Experimental Study On Local Treatment Of Traumatic Heterotopic Ossification With PLGA Loaded Celecoxib Microspheres

Background:Traumatic heterotopic ossification is a condition in which certain traumatic factors cause cartilage-like or bone-like tissue to appear in the non-ossified tissues of the body.The incidence is as high as 23%-100%after certain traumatic surgeries without preventive treatment.It often manifests as pain and limited joint movement,which seriously affects the patient’s postoperative recovery and quality of life.Once traumatic heterotopic ossification occurs,surgical treatment options are limited due to frequent recurrence after surgical resection and the fact that some patients may not be able to have it removed due to its sensitive location.Non-steroidal anti-inflammatory drugs are commonly used clinically to prevent the development of heterotopic ossification.However,long and frequent dosing cycles,often with significant drug side effects outside the lesion site,thus reducing patient compliance,have largely limited the use of NSAIDs.Therefore,we attempted to construct a long-term extended-release system of NSAIDs for the local treatment of traumatic heterotopic ossification.Objectives:1.To prepare celecoxib-PLGA microspheres by oil-in-water emulsion method and determine their physicochemical properties.2.To study the inhibitory effects of celecoxib-PLGA microspheres on cell survival,proliferation and osteogenic differentiation under in vitro conditions.3.To study the biocompatibility of celecoxib-PLGA microspheres under in vivo conditions and the inhibitory effect on ectopic ossification.Methods:1.Celecoxib-PLGA microspheres were prepared using the oil-in-water emulsion method.Their characterization was observed using scanning electron microscopy.The particle size distribution,drug loading capacity,encapsulation rate and drug release profile were determined.2.The effects of celecoxib-PLGA microspheres on the viability and morphology of L929 cells were studied using CCK 8 method and deadwood staining.3.The inhibitory effect of celecoxib-PLGA microspheres on osteogenic differentiation of bone marrow mesenchymal stem cells was studied by using alizarin red staining and real-time quantitative polymerase chain reaction.4.The therapeutic effects of celecoxib-PLGA microspheres on traumatic ectopic ossification in SD rats were studied by micro-CT scanning,Masson trichrome staining and HE staining.Results:1.The three groups of celecoxib-PLGA microspheres had different distribution of particle size and drug release profiles,but had similar drug loading and encapsulation rates,as well as uniform spherical shape.2.Celecoxib-PLGA microspheres co-cultured with L929 cells at concentrations of 10-1000 ppm showed no significant cell death and allowed the better active spindle-shaped L929 cells to adhere to the surface.3.Celecoxib-PLGA microspheres effectively reduced the formation of orange-red calcium nodules after 21-day induced osteogenic differentiation of bone marrow mesenchymal stem cells and inhibited the expression of ectopic ossification-related genes:HIF 1 α,BMP 2 and SOX 9 after 14-day induced osteogenic differentiation.4.celecoxib-PLGA microspheres inhibited ectopic ossification at the animal level:micro-CT,HE staining of tissue sections and Masson staining indicated a significant reduction in the amount of ectopic bone and a decrease in tissue ossification centers after 70 days compared to the control experimental group.Conclusions:1.celecoxib-PLGA microspheres prepared by oil-in-water emulsion method can present different particle size distribution and adjustable drug release profile by adjusting the parameters.2.Celecoxib-PLGA microspheres have good biocompatibility in vivo and in vitro.3.Topical application of celecoxib-PLGA microspheres can effectively inhibit ectopic bone formation in vivo and in vitro.