Detection of Apoptosis in Kidney Biopsies of Patients with D+ Hemolytic Uremic Syndrome

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Abstract
In this study we have investigated the presence of apoptotic cells in renal biopsy material of seven patients with hemolytic uremic syndrome (HUS) by using an improved and stringent terminal deoxynucleotidyl nick-end labeling (TUNEL) technique. Renal biopsy material was taken in the second or third week after onset of the disease. Renal biopsy material of patients with minimal lesions nephrotic syndrome or thin basement syndrome were used as control. It has been reported that nonapoptotic cells can be labeled nonspecifically due to proteinase K pretreatment or a delay in fixation when only TUNEL technique is used. In post mortem material this delay in fixation is seen. Moreover, it has been described that mainly nonapoptotic cells that shows signs of active gene transcription can be labeled in this nonspecific way. For this reason we used the TUNEL technique in combination with a label for RNA synthesis and splicing factor (SC-35). Indeed, we found nonspecific labeling of nonapoptotic nuclei in biopsy material of HUS patients, but not in control biopsy material. By using co-labeling with RNA synthesis factor SC-35, we were able to identify true apoptotic cells. There was a significant increase (p < 0.05) in the presence of apoptotic cells in biopsy material of HUS patients compared with material of controls. About 80% of apoptotic cells were detected in tubuli and only 20% in glomeruli of the renal biopsies of HUS patients. Furthermore, most apoptotic cells were detected in those patients that had received peritoneal dialysis suggesting that there is a relationship between severity of the disease and amount of apoptotic cells. The finding of apoptotic cells suggest that apoptosis plays a role in HUS.