MEBT/MEBO Regulates The Expression Of MMPs Through MTORC Signaling Pathway In Chronic Refractory Wounds

Objective: To explore the repair mechanism of moist exposure therapy/Moist burn ointment(MEBT/MEBO)promoting chronic refractory wounds on the body through m TORC signaling pathway matrix metalloproteinases(MMP3,MMP8).Methods:(1)60 SPF Wistar rats were taken as experimental objects,and these rats were divided into blank group,control group,model group,rb-b FGF group,and MEBO group,with12 rats in each group.Model group,rb-b FGF group,MEBO group,and control group made a circular wound on the back.Subsequently,the model group,rb-b FGF group,and MEBO group were injected with hydrocortisone acetate injection immediately to form chronic refractory wounds.The control group did not inject hydrocortisone acetate injection to form acute wounds.The model group did not apply any medicine,the rb-b FGF group applied rb-b FGF gel,the MEBO group applied MEBO,the control group was covered with gauze of sodium chloride solution,and the blank group only had the skin prepared without trauma treatment.(2)The wound healing of the rats in each group was observed on the 3rd,7th,and 14 th days after modeling,and the wound tissues of the rats were excised and embedded in paraffin at these three time points,followed by Masson staining and HE staining.(3)Western Blot was used to detect the wound of each group of rats MMP3,MMP8 and p-m TORC in the wound tissue.(4)The real-time PCR method was used to detect the wound of each group of rats MMP3,MMP8,m TORC m RNA gene transcription in tissues.Results:(1)Wound healing rate: The wound healing rate of the rats in the four groups on the 7th and 14 th days after modeling was significantly higher than that on the 3rd day(all P<0.05).On the 3rd day after modeling,there was no significant difference in the wound healing rate between the control group,the model group,the rb-b FGF group,and the MEBO group(F=2.474,P=0.074).On the 7th and 14 th days after modeling,the wound healing rates of the control group,rb-b FGF group and MEBO group were higher than those of the model group(all P<0.05).(2)Wound histopathology: HE and Masson staining results showed that on the 3rd day after modeling,except for the blank group,the four groups had a large number of inflammatory cells,less new capillaries,and disordered arrangement of fibroblasts.Skin appendages are not generated.On the 7th day after modeling,the inflammatory cells in the control group,MEBO group,and rb-b FGF group were significantly reduced compared with the previous ones,capillary angiogenesis was present,and fibroblasts were arranged neatly.In the model group,there were more new capillaries,fewer fibroblasts and disordered arrangement,more inflammatory cells,and no skin appendages.On the 14 th day after modeling,the new capillaries in the control group,MEBO group,and rb-b FGF groups were neatly arranged,no infiltrating inflammatory cells were found,skin appendages were formed,and collagen fibers were significantly increased and arranged neatly.A small amount of lymphocytes were still infiltrated in the tissue of the model group,and the arrangement of capillaries and collagen fibers was disordered.(3)Western blot results showed that the protein expressions of MMP3,MMP8,and p-m TORC were significantly higher on the 7th day after modeling in the MEBO group than on the 3rd day,and the protein expressions of MMP3,MMP8,and p-m TORC on the 14 th day were significantly higher.Compared with the 7th day and the 3rd day,the expression of these proteins increased first and then decreased over time(P<0.05).However,the expression levels of MMP3,MMP8,and p-m TORC proteins in the model group were significantly higher on the 7th and 14 th days after modeling than on the 3rd day,and the expression of these proteins continued to increase over time(P<0.05).On the 3rd day after modeling,the protein expression levels of MMP3 and MMP8 in the control group,rb-b FGF group,and MEBO group were obviously higher than those in the model group(P<0.05).The protein expression levels of MMP3 and MMP8 in the MEBO group were significantly higher than those in the rb-b FGF group(P<0.05),but the p-m TORC protein expression in the MEBO group was not obviously different from that in the rb-b FGF group(P>0.05).On the 7th day after modeling,the protein expression levels of MMP3 and MMP8 in the control group,rbb FGF group,and MEBO group were obviously higher than those in the model group(P<0.05).The protein expression levels of MMP3 and MMP8 in the MEBO group were obviously higher than those in the rb-b FGF group(P<0.05),but the p-m TORC protein expression in the MEBO group was not obviously different from that in the rb-b FGF group(P>0.05).(4)The results of real-time PCR showed that the transcription levels of MMP3 m RNA,MMP8 m RNA gene in MEBO group on the 7th day were obviously higher than those on day 3(P<0.05);with the passage of time,the expression of these genes increased first and then decreased.In the model group,the transcription levels of MMP3 m RNA,MMP8 m RNA were obviously higher on the7 th day than on the 3rd day(P<0.05).At day 3,7,and 14 of intervention,there was no significant difference in m TORC m RNA expression level in wound tissue among all groups(ALL P> 0.05).Conclusion:(1)MEBT/MEBO treatment can effectively promote the healing of chronic refractory wounds,reduce inflammatory cell reaction,and promote the generation of capillaries,collagen fibers,hair follicles and sebaceous glands.(2)MEBT/MEBO can up-regulate the protein expressions of MMP3,MMP8,p-m TORc and m RNA transcription levels of MMP3 and MMP8,thereby eliminating inflammation,promoting fibroblast and capillary formation,and ultimately promoting wound healing.(3)MEBT/MEBO can activate the m TORC signaling pathway,and then up-regulate the expressions of MMP3 and MMP8 to accelerate the healing speed.