Bone Morphogenetic Protein Group Research Articles

Osteointegration of functionalised high-performance oxide ceramics: imaging from micro-computed tomography

BackgroundThis study evaluated the osseointegration potential of functionalised high-performance oxide ceramics (HPOC) in isolation or coated with BMP-2 or RGD peptides in 36 New Zeeland female rabbits using micro-computed tomography (micro CT). The primary outcomes of interest were to assess the amount of ossification evaluating the improvement in the bone volume/ total volume (BV/TV) ratio and trabecular thickness at 6 and 12 weeks. The second outcome of interest was to investigate possible differences in osteointegration between the functionalised silanised HPOC in isolation or coated with Bone Morphogenetic Protein 2 (BMP-2) or RGD peptides.Methods36 adult female New Zealand white rabbits with a minimum weight of three kg were used. One-third of HPOCs were functionalised with silicon suboxide (SiOx), a third with BMP-2 (sHPOC-BMP2), and another third with RGD (sHPOC-RGD). All samples were scanned with a high-resolution micro CT (U-CTHR, MILabs B.V., Houten, The Netherlands) with a reconstructed voxel resolution of 10 µm. MicroCT scans were reconstructed in three planes and processed using Imalytics Preclinical version 2.1 (Gremse-IT GmbH, Aachen, Germany) software. The total volume (TV), bone volume (BV) and ratio BV/TV were calculated within the coating area.ResultsBV/TV increased significantly from 6 to 12 weeks in all HPOCs: silanised (P = 0.01), BMP-2 (P < 0.0001), and RGD (P < 0.0001) groups. At 12 weeks, the BMP-2 groups demonstrated greater ossification in the RGD (P < 0.0001) and silanised (P = 0.008) groups. Trabecular thickness increased significantly from 6 to 12 weeks (P < 0.0001). At 12 weeks, BMP-2 promoted greater trabecular thickness compared to the silanised group (P = 0.07), although no difference was found with the RGD (P = 0.1) group.ConclusionSinalised HPOC in isolation or functionalised with BMP-2 or RGD promotes in vivo osteointegration. The sinalised HOPC functionalised with BMP-2 demonstrated the greatest osseointegration.

(274) BMP2 Restores Erectile Dysfunction Through Neurovascular Regeneration and Fibrosis Reduction in Diabetic Mice

Abstract Introduction The odds of erectile dysfunction (ED) are thrice more prevalent in diabetes. Severe peripheral vascular and neural damage in diabetic patients responds poorly to PDE5 inhibitors. However, bone morphogenetic protein 2 (BMP2) is known to be involved in angiogenesis. Objective To assess the efficacy of BMP2 stimulating angiogenesis and augmenting nerve regeneration in a mouse model of diabetic-induced erectile dysfunction. Methods The induction of diabetes mellitus was done by streptozotocin (STZ, 50 mg/kg daily) administered intraperitoneally for five successive days to male C57BL/6 mice that were 8 weeks old. Eight weeks post inductions, animals were allocated to one of five groups: a control group, an STZ-induced diabetic mouse group receiving two intra-cavernous 20 μL PBS injections, or one of three BMP2 groups administered two injections of BMP2 protein (1μg, 5 μg, or 10 μg) diluted in 20 μL of PBS with three days interval between the first and second injection. The erectile functions were assessed two weeks after PBS or BMP2 protein injections by recording the ICP (intracavernous pressure) through cavernous nerve electrical stimulation. Angiogenic activities also nerve regenerating effects of BMP2 were determined in penile tissues, aorta, vena cava, the main pelvic ganglions, the dorsal roots, and from the primary cultured MCECs (mouse cavernous endothelial cells). Moreover, fibrosis-related factors protein expressions were evaluated by western blot. Results Erectile function recovery to 81% of the control value in diabetic mice was found with intra-cavernous BMP2 injection (5 μg/20 μL). Pericytes and also endothelial cells were extensively restored. It was confirmed that angiogenesis was promoted in the corpus cavernosum of diabetic mice treated with BMP2 through increased ex vivo sprouting of aortic rings, vena cava and penile tissues, and migration and tube formation of MCECs. BMP2 protein enhanced cell proliferation and reduced apoptosis in MCECs and penile tissues, and promoted neurite outgrowth in major pelvic ganglia and dorsal root ganglia under high-glucose conditions. Furthermore, BMP2 suppressed fibrosis by reducing MCEC fibronectin, collagen 1, and collagen 4 levels under high-glucose conditions. Conclusions BMP2 modulates neurovascular regeneration and inhibits fibrosis to revive the mice's erection function in diabetic conditions. Our findings propose that the BMP2 protein represents a novel and promising approach to treating diabetes-related erectile dysfunction (ED). Disclosure No.

