treated with TGF-β1 in vitro to induce liver fibrosis. In vivo studies showed that TJ-M2010-5 attenuated the CCl4-induced liver damage, collagen accumulation, and the activation of hepatic stellate cells by inhibiting the nuclear transfer of NF-κB. Moreover, in vitro experiments of LX2 cells stimulated with TGF-β1 further indicated that the NF-κB pathway is involved in the development of liver fibrosis. TJ-M2010-5 significantly inhibited the proliferation and activation of LX2 cells. In addition, TJ-M2010-5 upregulated the expression of bone morphogenetic protein and membrane-bound inhibitor (BAMBI) in LX2 cells by blocking the activation of MyD88/NF-κB, thereby inhibiting the phosphorylation of Smad2/3 and the expression of collagen I (COL1A1) induced by TGF-β1. In conclusion, this study illustrates the anti-hepatic fibrosis effect of TJ-M2010-5 and provides a new treatment method for liver fibrosis.