Alveolar fluid in acute respiratory distress syndrome promotes fibroblast migration: role of platelet-derived growth factor pathway

OBJECTIVES: Fibroblast migration is an initiating step in fibroproliferation; its involvement during acute lung injury and acute respiratory distress syndrome remains poorly understood. The aims of this study were: 1) to determine whether bronchoalveolar lavage fluids from patients with acute lung injury/acute respiratory distress syndrome modulate lung fibroblast migration; 2) to assess lung fibroblast migration\u27s clinical relevance; and 3) to evaluate the role of the platelet-derived growth factor pathway in this effect. DESIGN: Prospective cohort study. SETTING: Three intensive care units of a large tertiary referral center. PATIENTS: Ninety-three ventilated patients requiring bronchoalveolar lavage fluids were enrolled (48 with acute respiratory distress syndrome, 33 with acute lung injury, and 12 ventilated patients without acute lung injury/acute respiratory distress syndrome). INTERVENTIONS: After bronchoalveolar lavage fluids collection during standard care, the patients were followed up for 28 days and clinical outcomes were recorded. Migration assays were performed by using a Transwell model; bronchoalveolar lavage fluids platelet-derived growth factor and soluble platelet-derived growth factor receptor-alpha were characterized by Western blot and measured by ELISA. MEASUREMENTS AND MAIN RESULTS: Most of the bronchoalveolar lavage fluids inhibited basal fibroblast migration. Bronchoalveolar lavage fluids chemotactic index increased with severity of lung injury (28% in patients without acute lung injury/acute respiratory distress syndrome and with acute lung injury vs. 91% in acute respiratory distress syndrome patients; p = .016). In acute lung injury/acute respiratory distress syndrome patients, inhibition of basal fibroblast migration by bronchoalveolar lavage fluids below 52% was independently associated with a lower 28-day mortality (odds ratio [95% confidence interval] 0.313 [0.10-0.98], p = .046). Platelet-derived growth factor-related peptides and soluble platelet-derived growth factor-Ralpha were detected in all bronchoalveolar lavage fluids from acute lung injury/acute respiratory distress syndrome patients. The effect of bronchoalveolar lavage fluids stimulating migration was inhibited by a specific platelet-derived growth factor receptor inhibitor (AG1296). Bronchoalveolar lavage fluids inhibiting migration reversed the effect of rh-platelet-derived growth factor-BB and reduced by 40% the binding of 125I-platelet-derived growth factor-BB to fibroblast cell surface in favor of a role for platelet-derived growth factor-sRalpha. CONCLUSIONS: : Together, our results suggest that during acute lung injury, fibroblast migration is modulated by bronchoalveolar lavage fluids through a platelet-derived growth factor/platelet-derived growth factor-sRalpha balance. Migration is associated with clinical severity and patient 28-day mortality

Sphingosine-1-phosphate promotes the persistence of activated CD4 T cells in inflamed sites

Inflammation can be protective or pathogenic depending on context and timeframe. Acute inflammation, including the accumulation of CD4 T cells, accompanies protective immune responses to pathogens, but the presence of activated CD4 T cells at sites of inflammation is associated with chronic inflammatory disease. While significant progress has been made in understanding the migration of CD4 T cells into inflamed sites, the signals that lead to their persistence are poorly characterized. Using a murine ear model of acute inflammation and intravital two-photon imaging, we have dissected the signals that mediate CD4 T cell persistence. We report the unexpected finding that the bioactive lipid, sphingosine-1-phosphate (S1P), is both necessary and sufficient for the persistence of activated CD4 T cells at peripheral tissues in acute inflammation. S1P mediated the enhanced motility of CD4 T cells at inflamed tissues but did not affect their migration to the downstream draining lymph node. We found that sphingosine kinase-1, which regulates S1P production is increased at inflamed sites in mice and in patients with the chronic inflammatory disease, rheumatoid arthritis. Together, these data suggest that S1P, or its regulators, may be key targets to promote or disrupt accumulation of CD4 T cells at inflamed tissues

Deposit Growth in the Wetting of an Angular Region with Uniform Evaporation

Solvent loss due to evaporation in a drying drop can drive capillary flows and solute migration. The flow is controlled by the evaporation profile and the geometry of the drop. We predict the flow and solute migration near a sharp corner of the perimeter under the conditions of uniform evaporation. This extends the study of Ref. 6, which considered a singular evaporation profile, characteristic of a dry surrounding surface. We find the rate of the deposit growth along contact lines in early and intermediate time regimes. Compared to the dry-surface evaporation profile of Ref. 6, uniform evaporation yields more singular deposition in the early time regime, and nearly uniform deposition profile is obtained for a wide range of opening angles in the intermediate time regime. Uniform evaporation also shows a more pronounced contrast between acute opening angles and obtuse opening angles.Comment: 12 figures, submitted to Physical Review

