| Objective:Epidural fibrosis is one of the common complications after spinal surgery.Severe epidural fibrosis can lead to compression of the dura mater and nerve roots,cause the recurrence of chronic low back and leg pain,dysfunction or loss of lower limb function,and eventually result in Failed back surgery syndrome.In recent years,scholars at home and abroad have carried out a lot of researches on the prevention and treatment of postoperative epidural fibrosis,including oral or local application of drugs,intraoperative application of biomaterials,but these treatment methods have some disadvantages,which limit its clinical transformation.Therefore,it is of great significance to explore a new mode of treatment for epidural fibrosis.In this study,the anti-fibrotic drug resveratrol was encapsulated in PLGA microspheres and loaded with injectable GelMA hydrogel to prepare a drug loaded hydrogel system for intraoperative use to prevent postoperative epidural fibrosis.Methods:1.Human fibroblasts were cultured in vitro and treated with resveratrol.CCK-8 assay,EdU staining,Annexin V/PI double staining and scratch test were used to analyze the effects of drugs on the proliferation,apoptosis and migration of fibroblasts.Western blot was used to analyze the changes of protein expression levels related to cell proliferation.2.Different concentrations of hydrogels were prepared.Resveratrol was embedded in the PLGA microspheres and loaded into hydrogels to RSV@PLGAGelMA hydrogels.The morphology of the RSV@PLGA-GelMA hydrogels was analyzed by macroscopical observation and scanning electron microscopy.The swelling ability,degradation behavior and mechanical properties of RSV@PLGAGelMA hydrogels were detected.The characteristics of drug release were calculated.CCK-8 assay was used to evaluate the cytotoxicity of hydrogels.3.A rat laminectomy model was established,and the RSV@PLGA-GelMA hydrogels was implanted during operation.The effects of RSV@PLGA-GelMA on preventing epidural fibrosis were analyzed by HE and Masson staining.Immunohistochemical staining and Western blot were used to detect the difference in the expression of target proteins related to fibrosis.4.Resveratrol and TGF-β1 were used to intervene fibroblasts at the same time.The effects of resveratrol and TGF-β1 on the proliferation and apoptosis of fibroblasts were evaluated with EdU staining and Annexin V/PI double staining.The expression of TGF-β1-related downstream signaling proteins was detected with Western blot to explore the potential mechanism of resveratrol in preventing fibrosis.Results:1.The experimental results showed that resveratrol can effectively inhibit fibroblasts proliferation and migration and promote cell apoptosis.2.The surfaces of RSV@PLGA-GelMA hydrogels were smooth and porous,and the size and distribution of pores were homogeneous.Mechanical tests showed that 8%of the hydrogel system had moderate mechanical strength.Sustainable and stable drug release was achieved when resveratrol was loaded in hydrogel.The results of cytotoxicity analysis confirmed that hydrogels had good biocompatibility.3.Animal experiments showed that the RSV@PLGA-GelMA hydrogel could prevent epidural fibrosis after laminectomy,and the material had no obvious tissue toxicity.4.Resveratrol can inhibit the proliferation of fibroblasts induced by TGF-β1 and reduce the phosphorylation levels of Smad2,Smad2,PI3 K and Akt in its downstream signaling pathways.Conclusions:Resveratrol can inhibit the proliferation and migration of fibroblasts.Loading the resveratrol into the injectable GelMA hydrogel can realize the sustained release of the drug.The RSV@PLGA-GelMA(8%)hydrogel has good mechanical properties and biological safety,and can prevent epidural fibrosis after laminectomy in rats.The drugloaded hydrogel system has the characteristics of being injectable and convenient to use,and may provide a new treatment plan and inspiration for the clinical prevention of postoperative epidural fibrosis. |
PDF Full Text Request