Kamran Atabai

IL-17A produced by αβ T cells drives airway hyper-responsiveness in mice and enhances mouse and human airway smooth muscle contraction

Emerging evidence suggests that the T helper 17 (TH17) subset of αβ T cells contributes to the development of allergic asthma. In this study, we found that mice lacking the αvβ8 integrin on dendritic cells did not generate TH17 cells in the lung and were protected from airway hyper-responsiveness in response to house dust mite and ovalbumin sensitization and challenge. Because loss of TH17 cells inhibited airway narrowing without any obvious effects on airway inflammation or epithelial morphology, we examined the direct effects of TH17 cytokines on mouse and human airway smooth muscle function. Interleukin-17A (IL-17A), but not IL-17F or IL-22, enhanced contractile force generation of airway smooth muscle through an IL-17 receptor A (IL-17RA)–IL-17RC, nuclear factor κ light-chain enhancer of activated B cells (NF-κB)–ras homolog gene family, member A (RhoA)–Rho-associated coiled-coil containing protein kinase 2 (ROCK2) signaling cascade. Mice lacking integrin αvβ8 on dendritic cells showed impaired activation of this pathway after ovalbumin sensitization and challenge, and the diminished contraction of the tracheal rings in these mice was reversed by IL-17A. These data indicate that the IL-17A produced by TH17 cells contributes to allergen-induced airway hyper-responsiveness through direct effects on airway smooth muscle.

Essential role for MFG-E8 as ligand for αvβ5 integrin in diurnal retinal phagocytosis

The integrin receptor αvβ5 controls two independent forms of interactions of the retinal pigment epithelium (RPE) with adjacent photoreceptor outer segments that are essential for vision. αvβ5 localizes specifically to apical microvilli of the RPE and contributes to retinal adhesion that maintains RPE contacts with intact outer segments at all times. Additionally, αvβ5 synchronizes diurnal bursts of RPE phagocytosis that clear photoreceptor outer segment fragments (POS) shed in a circadian rhythm. Dependence of retinal phagocytosis and adhesion on αvβ5 receptors suggests that the extracellular matrix ensheathing RPE microvilli contains ligands for this integrin. Here we studied mice lacking expression of functional MFG-E8 to test the contribution of this integrin ligand to αvβ5 functions in the retina. Lack of MFG-E8 only minimally reduced retinal adhesion. In contrast, lack of MFG-E8, like lack of αvβ5 receptor, eliminated αvβ5 downstream signaling involving the engulfment receptor MerTK and peak POS phagocytosis, both of which follow light onset in wild-type retina. MFG-E8-deficient RPE in primary culture retained normal epithelial morphology and levels of apical αvβ5 receptors, but showed impaired binding and engulfment of isolated POS. Soluble or POS-bound recombinant MFG-E8 was sufficient to fully restore phagocytosis by MFG-E8-deficient RPE. Furthermore, MFG-E8 supplementation strongly increased POS binding by wild-type and MerTK-deficient RPE, but did not affect POS binding by RPE lacking αvβ5. Thus, MFG-E8 stimulates rhythmic POS phagocytosis by ligating apical αvβ5 receptors of the RPE. These results identify MFG-E8 as the first extracellular ligand in the retina that is essential for diurnal POS phagocytosis.

Mfge8 diminishes the severity of tissue fibrosis in mice by binding and targeting collagen for uptake by macrophages

Milk fat globule epidermal growth factor 8 (Mfge8) is a soluble glycoprotein known to regulate inflammation and immunity by mediating apoptotic cell clearance. Since fibrosis can occur as a result of exaggerated apoptosis and inflammation, we set out to investigate the hypothesis that Mfge8 might negatively regulate tissue fibrosis. We report here that Mfge8 does decrease the severity of tissue fibrosis in a mouse model of pulmonary fibrosis; however, it does so not through effects on inflammation and apoptotic cell clearance, but by binding and targeting collagen for cellular uptake through its discoidin domains. Initial analysis revealed that Mfge8-/- mice exhibited enhanced pulmonary fibrosis after bleomycin-induced lung injury. However, they did not have increased inflammation or impaired apoptotic cell clearance after lung injury compared with Mfge8+/+ mice; rather, they had a defect in collagen turnover. Further experiments indicated that Mfge8 directly bound collagen and that Mfge8-/- macrophages exhibited defective collagen uptake that could be rescued by recombinant Mfge8 containing at least one discoidin domain. These data demonstrate a critical role for Mfge8 in decreasing the severity of murine tissue fibrosis by facilitating the removal of accumulated collagen.

