Effects of exogenous recombinant human bone morphogenic protein-7 on the corneal epithelial mesenchymal transition and fibrosis

AIM：To evaluate the effect of exogenous recombinant human bone morphogenic protein-7（rhBMP-7）on transforming growth factor-β（TGF-β）-induced epithelial mesenchymal cell transition（EMT）and assessed its antifibrotic effect via topical application.METHODS：The cytotoxic effect of rhBMP-7 was evaluated and the EMT of human corneal epithelial cells（HECEs）was induced by TGF-β. HECEs were then cultured in the presence of rhBMP-7 and/or hyaluronic acid（HA）. EMT markers,fibronectin,E-cadherin,α-smooth muscle actin（α-SMA）,and matrix metaloproteinase-9（MMP-9）,were evaluated. The level of corneal fibrosis and the reepithelization rate were evaluated using a rabbit keratectomy model. Expression of α-SMA in keratocytes were quantified following treatment with different concentrations of rhBMP-7.RESULTS：Treatment with rhBMP-7 attenuated TGF-β-induced EMT in HECEs. It significantly attenuated fibronectin secretion（31.6%; P〈0.05）,the α-SMA protein level（72.2%; P〈0.01）,and MMP-9 expression（23.6%,P〈0.05）in HECEs compared with cells grown in the presence of TGF-β alone. E-cadherin expression was significantly enhanced（289.7%; P〈0.01）in the presence of rhBMP-7. Topical application of rhBMP-7 combined with 0.1% HA significantly reduced the amount of α-SMA~＋ cells by 43.18%（P〈0.05）at a concentration of 2.5 μg/mL and by 47.73%（P〈0.05）at 25 μg/mL,compared with the control group,without disturbing corneal reepithelization.CONCLUSION：rhBMP-7 attenuates TGF-β-induced EMT in vitro,and topical application of rhBMP-7 reduces keratocyte myodifferentiation during the early wound healing stages in vivo without hindering reepithelization. Topical rhBMP-7 application as biological eye drops seems to be feasible in diseases involving TGF-β-related corneal fibrosis with corneal reepithelization disorders.

Three-dimensional bioprinting of multicell-laden scaffolds containing bone morphogenic protein-4 for promoting M2 macrophage polarization and accelerating bone defect repair in diabetes mellitus

Critical-sized bone defect repair in patients with diabetes mellitus remains a challenge in clinical treatment because of dysfunction of macrophage polarization and the inflammatory microenvironment in the bone defect region.Three-dimensional(3D)bioprinted scaffolds loaded with live cells and bioactive factors can improve cell viability and the inflammatory microenvironment and further accelerating bone repair.Here,we used modified bioinks comprising gelatin,gelatin methacryloyl(GelMA),and 4-arm poly(ethylene glycol)acrylate(PEG)to fabricate 3D bioprinted scaffolds containing BMSCs,RAW264.7 macrophages,and BMP-4-loaded mesoporous silica nanoparticles(MSNs).Addition of MSNs effectively improved the mechanical strength of GelMA/gelatin/PEG scaffolds.Moreover,MSNs sustainably released BMP-4 for long-term effectiveness.In 3D bioprinted scaffolds,BMP-4 promoted the polarization of RAW264.7 to M2 macrophages,which secrete anti-inflammatory factors and thereby reduce the levels of pro-inflammatory factors.BMP-4 released from MSNs and BMP-2 secreted from M2 macrophages collectively stimulated the osteogenic differentiation of BMSCs in the 3D bioprinted scaffolds.Furthermore,in calvarial critical-size defect models of diabetic rats,3D bioprinted scaffolds loaded with MSNs/BMP-4 induced M2 macrophage polarization and improved the inflammatory microenvironment.And 3D bioprinted scaffolds with MSNs/BMP-4,BMSCs,and RAW264.7 cells significantly accelerated bone repair.In conclusion,our results indicated that implanting 3D bioprinted scaffolds containing MSNs/BMP-4,BMSCs,and RAW264.7 cells in bone defects may be an effective method for improving diabetic bone repair,owing to the direct effects of BMP-4 on promoting osteogenesis of BMSCs and regulating M2 type macrophage polarization to improve the inflammatory microenvironment and secrete BMP-2.

