Map2k6 is a potent genetic modifier of arterial rupture in vascular Ehlers-Danlos syndrome mice
Caitlin J. Bowen, Rebecca Sorber, Juan Francisco Calderón Giadrosic, Jefferson J. Doyle, Graham Rykiel, Zachary Burger, Xiaoyan Zhang, Wendy A. Espinoza Camejo, Nicole Anderson, Simone Sabnis, Chiara Bellini, Elena Gallo MacFarlane, Harry C. Dietz
Caitlin J. Bowen, Rebecca Sorber, Juan Francisco Calderón Giadrosic, Jefferson J. Doyle, Graham Rykiel, Zachary Burger, Xiaoyan Zhang, Wendy A. Espinoza Camejo, Nicole Anderson, Simone Sabnis, Chiara Bellini, Elena Gallo MacFarlane, Harry C. Dietz
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Research Article Cardiology Genetics Vascular biology

Map2k6 is a potent genetic modifier of arterial rupture in vascular Ehlers-Danlos syndrome mice

Abstract

Aortic dissection or rupture is a major cause of mortality in vascular Ehlers-Danlos syndrome (vEDS), a connective tissue disorder caused by heterozygous mutations in the collagen type III alpha 1 chain (COL3A1) gene. C57BL6/J (BL6) mice carrying the Col3a1G938D/+ mutation recapitulate the vEDS vascular phenotype and die suddenly of aortic rupture/dissection. However, 129S6/SvEvTac (referred to here as 129) mice expressing the same Col3a1G938D/+ mutation show near-complete lifelong protection from vascular rupture. To identify genetic modifiers of vascular risk in vEDS, we performed genome-wide genotyping of intercrossed BL6/129 vEDS mice stratified by survival and identified a significant protective locus encompassing a variant in Map2k6, encoding mitogen-activated protein kinase kinase 6 (M2K6), a p38-activating kinase. Genetic ablation of Map2k6 rendered previously protected 129 vEDS mice susceptible to aortic rupture, in association with reduced protein phosphatase 1 activity and increased PKC and ERK phosphorylation. Accelerated vascular rupture in vEDS mice treated with a pharmacological inhibitor of p38 was rescued by concomitant ERK antagonism, supporting an opposing role for ERK and p38 in the modification of aortic rupture risk in vEDS. These results suggest that pharmacologic strategies aimed at mimicking the effect of this natural protective pathway may attenuate aortic rupture risk in vEDS.

Authors

Caitlin J. Bowen, Rebecca Sorber, Juan Francisco Calderón Giadrosic, Jefferson J. Doyle, Graham Rykiel, Zachary Burger, Xiaoyan Zhang, Wendy A. Espinoza Camejo, Nicole Anderson, Simone Sabnis, Chiara Bellini, Elena Gallo MacFarlane, Harry C. Dietz

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Figure 1

The 129Sve background protects vEDS mice from premature death.

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The 129Sve background protects vEDS mice from premature death. (A) Kapla...
(A) Kaplan-Meier survival curve comparing control (Col3a1+/+) (n = 199) and vEDS (Col3a1G938D/+) mice (n = 191) on a BL6 background with control (n = 18) and vEDS mice (n = 31) on a 129 background. Significant differences were calculated using log-rank (Mantel-Cox) analysis (*** P < 0.001). (B) Kaplan-Meier survival curve comparing male BL6 vEDS male (n = 82) and female (n = 109) mice with 129 vEDS male (n = 15) and female (n = 16) mice. Significant differences were calculated using log-rank (Mantel-Cox) analysis (*** P < 0.001). (C) Verhoeff–Van Gieson (VVG), Masson’s trichrome, and Picrosirius red (PSR) staining of control and vEDS aortic cross sections at 2 months of age. White arrows indicate elastin fiber breaks. Scale bar is 50 microns. (D) Quantification of aortic wall thickness in aortic cross sections. P value refers to 2-way ANOVA with Holm-Šídák post hoc test (*** P < 0.001). (E) Quantification of elastin fiber breaks in VVG-stained aortic cross sections. Asterisks identify discovery (q < Q) by Kruskal-Wallis test with post hoc false discovery rate correction. (F) Quantification of collagen content in aortic cross sections, as measured by normalized PSR intensity. P value refers to 2-way ANOVA with Holm-Šídák post hoc test (*** P < 0.001). (G) Immunoblot of aortic lysates obtained from the proximal descending aortas of control and vEDS mice in the BL6 and 129 backgrounds at 2 months of age, probed with antibodies directed against phosphorylated PKCβ at residue Ser660 (pPKC) and phosphorylated ERK (pERK1/2). (H) Quantification for immunoblot shown in G. P value refers to 2-way ANOVA with Holm-Šídák post hoc test for p-PKC and Kruskal-Wallis test for p-ERK (* P < 0.05, *** P < 0.001). For D–F and H, each symbol represents an independent biological replicate, with unfilled symbols representing male samples. Error bars show mean ± SEM. Black circles represent control mice and red squares represent vEDS mice.