CD3+CD4-CD8- Double-negative αß T cells attenuate lung ischemia-reperfusion injury.

OBJECTIVE: Lung ischemia-reperfusion injury (IRI) is a common complication after lung transplantation, and immune cells have been implicated in modulating outcomes. We hypothesized that a newly described subset of αß T-cell receptor positive cells; that is, CD4-CD8- (double negative [DN]) T cells, are found in lungs and can protect against lung IRI. METHODS: Ischemia was induced in C57BL/6 mice by left pulmonary artery and vein occlusion for 30 minutes followed by 180 minutes of reperfusion. These mice were paired with sham hilar dissected surgical controls. In mice undergoing IRI, adoptive transfer of DN T cells or conventional T cells was performed 12 hours before occlusion. Flow cytometry was used to quantify T cells and inflammatory cytokines, and apoptotic signaling pathways were evaluated with immunoblotting. Lung injury was assessed with Evans blue dye extravasation. RESULTS: DN T cells were significantly higher (5.29% ± 1% vs 2.21% ± 3%; P < .01) in IRI lungs and secreted higher levels of interleukin-10 (30% ± 5% vs 6% ± 1%; P < .01) compared with surgical sham controls. Immunoblotting, hematoxylin and eosin staining and Evans blue dye demonstrated that adoptive transfer of DN T cells significantly decreased interstitial edema (P < .01) and attenuated apoptosis/cleaved caspase-3 expression in the lungs following lung IRI (P < .01). CONCLUSIONS: DN T cells traffic into lungs during IRI, and have tissue protective functions regulating inflammation and apoptosis. We propose a potential novel immunoregulatory function of DN T cells during lung IRI.

Mixing commonly used crystalloid solutions with red blood cells in five common additives does not negatively impact hemolysis, aggregometry, or deformability.

BACKGROUND: Literature is beginning to challenge the belief that it is unsafe to coinfuse red blood cells (RBCs) with solutions other than isotonic saline. We recently showed that additive-free RBCs tolerated coincubation with Plasma-Lyte or catecholamines dissolved in normal saline (NS), though 5% dextrose in water (D5W) promoted hemolysis. Herein, we evaluate the effect of coincubating crystalloids on additive-preserved RBC hemolysis, aggregation, and membrane deformability. STUDY DESIGN AND METHODS: RBCs were coincubated 5 minutes with plasma, NS, Plasma-Lyte, lactated Ringer's (LR) or D5W (1 mL PRBC +131.3 µL solution). Samples were then assessed for hemolysis (free hemoglobin), aggregation (critical shear stress [mPa]), and membrane deformability (elongation index [EI]). Significance (P ≤ .05) by t test or ANOVA with post-hoc Tukey-Kramer test. RESULTS: Additive-prepared RBCs coincubated with crystalloid instead of plasma demonstrated: (a) no increase in hemolysis as indicated by plasma free hemoglobin levels that is likely to be clinically relevant; (b) no increase, but in some cases a decrease, in aggregation as indicated by critical shear stress; and (c) in some combinations, a deterioration in deformability. When present, the deformability decrease was likely clinically insignificant in degree, and always returned to normal when the crystalloid was subsequently diluted out with plasma. CONCLUSION: Our data suggest that additive-prepared RBCs coincubated for 5 minutes with any of four common crystalloids demonstrate no clinically relevant increased lysis, increased aggregation, or decreased deformability.

Effect of incubation with crystalloid solutions or medications on packed red blood cells.

BACKGROUND: American Association of Blood Banks (AABB) guidelines suggest that packed red blood cells (PRBCs) be administered through a dedicated intravenous (IV) catheter. Literature supporting this broad-scope declaration are scarce. Obtaining additional IV access is painful, costly, and an infectious risk. We evaluated the effect of co-incubating PRBCs with crystalloids and medications on PRBC hemolysis, membrane deformability, and aggregation, as well as medication concentration. METHODS: PRBCs were co-incubated 5 minutes with plasma, normal saline (NS), 5% dextrose in water (D5W), Plasmalyte, epinephrine (epi), norepinephrine (norepi), dopamine (dopa), or Propofol (prop). Samples were then assessed for hemolysis (free hemoglobin, serum potassium), membrane deformability (elongation index [EI]), aggregation (smear, critical shear stress [mPa]) and drug concentration (High Performance Liquid Chromatography/Tandem Mass Spectrometry [LCMS-MS]). Significance (p ≤ 0.05) was determined by Wilcoxon-paired comparisons or Wilcoxon/Kruskall Willis with post-hoc Dunn's test. RESULTS: Compared to co-incubation with plasma: 1) co-incubation resulted in significantly increased hemolysis only when D5W as used (free hemoglobin, increased potassium); 2) EI trended lower when co-incubated with D5W and trended toward higher when co-incubated with prop; 3) aggregation was significantly lower when PRBCs co-incubated with NS, D5W, or Plasmalyte, and trended lower when co-incubated with epi, norepi, or dopa. Medication concentrations were between those predicted by distribution only in plasma and distribution through the entire intra- and extracellular space. CONCLUSION: Our data suggest that 5 minutes of PRBC incubation with isotonic crystalloids or catecholamines does not deleteriously alter PRBC hemolysis, membrane deformability, or aggregation. Co-incubation with D5W likely increases hemolysis. Propofol may promote hemolysis.