Anne-Claire Lukaszewicz

Monocytic HLA-DR expression in intensive care patients: interest for prognosis and secondary infection prediction.

Objectives: To test early measurement of human leukocyte antigen‐DR expression on circulating monocytes (mHLA‐DR) as prognostic marker, and the trend of mHLA‐DR recovery for the prediction of late secondary infection risk in a large intensive care unit population. Design: Prospective, observational study over 16 mos. Setting: Intensive care unit in a tertiary teaching hospital. Inclusion criteria: Simplified Acute Physiology Score II >15, age >18 yrs. Measurements and Main Results: The mHLA‐DR was measured by flow cytometry within the first 3 days and twice a week until discharge. We used a logistic regression model for outcome prediction, and a competing risk approach to test the relationship between mHLA‐DR recovery (log (mHLA‐DR) slope) and incidence of secondary infection. A total of 283 consecutive patients suffering from various pathologies were monitored (Simplified Acute Physiology Score II = 39, Sepsis‐related Organ Failure Assessment of 5 on day 0). Early mHLA‐DR was decreased in the whole population, however, more deeply in sepsis (p < .0001). Low mHLA‐DR was associated with mortality in the whole population (p = .003), as in subgroups (nonseptic, neurologic, and septic), but not when adjusted on Simplified Acute Physiology Score II. In patients with a length of stay of >7 days (n = 70), the lower the slope of mHLA‐DR recovery, the higher the incidence of the first secondary infection (adjusted on early mHLA‐DR, p = .04). Conclusions: For a given severity, mHLA‐DR proved not to a predictive marker of outcome, but a weak trend of mHLA‐DR recovery was associated with an increased risk of secondary infection. Monitoring immune functions through mHLA‐DR in intensive care unit patients therefore could be useful to identify a high risk of secondary infection.

Oxygen consumption of human peripheral blood mononuclear cells in severe human sepsis

Objective: During sepsis, after an initial stimulation immune cells down‐regulate their functions, leading to a state of immunosuppression. Because the mechanisms of such down‐regulation are unclear, we investigated the hypothesis of an energetic failure of immune cells to participate in immune dysfunction. Design: Cohort of septic shock patients to study peripheral blood mononuclear cells (PBMCs) biological energy in comparison to healthy volunteer cells. Setting: Critical care unit and laboratory, university hospital. Subjects: Eighteen severe sepsis or septic shock patients and 32 healthy volunteers. Interventions: Ex vivo measurement of oxygen consumption in PBMCs taken from patients. The PBMCs’ mitochondrial oxidative phosphorylation was investigated using adenosine diphosphate stimulation. The plasma factors implication was tested, using healthy cells incubated in septic plasma, or septic cells incubated in healthy plasma, at different time points of sepsis. The relationship between monocyte human leukocyte antigen‐DR expression and bioenergetic results was tested. Measurements and Main Results: Baseline oxygen consumption was higher in septic PBMCs (p < .01), with an attenuated response to adenosine diphosphate stimulation (p < .01). Oxygen consumption of healthy PBMCs incubated in septic plasma mimicked the septic cell response, with amplitude depending on the duration of sepsis (days 0–28). Septic cells incubated in healthy plasma partially recovered normal patterns. Septic plasma incubation increased the fraction of decoupling oxygen consumption (p = .021). A relationship between oxygen consumption (baseline or adenosine diphosphate stimulated) and human leukocyte antigen‐DR expression was observed for incubation with plasma sampled at different time points of septic shock. Conclusion: Energetic failure of PBMCs in sepsis may be a factor associated with the modulation of immune response and human leukocyte antigen‐DR phenotype, partially driven by plasma factors.

