| Background:As the largest organ and the first line of defense of the human body,the skin is most vulnerable to external aggressions,and the oral and maxillofacial areas are more susceptible to skin trauma because they are in a relatively exposed state.Facial defects resulting from maxillofacial skin injuries can cause both physical and psychological pain to patients.Although most common skin defects can be restored to their original shape within a short period of time through the body’s self-healing ability,it is difficult for adults to fully restore their original shape and function after skin injury,just like infants.Therefore,clinically,when facing patients with skin trauma,the healing time should be shortened as much as possible,while achieving the reconstruction of skin function.Wound dressing is a kind of material to cover the wound,which can play a barrier role to the wound,keep the wound moist and prevent the invasion of foreign bacteria,and also control the release of drugs or growth factors so that they can act on the wound site for a long time,so the reasonable use of wound dressing can accelerate the process of wound repair.Because of its good biocompatibility,hydrophilicity and three-dimensional pore structure,hydrogel can mimic extracellular matrix and facilitate the migration,adhesion and proliferation of cells,and the research on its use as wound dressing has gradually increased in recent years.However,hydrogel alone as a dressing may not achieve particularly ideal effects,and the repair of skin wounds can be roughly divided into four consecutive and coordinated stages:hemostasis,inflammation,new tissue proliferation and tissue remodeling,so we consider adding cells and bioactive substances that can damage regulate the healing process,and using hydrogel as a carrier,so as to achieve a continuous effect in each stage of wound repair.Many studies have shown that mesenchymal stem cells(MSCs)have a strong capacity for self-renewal and multilineage differentiation,and their ability to enhance skin wound healing has been well documented.Among them,adipose derived stem cells(ADSCs)have the advantages of high quantity,easy access and rapid proliferation compared with other sources such as bone marrow-derived and umbilical cord-derived stem cells,and have a wide prospect in the application of skin wounds.Substance P(SP)is a neuropeptide containing 11 amino acids,which is involved in regulating many pathological and physiological processes in the organism.In recent years,some researchers have suggested that SP is an"injury-inducing factor"that initiates endogenous repair after tissue damage,and that SP is able to recruit stem cells,regulate inflammation and promote vascularization,and has a regulatory effect on all stages of skin tissue regeneration.However,both ADSCs and SP are time-sensitive and cannot persist in vivo for a long period of time,so a combination of all three is considered for skin injury repair.Objective:To prepare methacrylate gelatin(Gel MA)mixed with silk fibroin(SF)photocrosslinked hydrogels loaded with adipose derived stem cells(ADSC)and substance p(SP)as dressing,and its role in the injury repair process was investigated by in vivo and in vitro experiments.Methods:1.Gel MA was synthesized using gelatin and methacrylic anhydride,and SF was extracted from cocoons to prepare 10%Gel MA and 10%SF solution;the two were mixed in different proportions and sonicated to prepare G-S hydrogel.2.The hydrogels were characterized by ~1H NMR,scanning electron microscopy(SEM),and solubilization,compression and degradation experiments were performed.3.Adipose tissue was extracted from the groin of SD rats,digested and cultured to obtain ADSC.4.The ratios of G-S hydrogels were screened by CCK-8 experiment.5.CCK-8 assay and live/dead cell staining verified the effect of SP-loaded G-S hydrogels on ADSC proliferation;scratch assay verified,the effect of G-S@SP hydrogels on migration of ADSCs;inoculation of ADSCs onto hydrogels for culture,staining of nuclei and cytoskeleton,and investigation of the effect of G-S@SP hydrogels on ADSC adhesion and cell extension.6.The anti-inflammatory effect of G-S@SP/ADSC hydrogels loaded with SP,ADSC was evaluated in vitro by q RT-PCR,cellular immunofluorescence staining.7.The in vitro vascularization-promoting effect of G-S@SP/ADSC hydrogels was assessed by scratch assay,transwell assay,q RT-PCR,tubule formation assay,immunofluorescence staining.8.The effect of G-S@SP/ADSC hydrogel on skin wound repair was evaluated by rat back skin trauma model.Results:1.Gel MA was successfully synthesized and SF was successfully extracted.G-S hydrogel was synthesized.2.The hydrogel with G7-S3 ratio was selected for subsequent experiments after combining CCK-8 results and material properties.3.CCK-8,scratch assay,adhesion assay and cytoskeleton staining demonstrated that G-S@SP hydrogel could promote ADSC proliferation,migration and adhesion,and facilitate cell extension.4.In vitro experiments demonstrated that G-S@SP/ADSC hydrogel has anti-inflammatory and vascularization-promoting effects in vitro.5.In vivo experiments showed that G-S@SP/ADSC hydrogel promoted the healing of skin wounds in rats,had certain inflammation regulation ability and could promote tissue vascularization.Conclusion:ADSC is a kind of stem cells commonly used in tissue engineering to promote tissue repair,and SP is a kind of"damage inducing factor"that can initiate endogenous repair.G-S hydrogel as a scaffold material has good biocompatibility and provides a growth environment for ADSC inoculated inside the hydrogel.By adding SP into the hydrogel,it can not only achieve the purpose of slow release,but also play a synergistic role with ADSC to promote tissue repair,and finally construct a wound dressing with anti-inflammation and promote tissue vascularization,which can effectively accelerate wound healing in rat skin. |
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