Transfusion of stored red blood cells adhere in the rat microvasculature.

BACKGROUND: Ex vivo storage of red blood cells (RBCS) for transfusions is associated with a "storage lesion," which decreases RBC deformability and increases RBC adhesiveness to vascular endothelium. This may impair microcirculatory flow with deleterious effects on oxygen delivery after transfusion. Previous studies have shown that human RBCs adhere to endothelial monolayers in vitro with prolonged storage and is reduced by prestorage leukoreduction (LR). The objective of this study was to determine whether duration of RBC storage and LR influence RBC adhesion in vivo in capillaries. STUDY DESIGN AND METHODS: Rat RBCs were collected and stored in CPDA-1 under standard blood bank conditions. Three RBC products were compared: 1) fresh RBCs, less than 24 hours of storage (n = 6); 2) non leukoreduced (NLR) RBCs stored for 7 days (n = 6); and 3) prestorage LR RBCs stored for 7 days (n = 6). RBCs were labeled with fluorescein isothiocyanate (FITC) 24 hours before transfusion and reinjected in an isovolemic manner into healthy rats. The FITC-labeled RBCs were visualized in the extensor digitorum longus muscle using intravital video microscopy (20 x magnification). The number of RBCs adherent in capillaries was counted 1 hour after transfusion in 10 random fields and the median values were compared with one-way analysis of variance. RESULTS: Stored RBCs showed increased levels of adherence in capillaries compared to their fresh counterparts (p < 0.05). Prestorage LR decreased RBC adherence to levels equivalent to those of fresh RBCs (p < 0.05 for stored LR vs. stored NLR). CONCLUSION: Rat RBCs stored under conditions that closely mimicked clinical transfusion adhere in capillaries. The decreased RBC adherence with LR suggest a direct effect of white blood cells or their byproducts on RBC deformability and/or adhesiveness to microvascular endothelium. Further study will examine the mechanism of adherence and the impact it has on microcirculatory flow and oxygen delivery in the critically ill host.

Flow cytometric assessment of monocyte activation markers and circulating endothelial cells in patients with localized or metastatic breast cancer.

BACKGROUND: Monocyte activation in cancer patients may be reflective of anticancer activity. However, studies indicate that recruitment of macrophages can actually promote tumor growth and angiogenesis. Assessment of other microenvironmental cells such as circulating endothelial cells (CECs) may provide additional information regarding disease progression. The objective of this study was to assess monocyte activation and CECs in breast cancer patients and determine the potential clinical relevance during disease progression. METHODS: Patients (n = 41) with localized or metastatic breast cancer who were not currently receiving treatment were eligible for study inclusion. Peripheral blood was collected and analyzed by flow cytometry for monocyte activation (Leuko64 assay kit), and for CECs (CD146(+)CD45(-) phenotype). RESULTS: Metastatic breast cancer patients demonstrated a higher monocyte CD64 index relative to normal donors and localized breast cancer patients (P < 0.05). Furthermore, breast cancer patients had a lower monocyte CD163 index relative to normal donors (P = 0.008). Localized breast cancer patients demonstrated higher levels of CD146(+)CD45(-) cells CECs relative to metastatic breast cancer patients and normal donors. Within the localized breast cancer population, levels of CD146(+)CD45(-) cells increased with disease stage (P < 0.05). CONCLUSIONS: These results suggest that monocyte activation and CECs may play a role in breast cancer progression. We speculate that monocyte activation may reflect a reaction to metastatic cells and/or response to tissue damage caused by metastatic growth in distant organs. Furthermore, the observation that CECs increase with disease stage in localized breast cancer suggests that CECs could be a useful surrogate marker for disease progression in this patient population.

Performance characteristics of the Coulter LH 500 hematology analyzer.

The challenge for modern hematology laboratories is to provide accurate and reproducible results, with seamless performance between facilities, in a cost-effective manner. Beckman Coulter recently developed the Coulter LH 500 to meet the needs of smaller laboratories or serve as a backup in larger laboratories. The principal goal of this study was to validate all parameters and performance specifications of the LH 500 compared to the Coulter LH 750 predicate analyzer. A total of 245 spent clinical samples from the London Health Sciences Centre (LHSC) and 251 from the University of Pittsburgh Medical Center Health System (UPMCHS) were analyzed during the study. The samples were selected to include 75% abnormal and 25% normal blood samples. According to the results of a rank sum test, there was no significant difference between the LH 500 and LH 750 for all complete blood count parameters (P > .05) except the red cell distribution width, which showed a slight negative bias on the LH 500. Differential parameters comparing the LH 500 to a 400-cell manual differential showed correlation coefficients (r2) from 0.75 to 0.99 for all parameters except basophils. Of the samples run on the LH 500 at LHSC, the false-positive differential flagging rate was 17.32% and the false-negative rate was 3.03%. Sensitivity was 82.93%, specificity 78.95%, and efficiency 79.65%. At UPMCHS, the false-positive differential flagging rate was 13.37% and false-negative rate 2.97%. Sensitivity was 91.89%, specificity 78.91%, and efficiency 83.66%. Overall, the LH 500 performed accurately and reproducibly compared to the LH 750 and the reference procedures. It would be an excellent second instrument for larger laboratories concerned with harmonization of instrumentation and reagents or as a primary instrument for smaller hematology laboratories with limited space.

WBC reduction reduces storage-associated RBC adhesion to human vascular endothelial cells under conditions of continuous flow in vitro.

BACKGROUND: The effects of storage duration, WBC reduction, and irradiation on RBC adherence to vascular endothelia are unknown and are investigated under conditions of continuous flow. STUDY DESIGN AND METHODS: Thirty-two RBC units were collected and divided into three groups, non-WBC-reduced (NWR), WBC-reduced (WR), and irradiated-WBC-reduced. Aliquots of RBCs were removed on Days 1, 15, and 28 of storage for analysis. The RBC suspensions were then perfused at a 1.5 percent Hct in a protein-poor medium under conditions of continuous flow over human umbilical vein endothelial cell monolayers. On each slide, 25 randomly chosen sites were videorecorded over 10 minutes, and the number of RBCs adherent to the endothelial cell monolayer was counted. RESULTS: NWR RBCs stored for 28 days demonstrated a greater degree of adherence to endothelial cells compared to Days 1 and 15 (p < 0.03). The WR group had significantly fewer adherent RBCs than the NWR group on day 28 (p < 0.01). Irradiation had no effect on RBC adherence. CONCLUSION: Prolonged storage of NWR RBCs increases RBC adherence to endothelial cells in vitro. WBC reduction before storage abrogates the effect of storage duration on increased adhesion. Studies to assess whether an in vivo effect occurs are required.