Effects Of Uremic Toxins On Erythropoietin Receptor

Objective: To explore the important role of erythropoierin receptor (EPOR) in the mechanisms of renal anemia via investigating (1) the changes of serum EPO concentration; (2) the inhibitory effect of EPORmRNA expression, EPOR protein and the Clone-forming unit-erythroid (CFU-E) growth of erythroid progenitors co-incubated with some uremic toxins in vitro; (3) the recovery of EPORmRNA expression and the EPOR protein content after uremic toxins were cleared totally following kidney transplantation.Methods:Part 1: Fourty two patients with chronic renal failure (CRF) who received cadaveric kidney transplantation were studied prospectively during the first 24 weeks postoperatively. Serum erythropoietin (EPO) levels were measured with ELISA. serum creatinine (SCr), haemoglobin (Hb) were assayed pre- and post-operatively at the same time.Part 2: Uremic toxins including creatinine, urea, Uric acid, spermidine, middle molecule substances (MMS) and Parathyroid hormone (PTH) was co-incubated with mononuclear cells of rat bone marrow respectively for 7 days. Counted the number of CFU-E with inverted microscope, examined the expression of EPORmRNA by RT-PCR technique and assayed the content of EPOR protein by Western Blot analysis.Part 3: The model of chronic renal failure of rat was surgically induced by partial (5/6) nephrectomy. CRF rats were recipients of renal allograft. The expression of EPORmRNA and the content of EPOR protein of bone marrow were examined by RT-PCR technique and Western Blot analysis pre- andpost-transplant. Correlations between EPORmRNA, the content of EPOR protein, SCr, and Hb were analysised.Results:Part1: Mean serum EPO concentration of CRF patients' showed significant lower than that of control. After transplantation, renal function recovered quickly while Hb level increased steadily in 37 patients with immediate graft function (IGF). Their EPO concentration increased significantly and double peaks of EPO level were obtained at day 4 and week 4. 4 cases in IGF suffered from post-transplant erythrocytosis (PTE). But their EPO concentration did not reach statistical significance. 5 cases developed delayed graft function (DGF), without EPO double peaks and their EPO concentrations showed signiffcant lower than that of IGF.Part 2: Creatinine, urea, uric acid and PTH had no effect on CFU-E growth of rat bone marrow in vitro. But Spermidine and MMS inhibited the growth of CFU-E significantly. All the uremic toxins except PTH had remarkably inhibitory effect on the expression of EPORmRNA and the amount of EPOR protein.Part 3: CRF model were successfully established on post operation days (POD) 90. The SCr and BUN decreased sharply immediately after allograft transplantation. Expression of EPORmRNA and content of EPOR protein of CRF rats were significantly lower than normal level. After transplantation, expression of EPOR mRNA and content of EPOR protein increased significantly on POD 7 and recovered on POD 14. Negative correlations were found between the expression of EPORmRNA or content of EPOR protein and SCr respectively, whereas statistically positive correlations were found between the expression of EPORmRNA or content of EPOR protein and Hb.Conclusions:1. The reduction of serum EPO concentration in CRF patients is an important mechanisms of renal anemia. It recovers depending on the recovery of graft function.2. Most uremic toxins inhibit the expression of EPORmRNA and the content of EPOR protein on CFU-E of rat's bone marrow significantly in vitro.3. EPORmRNA and the content of EPOR protein recover to normal level after allograft transplantation when uremic toxins are eliminated completely.4. We put forward a new viewpoint in renal anemia EPOR inhibition hypothesis. It may be an important complement to the EPO reduction theory in renal anemia. It may have potential significance in renal anemia therapy.