Study On Black Phosphorus Nanosheets/Nano Magnesium Oxide Incorporated Composite Hydrogels For Promoting Bone Repair And Related Mechanisms

Bone defects caused by trauma,tumor resection,infection and congenital diseases still face great challenges in clinical treatment.Although bone tissues have a certain self-healing property,when the bone defect exceeds the critical value,the self-repairing ability is not enough to complete bone regeneration,and intervention measures are often required to promote bone regeneration.Currently,clinical strategies for the treatment of large-scale bone defects include autologous bone grafting,allogeneic bone grafting,and artificial bone scaffolds.Autologous bone graft is the gold standard for the treatment of bone defects,but it has disadvantages such as limited sources and damage to the donor.Allogeneic bone graft also has shortcomings such as potential disease transmission and immunological rejection.The artificial bone scaffolds can effectively solve the above problems due to the advantages of a wide range of material sources,controllable components and high biosafety.The construction of artificial bone repair scaffolds with biomimetic bone tissue extracellular matrix structure and function to accelerate bone repair is the focus of research in the field of bone tissue engineering.Bone tissue is a natural nanocomposite featuring organic proteins（mainly collagen type I）,inorganic minerals（mainly calcium phosphate）and multiple cell types.When bone defects occur,mesenchymal stem cells play an important role in the repair of bone defects,they can migrate to the defect and differentiate into osteoblasts.Osteoblasts secrete unmineralized collagen and alkaline phosphatase,which promotes the deposition of minerals along collagen fibers,thereby achieving bone regeneration.Currently,methods of directly loading stem cells into biomaterials or promoting stem cell migration by growth factors have the effect of promoting bone regeneration,but such methods are often limited by complex regulatory requirements in clinical translational application.Therefore,improving the physical and chemical properties of bone repair materials to recruit the body’s own stem cells to promote bone repair is a current research hotspot.In this study,the multifunctional composite hydrogels loaded with black phosphorus nanosheets（BPNS）and nano magnesium oxide（Mg O）were prepared.Without the addition of additional biological components,this composite hydrogel can promote stem cell migration,osteogenic differentiation and biomineralization,thus accelerating the bone regeneration in situ.The main research contents are as follows:Firstly,the BPNS was prepared by liquid phase exfoliation method,and the morphology and size of BPNS were characterized by scanning electron microscope,atomic mechanical microscope and particle size analyzer.The results showed that the prepared BPNS surface layer was uniform in size and had typical two-dimensional material structure.Then,after loading with Mg O and BPNS in PVA/CS,the PVA/CS-Mg O-BPNS composite hydrogel was prepared by the physical cross-linking method of repeated freezing and thawing.The morphology,structure and mechanical strength of the composite hydrogel were characterized by Fourier transform infrared spectroscopy,scanning electron microscope,thermogravimetric analysis and universal test machine,respectively.The results showed that the PVA/CS-Mg O-BPNS composite hydrogel has good porous structure and can sustainably release Mg²⁺and phosphate.At the same time,the degradation of Mg O can cause a weak alkaline microenvironment,which can synergize with phosphate to promote PVA/CS-Mg O-BPNS biomineralization performance in vitro.Secondly,the photothermal conversion properties of PVA/CS-Mg O-BPNS composite hydrogels were analyzed by under NIR irradiation.Staphylococcus aureus and Escherichia coli were selected to test their in vitro antibacterial properties.The results showed PVA/CS-Mg O-BPNS composite hydrogel has good photothermal conversion and stability under NIR irradiation.At the same time,based on the dual effects of CS inherent antibacterial properties and photothermal therapy,the antibacterial rate of PVA/CS-Mg O-BPNS composite hydrogel against the above two bacteria can beyond 99.9%,which can effectively prevent the occurrence of implant-related infections.Thirdly,the phenotype of the isolated rat bone marrow mesenchymal stem cells was identified by multi-directional differentiation induction and flow cytometry.The PVA/CS-Mg O-BPNS composite hydrogel has good cytocompatibility by means of cell live and dead staining and cell proliferation detection.Transwell cell migration experiments and osteogenesis experiments proved that PVA/CS-Mg O-BPNS composite hydrogel can promote the migration and osteogenic differentiation of mesenchymal stem cells in vitro.Through RNA sequencing and to investigate the related molecular mechanisms,it was showed that the PVA/CS-Mg O-BPNS composite hydrogel can promote cell migration and osteogenic differentiation though activating the Pi3k-Akt signaling pathway.Finally,the PVA/CS-Mg O-BPNS were implanted into a rat cranial defect model.Micro-CT,HE staining,Masson staining,immunohistochemical staining and other methods proved that PVA/CS-Mg O-BPNS composite hydrogel can effectively promote the formation of new bone.At the same time,animal experiments also proved that the PVA/CS-Mg O-BPNS composite hydrogel does not have toxic effects on the main organs of rats,and has good in vivo biocompatibility.In conclusion,the PVA/CS-Mg O-BPNS composite hydrogel has good biocompatibility and osteo-inductivity,is a good scaffold material for bone tissue engineering,and has great application prospects in bone repair.