颗粒蛋白前体小干扰RNA提高小鼠皮肤创面修复的质量
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摘要
为探讨颗粒蛋白前体(progranulin,PGRN)小干扰RNA对提高小鼠切创损伤创面愈合质量的作用,通过选取33只健康雄性BALB/c小鼠,制作小鼠背部皮肤孔状切除模型,按照处理因素不同随机分为3组:健康小鼠对照组(Control组)、溶剂+皮肤孔状切除组(Vehicle组)和Si-m-PGRN+皮肤孔状切除组(Si-m-PGRN组)。分别于造模后0、4、8、12、16 d背部皮下注射30μL的生理盐水或Si-m-PGRN。各组小鼠分别于4、8、12、16、20 d麻醉处死取材,采用大体观察、常规组织学检查和免疫组织化学染色等方法研究了创面愈合情况。结果表明:注射Si-m-PGRN后,PGRN在伤口处表皮和真皮中表达较低。Si-mPGRN 组与 Vehicle 组在伤后4、8、12、16 d相比,创面面积缩小明显,愈合率较快,新生表皮的细胞层数较多,表皮较厚,分层明显,伤口的再上皮化加速,上皮层次明显;伤后20 d,Vehicle 组和Si-m-PGRN组的上皮厚度与Control组相比显著较厚,而两组之间创面面积、愈合率,新生表皮的厚度均无差异。可见PGRN小干扰RNA分子在皮肤损伤愈合前期能提高小鼠皮肤伤口愈合率、增加新生表皮厚度,加快伤口的再上皮化速度,而在皮肤损伤愈合后期无明显作用,表明PGRN小干扰RNA在一定程度上可以显著提高创面愈合的质量。
In order to investigate the effects of progranulin( PGRN) siRNA on the quality of wound healing in mice,according to different treatment factors,a total of 33 male,wild-type BALB/c mice with excisional wounds were randomly assigned into three groups: healthy control group( control group),physiological saline + pore resection group( vehicle group) and Si-m-PGRN + pore resection group( Si-m-PGRN group). 30 μL of saline or sim-PGRN were subcutaneously injected into the back at 0,4,8,12,16 d after injury. Mice from each group were sacrificed by intraperitoneal injection with an overdose of sodium pentobarbital on days 4,8,12,16,20 d after surgery. The area of the wounds was measured,the histological examination was performed,and the immunohistochemical examination was carried out to detect the positive staining of PGRN to evaluate the velocity and quality of wound healing. The results show that the expression of PGRN in the epidermis and dermis is lower after Si-m-PGRN injected. Compared with vehicle group,on 4,8,12,16 d after surgery,the wound area is significantly reduced,the healing rate is faster,the number of cell layers of new epidermis is more,the epidermis is thicker,the layers are obvious,the re-epithelialization of wound is accelerated,and the epithelial layer is obvious in the Si-m-PGRN group. At 20 d,the epithelial thickness of vehicle group and Si-m-PGRN group is significantly thicker,compared with those in the control group. There is no significant difference in the wound area,the healing rate and the thickness of neoepidermis between the Si-m-PGRN and vehicle group at 20 d post-injury. It is concluded that PGRN siRNA can improve the healing rate of skin wounds,increase the thickness of new epidermis and accelerate the reepithelialization of wounds in the early stage of skin wound healing,but has no obvious effect in the late stage. It shows that PGRN siRNA plays a certain role in improving the quality of wound healing.
In order to investigate the effects of progranulin( PGRN) siRNA on the quality of wound healing in mice,according to different treatment factors,a total of 33 male,wild-type BALB/c mice with excisional wounds were randomly assigned into three groups: healthy control group( control group),physiological saline + pore resection group( vehicle group) and Si-m-PGRN + pore resection group( Si-m-PGRN group). 30 μL of saline or sim-PGRN were subcutaneously injected into the back at 0,4,8,12,16 d after injury. Mice from each group were sacrificed by intraperitoneal injection with an overdose of sodium pentobarbital on days 4,8,12,16,20 d after surgery. The area of the wounds was measured,the histological examination was performed,and the immunohistochemical examination was carried out to detect the positive staining of PGRN to evaluate the velocity and quality of wound healing. The results show that the expression of PGRN in the epidermis and dermis is lower after Si-m-PGRN injected. Compared with vehicle group,on 4,8,12,16 d after surgery,the wound area is significantly reduced,the healing rate is faster,the number of cell layers of new epidermis is more,the epidermis is thicker,the layers are obvious,the re-epithelialization of wound is accelerated,and the epithelial layer is obvious in the Si-m-PGRN group. At 20 d,the epithelial thickness of vehicle group and Si-m-PGRN group is significantly thicker,compared with those in the control group. There is no significant difference in the wound area,the healing rate and the thickness of neoepidermis between the Si-m-PGRN and vehicle group at 20 d post-injury. It is concluded that PGRN siRNA can improve the healing rate of skin wounds,increase the thickness of new epidermis and accelerate the reepithelialization of wounds in the early stage of skin wound healing,but has no obvious effect in the late stage. It shows that PGRN siRNA plays a certain role in improving the quality of wound healing.
