SARS-CoV-2 pneumonia and bacterial pneumonia patients differ in a second hit immune response model

Secondary infections have been shown to complicate the clinical course and worsen the outcome of critically ill patients. Severe Coronavirus Disease 2019 (COVID-19) may be accompanied by a pronounced cytokine release, and immune competence of these patients towards most pathogenic antigens remains uncompromised early in the disease. Patients with bacterial sepsis also exhibit excessive cytokine release with systemic hyper-inflammation, however, typically followed by an anti-inflammatory phase, causing immune paralysis. In a second hit immune response model, leukocyte activation capacity of severely ill patients with pneumonia caused by SARS-CoV-2 or by bacteria were compared upon ICU admission and at days 4 and 7 of the ICU stay. Blood cell count and release of the pro-inflammatory cytokines IL-2, IFNγ and TNF were assessed after whole-blood incubation with the potent immune stimulus pokeweed mitogen (PWM). For comparison, patients with bacterial sepsis not originating from pneumonia, and healthy volunteers were included. Lymphopenia and granulocytosis were less pronounced in COVID-19 patients compared to bacterial sepsis patients. After PWM stimulation, COVID-19 patients showed a reduced release of IFNγ, while IL-2 levels were found similar and TNF levels were increased compared to healthy controls. Interestingly, concentrations of all three cytokines were significantly higher in samples from COVID-19 patients compared to samples from patients with bacterial infection. This fundamental difference in immune competence during a second hit between COVID-19 and sepsis patients may have implications for the selection of immune suppressive or enhancing therapies in personalized medicine

A high-precision polarimeter

We have built a polarimeter in order to measure the electron beam polarization in hall C at JLAB. Using a superconducting solenoid to drive the pure-iron target foil into saturation, and a symmetrical setup to detect the Moller electrons in coincidence, we achieve an accuracy of <1%. This sets a new standard for Moller polarimeters.Comment: 17 pages, 9 figures, submitted to N.I.

Antiinflammatory Therapy with Canakinumab for Atherosclerotic Disease

Background: Experimental and clinical data suggest that reducing inflammation without affecting lipid levels may reduce the risk of cardiovascular disease. Yet, the inflammatory hypothesis of atherothrombosis has remained unproved. Methods: We conducted a randomized, double-blind trial of canakinumab, a therapeutic monoclonal antibody targeting interleukin-1β, involving 10,061 patients with previous myocardial infarction and a high-sensitivity C-reactive protein level of 2 mg or more per liter. The trial compared three doses of canakinumab (50 mg, 150 mg, and 300 mg, administered subcutaneously every 3 months) with placebo. The primary efficacy end point was nonfatal myocardial infarction, nonfatal stroke, or cardiovascular death. RESULTS: At 48 months, the median reduction from baseline in the high-sensitivity C-reactive protein level was 26 percentage points greater in the group that received the 50-mg dose of canakinumab, 37 percentage points greater in the 150-mg group, and 41 percentage points greater in the 300-mg group than in the placebo group. Canakinumab did not reduce lipid levels from baseline. At a median follow-up of 3.7 years, the incidence rate for the primary end point was 4.50 events per 100 person-years in the placebo group, 4.11 events per 100 person-years in the 50-mg group, 3.86 events per 100 person-years in the 150-mg group, and 3.90 events per 100 person-years in the 300-mg group. The hazard ratios as compared with placebo were as follows: in the 50-mg group, 0.93 (95% confidence interval [CI], 0.80 to 1.07; P = 0.30); in the 150-mg group, 0.85 (95% CI, 0.74 to 0.98; P = 0.021); and in the 300-mg group, 0.86 (95% CI, 0.75 to 0.99; P = 0.031). The 150-mg dose, but not the other doses, met the prespecified multiplicity-adjusted threshold for statistical significance for the primary end point and the secondary end point that additionally included hospitalization for unstable angina that led to urgent revascularization (hazard ratio vs. placebo, 0.83; 95% CI, 0.73 to 0.95; P = 0.005). Canakinumab was associated with a higher incidence of fatal infection than was placebo. There was no significant difference in all-cause mortality (hazard ratio for all canakinumab doses vs. placebo, 0.94; 95% CI, 0.83 to 1.06; P = 0.31). Conclusions: Antiinflammatory therapy targeting the interleukin-1β innate immunity pathway with canakinumab at a dose of 150 mg every 3 months led to a significantly lower rate of recurrent cardiovascular events than placebo, independent of lipid-level lowering. (Funded by Novartis; CANTOS ClinicalTrials.gov number, NCT01327846.

