5-HTTLPR genotype significantly affected the ratio of IL-6 to IL-10 under resting conditions and after acute psychosocial stress, with SS individuals exhibiting a significantly higher ratio across timepoints. This inflammatory bias was present despite both groups being medically healthy and reporting similarly low exposure to early and recent life stress, which may be independently associated with inflammation (Cohen et al., 1999
; Danese et al., 2007
; Danese et al., 2008
). All participants reported strong subjective experiences of stress and negative emotionality immediately after the task, but these reports were also similar across genotype groups, suggesting that SS individuals do not exhibit a greater pro-inflammatory bias during the stress task simply because they perceive the task as more stressful. To our knowledge, this is the first report of a pro-inflammatory bias/phenotype in individuals carrying the SS genotype of 5-HTTLPR. Thus, even healthy SS-individuals may be chronically exposed to a pro-inflammatory burden under resting and stress conditions, which could increase their vulnerability to disorders like depression and other diseases that can be facilitated/exacerbated by a chronic pro-inflammatory state.
Given the current controversy over the relationships between 5-HTTLPR genotype, life stressors, and depression risk (Risch et al., 2009
), the potential implications of the findings described here are significant, and call for further investigation. The pro-inflammatory bias shown by SS individuals, both at baseline and during psychosocial stress, parallels the increased inflammatory markers seen under resting conditions in individuals with depression (Dantzer et al., 2008
; Capuron et al. 2008
). Importantly, this bias is shown by SS individuals in the absence of psychiatric, immune, or other disease conditions which suggests that the pro-inflammatory bias may precede (and partially mediate) the development of pathology in conjunction with other factors. Individuals undergoing cytokine therapy often become clinically depressed (Miller, 2009
; Miller et al., 2009
; Raison et al., 2009
) with individuals with weaker variants of 5-HTTLPR particularly at risk (Bull et al. 2008
; Lotrich et al. 2009
); a similar mechanism could lead to increased risk of depression in SS individuals after repeated mild to moderate stressors. Thus, the pro-inflammatory bias present under resting conditions that persists during stress, may cumulatively, after repeated stressful events, contribute to increased risk for development of depression in SS individuals.
Like other studies demonstrating acute stress-induced increases in pro-inflammatory cytokines (Altemus et al., 2001
; Steptoe et al., 2007
), the present study demonstrates an increase in circulating IL-6, and also demonstrates an increase in the IL-6/IL-10 ratio following exposure to the TSST. Moreover, our findings of a baseline pro-inflammatory bias in healthy SS individuals parallel similar findings in depressed populations: Pace et al. (2006)
reported higher baseline IL-6 concentrations in depressed individuals compared to healthy controls, Dhabhar et al., (2009)
reported lower IL-10 concentrations and higher IL-6/IL-10 ratio in depressed individuals relative to healthy controls, and Bob et al., (2009)
reported that baseline IL-6 is correlated with both depressive symptoms and life stress in depressed individuals. Interestingly, Pace et al., (2006)
also reported increased IL-6 reactivity to TSST in their depressed subjects, an effect that we did not observe in our young and healthy population of SS individuals. This difference may reflect the evolution of the stress-induced pro-inflammatory response over time, which might occur in parallel with the development of psychiatric symptoms after a critical level of exposure to stressors, or both. In an interesting and important study, Bull et al. (2008)
have shown that individuals expressing a low IL-6-synthesizing polymorphism showed fewer depressive symptoms following interferon-alpha treatment. Their study also showed that individuals expressing the LL genotype of the 5-HTTLPR polymorphism, also showed fewer depressive symptoms following interferon-alpha administration, but with a significantly smaller effect. Their results suggest that interactions between circulating cytokine levels and 5-HTTLPR genotype may be important for the development of depressive symptoms, which is in keeping with the results presented here.
Given the findings described here, the biological mechanisms by which the SS genotype might result in a pro-inflammatory bias require further investigation. As discussed previously (Dhabhar et al., 2009
), studies involving in vitro
activation of immune cells in the presence or absence of serotonin, provide clues as to how the availability of serotonin (that may be reduced in SS individuals) may affect the balance between pro- and anti-inflammation. These studies suggest that serotonin-replete conditions tilt the immune balance in favor of anti-inflammatory cytokines, and that conversely, serotonin-deficient conditions, tilt the balance towards pro-inflammation (Menard, Turmel, & Bissonnette, 2007
; Kubera, Kenis, Bosmans, Scharpé, & Maes, 2000
Importantly, SS individuals may be at particular risk of developing serotonin deficiency-related pathology once a significant pro-inflammatory bias is established. This is because in addition to serotonin deficiency potentially increasing pro-inflammatory cytokine concentrations, pro-inflammatory cytokines can in turn precipitate serotonin deficiency by increasing the activity of indoleamine 2,3 dioxygenase (IDO) which diverts tryptophan metabolism towards the kynurenine pathway and decreases tryptophan availability for serotonin synthesis (Dantzer et al., 2008
; Miller, 2009
; Miller et al., 2009
; Schiepers, Wichers, & Maes, 2005
; Raison et al., 2009
). Human and animal studies suggest that the interaction between serotonin metabolism and inflammatory cytokine activity, in particular via the IDO pathway, may play a key role in the development of depressive symptoms. In animal models, depression-like behavior induced by LPS, a common component of pathogenic bacterial cell walls, is mediated by IDO (O'Connor et al. 2009
). Human studies also suggest that tryptophan depletion associated with inflammation may be responsible for the depressogenic effects of inflammatory cytokines. Cancer patients treated with interferon have depressive symptoms that are associated with the magnitude of tryptophan depletion they experience (Capuron et al 2002b
), and two new studies have demonstrated that individuals with weaker 5-HTTLPR variants are more likely to experience depression after interferon-alpha therapy (Bull et al., 2008
; Lotrich et al., 2009
). Given that SS individuals already have a diminished capacity for serotonin uptake, the additional burden of depleted serotonin after even mild to moderate stressful events may act as a "second hit, " putting them at elevated risk for depression.