Bone morphogenic protein signaling in spinal cord injury

Spinal cord injury(SCI)is a debilitating injury that results from traumatic or non-traumatic insults to the spinal cord,causing significant impairment of the patient's activity and quality of life.Bone morphogenic proteins(BMPs)are a group of polyfunctional cytokines belonging to the transforming growth factor beta superfamily that regulates a wide variety of cellular functions in healthy and disease states.Recent studies suggest that dysregulation of BMP signaling is involved in neuronal demyelination and death after traumatic SCI.The focus of this article is to describe our current understanding of the role of BMP signaling in the regulation of cell fate,proliferation,apoptosis,autophagy,and inflammation in traumatic SCI.First,we will describe the expression of BMPs and pattern of BMP signaling before and after traumatic SCI in rodent models and in vitro.Next,we will discuss the role of BMP in the regulation of neuronal and glial cell differentiation,survival,functional recovery from traumatic SCI,and the gap in knowledge in this area that requires further investigation to improve SCI prognosis.

KDM6B epigenetically regulates odontogenic differentiation of dental mesenchymal stem cells

Mesenchymal stem cells （MSCs） have been identified and isolated from dental tissues, including stem cells from apical papilla, which demonstrated the ability to differentiate into dentin-forming odontoblasts. The histone demethylase KDM6B （also known as JMJD3） was shown to play a key role in promoting osteogenic commitment by removing epigenetic marks H3K27me3 from the promoters of osteogenic genes. Whether KDM6B is involved in odontogenic differentiation of dental MSCs, however, is not known. Here, we explored the role of KDM6B in dental MSC fate determination into the odontogenic lineage. Using shRNA-expressing lentivirus, we performed KDM6B knockdown in dental MSCs and observed that KDM6B depletion leads to a significant reduction in alkaline phosphate （ALP） activity and in formation of mineralized nodules assessed by Alizarin Red staining. Additionally, mRNA expression of odontogenic marker gene SP7 （osterix, OSX）, as well as extracellular matrix genes BGLAP （osteoclacin, OCN） and SPP1 （osteopontin, OPN）, was suppressed by KDM6B depletion. When KDM6B was overexpressed in KDM6B-knockdown MSCs, odontogenic differentiation was restored, further confirming the facilitating role of KDM6B in odontogenic commitment. Mechanistically, KDM6B was recruited to bone morphogenic protein 2 （BMP2） promoters and the subsequent removal of silencing H3K27me3 marks led to the activation of this odontogenic master transcription gene. Taken together, our results demonstrated the critical role of a histone demethylase in the epigenetic regulation of odontogenic differentiation of dental MSCs. KDM6B may present as a potential therapeutic target in the regeneration of tooth structures and the repair of craniofacial defects.

Supercritical fluid-assisted controllable fabrication of open and highly interconnected porous scaffolds for bone tissue engineering

Recently tremendous progress has been evidenced by the advancements in developing innovative three-dimensional(3 D)scaffolds using various techniques for addressing the autogenous grafting of bone. In this work, we demonstrated the fabrication of porous polycaprolactone(PCL) scaffolds for osteogenic differentiation based on supercritical fluid-assisted hybrid processes of phase inversion and foaming. This eco-friendly process resulted in the highly porous biomimetic scaffolds with open and interconnected architectures. Initially, a 2^3 factorial experiment was designed for investigating the relative significance of various processing parameters and achieving better control over the porosity as well as the compressive mechanical properties of the scaffold. Then, single factor experiment was carried out to understand the effects of various processing parameters on the morphology of scaffolds. On the other hand, we encapsulated a growth factor, i.e., bone morphogenic protein-2(BMP-2), as a model protein in these porous scaffolds for evaluating their osteogenic differentiation. In vitro investigations of growth factor loaded PCL scaffolds using bone marrow stromal cells(BMSCs) have shown that these growth factor-encumbered scaffolds were capable of differentiating the cells over the control experiments. Furthermore, the osteogenic differentiation was confirmed by measuring the cell proliferation, and alkaline phosphatase(ALP) activity, which were significantly higher demonstrating the active bone growth. Together, these results have suggested that the fabrication of growth factor-loaded porous scaffolds prepared by the eco-friendly hybrid processing efficiently promoted the osteogenic differentiation and may have a significant potential in bone tissue engineering.

