Influence of a High-Fat Diet on Cardiac iNOS in Female Rats

Abstract

Overexpression of inducible nitric oxide synthase (iNOS) is a key link between high-fat (HF) diet induced obesity and cardiovascular (CV) disease. Several studies have reported that oestradiol has cardioprotective effects that may be mediated through reduction of iNOS activity/expression. In the present study, female Wistar rats were fed a standard diet or a HF diet (balanced diet for laboratory rats enriched with 42% fat) for 10 weeks. Gene and protein expression of iNOS were measured in heart tissue. HF diet-fed rats exhibited a significant increase in cardiac iNOS mRNA by 695% (p<0.05), iNOS protein level by 248% (p<0.01), without changes in nitrate/nitrite levels. Expression of CD36 protein in plasma membranes was increased by 37% (p<0.05), while the concentration of free fatty acids (FFA) was reduced by 25% (p<0.01) in HF diet-fed rats. Expression of the p50 subunit of nuclear factor-κB (NFκB-p50) in heart lysate was increased by 77% (p<0.01) in HF diet-fed rats.Expression and phosphorylation of protein kinase B (Akt) and extracellular signal-regulated kinases 1/2 (ERK1/2) in control and HF diet-fed rats were also examined. Expression of Akt and ERK1/2 were unchanged between the groups. There was a significant increase in the ratio of phospho-Akt/total Akt but not for phospho-ERK1/2/total ERK1/2/ in HF-fed rats.Estrogen receptor-α levels (by 50%; p<0.05) and serum oestradiol concentrations (by 35%; p<0.05) were examined and shown to be significantly reduced in HF diet-fed rats. Our results revealed that a HF diet led to increased iNOS expression, most likely via a mechanism involving Akt and NFκB-p50 proteins. Decreased levels of oestradiol and ERα protein in the HF-fed group, in combination with increased iNOS levels are consistent with the hypothesis that oestradiol has a cardioprotective effect through its ability to regulate iNOS expression.Key words: cardioprotection, cardiovascular disease, oestradiol, inducible nitric oxide synthase, obesity Abbreviations: Akt, protein kinase B; CD36, cluster of differentiation 36; CHD, coronary heart disease; CV, cardiovascular; CVD, cardiovascular disease; DMT2, diabetes mellitus type 2; ERα, oestrogen receptor-α; ERs, oestrogen receptors; ERK1/2, extracellular signalregulated kinases 1/2; FFA, free fatty acids; HF, high fat; HOMA-IR, HOMA-index of insulin resistance; HOMA-β, HOMA-index of β-cell function; IκB, inhibitor of NFκB; iNOS, inducible nitric oxide synthase; IR, insulin resistance; NFκB-p50, the p50 subunit of nuclear factor-κB; TES, N-[Tris(hydroxymethyl)methyl]-2-aminoethanesulfonic acid; TG, triglycerides VSMCs, vascular smooth muscle cells.

Changes in cardiac Na+/K+-ATPase expression and activity in female rats fed a high-fat diet

“…we have previously demonstrated that a HF diet in the same rats used in this study caused an increase in cardiac iNOS mRNA and protein levels by a mechanism involving increased activation of Akt [60]. The mechanism by which iNOS may regulate RhoA expression in the heart appears to be a combination of transcriptional and translational upregulation of the RhoA gene and decreased degradation of the RhoA protein [56,25,61,62].…”

“…This may be a consequence of enhanced RhoA/ROCK signalling, since it has been shown that both partial deletion of ROCK2 and its inhibition by fasudil prevents Ser 307 phosphorylation of IRS-1 in mice fed a HF diet for 17 weeks [56]. In addition, we have previously reported that in the same rats used in this study, a HF diet altered the expression of cardiac cluster of differentiation 36 (CD36) and fatty acid metabolism [60], leading to the accumulation of intramyocellular lipids, which in turn may activate serine kinases such as protein kinase C and mammalian target of rapamycin to consequently induce serine phosphorylation of IRS-1 [69,70].…”

“…We have previously demonstrated that a HF diet decreases serum estradiol level, as well as cardiac estrogen receptor-α (ERα) signalling and believe that due to the lack of beneficial action of estradiol, some of HF diet effects on female heart are similar to those observed in male rats and they include the up-regulation of cardiac iNOS expression [60] and consequential stimulation of RhoA/ROCK signalling and decrease in IRS-1/PI3K association and the expression of ɑ1 subunit of Na + /K + -ATPase. However, why reduced estradiol affected the level of cardiac Na + /K + -ATPase ɑ1 subunit expression but not ɑ2 subunit expression remains to be elucidated.…”

Candidate SNP Markers of Atherogenesis Significantly Shifting the Affinity of TATA-Binding Protein for Human Gene Promoters Show Stabilizing Natural Selection as a Sum of Neutral Drift Accelerating Atherogenesis and Directional Natural Selection Slowing It

“…The promoter of human gene NOS2 carries a known SNP marker (NOS2:−51T>C) of malaria resistance [86,87], which is caused by overexpression of inducible nitric oxide synthase encoded by this gene. Due to PubMed [38], we know that an NO excess is an accelerator of atherogenesis according to both physiological [88] and nutritional [89] original research articles. Therefore, the known SNP marker (NOS2:−51T>C) of malaria resistance [86,87] could be additionally regarded as a candidate SNP marker of accelerated atherogenesis (Table 3).…”

Effects of estrogen on fatty-acid-induced cytotoxicity in mouse Neuro-2a neural cells

“…, 2006 ). In female rats, a high fat diet led to increased iNOS expression, and decreased levels of E2 and ERα protein ( Jovanovic et al . , 2017 ).…”