Kidney Week

Abstract: FR-PO319

Deletion of SPAK (Stk39) Reduces Hypertension, NKCC2 Phosphorylation and NKCC2 Activity in Dahl Salt-Sensitive Rats

Session Information

• Hypertension and CVD: Mechanisms - I
  October 26, 2018 | Location: Exhibit Hall, San Diego Convention Center
  Abstract Time: 10:00 AM - 12:00 PM

Category: Hypertension and CVD

• 1403 Hypertension and CVD: Mechanisms

Authors

• Garcia-Pedraza, Jose A., Henry Ford Hospital, Detroit, United States

Jose A. Garcia-Pedraza,

• Liao, Tang-Dong, Henry Ford Hospital, Detroit, Michigan, United States

Tang-Dong Liao,

• Ortiz, Pablo A., Henry Ford Hospital, Detroit, Michigan, United States

Pablo A. Ortiz,

Background

The Na/K/2Cl cotransporter NKCC2 mediates the NaCl reabsorption in the thick ascending limb (TAL), playing a crucial role in blood pressure regulation. NKCC2 can be phosphorylated by the SPAK kinase (Stk39) at Thr^96,101 and this is thought to activate NKCC2. We found that NKCC2 phosphorylation was 5-fold higher in TALs from Dahl salt-sensitive (DSS) rats. However, the role of SPAK in NKCC2 function and salt-sensitive hypertension remain unclear. We hypothesized that SPAK is involved in salt-sensitive hypertension in part by increasing NKCC2 phosphorylation and activity.

Methods

To test this, we generated SPAK knockout (KO) rats in a DSS genetic background via Crispr/Cas9 targeting. We used telemetry to measure systolic blood pressure (SBP) on normal (0.22% Na) or high salt diet (4% Na).

Results

At baseline, SPAK-KO had lower SBP than wild-type DSS (WT) rats (SPAK-KO: 139±1, WT: 153±2 mmHg; p<0.05). After 4 weeks on high salt, SBP rose reaching 162±3 in SPAK-KO and 183±3 mmHg in WT rats (p<0.05). The increase in SBP after feeding a high salt diet was lower in SPAK-KO than WT rats at 7 days (10±1 vs 15±1 mmHg, p<0.05), 14 days (14±2 vs 19±2 mmHg, p<0.05), and 28 days (22±3 vs 30±2 mmHg, p<0.05). We measured total and phospho-NKCC2 in isolated TALs. On normal salt diet, phospho-NKCC2 (Thr^96,101) over total NKCC2 was 61±25% lower in SPAK-KO (p<0.05), whereas total NKCC2 was not different. After 4 weeks on high salt, phospho-NKCC2 was 62±13% lower in SPAK-KO (p<0.05) whereas total NKCC2 was not different. SPAK-KO rats had a decreased bumetanide-induced natriuresis while on a normal salt diet (UNa WT: 1271±55; UNa SPAK-KO: 843±80 µmol/12h; p<0.05). Lymphocyte infiltration (CD3+) into the kidney after 4 weeks on high salt diet was not different between strains, whereas CD68+ macrophages were lower in SPAK-KO (WT: 266±35, SPAK-KO: 150±6 CD68+/mm², p<0.05).

Conclusion

We conclude that SPAK-KO DSS rats exhibit lower SBP at baseline and reduced salt-sensitivity in part caused by lower NKCC2 phosphorylation and activity. In addition, decreased macrophage infiltration in SPAK KO rats supports a potential role for SPAK in renal inflammation during salt-sensitive hypertension.

Funding

• NIDDK Support