A Combined Treatment of BMP2 and Soluble VEGFR1 for the Enhancement of Tendon-Bone Healing by Regulating Injury-Activated Skeletal Stem Cell Lineage

Background: Bone morphogenetic protein 2 (BMP2) is an appealing osteogenic and chondrogenic growth factor for promoting tendon-bone healing. Recently, it has been reported that soluble vascular endothelial growth factor (VEGF) receptor 1 (sVEGFR1) (a VEGF receptor antagonist) could enhance BMP2-induced bone repair and cartilage regeneration; thus, their combined application may represent a promising treatment to improve tendon-bone healing. Moreover, BMP2 could stimulate skeletal stem cell (SSC) expansion and formation, which is responsible for wounded tendon-bone interface repair. However, whether the codelivery of BMP2 and sVEGFR1 increases tendon enthesis injury–activated SSCs better than does BMP2 alone needs further research. Purpose: To study the effect of BMP2 combined with sVEGFR1 on tendon-bone healing and injury-activated SSC lineage. Study Design: Controlled laboratory study. Methods: A total of 128 C57BL/6 mice that underwent unilateral supraspinatus tendon detachment and repair were randomly assigned to 4 groups: (1) untreated control group; (2) hydrogel group, which received a local injection of the blank hydrogel at the injured site; (3) BMP2 group, which received an injection of hydrogel with BMP2; and (4) BMP2 with sVEGFR1 group, which received an injection of hydrogel with BMP2 and sVEGFR1. Histology, micro–computed tomography, and biomechanical tests were conducted to evaluate tendon-bone healing at 4 and 8 weeks after surgery. In addition, flow cytometry was performed to detect the proportion of SSCs and their downstream differentiated subtypes, including bone, cartilage, and stromal progenitors; osteoprogenitors; and pro-chondrogenic progenitors within supraspinatus tendon enthesis at 1 week postoperatively. Results: The repaired interface in BMP2 with sVEGFR1 group showed a significantly improved collagen fiber continuity, increased fibrocartilage, greater newly formed bone, and elevated mechanical properties compared with the other 3 groups. There were more SSCs; bone, cartilage, and stromal progenitors; osteoprogenitors; and pro-chondrogenic progenitors in the BMP2 with sVEGFR1 group than that in the other groups. Conclusion: Our study suggests that the combined delivery of BMP2 and sVEGFR1 could promote tendon-bone healing and stimulate the expansion of SSCs and their downstream progeny within the injured tendon-bone interface. Clinical Relevance: Combining BMP2 with sVEGFR1 may be a good clinical treatment for wounded tendon enthesis healing.

Bone morphogenetic protein and cancer in spinal fusion: a propensity score-matched analysis.

Bone morphogenetic protein (BMP) has been increasingly used in spinal surgery to promote arthrodesis. Because BMP stimulates cellular proliferation, its association with tumorigenesis is a concern. Previous research has generated conflicting conclusions on the risk of cancer in patients receiving BMP. The authors aimed to compare the incidence of solid organ and hematopoietic malignancies in patients undergoing spinal arthrodesis with or without BMP. The PearlDiver Mariner Patient Claims Database was queried for patients undergoing thoracolumbar fusion between 2015 and 2021. Patients with preexisting malignancy were excluded. Data were analyzed for incidence of solid organ malignancy and hematopoietic malignancy diagnosed after spinal surgery. Propensity score matching using age, sex, tobacco usage, and year of surgery was performed between patients who did and those who did not receive BMP. Among patients without prior solid organ malignancy, BMP was used in 22,139 patients and not used in 306,249. In the propensity score-matched group, 3.1% of the BMP group developed solid organ malignancy following surgery compared with 3.5% in the non-BMP group. The relative risk (RR) of developing solid organ malignancy after BMP exposure was 0.89 (95% CI 0.81-0.98, p = 0.02). Among patients without prior hematopoietic malignancy, BMP was used in 23,505 patients and not used in 328,796 patients. In the propensity score-matched group, 0.4% of the BMP group developed hematopoietic malignancy compared with 0.6% of the non-BMP group. The RR of developing hematopoietic malignancy after BMP exposure was 0.71 (95% CI 0.55-0.93, p = 0.015). BMP use in thoracolumbar fusion was not associated with an increased risk of new malignancy, which further supports emerging data on the lack of an association between BMP use and increased malignancy.

The Effect of Bone Morphogenetic Protein-2 (BMP-2) on Volumetric and Histometric Outcomes for Peri-Implant Defects in the Animal Model: A Systematic Review and Meta-Analysis.

