Transfused older stored red blood cells improve the clinical course and outcome in a canine lethal hemorrhage and reperfusion model.

BACKGROUND: In canine models, transfused older stored red blood cells (RBCs) hemolyze in vivo resulting in significantly increased intravascular cell-free hemoglobin (CFH) and non-transferrin-bound iron (NTBI). During canine bacterial pneumonia with septic shock, but not in controls, older stored RBCs were associated with significantly increased lung injury and mortality. It is unknown if in shock without infection transfusion of older RBCs will result in similar adverse effects. STUDY DESIGN AND METHODS: Two-year-old purpose-bred beagles (n = 12) were transfused similar quantities of either older (42-day) or fresher (7-day) stored universal donor canine RBCs 2.5 hours after undergoing controlled hemorrhage (55 mL/kg). RESULTS: With older transfused RBCs, CFH (p < 0.0001) and NTBI (p = 0.004) levels increased, but lung injury (p = 0.01) and C-reactive protein levels (p = 0.002) declined and there was a trend toward lower mortality (18% vs. 50%). All three deaths after transfused fresher RBCs resulted from hepatic fractures. Lowered exogenous norepinephrine requirements (p < 0.05) and cardiac outputs (p < 0.05) after older transfused RBCs were associated with increased CFH levels that have known vasoconstrictive nitric oxide scavenging capability. CONCLUSIONS: In hemorrhagic shock, older RBCs altered resuscitation physiology but did not worsen clinical outcomes. Elevated CFH may lower norepinephrine requirements and cardiac outputs ameliorating reperfusion injuries. With hemorrhagic shock, NTBI levels persist in contrast to the increased clearance, lung injury, and mortality in the previously reported infection model. These preclinical data suggest that whereas iron derived from older RBCs promotes bacterial growth, worsening septic shock mortality during infection, release of CFH and NTBI during hemorrhagic shock is not necessarily harmful.

A unique population of effector memory lymphocytes identified by CD146 having a distinct immunophenotypic and genomic profile.

BACKGROUND: CD146 is a well described homotypic adhesion molecule found on endothelial cells and a limited number of other cell types. In cells from the peripheral circulation, CD146 has also been reported to be on activated lymphocytes in vitro and in vivo. The function associated with CD146 expression on lymphoid cells is unknown and very little information is available concerning the nature of CD146+ lymphocytes. In the current study, lymphocytes from healthy donors were characterized based upon the presence or absence of CD146 expression. RESULTS: CD146 was expressed on a low percentage of circulating T lymphocytes, B lymphocytes, and NK cells in healthy individuals. CD146 expression can be induced and upregulated in vitro on both B cells and T cells, but does not correlate with the expression of other markers of T cell activation. CD146 positive T cells do not represent clonal expansions as determined with the use of anti Vbeta reagents. Data suggest that CD146 positive cells have enhanced adherence to endothelial monolayers in vitro. Gene profiling and immunophenotyping studies between CD146+ and CD146- T cells revealed several striking genotypic distinctions such as the upregulation of IL-8 and phenotypic differences including the paucity of CCR7 and CD45RA among CD146 positive T cells, consistent with effector memory function. A number of genes involved in cell adhesion, signal transduction, and cell communication are dramatically upregulated in CD146+ T cells compared to CD146- T cells. CONCLUSION: CD146 appears to identify small, unique populations of T as well as B lymphocytes in the circulation. The T cells have immunophenotypic characteristics of effector memory lymphocytes. The characteristics of these CD146+ lymphocytes in the circulation, together with the known functions in cell adhesion of CD146 on endothelial cells, suggests that these lymphocytes may represent a small subpopulation of cells primed to adhere to the endothelium and possibly extravasate to sites of inflammation.

Detection of circulating endothelial cells and endothelial progenitor cells by flow cytometry.

The finding of angiogenic and vasculogenic cells in the peripheral circulation may have profound effects on the course of a variety of diseases ranging from cancer to cardiovascular disease. These cells are ascribed to be endothelial in nature and are generally referred to as circulating endothelial cells if mature or as endothelial progenitor cells if immature. Different approaches have been used to detect these cells, including in vitro culture, magnetic bead isolation, and flow cytometry. We review flow cytometric methods for the detection and enumeration of these cells and provide technical suggestions to promote the accurate enumeration of circulating endothelial cells and endothelial progenitor cells.

Multiplex bead array assays for detection of soluble cytokines: comparisons of sensitivity and quantitative values among kits from multiple manufacturers.

BACKGROUND: Multiplex bead array assays permit simultaneous cytometric quantitation of multiple cytokines in solution by capturing these to spectrally distinct beads. Because several manufacturers offer reagents to quantitate the same cytokines on a single instrument, a comparison should be made to determine whether these kits yield similar data and whether these data are comparable to enzyme-linked immunosorbent assay (ELISA). METHODS: This study compared cytokine detection kits by using Luminex 100. Twenty-six serum samples from seven subjects were analyzed for interferon-gamma, interleukins 1beta, 6, and 8, and tumor necrosis factor-alpha by using multiplex kits from LINCO Research, Bio-Rad Laboratories, R&D Systems, and BioSource International. Each assay was performed according to the manufacturers' specifications. Standard curves were generated by using reference concentrations supplied by each manufacturer. ELISAs for interleukin-8 were performed by using kits from R&D and BioSource. RESULTS: Cytokine levels followed similar patterns, although absolute concentrations differed among kits. ELISA and Luminex values for interleukin-8 were similar in kits from the same manufacturer. CONCLUSIONS: Because relative cytokine measurements are often valuable when performed serially, it may be possible to make interlaboratory comparisons by using different kits. When comparison of absolute values is crucial, kits from the same supplier should be used. Within-vendor, bead array, and ELISA values appear comparable.