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Increasing NaCl Induces Superoxide Production in Macula Densa Cell Line
Author(s) -
Liu Ruisheng,
Lin Chunxia,
Carretero Oscar A.,
Ren YiLin,
Harding Pamela
Publication year - 2007
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.21.5.a501-a
Subject(s) - macula densa , tubuloglomerular feedback , chemistry , apocynin , nadph oxidase , nox4 , microbiology and biotechnology , biology , endocrinology , biochemistry , enzyme , renin–angiotensin system , blood pressure , renal function
To study whether the cultured macula densa cell line, MMDD1 shares the same characteristics of superoxide (O 2 − ) production as the isolated, perfused macula densa, we measured the O 2 − in the cell line. We also used laser capture microdissection (LCM) and PCR to examine which isoforms of NAD(P)H oxidase are expressed in the rat macula densa and compared with the cell line. The cultured cells were loaded with lucigenin (5 μM) and placed in a luminometer for O 2 − measurement. When NaCl was increased from 10 to 80 mM, O 2 − production rose from 0.27 ± 0.05 to 0.63 ± 0.08 units/10 5 cells ( p < 0.01 vs 10 mM NaCl). When the NAD(P)H oxidase inhibitor apocynin (10 −5 M) was added for 30 min, O 2 − production was 0.31 ± 0.07 units/10 5 cells ( p < 0.01 vs 10 mM NaCl). With RT‐PCR, we found NOX2 and NOX4 are expressed in the cell line. To determine which NOX isoforms are expressed in the macula densa, we performed LCM followed by single‐cell‐PCR on frozen sections of rat kidney. The macula densa cells were identified by their anatomical location and morphology and dissected with the laser. We confirmed that the captured cells are macula densa by nNOS expression. We found that NOX2 and NOX4 are expressed by the macula densa. In summary, we found that increasing luminal NaCl induces O 2 − production by the cultured macula densa cells, primarily from NAD(P)H oxidase. Both rat macula densa and the cell line express NOX2 and NOX4 isoforms.