An assessment of cortisol analysis in hair and its clinical applications

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Abstract

Hair analyses for exogenous compounds, specifically drugs of abuse, have been a useful tool in detecting long-term drug exposure. More recently, studies have delved into the exposure of endogenous compounds in hair.

Cortisol is synthesized in the adrenal cortex in response to stress-induced activation of the hypothalamic–pituitary–adrenal (HPA) axis. While catecholamines generally indicate acute stress, cortisol can be used as an indicator for sub-acute and chronic stress.

Studies on the effects of chronic stress are most often subjective in nature, relying on questionnaires asking the participant to recall on past stressors. This can lead to the issue of recall and reporting bias. A new objective measure of chronic stress is needed for a more accurate understanding of the effects of chronic stress on the body. This review uses emerging evidence to describe the usefulness of hair analysis for cortisol and discusses the current methods used.

Introduction

The use of hair analysis has become an increasingly widespread tool to allow for long-term measure of drug exposure, both exogenous and endogenous. The aim of this review is to discuss the studies on the use of hair analysis for the endogenously produced steroid hormone, cortisol. More specifically, the authors will describe the current methods used for measuring cortisol in hair and report research findings on the topic of hair cortisol as a biomarker of chronic stress. In conducting this review, a literature search was performed to include all studies on hair cortisol and its use to assess chronic stress. As hair cortisol has been investigated in more than one area of the body, it is important to note that the studies described in this review are limited to those using hair samples obtained from the scalp.

The interest for measuring substances in hair started with the recognition that glucocorticoids (and other steroids) are often abused by athletes to improve their performance. Since the 1960s, testing for such compounds has been performed in the blood and urine of athletes [1], [2]. However, these matrices offer only short-term information, allowing users to discontinue the compound only a few days prior to testing or the competition. Therefore, the issue of false negative test results remains a problem in doping control and other toxicological screening tests.

The ability to detect a drug in hair long after the drug use or exposure occurs has been proven to be very beneficial. Hair may provide a long-term “memory” that documents chronic substance abuse and can provide a positive result in cases where blood or urine tests may have been falsely negative. In 2000 Gaillard et al. reported several drugs of abuse (including steroids) in the hair and urine of cyclists. Several corticosteroids were detected in both urine (12 out of 30 cases) and hair (5 out of 12 cases) using a high performance-liquid chromatography–mass spectrometry (HPLC/MS/MS) method [1]. The rate of positive corticosteroid results was 40% (in urine) and 42% (in hair), and combination of the two matrices resulted in a complementary positive rate of 47%. Similarly, a second study by Bévalot et al. used liquid chromatography–electrospray ionization-mass spectrometry to detect corticosteroids in the hair of athletes [2]. While using a slightly different method, similar findings were obtained. Although urinalysis is typically seen as the method of choice, these studies demonstrate the potential usefulness of hair analysis for long-term corticosteroid abuse in athletes and suggest these two methods be used to compliment each other [1], [2]. Many studies have looked at the use of hair analysis to determine exposure to exogenous drugs within the clinical and forensic toxicological context. For a more detailed discussion we refer to the review by Kintz et al. (Ther. Drug Monit. 2006, 28:442–446).

The study of glucocorticoid hormones in hair began in 2000 with Cirimele et al. who investigated ten corticosteroids in human hair with the use of high-performance liquid chromatography–ionspray mass spectrometry [3]. Using both forensic and clinical cases, this group identified prednisone and beclomethasone (both exogenous corticosteroids) in human hair [3]. One of the first studies that looked at measuring endogenously produced steroid hormones in hair was that of Yang et al. [4]. Estradiol, progesterone and testosterone were detected in human hair using the radio-immunoassay method, and the changes in hair levels were found to be similar as the changes in blood [4].

Hair analysis for endogenously produced cortisol in humans was first documented by Raul et al. [5]. Using liquid chromatography–mass spectrometry (LC/MS), these investigators measured both cortisol and its metabolite cortisone in the hair of 44 individuals. They found cortisol and cortisone concentrations ranging from 5 to 91 pg/mg and 12 to 163 pg/mg, respectively [5]. The researchers suggest that one of the mechanisms involved in compound incorporation into hair is through passive diffusion through sweat [5]. They also suggested that the increase in cortisone:cortisol ratio found in hair could be the result of increased conversion of cortisol to cortisone by 11-beta hydroxysteroid dehydrogenase type 2 enzyme present in the sweat glands prior to deposition on the hair [5]. Raul et al. did not find any effect of hair colour or gender on the measured cortisol or cortisone content.