Purpose: To study the effect of BMP-2 on the volumetric and histometric changes in peri-implant defect treatments in animal models. Materials and Methods: An electronic search of four databases and a manual search of peer-reviewed journals for relevant articles were performed. Animal studies with data that compared the volumetric and/or histometric outcomes of peri-implant defect treatment with and without the use of BMP-2 were included. Meta-analyses were performed to analyze the weighted mean difference (WMD) and confidence interval (CI) for the recorded variables. Results: After completing the search process, 21 randomized controlled trials were included. The results of the meta-analyses showed that the WMD of bone-to-implant contact (%BIC) with 4 to 8 weeks and 12 to 24 weeks of follow-up was 15.50% (95% CI = 3.28% to 27.72%, P = .01) and 16.17% (95% CI = 11.17% to 21.16%, P < .00001), respectively, favoring the BMP-2 group. The WMD for the percentage of defect fill with 4 to 8 weeks and 12 to 24 weeks of follow-up was 15.88% (95% CI = 3.90% to 27.86%, P = .009) and 10.48% (95% CI = 0.95% to 20.02%, P = .03), respectively, favoring the BMP-2 group. The WMD for the vertical bone gain with 8 to 16 weeks of follow-up was 1.63 mm (95% CI = 0.58 to 2.67 mm, P = .002), also favoring the BMP-2 group. Conclusion: This review demonstrated that the use of BMP-2 in treating peri-implant defects showed better clinical and histometric outcomes than defects not treated with BMP-2 in animal models.

Mechanism of the Notch1 signaling pathway regulating osteogenic factor influences lumbar disc calcification

To explore the mechanism of the Notch1 signaling pathway in regulating osteogenic factors and influencing lumbar disc calcification. Primary annulus fibroblasts from SD rats were isolated and subcultured in vitro. The calcification-inducing factors bone morphogenetic protein-2 (BMP-2) and basic fibroblast growth factor (b-FGF) were added to separate groups to induce calcification, which were referred to as the BMP-2 group and the b-FGF group, respectively. A control group was also set up, which was cultured in normal medium. Subsequently, cell morphology and fluorescence identification, alizarin red staining, ELISA, and quantitative real-time polymerase chain reaction (QRT-PCR) were performed to determine the effect of calcification induction. Cell grouping was performed again, including the control group, the calcification group (adding the inducer BMP-2), the calcification + LPS group(adding the inducer BMP-2 and the Notch1 pathway activator LPS), and the calcification + DAPT group (adding the inducer BMP-2 and the Notch1 pathway inhibitor DAPT). Alizarin red staining and flow cytometry were used to detect cell apoptosis, ELISA was used to detect the content of osteogenic factors, and Western blot was used to detect the expression of BMP-2, b-FGF, and Notch1 proteins. The induction factor screening results showed that the number of mineralized nodules in fibroannulus cells in BMP-2 group and b-FGF group was significantly increased, and the increase was greater in the BMP-2 group Meanwhile, ELISA and Western blot results showed that BMP-2, b-FGF and mRNA expression levels of BMP-2, b-FGF and Notch1 in the induced group were significantly increased (P<0.01). The results of the mechanism of Notch1 signaling pathway affecting lumbar disc calcification showed that compared to calcified group, the number of fibroannulus cell mineralization nodules, apoptosis rate, BMP-2, b-FGF content, the expression levels of BMP-2, b-FGF, and Notch1 proteins were further increased significantly However, the number of mineralization nodules, apoptosis rate, BMP-2 and b-FGF levels, BMP-2, b-FGF and Notch1 protein expression levels were decreased in the calcified +DAPT group (P<0.05 or P<0.01). Notch1 signaling pathway promotes lumbar disc calcification through positive regulation of osteogenic factors.

Differentiation of rat bone marrow mesenchymal stem cells into neurons induced by bone morphogenetic protein 7 in vitro

ABSTRACT Objectives Spinal cord injury (SCI) is caused by external direct or indirect factors with high disability rate, which may even endanger the life of patients. To explore the role of bone morphogenetic protein 7 (BMP-7) in the differentiation of rat bone marrow mesenchymal stem cells (BMSCs) into neurons in vitro. Methods BMSCs were isolated and cultured by whole bone marrow adherence method. Adipogenic induction and osteogenic differentiation were used to test the multi⁃directional differentiation ability of BMSCs. Results After 28 days of adipogenic induction, BMSCs showed lipid droplets in the cytoplasm. After osteogenic induction, there were opaque lumps of mineral nodules in BMSCs. There were also orange-red or red mineral nodules in the extracellular matrix. The BMSCs in the 75 ng/ml BMP-7 group were morphologically similar to the neurons. After induction with BMP-7 for 2 h, the NF200 mRNA expression was higher, mRNA expression levels of SYN1, MAP2 and GFAP were higher. Positive rate of immunofluorescence staining in the BMP-7 group was notably increased. The positive rate of NSE immunofluorescence staining in the BMP-7 group was higher. Conclusion BMP-7 can induce rat BMSCs to differentiate into neurons in vitro.

