| Blackground:Trauma,infection,surgery and so on often lead to the defect or loss of periodontal supporting tissue.While traditional periodontal treatment generally cannot promote regeneration of periodontal tissue.In recent years,the development of tissue engineering is remarkable.How to use tissue engineering to promote the regeneration of the periodontal tissue and rebuild the natural periodontal tissue physiological structure and function of natural periodontium,has always been the hot issue of scholars,such as bone graft,guided tissue regeneration,enamel matrix protein derivative,growth factor,stem cell therapy,cell engineering and laser treatment,etc.The optimal healing of periodontal tissue defect is to achieve the complete functional regeneration of alveolar bone,cementum and periodontal membrane,so as to obtain the new periodontal attachment and achieve a good long-term effect.Therefore,to reconstruct the health of periodontal tissue is an important problem in the treatment of oral diseases.With the continuous development of material,Good bone integration is no longer the sole criterion for determining the success of oral diseases such as implant surgery,the aesthetic effect of gingival soft tissue is also the important indexes for evaluation of the effect.At present,a large number of studies have been made for the periodontal regeneration especially regeneration of periodontal membrane and bone,the application of all kinds of new materials has superior effect of tissue regeneration,but relatively few studies of gingival soft tissue repair regeneration.Gingival connective tissue contains abundant blood vessels and progenitor cells,which play a very important role in the regeneration of periodontal tissue.As an important factor influencing the early healing of periodontal tissue,the aesthetic effect and the long-term function,the shape and tissue of the gingival soft tissue are very important.Human tissue is a highly ordered tissue assembled in order by the most basic cell and extracellular matrix.The cells in the tissues such as nerve,muscle,tendon and blood vessel present an orientation structure,and the arrangement of cells has a certain effect on the phenotype of the cells.Therefore,how to realize the orderly arrangement of multi-cells has become an important research direction in the field of tissue engineering.In nearly a decade,electrostatic spinning technology has attracted great attention,it can be prepared different orientation and arrangement of scaffolds that is similar to natural extracellular matrix(ECM),which provide the suitable growth environment for the cells.Based on this,the aligned fiber scaffold fabricated by parallel electrode electrospinning technology has been widely concerned.Cell culture is one of the most basic operations of biological research.How to simulate the physiological environment in vitro is the answer that many scholars have been pursuing.Thus,three-dimensional cell culture emerged.Three-dimensional cell culture is mean to simulates the ECM of real tissue by cells aggregates or establishing cell spheres or embedding cells into scaffolds to simulate the internal environment to a certain extent.Scaffolds is the most commonly used material of three-dimensional cell culture,As one of the material for making cell scaffolds,polycaprolactone(PCL)has good biocompatibility,low cost,easy processing,slow hydrolysis degradation rate.After degradation,the human body reabsorbs it's hydrolysate and has small reaction to tissue.This experiment will observe whether the aligned PCL scaffolds can promote the proliferation and differentiation of gingival fibroblasts,and predict the ability of aligned PCL fibrous scaffold to promote gingival soft tissue defect repair and regeneration.Objective:In this experiment,the polycaprolactone fiber membrane was prepared by parallel electrode electrostatic spinning technology as scaffold material,and co-cultured with the gingival fibroblasts,compared the effects of two different topological structures of scaffold material on cell proliferation and related protein expression.Methods:1.Preparation of random and aligned scaffold:Using parallel electrode electrostatic spinning techniques to prepare random and aligned scaffold,scanning electron mirrors to observe.2.Cell culture:Select three days old SD rats,the tissue block method is adopted to extract the original generation of gingival fibroblasts,extend the purification and identification of cell source,4 to 6 generations of cells are accepted.3.Construct the cell-scaffold complex:After sterilization of two kinds of PCL scaffolds,gingival fibroblasts were seeded and cultured for 5-7days,After fixation and dehydration,observe by scanning electron microscope.4.The effects of two PCL fibers on proliferation of GF:The GF was seeded on the random and aligned scaffold,and detect cell proliferation in the 1st,4th,7th and 10 th days by CCK8.5.The effects of two PCL fibers on the expression of various proteins of GF:The GF was seeded on the random and aligned scaffold,and detect the mRNA expression of FN,FAK and COL-1 by qRT-PCR after 10 days.Results:1.Detect the cell proliferation in the two kinds of PCL scaffolds in the 1st,4th,7th and 10 th day respectively.The results showed that the proliferation of the aligned scaffold group cells was on the rise,and the random scaffold group showed a rising trend in 1-7d,and the decline began after 7d.There was a significant difference in cell proliferation between day 7 and 10(p < 0.05),Compared with the random scaffold,the aligned scaffold promoted the proliferation of GF.2.qRT-PCR tests in 10 days,compared with the random scaffolds,There was a significant difference in the mRNA expression(FN,FAK,COL-1)of the aligned scaffold group are raised,and statistically difference(p < 0.05).Conclusion:1.GF was successfully extracted,which grows strong and stable.2.the aligned PCL scaffolds were manufactured,with a uniform diameter and good ordering.3.Compared with the random scaffold,the aligned PCL scaffolds promote the proliferation and related protein expression of gingival fibroblasts. |
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