Dionne F T, Chevalier S, Bleau G, Roberts K D, Chapdelaine A
Mol Cell Endocrinol. 1983 Nov;33(1):113-26. doi: 10.1016/0303-7207(83)90060-6.
The increase in acid phosphatase (AP) activity in cultured canine prostatic epithelial cells was investigated as a biochemical marker of in vitro cellular differentiation. The enzyme was studied in secretory and non-secretory epithelial cell populations obtained from control and cycloheximide-treated cultures over a period of 3 weeks and compared to the AP present in tissue and cellular extracts from normal canine prostates. The progressive increase in AP activity with the duration of culture was strongly inhibited by cycloheximide in both cell populations. The degree of inhibition was more pronounced late in the culture when AP activity increased at a faster rate in secretory cells. Cycloheximide inhibited protein biosynthesis by 70-80% as evidenced by a reduction in the incorporation of amino acids into acid-insoluble material. However, the specific activities of AP in the cellular extracts were similar in control and cycloheximide-treated cultures and increased sharply by 3-4-fold in the secretory cells after 12 days of culture. When extracts derived from control and cycloheximide-treated cells of various duration were submitted to electrophoresis in polyacrylamide gels (PAGE), a unique pattern of three bands of AP activity with Rf values of 0.18, 0.27 and 0.38 was obtained. In controls the AP activity in the band with an Rf of 0.18 increased preferentially during the culture period and was more important quantitatively in secretory cells. In cycloheximide-treated cultures the increase of AP activity associated with the band with an Rf of 0.18 was more strongly inhibited. The addition of tartrate to the staining mixture inhibited all three bands of AP activity. Similar results were obtained when extracts derived from freshly dispersed cells as well as from normal canine prostatic tissue were submitted to PAGE; the AP activity was resolved into 3 bands with Rf values of 0.15-0.18, 0.23-0.27 and 0.33-0.38; all three bands were inhibited by the addition of tartrate and the first band was predominant. Thus, the increase in AP activity in prostatic epithelial cells in a culture medium supplemented with serum and deprived of sex steroids is due to the de novo synthesis of a major form of the enzyme by the secretory cells.
研究了培养的犬前列腺上皮细胞中酸性磷酸酶（AP）活性的增加，将其作为体外细胞分化的生化标志物。在3周的时间里，对从对照培养物和环己酰亚胺处理的培养物中获得的分泌性和非分泌性上皮细胞群体中的该酶进行了研究，并与正常犬前列腺组织和细胞提取物中的AP进行了比较。在两个细胞群体中，环己酰亚胺均强烈抑制了AP活性随培养时间的逐渐增加。当分泌细胞中AP活性以更快的速率增加时，培养后期的抑制程度更为明显。环己酰亚胺抑制蛋白质生物合成达70 - 80%，这可通过氨基酸掺入酸不溶性物质的减少来证明。然而，对照培养物和环己酰亚胺处理的培养物中细胞提取物中AP的比活性相似，培养12天后分泌细胞中的AP比活性急剧增加3 - 4倍。当将来自不同培养时间的对照细胞和环己酰亚胺处理细胞的提取物在聚丙烯酰胺凝胶（PAGE）中进行电泳时，获得了一种独特的AP活性三条带模式，其Rf值分别为0.18、0.27和0.38。在对照中，Rf为0.18的条带中的AP活性在培养期间优先增加，并且在分泌细胞中的数量上更重要。在环己酰亚胺处理的培养物中，与Rf为0.18的条带相关的AP活性增加受到更强的抑制。向染色混合物中加入酒石酸盐可抑制所有三条AP活性带。当将来自新鲜分散细胞以及正常犬前列腺组织的提取物进行PAGE时，也获得了类似的结果；AP活性被解析为三条带，其Rf值分别为0.15 - 0.18、0.23 - 0.27和0.33 - 0.38；加入酒石酸盐可抑制所有三条带，且第一条带占主导。因此，在补充血清并去除性类固醇的培养基中，前列腺上皮细胞中AP活性的增加是由于分泌细胞重新合成了该酶的一种主要形式。
