Rhoads J M, Chen W, Chu P, Berschneider H M, Argenzio R A, Paradiso A M
Department of Pediatrics, North Carolina State University School of Veterinary Medicine, Chapel Hill 27599.
Am J Physiol. 1994 May;266(5 Pt 1):G828-38. doi: 10.1152/ajpgi.1994.266.5.G828.
L-Glutamine (Gln) is a major respiratory fuel and substrate for nucleic acid synthesis in mammalian intestinal cells. The structurally related amino acid, L-asparagine (Asn), stimulates the proliferative enzyme ornithine decarboxylase in colonocytes, an effect that is blocked by the Na+-H+ exchange inhibitor amiloride. In an epithelial cell line derived from newborn piglet jejunum (IPEC-J2 cells), we determined intracellular pH (pHi) by computer-assisted microfluorimetry in single cells loaded with pH-sensitive dye 2',7'-bis(2-carboxyethyl)5-(6)- carboxyfluorescein. Resting pHi in N-2-hydroxyethylpiperazine-N'-2- ethanesulfonic acid-buffered NaCl Ringer was 7.06 +/- 0.02. Removal of external Na+ caused reversible acidification; recovery of pHi from NH+4-induced acid load was Na+ dependent, amiloride inhibitable, and Cl-independent. Asn and Gln had no measurable effect on resting pHi, but pretreatment with Asn or Gln induced a consistent twofold increase in pHi recovery from an acid challenge that was not seen with L-proline, D-glutamine, or L-phenylalanine. Inhibition of Gln metabolism by aminooxyacetate abolished the stimulatory effect of Gln on the exchanger. The tumor promotor phorbol 12-myristate 13-acetate (PMA) stimulated recovery rate from acid load and also increased resting pHi. The effects of PMA and Gln on Na+-H+ exchange from acid load were additive. Stimulation of Na+-H+ exchange by PMA, but not by Gln, was inhibited by protein kinase C (PKC) inhibitor 1-(5-isoquinolinylsulfonyl)-2-methylpeperazine. We conclude that Gln metabolism stimulates Na+-H+ exchange of acid-loaded porcine enterocytes by a mechanism not requiring activation of PKC.
L-谷氨酰胺（Gln）是哺乳动物肠道细胞中核酸合成的主要呼吸燃料和底物。结构相关的氨基酸L-天冬酰胺（Asn）可刺激结肠细胞中的增殖酶鸟氨酸脱羧酶，该作用可被Na⁺-H⁺交换抑制剂阿米洛利阻断。在源自新生仔猪空肠的上皮细胞系（IPEC-J2细胞）中，我们通过计算机辅助微量荧光测定法，对加载了pH敏感染料2'，7'-双（2-羧乙基）5-（6）-羧基荧光素的单细胞进行细胞内pH（pHi）测定。在N-2-羟乙基哌嗪-N'-2-乙磺酸缓冲的NaCl林格液中，静息pHi为7.06±0.02。去除细胞外Na⁺会导致可逆性酸化；从NH₄⁺诱导的酸负荷中恢复pHi依赖于Na⁺，可被阿米洛利抑制，且与Cl⁻无关。Asn和Gln对静息pHi无显著影响，但用Asn或Gln预处理可使酸刺激后pHi的恢复一致地增加两倍，而L-脯氨酸、D-谷氨酰胺或L-苯丙氨酸则无此现象。氨基氧乙酸抑制Gln代谢可消除Gln对交换体的刺激作用。肿瘤促进剂佛波酯12-肉豆蔻酸酯13-乙酸酯（PMA）可刺激酸负荷后的恢复率，还可提高静息pHi。PMA和Gln对酸负荷后Na⁺-H⁺交换的作用具有相加性。PMA而非Gln对Na⁺-H⁺交换的刺激作用可被蛋白激酶C（PKC）抑制剂1-（5-异喹啉磺酰基）-2-甲基哌嗪抑制。我们得出结论，Gln代谢通过一种不需要激活PKC的机制刺激酸负荷的猪肠细胞的Na⁺-H⁺交换。
