S100A9 adenovirus

The release of endogenous damage-associated molecular patterns (DAMPs), including members of the S100 family, is associated with infection, cellular stress, tissue injury, and cancer. Here, the researchers examined the ability of S100 proteins to induce proinflammatory cytokines and cell migration, and the role of RAGE and TLR4 in mediating these responses in vitro. They also investigated the potential of S100A9 homodimers to induce lung inflammation in vivo using adenoviral delivery of S100A9 in mice. S100A8, S100A9, and S100A12, but not S100A8/A9 heterodimers, induced modest levels of TLR4-mediated cytokine production by human peripheral blood mononuclear cells (PBMCs). In contrast, for most S100s, including S100A9, RAGE blockade inhibited S100-mediated cell migration of THP1 cells and major leukocyte populations, while TLR4 blockade had no such effect. Intranasal administration of murine S100A9 adenovirus induced a specific, time-dependent, macrophage-dominated infiltration that was concurrent with elevated levels of S100A9 and proinflammatory cytokines in bronchoalveolar fluid. TLR4-deficient mice produced significantly reduced inflammatory cytokines, but unexpectedly, loss of TLR4 signaling or RAGE deficiency did not significantly affect S100A9-mediated lung pathology or inflammatory cell infiltration in the alveolar space. These data suggest that physiological levels of S100A9 homodimers can elicit an inflammatory response in vivo, and that, although RAGE and TLR4 blockade inhibited the inflammatory response in vitro, the response was not primarily dependent on these two receptors.

To directly assess the inflammatory effects of S100A9 in vivo, researchers constructed adenovirus encoding murine S100A9 and administered it intranasally to uninfected mice. At 8 days post-infection,  the cellular infiltrates in the BAL fluids of the C57BL/6 wild-type mice were significantly increased in the adeno-mS100A9 treated group compared to the adeno-null group (Figure 1A). Of note, there were no significant differences in inflammatory infiltrates in the bronchoalveolar lavage (BAL) fluid between wild-type mice and Ager-/- (RAGE-deficient) mice after adenovirus-mS100A9 treatment (Figure 1A). Levels of IFNγ and IL-6 in the bronchoalveolar fluid of adeno-mS100A9 infected mice were significantly increased compared with the adeno-null controls, but again there were no differences between wild-type mice and ager-/- mice infected with mS100A9 adenovirus (Figure 1B). Western blot analysis of bronchoalveolar fluid showed that murine S100A9 was detected almost exclusively in the adeno-S100A9 challenge group on day 8 (Figure 1C). In lung tissue, there were mononuclear cell infiltrations (mainly macrophages) around multiple bronchioles and blood vessels, and the number of cells in the interstitium and air spaces was slightly increased in the adeno-null treated group, while inflammatory cells were absent or very few in the PBS control group (Figure 1D). The peribronchial, perivascular, and interstitial inflammatory cell infiltrations were significantly more severe in mice treated with adeno-mS100A9, mainly composed of macrophages and lymphocytes, and sometimes even extended into the alveolar space (Figure 1D). There were no significant differences in the results of histopathological evaluation between wild-type and RAGE-deficient mice treated with adeno-mS100A9 (Figure 1D). These results suggest that RAGE does not appear to play a role in S100A9-mediated cytokine induction and inflammation in this in vivo model.

Figure 1. Murine S100A9 induced lung inflammation is RAGE-independent. (Chen B, et al., 2015)