Extracorporeal life support for patients with acute respiratory distress syndrome: report of a Consensus Conference

The influenza H1N1 epidemics in 2009 led a substantial number of people to develop severe acute respiratory distress syndrome and refractory hypoxemia. In these patients, extracorporeal membrane oxygenation was used as rescue oxygenation therapy. Several randomized clinical trials and observational studies suggested that extracorporeal membrane oxygenation associated with protective mechanical ventilation could improve outcome, but its efficacy remains uncertain. Organized by the Société de Réanimation de Langue Française (SRLF) in conjunction with the Société Française d’Anesthésie et de Réanimation (SFAR), the Société de Pneumologie de Langue Française (SPLF), the Groupe Francophone de Réanimation et d’Urgences Pédiatriques (GFRUP), the Société Française de Perfusion (SOFRAPERF), the Société Française de Chirurgie Thoracique et Cardiovasculaire (SFCTV) et the Sociedad Española de Medecina Intensiva Critica y Unidades Coronarias (SEMICYUC), a Consensus Conference was held in December 2013 and a jury of 13 members wrote 65 recommendations to answer the five following questions regarding the place of extracorporeal life support for patients with acute respiratory distress syndrome: 1) What are the available techniques?; 2) Which patients could benefit from extracorporeal life support?; 3) How to perform extracorporeal life support?; 4) How and when to stop extracorporeal life support?; 5) Which organization should be recommended? To write the recommendations, evidence-based medicine (GRADE method), expert panel opinions, and shared decisions taken by all the thirteen members of the jury of the Consensus Conference were taken into account.

Exocytosis acts as a modulator of the ILT4-mediated inhibition of neutrophil functions

Significance Neutrophils are key components of inflammatory responses and immune defense against pathogens. Neutrophil functions are tightly controlled by surface receptors. Our study shows that the engagement of immunoglobulin-like transcript 4 (ILT4) inhibitory receptor impairs phagocytosis and respiratory burst of neutrophils. Moreover, we found that following induction of neutrophil granule exocytosis, ILT4 expression increases as a result of the rapid translocation of an intracellular pool to the cell surface. This increase of ILT4 expression enhances the ILT4-mediated inhibition of neutrophil activity. Finally, ILT4 up-regulation on neutrophils is impaired in sepsis patients. These results reveal a unique mechanism of regulation of neutrophil functions through ILT4 and its exocytosis that may have implications in inflammatory disorders and immune responses. Neutrophils play a major role in inflammatory responses and immune defense against pathogens. Even though expression of inhibitory receptors has been reported on neutrophils, their role remains poorly defined. Here we show that primary human neutrophils expressed immunoglobulin-like transcript 4 (ILT4) inhibitory receptor and that this expression was induced during differentiation of the myelomonoblast PLB-985 cell line into “neutrophil-like” cells. Functional assays indicated that human leukocyte antigen G, the preferred ligand of ILT4, inhibited the phagocytic function of neutrophils. ILT4 engagement also impaired reactive oxygen species production induced through CD32a and both receptors were found colocalized into neutrophil lipid rafts. Moreover, neutrophil degranulation induced through inflammatory stimuli increased ILT4 expression as a result of the rapid translocation of an intracellular pool to the cell surface. Consequently to this ILT4 up-regulation, the human leukocyte antigen G-mediated inhibition of neutrophil phagocytic function was enhanced. Finally, we found that ILT4 up-regulation induced on healthy donor neutrophils following stimulation was impaired in presence of plasma from patients with sepsis. Similarly, ILT4 up-regulation was inhibited in neutrophils from septic patients. Altogether, our results reveal a unique mechanism of regulation of neutrophil functions through ILT4 and its exocytosis that may have implications in inflammatory disorders.

Elevated production of radical oxygen species by polymorphonuclear neutrophils in cerebrospinal fluid infection

BackgroundCentral nervous system infection is a daily concern in neurointensive care; however, diagnosis remains difficult because classical criteria based on cerebrospinal fluid (CSF) analysis are difficult to interpret in post-trauma or neurosurgery patients after recent bleeding. A rapid, specific, sensitive test to diagnose CSF infection would help streamline therapeutic decisions in clinical practice and limit the risk of multiresistant bacteria. We hypothesized that polymorphonuclear neutrophil (PMN) phenotype and radical oxygen species (ROS) production in CSF may be specific to the presence of infection.MethodsThis study included 30 patients with suspected CSF infection with ventricular hemorrhage requiring external ventricular drainage, and 13 patients after trauma or surgery. Criteria for evaluating CSF infection included positive culture and > 100 leukocytes/mm3. Analysis of PMN phenotype was performed using flow cytometry (CD16, CD11b, and CD62L). ROS production was analyzed through luminometry (luminol).ResultsInfected CSF exhibited higher production of ROS compared with noninfected CSF. PMNs in CSF exhibited low CD16 and high annexin V expression, suggesting apoptosis.ConclusionsMeasurement of ROS production may discriminate infected from noninfected CSF. This simple test would be easy to employ in clinical practice to improve CSF infection management.

