Nagasawa K, Mak T W
Proc Natl Acad Sci U S A. 1980 May;77(5):2964-8. doi: 10.1073/pnas.77.5.2964.
At nanomolar concentrations, phorbol ester, a class of potent tumor promoters, can promote differentiation in the human malignant T-lymphoblastic cell line MOLT-3. The optimal dose for induction, as measured by the increase of the number of cells containing sheep erythrocyte receptors (E-rosette assay), is between 8 and 16 nM 12-O-tetradecanoylphorbol 13-acetate (TPA), although there were significant increases of E-rosette-positive (E(+)) cells at concentrations as low as 1.6 nM TPA. The induction was linear for 4 days, then it reached a plateau. This induction was independent of the cell densities of the cultures, and the viability of the E(+) cells remained high (95-100%) even after 10 days of culture in the presence of the tumor promoters. The E(+) cells, when measured with the more stable 2-aminoethylisothiouronium bromide E-rosette assay, indicated that virtually all (75-95%) of the MOLT-3 cells became E(+) by 4 days in culture. This induction by TPA was also accompanied by a dramatic drop in the plating efficiencies and a reduction in DNA synthesis. Examination of phorbol and other phrobol esters indicated that the ability to induce these cells correlated well with the tumor-promoting activities of these compounds, because only TPA and to a lesser extent phorbol 12,13,-dibenzoate induced E(+) cells, while phorbol and 4alpha-phorbol 12,13-didecanoate had no effect. Studies of MOLT-3 cells depleted of E(+) cells indicated that the induction of E(+) cells cannot be explained solely on the basis of enrichment or stimulation of the background E(+) cells in MOLT-3 cultures. Finally, we have shown that TPA also affected another differentiation marker, the loss of the enzyme terminal deoxyribonucleotidyl transferase. Terminal transferase activities and percentages of terminal-transferase-positive cells in these cultures were reduced to as low as 1/10th in 4 days in the presence of 16 nM TPA.
在纳摩尔浓度下，佛波酯（一类强效肿瘤促进剂）可促进人恶性T淋巴细胞系MOLT-3的分化。通过含有绵羊红细胞受体的细胞数量增加（E花环试验）来衡量，诱导的最佳剂量为8至16 nM的12-O-十四酰佛波醇-13-乙酸酯（TPA），尽管在低至1.6 nM TPA的浓度下E花环阳性（E(+)）细胞也有显著增加。诱导作用在4天内呈线性，然后达到平台期。这种诱导与培养物的细胞密度无关，即使在肿瘤促进剂存在的情况下培养10天后，E(+)细胞的活力仍保持在较高水平（95 - 100%）。用更稳定的2-氨基乙基异硫脲溴化物E花环试验检测E(+)细胞时发现，实际上所有（75 - 95%）的MOLT-3细胞在培养4天时都变成了E(+)细胞。TPA的这种诱导还伴随着接种效率的急剧下降和DNA合成的减少。对佛波醇和其他佛波酯的研究表明，诱导这些细胞的能力与这些化合物的肿瘤促进活性密切相关，因为只有TPA以及程度较轻的佛波醇12,13 - 二苯甲酸酯能诱导E(+)细胞，而佛波醇和4α - 佛波醇12,13 - 二癸酸酯则没有作用。对耗尽E(+)细胞的MOLT-3细胞的研究表明，E(+)细胞的诱导不能仅仅基于MOLT-3培养物中背景E(+)细胞的富集或刺激来解释。最后，我们已经表明TPA还影响另一个分化标志物，即末端脱氧核苷酸转移酶的丧失。在16 nM TPA存在的情况下，这些培养物中的末端转移酶活性和末端转移酶阳性细胞的百分比在4天内降低到了低至十分之一。
