摘要
目的 研究血红素加氧酶-1（HO-1）对原代培养大鼠肺泡Ⅱ型上皮细胞（AEC-Ⅱ）生长的直接影响.方法 免疫黏附法纯化SD大鼠AEC-Ⅱ,细胞进行活性和纯度鉴定后,随机分为七组：A组（正常对照组）;B组（H2O2组,0.5 mmol/L H2O2）;C1～5组（0.01～50 μmol/L HO-1）.各组进行细胞形态学观察、细胞计数和以MTT法测定OD值.结果 ①免疫黏附法可收获AEC-Ⅱ（2～2.5）×107个/鼠,活性和纯度均＞90%;电镜下可见AEC-Ⅱ特征性结构--嗜锇性板层小体（Lb）.②B组培养AEC-Ⅱ至29 h,发现贴壁细胞数量减少,胞内可见反光增强的空泡,上清液中可见较多的细胞碎片.③C组与A组比较细胞计数、OD值差异无统计学意义（P＞0.05）,细胞形态无明显改变;与B组比较细胞计数、OD值差异有统计学意义（P＜0.05）.结论 0.01～50 μmol/L HO-1对原代培养AEC-Ⅱ的生长没有直接的细胞毒性,可以在体外AEC-Ⅱ培养中使用.
Objective To study the direct effects of heme oxygenase - 1 （HO - 1 ） on the growth of rat primary cultured alveolar epithelial thetype Ⅱ cells （ AEC - Ⅱ）. Methods The AEC -Ⅱ were isolated from SD rats by immune adherence separation and purification. Cell viability was assessed trypan blue, cell purity by alkaline phosphatase, morphology and qualitative characteristic structure of cell were viewed with inverted microscope and electron microscope. The cells were randomly divided into seven groups： A（The normal control group）, B（ H202 group）, C1 - 5 （0.01～50μmol/L HO - 1 ）. The cells were vaccinated in 24 hole, 96 holes in cell culture plate, after being 24 h cultured, AEC - Ⅱ were treatment with HO - 1 （0. 01 -50 μmol/L） for 5 h, as evidenced by cells count and optical density （OD）. Results By purification immune adhesion method in the yields of primary cultured AEC -Ⅱ were （2 -2.5 ） × 10^7/rat, the viability and purity were 〉 90% , and electron microscopy showed the characteristic structure - osmiophilic lamellar body of the AEC -Ⅱ. AEC - Ⅱ were treated with H2 02 （0.5 mmol/L） for 3 h, cell viability decreased, reflecting of intracellular vacuoles were enhanced, a large number of cells off the wall, cell debris appeared in supernatant. Cells count and OD of treatment with HO - 1 （0.01 - 50 μmol/L） groups had no significant differences than those of the normal controls group （P 〉 0. 05 ）, and was higher than H2 02 group （ P 〈 0. 05 ）. Conclusion 0.01 - 50 μmol/L of HO- 1 in the primary cultured AEC - Ⅱ have no direct cytotoxicity, and HO -1 can be used in cultured AEC -Ⅱ in vitro.
出处
《中国急救医学》
CAS
CSCD
北大核心
2010年第12期1099-1102,I0010,共5页
Chinese Journal of Critical Care Medicine
