| Hypertrophic scars result from abnormal repair of body tissue after trauma and are inevitable products of trauma repair process.Hypertrophic scars are pathological scars which may lead to itching,pain and other uncomfortable symptoms,thus affecting the appearance,or even scar contracture that may result in dysfunction.The formation of hypertrophic scars biologically manifests as collagen metabolism disorders.Matrix components of scar tissue include collagen fibers,structural proteins and proteoglycans,etc.;wherein collagen fibers are the main component of scar tissue,most of which is collagen fiber type I.The synthesis and decomposition of collagen is a complex dynamic process in which a variety of biologically active enzymes are involved.Under normal circumstances,collagen synthesis and decomposition within the wound fall into a dynamic balance;in some cases,this balance is broken,leading to reduced decomposition of collagen fibers and collagen synthesis faster than collagen decomposition,resulting in collagen deposition,thereby leading to formation of scar tissue.Recent studies have shown that a variety of abnormal expression of cytokines is closely related to scar formation,especially the transforming growth factor?(TGF-?).TGF-?directly stimulates angiogenesis,provokes fibroblast hypertrophy,and promotes synthesis of collagen,fibronectin and laminin from fibroblasts;on the other hand,TGF-?may inhibit the expression and activity of matrix metalloproteinase-1(MMP1),promote the expression of TIMP-1,thus inhibiting decomposition of collagen and other extracellular matrix;therefore,TGF-?is considered to be one of the important factors affecting wound surface healing.Exploring the mechanism of wound surface healing and scar formation and its treatment has become a hot topic in molecular biology,biophysics,biomedical engineering,tissue engineering and clinical medicine,etc.,and people are paying more attention to it.The current treatments are mainly divided into surgical treatments and non-surgical treatments.Non-surgical treatments include infrared and magnetic field therapy,laser therapy,sonophoresis,recompression therapy,cryotherapy and silicone therapy and other physical therapies,as well as pharmacotherapy,gene therapy,etc.5-FU which is a commonly used anti-tumor drugs is a kind of anti-metabolites.At present,there are many reports about the treatment of keloids with 5-FU at home and abroad.Nanda and Reddy used 5-FU to treat 28 patients with keloid for 12 weeks and achieved good results.Apikian and Good-man adopted injection of 5-FU for the treatment of keloid,and the difference was significant before and after the injection.Domestic scholars have conducted studies on concentration and time of 5-FU in the treatment of keloid,and achieved good results,which plays a certain guiding role in clinical application of 5-FU.Through interfering with DNA replication and transcription of scar fibroblasts,5-FU may inhibit the hypertrophy of fibroblasts and over expression of collagen,so as to reduce scar hypertrophy and collagen deposition.In other words,5-FU must enter the scar tissue from the outside,and then enter the fibroblasts in order to play a role in the inhibition of scar.Our previous studies show that,electrets may act as an effective and safe physical penetration enhancer to promote the drug to penetrate the skin through the dense cuticle structure and then enter the skin to play a pharmacodynamic effect.Among so many scar treatment methods,physical therapy is currently the most commonly used method,in which the application of electric field(pressure)in wound surface healing and scar treatment shows great potential;in 1843,dubois-reymond found current of 1mA intensity in human skin wound.It was later confirmed that the presence and maintenance of wound currents played an important role in wound surface healing.The application of electric stimulation therapy in wound surface healing had been recognized in U.S.since 2002.Electret is a kind of functional dielectric material which can maintain long-term polarization charge and store space charge.The electrostatic field and microcurrent generated by electrets act as exogenous physical factors in biological tissue and biological macromolecules with biological electret state,which may play a good biological effect and promote tissue rehabilitation.Clinically,the main dosage forms applied in pharmacotherapy for hypertrophic scars are local injection,unguent,cream and other traditional dosage forms.But each has its own limitations,therefore in this study,we selected transdermal dosage form,adopted anti-tumor drug 5-FU which may exactly exert effect on hypertrophic scars as a model drug,and studied the effect and mechanism of 5-FU combined with electrets in the treatment of hypertrophic scars.This paper is divided into the following four parts:1.Preparation,formulation optimization and property characterization of 5-FU patchesThis paper takes 5-FU as the model drug,Eudragit E100 as the matrix,tributyl citrate as the plasticizer and adopts the solvent evaporation method to prepare pressure-sensitive adhesive patches.During the preparation of the patches,the amount of E100 and the ratio of the plasticizer and E100 may affect properties of the patches.Variables which affect properties of the patches are identified by the single-factor exploration;the experiment through orthogonal test design is used,takes the primary adhesive,the lasting adhesive and the nature release of patches as indexes;the comprehensive scoring method is adopted to optimize the formulation of the patches,to determine the optimized one,and to characterize the properties of the patches.Results have showed that both the mass of E100 and the mass ratio between tributyl citrate and E100 affect the properties of the patches;through the comprehensive scoring method,the optimum of 5-FU patches under the condition of the mass of E100 is 0.30g and the ratio of tributyl citrate is 55%is identified.And the the patches through above methods are characterized,the properties obtained are shown as follows:the primary adhesive is(4.03±0.53)g,the lasting adhesive is(122.5±7.13)min.Then nature release of the patches is considered;the effects different polarities and surface potentials of electrets on the nature release of the patches are explored,and the modeling of the nature release of 5-FU patches is identified.Results have showed that,The regularity of release of 5-FU was in accordance with Higuchi formulation;The release of the patches after the effect of electrets increased;the release amount of the drug increased as surface potentials of electrets increased;no significant difference was identified between positive and negative polarities,and the regularity was also accordance with Higuchi formulation;Modeling was then established based on the