The Src-Family Kinases Hck and Fgr Regulate Early Lipopolysaccharide-Induced Myeloid Cell Recruitment into the Lung and Their Ability To Secrete Chemokines

Abstract

Myeloid leukocyte recruitment into the lung in response to environmental cues represents a key factor for the induction of lung damage. We report that Hck and Fgr-deficient mice show a profound impairment in early recruitment of neutrophils and monocytes in response to bacterial lipopolysaccharide (LPS). The reduction in interstitial and airway neutrophil recruitment was not due to a cell-intrinsic migratory defect, because Hck and Fgr-deficient neutrophils were attracted to the airways by the chemokine CXCL2 as wild type cells. However, early accumulation of chemokines and TNFα in the airways was reduced In hck−/−fgr−/− mice. Considering that chemokine and TNFα release into the airways was neutrophil-independent, as suggested by a comparison between control and neutrophil-depleted mice, we examined LPS-induced chemokine secretion by neutrophils and macrophages in wild type and mutant cells. Notably, mutant neutrophils displayed a marked deficit in their capability to release the chemokines CXCL1, CXCL2, CCL3 and CCL4, and TNFα, in response to LPS. However, intracellular accumulation of these chemokines and TNFα in, as well as secretion of a wide array of cytokines, including IL-1α, IL-1β, IL-6 and IL-10, by hck−/−fgr−/− neutrophils was normal. Intriguingly, secretion of CXCL1, CXCL2, CCL2, CCL3, CCL4, RANTES and TNFα, but not IL-1α, IL-1β, IL-6, IL-10 and GM-CSF, was also markedly reduced in bone marrow-derived macrophages (BMDM). Consistently, the Src kinase inhibitors PP2 and dasatinib reduced chemokine secretion by neutrophils and BMDM. These findings identify Src kinases as critical regulator of chemokine secretion in myeloid leukocytes during lung inflammation.

Inhibition of Hematopoietic Cell Kinase Activity Suppresses Myeloid Cell-Mediated Colon Cancer Progression

“…Since Hck activity enhances migration of immune cells to sites of inflammation (Ernst et al, 2002; Mazzi et al, 2015), we quantified tumor-associated immune cells by flow cytometry. Surprisingly, in both CRC models we observed a comparable abundance between WT and Hck CA mice of CD45 + cells gated for CD19 + B lymphocytes (15.90% ± 0.93% versus 14.93% ± 0.73%; data refer to sporadic model), TCRβ + T lymphocytes (36.87% ± 1.88% versus 33.73%±1.53%),orCD11b + F4/80 + macrophages (17.8% ± 0.50% versus 19.36% ± 2.67%).…”

“…Although Hck and Fgr only show overlapping localization at the cell membrane, compound Hck KO ; Fgr KO mice are unable to clear Listeria infection (Lowell et al, 1994), and show blunted recruitment of myeloid cells to arterial plaques or to LPS-challenged lungs (Mazzi et al, 2015; Medina et al, 2015). By contrast, excessive activation of endogenous SFK results in a lethal autoimmune response in Lyn CA mice (Hibbs et al, 1995) and inflammatory lung disease in Hck CA mice (Ernst et al, 2002).…”

Kinases as Novel Therapeutic Targets in Asthma and Chronic Obstructive Pulmonary Disease

“…Hck overexpression in mice induces chronic neutrophilic inflammation and emphysema (Ernst, 2002). Hck is involved in neutrophil and myeloid cell recruitment into the lung, because Hck knockout mice fail to recruit these cells in response to TNF-a and LPS Kinase Inhibitors in Airway Disease 807 (Mazzi et al, 2015). There is increased expression of Hck in circulating neutrophils from patients with COPD (Yanagisawa et al, 2009), suggesting that this kinase may be a good target for inhibition.…”

Intracellular β2 integrin (CD11/CD18) interacting partners in neutrophil trafficking

“…Interestingly, in vitro chemotaxis assays using fibrinogen‐coated transwell chambers with fMLP as a chemoattractant reveal normal migratory behaviour of Hck −/− Fgr −/− Lyn −/− neutrophils . This observation is confirmed in multiple disease models, including LPS‐induced acute lung injury showing a comparable neutrophil accumulation at the site of inflammation between wild‐type and SFK knockout mice. Furthermore, bone marrow chimeric mice with a mixed Hck −/− Fgr −/− Lyn −/− and wild‐type hematopoietic system show also a normal recruitment of neutrophils to sites of inflammation, suggesting that SFKs are not required for the cell intrinsic migratory ability of neutrophils .…”

“…However, SFK‐deficient mice and wild‐type mice transplanted with Hck −/− Fgr −/− Lyn −/− bone marrow are protected from disease development. Here, SFK‐deficiency results in reduced release of pro‐inflammatory mediators revealing a crucial role of SFKs in disease progression by regulating chemokine secretion of myeloid cells …”