Increased STAT3 phosphorylation on CD27(+) B-cells from common variable immunodeficiency disease patients.

Maturation and differentiation of B-cells are driven by T-cells' help through IL-21/STAT3 axis in GC centers or through extrafollicular pathways, in a T-independent manner. B-cell differentiation is defective in common variable immunodeficiency disease (CVID) patients. We investigated if IL-21/STAT3 axis alterations could influence B-cell fate. We activated purified CVID B-cells with surrogate T-dependent (anti-CD40), T-independent (TLR-9 ligand) stimuli or through B-cell receptor engagement (anti-IgM) with or without IL-21. IL-21 mediated STAT3 activation was greater on CD27(-) than CD27(+) B-cells depending on the stimulus. IL-21 alone induced STAT3 phosphorylation (pSTAT3) only on CD27(-) B-cells and IL-21 induced higher pSTAT3 levels on CD27(-) than CD27(+) B-cells after anti-IgM or anti-CD40 activation. CVID CD27(+) B-cells showed selective STAT3 hyperphosphorylation after activation with anti-IgM or anti-CD40 alone and anti-IgM, anti-CD40 or ODN combined with IL-21. Increased STAT3 activation during immune responses could result in B-cell differentiation defects in CVID.

Expansion of myeloid-derived suppressor cells in chronic obstructive pulmonary disease and lung cancer: potential link between inflammation and cancer.

BACKGROUND: Chronic obstructive pulmonary disease (COPD) is a risk factor for lung cancer (LC). Myeloid-derived suppressor cells (MDSCs) down-regulate the T cell receptor ζ chain (TCR ζ) through L-arginine deprivation and lead to T cell dysfunction and deficient antitumor immunity. We hypothesized that abnormally high levels of MDSCs in COPD patients may alter tumor immunosurveillance. METHODS: We compared the proportion of circulating MDSCs (Lin-HLA-DR-/CD33+/CD11b+) (by flow cytometry), arginase I (ARG I) serum levels (by ELISA), and expression levels of TCR ζ on circulating lymphocytes (by flow cytometry) in 28 patients with LC, 62 subjects with COPD, 41 patients with both LC and COPD, 40 smokers with normal spirometry and 33 non-smoking controls. T cell proliferation assays were performed in a subgroup of participants (CFSE dilution protocol). RESULTS: We found that: (1) circulating MDSCs were up-regulated in COPD and LC patients (with and without COPD); (2) MDSCs expansion was associated with TCR ζ down-regulation in the three groups; (3) in LC patients, these findings were independent of COPD and tobacco smoking exposure; (4) TCR ζ down-regulation correlates with T cell hyporesponsiveness in COPD and LC patients. CONCLUSIONS: These results suggest that tumor immunosurveillance might be impaired in COPD and may contribute to the increased risk of LC reported in these patients.

Differential effects of smoking and COPD upon circulating myeloid derived suppressor cells.

BACKGROUND: Chronic obstructive pulmonary disease (COPD) is characterized by an enhanced and persistent innate and acquired immune response to tobacco smoking. Myeloid-derived suppressor cells (MDSCs) modulate T-cell responses by down-modulating the T cell receptor ζ chain (TCR ζ) through the catabolism of l-arginine. The effects of smoking on MDSCs and their potential participation in COPD immunopathogenesis have not been explored so far. METHODS: To investigate it, we compared the level of circulating Lineage-/HLA-DR-/CD33+/CD11b+ MDSCs, the serum concentration of arginase I (ARG I) and the expression of peripheral T-cell receptor ζ chain (TCR ζ) in never smokers, smokers with normal spirometry and COPD patients. Flow cytometry was used to quantify circulating MDSCs and TCR ζ expression. Serum ARG I levels were determined by ELISA. RESULTS: The main findings of this study were that: (1) current smoking upregulates and activates circulating MDSCs both in smoker controls and COPD patients; and, (2) at variance with the smokers with normal spirometry, in patients with COPD this effect persists after quitting smoking and is accompanied by a significant and specific down-regulation of the TCR ζ chain expression in circulating T lymphocytes. CONCLUSION: Smoking modulates circulating MDSCs. Their regulation appears altered in patients with COPD.

