Response of murine gamma delta T cells to the synthetic polypeptide poly-Glu50Tyr50

Random heterocopolymers of glutamic acid and tyrosine (pEY) evoke strong, genetically controlled immune responses in certain mouse strains. We found that pE50Y50 also stimulated polyclonal proliferation of normal gamma delta, but not alpha beta, T cells. Proliferation of gamma delta T cells did not require prior immunization with this Ag nor the presence of alpha beta T cells, but was enhanced by IL-2. The gamma delta T cell response proceeded in the absence of accessory cells, MHC class II, beta 2-microglobulin, or TAP-1, suggesting that Ag presentation by MHC class I/II molecules and peptide processing are not required. Among normal splenocytes, as with gamma delta T cell hybridomas, the response was strongest with V gamma 1+ gamma delta T cells, and in comparison with related polypeptides, pE50Y50 provided the strongest stimulus for these cells. TCR gene transfer into a TCR-deficient alpha beta T cell showed that besides the TCR, no other components unique to gamma delta T cells are needed. Furthermore, interactions between only the T cells and pE50Y50 were sufficient to bring about the response. Thus, pE50Y50 elicited a response distinct from those of T cells to processed/presented peptides or superantigens, consistent with a mechanism of Ig-like ligand recognition of gamma delta T cells. Direct stimulation by ligands resembling pE50Y50 may thus selectively evoke contributions of gamma delta T cells to the host response.     

Role of gammadelta T cells in protecting normal airway function

Since their discovery 15 years ago, the role of gammadelta T cells has remained somewhat elusive. Responses of gammadelta T cells have been found in numerous infectious and non-infectious diseases. New evidence points to gammadelta T cells' functioning in the airways to maintain normal airway responsiveness or tone. In the lung, distinct subsets of gammadelta T cell subsets seem to have specific roles, one subset promoting allergic inflammation, the other serving a protective role.     

Interferon-alpha acutely impairs sleep in healthy humans

We investigated the effects of two low doses of interferon-alpha (IFN-alpha) on nocturnal sleep in 18 healthy men by means of polysomnographic sleep recordings. At 1900h, human recombinant IFN-alpha (1000 or 10000 U/kg body weight) or placebo was administered subcutaneously. Between 2300h and 0700h subjects were allowed to sleep. In general effects were stronger at the dose of 10000 than 1000 U/kg body weight of IFN-alpha. Although, after IFN-alpha subjects experienced increased fatigue, the cytokine impaired the quality of nocturnal sleep. The higher dose of IFN-alpha suppressed slow wave sleep (17.8 +/- 2.0% vs 25.2 +/- 2.6% following placebo, P<0.003) but increased time spent in shallow sleep (P<0.05) during the first half of sleep time. Rapid eye movement (REM) sleep latency was postponed (P<0.02) and time spent in REM sleep was significantly decreased after IFN-alpha (P<0.04). The impairing influence of IFN-alpha on sleep in humans is in contrast with findings of sleep promoting effects of this cytokine in animals. Our data suggest that endogenous IFN-alpha may be a factor responsible for alterations of sleep, e.g. in the course of viral infections.     

The Laptev Sea walrus Odobenus rosmarus laptevi: an enigma revisited

The walrus ( Odobenus rosmarus ) is in some current systematic schemes divided into three subspecies: O. r. rosmarus in the North Atlantic, O.   r. divergens in the North Pacific and O.   r. laptevi in the Laptev Sea. These three subspecies have been described as differing in body size, but the taxonomic status of O.   r. laptevi is disputed. The current study applies molecular and morphometric methods to assess the taxonomic status of O.   r. laptevi and to analyse the systematic and phylogeographic relationships between the three purported walrus subspecies. Tusk length and tusk circumference were measured from the few skulls available of O.   r. laptevi , and the obtained values were within the ranges reported for Pacific walruses. Thus, morphologically, subspecies status for O.   r. laptevi is not supported according to the Amadon–Mayr '75% rule'. Phylogenetic analyses and haplotype networks based on mitochondrial nucleotide sequence data of NADH dehydrogenase 1, 16S rRNA, cytochrome oxidase I and the d -loop of the control region of the historic O.   r. laptevi bone material and contemporary O.   r. rosmarus and O.   r. divergens showed that the Laptev Sea walrus groups with individuals from the North Pacific. Thus, the mitochondrial sequence data do not support the recognition of three walrus subspecies as reciprocally monophyletic evolutionary units with independent evolutionary histories. Only O.   r. rosmarus and O.   r. divergens meet this criterion with the present sampling. Accordingly, we recommend that Odobenus r. laptevi be abandoned and the Laptev walrus instead be recognized as the westernmost population of the Pacific walrus, Odobenus r. divergens. However, further research is recommended to assess whether the Laptev walrus could be considered as a significant unit in terms of conservation and management, since it is unique in several ecological parameters.     

