Immunocytochemical distribution of H‐K‐ATPase beta‐subunit in the hyperglycemic mice

• Published in: The FASEB journal 2008-04, Vol.22 (S2), p.181-181
• Main Authors: Elmore, Zachary, Duley, Adam, Headford, Mikel, Muncie, Christopher, Phelps, Nathaniel, Nakamura, Suguru
• Format: Article
• Language: English
• ISSN: 0892-6638; 1530-6860
• DOI: 10.1096/fasebj.22.2_supplement.181

BACKGROUND: Two proton pumps, H‐K‐ATPase (HKA) and H‐ATPase play an important role in acid‐base balance in the kidney. Three H‐K‐ATPase subunits (HKAalpha1, HKAalpha2, HKAbeta) are expressed in the collecting duct. The beta‐subunit is essential for the functional expression of both the HKAalpha1 and HKAalpha2. It has been reported that the HKAalpha1 (gastric) antibody level is increased in type 1 diabetic patients. Glucose stimulates H‐ATPase and H‐K‐ATPase activities and has been demonstrated in the kidney cells. We examined whether hyperglycemia stimulates the HKA beta‐subunit in the outer medullary collecting duct (OMCD) in the mouse kidney. METHODS: The hyperglycemia was induced by an intraperitoneal injection of streptozotocin (65 mg/kg) and monitored for 5 days. The OMCD tubules were dissected and placed on the center of PLL‐coated cover glass and covered with a phosphate buffered saline (PBS). The OMCDs were fixed in 4% paraformaldehyde, permeabilized with 0.5% Triton X‐100, incubated with a specific monoclonal antibody (HKA beta‐subunit, 34 kDa) and stained with FITC‐conjugated antibody. RESULTS: In the normal mice OMCDs, the intercalated cells demonstrated the immunocytochemical distribution on HKA that was mainly polarized to apical pole and slightly polarized to basolateral pole. In the hyperglycemic mice OMCDs, there was a diffusely distributed staining throughout the cells. CONCLUSIONS: Results suggested that the HKA beta‐subunit is stimulated by hyperglycemia in the renal OMCDs. The HKA beta‐subunit is likely to associate with the HKAalpha1 and HKAalpha2 under normal and hyperglycemic conditions.