| Scar often occurs after trauma and is the product of tissue repair.It has a great impact on patients and is also one of the medical problems that need to be solved urgently in various disciplines.Pathological scars can be divided into hypertrophic scars and keloids.This paper mainly studies hypertrophic(HP)scars.HP scars generally grow after wound healing and are higher than surrounding skin tissues.They are different in thickness and irregular in shape.Early signs are hyperemia and swelling of scars,red or purple,accompanied by severe itching and tough texture.Studies have shown that the formation of hypertrophic scar is closely related to abnormal expression of various factors,excessive proliferation of inflammatory cells and fibroblasts,and excessive deposition of collagen type I and collagen type ?.At present,the main methods of scar treatment are surgical treatment and non-surgical treatment.Surgical treatment has limited efficacy and is prone to recurrence;non-surgical treatment includes physical therapy such as infrared,magnetic field,laser therapy,and other methods such as drugs.Among them,physical therapy is more commonly used,among which electric field(pressure)has great potential to promote wound healing and scar treatment.Electret is a kind of dielectric materials which can provide stable electrostatic field and microcurrent.The electrostatic field generated by electret can promote the microcirculation of wound site,promote the growth of fibroblasts,induce apoptosis of fibroblasts,inhibit growth of bacteria and promote the percutaneous penetration of drugs.Its biological effect will have a good effect on wound and scar treatment.5-fluorouracil(5-FU)is the earliest anticancer drugs.It can inhibit the synthesis of DNA after acting on cells,thus affecting the growth and proliferation of fibroblast cells.It has been reported that 5-FU can treat scars.In this study,electret and 5-FU were combined to make transdermal drug delivery patch.After successful establishment of scar model of rabbit ear,animal and immunohistochemistry experiments were carried out by means of scar hypertrophy index measuring,HE staining,Masson staining and immunohistochemistry analysis to study the effect of electret and electret in combination with 5-FU to inhibit the growth of scar.After the establishment of scar model of rat back skin,isolation of fibroblasts from normal and scar skin tissues,cytological experiments were carried out.The effects of electret and electrets in combination with 5-FU on cell proliferation,cell cycle,cell migration,apoptosis and apoptotic gene expression were investigated by flow cytometry in order to understand the mechanism of electrostatic field of electret and the combination of electret with 5-FU on scar formation at cell level.The main contents of this paper include:(1)Preparation of patchesEudragit E100,butyl triester citrate,5-Fu solid powder and 3% azone were dissolved in anhydrous ethanol.Then the system was cooled and coated on polypropylene(PP)film(backing layer),dried naturally at room temperature and covered with anti-sticking layer to obtain the 5-Fu patch.A series of electrets were fabricated by injecting PP film with different grid voltages in a grid-controlled constant voltage corona charging system.The electret 5-FU patches were prepared by combining the backing surface of 5-Fu patch with the charging surface of electret with different surface potential.(2)Animal experimentsAfter anesthesia of New Zealand white rabbits,a circular wound with a diameter of 1.0 cm was made on the inner ear of the perforator rabbits.The whole skin was removed by a scalpel to cartilage and healed naturally.Scar could be formed after 4 weeks.In vivo experiments were carried out using natural healing scar as control group,±5000V electret patch,5-FU,±5000V electret 5-FU patch as experimental groups.Scar hyperplasia index measuring,HE staining,Masson staining and immunohistochemical detection were carried out respectively.The results were as follows:The results of scar hyperplasia index measurement indicated that electret,5-Fu and their combination could reduce the index,and the order of reduction was electret 5-FU patch group > 5-FU patch group > electret group,and there was no significant difference between positive and negative electrets.HE staining results showed that:(1)The control group showed typical hypertrophic scar tissue structure.(2)5000V electret inhibited HP scar growth,and there is no significant difference between the effects of ±5000V electret.(3)5-FU patch had stronger inhibitory effect.(4)±5000V electret 5-FU patch had stronger inhibitory than 5-FU,and there was also no significant difference between the effect of positive and negative electrets.The results of Masson staining was the same as HE staining.Immunohistochemical results showed that:(1)The contents of collagen I,collagen ? and TGF-beta in normal skin tissue were very low.