The Impact of Anticoagulation Agent on the Composition and Phenotype of Blood Leukocytes in Dromedary Camels.

Jamal Hussen,Turke Shawaf,Sameer M Alhojaily

doi:10.3390/vetsci9020078

Abstract

For the analysis of several cellular biomarkers, blood samples are anticoagulated using different agents with different modes of action. However, for the most commonly used anticoagulants, EDTA and heparin, varying effects on blood components have been reported in different species. As little is known about the impact of anticoagulants on the immunological evaluation of camel leukocytes, the present study analyzed the leukogram, the immunophenotype, and the cell vitality of camel leukocytes separated from blood samples anticoagulated with EDTA or lithium heparin. Using flow cytometry and staining with monoclonal antibodies to several cell surface markers, the composition and immunophenotype of camel leukocytes separated from blood anticoagulated with EDTA or heparin were analyzed. In comparison to EDTA-anticoagulated blood, using lithium heparin as an anticoagulant resulted in reduced numbers of total leukocytes and reduced numbers of neutrophils, which led to a reduced neutrophil to lymphocyte ratio. The analysis of cell necrosis and apoptosis after the staining of leukocytes with the DNA-sensitive dye propidium iodide and the mitochondrial membrane potential probe JC1 revealed a higher fraction of necrotic neutrophils and higher fractions of apoptotic neutrophils and monocytes in heparin blood than in EDTA blood. In addition, monocytes from heparin blood showed higher expression levels of the cell surface markers CD14, CD163, and MHCII when compared to cells from EDTA blood. Similarly, in heparin blood, CD44 and CD172a were expressed higher on neutrophils, while CD11a was expressed higher on lymphocytes in comparison to cells from EDTA blood. The results of the current study indicate the importance of considering the type of anticoagulant when investigating the composition, vitality, and immunophenotype of camel leukocytes.

Highlights

The evaluation of immune cell composition and phenotype represents an effective procedure for the identification of disease biomarkers in human and veterinary medicine [1,2,3].The absolute and relative quantification of the main leukocyte populations including the neutrophilic, eosinophilic, and basophilic granulocytes, lymphocytes, and monocytes, which is called the leukogram, provides a cost-effective evaluation tool with species-specific patterns in health and disease [4]
For For several immunological testing procedures including several immunological testing procedures includingthe themeasurement measurementof ofleukoleukocyte cytecomposition, composition,the theimmunophenotyping, immunophenotyping,and andthe thefunctional functionalanalysis analysis different leuofof different leukocyte subpopulations, blood samples are are collected in tubes containing anticoagulation agents kocyte subpopulations, blood samples collected in tubes containing anticoagulation with different mode of actions
The present study identified the significant impact of the anticoagulation agent on the phenotype and vitality of camel blood leukocytes

Summary

Introduction

The evaluation of immune cell composition and phenotype represents an effective procedure for the identification of disease biomarkers in human and veterinary medicine [1,2,3].The absolute and relative quantification of the main leukocyte populations including the neutrophilic, eosinophilic, and basophilic granulocytes, lymphocytes, and monocytes, which is called the leukogram, provides a cost-effective evaluation tool with species-specific patterns in health and disease [4]. Several leukogram changes have been described in healthy and diseased dromedary camel with some discrepancies in the reported leukogram patterns [6]. In these studies, ethylenediaminetetraacetic acid (EDTA) and heparin, which have different modes of action, are the most common anticoagulation agents used for blood sample collection [7]. The mode of action for the anticoagulation effect of heparin depends on its ability to potentiate the inhibitory activity of the plasma protein antithrombin III, leading to the inhibition of several serine proteases of the coagulation system including factors IIa (thrombin), Xa, and IXa [8,9]. Heparin can act through modulating other protease inhibitors including heparin co-factor II and inhibitors of protein C and plasminogen [9,10]

Methods

Discussion

Conclusion