Whole-Blood Assay to Measure Oxidative Burst and Degranulation of Neutrophils for Monitoring Trauma Patients

AbstractBackground and Purpose: Polymorphonuclear neutrophils (PMNs) protect the host from invading microorganisms, but excessive PMN activation after trauma causes tissue injury. Rapid monitoring of PMN function is critical for the assessment of the inflammatory state of trauma patients. Here, the authors adapted two simple and rapid methods to measure oxidative burst and degranulation of human PMNs in whole blood to avoid potential interference of cell isolation procedures with the assessment of PMN function.Material and Methods: Heparinized blood was drawn from healthy volunteers or trauma patients, preincubated at 37 °C for 5 min, and stimulated with N-formyl-methionyl-leucyl-phenylalanine (fMLP). Four assays for oxidative burst were tested: (1) cytochrome C; (2) homovanillic acid (HVA); (3) Amplex^® Red; and (4) flow cytometry with dihydrorhodamine 123 (DHR). PMN degranulation was assessed with flow cytometry using antibodies to: (1) CD11b/Mac-1 (CD18); (2) CD63; and (3) CD66b (CD67).Results: With the exception of the DHR method, all methods to measure oxidative burst were found to be unsuitable in whole blood due to interference of plasma proteins and hemoglobin with the fluorimetric or photometric readouts. By contrast, all degranulation methods were suitable for whole-blood studies. However, for the assessment of formyl peptide-induced degranulation, anti-antibodies to CD11b/Mac-1 and CD66b were up to five times more sensitive than antibodies to CD63. Thus, the degranulation and DHR methods were optimized for increased sensitivity, speed, and specificity and their usefulness to measure PMN function in trauma patients was tested.Conclusion: The whole-blood methods based on flow cytometry with DHR, anti-CD11b/Mac-1, and anti- CD66b are rapid, simple, and reliable techniques to assess PMN function for trauma research.     

Thorakolumbaler Übergang der Wirbelsäule

Trauma und Berufskrankheit - In der überwiegenden Anzahl der Patienten mit posttraumatischen Fehlbildungen der Wirbelsäule geben Schmerzen Anlass zu einem Korrektureingriff. Die Ursachen...     

A mating deficient and temperature-sensitive lethal mutant of Schizosaccharomyces pombe defines a new fertility locus

Summary A recessive mutant allele, mef1-84, of a novel locus mapping on the left arm of chromosome I, between ade3 and ura1, 5 cM apart from lys5, confers temperature-sensitive growth and mating deficiency at the nonrestrictive temperatures for growth. Two other mutations suppress the phenotype conferred by mef1-84: sts1-1 suppresses the temperature-sensitive growth only, and smd1-35 suppresses both temperature-sensitive growth and mating deficiency.     

Preoperative and intraoperative scintigraphic detection of sentinel node

It is known that the malignant primary tumor site has a specific lymphatic drainage, all the time intermediate, however, by a first lymph node named sentinel node. This can be situated in some typical or atypical topographic area and can be or not clinically evident. If the sentinel node is not invaded by the metastatic cells there is no metastatic risk for other lymphatic nodes, so the surgical treatment is not necessary to involve the corresponding lymphatic area dissection. An imagine, noninvasive detection of the involvement of the sentinel node in the spread of cancer could be related with the surgical decision to perform (or not) fully regional lymphadenectomy (based on biopsy result of such positive detected nodes) allowing only a tumorectomy in the first stages of neoplasia. For the case of scintigraphic method to detect the sentinel node, the performance depend on the development of radiotracers, the modification of the lymphoscintigraphic mapping techniques protocols, and the development and improvement of detectors.     

Network of telocytes in the temporomandibular joint disc of rats

The phenotypes of the temporomandibular joint (TMJ) disc cells range from fibroblasts to chondrocytes. There are relatively few reported studies using transmission electron microscopy (TEM) to determine the ultrastructural features of these cells. It was hypothesized that at least a subpopulation of TMJ stromal cells could be represented by the telocytes, cells with telopodes. In this regard a TEM study was performed on rat TMJ samples. Collagen-embedded networks were found built-up by cells with telopodes with subplasmalemmal caveolae, moderate content in matrix secretory organelles and well-represented intermediate filaments. Appositions of cell bodies were found. Prolongations of such cells were closely related to nerves and microvessels. Our study indicates that the TMJ disc attachment seems equipped with telocytes capable of stromal signaling. However, further studies are needed to assess whether the telocytes belong to a renewed cell population derived from circulating precursors.     

Regenerative Potential of Human Schneiderian Membrane: Progenitor Cells and Epithelial‐Mesenchymal Transition

An innate osteogenic potential of the Schneiderian membrane (SM) is progressively assessed in studies ranging from non‐human species to human subjects. It has relevance for endosteal placement and osseointegration. Nestin‐expressing osteogenic progenitor cells are allegedly involved in bone formation and remodelling. Nestin phenotype was not assessed previously in human SM. We therefore aimed to fill that particular gap in the literature. Bioptic samples of human adult SM were obtained during surgery from eight adult patients, operated for non‐malignant pathologies. Immunohistochemistry on paraffin‐embedded tissue samples used primary antibodies against nestin, CD45, CD146, cytokeratin 7 (CK7), and alpha‐smooth muscle actin (α‐SMA). Nestin expression was consistently found in endothelial cells, and was scarcely encountered in pericytes, putative stromal stem/progenitor cells, as well as in glandular epithelial cells. Moreover, woven bone formation in the periosteal layer of the SM can also be regarded as evidence of the osteogenic potential of this membrane. Nestin and CD45 expression in cells of the primary bone supports the osteogenic potential of SM nestin‐expressing cells and a possible involvement of hematopoietic stem cells in maxillary sinus floor remodeling. CD146, a known inducer of epithelial‐mesenchymal transition (EMT), was expressed in epithelia, as was CK7. Isolated stromal cells were found expressing CD146, CK7 and α‐SMA, suggesting that regenerative processes happening in the SM may also involve processes of EMT which generate stem/progenitor cells. This study provides additional evidence for the regenerative potential of the Schneiderian membrane and identifies potential roles for cells of its stem niche in osteogenesis. Anat Rec, 298:2132–2140, 2015. © 2015 Wiley Periodicals, Inc.