PI3K-Akt signaling regulates BMP2-induced osteogenic differentiation of mesenchymal stem cells (MSCs): A transcriptomic landscape analysis

Bone morphogenetic protein 2 (BMP2) effectively induced mesenchymal stem cells (MSCs) osteogenic differentiation hold great potential for bone tissue engineering. However, a global mechanistic view of BMP2-induced osteogenic differentiation of MSCs remains to be fully elucidated. Here, human umbilical cord-derived MSCs (UC-MSCs) were induced with BMP2, three days and five days later, total RNA were extracted and subjected to RNA-sequencing (RNA-Seq) followed with bioinformatic analysis. Osteogenic differentiation abilities were evaluated with Alkaline phosphatase (ALP) staining and osteogenic differentiation marker expression at both mRNA and protein levels. We identified that adenoviral vectors effectively transduced in UC-MSCs and expressed BMP2 in high efficiency. Both on day 3 and day 5, differentially expressed genes (DEGs) were highly enriched in PI3K-Akt signaling pathway. As for the common DEGs among total BMP2 group vs control group, BMP2 (day 3) versus control (day 3) and BMP2 (day 5) versus control (day 5), there were 105 DGEs and highly enriched in PI3K-Akt signaling pathway. Finally, we found that PI3K-Akt signaling inhibitor dramatically inhibited BMP2-iduced osteogenic differentiation of UC-MSCs. We firstly identified that PI3K-Akt signaling pathway plays a pivotal role in BMP2-induced osteogenic differentiation of MSCs, which may apply a new perspective for BMP2 based bone tissue engineering.

Bone Morphogenetic Protein in Anterior Lumbar Interbody Fusions: A Propensity-Matched Medicare Outcome Analysis.

Bone morphogenetic protein (BMP) is a costly agent commonly used in spine surgery. Its effectiveness and complication profile have never been studied in a large, propensity-matched population following its approval by the Food and Drug Administration for use in single-level anterior lumbar interbody fusion (ALIF) surgeries. To investigate the rate of symptomatic pseudarthrosis or need for revision surgery after single-level stand-alone ALIFs with and without the use of BMP. Medicare Standard Analytic files derived from Medicare parts A and B were used to identify adult patients who underwent single-level ALIF procedures with and without use of BMP between 2004 and 2014. Patients were propensity matched based on their age, gender, and history of diabetes mellitus, hypertension, chronic kidney disease, body mass index greater than 30 kg/m2, smoking, rheumatoid arthritis, and osteoporosis. Sensitivity analysis using adjusted multivariate logistic regression models was also performed. The primary outcomes were the rates of symptomatic pseudarthrosis or need for revision surgery. The propensity-matched population analyzed in this study contained 22,380 patients undergoing single-level ALIF (8971 [40.6%] with BMP and 13,139 [59.4%] without BMP). Both patient groups were balanced at baseline. The rate of symptomatic pseudarthrosis in the propensity-matched analysis was higher in the BMP group (1.9% vs 1.4%, P < 0.05). BMP use during single-level ALIFs was associated with 44% increased odds of developing pseudarthrosis (OR 1.44, 95% CI 1.16-1.76). However, there was no statistically significant difference in the rate of revision surgery between groups (3.7% vs 3.5%, P = 0.49). BMP use in single-level ALIFs may be associated with increased risk of symptomatic pseudarthrosis. Large prospective pragmatic trials are needed to corroborate our findings.

Cytokine co-stimulation effect on odontogenic differentiation of stem cells

The study aims to evaluate the effect of bone morphogenetic protein-2 (BMP-2) and transforming growth factor-beta 1 (TGF-β1) co-stimulation on odontogenic differentiation of human dental pulp stem cells (hDPSCs). The viability/proliferation of hDPSCs treated with BMP-2 (group B), TGF-β1 (group T), or BMP-2/TGF-β1 (group BT) were evaluated. The experiments on odontogenic differentiation were done for 14days. The following subgroups were added to investigate the effect of co-stimulation with different timing: subgroup B1, TGF-β1 co-stimulation in the first week; subgroup B2, TGF-β1 co-stimulation in the second week; subgroup T1, BMP-2 co-stimulation in the first week; and subgroup T2, BMP-2 co-stimulation in the second week. The mineralization was assessed using alizarin red staining. The expression of following genes was assessed using quantitative real-time polymerase chain reaction: dentin sialophosphoprotein (DSPP), dentin matrix protein-1 (DMP1), osteopontin (OPN), and alkaline phosphatase. All groups showed viability similar to the control group (P > .05). The greater mineralization was detected in B groups on day 14. The expressions of DSPP, DMP-1, and OPN increased on day 14 (P < .05). In the combination groups, the higher expressions of DSPP and DMP-1 were observed in subgroups B1 and B2 than groups B and T (P < .05). BMP-2 was the key in odontogenic differentiation of hDPSCs, which was further enhanced by co-stimulation with TGF-β1. Continuous stimulation with TGFβ-1 did not improve the differentiation of hDPSCs. Combined use of the BMP-2 and TGFβ-1 at the specific sequence can provide a tissue engineering approach for the future guided dentin regeneration.

The Effect of Genetically Modified Lactobacillus plantarum Carrying Bone Morphogenetic Protein 2 Gene on an Ovariectomized Rat.

