Question: 

The epithelial sodium channel (ENaC) plays a critical role in the regulation of renal sodium excretion. Recent evidence indicates that proteolytic processing of ENaC contributes to its activation. In renal inflammatory disease, damage of the glomerular filter can result in pathological excretion of plasma proteins (proteinuria). Filtered plasma proteases or protease precursors may appear in the urine and activate ENaC resulting in sodium retention and oedema formation. In this study, we investigated whether urine samples from doxorubicin treated nephrotic mice can stimulate the activity of heterologously expressed mouse ENaC.

Methodology: 

Urine samples from wild-type mice of the strain 129S1/SvImJ were collected daily before (two days) and after (26 days) induction of nephrotic syndrome by doxorubicin (single dose of 15 mg/g bw i.v.). Urinary protease activity was determined by gelatin zymography. Mouse abg ENaC was heterologously expressed in Xenopus laevis oocytes and amiloride-sensitive whole-cell currents (DI_ami) were determined by the two-electrode voltage-clamp technique before and after 30 minutes incubation of the oocytes in mouse urine samples.

Result: 

As expected, urine samples collected before induction of the nephrotic syndrome contained very little protein and protease activity was very low. Following doxorubicin treatment, the protein content and protease activity in the urine samples increased dramatically in a time-dependent manner. Exposure of ENaC expressing oocytes to control urine samples obtained before doxorubicin treatment caused a small but significant increase in DI_ami. Incubating oocytes with urine collected in the initial phase following induction of nephrotic syndrome caused a further increase of DI_ami in most experiments (10 out of 11). The additional increase of DI_ami was variable in onset and duration. Surprisingly, urine samples with a high protein concentration and protease activity collected from mice at a later stage of the disease had no pronounced stimulatory effect on ENaC compared to control urine samples.

Conclusion: 

Our findings demonstrate that ENaC can be stimulated by mouse urine. This stimulation may be mediated by urinary proteases and appears to be more prominent in the initial phase of nephrotic syndrome. However, the lack of pronounced ENaC stimulation by nephrotic urine samples with high protein content and protease activity suggests that proteolytic activation of ENaC may be prevented by compensatory mechanisms e.g.via upregulation of protease inhibitors.