Both human and animal studies have suggested that, in addition to high levels of pro-inflammatory cytokines, low levels of anti-inflammatory cytokines are associated with depressive symptoms and depression-like behavior (Dhabhar et al., 2009
; Song et al., 2009
; Mesquita et al., 2008
). Interestingly, the higher IL-6/IL-10 ratio shown by SS individuals under resting conditions and during acute stress suggests that even under healthy conditions, these individuals show increased levels of immune factors that are associated with the development of depression. These factors may reflect a higher-level disturbance in overall immune balance in SS individuals.
The approach taken in this study builds on the imaging genetics literature, which has identified a robust S allele-related effect of increased amygdala reactivity and decreased fronto-limbic functional coupling (Hariri et al., 2005
). We identify a pro-inflammatory bias in SS individuals, highlighting the effects of the polymorphism on the immune system whose dysregulation can clearly lead to increased depression risk. This work highlights the need for examination of multiple, potentially interrelated biological systems in the search for proximal effects of genetic variants of interest.
This study has several limitations. First, given our small sample size, it will be important to replicate these findings in larger samples. However, the fact that we were able to detect between-group differences in IL-6/IL-10 ratio in our small sample speaks to the strength of these effects. Second, we chose a young and healthy sample in order to minimize confounding factors and to examine the inflammatory effects of a genetic risk factor that had not yet led to adverse medical or psychiatric outcomes. However, the homogeneity of our sample may limit the generalizability of our results. Future prospective studies should elucidate the association between a pro-inflammatory bias in healthy young individuals and their risk of developing depression later on. Third, due to the prolonged inflammatory cytokine response to acute stress, we were not able to examine genotype differences in the duration / resolution of this inflammatory stress response. Future studies that follow inflammatory markers for extended periods of time after acute stress may help broaden our understanding of genotype-related differences in cytokine response. Fourth, given that some studies have reported differences between SS and LL individuals in post-awakening (Chen et al., 2009
) and stress-induced (Gotlib et al., 2008
) salivary cortisol concentrations, while others have only found differences in post-awakening but not stress-induced cortisol (Wust et al., 2009
), an investigation of circulating cortisol concentrations simultaneously with cytokines and other factors is likely to provide useful and important information. Such an investigation merits future consideration. Finally, any laboratory stressor can only approximate the effect of a real-life psychosocial stressor, and cannot predict the effects of multiple exposures to stressful events. We chose the TSST because the psychosocial stress it elicits is similar to the social rejection stress that has been especially associated with depression onset (Kendler et al., 1995). However, studies that measure the cumulative effects of multiple stressful events, both in and out of the laboratory setting, will increase our understanding of the cumulative impact of stressors on inflammation, and on subsequent psychiatric and medical health outcomes.
To our knowledge, these results are the first to show that even in the absence of a disease state, compared to LL individuals, SS individuals are exposed to higher levels of pro-inflammatory cytokines under resting conditions and during stress. Since life generally involves exposure to a series of stressors, this pro-inflammatory physiological bias is likely to result in a higher chronic inflammatory burden for SS individuals over time, which may increase their vulnerability to stress-related disorders like depression and other diseases that can be facilitated/exacerbated by a chronic pro-inflammatory state. Several studies have demonstrated that positive coping strategies such as values affirmation (Creswell et al., 2005
), meditative practice (Pace et al., 2009
) and social support (e.g., presence of a friend during TSST, Heinrichs et al., 2003
) can decrease the neuroendocrine response to stressful tasks. These studies offer a hopeful perspective on the ability of individuals to cope more effectively with life stressors. They point to the need for further studies to assess whether individuals at increased risk for inflammation-induced depression due to genetic, cytokine treatment-related, or inflammatory disorder-associated causes, can use such coping strategies to reduce the negative impact of life stressors on their health. Clearly, future studies are needed to replicate the findings described here, investigate the biological mechanisms mediating the pro-inflammatory bias/phenotype shown by SS individuals, and explore the potential for using a three-pronged intervention involving behavioral, neurotransmitter-directed and immuno-modulatory approaches for the treatment of depression and other stress-related disorders.