The relevance of pupillometry for evaluation of analgesia before noxious procedures in the intensive care unit.

BACKGROUND:Many patients in the intensive care unit are unable to communicate verbally. Accurately predicting whether such patients will exhibit painful behaviors during noxious procedures and assessing the adequacy of analgesia during those procedures is a challenge. In addition to observational pain assessment tools such as the Behavioral Pain Scale, physiologic indicators such as the pupillary response have been proposed. The pupil is innervated by both divisions of the autonomic nervous system and is affected by pain and analgesic medications. We evaluated the pupillary response to a light stimulus before noxious procedures as a method to predict pain during the procedure. METHODS:We correlated different aspects of the pupillary light reflex with established strategies for pain assessment to evaluate the adequacy of analgesia before surgical dressing changes performed in the intensive care unit in patients with cellulitis associated with mediastinitis or not. RESULTS:We found that a percentage of variation in pupil size >19% predicted the presence of pain as assessed by a Behavioral Pain Scale score of >3 with a sensitivity of 100% (95% confidence interval, 100%–100%) and a specificity of 77% (95% confidence interval, 54%–100%). CONCLUSIONS:In patients unable to communicate verbally, pupillometry may potentially guide caregivers to adjust analgesia before noxious procedures.

Modulation by Polymyxin-B Hemoperfusion of Inflammatory Response Related to Severe Peritonitis.

ABSTRACT Conflicting results have been reported on the influence of Polymyxin-B hemoperfusion treatment on systemic inflammation markers. The aim of the study was to assess in a randomized control trial the influence on plasma cytokine concentrations of Polymyxin-B hemoperfusion in septic shock due to peritonitis. A panel of 10 pro- or anti-inflammatory cytokines was measured in 213 patients with peritonitis-induced septic shock enrolled in the randomized trial ABDOMIX testing the impact of 2 Polymyxin-B hemoperfusion sessions with standard treatment. Gram-negative bacteria were identified in 69% of patients. In the overall population, baseline plasma cytokine concentrations were not different between the two groups. Circulating tumor necrosis factor-&agr;, interleukin (IL)-1&bgr;, IL-10, IL-6, and IL-1RA decreased significantly over time in both groups (P <0.0001 for all in controls, and P = 0.0002, 0.003, and <0.0001 in patients treated with Polymyxin-B hemoperfusion). IL-17A decreased significantly in patients treated with Polymyxin B hemoperfusion (P = 0.045) but not in controls. At the end of the second Polymyxin-B hemoperfusion session or at corresponding time in controls, plasma levels of cytokines did not differ between the two groups. Similar results were found in the subgroup of patients with gram-negative peritonitis who completed two Polymyxin-B hemoperfusion sessions. These results do not support a significant influence of Polymyxin-B hemoperfusion on circulating cytokines assessed except for IL-17A which clinical significance remains to be elucidated.

Downregulation of Blood Monocyte HLA-DR in ICU Patients Is Also Present in Bone Marrow Cells