A new method for sustained generation of ultra-pure nitric oxide-containing gas mixtures via controlled UVA-photolysis of nitrite solutions

Exogenous gaseous nitric oxide (gNO) is an FDA approved drug for treatment of a variety of human pathologies like Persistent Pulmonary Hypertension in neonates and premature babies, skin lesions and fungal dermatophyte infections. Substantial disadvantages of current gNO-based therapies are the high therapy costs, high storage costs of the gas cylinders, and the rapid contamination of compressed NO gases with various decomposition products. Here we describe a new, very simple, and inexpensive photolytic generator of uncontaminated NO-containing gas mixtures at therapeutic concentrations. The new method bases on UVA-induced and redox-assisted decomposition of nitrite ions in aqueous solutions. NO formation via UVA-induced photolysis of nitrite is accompanied by an OH radical-dependent production of NO₂ that beside its toxic character additionally strongly reduces the NO yield by consuming NO in its reaction to N₂O₃. During the UVA-induced photodecomposition process both, inhibition of NO₂ formation or NO₂ depletion by antioxidants hinders the NO-consuming reaction with NO₂ and ensured a maximal purity and maximal yield of NO-containing gas mixtures. Therefore, NO-containing gas mixtures generated by the described method are suitable for medical applications like inhalation or gassing of chronic non-healing wounds. Control of temperature, UVA intensity and composition of the reaction mixture allows facile control over the final NO level in the carrier gas over a wide concentration range. We demonstrate the sustained and stable release of NO over a wide dynamic range (10–5000 ppm NO) for many hours. The method avoids contamination-prone long time storage of NO gas. As such, it appears particularly relevant for applications involving the additional presence of oxygen (e.g. inhalation).     

Oral cancer,cigarette smoke and mitochondrial 18 kDa translocator protein (TSPO) — In vitro,in vivo,salivary analysis

Oral cancer features high rates of mortality and morbidity, and is in dire need for new approaches. In the present study we analyzed 18 kDa translocator protein (TSPO) expression in oral (tongue) cancer tumors by immunohistochemistry. We also assayed TSPO binding in human tongue cancer cell lines and in the cellular fraction of saliva from tongue cancer patients, heavy cigarette smokers, and non-smoking healthy people as controls. Concurrently, TSPO protein levels, cell viability, mitochondrial membrane potential (Δψ_m), and general protein levels were analyzed. TSPO expression could be significantly enhanced in oral cancer tumors, compared to unaffected adjacent tissue. We also found that five-year survival probability dropped from 65% in patients with TSPO negative tumors to 7% in patients with highly expressed TSPO (p < 0.001). TSPO binding capacity was also pronounced in the human oral cancer cell lines SCC-25 and SCC-15 (3133 ± 643 fmol/mg protein and 6956 ± 549 fmol/mg protein, respectively). Binding decreased by 56% and 72%, in the SCC-25 and SCC-15 cell lines, respectively (p < 0.05) following CS exposure in cell culture. In the cellular fraction of saliva of heavy smokers TSPO binding was lower than in non-smokers (by 53%, p < 0.05). Also the cellular fraction of saliva exposed to CS in vitro showed decreased TSPO binding compared to unexposed saliva (by 30%, p < 0.001). Interestingly, oral cancer patients also displayed significantly lower TSPO binding in the cellular fraction of saliva compared to healthy controls (by 40%, p < 0.01). Our results suggest that low TSPO binding found in the cellular fraction of saliva may depend on genetic background as well as result from exposure to CS. We suggest that this may be related to a predisposition for occurrence of oral cancer.     

Intestinal crypt homeostasis results from neutral competition between symmetrically dividing Lgr5 stem cells

Intestinal stem cells, characterized by high Lgr5 expression, reside between Paneth cells at the small intestinal crypt base and divide every day. We have carried out fate mapping of individual stem cells by generating a multicolor Cre-reporter. As a population, Lgr5(hi) stem cells persist life-long, yet crypts drift toward clonality within a period of 1-6 months. We have collected short- and long-term clonal tracing data of individual Lgr5(hi) cells. These reveal that most Lgr5(hi) cell divisions occur symmetrically and do not support a model in which two daughter cells resulting from an Lgr5(hi) cell division adopt divergent fates (i.e., one Lgr5(hi) cell and one transit-amplifying [TA] cell per division). The cellular dynamics are consistent with a model in which the resident stem cells double their numbers each day and stochastically adopt stem or TA fates. Quantitative analysis shows that stem cell turnover follows a pattern of neutral drift dynamics.     

Evaluation of ultrasound velocity measurements for estimating protease activities using casein as substrate

Ultrasonic resonator technology (URT) was compared with the well established UV–Vis/ninhydrin assay to estimate protease activities in defined buffer systems. Hydrolysis of casein was measured using subtilisin, trypsin, halophilic protease from Haloferax mediterranei and Bacillus lentus alkaline protease. Sensitivity, reproducibility, working range as well as the limit of detection and the limit of quantification were comparable for both methods. Salt concentrations (0.5 M NaCl) interfered with the URT method. The quantification of protease activity by URT was possible when the product concentration measured by the UV–Vis/ninhydrin assay was correlated to the corresponding ultrasonic velocity signals.