Abstract/Description

Introduction: Atherosclerosis is a chronic disease characterized by plaque buildup within arteries, leading to cardiovascular diseases. Sleep fragmentation (SF), defined as less than seven hours of uninterrupted sleep, is a recognized risk factor and is commonly observed in menopausal women. Menopause causes a decline in estrogen and ovarian follicles, resulting in the loss of estrogen’s protective anti-atherogenic effects. SF exacerbates atherosclerosis by promoting oxidative stress and inflammation. Since SF contributes to atherosclerosis, and menopausal women are particularly affected by disturbed sleep, we aim to examine how menopause affects atherogenesis in the conditions of disturbed sleep.

Methods: We used the 4-vinylcyclohexene diepoxide (VCD) model to mimic human menopause. Six- to eight-week-old Apoe-/- mice were injected intraperitoneally with VCD for 20 days to deplete ovarian follicles and produce a hormone profile comparable to natural menopause. Vaginal cytology, cell staining and microscopy were performed to assess menopausal transition. At day 52, mice were placed on a high-fat diet, and sleep fragmentation (SF) was induced. Aged mice also underwent cytology. Ovaries, aorta, and hearts were collected. Hearts were stained with Picrosirius Red and MOVAT to assess plaques, fibrous caps, and necrotic cores.

Results: SF increased plaque and necrotic core areas, and reduced collagen content in SF Apoe-/- mice compared to home cage controls. Aged mice displayed persistent diestrus, characterized by neutrophil and epithelial cell abundance. 20-day VCD-treatment caused irregular estrous cycles but not full menopause, whereas 25-day treatment induced persistent diestrus. Ovarian histology of 20-day VCD-injected Apoe-/- mice might show structural changes of follicles. Notably, at 40 days after VCD-injection for 25 days, Apoe-/- mice already exhibited constant diestrus. Atherosclerotic plaques were present in VCD-treated, SF mice.

Discussion: Our results indicate SF exacerbates atherogenesis by increasing plaque vulnerability, and the VCD model effectively mimics menopausal transition. Persistent diestrus after 25 days of VCD treatment reliably indicates menopause. These findings support using VCD and aged mice to study postmenopausal atherosclerosis and SF-related cardiovascular risk.

Comments

Key Words:

Atherosclerosis, Menopause, Postmenopausal, Inflammation, Sleep Fragmentation

Presenting Author Name/s

Sarah Shackelford

Faculty Advisor/Mentor

Elena Galkina

Faculty Advisor/Mentor Email

galkinev@odu.edu

Faculty Advisor/Mentor Department

Biomedical and Translational Sciences, Cardiopulmonary and Metabolic Diseases Focus Group

College/School/Affiliation

Eastern Virginia Medical School (EVMS)

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VCD-Induced Model of Menopause to Study Effects of Sleep Fragmentation on Atherogenesis

Introduction: Atherosclerosis is a chronic disease characterized by plaque buildup within arteries, leading to cardiovascular diseases. Sleep fragmentation (SF), defined as less than seven hours of uninterrupted sleep, is a recognized risk factor and is commonly observed in menopausal women. Menopause causes a decline in estrogen and ovarian follicles, resulting in the loss of estrogen’s protective anti-atherogenic effects. SF exacerbates atherosclerosis by promoting oxidative stress and inflammation. Since SF contributes to atherosclerosis, and menopausal women are particularly affected by disturbed sleep, we aim to examine how menopause affects atherogenesis in the conditions of disturbed sleep.

Methods: We used the 4-vinylcyclohexene diepoxide (VCD) model to mimic human menopause. Six- to eight-week-old Apoe-/- mice were injected intraperitoneally with VCD for 20 days to deplete ovarian follicles and produce a hormone profile comparable to natural menopause. Vaginal cytology, cell staining and microscopy were performed to assess menopausal transition. At day 52, mice were placed on a high-fat diet, and sleep fragmentation (SF) was induced. Aged mice also underwent cytology. Ovaries, aorta, and hearts were collected. Hearts were stained with Picrosirius Red and MOVAT to assess plaques, fibrous caps, and necrotic cores.

Results: SF increased plaque and necrotic core areas, and reduced collagen content in SF Apoe-/- mice compared to home cage controls. Aged mice displayed persistent diestrus, characterized by neutrophil and epithelial cell abundance. 20-day VCD-treatment caused irregular estrous cycles but not full menopause, whereas 25-day treatment induced persistent diestrus. Ovarian histology of 20-day VCD-injected Apoe-/- mice might show structural changes of follicles. Notably, at 40 days after VCD-injection for 25 days, Apoe-/- mice already exhibited constant diestrus. Atherosclerotic plaques were present in VCD-treated, SF mice.

Discussion: Our results indicate SF exacerbates atherogenesis by increasing plaque vulnerability, and the VCD model effectively mimics menopausal transition. Persistent diestrus after 25 days of VCD treatment reliably indicates menopause. These findings support using VCD and aged mice to study postmenopausal atherosclerosis and SF-related cardiovascular risk.