Ibrutinib inhibits SDF1/CXCR4 mediated migration in AML

Pharmacological targeting of BTK using ibrutinib has recently shown encouraging clinical activity in a range of lymphoid malignancies. Recently we reported that ibrutinib inhibits human acute myeloid leukemia (AML) blast proliferation and leukemic cell adhesion to the surrounding bone marrow stroma cells. Here we report that in human AML ibrutinib, in addition, functions to inhibit SDF1/CXCR4-mediated AML migration at concentrations achievable in vivo. It has previously been shown that SDF1/CXCR4-induced migration is dependent on activation of downstream BTK in chronic lymphocytic leukaemia (CLL) and multiple myeloma. Here we show that SDF-1 induces BTK phosphorylation and downstream MAPK signalling in primary AML blast. Furthermore, we show that ibrutinib can inhibit SDF1-induced AKT and MAPK activation. These results reported here provide a molecular mechanistic rationale for clinically evaluating BTK inhibition in AML patients and suggests that in some AML patients the blasts count may initially rise in response to ibrutinib therapy, analgous to similar clinical observations in CLL

Intracapillary leucocyte accumulation as a novel antihaemorrhagic mechanism in acute pancreatitis in mice

Background: Pancreatic infiltration by leucocytes represents a hallmark in acute pancreatitis. Although leucocytes play an active role in the pathophysiology of this disease, the relation between leucocyte activation, microvascular injury and haemorrhage has not been adequately addressed.Methods: We investigated intrapancreatic leucocyte migration, leucocyte extravasation and pancreatic microperfusion in different models of oedematous and necrotising acute pancreatitis in lys-EGFP-ki mice using fluorescent imaging and time-lapse intravital microscopy.Results: In contrast to the current paradigm of leucocyte recruitment, the initial event of leucocyte activation in acute pancreatitis was represented through a dose- and time-dependent occlusion of pancreatic capillaries by intraluminally migrating leucocytes. Intracapillary leucocyte accumulation (ILA) resulted in dense filling of almost all capillaries close to the area of inflammation and preceded transvenular leucocyte extravasation. ILA was also initiated by isolated exposure of the pancreas to interleukin 8 or fMLP, demonstrating the causal role of chemotactic stimuli in the induction of ILA. The onset of intracapillary leucocyte accumulation was strongly inhibited in LFA-1-/- and ICAM-1-/- mice, but not in Mac-1-/- mice. Moreover, prevention of intracapillary leucocyte accumulation led to the development of massive capillary haemorrhages and transformed mild pancreatitis into lethal haemorrhagic disease.Conclusions: ILA represents a novel protective and potentially lifesaving mechanism of haemostasis in acute pancreatitis. This process depends on expression of LFA-1 and ICAM-1 and precedes the classical steps of the leucocyte recruitment cascade

Targeting PI3Kδ and PI3Kγ signalling disrupts human AML survival and bone marrow stromal cell mediated protection

Phosphoinositide-3-kinase (PI3K) is an enzyme group, known to regulate key survival pathways in acute myeloid leukaemia (AML). It generates phosphatidylinositol-3,4,5-triphosphate, which provides a membrane docking site for protein kinaseB activation. PI3K catalytic p110 subunits are divided into 4 isoforms; α,β,δ and γ. The PI3Kδ isoform is always expressed in AML cells, whereas the frequency of PI3Kγ expression is highly variable. The functions of these individual catalytic enzymes have not been fully resolved in AML, therefore using the PI3K p110δ and p110γ-targeted inhibitor IPI-145 (duvelisib) and specific p110δ and p110γ shRNA, we analysed the role of these two p110 subunits in human AML blast survival. The results show that PI3Kδ and PI3Kγ inhibition with IPI-145 has anti-proliferative activity in primary AML cells by inhibiting the activity of AKT and MAPK. Pre-treatment of AML cells with IPI-145 inhibits both adhesion and migration of AML blasts to bone marrow stromal cells. Using shRNA targeted to the individual isoforms we demonstrated that p110δ-knockdown had a more significant anti-proliferative effect on AML cells, whereas targeting p110γ-knockdown significantly inhibited AML migration. The results demonstrate that targeting both PI3Kδ and PI3Kγ to inhibit AML-BMSC interactions provides a biologic rationale for the pre-clinical evaluation of IPI-145 in AML

Beneficial effects of reconstituted high-density lipoprotein (rHDL) on circulating CD34+ cells in patients after an acute coronary syndrome

