Why new models are a must to understand female aging

A new study highlights the inability of most animal models to accurately translate features of female aging.

Knowledge of the basic biology of aging has significantly improved due to the use of the usual suspects of model organisms such as Caenorhabditis elegans (roundworms), Drosophila melanogaster (fruit flies) and Rodentia (rodents). Such model organisms have provided – and still provide – important information on how to monitor, manipulate and measure biological systems over their brief lifespans, and, moreover, the systems in these models share physiological characteristics with humans, thereby providing insights into human physiology and pathophysiology. However, significant inequities between men and women have existed for several years and continue to exist.

With age, both men and women face problems in carrying out their daily activities due to functional disabilities. However, studies indicate that such disabilities most often are not distributed equally – they often affect more women as compared with men [1]. Moreover, most of the animal models are unable to replicate female aging. For instance, females are four times more susceptible to osteoporosis as compared with males, but aging female rodents do not show a decline in bone mass. The incidence of Alzheimer’s disease and non-AD dementia is higher in females when compared with males, but this is not replicated in animal models. Along with these, several more disconnects exist between animal models and female aging but they are rarely identified due to the extensive use of male models in aging studies.

Longevity.Technology: The knowledge gap on how aging impacts the onset and progression of diseases in females and can lead to worsened health outcomes. As per the meta-analyses on disability status of the last 20 years in France, the US and Spain, females live with higher morbidity as compared with similarly-aged males. Females were also reported to be 50 percent more likely to undergo heart attack misdiagnosis and 33 percent more likely to undergo a stroke misdiagnosis as compared with males. 80 percent of drugs were removed from markets due to adverse events in females between 1997 and 2000 [2]. Even in 2020, females reported more side effects to prescription drugs for age-related diseases as compared with males. Lack of understanding of aging female physiology causes difficulty in diagnosis, treatment and prevention of diseases in older females, so on International Women’s Day, we look at how research into women’s longevity is affected by limitations of animal models.

A new study published in Nature Aging indicated the limitations associated with commonly used aging models and recommended better ones that would throw more light into female aging and age-related diseases and address the gap that currently exists in geroscience research.

Lack of studies focused on female aging

Female aging in humans is closely associated with reproductive senescence and menopause; this could affect tissues and organs throughout the body. Pregnancy and breastfeeding are a few other factors that may also impact female aging.

Female aging post menopause

Females mostly spend over one-third of their lives post-menopause, and this period is also associated with an increased disease onset. The onset of menopause is defined as when the menstrual cycles have stopped for at least 12 months along with loss of ovarian follicular function and changes in circulating sex hormone profiles. The role of reproductive senescence has been largely overlooked in the onset of many other age-related diseases in preclinical studies mostly because most animal models did not show human menopausal phenotypes.

Studies indicate that approximately over 70 percent of age-related diseases were found to be associated with or affected by menopause; however, only a few of the studies considered menopause to play a role in aging. Similarly, pregnancy, birthing, and breastfeeding were found to play roles in the increased or decreased incidence of age-related diseases but were not considered in aging biology research.

Representative models of female aging

A 2013 study reported that mice were unable to mimic the responses of inflammatory diseases seen in humans. This led to the increased selection of ‘humanized’ or ‘dirty’ mice which could better replicate the intricacies of human immune responses. Aging biology also came up with different animal models for the study of female aging [2].

Ovariectomy model of menopause

The ovariectomy model (OVX) removes the ovaries from animals surgically causing loss of sex hormones that mimic human menopause. It provides an opportunity to study the signaling cascades and phenotypes from individual sex hormones, but the model has several limitations. The first is that the ovaries remain intact in case of natural menopause which is not the same in OVX. This in turn causes abrupt stopping of ovarian function as compared with the progressive loss of ovarian function that occurs during natural menopause. This progressive loss is important since several menopause-related symptoms are present during the transient period, also termed perimenopause. OVX also leads to a drop in all ovarian-produced sex hormones including those not directly involved in menopause such as testosterone.

VCD model of menopause

The 4-vinyl cyclohexene diepoxide (VCD) model of menopause allows the progressive loss of ovarian sex hormones and preserves intact ovaries. VCD is an ovarian-specific toxin that causes apoptosis of primordial and primary ovarian follicles. VCD models also enable the evaluation of both perimenopause and menopause phases. However, this model is costly, time-consuming, and labor-intensive as compared with OVX. Moreover, the VDC has mostly not taken place in young animals, thereby making tissue and systemic cell responses age-dependent.

Spiny mice as naturally occurring menopause model

The African spiny mouse, Acomys cahirinus was the first mouse species that was reported to menstruate and undergo a gradual menopausal transition similar to humans. These mice show menopausal phenotype typically at 36 months. A drop in estradiol was observed between 1 and 2 years, however, no changes were observed in the case of testosterone which is similar to natural menopause in humans. However, further studies are required to ensure whether the menopause phenotype in these mice can be used to study menopause-induced systemic changes in humans.

Menopause status in rodents

It is difficult to know which rodents show premenopause, perimenopause, or menopause state due to the low availability of collectible serum from rodents, low levels of circulating hormones, and insensitivity of commercially available assays. Although mass spectrometry has been observed to be effective in determining sex hormone profiles in mice and rats, the technique is expensive and requires large samples. The development of more cost-effective, reliable and reproducible assays is required for the quantification of female sex hormones in rodents. Until then, other procedures can be used for the tracking of the estrous cycle and estropause status such as histological and postmortem evaluations.

Pregnancy, birthing and breastfeeding

Although several studies have reported pregnancy, childbirth, and breastfeeding to impact both longevity and healthspan, they are mostly included in studies focusing only on reproductive aging. Female breeders are mostly excluded from research studies, most of the female rodents obtained from animal vendors have never produced a litter as well as never breastfed. Recent studies have indicated several pregnancy-related changes occur that can help to understand aging mechanisms as well as age-related diseases.

Cell-culture systems

Studies have reported cell-culture systems to be important in evaluating aging mechanisms, but there has been little consideration of sex in such systems. However, cell culture conditions can identify sex-dependant changes associated with aging. Several changes from conventional methods can help to better understand female cellular aging such as culturing of cells in phenol red free media and the presence of serum isolated from age-matched male or female organisms.

Four core genotypes model

The four core genotypes model involves mice with XX chromosomes with male or female gonads or mice with XY chromosomes with male or female gonads. It helps to understand the chromosomal and hormonal differences in the development of age-related diseases. For example, this model showed that XX chromosomes increased lifespan which was independent of gonad. However, one main limitation of this model is that such mice are not available commercially so are not bred and aged in-house.

Time for change

Very few studies outside reproductive biology have used models that reiterated key features of female aging. This resulted in a lack of understanding of how to treat age-related diseases in females. Research can do, and must do, better. More studies need to incorporate representative models of female aging such as pregnancy, menopause and breastfeeding – evaluating features unique to female aging will help understand the fundamental biology of aging, and will, in turn, improve women’s healthspan and longevity.

[1] https://www.tandfonline.com/doi/full/10.1080/09581596.2018.1492092
[2] https://www.nature.com/articles/s43587-023-00509-8