ObjectiveOsteoporosis result from age-related decline in the number of osteoblast progenitors in the bone marrow. Probiotics have beneficial effects on the host, when administered in appropriate amounts. This study investigated the effects of probiotics expressing specific genes, especially the effects of genetically modified bone morphogenetic protein (BMP)-2-expressing Lactobacillus plantarum CJNU 3003 (LP) on ovariectomized rats. MethodsTwenty-eight female Wistar rats (250–300 g, 12 weeks old) were divided into four groups : the sham (control), the ovariectomy (OVX)-induced osteoporosis group (OVX), the OVX and LP (OVX/LP), OVX and genetically modified BMP-2-expressing LP (OVX/LP with BMP) groups. The three groups underwent bilateral OVX and two of these groups were administered two different types of LP via oral gavage daily. At 16 weeks post-OVX, blood was collected from the heart and the bilateral tibiae were extracted and were scanned by ex-vivo micro-computed tomography and stained with hematoxylin-and-eosin (H&E) and Masson’s trichrome stain for pathological assessment. The serum levels of osteocalcin (OC), rat C-telopeptide of type I collagen (CTX-I), BMP-2, and receptor activator of nuclear factor-ĸB ligand (RANKL) were measured. ResultsThe 3D-micro-computed tomography images showed that the trabecular structure in the OVX/LP with BMP group was maintained compared with OVX and OVX/LP groups. No significant differences were detected in trabecular thickness (Tb.Th) between control and OVX/LP with BMP groups (p>0.05). Furthermore, a tendency toward increased BMD, trabecular bone volume, Tb.Th, and trabecular number and decreased trabecular separation was found in rats in the OVX/LP with BMP groups when compared with the OVX and OVX/LP groups (p>0.05). The H&E and Masson’s trichrome stained sections showed a thicker trabecular bone in the OVX/LP with BMP group compared with the OVX and OVX/LP groups. There was no difference in serum levels of OC, CTX and RANKL control and OVX/LP with BMP groups (p>0.05). In contrast, significant differences were found in OC and CTX-1 levels between the OVX and OVX/LP with BMP groups (p<0.05). ConclusionOur results showed that the expression of genetically modified BMP-2 showed inhibition effect for bone loss in a rat model of osteoporosis.

Comparative Evaluation of Efficacy between Recombinant Human Bone Morphogenetic Protein-2 Impregnated with Absorbable Sponge and Platelet-Rich Fibrin in the Treatment of Grade II Furcation Defects: A Randomized Controlled Trial.

Aim and Objectives:The objective of the study was to clinically and radiographically compare and evaluate the regenerative potential of recombinant human bone morphogenetic protein-2 (RhBMP-2) impregnated with absorbable collagen sponge and platelet-rich fibrin (PRF) in the treatment of Grade II furcation defects.Patients and Methods:Thirty-two subjects were randomly assigned to each of the following groups: bone morphogenetic protein (BMP) group and PRF group, with one defect/subject. Sixteen Grade II furcation defects were treated with RhBMP-2 impregnated with absorbable collagen sponge in the BMP group and the remaining 16 defects were treated with PRF in the PRF group. Clinical and radiographic parameters which were evaluated at baseline, postoperative 1 week, 3 months, and 6 months were probing pocket depth, clinical attachment level, scoring of plaque index, and gingival index, and the bone fill was evaluated using Digital Subtraction technique and morphometric area analysis with ImageJ® software.Results:RhBMP-2 in absorbable collagen sponge was effective in increasing the bone fill in Grade II furcation defects when compared to PRF alone (P = 0.05). In relation to clinical parameters, both the groups showed no statistical significance between them.Conclusion: The unique regenerative potential of RhBMP-2 impregnated with absorbable collagen sponge makes it a potential agent to be used as a graft material for the treatment of Grade II furcation defects.

Effect of bone morphogenetic protein-2 on diabetic retinopathy and its mechanism of action

Purpose: To investigate the effect of bone morphogenetic protein-2 (BMP-2) on human retinal vascular endothelial cells (RECs) and human retinal pigment epithelial cells (RPE) cultured in high glucose (HG) in vitro, and the underlying mechanism.&#x0D; Methods: Cell counting kit-8 (CCK-8) was used to determine cell proliferation while Western blot was used to assay the expressions of extracellular matrix and angiogenesis-related factors, Expressions of cytokines and chemokines were assessed by quantitative real time polymerase chain reaction (qRTPCR) and enzyme-linked immunosorbent assay (ELISA). Changes in Smad, ERK, JNK and p38MAPK signal pathway were measured by transfection and interference.&#x0D; Results: The level of expression of BMP-2 in HG group was higher than that in normal glucose (NG) culture group. The expressions of angiogenesis-related factors i.e. vascular endothelial growth factor (VEGF) and intercellular cell adhesion molecule-1 (ICAM1), pro-inflammatory factors i.e. IL-6 and chemokine monocyte chemokine protein-1 (MCP1), increased significantly in HG group compared to NG and HG + BMP-2 groups. Phosphorylation of Smad1/5/8 and activation of ERK, JNK and p38MAPK signaling pathways were enhanced by BMP-2.&#x0D; Conclusion: These results suggest that BMP-2 promotes angiogenesis and enhances the expressions of inflammatory cytokines via Smad signaling pathway.

