Vascular dysfunction in young, mid-aged and aged mice with latent cytomegalovirus infections

Abstract

Human cytomegalovirus (HCMV) is associated with vascular diseases in both immunosuppressed and immunocompetent individuals. CMV infections cycle between active and latent phases throughout life. We and others have shown vascular dysfunction during active mouse CMV (mCMV) infections. Few studies have examined changes in physiology during latent CMV infections, particularly vascular responses or whether the negative effects of aging on vascular function and fertility will be exacerbated under these conditions. We measured vascular responses in intact mesenteric and uterine arteries dissected from young, mid-aged, and aged latently mCMV-infected (mCMV genomes are present but infectious virus is undetectable) and age-matched uninfected mice using a pressure myograph. We tested responses to the α(1)-adrenergic agonist phenylephrine, the nitric oxide donor sodium nitroprusside, and the endothelium-dependent vasodilator methacholine. In young latently mCMV-infected mice, vasoconstriction was increased and vasodilation was decreased in mesenteric arteries, whereas both vasoconstriction and vasodilation were increased in uterine arteries compared with those in age-matched uninfected mice. In reproductively active mid-aged latently infected mice, mesenteric arteries showed little change, whereas uterine arteries showed greatly increased vasoconstriction. These vascular effects may have contributed to the decreased reproductive success observed in mid-aged latently mCMV-infected compared with age-matched uninfected mice (16.7 vs. 46.7%, respectively). In aged latently infected mice, vasodilation is increased in mesenteric and uterine arteries likely to compensate for increased vasoconstriction to mediators other than phenylephrine. The novel results of this study show that even when active mCMV infections become undetectable, vascular dysfunction continues and differs with age and artery origin.

Fig. 1.

Immunofluorescent staining for β-galactosidase protein expression in tissues from young and aged latently mouse cytomegalovirus (mCMV)-infected and uninfected mice. Heart, kidney, liver, lung, and spleen tissues from young (A) and aged (B) mice that were uninfected or infected at 1 to 2 mo of age were collected 2 wk postinfection (acute infection; A) or >2 mo postinfection (latent infection: young, A; and aged, B). Tissues were stained for β-galactosidase protein expression (green) to test for active infection. Nuclei were stained with 4′, 6-diamidino-2-phenylindole (DAPI, blue).

Fig. 2.

Responses to phenylephrine (PE) by mesenteric and uterine arteries. PE-induced vasoconstriction was measured in mesenteric (A–C) and uterine (D–F) arteries from young (A and D), mid-aged (B and E), and aged (C and F) latently mCMV-infected and age-matched uninfected mice. Results for each curve were summarized and expressed as means ± SE percent decrease in lumen diameter compared at each PE concentration with the initial equilibrated diameter. Significant differences between the curves was calculated using a repeated-measures 2-way ANOVA (P < 0.05). This was followed by Holm-Sidak's post hoc analysis to determine significance between points within the curves (P < 0.05). *P < 0.05, significant differences; n = number of animals.

Fig. 3.

Responses to methacholine (ME) by mesenteric and uterine arteries. ME-induced vasodilation was measured in mesenteric (A–C) and uterine (D–F) arteries from young (A and D), mid-aged (B and E), and aged (C and F) latently mCMV-infected and age-matched uninfected mice. Results for each curve were summarized and expressed as means ± SE percent increase in lumen diameter compared with the initial preconstricted diameter and normalized to the passive lumen diameter. Significance was assessed as for Fig. 2. *P < 0.05, significant differences; n = number of animals.

Fig. 4.

Nitric oxide contribution to ME-induced vasodilation in mesenteric arteries from young, mid-aged, and aged uninfected and latently mCMV-infected mice. ME-induced vasodilation was measured in mesenteric arteries from young (A–C), mid-aged (D–F), and aged (G–I) latently mCMV-infected (B, E, and H) and uninfected (A, D, and G) mice in the presence and absence of N^G-nitro-l-arginine methyl ester (l-NAME). Results for each curve were expressed as means ± SE, and the dose-response curves were compared as in Fig. 2. The area under each curve was calculated, and the difference between 2 curves on the same graph was obtained. These differences in area under the curve were compared between latently mCMV-infected and uninfected groups (C, F, and I) with a Student's t-test. *P < 0.05, significant differences; n = number of animals.

Fig. 5.

Responses to sodium nitroprusside (SNP) by mesenteric and uterine arteries. SNP-induced vasodilation was measured in mesenteric (A–C) and uterine (D–F) arteries from young (A and D), mid-aged (B and E), and aged (C and F) latently mCMV-infected and age-matched uninfected mice. Results were summarized and presented as means ± SE, and significant differences were assessed as for Fig. 2. *P < 0.05, significant differences; n = number of animals.

Fig. 6.

Distensibility curves for mesenteric and uterine arteries. Changes in lumen diameter were measured after stepwise increases in intraluminal pressure in Ca²⁺-free EGTA physiological saline solution in the presence of papaverine in mesenteric (A–C) and uterine (D–F) arteries from young (A and D), mid-aged (B and E), and aged (C and F) latently mCMV-infected and age-matched uninfected mice. Results were expressed as means ± SE percent increase in lumen diameter compared with the initial diameter at 4 mmHg. Significant differences were assessed as for Fig. 2. Where error bars are not visible, the errors are too small to be seen at this scale. *P < 0.05, significant differences; n = number of animals.

Fig. 7.

Comparison of vascular responses in mesenteric and uterine arteries from young, mid-aged, and aged uninfected mice. PE-induced vasoconstriction for mesenteric (A) and uterine (D) arteries, ME-induced vasodilation for mesenteric (B) and uterine (E) arteries, and SNP-induced vasodilation for mesenteric (C) and uterine (F) arteries were measured and compared among the 3 age groups. Results were summarized and are presented as means ± SE. Significant differences between the curves were calculated using a repeated-measures 2-way ANOVA with Holm-Sidak's post hoc analysis (P < 0.05). Significant differences: *P < 0.05, mid-aged compared with young; #P < 0.05, aged compared with young.