Arginase II inhibition prevents interleukin-8 production through regulation of p38 MAPK phosphorylation activated by loss of mitochondrial membrane potential in nLDL-stimulated hAoSMCs

Arginase inhibition exhibits beneficial effects in vascular endothelial and smooth muscle cells. In human aortic smooth muscle cells (hAoSMCs), native low-density lipoprotein (nLDL) induced the production of interleukin-8 (IL-8) that is involved in the pathogenesis of cardiovascular diseases. Therefore, we examined the effect of arginase inhibition on IL-8 production and the underlying mechanism. In hAoSMCs, reverse transcription-PCR, western blotting and immunocytochemistry with MitoTracker confirmed that arginase II was confined predominantly to mitochondria. The mitochondrial membrane potential (MMP) was assessed using tetramethylrhodamine ethyl ester. The MMP decreased upon nLDL stimulation but was restored upon arginase inhibition. MMP loss caused by nLDL was prevented by treatment with the intracellular Ca²⁺ chelator BAPTA-AM. In mitochondrial Ca²⁺ measurements using Rhod-2 AM, increased mitochondrial Ca²⁺ levels by nLDL were inhibited upon preincubation with an arginase inhibitor. Among the polyamines, spermine, an arginase activity-dependent product, caused mitochondrial Ca²⁺ movement. The nLDL-induced MMP change resulted in p38 mitogen-activated protein kinase (MAPK) phosphorylation and IL-8 production and was prevented by the arginase inhibitors BAPTA and ruthenium 360. In isolated AoSMCs from ApoE^-/- mice fed a high-cholesterol diet, arginase activity, p38 MAPK phosphorylation, spermine and mitochondrial Ca²⁺ levels and keratinocyte-derived chemokine (KC) production were increased compared with wild-type (WT) mice. However, in AoSMCs isolated from arginase II-null mice, increases in MMP and decreases in mitochondrial Ca²⁺ levels were noted compared with WT and were associated with p38 MAPK activation and IL-8 production. These data suggest that arginase activity regulates the change in MMP through Ca²⁺ uptake that is essential for p38 MAPK phosphorylation and IL-8 production.