The effect of EDTA and Heparin anticoagulants on the survival of Neutrophils and T-cell responses among Visceral Leishmaniasis Patients in North West Ethiopia

Abstract

Background: The leishmaniasis are a group of diseases caused by protozoan parasites of the genus Leishmania. Leishmania are, obligate intracellular kinetoplastid protozoan parasites, that are transmitted through the bite of a vector called sandy fly. The disease ranges from self-healing cutaneous to visceral leishmaniasis (VL), which is severe and can causes death when untreated. Following the delivery of parasites by sand fly bite or by needle injection, neutrophils have been found the first recruited to the infection site. The role of neutrophils in either promoting or suppressing host immune response remains controversial. Objective: To assess the effect of EDTA and heparin anticoagulants on the survival of LDGs in PBMCs and the release of interferon-γ and IL-10 in the WBA. Methods: A cross-sectional study was conducted on clinically and laboratory confirmed VL patients from December 2014 to May 2015 at University of Gondar Hospital. Convenience sampling technique was used to select study participants and all consecutive VL cases who came to the hospital during the study period and controls were included. 6 ml of blood using EDTA and 6 ml using heparin was collected from all study participants before and after treatment and from controls to assess the effect on LDGs using FACs caliber, arginase activity using Enzymatic assay and production of IFN-γ and IL-10 using ELISA. Data were evaluated using GraphPad Prism 6 and differences were considered statistically significant at p < 0.05. FACS data were analyzed using Summit v4.3 software. Results: The frequency of LDGs is significantly lower when the blood is collected with heparin as compared to EDTA. Anticoagulants does not have an impact on the levels of arginase released. And results show that EDTA prevents the production of IFN-γ. Cells from active VL patents produce no or low levels of IFN-γ and IL-10, however, after successful treatment, these cells gradually regain their capacity to produce IFN-γ, but not IL-10. Conclusion and recommendations: In this study results suggest that, VL patients have lost their ability to mount a Th1 response during active VL and that active disease is not associated with sustained levels of IL-10. For the future in vitro experiment, For the future in vitro experiment, For the future in vitro experiment, For the future in vitro experiment, For the future in vitro experiment, For the future in vitro experiment, For the future in vitro experiment, For the future in vitro experiment, For the future in vitro experiment, For the future in vitro experiment, For the future in vitro experiment, For the future in vitro experiment, For the future in vitro experiment, For the future in vitro experiment, For the future in vitro experiment, choosing different anticoagulants to study about different cells and T cell response against leishmania is important