The functional significance of the lung-liver axis during pneumonia

The hepatic acute phase response (APR), stimulated by injury or inflammation, is characterized by significant changes in circulating acute phase protein (APP) concentrations. While individual functions of liver-derived APPs are known, the net consequence of APP changes is unclear. Pneumonia and sepsis elicit systemic inflammation and induce a robust APR. Although APR activation is regarded as a hallmark of infection, direct contributions of liver activation to pulmonary defense during pneumonia and sepsis-induced pneumonia remain unclear. Pneumonia causes a pulmonary inflammatory response coordinated largely by alveolar macrophages, and is typified by cytokine production, leukocyte recruitment and plasma extravasation, the latter of can enable delivery of hepatocyte-derived APPs to the infection site. To determine the functional significance of the hepatic APR during pneumonia, we challenged APR-null mice lacking hepatocyte signal transducer and activator of transcription-3 (STAT3) and RelA with 106 colony-forming units (CFU) Escherichia coli intratracheally. HepSTAT3/RelA-/- mice displayed ablated APP induction, significantly increased mortality, tumor necrosis factor-dependent hepatotoxicity, and pulmonary bacterial burdens. Following a lower (4x105 CFU) E. coli inoculum, hepSTAT3/RelA-/- mice had decreased APP concentrations with reduced pulmonary inflammation and diminished airspace macrophage activation. Similar results were obtained in the context of endotoxemia and pneumonia. We employed an endotoxemia/pneumonia model, whereby 18 hours of intraperitoneal E. coli lipopolysaccharide (5 mg/kg) was followed by intratracheal E. coli (106 CFU) in mice lacking hepatocyte STAT3 (hepSTAT3-/-) or control hepSTAT3+/+ mice. Following endotoxemia and pneumonia, hepSTAT3-/- mice, with significantly reduced levels of circulating and airspace APPs, exhibited significantly elevated lung and blood bacterial burdens and mortality. While neither recruited airspace neutrophils nor lung injury were altered in endotoxemic hepSTAT3-/- mice, in vivo production of reactive oxygen species in alveolar macrophage was significantly decreased. Additionally, bronchoalveolar lavage fluid from this group of hepSTAT3-/- mice allowed greater bacterial growth ex vivo. These results identify a lung-liver axis, whereby the liver response enhances macrophage activation and pulmonary host defense during pneumonia and sepsis-induced pneumonia. Taken together, induction of liver acute phase gene expression programs contributes to countering the deleterious consequences of pneumonia, whether it is alone or in the context of sepsis-induced infection

Myeloid ZFP36L1 does not regulate inflammation or host defense in mouse models of acute bacterial infection.

Zinc finger protein 36, C3H type-like 1 (ZFP36L1) is one of several Zinc Finger Protein 36 (Zfp36) family members, which bind AU rich elements within 3' untranslated regions (UTRs) to negatively regulate the post-transcriptional expression of targeted mRNAs. The prototypical member of the family, Tristetraprolin (TTP or ZFP36), has been well-studied in the context of inflammation and plays an important role in repressing pro-inflammatory transcripts such as TNF-α. Much less is known about the other family members, and none have been studied in the context of infection. Using macrophage cell lines and primary alveolar macrophages we demonstrated that, like ZFP36, ZFP36L1 is prominently induced by infection. To test our hypothesis that macrophage production of ZFP36L1 is necessary for regulation of the inflammatory response of the lung during pneumonia, we generated mice with a myeloid-specific deficiency of ZFP36L1. Surprisingly, we found that myeloid deficiency of ZFP36L1 did not result in alteration of lung cytokine production after infection, altered clearance of bacteria, or increased inflammatory lung injury. Although alveolar macrophages are critical components of the innate defense against respiratory pathogens, we concluded that myeloid ZFP36L1 is not essential for appropriate responses to bacteria in the lungs. Based on studies conducted with myeloid-deficient ZFP36 mice, our data indicate that, of the Zfp36 family, ZFP36 is the predominant negative regulator of cytokine expression in macrophages. In conclusion, these results imply that myeloid ZFP36 may fully compensate for loss of ZFP36L1 or that Zfp36l1-dependent mRNA expression does not play an integral role in the host defense against bacterial pneumonia

ZFP36L1 induction is partially dependent on NF-κB RelA.

<p>(A) Primary alveolar macrophages were obtained from control mice (WT) or mice with a myeloid specific deficiency of RelA (RelA^Δ/Δ) and stimulated in culture with <i>E. coli</i> for 3 hours. (B) Densitometric analysis of immunoblot results from (A).</p

Myeloid ZFP36L1 deficiency does not result in spontaneous immunopathology.

<p>(A) Average weight (n = 12–16) of 8–20 week old mice measured at time of experimentation (B) BAL differential cell counts (n = 7,7) or (C) blood cell counts (n = 7,7) in mice with and without myeloid deficiency of ZFP36L1 (D) TNF- α and IL-6 mRNA in alveolar macrophages obtained from uninfected mice (n = 10,10) (E) TNF-α and IL-6 protein levels measured by ELISA from BAL fluid of uninfected mice (n = 6,6) (F) qRT-PCR analysis of <i>Zfp36</i> mRNA from alveolar macrophages collected from uninfected mice (n = 6,6 p<0.05). Results indicate fold change relative to Cre (−).</p

Myeloid ZFP36L1 does not regulate pulmonary host defense or lung injury.

<p>Lung bacterial burden from Cre(−) or Cre(+) mice after i.t. instillation of (A) <i>E. coli</i> or (B) after 24 hours <i>S. pneumoniae</i> serotype 19 infection. (C) BAL differential cell counts in Cre (−) and Cre (+) mice (n = 6,5) after 24 hours i.t. <i>E coli</i>. Bacterial burden in (D) blood and (E) lung 24 hours after intraperitoneal infection with <i>E. coli</i>. (F) Total protein concentration in BAL fluid as a measurement of lung injury 24 or 72 hours after infection with <i>E. coli</i> (n = 5–8).</p