Sustained Release of Bone Morphogenetic Protein-2 through Alginate Microbeads Enhances Bone Regeneration in Rabbit Tibial Metaphyseal Defect Model.

Bone morphogenetic protein-2 (BMP-2) is widely used to enhance bone regeneration. However, because of its short half-life and rapid disappearance, large amounts of BMP-2 are needed, leading to unintended side effects. In this study, BMP-2-encapsulated alginate microbeads (AM) were used to enhance bone regeneration. Enzyme-linked immunosorbent assay confirmed the sustained release of BMP-2 from AM. Vascular endothelial growth factor (VEGF)-adsorbing aptamer-conjugated hydroxyapatite (Apt-HA) was used for osteoconduction and dual delivery of VEGF and BMP-2. For in vivo bone regeneration evaluation, the grafts (1) Apt-HA + phosphate-buffered saline (PBS), (2) Apt-HA + AM without BMP-2, (3) Apt-HA + BMP-2, and (4) Apt-HA + AM encapsulated with BMP-2 were implanted into rabbit tibial metaphyseal defects. After four weeks, micro-computed tomography (CT), histological, and histomorphometric analyses were performed to evaluate bone regeneration. The Apt-HA + AM with BMP-2 group revealed a significantly higher new bone volume and bone volume/total volume (BV/TV) in both cortical and trabecular bone than the others. Furthermore, as evaluated by histomorphometric analysis, BMP-2 AM exhibited a significantly higher bone formation area than the others, indicating that AM could be used to efficiently deliver BMP-2 through sustained release. Moreover, the combined application of BMP-2-encapsulated Apt-HA + AM may effectively promote bone regeneration.

Recombinant human bone morphogenetic protein-4 enhances tendon-to-bone attachment healing in a murine model of rotator cuff tear.

Injuries of tendon-to-bone attachments (TBA) are common clinical challenges. Bone morphogenetic protein-4 (BMP-4) is potent in chondrogenesis. However, studies focusing on the influence of BMP-4 on the healing of TBA are scarce. Thus, this study's objective was to explore the effect of BMP-4 on the healing of TBA in a murine model of rotator cuff tear. An injury model of the supraspinatus tendon (SST) insertion was established on a total of 120 mature C57 black (BL)/6 mice (12 weeks old), who were then randomly allocated into 3 groups: BMP-4, noggin (an inhibitor of all BMP activities), and control, At weeks 2 and 4 after surgery, the supraspinatus tendon-humerus complexes (SSTHC) were harvested for microradiographic, histologic, immunofluorescent, and biomechanical evaluations. Radiographic data showed that BMP-4 was able to improve the quality of subchondral bone, manifested as higher bone volume fraction (BV/TV), trabecular number (Tb.N), trabecular thickness (Tb.Th), and lower trabecular spacing (Tb.Sp). Histologically, the BMP-4 group at week-2 and -4 showed a better TBA healing interface, characterized by better organizational integration and remodeling, thicker fibrocartilage layer, and more fibrocartilage cells. Immunofluorescence evaluation demonstrated that the number of SOX 9 positive cells in the BMP-4 group was significantly more than that in the control or noggin groups at postoperative weeks 2 and 4 (P<0.05 for all). Mechanical testing results at postoperative weeks 4 demonstrated the failure load, and stiffness in the BMP-4 group were significantly higher (P<0.05 for both), while in the noggin group were significantly lower (P<0.05 for both), compared to the control group. The BMP-4 might enhance TBA healing by promoting the regeneration of fibrocartilaginous enthesis and mineralization, while this process was inhibited by noggin. Thus, BMP-4 may be a potential therapy to augment TBA healing and finally lead to more rapid rehabilitation and reduce recurrent injury risk.

Improved osseointegration using porcine xenograft compared to demineralized bone matrix for the treatment of critical defects in a small animal model.

