Soluble Protein Oligomers induce Endoplasmic Reticulum Stress in Acute Conditions in Mesenteric Resistance Arteries from Male and Female Mice

Abstract

Background While amyloid β proteins, including toxic soluble oligomers, are a well established pathogenic mechanism of alzheimer's disease, SPOs are also found in the peripheral vascular system where their accumulation is considered to be a fundamental mechanism that underlies vascular dysfunction and cell death. However, the precise mechanism linking increased circulating levels of SPOs and vascular dysfunction remains unknown. We therefore hypothesized that SPOs are recognized by the vasculature leading to endoplasmic reticulum (ER) stress which further induces the release of SPOs and vascular injury. Methods & results Mesenteric resistance arteries (MRAs) from male and female wild type C57BL/6 mice (14-20 weeks old) were isolated to measure vascular function via a wire myograph. Concentration response curves to acetylcholine (10-9-10-5mol/L) and phenylephrine (10-9-10-5 mol/L) were performed. We observed that in acute conditions, and in arteries from both male and female mice, incubation with SPOs (0.1mM) caused exacerbated endothelium-dependent vasodilation compared to vehicle (F12 media, 5mL) [Male: EC50: Vehicle -6.6 ± 0.1 (n=7) vs. SPOs: -7.0 ± 0.1 (n=4), p=0.03; Female: EC50: Vehicle -6.7 ± 0.1 (n=6) vs. SPOs: -7.3 ± 0.06 (n=5), p=0.001]. A similar phenotype was observed in arteries treated with ER stress inducer tunicamycin (5mg/ml, positive control). This suggests that SPOs and ER stress exacerbate endothelium-dependent vasodilation, independent of sex. To determine whether SPO's are a cause of ER stress, arteries were treated with SPOs or vehicle for 15 min prior to incubation with an ER stress inhibitor 4-Phenylbutyric acid (4-PBA) (2mM) for 30 min. We observed that incubation with 4-PBA prevented the exacerbated vasodilation induced by SPOs in both male and female arteries, indicating that SPOs trigger ER stress in acute conditions and this is also independent of sex (Fig 1). On the other hand, to determine whether SPOs are a consequence of ER stress, arteries were incubated with ER stress inducer tunicamycin (5mg/ml) or vehicle for 15 min prior to incubation with the oligomer inhibitor K01-162 (10mM) for 30 min. Interestingly, the SPO inhibitor K01-162 did not prevent the exacerbated vasodilation induced by tunicamycin in arteries from male mice. However, this response was restored in arteries from female mice suggesting that ER stress induces the release of SPOs in arteries from female but not from male mice in acute conditions. Phenylephrine-induced vascular contraction did not change in the presence of tunicamycin or SPOs irrespective of sex. Conclusion We concluded that SPOs induce ER stress and subsequently exacerbate endothelium-dependent vasodilation in acute conditions irrespective of sex. However, ER stress induction in arteries from females also leads to the release of SPOs, escalating a feed-forward mechanism of further SPO release. Collectively, these results demonstrate that SPO's exacerbate endothelium-dependent vasodilation acutely and may contribute to vascular damage.