Abstract HL60 myeloid leukemia cells are extensively used as a differentiation model. We investigated a variant of HL60 which is resistant to differentiation induction (HL60-R) by standard differentiation inducers such as retinoic acid and dimethylsulfoxide (DMSO). To find an explanation for this resistance, we examined nucleotide (NTP) and deoxynucleotide (dNTP) pools in HL60-R and its parent cell line, sensitive to differentiation, HL60-S. We also explored whether these differences led to a difference in sensitivity to various antimetabolites. Drug sensitivity was measured with the tetrazolium (MTT) assay, while nucleotides were measured with anion-exchange HPLC. HL60-R cells were between 2- and 5-fold resistant to the antimetabolites 5-fluorouracil, Brequinar, hydroxyurea and N-(phosphonacetyl)-L-aspartate (PALA), but more sensitive to aza-2’-deoxycytidine (DAC), cytarabine and thymidine (5- to 10-fold). The NTP pools in both HL60 variants showed a normal pattern with ATP being the highest (2530–2876 pmol/106 cells) and CTP being lowest. However, UTP pools were 2-fold higher in the HL60-S cells (p < .01), while CTP and GTP pools were 30% higher (p < .01) compared to HL60-R cells. For the dNTP pools, larger differences were observed, with dATP (50 pmol/106 cells) being highest in HL60-R cells, but dATP was 4-fold lower in HL60-S cells. In HL-60-R, the triple combination retinoic acid, DMSO and DAC increased all NTPs almost 2-fold in contrast to HL60-S. Uridine increased UTP (1.4-fold), CTP (2-fold) and dCTP (1.4.-fold) pools in both cell lines, but thymidine increased only dTTP pools (4- to 7-fold), with a depletion of dCTP. PALA decreased UTP and CTP in both cell lines, but increased ATP (only in HL60-R). Hydroxyurea decreased dNTP especially in HL60-S cells. In conclusion, the pronounced differences in NTP and dNTP pools between HL60-S and HL60-R possibly play a role in the induction of differentiation and drug sensitivity.

中文翻译：

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分化抗性影响核糖和脱氧核糖核苷酸库以及对 HL60 细胞中嘧啶代谢拮抗剂的敏感性

摘要 HL60 髓系白血病细胞被广泛用作分化模型。我们研究了 HL60 的一种变体，它对标准分化诱导剂如视黄酸和二甲基亚砜 (DMSO) 的分化诱导 (HL60-R) 具有抗性。为了找到这种抗性的解释，我们检查了 HL60-R 及其对分化敏感的亲本细胞系 HL60-S 中的核苷酸 (NTP) 和脱氧核苷酸 (dNTP) 库。我们还探讨了这些差异是否导致对各种抗代谢物的敏感性差异。使用四唑 (MTT) 测定法测量药物敏感性，而使用阴离子交换 HPLC 测量核苷酸。HL60-R 细胞对抗代谢物 5-氟尿嘧啶、Brequinar、羟基脲和 N-(膦乙酰基)-L-天冬氨酸 (PALA) 的耐药性介于 2 到 5 倍之间，但对 aza-2'-脱氧胞苷 (DAC)、阿糖胞苷和胸苷更敏感（5 至 10 倍）。两种 HL60 变体中的 NTP 池均显示正常模式，ATP 最高（2530-2876 pmol/106 个细胞），CTP 最低。然而，与 HL60-R 细胞相比，HL60-S 细胞中的 UTP 池高 2 倍（p < .01），而 CTP 和 GTP 池高 30%（p < .01）。对于 dNTP 库，观察到更大的差异，HL60-R 细胞中的 dATP（50 pmol/106 个细胞）最高，但 HL60-S 细胞中的 dATP 低 4 倍。在 HL-60-R 中，与 HL60-S 相比，视黄酸、DMSO 和 DAC 的三重组合使所有 NTP 增加了近 2 倍。尿苷增加了两种细胞系中的 UTP（1.4 倍）、CTP（2 倍）和 dCTP（1.4 倍）池，但胸苷仅增加了 dTTP 池（4 到 7 倍），dCTP 耗尽。PALA 降低了两种细胞系中的 UTP 和 CTP，但增加了 ATP（仅在 HL60-R 中）。羟基脲降低 dNTP，尤其是在 HL60-S 细胞中。总之，HL60-S 和 HL60-R 之间 NTP 和 dNTP 库的显着差异可能在诱导分化和药物敏感性中起作用。