The percentage of horses considered “aged” has been increasing, with estimates of all horses older than 15 years of age ranging from 22 to 34% (1, 2). In older horses, basal metabolic rate decreases (3), and plasma metabolite and hormone concentrations related to energy metabolism change with age (4). The occurrence of age-related diseases increases in old animals (5, 6), and enhanced inflammatory reactivity known as inflammaging may arise with their onset (7, 8). Franceschi et al. termed the persistence of low-grade chronic inflammatory status as inflammaging (9), which involves the accumulation of damaged macromolecules and cellular debris because of increased production and chronically inhibited damage surveillance and repair functions in multiple tissues (10). The secretion of proinflammatory cytokines from senescent cells accumulates in tissues with age, termed “senescence-associated secretory phenotype (SASP),” and contributes to the onset of inflammaging (11). As plasma concentrations of macromolecules, metabolites, hormones, and enzymes involved in energy metabolism change in animals with age, monitoring these concentrations can aid in detecting inflammaging. Appropriate treatment for inflammaging at an early stage may prevent various age-related diseases. Arfuso et al. reported that serum C-reactive protein (CRP) and α2 macroglobulin concentrations increased in horses with aging, and acute phase proteins (APPs) such as CRP could be good parameters to assess inflammaging in horses (12). In this study, changes in metabolite and hormone concentrations and enzyme activities involved in energy metabolism in the plasma of clinically healthy riding horses of various ages were measured. Macromolecules that vary with age are considered to be useful biomarkers for diagnosing inflammaging in riding horses.

All horses were clinically healthy, and their BCS values were 4 or 5 on the 9-point scale. Plasma glucose, TP, creatinine, and NEFA concentrations in horses remained unchanged through adolescence, middle age, and old age. TG, T-Cho, BUN, and insulin concentrations increased with age, while adiponectin concentrations decreased. MDA and SAA concentrations did not change between adolescence and middle age but increased with aging beyond middle age. The MDA concentrations in the old age group were significantly higher than those in the adolescent group (Table 1). Plasma AST, ALT, and GPx activities did not change in the horses with age, while plasma SOD and leukocyte AMPK activities decreased. The plasma M/L ratio did not change with aging (Table 2).

According to their plasma ALT and AST activities and creatinine and BUN concentrations, it could be presumed that no riding horses in this study had any specific lesions in their livers, kidneys, or skeletal muscles. Although the BCS of riding horses did not increase with aging, they showed symptoms of insulin resistance, including increases in both plasma insulin and TG. In healthy, middle-aged humans, insulin resistance and dyslipidemia associated with the accumulation of excess visceral adipose tissue are frequently observed (19, 20). A horse at 17 years of age is equivalent to a human at 50 years of age (21). Dysregulation of lipid metabolism became clear in riding horses aged over 17 years. The increases in plasma MDA and SAA concentrations and decreases in SOD activities were notable. White adipose tissue dysfunction and the accumulation of visceral fat occur in elderly humans (22, 23). Although older riding horses did not show marked obesity, they accumulated more dysregulated adipocytes (enlarged adipocytes) as visceral fat compared to younger horses (4). In older riding horses, plasma adiponectin concentrations decreased, as is the case for aging humans. The molecular mechanisms responsible for adiponectin concentrations may be the direct result of inflammatory cells suppressing reactive oxygen species (ROS) and cytokines, inhibiting the NF-κB inflammatory signaling pathway, and downregulating inflammatory responses (24, 25). Decreasing adiponectin concentrations lead to dyslipidemia and insulin resistance (26). Obesity is known to affect immunity and inflammation in horses, and failure to control for BCS can obscure the interpretation of these results (27, 28). Increased plasma SAA concentrations in obese animals could be the result of normal fat mass and/or increased expression and secretion of SAA from dysfunctional adipose tissue or other tissues (26). Concentrations of MDA as a product of lipid peroxidation (29) increase with chronic inflammation as a result of ROS. Increased MDA in older riding horses is considered to be induced by lower adiponectin concentrations and decreased SOD activities.

SAA from dysfunctional adipose tissue may act locally to alter cytokine production and fat metabolism. Additionally, it may act systemically on the liver, muscle, cells of the immune system, and the vasculature to impact insulin resistance and atherosclerosis (26). Although SAA is commonly used as a marker for APP in cats and horses (30, 31), it may be used as a marker of inflammaging in riding horses when combined other lipid metabolism markers.

Although riding horses with sufficient exercise continually seem to be less likely to develop inflammaging, horses older than 17 years tend to show a proinflammatory status with dysregulated lipid metabolism. Current SAA reagents do not have enough sensitivity to confirm a significant difference; however, it is expected that improved reagent performance will allow SAA to become a biomarker that can differentiate the presence or absence of inflammaging with dysregulated lipid metabolism in riding horses.

This study has some limitations. First, the number of horses in each group was small. Second, the insulin, adiponectin, and SAA antibodies for measurements were not specific to the equine species. An age- and sex-matched control group, fed on a similar diet, would be required to help eliminate the possible influence of the aforementioned factors. Further studies should be performed to evaluate the usefulness of SAA and other macromolecules such as TG, NEFA, MDA, and SOD as biomarkers for diagnosing inflammaging in riding horses.

Changes in metabolite and hormone concentrations and enzyme actiMalate dehydrogenase UV assayities relating to energy metabolism in the plasma of clinically healthy riding horses of various ages were measured. All horses had BCS values of 4 or 5, without obesity. Plasma TG, T-Cho, BUN, and insulin concentrations increased gradually with aging, whereas plasma adiponectin concentrations decreased. MDA and SAA concentrations increased with aging, while plasma AST, ALT, and GPx activities did not change. Furthermore, plasma SOD and leukocyte AMPK activities decreased. Although riding horses with sufficient and continuous exercise seem to be less likely to develop inflammaging, horses over the age of 17 years tend to show a proinflammatory status with the disorder of lipid metabolism. SAA, in combination with other lipid metabolism markers, may be a useful biomarker for inflammaging with dysregulated lipid metabolism in riding horses.

The original contributions presented in the study are included in the article/supplementary material, further inquiries can be directed to the corresponding author.

The animal study was approved by the Nippon Veterinary and Life Science University Animal Research Committee. The study was conducted in accordance with the local legislation and institutional requirements.

YA: Writing – original draft, Validation, Resources, Methodology, Investigation, Formal analysis, Data curation, Conceptualization. TA: Writing – review & editing, Validation, Supervision, Project administration, Formal analysis, Data curation, Conceptualization. YT: Writing – review & editing, Validation, Resources, Methodology, Data curation.