(2)Compared with the normal skin,the contents of collagen I,collagen ? and TGF-beta in contro group were increased significantly;After application of ±5000V electrets,the expression of the three factors was slightly lowered;there was no significant difference in the effect between positive and negative electrets.(3)Compared with electret group,the contents of the three factors in 5-FU group were significantly lowered.(4)±5000V electret 5-FU group had the lowest content,and there was no significant difference in the effect between positive and negative electrets.(3)Cytological experimentsScar can be formed four weeks after creation of wound on back of the SD rats.The fibroblasts of normal and hypertrophic scar skin tissue cultured by adherent method grew well,which could be used for follow-up experimental study.The results are as follows:The results of cell proliferation test showed that positive electret and 5-FU reduced the proliferation of the two kinds of cells,and 5-FU had stronger effect;negative electret promoted the proliferation of the two kinds of cells;positive electret combined with 5-FU had stronger inhibitory effect;negative electret combined with 5-FU had lower inhibitory effect than 5-FU group.The results of cell cycle test showed that the proportion of G1 phase cells decreased and that of G2 phase cells increased in-5000 V electret;the proportion of G1 phase cells increased in different degrees in other groups,and that of G2 phase cells decreased in different degrees;the proportion of G1 phase cells in each group was in the order of + 5000 V electret 5-FU group > 5-FU group > 5-5000 V electret 5-FU group > + 5000 V electret > control group > 5000 V electret group.The proportion of G2 phase increased in this order,and the change of S phase was irregular.The results of cell migration test showed that +5000V electret decreased the mobility of the cell,while-5000 V electret increased the cell mobility,and 5-FU group had lower mobility than that of the +5000V electrets group.The order of cell mobility in each group was-5000 V electrets group > control group > 5000 V electret group>-5000 V electrets 5-FU group > 5-FU group > 5000 V electret 5-FU group.The results of apoptotic test showed that there were no apoptotic cells or few apoptotic cells in the control group;+5000V electret showed a few apoptotic cells;-5000 V electret showed no apoptotic cells;5-FU group had more apoptotic cells;+5000V electret 5-FU group had more apoptotic cells than 5-FU group;-5000 V electret 5-FU group had lower apoptotic cells than 5-FU group,but higher apoptotic cells than +5000V electret group.The two kinds of cells have the same regularity.The apoptotic genes,including P53,fas/fasl,Bax,were detected in this study.The results showed that the relative expression of four apoptotic genes increased in +5000V electret group,but decreased in 5000 V electret group.The expression of four genes increased significantly in 5-FU group.After application of +5000V electret 5-FU patch,the expressions were more than those of in 5-FU group.However,combination of-5000 V electret with 5-FU led to lower expression of the gene than that of in the 5-FU group.In conclusion,animal experiments in vivo confirmed that ±5000V electret,5-FU and ±5000V electret combined with 5-FU could inhibit the growth of scar tissue in rabbit ears,among which ±5000V electret combined with 5-FU had the best effect.Cytological experiments confirmed that negative electret could promote the growth of fibroblasts in normal and scar skin tissues of rats,and reduce the expressions of four apoptotic genes,P53,fas/fasl,Bax,to inhibit the apoptosis of two kinds of cells;5-FU mainly inhibits the division of G1 phase fibroblasts,thus delaying the proliferation of two kinds of fibroblasts,inhibiting the growth of scars,and can promote the apoptosis of two kinds of fibroblasts by promoting the expression of the four apoptotic genes mentioned above;5-FU combined with positive electret enhances the apoptosis of G1 phase cells.The inhibitory effect of fissure further delayed the proliferation of the two fibroblasts,and enhanced the apoptotic effect of 5-FU on the two fibroblasts by promoting the expression of four apoptotic genes,that is,5-FU and positive electrets had synergistic effect on the growth inhibition of hypertrophic scars.Negative electret can not only inhibit the growth of scar,but also regulate the toxicity of 5-FU to cells and control the growth rhythm of scar tissue.Negative electret may inhibit the growth of scar tissue by inhibiting inflammation and improving microcirculation of scar tissue. |
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