Keating Elisa, Gonçalves Pedro, Lemos Clara, Costa Fernanda, Campos Isabel, Smith Sylvia B, Bridges Christy C, Martel Fátima
Department of Biochemistry (U38-FCT), Faculty of Medicine of Porto, University of Porto, Porto 4200-319, Portugal.
J Membr Biol. 2007 Apr;216(2-3):143-52. doi: 10.1007/s00232-007-9057-5. Epub 2007 Aug 9.
The aim of this work was to test the putative involvement of members of the ABC superfamily of transporters on folic acid (FA) cellular homeostasis in the human placenta. [(3)H]FA uptake and efflux in BeWo cells were unaffected or hardly affected by multidrug resistance 1 (MDR1) inhibition (with verapamil), multidrug resistance protein (MRP) inhibition (with probenecid) or breast cancer resistance protein (BCRP) inhibition (with fumitremorgin C). However, [(3)H]FA uptake and efflux were inhibited by progesterone (200 microM). An inhibitory effect of progesterone upon [(3)H]FA uptake and efflux was also observed in human cytotrophoblasts. Moreover, verapamil and ss-estradiol also reduced [(3)H]FA efflux in these cells. Inhibition of [(3)H]FA uptake in BeWo cells by progesterone seemed to be very specific since other tested steroids (beta-estradiol, corticosterone, testosterone, aldosterone, estrone and pregnanediol) were devoid of effect. However, efflux was also inhibited by beta-estradiol and corticosterone and stimulated by estrone. Moreover, the effect of progesterone upon the uptake of [(3)H]FA by BeWo cells was concentration-dependent (IC(50 )= 65 [range 9-448] microM) and seems to involve competitive inhibition. Also, progesterone (1-400 microM) did not affect either [(3)H]FA uptake or efflux at an external acidic pH. Finally, inhibition of [(3)H]FA uptake by progesterone was unaffected by either 4-acetamido-4'-isothiocyanato-2,2'-stilbenedisulfonic acid (SITS), a known inhibitor of the reduced folate carrier (RFC), or an anti-RFC antibody. These results suggest that progesterone inhibits RFC. In conclusion, our results show that progesterone, a sterol produced by the placenta, inhibits both FA uptake and efflux in BeWo cells and primary cultured human trophoblasts.
这项工作的目的是测试转运蛋白ABC超家族成员对人胎盘中叶酸(FA)细胞内稳态的假定影响。维拉帕米抑制多药耐药蛋白1(MDR1)、丙磺舒抑制多药耐药相关蛋白(MRP)或烟曲霉毒素C抑制乳腺癌耐药蛋白(BCRP),对BeWo细胞中[³H]FA的摄取和流出没有影响或几乎没有影响。然而,孕酮(200 μM)抑制了[³H]FA的摄取和流出。在人细胞滋养层细胞中也观察到孕酮对[³H]FA摄取和流出的抑制作用。此外,维拉帕米和β-雌二醇也降低了这些细胞中[³H]FA的流出。孕酮对BeWo细胞中[³H]FA摄取的抑制作用似乎非常特异,因为其他测试的类固醇(β-雌二醇、皮质酮、睾酮、醛固酮、雌酮和孕二醇)没有作用。然而,β-雌二醇和皮质酮也抑制了流出,雌酮则刺激了流出。此外,孕酮对BeWo细胞摄取[³H]FA的作用具有浓度依赖性(IC₅₀ = 65 [范围9 - 448] μM),似乎涉及竞争性抑制。同样,在外部酸性pH值下,孕酮(1 - 400 μM)对[³H]FA的摄取或流出均无影响。最后,孕酮对[³H]FA摄取的抑制作用不受还原型叶酸载体(RFC)的已知抑制剂4-乙酰氨基-4'-异硫氰酸基-2,2'-二苯乙烯二磺酸(SITS)或抗RFC抗体的影响。这些结果表明孕酮抑制RFC。总之,我们的结果表明,胎盘产生的一种固醇孕酮可抑制BeWo细胞和原代培养的人滋养层细胞中FA的摄取和流出。