The Metabolomics of Biological Stress

Stress, as with most biological phenomena, must be maintained in a delicate balance. A little bit of stress is a good thing—it’s what drives adaptation and enables organisms to respond appropriately to threatening situations or to heal in response to injury. But too much stress for too long is a critical contributor to disease.  

The biological stress response is an organism’s response to internal and external stressors, which can include injury, infection, toxic exposure, or normal physiological functions such as eating. Appropriate responses to these stressors are mediated by tightly controlled cell signaling and involve complex interconnections between the nervous, endocrine, immune, and circulatory systems^1,2. 

Metabolites play a central role in biological stress and the body’s response to stressors, and stressors often induce large-scale metabolic changes³. Blood and salivary tests, brain imaging, behavior tests, gene expression analysis, and DNA methylation sequencing have all been used to examine biological stress, though these approaches are limited to only a small subset of metabolites or specific biochemical pathways.  

With the increasing popularity of ‘omics techniques, metabolomics is emerging as a tool that can yield more sensitive detection and quantification of a wide array of metabolites and pathways associated with and impacted by biological stress. These metabolites, and the pathways they comprise, may serve as biomarkers of disease states or represent therapeutic targets for a variety of conditions³. 

Generally, biological stress-related metabolites can be grouped into seven classes, depending on the physiological system or response they belong to/participate in. 

Stress Signaling 

When exposed to a stressor or set of stressors, the body’s job is to return the body to a state of homeostasis as quickly as possible. To achieve this, the acute stress response causes the release of stress hormones (catecholamines such as epinephrine and glucocorticoids such as cortisol) produced by the sympathetic nervous system (SNS) and the hypothalamic-pituitary adrenocortical (HPA) axis^1,2. Their release makes energy immediately available and redirects that energy to tissues that become more active during stress, raising blood pressure. Immune cells are also activated and, when present, recruited to areas of injury or infection.  

Repeated or continuous activation of the acute stress response can lead to serious cardiovascular damage and can suppress immunity (discussed further below)^1,2. Additionally, elevated stress hormones and chronic activation of the HPA axis are linked to mental health and mood disorders^4,5, and the impact of cortisol on neurotransmitter function (e.g., serotonin, 5HIAA, etc.) is well-recognized⁴.  

Several metabolomics studies in animal models and in humans have identified a variety of metabolites associated with HPA axis activation and psychoneurological system diseases, including cortisol, hydroxyhemopyrrolin-2-one (HPL), and pyrrole, as well as disturbances in several metabolic pathways including glycolysis/gluconeogenesis^6,7. Others have reported significant changes in several metabolites and pathways, including amino acid biosynthesis, protein translation, and glycolysis pathways, in response to both acute and chronic stress⁸. 

Inflammation and Immunity 

The biological stress response significantly impacts immune system function, and perturbations caused by injury or infection are themselves sources of stress that can lead to additional immune system involvement. When the stressor is injury or infection, cytokine release attracts immune cells to the site of infection or injury, activating the HPA axis and the acute stress response and leading to the release of the small molecules discussed above⁹. These events have significant impacts on metabolic processes.  

Studies have begun to unravel how acute and chronic inflammation differentially impact metabolites and to distinguish between local and systemic metabolic consequences to identify biomarkers and potential therapeutic targets for several diseases¹⁰.  

Lipid mediators such as arachidonic acid, amino acid mediators such as alanine, leucine, isoleucine, lysine, and valine, and short chain fatty acids such as butyrate and acetate have been identified as key players in the immune system’s response to biological stress and as biomarkers of disease states ranging from neurological to gut disorders¹⁰. Other molecules, including creatinine, citrate, succinate, lactate, and fructose have also been identified. Additionally, adenosine and itaconate have been identified as anti-inflammatory metabolites that also impact the pathogenesis of infection¹¹. 

Injury and Circulation 

As discussed briefly above, the biological stress response to injury has a direct impact on the cardiovascular system, raising blood pressure and selectively dilating or constricting blood vessels. When the injury occurs to the vasculature itself, a complex and elegant cellular response ensures wounds are closed quickly to prevent excessive blood loss and maintain tissue-level boundaries.  

Multiple cells, including platelets, are involved both in wound healing and in the inflammatory response to injury. Compromised vasculature not only directly causes an inflammatory response but can also slow the delivery of molecules involved in wound healing to areas of injury in other bodily locations. 

Metabolomics studies have revealed a rich network of metabolites that promote, are impacted by, or hinder wound healing and vascular injury. Alterations in arginine metabolism, the kynurenine pathway, and several metabolites including lactate, citrate, itaconate, and others have been reported^12-14.  

