Elsevier

Research in Veterinary Science

Volume 138, September 2021, Pages 161-166
Research in Veterinary Science

Original Article
Metabolic profiling of serum from dogs with pituitary-dependent hyperadrenocorticism

https://doi.org/10.1016/j.rvsc.2021.06.011Get rights and content

Highlights

  • Pituitary-dependent hyperadrenocorticism (PDH) alters serum metabolome in dogs.

  • Metabolites more affected were fatty acids, amino acids and glycerophospholipids.

  • Metabolite changes could be related to the physiopathology of this disease.

Abstract

Hyperadrenocorticism (HAC) is one of the most common endocrine diseases in dogs characterized by excessive cortisol production caused by an adrenocorticotropic hormone (ACTH)-secreting tumor, namely pituitary-dependent HAC (PDH) or cortisol-secreting adrenal tumor. Metabolomics presents the ability to identify small molecule metabolites. Thus, the use of metabolomics techniques in canine PDH can provide information about the pathophysiology and metabolic changes in this disease. This study aimed to identify and compare differences in serum metabolites between dogs with PDH and healthy dogs. The metabolomic profile of 20 dogs diagnosed with PDH was compared with 20 healthy dogs using liquid chromatography/mass spectrometry (LC/MS), and metabolite discrimination was performed using partial least squares-discriminant analysis (PLS-DA), the variable important in projection (VIP) and fold changes (FC) group-wise comparisons. The hypergeometric test identified the significantly altered pathways. A total of 21 metabolites were found to be significantly different between the two groups. The major alterations were found in arachidonic and decanoic acid, and phospholipids related to phosphatidylcholine (PC), phosphatidylethanolamine (PE) and phosphatidylinositol (PI). These metabolites are related to insulin resistance and other complications (i.e. hypertension). Our results indicate that PDH produces changes in serum metabolites of dogs, and the knowledge of these changes can aid to better understanding of pathophysiological processes involved and contribute to potentially detect new biomarkers for this disease.

Introduction

Hyperadrenocorticism (HAC) is one of the most common endocrine diseases in dogs, with an estimated prevalence of 0.27% to 1.46% (Carotenuto et al., 2019; O'Neill et al., 2016). HAC can be caused by an excessive cortisol production due to an adrenocorticotropic hormone (ACTH)-secreting pituitary tumor, namely pituitary-dependent HAC (PDH), or by a cortisol-secreting tumor arising in the adrenal cortex (Pérez-Alenza and Melián, 2017). The most common clinical signs consistently reported in both HAC types are polyuria, polydipsia and polyphagia, abdominal enlargement, and hepatomegaly (Ling et al., 1979; Reusch and Feldman, 1991).

Untargeted metabolomics allows a rapid and global description of many metabolites in a single sample, being of high interest to detect differences between physiological and pathological conditions (Emwas, 2015; Nicholson and Lindon, 2008). The number of studies carried out in veterinary medicine is small compared to humans (Carlos et al., 2020). However, the untargeted metabolomic analysis allowed the detection of metabolites that vary in dogs with diabetes compared to healthy animals (O'Kell et al., 2019). While, to the author's knowledge, metabolomics has not been applied to HAC in dogs. The analysis of the metabolic processes that occur in the serum can potentially help to understand the pathophysiological processes involved in a disease and it can also help to discover new biomarkers (Carlos et al., 2020). The importance of using reliable biomarkers in HAC patients (Hasenmajer et al., 2020; Ragnarsson, 2020) has been documented in humans and dogs (Jaffey et al., 2020). This study aimed to identify and compare differences in serum metabolites between dogs with PDH and healthy dogs.

Section snippets

Study Design and Case Selection

The individual characteristics of dogs enrolled in this study are presented in the supplementary information (Tables S1 and S2). These samples corresponded to two groups of dogs; one group of dogs diagnosed with PDH and one group of healthy dogs. Samples were collected between December 2018 and January 2020 and were stored at −80 °C until the analysis. In the case of dogs with PDH, the inclusion criteria were: (1) not presenting other pathology, (2) not having received treatment before

Metabolomics Analysis of Dogs With Pituitary-Dependent Hyperadrenocorticism and Healthy Dogs

A total of 314 different compounds were found in both groups. The different serum fingerprints between PDH and healthy dogs were shown in the PLS-DA model (R2 = 99%, Q2 = 41%, Fig. 1). Considering the screening of identified metabolites under the conditions of VIP > 1 and FC > 1.5 (Fig. 2), a total of 21 discriminative metabolites were found. Among them, two were identified, and 19 were recognized based on the Fragment Annotation Kit (FrAnK). This Django-based application implements Celery to

Discussion

This work describes alterations in serum metabolome that occur in dogs with PDH by applying metabolomics using a LC/MS analytical platform. A total of 21 metabolites showed significant alterations; 10 being downregulated and 11 being upregulated in the PDH group. The changes of higher magnitude were found in the fatty acids decanoic and arachidonic acid, and in the amino acids tryptophan and l-glutamine. In addition, alterations in phospholipids such as phosphatidylcholine (PC),

Conclusions

In conclusion, dogs with PDH showed alterations in the concentration of various metabolites compared to healthy dogs when serum was analyzed by LC/MS. The major variations were associated to arachidonic acid metabolism with the decrease of arachidonic acid and to glycerophospholipids metabolism with the decrease of PC(24:1(15Z)/14:1(9Z) and PI(18:0–20:4), and the increase of PE(20:0/16:1(9Z)). These metabolites could be related to insulin resistance and other complications (i.e. hypertension)

Acknowledgements

This research was supported by the European Structural and Investment Funds (Grant Agreement KK.01.1.16.0004). A.M.-P. was funded by Seneca Foundation through a post-doctoral grant (21239/PD/19). A.T. has a post-doctoral fellowships Ramón y Cajal (RYC-2017-22992) supported by the Ministerio de Economía y Competitividad, Agencia Estatal de Investigación (AEI), Spain, and The European Social Fund (ESF).er separation can be observed with the peak

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