Mount Peter F, Fraser Scott A, Watanabe Yasuo, Lane Natalie, Katsis Frosa, Chen Zhi-Ping, Kemp Bruce E, Power David A
The Austin Research Institute, Austin Hospital, Heidelberg, Australia.
Nephron Physiol. 2006;102(2):p36-50. doi: 10.1159/000089092. Epub 2005 Oct 19.
Renal nitric oxide (NO) synthesis increases with increasing salt intake, however, the mechanisms underlying this are poorly understood. We hypothesized that activating or inhibitory phosphorylation of neuronal and endothelial nitric oxide synthase (nNOS, eNOS) regulates renal NO production in response to altered dietary salt.
Sprague-Dawley rats were fed low, normal or high salt diets for 12 h or 2 weeks, and kidney NOS phosphorylation was analyzed by Western blot using phosphopeptide antibodies against the sites nNOS-Ser(1412), nNOS-Ser(847), eNOS-Ser(1176) and eNOS-Thr(494).
At 12 h, total nNOS increased 1.4-fold (p < 0.01) in the high salt group and decreased by 26% (p < 0.05) in the low salt group. Changes in expression of phospho-nNOS at 12 h were accounted for by the changes in total nNOS. No change in total or phospho-eNOS was seen at 12 h. At 2 weeks, in the low salt group expression of total nNOS increased 1.8-fold (p < 0.05) whereas expression of nNOS phosphorylated at the inhibitory site Ser(847) increased 4.3-fold (p < 0.01). Total eNOS was increased 3-fold in the low salt group (p < 0.01), with parallel increases in eNOS phosphorylated at both activating and inhibitory sites (p < 0.05). In the 2-week high salt group no changes in NOS expression or phosphorylation were seen, despite the observed increased excretion of urinary NO metabolites.
In summary, changes in phospho-nNOS and phospho-eNOS expression occurred in parallel with changes in total expression, thus, the overall activating and inhibitory effects of nNOS and eNOS phosphorylation at the sites studied were not changed by altered dietary salt.
随着盐摄入量的增加，肾脏一氧化氮（NO）的合成也会增加，然而，其潜在机制尚不清楚。我们推测，神经元型和内皮型一氧化氮合酶（nNOS、eNOS）的激活或抑制性磷酸化可调节肾脏NO的产生，以应对饮食中盐的变化。
将Sprague-Dawley大鼠分别喂食低、正常或高盐饮食12小时或2周，使用针对nNOS-Ser(1412)、nNOS-Ser(847)、eNOS-Ser(1176)和eNOS-Thr(494)位点的磷酸化肽抗体，通过蛋白质印迹法分析肾脏中一氧化氮合酶的磷酸化情况。
12小时时，高盐组总nNOS增加1.4倍（p<0.01），低盐组降低26%（p<0.05）。12小时时磷酸化nNOS表达的变化是由总nNOS的变化引起的。12小时时总eNOS或磷酸化eNOS未见变化。2周时，低盐组总nNOS表达增加1.8倍（p<0.05），而在抑制位点Ser(847)磷酸化的nNOS表达增加4.3倍（p<0.01）。低盐组总eNOS增加3倍（p<0.01），在激活和抑制位点磷酸化的eNOS也平行增加（p<0.05）。在2周的高盐组中，尽管观察到尿NO代谢产物排泄增加，但一氧化氮合酶的表达或磷酸化未见变化。
总之，磷酸化nNOS和磷酸化eNOS表达的变化与总表达的变化平行，因此，在所研究的位点，nNOS和eNOS磷酸化的总体激活和抑制作用不会因饮食中盐的改变而改变。