Oxidative Stress 

Reactive oxygen species (ROS) result from both injury/infection and from the normal process of energy production by cells. Therefore, the body must maintain a delicate balance between ROS and ROS scavengers to ensure that oxidative stress—the body’s inability to detoxify ROS, resulting in oxidative injury to cells—is kept in check. Oxidative stress has been associated with a range of health conditions, including various cancers, and is a key player in aging. Understanding the metabolites involved in controlling oxidative stress has the potential to aid our understanding of and treatment strategies for a large array of diseases. 

One of the most significant sources of ROS is normal energy production by the mitochondria, and many diseases associated with metabolic dysfunction—particularly obesity and diabetes comorbidities—have been associated with an altered balance between ROS production and ROS scavenging¹⁵. This relationship seems at least partly due to the ability of glucose metabolism to produce ROS via sorbitol metabolism, hexosamine metabolism, α-ketoaldehyde production, PKC activation, glycation, and oxidative phosphorylation¹⁵, discussed more in detail below.  

More generally, markers of platelet apoptosis, lipid, DNA, and protein modifications, and end products of lipid oxidation (most often malondialdehyde (MDA) and 4-hydroxy-2-nonenal (HNE)) are reliable markers of oxidative stress^15,16. Several markers of DNA and protein modifications have also been identified, including xanthine, pentosidine, and carboxymethyl lysine, among many others¹⁵. 

Metabolism and Glycemic Stress 

As mentioned above, glucose metabolism can contribute to ROS production via an array of metabolic pathways. Persistently elevated blood glucose levels lead to non-enzymatic protein glycation and oxidative degeneration, both of which can be measured via the presence of several of the protein modifications discussed above¹⁶. Additionally, alterations in several metabolites, including choline, fructose-glucose-galactose, kynurenic acid, and several others have been associated with glycemic index, glycemic load, and insulinemic responses to food¹⁷.  

Other research has also shown that several metabolites including amino acid metabolites, tricholoracetic acid, and free fatty acids and metabolic pathways including glycolysis, gluconeogenesis, bile acids, and vitamin D metabolism are altered in type 1 diabetes¹⁸, which, like type 2 diabetes, is characterized by blood glucose swings that must be carefully controlled to avoid the development of other conditions.  

Protein Catabolism and Decay 

Skeletal muscle atrophy (i.e., wasting) is a natural part of aging, but it can also result from various stressors including diseases, poor nutrition, or lack of muscle utilization. Although muscle wasting can be a short-term, reversible stress response, it can also become a more serious chronic condition when it results in non-reversible weight loss (cachexia). 

Skeletal muscle plays a critical role in several metabolic functions including energy homeostasis, insulin sensitivity, and amino acid metabolism¹⁹. It also acts as a reserve for glucose and amino acids for other tissues and organs, even providing compensatory metabolic function to failing organs¹⁹. Therefore, many studies have sought to characterize the metabolic consequences of muscle wasting associated with aging, cancer, and other diseases. Dysregulated amino acid metabolism is a hallmark of muscle wasting, with alterations in both essential amino acids and branched-chain amino acids observed¹⁹, with alterations in several amino acids/amino acid metabolites observed in muscle wasting patients suffering from chronic diseases^20,21. 

Asymmetric dimethyl arginine (ADMA), a metabolic byproduct of protein modification closely related to L-arginine, has been shown to be increased both with decreased muscle strength generally and, in animal models, in cancer-associated cachexia^21,22. Altered lipid metabolism and dysregulated energy metabolism are also observed in muscle wasting¹⁹, with signatures of altered mitochondrial function often observed in muscle wasting.  

Vitamins 

Vitamins are essential for human health and well-being, as they act as co-factors, antioxidants, and even signaling molecules. Vitamins are mostly obtained through diet. If dietary habits do not provide sufficient nutrition, vitamin deficiencies can occur, resulting in several negative health effects, including a compromised immune system²³ and increased susceptibility to environmental stressors.  

Vitamins can impact other metabolic processes, such as amino acid, carbohydrate, organic acid, and one-carbon metabolism²⁴. Several studies have utilized metabolomics to explore the metabolic consequences of deficiencies in or supplementation with several vitamins, including B6, A, E, and D, with alterations observed in several metabolites including acetate, pyruvate, trimethylamine-N-oxide, lysophosphatidylcholine, and others^24-28. 

Conclusions 

The biological stress response is a critical response required to maintain homeostasis in the face of a variety of stressors, both environmental and internal. Biological stress impacts every system in the body, and the effects of biological stress on metabolic pathways are central to the physiological impacts experienced by the body. Metabolon’s Biological Stress Discovery Panel was designed to detect the most relevant metabolites and pathways associated with and altered during metabolic stress. This panel can prove a critical addition to any research efforts in biological stress or any disease state influenced by biological stress, from cancer to infection to aging. Contact us today to learn more about our Biological Stress Discovery Panel and how you can use it to accelerate your own research. 

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