Yokota S, Fahimi H D
Histochem J. 1978 Jul;10(4):469-87. doi: 10.1007/BF01003010.
The effects of detergents, organic lipid solvents, and several adjuvants used in cell fractionation on the ultrastructure of the peroxisomal (microbody) membrane and its permeability to catalase have been investigated. Chopper sections of glutaraldehyde-fixed liver were incubated in the presence of various agents, followed by cytochemical staining for catalase and processed for electron microscopy. Catalase activity was also determined biochemically in the incubation medium. Marked catalase diffusion was found after treatment with 1% or 0.5% Triton X-100 or deoxycholate, as well as with 50% ethanol or acetone or 20% propanol or t-butanol. In contrast, 1% digitonin and lower concentrations of the above agents, as well as sucrose or glycerine caused selective diffusion of catalase from a limited population of peroxisomes. Treatment with 10% polyvinylpyrrolidone (PVP), which has been used as a protective agent in the isolation of microbodies, did not produce any alteration in the fine structure and cytochemical appearance of peroxisomes. These findings concur with earlier biochemical studies on freshly isolated peroxisomes and demonstrate the susceptibility of microbodies, even in glutaraldehyde-fixed rat liver to the effects of various agents which affect the microbody membrane. A close correlation between the ultrastructural integrity of the microbody membrane and its permeability to catalase has been found. The significance of these observations for the assessment of the permeability characteristics of the microbody membrane is discussed.
研究了去污剂、有机脂质溶剂以及细胞分级分离中使用的几种佐剂对过氧化物酶体(微体)膜超微结构及其对过氧化氢酶通透性的影响。将戊二醛固定的肝脏切碎切片在各种试剂存在下孵育,随后进行过氧化氢酶的细胞化学染色并制备用于电子显微镜观察的样本。还对孵育培养基中的过氧化氢酶活性进行了生化测定。在用1%或0.5%的 Triton X-100或脱氧胆酸盐以及50%乙醇或丙酮或20%丙醇或叔丁醇处理后,发现过氧化氢酶有明显扩散。相比之下,1%洋地黄皂苷以及上述试剂的较低浓度,以及蔗糖或甘油会导致过氧化氢酶从有限数量的过氧化物酶体中选择性扩散。用10%聚乙烯吡咯烷酮(PVP)处理,PVP在微体分离中用作保护剂,对过氧化物酶体的精细结构和细胞化学外观没有产生任何改变。这些发现与早期对新鲜分离的过氧化物酶体的生化研究一致,并表明即使在戊二醛固定的大鼠肝脏中,微体也容易受到影响微体膜的各种试剂作用。已发现微体膜的超微结构完整性与其对过氧化氢酶的通透性之间存在密切相关性。讨论了这些观察结果对评估微体膜通透性特征的意义。