Preparation Of Dex-loaded Shape Memory Composite Nanofibers Of NHAp/PLMC For Osteogenic Differentiation

Recently, how to develop biomimetic biomaterial scaffolds to repair damaged or deseasedbone defects has drawn a great deal of attention in the community of tissue engineering andregenerative medicine (TERM). Considering that the native bone tissue is a complexmicroenvironment system composed of various components and factors such as multiple types ofcells, extracellular matrix, biomolecule signaling factors and mechanical stimulation, it is essentialto design and fabricate biomaterial scaffolds to precisely imitate the matrix microenvironment,biomechanical microenvironment and biological factors microenvironment of the natural bonetissue, which could ultimately lead to effective repair and regeneration of different bone defects.In recent years, electrospinning has been widely employed for fabricating many types oftissue scaffolds. This technique is certainly able to provide an enabling platform for biomimickingthe matrix microenvironment of natural bone tissue at the composition and nanostructure levels.Biodegradable shape memory polymers (SMPs), as a kind of smart material, has received muchconcern due to a wealth of merits noted such as appropriate biocompatibility, high strain rate,adjustable deformation temperature, easy to shape into different configurations, and so on. Usingshape memory effect of the SMPs for scaffolding purpose is most likely able to exert lastingbiomechanical forces onto the surrounding bone tissue during their recovery processes. This couldtherefore provide an ideal biomechanical stimuliation for bone tissue remodeling. As anosteogenic inducing supplement, dexamethasone (Dex) has been widely used in bone tissueengineering. Taken together, it is thus reasonable to fabricate SMPs into nanofibers through theelectrospinning technique along with Dex-loading to achieve a comprehensive biomimicry to thematrix, biological factors and biomechanics environments of the natural bone.The aim of this thesis is to prepare shape memory-capable nanofibrous scaffold of nanohydroxyapatite/poly(D,L-lactide-co-trimethylene carbonate) loaded with Dex (denoted asDex/nHAp/PLMC), and to study its effects on the differentiation of bone mesenchymal stem cells(BMSCs) into osteoblasts under the culturing conditions of basic culture medium and osteogenesis medium, respectively. To this end, firstly we preparated the Dex/nHAp nanoparticles by using thenHAp to adsorb Dex in PBS; and then incorporated the Dex/nHAp nanoparticles into the shapememory polymer PLMC through electrospinning to produce the composite nanofibers ofDex/nHAp/PLMC. Morphology, structure, thermal, mechanical and shape memory properties ofthe nanofibers were thoroughly characterized. Ultrasound was employed as a new thermalstimulator to trigger shape recovery process and regulate the release kinetitics of the Dex from thenanofibrous Dex/nHAp/PLMC scaffold. The results show that Dex can be well adhered onto thesurface of nHAp; the Dex/nHAp/PLMC nanofibers have smooth surface morphology, and theloaded Dex/nHAp nanoparticles are uniformly dispersed within the nanofibers. Due to theincorporation of Dex/nHAp, the fiber diameters are slightly increased from542144nm to799180nm; the glass transition temperature (Tg, also is the transation temperature Ttransfor actuatingshape memory) of the nanofibers increased from38.6°C to43.4°C. The observed Ttransdata are inthe safe range of body temperature, which also implicates an external trigger (e.g., ultrasound) isneeded to realize shape recovery process of the nanofibrous Dex/nHAp/PLMC scaffold with shapememory effect. In addition, mechanical properties at room temperature and body temperature areboth improved. For instance, at room temperature and37C the tensile strength of theDex/nHAp/PLMC scaffold increased88.1%(from2.19MPa to4.12MPa with respect to thepristine PLMC) and31.9%(from6.9MPa to9.1MPa with respect to pristine PLMC),respectively. Shape memory tests show that with the incorporation of Dex/nHAp, the shape fixedrate and shape recovery rate of the composite nanofibers have reached to99.06%and99.99%,respectively. Besides, ultrasound mediated drug release results show that the amount of drugrelease could be maximized at the power of100w and the treat time of10s.In order to evaluate the osteogenesis capacity of the newly developed nanofibrousDex/nHAp/PLMC scaffold, BMSCs were used to study the influence of Dex/nHAp/PLMCnanofibers on the growth and osteogenic differentiation of the stem cells. The cytocompatibility ofthe composite nanofibers was evaluated by cell counting kit-8(CCK-8), SEM and fluorescencestaining, and the osteogenesis is evaluated by alkaline phosphatase (ALP) quantification, collagen(Col) quantification and ALP staining. The results show that the adhesion, proliferation and spreadability of BMSCs on the nanofibrous Dex/nHAp/PLMC scaffold are better than that on PLMCnanofibrous substrate. Even in the the case of basal medium condition used, data from proteinlevel assays including the alkaline phosphatase (ALP) activity and collagen detection both indicatethat the Dex/nHAp/PLMC composite nanofibers significantly promoted the osteogeneticdifferentiation of the BMSCs.In short, the results show that the biomimetic and bioactive Dex/nHAp/PLMC scaffolds cansignificantly promote the adhesion, proliferation and differentiation of BMSCs. In consideration of its high shape recovery ratio, this study sets the foundation to further explore the correlationbetween its shape memory effect and the generated mechanical force, and this study also forms thebasis for performing comprehensive simulation of the matrix, biological factors, andbiomechanical microenvironments of the natural bone tissue through animal test in vivo.