Research Projects

Overview: Studies conducted by our group and others have shown that even moderate exposures to radiation (up to several Gy) can have significant effects on immune responses, including cell-mediated cytotoxicity, mitogen-induced and spontaneous blastogenesis, cytokine expression, hematopoiesis, hematological parameters, and leukocyte phenotype distribution patterns. This has been demonstrated across a wide range of radiation types, including gamma-rays, protons, iron, carbon and silicon ions. However, there is little in the literature addressing the functional consequences of these changes. We propose that clinically and spaceflight relevant doses of radiation will have a detrimental impact on the ability of the immune system to respond to an in vivo immune challenge. Our goal is to determine the functional level at which irradiation impacts peripheral innate immunity utilizing a mouse model. Our endpoints include those involving both immunological and behavioral responses.

We have four main research priorities:

Innate Immunity & Phagocyte Function: As in vivo immune responses involve complex interactions between a variety of cell types (e.g. phagocytes, lymphocytes, etc), inter- and intra-cellular communication factors (e.g. cytokines, chemokines, "danger signals," etc.), and hormones (e.g. catecholamines, corticosterone, etc), an all-encompassing characterization of the effects of radiation on this system is somewhat daunting. Therefore, we limit the scope of this work to mechanisms that directly involve phagocytic populations, including both macrophages and neutrophils. For our disease model, we selected a well-characterized immune challenge (E. coli) known to be dominated by a robust phagocytic response. Functional assessment include phagocytosis and oxidative burst, surface receptor expression, and communication with accessory cell populations.

The response to a bacterial infection can be broken down into a series of steps ranging from pathogen recognition and phagocytosis to superoxide release and cytokine expression. These mechanisms are non-specific, involving complement-mediated lysis and destruction by neutrophils and macrophages via degradative enzymes (e.g. lysozyme, acid hydrolases, neutral proteases) and reactive oxygen and nitrogen intermediates (e.g. superoxide, hydrogen peroxide, hydroxyl radical, and nitric oxide (NO)). Once activated, phagocytes further enhance the inflammatory response via arachidonic acid metabolites and regulatory cytokines, including TNF-alpha, IL-1beta, IL-6, IL-8 and colony-stimulating factors. These cytokines, in turn, attract additional leukocyte populations to the site of infection, as well as activate a number of pathways, including the sympathetic nervous system (SNS). This typically results in a cascade of well-documented behavioral and physiological responses (e.g. fever, sleep, loss of appetite).

We are particularly interested in three inflammatory cytokines: IL-1beta, IL-6, and TNF-alpha. All three cytokines are released by activated macrophages in response to a pathogenic challenge. In small localized amounts, these cytokines contribute to the immune response by regulating activity, enhancing phagocytosis, promoting coagulation, and stimulating growth and repair in damaged tissues. Additionally, all three have been linked to "sickness behavior" and, therefore, CNS-immune communication. For example, TNF-alpha is released in the spleen during endotoxemia and drains into the liver via the splenic vein before entering the circulation. Vagal stimulation inhibits splenic TNF-alpha and IL-1betasynthesis, likely through vagal afferent pathways. Similarly, inhibiting the ACTH or glucocorticoid responses can lead to an increase in circulating TNF-alpha expression during endotoxic shock.

CNS-Immune Communication: The two-way communication between the brain and peripheral immunity has come into vogue only within the last two decades. Early reports indicate that inflammation in peripheral tissues activates the hypothalamic-pituitary-adrenal axis. Subsequent research has shown that this communication is crucial for maintaining and/or resetting homeostasis, depending on disease or psychological state. Many of the behaviors associated with "sickness" (e.g. lethargy, fever, hyperalgesia) and depression (e.g. anxiety, sleep disorders, cognitive dysfunction) are mediated by the expression of inflammatory cytokines in the periphery. There are also several reports that suggest that peripheral and central cytokine expression can influence memory consolidation.

The hypothalamus-pituitary- adrenal (HPA) axis exerts most of its influence systemically through a variety of hormones. The first part of this axis, the hypothalamus, is located in the forebrain. Although the neuroendocrine cells in the hypothalamus release a multitude of hormones, corticotropin-releasing hormone (CRH) is of particular interest as its secretion leads to the release of adrenocorticotropic hormone (ACTH) from the anterior pituitary. ACTH, in turn, circulates through the bloodstream to the third and final part of this axis, the adrenal gland. The adrenal cortex responds to ACTH by releasing the glucocorticoid, corticosterone (or cortisol in humans), into the blood. Similar to catecholamines, corticosterone has been shown to modulate several immune functions with typically suppressive results. For traditional stress models, activation of the HPA axis can occur within minutes (or less) of the stimulus.

A second communication pathway is via the sympathetic nervous system (SNS). This functionally discrete unit of the autonomic nervous system regulated primarily by the hypothalamus, including the paraventricular nucleus (PVN), through a complex interaction between multiple brainstem nuclei. Through a two neuron chain, sympathetic information is conveyed to peripheral targets, including all primary and secondary lymphoid organs. For example, the splenic nerve consists almost exclusively of sympathetic nerve fibers. These fibers are typically noradrenergic and exert their influence by releasing the catecholamine, norepinephrine (NE), as their major neurotransmitter. Preganglionic sympathetic fibers also terminate in the adrenal medulla. However, unlike the postganglionic fibers that terminate in immune organs, the adrenal releases mostly epinephrine (along with some NE) into the circulation. Both of these catecholamines have been reported to modulate (typically, suppress) many immune parameters. Therefore, the SNS can influence peripheral immunity locally, by releasing NE from "hardwired," noradrenergic nerve terminals, and/or systemically, by releasing epinephrine from the adrenal medulla. Additionally, NE released from sympathetic nerves spills into circulation to augment the effects of epinephrine. Chronic activation of one or both of these pathways can lead to long-term immunosuppression and behavioral deficits.

Communication of peripheral immune events to the brain involves, among other factors, the secretion of inflammatory cytokines. Although cytokines are large proteins and are unlikely to passively cross the blood-brain barrier, there are several mechanisms which allow circulating cytokines to influence brain activity. These include crossing through the circumventricular organs, active transport across capillary endothelium, or binding to receptors on capillary endothelium to trigger a response in the brain. There are also situations where sickness behavior occurs despite the lack of measurable circulating cytokines, suggesting that there are other routes of communication.

One particularly important pathway that does not depend on circulating cytokine levels is local cytokine interactions with sensory nerves such as the vagus, consisting of afferent fibers that carry signals to the brain from several pathogen ports of entry (e.g. gut and lungs). Directly relevant to this proposal are reports that vagal fibers express mRNA for IL-1 receptors, and that subdiaphragmatic vagotomy can inhibit many sickness responses exhibited after peripheral treatment with lipopolysaccharide (LPS) or IL-1beta. Furthermore, vagal fibers in the gut are surrounded by dense populations of dendritic cells. These particularly potent antigen presenting cells constitutively express IL-1beta and their close proximity to vagus suggest a clear and immediate path of communication. There are also specialized sensory structures scattered throughout the gut, called paraganglia, which form synapses with the vagus. Paraganglia are similar to taste receptors, "sensing" cytokines and other immune products released by nearby tissues.