Study Of Oxygen Generating Sodium Alginate Composite Hydrogels For Promoting Bone Regeneration And Treatment Of Periodontitis

Bone defect caused by tumor,inflammation or trauma is a common clinical problem.Autogenous bone,allogeneic bone or xenogeneic bone is a common clinical method,but its application is limited by the source,immune rejection and potential virus transmission.Tissue engineered bone scaffolds can avoid the above risks and have a wide range of sources,which is considered to be the most potential method for bone defect repair.However,tissue engineered bone scaffolds lack functional vascular system,resulting in hypoxia in the central area of scaffolds,which affects cell survival,differentiation and osteogenesis.It has been proven that providing enough oxygen for cells before blood vessels grow into scaffolds is of great significance for cell survival and bone regeneration.However,uncontrolled and excessive oxygen will produce excess reactive oxygen species,destroy the redox balance,lead to cell dysfunction or death,and hinder bone regeneration.Therefore,controlling oxygen release and antioxidation of bone scaffold materials are equally important for maintaining redox balance,cell survival and bone regeneration.Based on the above background,in the second chapter,we synthesized a hydrogel scaffold which could release oxygen for a long time for repairing bone defects.First,we synthesized a hydrogel scaffold with nontoxic sodium alginate.Scanning electron microscopy（SEM）showed that the hydrogel was composed of pores and skeletons with a high porosity and large pore size.We synthesized calcium peroxide nanoparticles and added them into hydrogels as oxygen producing materials.Calcium peroxide reacts with water to form hydrogen peroxide and calcium hydroxide.The former is a strong oxidant and continues to decompose into oxygen and water;the latter is a strong base.Therefore,if hydrogen peroxide and calcium hydroxide produce too much,the redox balance and acid-base balance of surrounding cells will be destroyed,resulting to cell death and affect bone regeneration.For this reason,we added an acid antioxidant,ascorbic acid,in the hydrogel to overcome the excessive reactive oxygen species and alkaline substances produced during the release of oxygen from calcium peroxide.Scanning electron microscopy（SEM）showed that the self generated oxygen composite hydrogel had porous three-dimensional structure and large porosity,and the composite hydrogel was successfully loaded with calcium peroxide and ascorbic acid by elemental distribution and infrared spectroscopy.The composite hydrogel can degrade slowly.During the degradation period,it can release the oxygen steadily and continuously,and the degradation product can maintain the neutral environment.In the third chapter,we evaluated the bone regeneration effect of oxygen generating hydrogel synthesized in the last chapter.Cytotoxicity experiments showed that the hydrogel was almost non-toxic.When the concentration reached 200μg/ml,the survival rate of bone marrow mesenchymal stem cells（r BMSCs）was still over 90%.It was further found that hypoxia（2%O₂）did not significantly inhibit the proliferation of r BMSCs,but the proliferation rate of r BMSCs in the composite hydrogel group increased significantly after 48 hours.Moreover,the composite hydrogel could promote the expression of osteogenic differentiation genes（such as OCN,Col,Sp-7,Runx2）,increased alkaline phosphatase activity and increased calcium nodule formation in hypoxia（2%O₂）r BMSCs,suggesting that the composite hydrogels can promote the osteogenic differentiation of r BMSCs in hypoxic state.In vivo animal experiments also showed that the composite hydrogel could significantly promote the regeneration of rat skull defects.In addition,no obvious abnormality was found in the main organ tissue staining and serum examination,indicating that the composite hydrogel had good safety.In order to expand the application scope of the oxygen generating composite hydrogel,in the fourth chapter,we synthesized injectable,mobile oxygen producing composite hydrogel for the treatment of periodontitis.Periodontitis is essentially a bone defect disease caused by bacterial infection.Therefore,bacteriostasis and bone regeneration are the main strategies for the treatment of periodontitis.Anaerobic bacteria is the main pathogenic bacteria of periodontitis,which is sensitive to oxygen.Increasing the oxygen content in the periodontal pocket can significantly inhibit the proliferation of bacteria;and increasing the oxygen content in the periodontal pocket is conducive to bone regeneration.Similar to the composite hydrogel made in the second chapter,the injectable composite hydrogel also has a three dimensional structure with high porosity and larger pore size.It can degrade slowly to produce oxygen,and the neutral environment is maintained during the degradation process.Bacteriostasis experiments showed that this composite hydrogel could obviously inhibit the proliferation and reproduction of Porphyromonas gingivalis under anaerobic environment.In vivo animal experiments showed that composite hydrogel could alleviate inflammation of gingival tissue and promote regeneration of alveolar bone in rats with periodontitis.In addition,no obvious toxicity was found by histological staining and serum examination.