release regularity of the patches,and results showed the modeling of the release regularity of the patches in this paper as follows:???Note:Q is referred as the accumulated amount of release,V is referred as surface potentials of electrets,t is referred as the time2.Research on the animal experiment dosage regimen of 5-fluorouracil patchesElectrets possess the function of improving local blood supply as well as growing conditions of epithelial cells;further,micro-currents conducted by electrets can stimulate the differentiation of epithelial cells.5-FU,however,possesses the function of inhibiting the replication of both DNA and RNA and the hypertrophy of cells.Thus,proper dosage regimen of electret 5-FU patches should be explored.A new dosage regimen should be explored through the method of measuring the healing rate of the wound,the variation in the thickness of wounded skin and the HE staining method to observe the changes of histomorphology.The results show that,Electrets can promote the wound contraction and present the positive correlation with the surface potentials of electrets,and no significant difference was identified between positive and negative polarities;The wounds of rat skin in 5-FU patch group healed slowly,which presented inhibitory effects of 5-FU on wound healing.HE staining method used to observe the changes of histomorphology showed the same conclusion with the healing rate of the wounds,the variation in the thickness of wounded skin.Thus,final dosage regimen was confirmed,and that was,electrets was applied on the wounds during the formation period of 0-14 days;electret 5-FU patches are applied on the wounds during the formation period of 14-42 days.3.Research on the inhibitory effects of electret 5-FU patches on the growth of hypertrophic scars of rat skin through zoology experimentsAfter the preparation of 5-FU patches and dosage regimen are identified,the zoology research on the inhibitory effect of 5-FU patches on the growth of hypertrophic scars is conducted.Samples are taken respectively from the dates after 1 week,two weeks,three weeks and four weeks;the hypertrophy index of hypertrophic scars is measured and is adoptedthroughtheHEstainingmethod,Massonmethodaswellas immunohistochemistry to explore inhibitory effects of 5-FU patches on the growth of hypertrophic scars of rat skin.The results of the hypertrophy index of hypertrophic scars show that,The hypertrophy index of hypertrophic scars from the control group gradually increased;The hypertrophy index of hypertrophic scars from the electret group presented the trend of decrease;The hypertrophy index of hypertrophic scars from the administration group decreased and the one from electret 5-FU group showed the best effect.The results of HE staining method show that,Hypertrophic tissue of rat skin from the control group continually proliferated,which were mainly characterized in the hypertrophy of epidermal and corium layer.Corium layer contained a large number of fibroblasts;the deposition of collagen fiber increased,presented the characteristics of thickness and densification with irregular order;Compared the 5-FU patches group with the control group,both epidermal and corium layer of hypertrophic tissue became thinner,the amount of fibroblasts of corium layer decreased,the deposition of collagen fiber decreased and the effect on the inhibitory effect of the hypertrophic scar hypertrophy was significant;Compared the patch group of different polarities and different surface potentials electrets with the 5-FU patch group,the effect was more significant,as surface potentials of electrets increased,the effect of hypertrophy of hypertrophic scars was more significant,the difference between groups of positive and negative electrets was not significant.The results of the immune group showed that,The expression of TGF-?contained in the hypertrophic tissue on rat in administration groups decreased,and that of Smad2 and Smad3 also decreased,but the expression of Smad7 increased,and that of ColI and ColIII decreased;The effect in electret 5-FU patch groups was more significant;The expression of TGF-?contained in the hypertrophic tissue on rat in administration groups decreased,but that of MMP1 increased,and the expression of MMP2?TIMP1?Col??Col?also increased,effects in electret 5-FU patch groups were more significant;Electret 5-FU patches significantly affected TGF-?/Smads?TGF-?/MMPs signal pathways formed by hypertrophic scars.4.Research on inhibitory effects of electret 5-FU patches on the growth of hypertrophic scars of rat skin through cytology experimentsZoology experiments has proved that electret 5-FU patches significantly affected TGF-?/Smads?TGF-?/MMPs signal pathways formed by hypertrophic scars.In order to further explore and verify the mechanism of inhibitory effects of electret 5-FU patches on the growth of hypertrophic scars,we conducted cytology experiments and studied through the methods of MMT,PCR and Western-Bolt.The experimental results of MMT show that,Electrets can promote cell proliferation;5-FU showed significant inhibitory effects on fibroblasts of hypertrophic scars.The experimental results of PCR show that,The expression of TGF-?in administration groups decreased,and that of Smad2 and Smad3 also decreased,but the expression of Smad7 increased,and that of ColI and ColIII decreased;The effect in electret 5-FU patch groups was more significant;The expression of TGF-?in administration groups decreased,but that of MMP1increased,and the expression of MMP2?TIMP1?Col??Col?decreased,effects in electret 5-FU patch groups were more significant;Electret 5-FU patches significantly affected TGF-?/Smads?TGF-?/MMPs signal pathways formed by hypertrophic scars.The experimental results of Western-Bolt show that,The expression of TGF-?in administration groups decreased,and that of Smad2 and Smad3 also decreased,but the expression of Smad7 increased,and that of ColI and ColIII decreased;The effect in electret 5-FU patch groups was more significant;The expression of TGF-?in administration groups decreased,but that of MMP1increased,and the expression of MMP2?TIMP1?Col??Col?decreased,effects in electret 5-FU patch groups were more significant;Electret 5-FU patches significantly affected TGF-?/Smads?TGF-?/MMPs signal pathways formed by hypertrophic scars.The results of these above experiments are in correspondence with those of zoology experiments,which indicates electret 5-FU patches can inhibit the growth of hypertrophic scars by inhibiting TGF-?/Smads?TGF-?/MMPs signal pathways formed by hypertrophic scars.In a word,this paper explores the mechanism of inhibitory effects of electret 5-FU patches on the growth of hypertrophic scars and provides new thoughts and methods for clinical therapy of hypertrophic scars. |
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