Lack of correlation among intracerebral cytokines, intracranial pressure, and brain tissue oxygenation in patients with traumatic brain injury and diffuse lesions.

OBJECTIVES: To determine the evolution of cytokine patterns using microdialysis in patients with traumatic brain injury with diffuse lesions and to study the relationship between cytokines and intracranial pressure, brain tissue oxygenation and lesion type on the computed cranial tomography scan (patients with and without brain swelling). DESIGN: Prospective and observational study. SETTING: Third-level university hospital. PATIENTS: Patients between 15 and 65 yrs with severe traumatic brain injury and a diffuse lesion requiring intracranial pressure and brain tissue oxygenation monitoring were eligible. INTERVENTIONS: Microdialysis catheters with a high-cutoff membrane of 100 kDa were inserted. RESULTS: Sixteen patients were included in the analysis. There was a substantial interindividual variability between cytokine values. The highest concentrations for the interleukin-1ß, interleukin-6, and interleukin-8 were measured during the first 24 hrs followed by a gradual decline. The average concentration for interleukin-10 did not vary over time. This pattern is the most frequent in patients with traumatic brain injury with diffuse lesions. The intracranial pressure-cytokines correlation coefficients for the 16 patients varied substantially: interleukin-1ß-intracranial pressure (-0.76 to 0.63); interleukin-6-intracranial pressure (-0.83 to 0.78); interleukin-8-intracranial pressure (-0.86 to 0.84); and interleukin-10-intracranial pressure (-0.36 to 0.65). The brain tissue oxygenation-cytokine correlation coefficients, like with intracranial pressure, also varied between patients: interleukin-1ß-brain tissue oxygenation (-0.49 to 0.68), interleukin-6-brain tissue oxygenation (-0.99 to 0.84); interleukin-8-brain tissue oxygenation (-0.65 to 0.74); and interleukin-10-brain tissue oxygenation (-0.34 to 0.52). Similarly, we found no difference in the cytokine values inpatient microdialysis with and without swelling in the computed tomographic scan. CONCLUSIONS: No clear relationship was found between the temporal pattern of cytokines and the behavior of the intracranial pressure, brain tissue oxygenation, and the presence or absence of swelling in the computed tomography scan. This study demonstrates the feasibility of microdialysis in recovering cytokines for a prolonged time, although there may be some nonresolved methodologic problems with this technique when we try to study the inflammation during traumatic brain injury that could affect the results and make interpretation of microdialysis data prone to difficulties.

Expression of toll-like receptors 2 and 4 is upregulated during hospital admission in traumatic patients: lack of correlation with blunted innate immune responses.

BACKGROUND: There are reports with conflicting results on the expression of toll-like receptors (TLRs) in trauma patients. In addition, these studies analyzed TLR expression only at patients' hospital admission but not later when complications usually arise. OBJECTIVES: To analyze the surface expression of TLR2 and TLR4 on circulating monocytes from trauma patients during the hospitalization period and to correlate this with cytokine production after stimulation with TLR2 and TLR4 agonists. The phagocytic capacity of monocytes was analyzed at the same time points of TLR expression analysis; to correlate these molecular findings with the presence or absence of infections. METHODS: Prospective and observational study from June 2005 to June 2007. In all analysis, a control group composed of healthy subjects was included. RESULTS: We studied 70 trauma patients admitted to the intensive care unit (ICU) of a tertiary hospital, and 30 healthy volunteers. Blood samples were collected at hospital admission, on day 7 and 14. Forty-four patients (63%) developed at least one episode of infection. Monocytes from trauma patients expressed higher levels of TLR2 and TLR4 than monocytes from control subjects at all time points. Expression of TLR2 and TLR4 in monocytes from those patients who developed any infection was significantly lower than in those patients without infection but still significantly higher than in control subjects. Cellular responses to TLR4 agonist were impaired. Monocytes from traumatic patients phagocytosized less efficiently than monocytes from control subjects. CONCLUSIONS: These results indicate that trauma patients present a dysregulation of the innate immune system that persists during the first 14 days after hospital admission.

