CD8+ and Regulatory T cells Differentiate Tumor Immune Phenotypes and Predict Survival in Locally Advanced Head and Neck Cancer

Background: The tumor immune status “inflamed”, “immune excluded”, and “desert” might serve as a predictive parameter. We studied these three cancer immune phenotypes while using a simple immunohistochemical algorithm. Methods: Pre-treatment tissue samples of 280 patients with locally advanced HNSCC treated with radiochemotherapy were analyzed. A double staining of CD8+ cytotoxic T cells (CTL) and FoxP3+ (Treg) was performed and the cell density was evaluated in the intraepithelial and stromal compartment of the tumor. Results: The classification of tumors as “immune desert” when stromal CTL were ≤ 50 cells/mm2, “inflamed” when intraepithelial CTL were > 500 cells/mm2, and as “excluded” when neither of these definitions met these cut off values allowed the best discrimination regarding overall survival. These groups had median OS periods of 37, 61, and 85 months, respectively. In “immune desert” and “immune excluded” tumors high Treg tended to worsen OS, but in “inflamed” tumors high Treg clearly improved OS. Conclusions: We propose that, in locally advanced HNSCC, the tumor immune state “inflamed”, “immune excluded”, and “immune desert” can be defined by intraepithelial and stromal CTL. Tregs can further subdivide these groups. The opposing effects of Tregs in the different groups might be the reason for the inconsistency of Tregs prognostic values published earlier

Impact of the rpoS genotype for acid resistance patterns of pathogenic and probiotic Escherichia coli

BACKGROUND: Enterohemorrhagic E. coli (EHEC), a subgroup of Shiga toxin (Stx) producing E. coli (STEC), may cause severe enteritis and hemolytic uremic syndrome (HUS) and is transmitted orally via contaminated foods or from person to person. The infectious dose is known to be very low, which requires most of the bacteria to survive the gastric acid barrier. Acid resistance therefore is an important mechanism of EHEC virulence. It should also be a relevant characteristic of E. coli strains used for therapeutic purposes such as the probiotic E. coli Nissle 1917 (EcN). In E. coli and related enteric bacteria it has been extensively demonstrated, that the alternative sigma factor σ(S), encoded by the rpoS gene, acts as a master regulator mediating resistance to various environmental stress factors. METHODS: Using rpoS deletion mutants of a highly virulent EHEC O26:H11 patient isolate and the sequenced prototype EHEC EDL933 (ATCC 700927) of serotype O157:H7 we investigated the impact of a functional rpoS gene for orchestrating a satisfactory response to acid stress in these strains. We then functionally characterized rpoS of probiotic EcN and five rpoS genes selected from STEC isolates pre-investigated for acid resistance. RESULTS: First, we found out that ATCC isolate 700927 of EHEC EDL933 has a point mutation in rpoS, not present in the published sequence, leading to a premature stop codon. Moreover, to our surprise, one STEC strain as well as EcN was acid sensitive in our test environment, although their cloned rpoS genes could effectively complement acid sensitivity of an rpoS deletion mutant. CONCLUSION: The attenuation of sequenced EHEC EDL933 might be of importance for anyone planning to do either in vitro or in vivo studies with this prototype strain. Furthermore our data supports recently published observations, that individual E. coli isolates are able to significantly modulate their acid resistance phenotype independent of their rpoS genotype

Complement activation via the lectin and alternative pathway in patients with severe COVID-19

Complement plays an important role in the direct defense to pathogens, but can also activate immune cells and the release of pro-inflammatory cytokines. However, in critically ill patients with COVID-19 the immune system is inadequately activated leading to severe acute respiratory syndrome (SARS) and acute kidney injury, which is associated with higher mortality. Therefore, we characterized local complement deposition as a sign of activation in both lungs and kidneys from patients with severe COVID-19. Using immunohistochemistry we investigated deposition of complement factors C1q, MASP-2, factor D (CFD), C3c, C3d and C5b-9 as well as myeloperoxidase (MPO) positive neutrophils and SARS-CoV-2 virus particles in lungs and kidneys from 38 patients who died from COVID-19. In addition, tissue damage was analyzed using semi-quantitative scores followed by correlation with complement deposition. Autopsy material from non-COVID patients who died from cardiovascular causes, cerebral hemorrhage and pulmonary embolism served as control (n=8). Lung injury in samples from COVID-19 patients was significantly more pronounced compared to controls with formation of hyaline membranes, thrombi and edema. In addition, in the kidney tubular injury was higher in these patients and correlated with lung injury (r=0.361*). In autopsy samples SARS-CoV-2 spike protein was detected in 22% of the lungs of COVID-19 patients but was lacking in kidneys. Complement activation was significantly stronger in lung samples from patients with COVID-19 via the lectin and alternative pathway as indicated by deposition of MASP-2, CFD, C3d and C5b9. Deposits in the lung were predominantly detected along the alveolar septa, the hyaline membranes and in the alveolar lumina. In the kidney, complement was significantly more deposited in patients with COVID-19 in peritubular capillaries and tubular basement membranes. Renal COVID-19-induced complement activation occurred via the lectin pathway, while activation of the alternative pathway was similar in both groups. Furthermore, MPO-positive neutrophils were found in significantly higher numbers in lungs and kidneys of COVID-19 patients and correlated with local MASP-2 deposition. In conclusion, in patients who died from SARS-CoV-2 infection complement was activated in both lungs and kidneys indicating that complement might be involved in systemic worsening of the inflammatory response. Complement inhibition might thus be a promising treatment option to prevent deregulated activation and subsequent collateral tissue injury in COVID-19