引文
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19 Alquézar C,de la Encarnación A,Moreno F,et al.Progranulin deficiency induces overactivation of WNT5A expression via TNF-α/NF-κB pathway in peripheral cells from frontotemporal dementialinked granulin mutation carriers[J].Journal of Psychiatry&Neuroscience,2016,41(4):225-239
2 Castellano-Pellicena I,Uzunbajakava N E,Mignon C,et al.Does blue light restore human epidermal barrier function via activation of Opsin during cutaneous wound healing?[J].Lasers in Surgery and Medicine,2018,doi:10.1002/lsm.23015
3 Hassanzadeh H,Matin M M,Naderi-Meshkin H,et al.Using paracrine effects of Ad-MSCs on keratinocyte cultivation and fabrication of epidermal sheets for improving clinical applications[J].Cell and Tissue Banking,2018,19(4):531-547
4 Jing H,Tan M S,Yu J T,et al.The role of PGRN in alzheimer's disease[J].Molecular Neurobiology,2016,53(6):4189-4196
5 Ichimura Y,Asano Y,Akamata K,et al.Progranulin overproduction due to Fli-1 deficiency contributes to the resistance of dermal fibroblasts to tumor necrosis factor in systemic sclerosis[J].Arthritis&Rheumatology,2015,67(12):3245-3255
6 Wei J,Hettinghouse A,Liu C.The role of progranulin in arthritis[J].Annals of the New York Academy of Sciences,2016,1383(1):5-20
7 Korolczuk A,Betowski J.Progranulin,a new adipokine at the crossroads of metabolic syndrome,diabetes,dyslipidemia and hypertension[J].Current Pharmaceutical Design,2017,23(10):1533-1539
8 Kong L,Zhao Y P,Tian Q Y,et al.Extracellular matrix protein 1,a direct targeting molecule of parathyroid hormone-related peptide,negatively regulates chondrogenesis and endochondral ossification via associating with progranulin growth factor[J].FASEB Journal,2016,30(8):2741-2754
9 He Z,Ong C H,Halper J,et al.Progranulin is a mediator of the wound response[J].Nature Medicine,2003,9(2):225-229
10 Li S S,Wang L L,Liu M,et al.Cannabinoid CB2 receptors are involved in the regulation of fibrogenesis during skin wound repair in mice[J].Molecular Medicine Reports,2016,13(4):3441-3450
11 Wang L L,Zhao R,Li J Y,et al.Pharmacological activation of cannabinoid 2 receptor attenuates inflammation,fibrogenesis,and promotes re-epithelialization during skin wound healing[J].European Journal of Pharmacology,2016,786:128-136
12 Nagelschmidt M,Becker D,B9nninghoff N,et al.Effect of fibronectin therapy and fibronectin deficiency on wound healing:A study in rats[J].The Journal of Trauma,1987,27(11):1267-1271
13 Kiritsi D,Nystr9m A.The role of TGFβin wound healing pathologies[J].Mechanisms of Ageing and Development,2018,172:51-58
14 Eming S A,Wynn T A,Martin P.Inflammation and metabolism in tissue repair and regeneration[J].Science,2017,356(6342):1026-1030
15 Gurtner G C,Werner S,Barrandon Y,et al.Wound repair and regeneration[J].Nature,2008,453(7193):314-321
16 Seo G Y,Lim Y,Koh D,et al.TMF and glycitin act synergistically on keratinocytes and fibroblasts to promote wound healing and antiscarring activity[J].Experimental and Molecular Medicine,2017,49(3):e302
17 Ansell D M,Campbell L,Thomason H A,et al.A statistical analysis of murine incisional and excisional acute wound models[J].Wound Repair and Regeneration,2014,22(2):281-287
18 Galiano R D,Michaels J T,Dobryansky M,et al.Quantitative and reproducible murine model of excisional wound healing[J].Wound Repair and Regeneration,2004,12(4):485-492
19 Alquézar C,de la Encarnación A,Moreno F,et al.Progranulin deficiency induces overactivation of WNT5A expression via TNF-α/NF-κB pathway in peripheral cells from frontotemporal dementialinked granulin mutation carriers[J].Journal of Psychiatry&Neuroscience,2016,41(4):225-239