Poster L21 - Post-menopausal levels of follicle stimulating hormone are associated with muscle weakness in aged female mice and adipogenesis of fibro-adipogenic progenitors

Friday, October 25, 2024

3:30 PM - 4:30 PM EST

Location: Exhibit Hall E, F & G

Presenting Author(s)

Suyash Sinkar (he/him/his)

Undergraduate Student
University of Pittsburgh
Sewickley, Pennsylvania, United States

Co-Author(s)

ZH

Zachary Hettinger

Postdoctoral Fellow
Harvard Medical School, United States

Primary Investigator(s)

GG

Gabrielle Gilmer

MD/Ph.D Student
University of Pittsburgh, United States

Co-Author(s)

RK

Rylee Kopchak

Doctoral Student
Mayo Clinic, United States
HH

Hannah Houston

Research Technician
Spaulding Rehabilitation Hospital, United States
KW

Kai Wang

Instructor in Physical Medicine and Rehabilitation
Spaulding Rehabilitation Hospital
Charlestown, Massachusetts, United States
JB

Juliana Bergmann

Research Technician
Spaulding Rehabilitation Hospital, United States
EC

Ekaterina Creed

Research Laboratory Manager
Spaulding Rehabilitation Hospital, United States
NS

Nafiseh Shashahan

Research Technician
Spaulding Rehabilitation Hospital, United States
Michael T. Hermanto, MS

Medical Student
University of Vermont College of Medicine
Burlington, Vermont, United States
TM

Tabo Mkandawire

Undergraduate Student
Ohio State University, United States
AM

Amanda Miller

Research Technician
University of Pittsburgh, United States

Last Author(s)

FA

Fabrisia Ambrosio

Associate Professor of Physical Medicine and Rehabilitation
Spaulding Rehabilitation Hospital, United States

Introduction: While both men and women experience age-related loss of muscle mass and strength (sarcopenia), post-menopausal women are significantly more likely to experience sarcopenia than men. Despite the observed association between menopause and sarcopenia, the mechanisms through which menopause may contribute to muscle weakness have not yet been determined and thus, we lack female-specific treatments. Studies of menopause effects on muscle have been limited as current animal models of aging have primarily employed male rodents. Of the few studies that include female animals, translation has been limited by the fact that rodents spontaneously rejuvenate their ovarian follicles following estropause (menopause equivalent for animals with an estrous cycle), and therefore do not maintain a menopausal phenotype. Further, efforts to mimic menopause-like changes in animals have primarily been used on young rodents, contrasting when menopause naturally occurs in humans and ultimately precluding an understanding of the interaction between menopause and aging on muscle weakness. To help close this gap in knowledge, our study employed an in-vivo model by simulating menopause in aged female mice to study its effects on skeletal muscle. Fat accumulation, known to affect muscle quality, was also studied to determine if menopause affects fat content. In addition, fibro-adipogenic progenitor cells (FAPs), the main cell population involved in skeletal muscle extra-cellular matrix (ECM) function, were isolated for an in-vitro analysis to determine if menopause-associated sex hormone changes induce an adipogenic state, and thus, fat accumulation in skeletal muscle.

Materials and

Methods: For in vivo experiments, the ovarian toxin 4-vinylcyclohexene diepoxide (VCD) was injected into middle-aged female mice to induce menopause (menopause group). Mice were divided into two groups and received daily injections of either VCD or sesame oil, a control (non-menopause group), for 10 days. Muscle contractile force was measured at day 195 (late menopause) using a force transducer, and tibialis anterior (TA) muscle sections were collected for histological analyses at days 25, 115, 155, and 195. Lipid accumulation of the TA was quantified using Oil Red O (ORO) staining and a triglyceride assay. For in vitro experiments, fibro-adipogenic progenitor cells (FAPs) were isolated from naïve (no injection) mice and sorted using Fluorescent Activated Cell Sorting. To better understand the effects of sex hormones on FAP function, FAPs were cultured for 7 days in estradiol (decreased in menopause), progesterone (decreased in menopause), follicle-stimulating hormone (FSH, increased in menopause), or luteinizing hormone (LH, increased in menopause). FAP adipogenic transition was quantified by relative proportions of PDGFRα+ (related to stem cell state) and CD10+ cells. Previous research work has established that FAPs lose CD10+ expression upon conversion to the adipogenic state. In this regard, a double positive indicated a stem cell state, single positive indicated an actively transforming state, and double negative indicated an adipogenic state.

Results, Conclusions, and Discussions: From the in-vivo study, muscle contractile testing demonstrated that menopause mice had decreased muscle contractile force relative to the non-menopause group (Figure 1). However, there was no significant difference in the myofiber cross-sectional area when comparing menopause and non-menopause groups (Figure 2), suggesting that muscle weakness was unrelated to muscle size. Quantification of ORO area relative to total muscle area revealed increased lipid accumulation, and the triglyceride content assay revealed increased triglyceride count in menopausal mice (Figures 3/4). These data collectively indicate an association between muscle weakness and lipid accumulation in the skeletal muscle of menopausal mice. From the in-vitro study, treatment of FAPs with FSH resulted in decreased proportion of PDGFRα+/CD10+ and increased proportion of PDGFRα-/CD10- (double negative), thus indicating adipogenic transition of FAPs with exposure to FSH (Figure 5). As FSH levels increase at menopause onset, we hypothesize that menopause may induce fat accumulation and muscle weakness by FSH-induced FAP adipogenesis. In this regard, FSH-targeted therapies may provide a possible a means for the treatment of menopause-associated sarcopenia in older women.

Acknowledgements (Optional): I would like to thank Dr. Gilmer, Dr. Hettinger, Dr. Ambrosio and all of my mentors in the Ambrosio lab for their support and guidance in the undergraduate research experience.