Calorie restriction (CR) extends lifespan and retards age-related chronic diseases in most species. There is growing evidence that the gut microbiota has a pivotal role in host health and age-related pathological conditions. Yet, it is still unclear how CR and the gut microbiota are related to healthy aging. Here, we report findings from a small longitudinal study of male C57BL/6 mice maintained on either ad libitum or mild (15%) CR diets from 21 months of age and tracked until natural death. We demonstrate that CR results in a significantly reduced rate of increase in the frailty index (FI), a well-established indicator of aging. We observed significant alterations in diversity, as well as compositional patterns of the mouse gut microbiota during the aging process. Interrogating the FI-related microbial features using machine learning techniques, we show that gut microbial signatures from 21-month-old mice can predict the healthy aging of 30-month-old mice with reasonable accuracy. This study deepens our understanding of the links between CR, gut microbiota, and frailty in the aging process of mice.

Five years ago, the National Institute of Health (NIH) introduced a mandate to revolutionize the way sex as a biological variable (SABV) is considered in NIH-funded preclinical research. Given the known effects of sex on aging physiology, pathology, treatment response, and the effectiveness of interventions it is particularly important that SABV be considered in basic biology of aging research. Five years after this mandate, a significant amount of published work funded by the National Institute on Aging (NIA) is still not including mice of both sexes and/or not considering sex differences or comparisons in preclinical studies. Here we review a cross-section of recently published NIA-funded research to determine adherence to this mandate. We discuss the state of the preclinical aging field in terms of SABV and suggest strategies for improving adherence to the NIH mandate. It is imperative that we consider SABV and include males and females in all aspects of aging biology research to improve health outcomes for all.

With an increasing aging population, the burden of age-related diseases magnifies. To alleviate this burden, geroprotection has been an area of intense research focus with the development of pharmacological interventions that target lifespan and/or healthspan. However, there are often sex differences, with compounds mostly tested in male animals. Given the importance of considering both sexes in preclinical research, this neglects potential benefits for the female population, as interventions tested in both sexes often show clear sexual dimorphisms in their biological responses. To further understand the prevalence of sex differences in pharmacological geroprotective intervention studies, we performed a systematic review of the literature according to the PRISMA guidelines. Seventy-two studies met our inclusion criteria and were classified into one of five subclasses: FDA-repurposed drugs, novel small molecules, probiotics, traditional Chinese medicine, and antioxidants, vitamins, or other dietary supplements. Interventions were analyzed for their effects on median and maximal lifespan and healthspan markers, including frailty, muscle function and coordination, cognitive function and learning, metabolism, and cancer. With our systematic review, we found that twenty-two out of sixty-four compounds tested were able to prolong both lifespan and healthspan measures. Focusing on the use of female and male mice, and on comparing their outcomes, we found that 40% of studies only used male mice or did not clarify the sex. Notably, of the 36% of pharmacologic interventions that did use both male and female mice, 73% of these studies showed sex-specific outcomes on healthspan and/or lifespan. These data highlight the importance of studying both sexes in the search for geroprotectors, as the biology of aging is not the same in male and female mice.Systematic Review Registration: [website], identifier [registration number].