LXRα and LXRβ are oxysterol-activated NRs. Although perhaps best known for their role in triggering transcriptional programs that promote reverse cholesterol transport, in recent years there has been a growing appreciation of their complex roles in innate immunity. In this capacity, LXRs are recognized to participate in bidirectional negative cross-talk with TLR3/4 and to inhibit proinflammatory gene expression in large part through their inhibition of NF-κB. In the present work, we extend the sphere of influence of LXRs to the lung and the PMN. Our finding in this manuscript of a relative pattern of liver
lung>spleen for LXRα protein expression and lung~spleen>liver for LXRβ protein expression directly parallels a previous report of relative LXRα and LXRβ mRNA abundance in murine tissues (22
). More specifically, we demonstrate expression of LXR in alveolar macrophages, alveolar type II cells, and PMNs and proceed to show potent anti-inflammatory and antihost defense effects of synthetic LXR agonists in the lung. These anti-inflammatory and antihost defense effects share in common impairment of PMN recruitment to the lung and attenuation of lung TNFα expression. LXR stimulation appears to attenuate PMN migration to the lung, at least in part, through inhibition of pulmonary TNFα expression and through impairment of PMN motility. Of note, although the effects of TO-901317 on TNFα parallel those previously reported for glucocorticoid treatment in a rodent LPS lung injury model (56
), our finding that LPS-induced LIX expression is insensitive to LXR agonism represents an important difference, since LIX was originally cloned as a GR-sensitive target (57
). We provide circumstantial evidence that the effect on PMN motility, in turn, may reflect underlying inhibition of RhoA.
We report that synthetic LXR agonists inhibit PMN motility and that this may reflect inhibition of Rho GTPases (). Cellular migration requires precisely timed and spaced cycling of Rho GTPase activation within the cell (53
). Although RhoA has been reported to inhibit LXR (58
), suggesting integration between Rho-dependent functions and LXR activation, we are aware of no previous reports of LXR activation inhibiting RhoA. As for other NR agonists, mixed effects upon Rho signaling have been reported. Androgens have been reported to stimulate Rho signaling (59
). PPARγ agonists have been reported to stimulate motility in intestinal epithelial cells, at least in part, through activation of the Rho GTPase Cdc42 (60
), to inhibit migration in leukocytes (51
), and to inhibit Rho activation in smooth muscle (63
). Our study design, which used a 4-h preincubation of PMNs with TO-901317 preceding KC exposure, does not fully discount the possibility that LXR activation may even activate Rho GTPases acutely and transiently. Such “nongenomic,” rapid signaling events have been described for other NR ligands (64
). This notwithstanding, our data do clearly show that prolonged pharmacologic activation of LXR in the PMN (mimicking our in vivo-dosing regimen) does markedly impair proper activation of RhoA triggered by acute exposure to KC. Of interest, this effect upon motility is similar to what we have previously reported for treatment of PMNs with hydroxymethylglutaryl CoA reductase inhibitors (29
), agents that have been reported to activate LXR in the leukocyte (58
). A previous report that the LXR ligand 25-hydroxycholesterol impairs phagocytosis in macrophages (65
) suggests that LXR may impact additional Rho-dependent macrophage functions that are relevant to host defense. Future studies will need to dissect further the responsible molecular mechanisms.
The present work raises several new issues that will require future investigation. Although synthetic LXR agonists appear to be promising therapeutic candidates for modulation of lung innate immunity, the present work does not address whether the observed effects were specifically LXR dependent, whether some of the in vivo effects might be “second-tier” consequences of LXR activation (e.g., secondary consequences of LXR-dependent changes in lipid metabolism), nor whether LXRα and LXRβ may play distinct roles as has been reported with other phenotypes (12
). Because synergistic effects have been reported among different NR agonists in cell culture (19
), we speculate that there may be a role for LXR agonists in combination therapy of inflammatory lung disease, e.g., as “steroid-sparing” agents. Although the apparent adverse effects of LXR stimulation upon host defense against extracellular bacteria (Figs. and ) will require further careful study, it is worth noting that glucocorticoids are used widely to good clinical effect despite their untoward effects upon host defense (66
). In our study, although conclusions should be tempered as lung histopathology was quantified at only one time point following K. pneumoniae
inoculation (48 h), we speculate that the enhanced mortality seen with the LXR agonist () was not due to aggravated lung injury. Instead, we speculate that it may reflect worsened septicemia due to enhancement of bacterial dissemination with the LXR agonist (). Of interest, it has been reported that LXR-null mice are more susceptible to bloodstream infection with the Gram-positive intracellular bacterium L. monocytogenes
). Because microbial virulence factors, host compartments, and host defense mechanisms differ markedly between these two bacteria, these observations, taken together, indicate that LXR may exert mixed influences upon the different effector arms of host defense. A report that a fungal metabolite, paxilline, is a potent LXR agonist (67
) suggests that pathogen-specific molecules may add a further layer of complexity to the role of LXR in host defense.
Finally, in a broader sense, we speculate that LXR may possibly underlie associations that have been noted between metabolic disorders such as obesity, in which a role for LXR has been identified (68
), and inflammatory lung diseases, such as asthma (69
). The present work suggests that cholesterol metabolism may play a much more important role in lung biology than has previously been recognized, and that therapies targeting disordered cholesterol metabolism may well impact lung biology.