Circadian Immune Measures in Healthy Volunteers: Relationship to Hypothalamic-Pituitary-Adrenal Axis Hormones and Sympathetic Neurotransmitters ZIAD KRONFOL, MD, MADHAVAN NAIR, PHD, QUN ZHANG, PHD, ELIZABETH E. HILL, PHD, AND MORTON B. BROWN, PHD Objective: The purpose of this study is to examine the circadian pattern of specific immunologic measures and to compare observed circadian rhythms of these measures with the well-established circadian rhythms of hypothalamic-pituitary-adrenal axis hormones and sympathetic neurotransmitters. Methods: Blood samples were collected every 2 hours for a total of 24 hours from nine healthy volunteers. The blood samples were assayed for hormones and immune measures, including adrenocorticotropic hormone (ACTH), cortisol, norepinephrine, and epinephrine. The immune measures included percentage and absolute number of neutrophils, lymphocytes, the lymphocyte subsets CD3+ (T cells), CD4+ (T helper/inducer), CD8+ (T suppressor/cytotoxic), CD56+ (natural killer [NK] cells) and NK cell activity (NKCA). Results: The following immune measures exhibited a significant circadian rhythm: the percentages of neutrophils, CD4+ cells, and CD56+ cells; the absolute numbers of total lymphocytes, CD3+ cells, CD4+ cells, and CD8+ cells; and NKCA. Cross-correlations between the circadian rhythms of selected hormones and immune measures indicated a strong inverse association between the circadian rhythms of cortisol and the different T cell subsets on the one hand, and a strong direct association between the rhythms of cortisol and the percentage of CD56+ and NKCA on the other. Cross-correlations involving the circadian rhythms of norepinephrine and the same immune measures were in general much weaker and statistically nonsignificant. Conclusion: In healthy individuals, both enumerative and functional immune measures exhibit circadian rhythms that seem to be associated most closely with the circadian rhythm of cortisol. Key words: circadian, cortisol, norepinephrine, T cells, NK cells, psychoneuroimmunology. INTRODUCTION It is now well established that the central nervous system and the immune system are closely interre- lated (1). Although mediators of such interactions have not always been identified with certainty, the hypothalamic-pituitary-adrenal (HPA) axis (2-4) and the sympathetic nervous system (SNS) (5-7) have often been assumed to play key roles. Studies have shown that glucocorticoids (8) and cat- echolamines (9), when added in vitro to cultures of human lymphocytes, were associated with inhibi- tion of various immunoregulatory activities. When the same products were assessed in vivo, the results varied. Corticosteroid administration resulted in an increase in natural killer (NK) cell activity (10), From the Departments of Psychiatry (Z.K., Q.Z., E.E.H.) and Biostatistics (M.B.B.), University of Michigan Medical Center, Ann Arbor, Michigan; and Department of Medicine, State Univer- sity of New York at Buffalo, New York (M.N.). Address reprint requests to: Ziad Kronfol, MD, Department of Psychiatry, University of Michigan Medical Center, 1500 E. Med- ical Center Drive, Ann Arbor, MI 48109-0722. Received for publication March 29, 1995; revision received February 12, 1996. whereas epinephrine injections were accompanied by a decrease in mitogen-induced lymphocyte pro- liferation (11). Similarly, there were conflicting re- sults regarding the effects of corticosteroids and catecholamines on circulating lymphocyte subsets (12,13). Part of the discrepancy may be related to the dose of hormone given, the route of administration, the duration of treatment, and the specific com- pound being used (ie, natural or synthetic). Al- though these studies of "pharmacologic" interven- tion have helped advance our knowledge in this area, they provided little information on day-to-day interactions between HPA-axis hormones, cat- echolamines, and specific immune measures under "physiologic" conditions. One research strategy to address this issue has been the study of circadian rhythmicity. HPA-axis hormones such as cortisol and adrenocorticotropic hormone (ACTH) have well-established circadian rhythms (14, 15). Similarly, although perhaps not as widely known, plasma catecholamines show circa- dian fluctuations (16, 17). Several studies have therefore examined the circadian rhythmicity of spe- cific immune measures in relation to HPA hor- mones, especially cortisol. Ritchie et al. (18), for instance, were able to document a circadian rhythm 42 0033-3174/97/5901 -0042S03.00/0 Copyright © 1997 by the American Psychosomatic Society Psychosomatic Medicine 59:42-50 (1997)