Contributing factors affecting erythropoiesis and analysis of erythropoiesis bioassay in renal patients in KwaZulu-Natal
Benjamin, Sherilene Cheryl
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Erythropoietin (EPO) is widely used in patients with chronic renal failure and is a necessity. However, due to the cost implications and the medical complications in our population it is imperative to review the factors affecting the process of erythropoiesis and the analysis of cell proliferation and cell viability in the bioassay. Complications such as hypertension and risk of worsening a malignancy cannot be ignored. We had previously analysed variations of erythropoietin levels in haemodialysis patients over a six month period. This study aims to evaluate erythropoiesis in conjunction with various laboratory, demographic, clinical parameters and inflammatory markers, in the population of haemodialysis patients. EPO, antibody level and antibody activity were analysed in the population groups as EPO responsive and EPO sensitive patients. This is a prospective, experimental and controlled study. Fifty nine patients were randomly selected from haemodialysis units of Addington and King Edward VIII Hospitals following an informed consent and 15 healthy individuals were also selected as controls. Demographic parameters (age, sex), clinical parameters (weight, height, skin folding, EPO doses and blood pressures (BP) were recorded. Pre-dialysis serum was used to measure laboratory markers (haemoglobin, transferrin, ferritin, albumin, ESR, C reactive protein, creatinine and urea). EPO levels and antibody levels were measured by ELISA, the optical density of each well was determined within fifteen minutes using the microplate reader set at 450 nm. All results were statistically analysed using SPSS statistical package version 21 (IBMR). Patients requiring very high doses of EPO to reach Hb of 11g/dL, and they remained anaemic after at least three months of adequate EPO doses were considered to be EPO resistant. Those who responded to the usual EPO doses were labelled EPO sensitive. The bioassay was used to quantify cell proliferation and cell viability in the presence of EPO. The UT 7 cells were cultured in medium, in the presence of serum from the EPO resistant, EPO sensitive patients and the healthy, control subjects. Luminescence was read with the Glorunner Microplate Luminometer and was recorded in relative light units (RLU). The analysis revealed: a non-significant positive correlation between haemoglobin and erythropoietin levels. However, a strong negative correlation was found between CRP and albumin level (R= -0.591; (p=0.001), which was not significant. No correlation was found between haemoglobin or erythropoietin levels and CRP or albumin. There was a positive correlation with systolic and diastolic blood pressures and mean arterial pressures which was statistically significant (p <0.05). EPO dosages and Hb levels were correlated significantly (p < 0.05). No correlation of EPO levels and Hb; age and Hb was found to be significant (p = 0.08). The UT 7 cells cultured in serum in medium alone with RHuEPO containing cells were statistically significant (p <0.01)). Reduction of ATP stimulation between medium and serum was observed. However, mean arterial pressures had a significant association with EPO resistance (p = 0.041) odd ratio- 1.066. In conclusion, EPO level is not a useful tool for the monitoring of its use as it does not correlate with EPO goal of red blood production in our patients. The neutralizing antibodies did not correlate with any of our variables contributing to erythropoiesis, and are therefore not confirmed as playing a major role in erythropoiesis. From the analysis of our results the key contributing factors of EPO doses, malnutrition and age were more significant in erythropoiesis. However the higher doses of EPO significantly increased the blood pressures and the mean arterial pressures (MAP). The analysis of the bioassay showed lack of difference between EPO responsive and EPO sensitive patients. This observation warrants further studies to clarify the role of serum of haemodialysis patients in erythropoiesis.