C3 promotes clearance of Klebsiella pneumoniae by A549 epithelial cells.

The airway epithelium represents a primary site for contact between microbes and their hosts. To assess the role of complement in this event, we studied the interaction between the A549 cell line derived from human alveolar epithelial cells and a major nosocomial pathogen, Klebsiella pneumoniae, in the presence of serum. In vitro, we found that C3 opsonization of poorly encapsulated K. pneumoniae clinical isolates and an unencapsulated mutant enhanced dramatically bacterial internalization by A549 epithelial cells compared to highly encapsulated clinical isolates. Local complement components (either present in the human bronchoalveolar lavage or produced by A549 epithelial cells) were sufficient to opsonize K. pneumoniae. CD46 could competitively inhibit the internalization of K. pneumoniae by the epithelial cells, suggesting that CD46 is a receptor for the binding of complement-opsonized K. pneumoniae to these cells. We observed that poorly encapsulated strains appeared into the alveolar epithelial cells in vivo but that (by contrast) they were completely avirulent in a mouse model of pneumonia compared to the highly encapsulated strains. Our results show that bacterial opsonization by complement enhances the internalization of the avirulent microorganisms by nonphagocytic cells such as A549 epithelial cells and allows an efficient innate defense.

Gammadelta T lymphocytes from cystic fibrosis patients and healthy donors are high TNF-alpha and IFN-gamma-producers in response to Pseudomonas aeruginosa.

BACKGROUND: Gammadelta T cells have an important immunoregulatory and effector function through cytokine release. They are involved in the responses to Gram-negative bacterium and in protection of lung epithelium integrity. On the other hand, they have been implicated in airway inflammation. METHODS: The aim of the present work was to study intracytoplasmic IL-2, IL-4, IFN-gamma and TNF-alpha production by gammadelta and alphabeta T lymphocytes from cystic fibrosis patients and healthy donors in response to Pseudomonas aeruginosa (PA). Flow cytometric detection was performed after peripheral blood mononuclear cells (PBMC) culture with a cytosolic extract from PA and restimulation with phorbol ester plus ionomycine. Proliferative responses, activation markers and receptor usage of gammadelta T cells were also evaluated. RESULTS: The highest production of cytokine was of TNF-alpha and IFN-gamma, gammadelta being better producers than alphabeta. No differences were found between patients and controls. The Vgamma9delta2 subset of gammadelta T cells was preferentially expanded. CD25 and CD45RO expression by the alphabeta T subset and PBMC proliferative response to PA were defective in cystic fibrosis lymphocytes. CONCLUSION: Our results support the hypothesis that gammadelta T lymphocytes play an important role in the immune response to PA and in the chronic inflammatory lung reaction in cystic fibrosis patients. They do not confirm the involvement of a supressed Th1 cytokine response in the pathogenesis of this disease.

Skewed inhibitory receptors expression in a TAP2-deficient patient.

Most of patients suffering from HLA class I deficiency due to mutations in TAP genes show a significative increase of the peripheral minor Vdelta1+ subpopulation of gammadelta T cells. Surface expression of inhibitory receptors (IR) for HLA class I molecules have been mainly attributed to Vdelta2+ gammadelta T clones. In this study we have analysed the expression of these receptors in both subsets of gammadelta T peripheral lymphocytes. We studied 16 healthy controls and a reported case of homozygous TAP2 mutation with a marked increase of Vdelta1+ gammadelta T cells. MICA/B presence in monocytes was also evaluated. In healthy subjects, the expression of CD94 and CD94/NKG2A was higher in the Vdelta2+ subset but cells bearing the IR ILT2 were found increased in the Vdelta1+. The patient Vdelta2+ gammadelta T cells showed the same IR expression than normal controls, in contrast the Vdelta1+ subset presented a special pattern of very high expression of CD94 and ILT2 and low of CD94/NKG2A. The presence of a new IR poorly represented in healthy individuals could account for the selective increase of Vdelta1+ gammadelta T in TAP-deficient patients. MICA/B surface expression in monocytes was not shifted in our patient.