Metabolic acidosis stimulates intestinal arginine absorption

Abstract

Introduction: L-Arginine is the exclusive precursor for nitric oxide (NO) biosynthesis and an essential nutrient for intestinal integrity during stress states. However, the effect of extracellular pH on intestinal arginine uptake is poorly understood. The purpose of this in vitro study was to investigate the regulation of intestinal arginine transport during acidosis. Methods:3H-L-arginine transport activity was measured in intestinal epithelial Caco-2 cells at various ambient pHs (pH 6.6–7.6) ± actinomycin-D (1 μM), cycloheximide (20 μM), phosphatidylinositide-3 kinase (PI-3) inhibitor wortmannin (1 uM), and the NO synthetase inhibitor N-ω-Nitro-L-Arginine (L-NA, 100 μM). Data were analyzed by ANOVA. Results: Decreasing the extracellular pH stimulated arginine transport activity in a time and pH-dependent fashion. Chronic acidosis (pH 6.6 for 48 hours) resulted in a 4-fold increase in arginine transport activity (0.94 ± 0.10 acidosis vs. 0.23 ± 0.05 control, nmole/mg/min, p < 0.01) and a 3-fold increase of arginine transporter mCAT-1 mRNA levels (p < 0.01). This acidosis-induced increase in arginine transport activity was due to a stimulation of transporter maximal transport capacity (Vmax 1.53 ± 0.08 acidosis vs. 0.56 ± 0.05 control, nmole/mg /min, p < 0.01) rather than a change in transporter affinity (Km = 54 ± 10 acidosis vs. 43 ± 6 control, μM arginine, p = NS). This acidosis-stimulated arginine transport activity was individually blocked by actinomycin-D, cycloheximide, and L-NA. The acidosis-induced arginine transport activity was further stimulated by wortmannin (1.4 ± 0.25 acidosis + wortmannin vs. 0.94 ± 0.10 acidosis vs. 0.21 ± 0.03 wortmannin vs. 0.23 ± 0.05 control, nmole/mg /min). Conclusion: Acidosis stimulates arginine transport in Caco-2 cells via a mechanism that leads to transcription of the gene and translation of arginine transporter. Phosphatidylinositide-3 kinase is a key intracellular regulator involved in this signal transduction cascade. The nitric oxide synthetase activity plays a pivotal role in this stimulation. An increased availability of arginine to cells during acidosis provides substrate for NO biosynthesis and may help in maintaining cellular integrity.