Differential expression of P2X7 receptor and IL-1 beta in nociceptive and neuropathic pain

Background: Despite substantial progress, pathogenesis and therapy of chronic pain are still the focus of many investigations. The ATP-gated P2X7 receptor (P2X7R) has previously been shown to play a central role in animal models of nociceptive inflammatory and neuropathic pain. Recently, we found that the adaptive immune system is involved in the pathophysiology of chronic nociceptive and neuropathic pain in humans. So far, data regarding P2X7R expression patterns on cells of the adaptive immune system of pain patients are scarce. We therefore analyzed the P2X7R expression on peripheral blood lymphocytes and monocytes, as well as serum levels of IL-1 beta in patients suffering from chronic nociceptive and neuropathic pain in comparison to healthy volunteers in order to identify individuals who might benefit from a P2X7R modulating therapy. Methods: P2X7R messenger RNA (mRNA) and protein expression were determined in patients with either chronic nociceptive low back pain (CLBP) or neuropathic pain (NeP), and in healthy volunteers by quantitative real-time PCR (qPCR) and by fluorescence-assisted cell-sorting (FACS), respectively. IL-1 beta serum levels were measured with a multiplex cytokine assay. Results: Compared to healthy volunteers, P2X7R mRNA (1.6-fold, p = 0.038) and protein levels were significantly increased on monocytes (NeP: 24.6 +/- 6.2, healthy volunteers: 17.0 +/- 5.4;p = 0.002) and lymphocytes (NeP: 21.8 +/- 6.5, healthy volunteers: 15.6 +/- 5.2;p = 0.009) of patients with NeP, but not in patients with CLBP. Similarly, IL-1 beta serum concentrations were significantly elevated only in NeP patients (1.4-fold, p = 0.04). Conclusions: A significant upregulation of P2X7R and increased IL-1 beta release seems to be a particular phenomenon in patients with NeP. P2X7R inhibitors may therefore represent a potential option for the treatment of this frequently intractable type of pain

ATP release and autocrine signaling through P2X4 receptors regulate gamma delta T cell activation

Purinergic signaling plays a key role in a variety of physiological functions, including regulation of immune responses. Conventional alpha beta T cells release ATP upon TCR cross-linking; ATP binds to purinergic receptors expressed by these cells and triggers T cell activation in an autocrine and paracrine manner. Here, we studied whether similar purinergic signaling pathways also operate in the ``unconventional'' gamma delta T lymphocytes. We observed that gamma delta T cells purified from peripheral human blood rapidly release ATP upon in vitro stimulation with anti-CD3/CD28-coated beads or IPP. Pretreatment of gamma delta T cells with (10)panx-1, CBX, or Bf A reversed the stimulation-induced increase in extracellular ATP concentration, indicating that panx-1, connexin hemichannels, and vesicular exocytosis contribute to the controlled release of cellular ATP. Blockade of ATP release with (10)panx-1 inhibited Ca2+ signaling in response to TCR stimulation. qPCR revealed that gamma delta T cells predominantly express purinergic receptor subtypes A2a, P2X1, P2X4, P2X7, and P2Y11. We found that pharmacological inhibition of P2X4 receptors with TNP-ATP inhibited transcriptional up-regulation of TNF-alpha and IFN-gamma in gamma delta T cells stimulated with anti-CD3/CD28-coated beads or IPP. Our data thus indicate that purinergic signaling via P2X4 receptors plays an important role in orchestrating the functional response of circulating human gamma delta T cells. J. Leukoc. Biol. 92: 787-794; 2012

