One hour after the second inhibitor treatment, mice were challenged with i.t. hours. In LDC1267 contrast, everolimus did not affect IL-6 and TNF- levels. Interestingly, both inhibitors reduced inflammatory cytokines in an LPS/oleic acid-induced lung injury model. In conclusion, the mTOR inhibitors did not worsen the overall histopathological severity, but they exerted distinct effects on proinflammatory cytokine expression in the lung depending on the lung injury model applied. 1.?Introduction The serine/threonine kinase mammalian target of rapamycin (mTOR) pathway belongs to a critical cellular signaling pathway that affects broad aspects of cellular functions including metabolism, growth, and survival [1,2]. mTOR complex 1 (mTORC1) is linked to the phosphatidylinositol-3 kinase (PI3K) via the serine/threonine kinase Akt . The PI3K/Akt/mTOR pathway is best known for having important functions in regulating adaptive immune cell activation, proliferation, and survival [4,5]. Rapamycin (also called sirolimus) and everolimus, two prototypical inhibitors of mTORC1, have potent immunosuppressive and anti-tumor activities by preventing proliferation and cell-cycle progression [6,7]. Hence, inhibition of mTORC1 was introduced in clinical transplantation and is currently employed as an immunosuppressive treatment to ameliorate chronic allograft damage and prevent immunosuppression-associated malignancies [8,9]. The cell-cycle arrest induced by mTOR inhibitors might in part explain LDC1267 its potent anti-tumor action especially in cancers with an up-regulated PI3K/Akt/mTOR pathway including anti-metastatic and anti-angiogenic effects [10,11]. In addition, mTOR inhibitors have shown promising results in advanced clinical trials against certain malignancies like renal cell carcinoma, mantle cell lymphoma, and endometrial cancers [12,13]. Transplant patients and renal cell carcinoma patients treated with the mTOR inhibitors rapamycin or everolimus experience noninfectious drug-induced pneumonitis (DIP) [14C17]. Moreover, breast cancer patients treated with everolimus similarly develop DIP  suggesting that DIP is common to all mTOR inhibitors. The frequency of DIP varies from 3-30% depending on the study analysis and seems independent from drug trough levels. Histologically, the patients present with diffuse interstitial infiltrates and bronchoalveolar lavage as well as lung biopsy reveals features of lymphocytic alveolitis, lymphocytic interstitial pneumonitis, bronchoalveolar obliterans organizing pneumonia, focal fibrosis, pulmonary alveolar hemorrhage, or a combination thereof . Bronchoalveolar fluid microbiological evaluation is negative for many bacteria, fungi, parasites, or viruses indicating a noninfectious origin of DIP . However, the underlying molecular causes of this form of acute lung injury (ALI) induced by the mTOR inhibitors are unclear. The pathogenesis of acute lung injury (ALI) is complex and may be triggered by air pollutants or environmental cues such as asbestos , nickel , or tobacco smoke  but also by chemotherapeutics. The pathology involves the recruitment of neutrophils, the expression of inflammatory cytokines, and the apoptosis of epithelial cells but also of neutrophils finally leading to a disruption of the alveolar epithelial lung barrier, pulmonary edema, and abnormalities in gas exchange [24,25]. Lung macrophages of the innate immune system play a key role for the pathogenesis of ALI. For example, in acid-induced lung injury, oxidized phospholipids generated in the lung induce injury via Toll-like receptor 4 (TLR4) activation on macrophages . This induces the expression of proinflammatory cytokines such as TNF-, IL-6 or IL-1. In addition, the TLR4 ligand lipopolysaccharide (LPS) can induce ALI in the mouse , which is dependent on TLR4 . These data together suggest that the innate immune system is critically involved in the pathogenesis of pneumonitis or ALI. While mTOR inhibitors were initially thought to globally dampen innate immune responses  and SIGLEC6 thereby contribute to their potent immunosuppressive properties, we LDC1267 and others could recently show that inhibition of mTOR is able to promote expression of the proinflammatory cytokines IL-12, TNF-, IL-6, and IL-1 after stimulation with the TLR4 ligand LPS [30C36]. Recently, it has been shown that activation of mTOR inhibits inflammatory responses and apoptosis in the lung of mice due to cigarette smoke or LPS [37C39]. Therefore, we speculated that inhibition of mTOR in the innate immune system might exaggerate an LDC1267 initial insult in the lung leading to augmented.