Human eosinophils induce mucin production in airway epithelial cells via epidermal growth factor receptor activation.

Eosinophil recruitment and mucus hypersecretion are characteristic of asthmatic airway inflammation, but eosinophils have not been shown to induce mucin production. Because an epidermal growth factor receptor (EGFR) cascade induces MUC5AC mucin in airways, and because EGFR is up-regulated in asthmatic airways, we examined the effect of eosinophils on MUC5AC mucin production in NCI-H292 cells (a human airway epithelial cell line that produces mucins). Eosinophils were isolated from the peripheral blood of allergic patients, and their effects on MUC5AC mucin gene and protein synthesis were assessed using in situ hybridization and ELISAs. When IL-3 plus GM-CSF or IL-3 plus IL-5 were added to eosinophils cultured with NCI-H292 cells, MUC5AC mucin production increased; eosinophils or cytokines alone had no effect. Eosinophil supernatant obtained by culturing eosinophils with IL-3 plus GM-CSF or IL-3 plus IL-5 also increased MUC5AC synthesis in NCI-H292 cells, an effect that was prevented by selective EGFR inhibitors (AG1478, BIBX1522). Supernatant of activated eosinophils induced EGFR phosphorylation in NCI-H292 cells. Supernatant of activated eosinophils contained increased concentrations of TGF-α protein (an EGFR ligand) and induced up-regulation of TGF-α expression and release in NCI-H292 cells. A blocking Ab to TGF-α reduced activated eosinophil-induced MUC5AC synthesis in NCI-H292 cells. These results show that activated eosinophils induce mucin synthesis in human airway epithelial cells via EGFR activation, and they implicate TGF-α produced by eosinophils and epithelial cells in the EGFR activation that results in mucin production in human airway epithelium.

The pulmonary physician in critical care • 5: Acute lung injury and the acute respiratory distress syndrome: definitions and epidemiology

An understanding of the epidemiology of ALI/ARDS and the effects of treatment have been hampered by the lack of a uniform definition of the syndrome. Various definitions have been proposed, and these are reviewed with particular attention to how changes in definition have affected our understanding of the natural history and treatment options for the condition.

Always cleave up your mess: targeting collagen degradation to treat tissue fibrosis

Pulmonary fibrosis is a vexing clinical problem with no proven therapeutic options. In the normal lung there is continuous collagen synthesis and collagen degradation, and these two processes are precisely balanced to maintain normal tissue architecture. With lung injury there is an increase in the rate of both collagen production and collagen degradation. The increase in collagen degradation is critical in preventing the formation of permanent scar tissue each time the lung is exposed to injury. In pulmonary fibrosis, collagen degradation does not keep pace with collagen production, resulting in extracellular accumulation of fibrillar collagen. Collagen degradation occurs through both extracellular and intracellular pathways. The extracellular pathway involves cleavage of collagen fibrils by proteolytic enzyme including the metalloproteinases. The less-well-described intracellular pathway involves binding and uptake of collagen fragments by fibroblasts and macrophages for lysosomal degradation. The relationship between these two pathways and their relevance to the development of fibrosis is complex. Fibrosis in the lung, liver, and skin has been associated with an impaired degradative environment. Much of the current scientific effort in fibrosis is focused on understanding the pathways that regulate increased collagen production. However, recent reports suggest an important role for collagen turnover and degradation in regulating the severity of tissue fibrosis. The objective of this review is to evaluate the roles of the extracellular and intracellular collagen degradation pathways in the development of fibrosis and to examine whether pulmonary fibrosis can be viewed as a disease of impaired matrix degradation rather than a disease of increased matrix production.

Polarization of Prostate Cancer Associated Macrophages is Induced by Milk-Fat Globule-EGF Factor 8 (MFG-E8) Mediated Efferocytosis