Background The downregulation of blood monocyte HLA-DR expression also occurs in tissue infiltrative cells in a context of acute clinical inflammation, especially sepsis. This context favors the development of secondary infections and results from various mechanisms. Little is known about HLA-DR expression on bone marrow (BM) cells of the monocyte lineage, the source of circulating monocytes. This study analyzed the BM HLA-DR expression in ICU patients compared to BM monocytes from non-ICU patients and to blood monocytes of control healthy donors. A potential dysfunction of myeloid differentiation was investigated in a sub-population of these ICU patients to characterize the phenotype of the immature forms of monocytes and granulocytes in BM. Methods and Findings BM and blood were drawn from 33 ICU and 9 non-ICU patients having a BM analysis to precise the etiology of abnormal low count in blood cells. The data were compared with blood cells of 28 control donors. Flow cytometry was used for both HLA-DR expression and phenotyping of immature forms of monocytes and granulocytes. HLA-DR expression was downregulated in both blood and BM monocyte in ICU patients compared to BM of non-ICU patients and blood of control donors. Amplitude of HLA-DR downregulation was comparable in septic and non-septic ICU patients. The phenotype of immature forms of monocytes and granulocytes in BM (n = 11) did not show abnormal myeloid (monocyte + granulocyte) differentiation. Conclusion The downregulation of HLA-DR in BM monocyte lineage is present in ICU patients without major changes in myeloid cells. It may result from a regulation mediated by soluble and/or neuro-endocrine factors present in BM cell microenvironment.

Multicenter testing of the rapid quantification of radical oxygen species in cerebrospinal fluid to diagnose bacterial meningitis.

Purpose Meningitis is a serious concern after traumatic brain injury (TBI) or neurosurgery. This study tested the level of reactive oxygen species (ROS) in cerebrospinal fluid (CSF) to diagnose meningitis in febrile patients several days after trauma or surgery. Methods Febrile patients (temperature > 38°C) after TBI or neurosurgery were included prospectively. ROS were measured in CSF within 4 hours after sampling using luminescence in the basal state and after cell stimulation with phorbol 12-myristate 13-acetate (PMA). The study was conducted in a single-center cohort 1 (n = 54, training cohort) and then in a multicenter cohort 2 (n = 136, testing cohort) in the Intensive Care and Neurosurgery departments of two teaching hospitals. The performance of the ROS test was compared with classical CSF criteria, and a diagnostic decision for meningitis was made by two blinded experts. Results The production of ROS was higher in the CSF of meningitis patients than in non-infected CSF, both in the basal state and after PMA stimulation. In cohort 1, ROS production was associated with a diagnosis of meningitis with an AUC of 0.814 (95% confidence interval (CI) [0.684–0.820]) for steady-state and 0.818 (95% CI [0.655–0.821]) for PMA-activated conditions. The best threshold value obtained in cohort 1 was tested in cohort 2 and showed high negative predictive values and low negative likelihood ratios of 0.94 and 0.36 in the basal state, respectively, and 0.96 and 0.24 after PMA stimulation, respectively. Conclusion The ROS test in CSF appeared suitable for eliminating a diagnosis of bacterial meningitis.

Postoperative lymphopenia: An independent risk factor for postoperative pneumonia after lung cancer surgery, results of a case-control study

Objective Postoperative lymphopenia has been proposed as a risk factor for postoperative infections but has never been identified as such in a multivariate analysis. Postoperative pneumonia (POP) is one of the most common complications after lung cancer surgery and is associated with a worse outcome. We aimed to evaluate the association between postoperative lymphopenia and POP after lung cancer surgery. Methods Patients admitted for lung cancer surgery (lobectomy, bilobectomy, or pneumonectomy) aged ≥ 18 years and with no history of an immunosuppressive state were eligible for inclusion. Lymphocyte counts were determined in blood drawn on the day before surgery and at postoperative days 1, 3 and 7. POP diagnosis was based on clinical, biological and radiological data. A logistic regression model adjusted on currently described risk factors for POP was used to explain the onset of this condition. Results Two hundred patients were included, of whom 43 (21.5%) developed POP. Preoperative lymphocyte count was 1.8±0.6x109 cells/L and 2.0±0.7x109 cells/L in patients with and without POP, respectively (P = .091). In both groups, the lymphocyte count nadir occurred at postoperative day 1. In multivariate analysis, lymphopenia at postoperative day 1 was significantly associated with increased risk of POP (odds ratio: 2.63, 95% CI [1.03–5.40]). POP rate at postoperative day 7 was higher in patients presenting low lymphocyte counts (≤1.19x109 cells/L) at postoperative day 1 (P = .003). Conclusions Our study showed that lymphopenia following lung cancer surgery was maximal at postoperative day 1 and was associated with POP.