Plopper C G, Weir A J, Morin D, Chang A, Philpot R M, Buckpitt A R
Department of Veterinary Anatomy and Cell Biology, School of Veterinary Medicine, University of California, Davis 95616.
Mol Pharmacol. 1993 Jul;44(1):51-61.
Previous studies have indicated that both cytodifferentiation of Clara cells and the onset of pulmonary cytochrome P450 activity are postnatal events. However, the relationship between these two events during lung development remains poorly understood. To determine how these events interrelate, we examined rabbit Clara cells during postnatal differentiation, with the following goals in mind: 1) to identify the patterns of intracellular expression of cytochrome P450 monooxygenase isozymes 2B and 4B and cytochrome P450 reductase, 2) to describe the biogenesis of the organelles with which these isozymes are associated, namely smooth and rough endoplasmic reticulum, and 3) to compare the patterns of expression with cytochrome P450 activity in the whole lung over the same period. Lungs of rabbits ranging in age from 24 days gestational age (DGA) to 25 weeks postnatally were studied. Ultrastructural morphometry showed that smooth endoplasmic reticulum averaged < 5% of the Clara cell volume in late gestational (24-30 DGA) and neonatal rabbits [0-7 days postnatally (DPN)], grew to 20-30% of the cell volume in 14-21-DPN animals, and approximated adult levels (> 40%) in 28-DPN rabbits. In contrast, rough endoplasmic reticulum decreased from > 10% of the cell volume at 27 DGA to < 5% in adults. All postnatal animals showed considerable heterogeneity in the abundance of smooth endoplasmic reticulum among individual cells. Immunohistochemistry revealed that cytochrome P450 reductase appeared in Clara cells earlier (28 DGA) than did either isozyme 2B or 4B (1 DPN). Each antigen was detected first in the apical borders of the cells, then throughout the cytoplasm in a few cells by 7 DPN, and finally in adult abundance by 28 DPN. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blotting showed that cytochrome P450 protein concentrations increased postnatally. Cytochrome P450 heme protein was not detected spectrophotometrically in the lungs of animals younger than 3 DPN but increased to approximately 70% of adult levels by 28 DPN. Likewise, cytochrome P450 activity (measured as ethoxy- and pentoxyresorufin O-dealkylation) was not detected in animals younger than 2 DPN but increased to approximately 75% of adult levels by 28 DPN.(ABSTRACT TRUNCATED AT 400 WORDS)
先前的研究表明，克拉拉细胞的细胞分化和肺细胞色素P450活性的出现均为出生后的事件。然而，在肺发育过程中这两个事件之间的关系仍知之甚少。为了确定这些事件是如何相互关联的，我们在出生后分化过程中检查了兔克拉拉细胞，心中有以下目标：1）确定细胞色素P450单加氧酶同工酶2B和4B以及细胞色素P450还原酶的细胞内表达模式，2）描述与这些同工酶相关的细胞器的生物发生，即光滑和粗糙内质网，3）比较同期整个肺中细胞色素P450活性的表达模式。研究了胎龄从24天到出生后25周的兔肺。超微结构形态测量显示，在妊娠晚期（24 - 30天胎龄）和新生兔[出生后0 - 7天（DPN）]中，光滑内质网平均占克拉拉细胞体积的<5%，在14 - 21天DPN的动物中增长到细胞体积的20 - 30%，在28天DPN的兔中接近成年水平（>40%）。相反，粗糙内质网从27天胎龄时细胞体积的>10%降至成年时的<5%。所有出生后的动物在单个细胞中光滑内质网的丰度方面表现出相当大的异质性。免疫组织化学显示，细胞色素P450还原酶在克拉拉细胞中出现的时间（28天胎龄）早于同工酶2B或4B（出生后1天）。每种抗原首先在细胞的顶端边界被检测到，然后到7天DPN时在一些细胞的整个细胞质中被检测到，最后到28天DPN时达到成年丰度。十二烷基硫酸钠 - 聚丙烯酰胺凝胶电泳和蛋白质印迹显示，细胞色素P450蛋白浓度在出生后增加。在3天DPN以下的动物肺中，用分光光度法未检测到细胞色素P450血红素蛋白，但到28天DPN时增加到成年水平的约70%。同样，在2天DPN以下的动物中未检测到细胞色素P450活性（以乙氧基和戊氧基试卤灵O - 脱烷基化测量），但到28天DPN时增加到成年水平的约75%。（摘要截短为400字）