Autograft (AG) is the gold standard bone graft due to biocompatibility, osteoconductivity, osteogenicity, and osteoinductivity. Alternatives include allografts and xenografts (XG). We investigated the osseointegration and biocompatibility of a decellularized porcine XG within a critical defect animal model. We hypothesized that the XG will result in superior osseointegration compared to demineralized bone matrix (DBM) and equivalent immune response to AG. Critical defects were created in rat femurs and treated with XG, XG plus bone morphogenetic protein (BMP)-2, DBM, or AG. Interleukin (IL)-2 and IFN-gamma levels (inflammatory markers) were measured from animal blood draws at 1week and 1month post-operatively. At 1month, samples underwent micro-positron-emission tomography (microPET) scans following 18-NaF injection. At 16weeks, femurs were retrieved and sent for micro-computerized tomography (microCT) scans for blinded grading of osseointegration or were processed for histologic analysis with tartrate resistant acid phosphatase (TRAP) and pentachrome. Enzyme linked immunosorbent assay testing demonstrated greater IL-2 levels in the XG vs. AG 1week post-op; which normalized by 28days post-op. MicroPET scans showed increased uptake within the AG compared to all groups. XG and XG+BMP-2 showed a trend toward increased uptake compared with DBM. MicroCT scans demonstrated increased osseointegration in XG and XG+BMP groups compared to DBM. Pentachrome staining demonstrated angiogenesis and endochondral bone formation. Furthermore, positive TRAP staining in samples from all groups indicatedbone remodeling. These data suggest that decellularized and oxidized porcine XG is biocompatible and at least equivalent to DBM in the treatment of a critical defect in a rat femur model.

Joint regulation of Wnt and bone morphogenetic protein signaling pathways to promote differentiation of human induced pluripotent stem cells into cardiomyocytes

To explore the role of joint regulation of Wnt and bone morphogenetic protein (BMP) signaling pathways in the differentiation of human induced pluripotent stem cells (hiPSCs) into cardiomyocytes. HiPSCs were cultured and observed under inverted phase contrast microscope. Immunofluorescence staining was used to observe the expressions of hiPSCs pluripotent markers (OCT3/4, NANOG, and TRA-1-60). HiPSCs were passaged which were taken for subsequent experiments within the 35th passage. When the fusion degree of hiPSCs was close to 100%, the CHIR99021 (Wnt pathway activator) was added on the 0th day of differentiation. Different concentrations of IWP4 (inhibitor of Wnt production) were added on the 3rd day of differentiation, and the best concentration of IWP4 was added at different time points. The optimal concentration and the best effective period of IWP4 were obtained by detecting the expression of troponin T (TNNT2) mRNA by real-time fluorescence quantitative PCR. Then, on the basis of adding CHIR99021 and IWP4, different concentrations of BMP-4 were added on the 5th day of differentiation, and the best concentration of BMP-4 was added at different time points. The optimal concentration and best effective period of BMP-4 were obtained by detecting the expression of TNNT2 mRNA. Finally, hiPSCs were divided into three groups: Wnt group, BMP group, and Wnt+BMP group. On the basis of adding CHIR99021 on the 0th day of differentiation, IWP4, BMP-4, and IWP4+BMP-4 were added into Wnt group, BMP group, and Wnt+BMP group respectively according to the screening results. Cells were collected on the 7th and the 15th days of differentiation. The expressions of myocardial precursor cell markers [ISL LIM homeobox 1 (ISL1), NK2 homeobox 5 (NKX2-5)] and cardiomyocyte specific markers [myocyte enhancer factor 2C (MEF2C), myosin light chain 2 (MYL2), MYL7, and TNNT2] were detected by real-time fluorescent quantitative PCR. Cells were collected on the 28th day of differentiation, and the expression of cardiac troponin T (cTnT) was detected by flow cytometry and immunofluorescence staining. The results of cell mophology and immunoflurescence staining showed that the OCT3/4, NANOG, and TRA-1-60 were highly expressed in hiPSCs, which suggested that hiPSCs had characteristics of pluripotency. The optimal concentration of IWP4 was 10.0 μmol/L ( P<0.05) and the best effective period was the 3rd day ( P<0.05) in inducing hiPSCs to differentiate into cardiomyocytes. The optimal concentration of BMP-4 was 20.0 ng/mL ( P<0.05) and the best effective period was the 3rd day ( P<0.05). The relative expressions of ISL1, NKX2-5, MEF2C, MYL2, MYL7, and TNNT2 mRNAs, the positive expression ratio of cTnT detected by flow cytometry, and sarcomere structure detected by immunofluorescence staining of Wnt+BMP group were superior to those of Wnt group ( P<0.05). Joint regulation of Wnt and BMP signaling pathways can improve the differentiation efficiency of hiPSCs into cardiomyocytes.

Serum miRNA-142 and BMP-2 are markers of recovery following hip replacement surgery for femoral neck fracture.

