Tamura Tsunenobu, Baggott Joseph E, Johnston Kelley E, Li Qing-Jun, Antony Asok C
Department of Nutrition Sciences, University of Alabama at Birmingham, Birmingham, AL 35294, USA.
Department of Medicine, Indiana University School of Medicine, Indianapolis, IN 46202, USA.
Microbiology (Reading). 1997 Aug;143 ( Pt 8):2639-2646. doi: 10.1099/00221287-143-8-2639.
Several mechanisms have been described to explain the resistance of cells to methotrexate (MTX); however, the basis for the heterogeneity of mechanisms has been obscure. It was hypothesized that the type of MTX resistance in a single species can be influenced by the form of extracellular folate supplied during the development of resistance. Two strains of MTX-resistant Enterococcus hirae [corrected] were developed by transferring the bacteria to media containing increasing concentrations of MTX in the presence of constant concentrations of either 5-formyl-5,6,7,8-tetrahydropteroylglutamic acid (5-HCO-H4PteGlu) or pteroylglutamic acid (PteGlu). These resistant strains were designated E. hirae/MTX/5-HCO-H4PteGlu and E. hirae/MTX/PteGlu, respectively [corrected]. The mechanisms of MTX resistance included: (1) increased folic acid reductase (FAR) activity in both resistant strains but increased dihydrofolate reductase (DHFR) activity only in E. hirae/MTX/PteGlu [corrected]; (2) decreased synthesis and intracellular retention of MTX containing two glutamyl residues; (3) decreased uptake of MTX accompanied by decreased uptake of folates; and (4) reduction of folate-binding capacity. Among these, the form of folate present in the media during the development of resistance affected DHFR and FAR activities and the transport of folates. These findings, together with data from other laboratories, suggest that it may be important to use a reduced form of folate, a more physiological form than oxidized PteGlu, in the media during the development of resistance for the study of the mechanisms of MTX resistance in cultured cells.
已经描述了几种机制来解释细胞对甲氨蝶呤（MTX）的耐药性；然而，机制异质性的基础一直不清楚。据推测，单一物种中甲氨蝶呤耐药性的类型可能受到耐药性产生过程中所提供的细胞外叶酸形式的影响。通过将细菌转移到含有不断增加浓度MTX且分别存在恒定浓度的5-甲酰基-5,6,7,8-四氢蝶酰谷氨酸（5-HCO-H4PteGlu）或蝶酰谷氨酸（PteGlu）的培养基中，培育出了两株耐MTX的平氏肠球菌[已修正]。这些耐药菌株分别被命名为平氏肠球菌/MTX/5-HCO-H4PteGlu和平氏肠球菌/MTX/PteGlu[已修正]。MTX耐药机制包括：（1）两株耐药菌株中的叶酸还原酶（FAR）活性均增加，但仅在平氏肠球菌/MTX/PteGlu中[已修正]二氢叶酸还原酶（DHFR）活性增加；（2）含两个谷氨酰残基的MTX的合成及细胞内潴留减少；（3）MTX摄取减少，同时叶酸摄取也减少；（4）叶酸结合能力降低。其中，耐药性产生过程中培养基中存在的叶酸形式影响DHFR和FAR活性以及叶酸的转运。这些发现，连同其他实验室的数据表明，在耐药性产生过程中，在培养基中使用还原型叶酸（一种比氧化型PteGlu更具生理性的形式）对于研究培养细胞中甲氨蝶呤耐药机制可能很重要。
