The metabolic profiling of cheetahs (Acinonyx jubatus): A systems biology approach to understanding the chronic diseases they suffer in captivity. – By Dr. Adrian Tordiffe in collaboration with the AfriCat Foundation
In captivity, cheetahs (Acinonyx jubatus) are known to suffer from several chronic diseases that do not occur in their wild-living counterparts. These include lymphoplasmacytic gastritis, glomerulosclerosis, renal amyloidosis, veno-occlusive disease of the liver, adrenal hyperplasia and several ill-defined neurological disorders. Many factors have been proposed as possibly being the cause of this phenomenon – factors such as stress, lack of exercise, low genetic variability and the provision of unnatural diets in captive facilities – but to date convincing pathophysiological explanations for these diseases have been lacking or unsatisfactory. Doctor Adrian Tordiffe investigated this problem using a systems biology approach – in other words, he attempted to understand as many of the components of the cheeatah’s metabolic system as possible. He did this by using untargeted metabolomic analysis of serum and urine from captive and free-ranging cheetahs, thereby generating new physiological data for this species in the hope of developing a better understanding of their metabolism.
Before the actual quantification of serum or urine metabolites, he created a new, more objective, method of approximating the body condition in cheetahs by developing a body mass index value, similar in concept to the same index used in humans. Unsurprisingly the body mass indices obtained in the study population showed significant differences between males and females, but importantly, all the animals fell within a healthy body mass index range. The impact of body condition on various serum and urine metabolites could thus be objectively assessed. He also evaluated the use of either urine creatinine concentrations or urine specific gravity values for the correction of spot urine samples obtained from the cheetahs. Creatinine is used in other species (including humans) to correct for differences in urine concentration between different study subjects because of differing levels of hydration. However in cheetahs, Adrian as able to show that there are other factors which affect creatinine concentration and therefore this method is unreliable in this species. He, instead, found that measuring the specific gravity of the urine in each sample provided a better indication of urine dilution. Urine specific gravity was therefore used to correct urine metabolite concentrations in this study.
Using gas chromatography-mass spectrometry, 339 different organic acids were identified and quantified in the urine of 56 captive and two free-ranging cheetahs. Phenolic compounds, which are thought to be produced by the anaerobic fermentation of aromatic amino acids by bacteria in the colon, as well as their corresponding glycine conjugates, were present in high concentrations in the urine of the captive cheetahs. Because detoxification of these phenolic compounds occurs through glycine conjugation, this could result in the chronic depletion of both glycine and sequestration of Coenzyme A, with associated negative metabolic consequences. Adrian has suggested that the high urine levels of these phenolic compounds may be caused by an excess in dietary protein, as most captive cheetahs are fed a diet rich in muscle meat and low in fat and other so-called animal fibre – for example skin, bone and cartilage.
Concentrations of these phenolic compounds also correlated negatively with the end-stage metabolites of dopamine and catecholamines – in other words, in cheetahs where the concetrations of these phenolic compounds were there tended to be lower concentrations of dopamine and catecholamines. These are all important neurotransmitters. This could potentially, thus, also provide an explanation for some of the neurological disorders seen in the captive cheetahs. Dopamine, in addition to being a neurotransmitter also plays important roles in the gastrointestinal system and the kidneys, and so dopamine depletion may also be playing a central role in the pathophysiology of both the gastric and renal diseases prevalent in captive cheetahs.
Using gas chromatography-mass spectrometry as well as liquid chromatography-tandem mass spectrometry Adrian established serum and urine amino acid profiles in captive cheetahs. Although the serum concentrations of most of the amino acids in cheetahs were comparable to those in published data for domestic cats, the serum arginine and ornithine concentrations were substantially higher.
Finally, the serum fatty acid and acylcarnitine profiles of 35 captive and 43 free-ranging cheetahs were evaluated through the use of gas chromatography-mass spectrometry and liquid chromatography-tandem mass spectrometry. The profiles obtained from the free-ranging animals provide a unique, healthy control group for comparison. Significant differences were noted for most of the fatty acid and acylcarnitine concentrations between these two populations, indicating dramatic differences in the dietary fat intake, composition and/or metabolism of these nutrients. Most of the serum polyunsaturated fatty acid and mono-unsaturated fatty acid concentrations were significantly lower in the free-ranging cheetahs, compared to the captive animals, suggesting that the fatty acids in the wild cheetah diet are largely saturated. Fatty acids not only provide a valuable source of energy, but also perform other vital functions in the body, including hormone production, cellular signalling and the provision of structural components of biological membranes. Altered serum fatty acids could thus have a dramatic impact on health and, since their concentrations are largely influenced by diet, the values obtained from free-ranging cheetahs potentially provide valuable healthy target values for their captive counterparts.
Through this unique approach, Adrian has established new baseline data for a large range of serum and urine metabolites in cheetahs. The results raise many questions and provide valuable new insights and hypotheses into the potential mechanisms of metabolic disorders in captive cheetahs, creating a platform for future research in this species.