Glucose-Induced Changes in Turnover of Na⁺/H⁺ Exchanger of Immortalized Lymphoblasts From Type I Diabetic Patients With Nephropathy

Increased cellular Na⁺/H⁺ exchanger (NHE) activity has been demonstrated in type I diabetic patients with nephropathy. Such patients also have a previous history of poor glycemic control. The interaction between hyperglycemia and changes in NHE activity remains obscure. Therefore, we examined the effects of media containing 5 and 25 mmol/l glucose on the increased NHE activity and turnover number in Epstein-Barr virus-transformed lymphoblasts from patients with diabetic nephropathy compared with normoalbuminuric diabetic and nondiabetic control subjects. NHE activity was determined fluorometrically, and NHE isoform 1 (NHE-1) density was measured with specific polyclonal antibodies. In the presence of 5 mmol/l glucose, cells from patients with diabetic nephropathy exhibited higher NHE activity with intracellular pH clamped to 6.0 compared with diabetic and nondiabetic control subjects (P < 0.005 for both), due to a higher turnover number of NHE-1. Incubation in 25 mmol/l glucose for 48 h caused an increase in NHE activity (P < 0.001) and turnover number (P < 0.01) in the diabetic nephropathy group only, with no significant change in the diabetic or nondiabetic control groups. The rate constants for cell proliferation and NHE activity or turnover number were correlated when cells were cultured in 5 mmol/l glucose (r = 0.34 and 0.32, respectively; P < 0.05) or 25 mmol/l glucose media (r = 0.66 and 0.65, respectively; P < 0.001). We conclude that only lymphoblasts from the diabetic nephropathy group show an increase in NHE activity and turnover number under conditions mimicking hyperglycemia. Thus, high glucose levels exaggerate the differences in NHE activity, turnover number, and cell proliferation rate already present between cells from diabetic nephropathy patients and those from diabetic and nondiabetic control subjects.