Role of Urokinase in Chronic Hypertension with Albuminuria in DOCA-Salt Murine Model

Background: In proteinuria, aberrant filtration of urokinase-type plasminogen activator (uPA) and plasminogen (plg) leads to intratubular activation of plasmin with potential proteolytic activation of the epithelial sodium channel (ENaC). The ENaC and uPA inhibitor amiloride, reduces blood pressure in patients with treatment resistant hypertension and albuminuria. However, the non-redundant role for uPA is unresolved. Objective: The study aimed to test the hypothesis that uPA is necessary for the development of chronic hypertension in DOCA-salt murine model with albuminuria. Methods: FVB/uPA wildtype (WT) and gene-targeted (KO) mice were anesthetized (xylazine 10 mg/kg, ketamine 100 mg/kg i.p.) and subjected to unilateral nephrectomy or sham operation. 14-days post-surgery, 4%NaCl diet was introduced and DOCA/sham-pellets were inserted subcutaneously. Blood pressure was measured by indwelling chronic femoral catheters for 15 days. A subset of mice was housed in metabolic cages for 24 hr urine collection. Kidney tissue was collected and used for immunoblotting for γENaC abundance. In a second series, angiotensin II (ANGII) (60ng/min kg) was used to compare blood pressure to low ANGII DOCA-salt mice. Normally distributed data was analyzed by two-way ANOVA followed by post-hoc Bonferroni t-test. Results: Urine from DOCA-salt treated mice showed glomerular proteinuria with significantly increased albumin excretion (0,4 mg/24 hr/g bodyweight(BW)) compared to control (0,0007 mg/24hr/gBW) with no difference between genotypes. DOCA-salt WT urine showed plg and plasmin abundance, while KO mice only showed plg abundance. Kidney tissue showed increased cleaved γENaC/full-length γENaC abundance in DOCA-salt treated mice with no difference between uPA KO and WT. DOCA-salt increased mean arterial pressure (133 ±4 mmHg), systolic (152±4 mmHg) and diastolic (111±3mmHg) blood pressure significantly from day 1 to 15 compared to control (99±4 mmHg, 108±7 mmHg, 79±4 mmHg) with no difference between genotypes. Heart rate was unchanged (586 bpm), and no difference between day and night pressure. ANGII infusion for 7d led to similar blood pressure elevation in WT and uPA KO mice. Conclusion: In chronic hypertension with albuminuria, uPA is necessary for activation of plasminogen to plasmin in urine, but does not contribute to hypertension, albuminuria, or kidney γENaC cleavage. Funding: Private Foundation Support, Government Support – Non-U.S.