The present study aimed to investigate the changes in miRNA-142 and bone morphogenetic protein-2 (BMP-2) expression before and after hip replacement surgery, and to determine their association with receptor activator of NF-κB ligand (RANKL) and osteoprotegerin (OPG). For this purpose, 142 cases of hip arthroplasty in patients with femoral neck fracture were selected as the research group, and 50 cases of healthy individuals who underwent a physical examination during the same time period were selected the control group. Serum miR-142 and BMP-2 levels were measured by RT-qPCR before and after surgery in the research group and the control group. Serum RANKL and OPG levels were detected before and after surgery by enzyme-linked immunosorbent assay (ELISA). The levels of serum miR-142 and BMP-2 in the research group were significantly lower than those in the control group. At 1 month after surgery, the levels of serum miR-142 and BMP-2 in the research group were significantly higher than those before surgery, and at 6 months after surgery, and they were higher than those in the control group. Pearson's correlation analysis revealed that the serum levels of miR-142 positively correlated with the BMP-2 levels before surgery, at 1 month after surgery, and at 6 months after surgery in the research group. The results of ROC curve analysis revealed that the AUC values of serum miR-142 and BMP-2 were 0.911 and 0.861, respectively. At 1 month after surgery, the levels of serum miR-142 and BMP-2 in group A (patients with a good or excellent recovery) were significantly higher than those in group B (patients with a fair or poor recovery). The levels of serum RANKL and OPG in the research group significantly increased at 1 month after surgery. The serum levels of miR-142 and BMP-2 positively correlated with those of RANKL and OPG before surgery and at 1 month after surgery. On the whole, the findings of the present study indicate that pre-operative serum miR-142 and BMP-2 levels are valuable for evaluating the post-operative recovery of patients with femoral neck fracture undergoing hip replacement surgery. In addition, in the present study, at 1 month after surgery, the levels of both miR-142 and BMP-2 were related to the recovery of the patients, and positively correlated with the RANKL and OPG levels.

Bone morphogenetic protein 2-enhanced osteogenic differentiation of stem cell spheres by regulation of Runx2 expression.

Bone morphogenetic protein 2 (BMP-2) is a growth factor that is used to induce osteogenic differentiation in stem cells. The present study assessed the effects of BMP-2 on stem cell spheroid morphology, viability and osteogenic differentiation. Stem cell spheres were constructed and treated with BMP-2 at predetermined concentrations (0-100 ng/ml) using concave microwells. Cell viability was qualitatively and quantitatively analyzed via microscopy and a water-soluble tetrazolium salt assay kit, respectively. Alkaline phosphatase activity was assessed and an anthraquinone dye for calcium deposit evaluation was performed to determine osteogenic differentiation. The expressions of (runt-related transcription factor 2) and collagen 1 were also determined via quantitative PCR. Spherical shapes were formed using concave microwells on day 1, which were maintained up to day 21. On day 1, the relative cell viability of 0, 10 and 100 ng/ml BMP-2 treated cells was 100.0±1.9, 97.3±4.4 and 101.3±2.6%, respectively. Significantly higher values for alkaline phosphatase activity were determined in the 100 ng/ml treated group when compared with the control group. Additionally, Runx2 mRNA levels were significantly higher in the 100 ng/ml BMP-2 group compared with the control group, as determined via quantitative PCR. The results of the present study indicated that BMP-2 enhanced the differentiation of stem cell spheres, which was demonstrated by increased alkaline phosphatase activity and Runx2 expression.

Electrospun Fibers Immobilized with BMP-2 Mediated by Polydopamine Combined with Autogenous Tendon to Repair Developmental Dysplasia of the Hip in a Porcine Model.

PurposeDevelopmental dysplasia of the hip (DDH) can increase the pressure between the joints, which causes secondary hip osteoarthritis. The aim of the present study was to fabricate poly(D, L-lactic acid)-poly(ethylene glycol)-poly(D, L-lactic acid) (PELA) electrospun fibrous scaffolds, immobilized with bone morphogenetic protein-2 (BMP-2), to repair the acetabulum defects.MethodsThe characteristics of PELA electrospun were analyzed using scanning electron microscopy, the release kinetics of BMP-2 in vitro were analyzed using enzyme-linked immunosorbent assays. Human mesenchymal stem cells (hMSCs) were used for in vitro experiments, the biocompatibility of the electrospinning materials was investigated using a cell counting kit-8 (CCK-8) kit, and osteogenic differentiation was tested via alkaline phosphatase (ALP) activity and relative gene expression. Eighteen miniature pig animal models were used in the in vivo experiment. The pigs were sacrificed at 24 weeks after surgery, and the reconstructed acetabulum was evaluated using histological sections.ResultsStructural analysis revealed that the diameter of the PELA electrospun fiber was 0.8195 ± 0.16 μm. The PELA electrospun fiber materials showed good hydrophilicity and biocompatibility and could continuously release BMP-2 within 27 days: 16.07 ± 0.11 ng of BMP-2 was released from the scaffold. A total of sixteen implants fully filled the defects, and hematoxylin and eosin staining and Goldner’s trichrome staining showed that the simple tendon group (T group) was mostly fibrous tissues, lots of fibroblasts and small amounts of chondrocytes were observed in the polydopamine-coated electrospun fiber group (DP group). The DP plus BMP-2 (DPB) group showed a large number of chondrocytes and partial new bone tissues.ConclusionPELA electrospun fibrous scaffolds are good sustained-release carriers, which can not only induce implant differentiation into cartilage and bone but also are completely degraded without toxicity. Therefore, the material can be used for bone and cartilage regeneration.