The impact of sub-thermoneutral housing temperature on models of cancer and allogeneic hematopoietic cell transplant in laboratory mice
Kokolus, Kathleen Marie
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Laboratory mice are commonly used to model human diseases and disorders. They are intended to mimic the biological and physiological processes that occur in humans and are used not only to study disease progression but also to test the efficacy of new treatments and immunotherapies. However, often data obtained from mouse models does not accurately predict what occurs in humans thereby threatening the ability to translate pre-clinical findings into the clinic. The research in this thesis reveals how a reversible environmental housing condition has been an underappreciated factor that can significantly affect immune function in two separate mouse models of human disease: cancer and graft versus host disease. At ambient temperatures of 30-31°C, which is considered the thermoneutral temperature (TT) for mice, resting metabolic rate is sufficient to generate enough heat to maintain body temperature. However, mice in laboratory animal facilities are kept at relatively cool standard temperatures (ST; 20-26°C) as mandated by the National Research Council. The data contained in this thesis strongly demonstrates "heat-seeking" behavior in tumor-bearing mice housed at ST. This indicates the presence of a physiological drive for additional warmth, likely needed to lessen the energy burden of maintaining thermal homeostasis. The cool ambient temperature at which laboratory mice are housed elicits a stress, which is driven by activation of the sympathetic nervous system and subsequent norepinephrine release. Scientific literature is full of work demonstrating that cool ambient temperatures are associated with a weakened immune response. However, until now no one has looked at the effects of ambient on mouse models of cancer or allogeneic hematopoietic cell transplant. In this thesis, we will provide evidence of negative effects of cool housing temperatures on the immune response in these two models, both of which involve cytotoxic T cell activation. We compared tumor growth in mice housed at ST and TT and found significantly reduced tumor incidence and suppressed tumor growth rates in animals housed at TT versus ST. Because this difference was not seen in SCID or NUDE mice as well as in mice depleted for CD8 + T cells, a prominent role of the adaptive immune response was suggested. Further analysis revealed more CD8 + T cells within the tumor microenvironment animals housed at TT compared to those housed at ST. Additionally, using a MHC pentamer loaded with CT26 tumor antigen revealed an increase in antigen-specific CD8 + T cells in the lymph nodes and tumors of mice maintained at TT versus ST. We also show that the CD8 + which are increased in the tumor microenvironment of mice at TT compared to ST show a more activated phenotype based on surface marker expression of various activation markers. We also examined the frequency of dendritic cells (DCs) in the spleens of mice maintained at ST and TT and found that mice at ST have relatively fewer mature DCs. Our data has also revealed an increased infiltration of immunosuppressive cells including Gr-1 + CD11b + and Foxp3 + cells. In addition to models of cancer, we also examined mouse models of graft versus host disease (GVHD), a disease which results from host T cell function. Since cold stressed mice exhibit weakened T cell dependent immune responses, we hypothesize that baseline GVHD would be reduced in mice at ST compared to that seen in mice at TT. We found that following allogeneic hematopoietic cell transplant (allo-HCT) mice housed at ST show no signs of GVHD; however, mice at TT displayed significant weight loss and lethal GVHD. Interestingly, the reduced GVHD seen at ST was found to be mediated through β-adrenergic receptors which become activated by signals from the sympathetic nervous system in order to generate additional heat to support body temperature when normal mice are cold-stressed. These studies provide insight into translational discrepancies of clinical and murine allo-HCT and demonstrate lethal GVHD in a mouse model generated by un-modified bone marrow. The full implications of chronic cold stress on either the anti-tumor immune response or the host response to allogeneic hematopoietic cell transplantation in mice are as yet unknown, but these data strongly suggest the existence of a fundamental relationship between cold stress and impaired CD8 + T cell-dependent immunity. Throughout this thesis we will propose various potential mechanisms and pathways through which cold stress induced by ambient temperature may be related to the immune response. For example, we will discuss possible alterations in the balance of immune promoting (i.e. activated T cells) versus immunosuppressive (i.e myeloid derived suppressor cells) cells as well as the balance of pro- and anti- inflammatory cytokine expression. We will also suggest the potential of the β-adrenergic pathway (which is activated when mice are under cold stress) mediating some of the immunological endpoints effected by ambient temperature.