The Preparation And Cytological Studies Of Electrospun Fiber Membrane Carrying RhPTH (1-34)

Background and Objective:Implant denture has been widely applied in clinical field to repair themissing teeth for patients. The bone defects around implants will affectrapid healing of implants-osseointegration interface. Currently, treatmentof bone defects by using bone autograft, allograft, biological material anddistraction osteogenesis technology has made a certain progress. However,because of secondary injury, immune rejection, high technicalrequirements, limit their more widespread application. With thedevelopment of biomaterial science, building of artificial bone by tissueengineering technology for bone defect is becoming a hot spot of currentresearch.rhPTH（1-34）, a drug for increasing the bone formation, canpromote the bone formation and increase the bone density by increasingthe number and prolonging the lifespan of osteoblasts. Thus, wespeculated that it can increase the bone mass and improve theosseointegration of the planting area. In the past clinical trials andresearch, rhPTH （1-34） were mainly administered by low-dose,intermittently, subcutaneously and systemically. However, thedeficiencies of PTH, such as volatility and short half-life, limit its furtherapplication in the field of oral Implantology. Thus, it will bring broader application prospects in the field of oral Implantology to develop adelivery system to improve the bioavailability of rhPTH（1-34）for itsgood function to promote bone formation.With high surface area and high porosity, electrospun fiber has theadvantages of complicated porous structure, it has a widely applicationsuch as tissue engineering and drug carrier. Composite fiber could beused as a kind of effective control of local slow-release drug carrier. Thisstudy aims to preparation loading rhPTH （1-34） to achieve thecontrollable release of rhPTH（1-34）. Furthermore, by studying the effectof this system in promoting cell proliferation and differentiation in vitro,we can provide a theoretical guidance for further applications.Methods:PLGA, nHA, rhPTH (1-34) according to the different componentsdissolved in chloroform, magnetic stir well, spinning. Preparation ofPLGA, PLGA\HA, PLGA\PTH, PLGA\HA\PTH components of differentmembrane. Using of scanning electron microscopy (SEM) to observematerial surface morphology; Using of universal testing machine to testthe material tensile mechanical properties; By measuring theweightlessness rate, pH change, and the water absorption to evaluationthe physical and chemical properties of material; The cumulative releaseof rhPTH (1-34) tests by MicroBCA method, reflect the in vitro release ofmaterials. By FITC\PI after dyeing, using fluorescence microscope celladhesion in the material surface; By MTT method to detect the materialimpact on MC3T3-E1cell proliferation; Through ALP activity experiment testing influence of MC3T3-E1cell differentiation.Results:1.The physical and chemical properties:Electrospun fiber membrane electrospun fiber membrane preparationby uniform texture, a porous structure, PLGA\HA\PTH and PLGA/HA,PLGA\PTH similar morphology, PLGA diameter reduced. PLGA tensilestrength best, PLGA\PTH times intensity, PLGA\HA, PLGA\HA\PTHthe strength of the two materials. PLGA\PTH, PLGA\HA\PTHweightlessness rate significantly faster than PLGA, PLGA\HA grouprespectively. In the process of the degradation of PLGA, PLGA\PTH pHdecline appear similar, similar in the other two groups. After soaking24dPBS, PLGA\HA bibulous rate is lowest about9.3%, PLGA\PTHbibulous rate is highest about28.1%. PLGA\PTH, PLGA\HA releaserhPTH(1-34) rate has no obvious differences, before one week is thequick release phase,4week later, the cumulative release was in the end.2.Electrospun fiber membrane cells biocompatibility:Cell adhesionCell cultured after6h, the amount of cell adhesion ofPLGA\HA\PTH group was more than PLGA\PTH, PLGA\HA group, andthe cells packed closely. The adhesive capacity of PLGA group was lessthan the other three groups significantly, At the same time cells not onlyadhere to the material surface but also immersed into material.Cell proliferationAt first day, PLGA\HA and PLGA\PTH PLGA is obviously better than the cell proliferation group(P<0.01), cell proliferationPLGA\PTH\HA group was obviously better than PLGA/PTH, PLGA/HAgroup (P<0.01). The trend of3days,5days was similar to1day.Cell differentiationAt first day, Compared the ALP content express of PLGA gourpwith PLGA\PTH group was no significant difference(P>0.05); ComparedPLGA\HA group with PLGA\PTH\HA group had a significantlydifference(P<0.05).At7day, the PLGA\HA, PLGA\PTH andPLGA\PTH\HA group had a significantly higher expression of ALPcompared to PLGA group(P<0.01), PLGA\PTH\HA of ALP activity ofthe most obvious (P<0.01). The result of14day was similar to7day.Conclusion:1. Loading rhPTH (1-34) electrospun fiber can effectively controlthe release of drugs.2. The addition of nHA into PLGA membrane could reduce acid(side effect) in the process ofdegradation.3. Both loading rhPTH(1-34) and nHA of electrospun fibermembrane can promote MC3T3-E1cell proliferation and differentiation.rhPTH (1-34) and nHA performance for synergy.