Preparation Of Oxygen-Carrying Bone Implant Materials And Effects On The Cell Response

The balance of oxygen in the bone repair microenvironment plays a multiple functions in bone remodeling process,which involves 1)cellular aerobic metabolism,2)collagen synthesis,3)expression of some angiogenic genes,4)osteogenic activity of osteoblast-related cells.However,severe bone damage is likely to cause destruction of the vascular network in the affected area,resulting in local hypoxic state of the bone.Regional hypoxia reduces the effective exchange of nutrients and metabolites,worsening bone repair behavior of the implant.Moreover,in the area of plateau hypoxia,human hemoglobin has less oxygen-carrying capacity,slowing down blood flow,and further aggravating the degree of hypoxia in damaged areas.Under this series of factors,the implants cause abnormal osteoblast proliferation.In addition,the weight-bearing bones of the lower limbs extremities can stimulate the recessive osteosarcoma proto-oncogene under the stimulation of bone pathological conditions such as bone injury,leading to cell mutations,resulting in a pathological fracture?osteosarcoma?.Overgrowth of osteosarcoma and abnormal vascular structure in the tumor will further aggravate the hypoxic state of the microenvironment,which reduce the effectiveness of anti-tumor treatment and easily worsen the behavior of bone repair.Therefore,appropriate oxygen concentration in the bone repair microenvironment is of great significance to promote implant integration.Based on the above background,the new bone implant with oxygen-releasing materials that could produce oxygen in situ or carry oxygen in vitro were selected to prepare.The effect of these materials to osteoblasts,osteosarcoma cells,etc.was investigated.The main results obtained were as follows:?1?Calcium peroxide?Ca O₂?was used as the oxygen releasing material,while polyvinylpyrrolidone?PVP?and polylactic acid-glycolic acid copolymer?PLGA?were simultaneously introduced to construct PLGA-PVP/Ca O₂ core-shell microspheres.The three-dimensional spherical shape of the PVP assembly was helpful for the subsequent PLGA packaging.The PLGA shell encapsulating PVP/Ca O₂ could reduce the contact rate of the material with body fluids and regulated the oxygen generation reaction.Next,the porous titanium coating?TC?was subjected to alkali heat and alkylation surface treatment,and an organic transition layer containing amino groups was formed on the surface of TC by using covalent bond grafting and fixing method,which formed a peptide amine chemical bond with the functional group?-COOH?of the PLGA end chain of the PLGA-PVP/Ca O₂ core-shell microsphere.This could strengthen the combination of the PLGA-PVP/Ca O₂ layer and TC to build a TC bone implant material with stable oxygen release function?TC-AAO?.?2?We evaluated the effects of TC-AAO on survival,adhesion and differentiation of MC3T3-E1 in simulated hypoxia?1%O₂,5%CO₂,37°C?and normal oxygen?21%O₂,5%CO₂,37°C?environments.Experimental results showed that PVP/Ca O₂promoted cell adhesion,proliferation and differentiation under hypoxic conditions.Compared with TC-B prepared by physical adsorption,TC-AAO immobilized by covalent grafting method could effectively regulate the release of oxygen and the accumulation of dissolved oxygen in the injured area.This area had a higher focal adhesion protein and actin strength,which could promote cell adhesion.Under hypoxic conditions,the alkaline phosphatase?ALP?activity of MC3T3-E1 cells on the surface of TC-AAO was about 1.5 times that of TC and TC-B?14 days?,indicating that the differentiation behavior was also significantly improved.Preliminary analysis by RT-PCR found that the introduction of Ca O₂ under hypoxic atmosphere promoted the normal expression of related osteogenic genes such as TGF-?1,and the expression level of TGF-?1 on the surface of TC was only half of that of TC-AAO.This showed that the slow and continuous oxygen release could effectively ensure the appropriate oxygen concentration in the microenvironment in the early stage of bone healing,thereby promoting the response of osteoblast-related cells and genes on the surface of the implant.?3?Using?-Fe₂O₃ as a magnetic material,the effect of oxygen release on tumor magnetocaloric therapy was studied.By controlling the flow of oxygen in the sheath gas,?-Fe₂O₃ nanoparticles with high magnetic induction intensity were prepared by using the inductive plasma method.Compared with the traditional solvothermal method,the induction plasma spraying only used ferric nitrate and absolute ethanol,which greatly reduced the residual risk of surfactants and had the advantage of easy operation.Meanwhile,PVA@?-Fe₂O₃-PFH oxygen self-enriched magnetic material could be prepared by complexing?-Fe₂O₃ magnetic particles with perfluorohexane?PFH?at the oil-water interface,which could solve the resistance of hypoxic tumor microenvironment to magnetothermal therapy.Cytotoxicity experiments showed that PVA@?-Fe₂O₃-PFH oxygen self-enriched magnetic material still had good activity and no obvious toxic and side effects on human osteosarcoma cells?MG-63?.?4?To evaluate the magnetocaloric therapeutic performance of PVA@?-Fe₂O₃-PFH,we investigated the damage of PFH,?-Fe₂O₃,PVA@?-Fe₂O₃-PFH to the DNA of MG-63.It was found that MG-63 tumor cells treated with PVA@?-Fe₂O₃-PFH showed further increased DNA damage,which was much higher than the results after treatment with?-Fe₂O₃ or PFH alone.