The benefits of hair analysis are numerous. The ability to detect past drug exposure is a unique feature of this matrix, as it provides researchers with a “window to the past.” Assuming hair grows approximately 1 cm per month [6], analysis of hair may document a historical timeline of drug exposure. Secondly, hair sampling is non-invasive and painless compared to traditional blood sample collection. The procedure is quite simple and the sample can be taken by a non-professional. Once taken, the sample can be stored at room temperature and sent through the mail. Hair analysis has emerged as a beneficial tool in forensic toxicology, environmental toxicology and in many research laboratories.

The anatomy of a growing hair strand is quite complex. The hair shaft is composed of the outer cuticle, cortex and inner medulla [7]. The hair follicle lies 3–4 mm below the surface of the skin and is closely associated with arterial capillaries, sebaceous glands and apocrine sweat glands [7], [8], [9]. These glands empty their sebum and sweat, respectively, into the hair follicle whereas eccrine sweat glands discharge their products onto the skin surface [8], [9].

While the mechanism of incorporation of substances in hair has not been clearly defined, there are several potential mechanisms that are generally accepted in the scientific community. Internally, drugs have been proposed to enter the hair via the blood capillaries during formation, as well as through sebum and sweat secretions into the hair follicle after formation (Fig. 1) [8], [9]. Passive (or active) diffusion from blood to the hair follicle is thought to be the primary mechanism [9].

Externally, drugs can deposit onto hair from the environment via smoke, pollution or physical contact, chemicals, etc. (Fig. 1) [8], [9]. For example, secondary smoke can be detected in hair leading to false positive results [8], [9]. Regarding cortisol, external contamination can potentially occur through the use of cortisol-containing creams, ointments, etc. that can reside on the hands and subsequently become bound to the outside of the hair shaft [10], [25], for example, when an individual scratches his/her head. Additionally, sweat and sebum can coat the outside of the hair shaft as it emerges from the scalp (Fig. 1) [8], [9].

Section snippets

Hair sampling

Hair samples are taken from the vertex posterior of the head and cut (not pulled out) with clean scissors as close to the scalp as possible. It is important that the hair follicle itself is not included in the analysis, as a previous study found that hair follicles are capable of producing cortisol in response to corticotropin-releasing hormone stimulation [11] and thus may skew the results. Additionally, prior studies have shown that this area of the scalp has the lowest coefficient of

Cortisol

Cortisol is a steroid hormone that is produced in the body in response to stress. Upon activation by an emotional or physical stressor, corticotropic releasing hormone (CRH) is released from the paraventricular cells of the hypothalamus and exerts its effect on the anterior pituitary gland [19], [20]. Here, CRH stimulates the synthesis of adrenocorticotropic hormone (ACTH) and its release into the bloodstream [19], [20]. In turn, ACTH acts on receptors of the adrenal cortex to stimulate the

Considerations

While the analysis of cortisol in hair is an exciting and informative tool to assess chronic systemic cortisol exposure, one must be aware of its potential limitations. Many different factors could influence the results of hair testing. These may include hair growth rates, gender, age, hair colour, environmental exposures and others. Further studies are needed to determine if and to what extent these factors affect hair cortisol concentrations.

For example, in regards to gender variability,

Conclusions

The measurement of cortisol in hair has the potential to serve as a biomarker of chronic stress in a variety of health conditions. In initial studies, hair cortisol analysis has been shown to be quite promising. As it provides us with long-term, retrospective data and is non-invasive in nature, hair constitutes a unique matrix with a great potential for research applications.

Future studies will expand the use of this technique to a variety of stressed populations, in the hopes of increasing our

Conflict of interest

The authors have no conflict of interest to disclose.

Acknowledgments

The authors would like to acknowledge their funding from the Physicians’ Services Incorporated Foundation and the Canadian Institute for Health Research. They had no involvement in the research study itself.

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