4种品系小鼠角质形成细胞原代培养比较研究  

Comparative study on primary culture of four strains ofmouse keratinocytes

作　　者：关艳玲 章萍萍 陈秀 蒋励萍 魏伟[1] 马旸[1] Guan Yanling;Zhang Pingping;Chen Xiu;Jiang Liping;Wei Wei;Ma Yang(Institute of Clinical Pharmacology,Anhui Medical University,Key Laboratory of Anti-inflammatory and Immune Medicine,Ministry of Education,Anhui Collaborative Innovation Center for Anti-inflammatory and Immune Drugs,Rheumatoid Arthritis Research Center,Anhui Medical University,Hefei 230032)

机构地区：[1]安徽医科大学临床药理研究所,抗炎免疫药物教育部重点实验室,安徽抗炎免疫药物协同创新中心,安徽医科大学类风湿关节炎研究中心,合肥230032

出　　处：《安徽医科大学学报》2023年第6期989-995,共7页Acta Universitatis Medicinalis Anhui

基　　金：国家自然科学基金(编号:81502123、81673444);安徽省高校科学研究项目(编号:KJ2019A0234);出生人口健康教育部重点实验室开放课题(编号:JKZD20212);抗炎免疫药物教育部重点实验室开放课题(编号:KFJJ-2020-12、KFJJ-2021-09);皮肤病学教育部重点实验室开放课题(编号:AYPYS2021-2)。

摘　　要：目的体外分离、培养并鉴定成年小鼠背部皮肤角质形成细胞(KC);对比4种品系小鼠KC的增殖与分化能力。方法剪取4种品系成年小鼠背部皮肤,剥离皮下脂肪组织,0.25%胰蛋白酶消化获得KC;免疫荧光法鉴定KC;显微镜下观察细胞生长状况;设计不同的种板时间和密度,使用CCK-8法、EdU和高内涵法检测细胞增殖活力;通过Western blot检测4种成年小鼠KC分化情况。结果对4种品系成年小鼠背部相同面积的皮肤组织进行胰蛋白酶消化,昆明(Kunming,KM)小鼠提取的KC数量最低,C57BL/6提取的细胞数量最多;BALB/c、KM、裸小鼠分别以8×10^(3)个/孔的密度铺板增殖效率较高,C57BL/6以1.6×10^(4)个/孔的密度铺板增殖效率较高;4种品系小鼠,BALB/c小鼠原代KC较C57BL/6小鼠原代KC增殖能力弱,分化程度差异无统计学意义。结论采用简便易行的方法成功分离并比较4种成年小鼠背部皮肤原代KC的增殖和分化能力,为皮肤相关疾病模型小鼠的品系选择提供一定实验依据。Objective To isolate,culture and identify adult mouse dorsal keratinocyte(KC)in vitro and to compare of the proliferation and differentiation capabilities of four adult mouse dorsal skin KC.Methods Four kinds of adult mouse dorsal skin were taken,the subcutaneous adipose tissue was peeled,and KC was obtained by 0.25%trypsin digestion;KC were identified using immunofluorescence method,cell growth status was observed under microscopy,different seed plate times and densities were designed,and cell proliferation viability was detected by CCK-8,EdU and high connotation method.Four adult mouse KC differentiation was detected by Western blot.Results Trypsin digestion was used to extract four strains of adult mouse back skin KC,and the Kunming(KM)mice had the lowest number of KC and C57BL/6 to extract the highest number of cells for the same area of skin tissue.BALB/c,KM,and nude mouse had higher plate-laying efficiency at a density of 8×10^(3)/well,respectively,and C57BL/6 had higher plate-laying efficiency at a density of 1.6×10^(4)/well.Among the four strains of mice,the primary KC of BALB/c mice was weaker than that of C57BL/6 mice,and there was no statistical difference in the degree of differentiation.Conclusion The proliferation and differentiation capacity of four adult mouse primary KC on the dorsal skin is successfully isolated and compared by a simple and easy method,which provided a certain experimental basis for the strain selection of mice with skin-related diseases.

关 键 词：角质形成细胞 原代培养 小鼠表皮 增殖 分化 

分 类 号：R322.99[医药卫生—人体解剖和组织胚胎学]