Characteristics of neutrophils and monocyets in patients with end stage kidney disease and those undergoing hemodialysis

Abstract

Chronic kidney disease is characterised by irreversible impairment of kidney function due to compromise in glomerular filtration. Clinical presentation progresses from asymptomatic initial stages to end stage kidney disease (ESKD) with complications. Clinical management is mainly focussed on slowing the progression and associated complications. However, in advanced stage of CKD, renal replacement therapy is sought; the most common among them being hemodialysis. HD involves diffusive transport of metabolic waste products across a semipermeable membrane into a dialysate solution. The procedure employs long tubings and a dialyzer which acts as artificial kidney into which blood is made to circulate extracorporeally and then shunted back into the body through another set of tubings after dialysis. Immune cells come into contact with these foreign materials at regular intervals and their function is altered. There is also a state of hypercytokinemia due to their decreased elimination creating an environment of chronic inflammation. One of the leading causes of mortality in HD is infections. Neutrophils and monocytes are inflammatory responders of the innate immune system playing a vital role in combating infections. While the various cytokines involved in inflammation have been studied, there is a paucity in knowledge on neutrophil phenotype and function and the changes that take place at different stages of HD. Similarly conflicting results from monocyte phenotype studies also needed further relook. With this background, we proceeded to characterize the phenotype and function of neutrophils and monocytes in patients at different stages of CKD and compared them to healthy controls. To achieve this, we recruited four groups of individuals; those with stage 5 CKD, not on dialysis (pre-dialysis/ pre-HD); those who are in their 6 to 12 months of HD (short term HD); those who received more than 2 years of HD (long-term HD); age matched apparently healthy controls (HCs). Peripheral venous blood samples were drawn before the start of HD and immune cells were isolated to characterize their phenotype and function using flow cytometry-based experiments. Firstly, we compared the characteristics of circulating neutrophils and monocytes obtained from subjects prior to dialysis (pre-HD) to healthy controls (HCs), followed by comparisons within the CKD sub-groups. Our findings showed that there were more immature neutrophils in the CKD group not on dialysis (pre-HD). The neutrophils in the pre-HD group showed specific functional alterations, such as lowered phagocytic ability. We also studied the low-density neutrophils (LDNs), which are found commonly in chronic conditions and malignancies, and found that the mean LDN percentage among the PBMCs are higher in the pre-HD group. Together, this data suggests that CKD results in impaired neutrophil numbers, phenotype and function when compared to HCs. Between the CKD sub-groups, the group on long-term HD showed neutrophils with the highest phagocytic ability, but had the lowest ability to produce reactive oxygen species. We also studied the CD36 (scavenger molecule) expression which is shown to be associated with atherosclerosis; it was higher in both the HD groups of CKD when compared to pre dialysis group. We did not prospectively follow up the patients from pre-HD stage to long-term HD. Hence, definitive conclusions cannot be made on the progression of marker expression in CKD. The study gathered other clinical and demographic details generating multiparametric data. While large-scale studies can reveal the distribution of variables, correlations, and trends using dimensionality reduction tools, this pilot study serves as a foundational step. It can inform power analysis for designing future large, multicentric cohort studies. The data generated from such studies can yield meaningful conclusions that help in modulating therapeutics for improved clinical outcomes.