Sleep-disordered breathing in patients with newly diagnosed lung cancer

Abstract Background There are currently no data on the prevalence of sleep-disordered breathing (SDB) in patients with newly-diagnosed lung cancer. This might be of interest given that SDB is associated with increased cancer incidence and mortality. Furthermore, intermittent hypoxia has been linked with tumor growth and progression. The aim of the current study was to investigate the prevalence of SDB in patients with newly-diagnosed lung cancer. Methods Patients with newly-diagnosed lung cancer from three centers in Germany were screened for SDB using a two-channel screening system (ApneaLink™). SDB was defined as an apnea-hypopnea index of > 5/h, and was classified as mild if the AHI was 5–15/h whereas an AHI ≥15/h was classified as severe SDB. The presence of SDB-related symptoms was assessed using the Epworth Sleepiness Scale (ESS) and the Pittsburgh Sleep Quality Index (PSQI). Results A total of 100 patients were included. The overall prevalence of SDB was 49%; 32 patients (32%) had mild SDB with a median AHI of 7.7/h (quartile [Q1 5.4/h, Q3 10.4/h]) and a median oxygen desaturation index of 8.5 [Q1 4.2/h; Q3 13.4/h] and seventeen patients (17%) had moderate to severe SDB with a median AHI of 25.2 [Q1 18/h, Q3 45.5/h] and a median oxygen desaturation index of 20.6/h [Q1 9.6/h, Q3 36.6/h]. Patients with moderate to severe SDB had mild daytime sleepiness (ESS score 8.24 ± 3.96 vs. 5.74 ± 3.53 in those without SDB vs. 6.22 ± 2.72 in those with mild SDB; p = 0.0343). The PSQI did not differ significantly between the three groups (p = 0.1137). Conclusions This study showed a high prevalence of SDB in patients with newly-diagnosed lung cancer. In these patients SDB was associated with intermittent hypoxia and increased daytime sleepiness. Additional research is needed to determine whether SDB influences prognosis and morbidity in patients with lung cancer. Trial registration NCT02270853 (ClinicalTrials.gov), date of registration: 14th October 2014

ACE2-independent SARS-CoV-2 virus entry through cell surface GRP78 on monocytes – evidence from a translational clinical and experimental approachResearch in context

Summary: Background: SARS-CoV-2 infects host cells via an ACE2/TMPRSS2 entry mechanism. Monocytes and macrophages, which play a key role during severe COVID-19 express only low or no ACE2, suggesting alternative entry mechanisms in these cells. In silico analyses predicted GRP78, which is constitutively expressed on monocytes and macrophages, to be a potential candidate receptor for SARS-CoV-2 virus entry. Methods: Hospitalized COVID-19 patients were characterized regarding their pro-inflammatory state and cell surface GRP78 (csGRP78) expression in comparison to healthy controls. RNA from CD14+ monocytes of patients and controls were subjected to transcriptome analysis that was specifically complemented by bioinformatic re-analyses of bronchoalveolar lavage fluid (BALF) datasets of COVID-19 patients with a focus on monocyte/macrophage subsets, SARS-CoV-2 infection state as well as GRP78 gene expression. Monocyte and macrophage immunohistocytochemistry on GRP78 was conducted in post-mortem lung tissues. SARS-CoV-2 spike and GRP78 protein interaction was analyzed by surface plasmon resonance, GST Pull-down and Co-Immunoprecipitation. SARS-CoV-2 pseudovirus or single spike protein uptake was quantified in csGRP78high THP-1 cells. Findings: Cytokine patterns, monocyte activation markers and transcriptomic changes indicated typical COVID-19 associated inflammation accompanied by upregulated csGRP78 expression on peripheral blood and lung monocytes/macrophages. Subsequent cell culture experiments confirmed an association between elevated pro-inflammatory cytokine levels and upregulation of csGRP78. Interaction of csGRP78 and SARS-CoV-2 spike protein with a dissociation constant of KD = 55.2 nM was validated in vitro. Infection rate analyses in ACE2low and GRP78high THP-1 cells showed increased uptake of pseudovirus expressing SARS-CoV-2 spike protein. Interpretation: Our results demonstrate that csGRP78 acts as a receptor for SARS-CoV-2 spike protein to mediate ACE2-independent virus entry into monocytes. Funding: Funded by the Sino-German-Center for Science Promotion (C-0040) and the Germany Ministry BMWi/K [DLR-grant 50WB1931 and RP1920 to AC, DM, TW]