Prolonged lipopolysaccharide-induced illness elevates glucagon-like peptide-1 and suppresses peptide YY:A human-randomized cross-over trial

Severe systemic inflammation is associated with nausea, loss of appetite, and delayed gastric emptying, which increases hospitalization admission length and mortality rate. There is a lack of human controlled studies exploring gastric emptying rates and underlying mechanisms during inflammatory conditions. We aimed to investigate if systemic inflammation in young men delays gastro‐intestinal transit times, lowers motility, and affects gastrointestinal hormone secretion. This substudy of a randomized crossover trial investigated eight healthy young men on two separate occasions; (I) following an overnight fast (healthy conditions/HC) and (II) fasting and bedrest combined with two lipopolysaccharide (LPS) injections of 1 ng kg(−1) following an overnight fast and 0.5 ng kg(−1) following another 24 h (systemic inflammation/SI). A standardized protein beverage and a SmartPill capsule (a wireless gastrointestinal monitoring system) were swallowed during each occasion. Whole gut transit time was comparable between HC and SI. SI decreased gastric mean pressure peak amplitude (p = 0.04) and increased pH rise across the pylorus and small bowel pH (p = 0.02) compared with HC. Glucagon‐like peptide‐1 was elevated during SI compared with HC (p = 0.04). Peptide YY was lower during SI compared with HC (p = 0.007). Prolonged LPS exposure combined with fasting and bedrest elevated glucagon‐like peptide 1 concentrations, which may play a role for the nausea and loss of appetite typically associated with SI

Non-professional phagocytosis: a general feature of normal tissue cells

Non-professional phagocytosis by cancer cells has been described for decades. Recently, non-professional phagocytosis by normal tissue cells has been reported, which prompted us to take a closer look at this phenomenon. Non-professional phagocytosis was studied by staining cultured cells with live-cell staining dyes or by staining paraffin-embedded tissues by immunohistochemistry. Here, we report that each of 21 normal tissue cell lines from seven different organs was capable of phagocytosis, including ex vivo cell cultures examined before the 3rd passage as well as the primary and virus-transformed cell lines. We extended our analysis to an in vivo setting, and we found the occurrence of non-professional phagocytosis in healthy skin biopsies immediately after resection. Using dystrophin immunohistochemistry for membrane staining, human post-infarction myocardial tissue was assessed. We found prominent signs of non-professional phagocytosis at the transition zone of healthy and infarcted myocardia. Taken together, our findings suggest that non-professional phagocytosis is a general feature of normal tissue cells

Simultaneous Effects of Light Intensity and Phosphorus Supply on the Sterol Content of Phytoplankton

Sterol profiles of microalgae and their change with environmental conditions are of great interest in ecological food web research and taxonomic studies alike. Here, we investigated effects of light intensity and phosphorus supply on the sterol content of phytoplankton and assessed potential interactive effects of these important environmental factors on the sterol composition of algae. We identified sterol contents of four common phytoplankton genera, Scenedesmus, Chlamydomonas, Cryptomonas and Cyclotella, and analysed the change in sterol content with varying light intensities in both a high-phosphorus and a low-phosphorus approach. Sterol contents increased significantly with increasing light in three out of four species. Phosphorus-limitation reversed the change of sterol content with light intensity, i.e., sterol content decreased with increasing light at low phosphorus supply. Generally sterol contents were lower in low-phosphorus cultures. In conclusion, both light and phosphorus conditions strongly affect the sterol composition of algae and hence should be considered in ecological and taxonomic studies investigating the biochemical composition of algae. Data suggest a possible sterol limitation of growth and reproduction of herbivorous crustacean zooplankton during summer when high light intensities and low phosphorus supply decrease sterol contents of algae

Czochralski Silicon as a Detector Material for S-LHC Tracker Volumes

4 pages, 6 figures, 12th Vienna Conference on InstrumentationWith an expected ten-fold increase in luminosity in S-LHC, the radiation environment in the tracker volumes will be considerably harsher for silicon-based detectors than the already harsh LHC environment. Since 2006, a group of CMS institutes, using a modified CMS DAQ system, has been exploring the use of Magnetic Czochralski silicon as a detector element for the strip tracker layers in S-LHC experiments. Both p+/n-/n+ and n+/p-/p+ sensors have been characterized, irradiated with proton and neutron sources, assembled into modules, and tested in a CERN beamline. There have been three beam studies to date and results from these suggest that both p+/n-/n+ and n+/p-/p+ Magnetic Czochralski silicon are sufficiently radiation hard for the $R>25$ cm regions of S-LHC tracker volumes. The group has also explored the use of forward biasing for heavily irradiated detectors, and although this mode requires sensor temperatures less than -50\,$^\circ$C, the charge collection efficiency appears to be promising.Peer reviewe