| Resveratrol is a naturally occurring polyphenol compounds, which has been foud to have anti-inflammatory, anti-tumor, cardiovascular protection,liver protection, neuroprotection, immunoregulation,anti-aging and other pharmacological effects. The existing research results show that resveratrol have certain effects to prevent or treat skin disorders.Resveratrol has two structures:the cis- structure and the trans- structure.The bi ology activity of the trans- resveratrol is better than that of the cis-resveratrol, while trans-resveratrol is more likely to transform into cis-resveratrol under the condition of high temperature, illumination and so on. Resveratrol, poorly soluble in water, chemic ally unstable, sensitive to oxidative decomposition,rapidly metabolied in the body after oral administration, low bioavailability in vivo, all which restrict its wide application. Hence, research about resveratrol formulations is on growing in recent years. Existing resveratrol formulations include liposomes, nanoemulsion, cyclodextrin complex, emulso me,nanosuspension,microspheres, nanoparticles, gel, etc.Psoriasis is a kind of immune-mediated chronic inflammatory skin disease, and th e histopathologic features of psoriasis include:hyperkeratosis and parakeratosis of kerat inocytes, distortion of blood vessels in the dermis, infiltration of inflammatory cells.Ke ratinocytes, T cells and dendritic cells are involved in psoriasis, but specific mechanis ms of pathogenesis are still unclear. Many cytokines are involved in the pathogenesis of psoriasis, among which the TNF alpha plays an important role in the pathogenesis of psoriasis. TNF alpha can promote proliferation of keratinocytes and regulate the inf lammatory response. Biological agents against TNF alpha for treatment of psoriasis h ave very good therapeutic effect.In the past few years the imiquimod-induced mouse model of psoriasis has been widely used. Many of the pathways involved in human psoriasis disease appear to be mirrored in the imiquimod-induced model. Researchers have found that the imiquimod-induced model has some relationship within the enzyme system of oxidative stress system. Lipid, protein and DNA oxidative damage are existing in psoriasis patients,and abnormally increased antioxidant system and reactive oxygen species might be the key factors in the pathogenesis of psoriasis.Reactive oxygen species can act as the second messenger regulating the MAPK/AP-1 NF-kB, JAK-STAT signal transduction pathway, and all these cellular pathways in psoriasis skin lesions are abnormally activated. These pathways can trigger reactions of cells involved in psoriasis to cytokines, make some inflammatory cytokines and chemokines to over express, which in turn affect cell proliferation and differentiation.The main research purpose of this subject is to discuss the effect of resveratrol on TNF alpha induced proliferation and secretion of Th1/Th2 type cytokines of HaCaT. Resveratrol liposome gels was made and applied to imiquimod-induced psoriatic mouse model to investigate the mechanism of how resveratrol acts, percutaneous pharmacokinetic of resveratrol liposome gel was investigated to gain more evidence for the further development and utilization of this compound.Percutaneous microdialysis technology is considered as a effective tool for the evaluation of drug absorption and metabolism in the skin. Minimal tissue damage, real-time, on-line sampling in animal body, all which make this technique widely used in percutaneous pharmacokinetic research.The contents of this research include four parts.The first part:Effect of resveratrol on TNF alpha induced proliferation of HaCaT was determined by MTT method,and the results show that resveratrol has inhibitory effect on cell proliferation. By ELISA detection,we found that resveratrol had inhibitory effect on secretion of IL-6 and IL-8 and promoting effect on the secretion of IL-10. Results of this part show that resveratrol can inhibit excessive proliferation of keratinocytes, and affect the secretion of Thl/Th2 cytokines in vitro.Resveratrol can inhibit secretion of Thl cytokine, promote secretion of Th2 cytokines.The second part:Preparation of resveratrol liposome gel and its transdermal performance in vitroFirst we determine the resveratrol liposome preparation methods, and then through the orthogonal design the best weight ratio was determined,and optimize the preparation technology of resveratrol liposome. Results of this section show that the best preparation technology are as follows:film dispersion method;soybean lecithin:cholesterol:resveratrol (20:4:5,w/w/w), suitable amount of ethanol to dissolve, ultrasound after 10 min, evaporate solvent under rediced pressure at40℃, adding suitable amount of pH7.4 PBS after soybean phospholipids in calabash wall forming a layer of uniform film, rotate 30 min, place 2 h to make it fully hydrated, ice bath under the condition of ultrasonic 30 min. Average coating rate 5 batch of sample was 90.27%, the RSD was 0.95%.The validation test results show that the preparation technology are stable and reliable.Liposome gels was made and Franze diffusion pool was used to investigate s in vitro transdermal performance of resveratrol liposome gel.In vitro drug release results show that resveratrol liposome gel has obvious sustained release and good transdermal absorption effect, compared with resveratrol gel. Liposome gel has more advantages than normal gel as a carrier of transdermal absorption.The third part:Therapy of resveratrol on imiquimod-induced psoriatic mouse modelFirst effect of resveratrol on ear thickening and inflammatory response of imiquimod induced mouse model was investigated. According to clinical PASI scale we evaluate severity of inflammation.Combined with biopsy staining, we can inspect ear tissue hyperplasia of epidermis and inflammatory response more visually. Results indicate that the imiquimod induced mouse model exihibit different degree of swelling, thickening, scales, which have some similarity with psoriasis disease lesion.Resveratrol can obviously relieve the sympotoms of psoriatic lesion, reduce PASI score significantly; changes morphological performance to be normal, inhibit epidermis proliferation, reduce parakeratosis of epidermal cells and inflammatory cell Since the imiquimod induced psoriasis mouse model are related to enzyme system of the oxidative stress system, so the effect of resveratrol on SOD activity and MDA content of the skin tissue was discussed.Results show that after application of imiquimod,SOD activity of mice ear skin were reduced while while MDA content were increased.After treatment with resveratrol, SOD activity and MDA content in mice ear skin tissue can be returned to normal.All these observations indicate that resveratrol can correct abnormal oxidative stress which might be the possible therapeutic mechanism.The forth part:Pharmacokinetic study of resveratrol liposome gel after transdermal administrationMicrodialysis technology combined with high performance liquid chromatography was used to study pharmakokinetic analysis of resveratrol after transdermal administration.Since resveratrol is lipophilic,the perfusion fluid was optimized.We selected the appropriate perfusion fluid in order to improve the probe recovery of resveratrol.From the perspective of stability, the pH9 PBS was chosen as the final perfusion fluid. Effect of perfusion flow rate on in vitro and in vivo probe recovery was investigated,and the 2μl/min was established as perfusion rate. Effect of concentration on in vitro and in vivo probe recovery was investigated, the results showed that concentration had no influence on probe recovery.We also investigated stability of the home-made probe to confirm that MD is suitable for pharmacokinetic analysis of resveratrol.Resveratrol liposome gel or ordinary gel were topically administrated to SD rats(0.5g/100g),and microdialysis sampling were collected at different time points.The concentration of resveratrol in dialysate were determined by HPLC and the real concentration of resveratrol in rat skin were calculated by in vivo probe recovery. According to noncompartmental analysis, the pharmacokinetic parameters of resveratrol liposome gel and ordinary gel were calculated using 3P97 software. Tmax、Cmax、T1/2、AUC0-t、MRT of resveratrol liposome gel are 327.2±9.4min、91.7±3.2 ng.ml-1、453.8±10.9min、688323.6 ±2274.5 ng.min.ml-1、395.3±11.6 min respectively. Tmax、Cmax、T1/2、AUC0-t、MRT of resveratrol gel are 290.8±8.2 min、75.8±5.1 ng.ml-1、389.5±14.5 min、558455.2±3481.6 ng.min.ml-1、368.5±14.1 min respectively.The results showed that retention time and drug concentration in the skin of resveratrol liposome gel were significantly higher than that of the ordinary gel.The liposome gel has a good slow release effect.This study evaluated the mechanism and therapeutic effect of resveratrol for treatment of psoriasis from the level of cell and animal.Resveratrol liposome gel was firstly prepared, and the pharmacokinetic analysis were studied using microdialysis techonology All these laid a foundation for the future development and utilization of this compound. |
PDF Full Text Request