CAJ | 학술논문

目的探讨缺氧诱导丝裂原因子(hypoxia-induced mitogenic factor,HIMF)对胎肺形态发育及表面活性蛋白B(surfactant protein B,SP-B)、表面活性蛋白C(snrfactant protein C,SP-C)表达的调控作用.方法体外分离、培养小鼠第13.5天胎肺组织,分为HIMF处理组:在培养基中添加重组HIMF蛋白,终浓度为100 nmol/L;对照组培养基中不加任何处理.分别在培养0、48、72 h后,每组每时间点选取20例胎肺采用倒置显微镜和HE染色观察胎肺形态学和组织学的改变,采用Western印迹、免疫组化、实时逆转录-聚合酶链反应技术检测胎肺组织中SP-B和SP-C的蛋白、mRNA表达水平变化.结果 HIMF处理组培养48、72 h后,肺泡数目分别为(14.37±0.85)和(18.41±1.24)个/高倍视野,肺泡分支的数目分别为(2.51±0.35)和(3.28±0.51)个/高倍视野,均较对照组相应时间点[分别为(8.09±0.92)、(9.54±0.78)、(1.45±0.32)和(1.69±O.43)个/高倍视野]增加,差异有统计学意义(P<0.05).对照组胎肺体外培养72 h后,SP-B、SP-C蛋白表达较少,染色强度弱,主要定位于Ⅱ肺泡上皮细胞;HIMF处理组可见Ⅱ型肺泡上皮细胞内SP-B、SP-C弥漫表达,以靠近肺组织边缘为甚.HIMF处理组培养48和72 h后,胎肺组织中SP-B、SP-C蛋白和mRNA水平均较对照组相应时间点增加,差异有统计学意义(P<0.05).结论 HIMF可能通过上调胚胎组织中SP-B和SP-C的表达,促进胎肺形态发育,为进一步研究HIMF在胚胎肺泡发育和成熟中的作用奠定了基础.
목적 탐토결양유도사렬원인자(hypoxia-induced mitogenic factor,HIMF)대태폐형태발육급표면활성단백B(surfactant protein B,SP-B)、표면활성단백C(snrfactant protein C,SP-C)표체적조공작용.방법 체외분리、배양소서제13.5천태폐조직,분위HIMF처리조:재배양기중첨가중조HIMF단백,종농도위100 nmol/L;대조조배양기중불가임하처리.분별재배양0、48、72 h후,매조매시간점선취20례태폐채용도치현미경화HE염색관찰태폐형태학화조직학적개변,채용Western인적、면역조화、실시역전록-취합매련반응기술검측태폐조직중SP-B화SP-C적단백、mRNA표체수평변화.결과 HIMF처리조배양48、72 h후,폐포수목분별위(14.37±0.85)화(18.41±1.24)개/고배시야,폐포분지적수목분별위(2.51±0.35)화(3.28±0.51)개/고배시야,균교대조조상응시간점[분별위(8.09±0.92)、(9.54±0.78)、(1.45±0.32)화(1.69±O.43)개/고배시야]증가,차이유통계학의의(P<0.05).대조조태폐체외배양72 h후,SP-B、SP-C단백표체교소,염색강도약,주요정위우Ⅱ폐포상피세포;HIMF처리조가견Ⅱ형폐포상피세포내SP-B、SP-C미만표체,이고근폐조직변연위심.HIMF처리조배양48화72 h후,태폐조직중SP-B、SP-C단백화mRNA수평균교대조조상응시간점증가,차이유통계학의의(P<0.05).결론 HIMF가능통과상조배태조직중SP-B화SP-C적표체,촉진태폐형태발육,위진일보연구HIMF재배태폐포발육화성숙중적작용전정료기출.
Objective To explore the regulatory effects of hypoxia-induced mitogenie factor(HIMF)on lung morphogenesis and expression o{surfactant protein B(SP-B)and surfactant protein C(SP-C)in embryonic lungs of mouse.Methods The mouse lungs at embryonic day 13.5 were separated and cultured in vitro,and divided into HIMF treatment group with final concentration as 100 nmol/L and control group with no HIMF.After incubation for 0,48 and 72 hours,cultured embryonic lungs from each group were harvested for HE staining-mediated morphological observatiom The changes of SP-B and SP-C expression in embryonic lungs were explored by western blot,immunohistoehemieal staining and real-time RT-PCR Results Administration of HIMF for 48 and 72 hours,morphogenesis were(14.37±0.85)and(18.41±1.24)/HP,and branching of the buds were(2.51±0.35)and(3.28±0.51)/HP,higher than those at the relative time points in control group,which were(8.09±0.92),(9.54±0.78),(1.45±0.32)and(1.69±0.43)/HP,respectively(P<0.05).In control embryonic lungs cultured for 72 hours.the expression of SP-B and SP-C was reduced,mainly locating at typeⅡ lung epithelial cells.Within the HIMF-treated embryonic lungs,there were intensive and diffuse expression of SP-B and SP-C,mainly around the edge of lung tissues.Compared with controls,the levels of SP-B protein and mRNA,SP-C protein and mRNA within HIMF-treated embryonic lungs for 48 and 72 hours were increased(P<0.05). Gonclusions HMF may promote lung morphogenesis via enhancing the expression of SP-B and SP-C in cultured embryonic lungs.which establishes a basis for further studying the roles of HIMF in lung alveolar development and maturation.