Model-based Comparative Prediction of Transcription-Factor Binding Motifs in Anabolic Responses in Bone

Understanding the regulatory mechanism that controls the alteration of global gene expression patterns continues to be a challenging task in computational biology. We previously developed an ant algorithm, a biologically-inspired computational technique for microarray data, and predicted putative transcription-factor binding motifs （TFBMs） through mimicking interactive behaviors of natural ants. Here we extended the algorithm into a set of web-based software, Ant Modeler, and applied it to investigate the transcriptional mechanism underlying bone formation. Mechanical loading and administration of bone morphogenic proteins （BMPs） are two known treatments to strengthen bone. We addressed a question： Is there any TFBM that stimulates both ＂anabolic responses of mechanical loading＂ and ＂BMP-mediated osteogenic signaling＂？ Although there is no significant overlap among genes in the two responses, a comparative model-based analysis suggests that the two independent osteogenic processes employ common TFBMs, such as a stress responsive element and a motif for peroxisome proliferator-activated receptor （PPAR）. The post-modeling in vitro analysis using mouse osteoblast cells supported involvements of the predicted TFBMs such as PPAR, Ikaros 3, and LMO2 in response to mechanical loading. Taken together, the results would be useful to derive a set of testable hypotheses and examine the role of specific regulators in complex transcriptional control of bone formation.

Differential expression of Bmp2,Bmp4 and Bmp3 in embryonic development of mouse anterior and posterior palate

Background The palate is differently regulated and developed along the anterior-posterior axis. The Bmp signal pathway plays a crucial role in palatogenesis. Conditioned-inactivation of Bmp type I receptor Alk2 or Alk3 in the neural crest or craniofacial region leads to palatal cleft in mice. However, how different Bmp members are involved in palatogenesis remains to be elucidated. In the present study, mRNA expression patterns of Bmp2, Bmp3 and Bmp4 in the developing anterior and posterior palates were examined and compared, focusing on the fusion stage.Methods To detect the expression of Bmp mRNA, antisense riboprobes were synthesized by in vitro transcription. Radioactive in situ hybridization was performed on sagital and coronal sections of mice head from E13 to E18.Results The expression of these Bmps were developmentally regulated in the anterior and posterior palates prior to, during and after palatal fusion. During palatal fusion, Bmp4 expression shifted from the anterior to the process, pattern whereas in their palates regulatingConclusions Bmp signalling is involved in palatogenesis in muhiple stages and has muhiple roles in regulating anterior and posterior palatal development. Disturbances of Bmp signalling during palatogenesis might be a possible mechanism of cleft palate.

Tissue engineering in mandibular reconstruction: osteogenesis-inducing scaffolds

Currently, the gold standard for aesthetic and functional reconstruction of critical mandibular defects is an autologous fibular flap;however, this carries risk of donor site morbidity, and is not a promising option in patients with depleted donor sites due to previous surgeries. Tissue engineering presents a potential solution in the design of a biomimetic scaffold that must be osteoconductive, osteoinductive, and support osseointegration. These osteogenesis-inducing scaffolds are most successful when they mimic and interact with the surrounding native macro- and micro-environment of the mandible. This is accomplished via the regeneration triad: (1) a biomimetic, bioactive osteointegrative scaffold, most likely a resorbable composite of collagen or a synthetic polymer with collagen-like properties combined with beta-tri calcium phosphate that is 3D printed according to defect morphology;(2) growth factor, most frequently bone morphogenic protein 2 (BMP-2);and (3) stem cells, most commonly bone marrow mesenchymal stem cells. Novel techniques for scaffold modification include the use of nano-hydroxyapatite, or combining a vector with a biomaterial to create a gene activated matrix that produces proteins of interest (typically BMP-2) to support osteogenesis. Here, we review the current literature in tissue engineering in order to discuss the success of varying use and combinations of scaffolding materials (i.e., ceramics, biological polymers, and synthetic polymers) with stem cells and growth factors, and will examine their success in vitro and in vivo to induce and guide osteogenesis in mandibular defects.