Background: High-density lipoproteins (HDL) favorably affect endothelial progenitor cells (EPC). Circulating progenitor cell level and function are impaired in patients with acute coronary syndrome (ACS). This study investigates the short-term effects of reconstituted HDL (rHDL) on circulating progenitor cells in patients with ACS. Methods and Findings: The study population consisted of 33 patients with recent ACS: 20 patients from the ERASE trial (randomized to receive 4 weekly intravenous infusions of CSL-111 40 mg/kg or placebo) and 13 additional patients recruited as controls using the same enrolment criteria. Blood was collected from 16 rHDL (CSL-111)-treated patients and 17 controls at baseline and at 6–7 weeks (i.e. 2–3 weeks after the fourth infusion of CSL-111 in ERASE). CD34+ and CD34+/kinase insert domain receptor (KDR+) progenitor cell counts were analyzed by flow cytometry. We found preserved CD34+ cell counts in CSL-111-treated subjects at follow-up (change of 1.6%), while the number of CD34+ cells was reduced (-32.9%) in controls (p = 0.017 between groups). The level of circulating SDF-1 (stromal cell-derived factor-1), a chemokine involved in progenitor cell recruitment, increased significantly (change of 21.5%) in controls, while it remained unchanged in CSL-111-treated patients (p = 0.031 between groups). In vitro exposure to CSL-111 of early EPC isolated from healthy volunteers significantly increased CD34+ cells, reduced early EPC apoptosis and enhanced their migration capacity towards SDF-1. Conclusions: The relative increase in circulating CD34+ cells and the low SDF-1 levels observed following rHDL infusions in ACS patients point towards a role of rHDL in cardiovascular repair mechanisms

Genome-wide gene expression profiling of stress response in a spinal cord clip compression injury model.

BackgroundThe aneurysm clip impact-compression model of spinal cord injury (SCI) is a standard injury model in animals that closely mimics the primary mechanism of most human injuries: acute impact and persisting compression. Its histo-pathological and behavioural outcomes are extensively similar to human SCI. To understand the distinct molecular events underlying this injury model we analyzed global mRNA abundance changes during the acute, subacute and chronic stages of a moderate to severe injury to the rat spinal cord.ResultsTime-series expression analyses resulted in clustering of the majority of deregulated transcripts into eight statistically significant expression profiles. Systematic application of Gene Ontology (GO) enrichment pathway analysis allowed inference of biological processes participating in SCI pathology. Temporal analysis identified events specific to and common between acute, subacute and chronic time-points. Processes common to all phases of injury include blood coagulation, cellular extravasation, leukocyte cell-cell adhesion, the integrin-mediated signaling pathway, cytokine production and secretion, neutrophil chemotaxis, phagocytosis, response to hypoxia and reactive oxygen species, angiogenesis, apoptosis, inflammatory processes and ossification. Importantly, various elements of adaptive and induced innate immune responses span, not only the acute and subacute phases, but also persist throughout the chronic phase of SCI. Induced innate responses, such as Toll-like receptor signaling, are more active during the acute phase but persist throughout the chronic phase. However, adaptive immune response processes such as B and T cell activation, proliferation, and migration, T cell differentiation, B and T cell receptor-mediated signaling, and B cell- and immunoglobulin-mediated immune response become more significant during the chronic phase.ConclusionsThis analysis showed that, surprisingly, the diverse series of molecular events that occur in the acute and subacute stages persist into the chronic stage of SCI. The strong agreement between our results and previous findings suggest that our analytical approach will be useful in revealing other biological processes and genes contributing to SCI pathology

Essential Role of Lyn in Fibrosis.

Fibrotic disorders involve replacement of normal parenchyma with myofibroblasts, which deposit connective tissue, leading to obliteration of the function of the underlying organ. The treatment options are inadequate and reflect the fact that signaling targets in myofibroblasts are unknown. Here we identify the hyperactive Lyn signaling in myofibroblasts of patients with chronic pancreatitis-induced fibrosis. Lyn activation coexpress with markers of activated myofibroblasts, and is increased ~11-fold in chronic pancreatitis compared to normal tissue. Inhibition of Lyn with siRNA or INNO-406 leads to the substantial decrease of migration and proliferation of human chronic pancreatitis myofibroblasts in vitro, while leaving migration and proliferation of normal myofibroblasts only slightly affected. Furthermore, inhibition of Lyn prevents synthesis of procollagen and collagen in myofibroblasts in a mouse model of chronic pancreatitis-induced fibrosis. We conclude that Lyn, as a positive regulator of myofibroblast migration, proliferation, and collagen production, is a key target for preventing fibrosis