Background: The growing body of data on tumor-associated macrophages largely neglects phagocytosis of apoptotic cells. Results: MFG-E8, induced during efferocytosis, contributes to macrophage polarization with STAT3/SOCS3 pathway involvement. Conclusion: Efferocytosis induces macrophage polarization into tumor-associated macrophages mediated by MFG-E8. Significance: A novel tumor-promoting mechanism for macrophage polarization through efferocytosis and MFG-E8 and its corresponding signaling pathway were identified. Tumor cells secrete factors that modulate macrophage activation and polarization into M2 type tumor-associated macrophages, which promote tumor growth, progression, and metastasis. The mechanisms that mediate this polarization are not clear. Macrophages are phagocytic cells that participate in the clearance of apoptotic cells, a process known as efferocytosis. Milk fat globule- EGF factor 8 (MFG-E8) is a bridge protein that facilitates efferocytosis and is associated with suppression of proinflammatory responses. This study investigated the hypothesis that MFG-E8-mediated efferocytosis promotes M2 polarization. Tissue and serum exosomes from prostate cancer patients presented higher levels of MFG-E8 compared with controls, a novel finding in human prostate cancer. Coculture of macrophages with apoptotic cancer cells increased efferocytosis, elevated MFG-E8 protein expression levels, and induced macrophage polarization into an alternatively activated M2 phenotype. Administration of antibody against MFG-E8 significantly attenuated the increase in M2 polarization. Inhibition of STAT3 phosphorylation using the inhibitor Stattic decreased efferocytosis and M2 macrophage polarization in vitro, with a correlating increase in SOCS3 protein expression. Moreover, MFG-E8 knockdown tumor cells cultured with wild-type or MFG-E8-deficient macrophages resulted in increased SOCS3 expression with decreased STAT3 activation. This suggests that SOCS3 and phospho-STAT3 act in an inversely dependent manner when stimulated by MFG-E8 and efferocytosis. These results uncover a unique role of efferocytosis via MFG-E8 as a mechanism for macrophage polarization into tumor-promoting M2 cells.

Roles of the Innate Immune System in Mammary Gland Remodeling During Involution

Mammary gland involution is a period of intensive tissue remodeling. Over the course of a relatively brief period, a large proportion of the mammary gland epithelium undergoes programmed cell death and is removed by phagocytes. In addition, the gland is cleared of residual milk fat globules as well as milk and adipocytes become the predominant cell type. The role of the immune system in this process has not been clearly defined. Professional phagocytes derived from the immune system can participate in the clearance of apoptotic and autophagic cells, the removal of residual milk components, and the prevention of mastitis during mammary gland involution. However, many of these functions can also be performed by non-professional phagocytes (e.g. mammary epithelial cells). This review will discuss the evidence that supports a role for innate immune cells in mammary gland remodeling during involution.

Mfge8 promotes obesity by mediating the uptake of dietary fats and serum fatty acids

Fatty acids are integral mediators of energy storage, membrane formation and cell signaling. The pathways that orchestrate uptake of fatty acids remain incompletely understood. Expression of the integrin ligand Mfge8 is increased in human obesity and in mice on a high-fat diet, but its role in obesity is unknown. We show here that Mfge8 promotes the absorption of dietary triglycerides and the cellular uptake of fatty acid and that Mfge8-deficient (Mfge8−/−) mice are protected from diet-induced obesity, steatohepatitis and insulin resistance. Mechanistically, we found that Mfge8 coordinates fatty acid uptake through αvβ3 integrin– and αvβ5 integrin–dependent phosphorylation of Akt by phosphatidylinositide-3 kinase and mTOR complex 2, leading to translocation of Cd36 and Fatp1 from cytoplasmic vesicles to the cell surface. Collectively, our results imply a role for Mfge8 in regulating the absorption and storage of dietary fats, as well as in the development of obesity and its complications.

Mfge8 suppresses airway hyperresponsiveness in asthma by regulating smooth muscle contraction

Airway obstruction is a hallmark of allergic asthma and is caused primarily by airway smooth muscle (ASM) hypercontractility. Airway inflammation leads to the release of cytokines that enhance ASM contraction by increasing ras homolog gene family, member A (RhoA) activity. The protective mechanisms that prevent or attenuate the increase in RhoA activity have not been well studied. Here, we report that mice lacking the gene that encodes the protein Milk Fat Globule-EGF factor 8 (Mfge8−/−) develop exaggerated airway hyperresponsiveness in experimental models of asthma. Mfge8−/− ASM had enhanced contraction after treatment with IL-13, IL-17A, or TNF-α. Recombinant Mfge8 reduced contraction in murine and human ASM treated with IL-13. Mfge8 inhibited IL-13–induced NF-κB activation and induction of RhoA. Mfge8 also inhibited rapid activation of RhoA, an effect that was eliminated by an inactivating point mutation in the RGD integrin-binding site in recombinant Mfge8. Human subjects with asthma had decreased Mfge8 expression in airway biopsies compared with healthy controls. These data indicate that Mfge8 binding to integrin receptors on ASM opposes the effect of allergic inflammation on RhoA activity and identify a pathway for specific inhibition of ASM hypercontractility in asthma.