# Transcriptional Factors Related to Cellular Kinetics, Apoptosis, and Tumorigenicity in Equine Adipose-Derived Mesenchymal Stem Cells (ASCs) Are Influenced by the Age of the Donors

## TL;DR

Older horses have stem cells that grow more slowly and may be less safe for treatments due to age-related changes in gene activity.

## Contribution

The study introduces a Gompertzian model and Bax/Bcl2 ratio as predictive tools for assessing the functional state of equine ASCs based on donor age.

## Key findings

- ASCs from older horses show slower growth and reduced survival.
- Older ASCs have altered gene expression related to tumor formation and apoptosis.
- Prolonged culture increases tumorigenic and pluripotency marker expression in ASCs.

## Abstract

Donors’ aging affects stem cells’ ability to regenerate damaged tissues, which is crucial for treating injuries in horses. This study examined how the age of donor horses influences the growth, survival, and potential tumor risks of adipose-derived mesenchymal stem cells (ASCs). Cells from young (<5 years old), middle-aged (5–15 years old), and elderly (>15 years old) horses were analyzed to assess their ability to multiply, maintain function, and avoid harmful genetic changes over time. The proposed Gompertzian model illustrates the proliferative ability of cells and, together with the Bax/Bcl2 ratio, could be a good predictive algorithm concerning the functional state of the isolated cells for detecting potential senescence. The findings show that ASCs from older horses have slower growth rates, a reduced ability to survive, and an altered expression of genes linked to tumor formation and apoptotic processes. These age-related changes raise concerns about the effectiveness and safety of using autologous horse stem cells for treatments later in life. However, the study also highlights potential ways to improve stem cell therapies by identifying factors influencing cell aging. This research provides valuable insights for veterinarians and scientists working on regenerative medicine, helping to develop safer and more effective treatments for equine injuries and age-related conditions.

The impact of donor age on Adipose-derived mesenchymal stem cell (ASC) functionality and safety remains insufficiently characterized, particularly in equine models. This study investigates the influence of age on ASCs proliferation dynamics and the expression of tumorigenic and apoptosis-related markers. Equine ASCs were isolated from juvenile (<5 years), middle-aged (5–15 years), and geriatric (>15 years) horses and assayed across multiple passages. The relative mRNA expressions of pluripotency (Oct4), tumorigenic (CA9), and apoptosis-related (Bax and Bcl 2) markers were evaluated. The Gompertz growth model, population doubling time (PDT), and tissue non-specific ALP activity also followed. The expression of pluripotency and tumorigenic markers showed passage-dependent up-regulation, raising concerns about prolonged culture expansion. Apoptotic regulation displayed a shift with aging, as evidenced by alterations in the Bax/Bcl2 ratio, suggesting compromised cell survival in older ASCs. An age-associated decline in proliferation rates was established, as evidenced by declining alkaline phosphatase (ALP) activity. These findings underscore the necessity for stringent age-based selection criteria in equine stem cell therapies and the challenges associated with using autologous stem cells for regenerative therapies in aged horses. Future research should focus on molecular interventions to mitigate age-related functional decline, ensuring the safety and efficacy of ASCs-based regenerative medicine in equine practice.