Expression and localization of N-domain ANG I-converting enzymes in mesangial cells in culture from spontaneously hypertensive rats.
The angiotensin-converting enzyme (ACE) profile in urine of hypertensive patients and spontaneously hypertensive rats (SHR; 90- and 65-kDa N-domain ACEs) is different from that of healthy subjects and Wistar rats (190 and 65 kDa). In addition, four ACE isoforms were purified from mesangial cells (MC) of Wistar rats in the intracellular compartment (130 and 68 kDa) and as secreted forms (130 and 60 kDa). We decided to characterize ACE forms from SHR MC in culture. Analysis of the ACE gene showed that SHR MC are able to express ACE mRNA. The concentrated medium and cell homogenate were separately purified by gel filtration and then subjected to lisinopril-Sepharose chromatography. The molecular masses of purified enzymes, 90 kDa for ACEm1A and 65 kDa for ACEm2A (secreted enzymes) and 90 kDa for ACEInth1A and 65 kDa for ACEInth2A (intracellular), were different from those of Wistar MC. The purified enzymes are Cl- dependent, inhibited by enalaprilat and captopril, and able to hydrolyze AcSDKP. Immunofluorescence and cell fractionation followed by Western blotting showed predominant immunoreaction of the 9B9 antiserum for N-domain ACE in the nuclei. The N-domain ACE was localized in the glomerulus from Wistar rats and SHR. ANG II and ANG-(1-7) were localized in the cell cytoplasm and nuclei. The 90-kDa N-domain ACE, described recently as a possible genetic marker of hypertension, was found inside the cell nuclei of SHR MC colocalized with ANG II and ANG-(1-7). The presence of ANG II in the cell nuclei could suggest an important role for this peptide in the transcription of new genes.
Camargo de Andrade
Maria Claudina
MC
Universidade Federal de São Paulo, Escola Paulista de Medicina, Departamento de Medicina, Disciplina de Nefrologia, Rua Botucatu, 740, CEP 04023-900, São Paulo, SP, Brazil.
Di Marco
Giovana Seno
GS
de Paulo Castro Teixeira
Vicente
V
Mortara
Renato Arruda
RA
Sabatini
Regiane Angélica
RA
Pesquero
João Bosco
JB
Boim
Miriam Aparecida
MA
Carmona
Adriana Karaoglanovic
AK
Schor
Nestor
N
Casarini
Dulce Elena
DE
D000804:Angiotensin II / Q000378:metabolism*; D000818:Animals; D002478:Cells, Cultured; D006973:Hypertension / Q000201:enzymology*; D007120:Immunochemistry; D050527:Mesangial Cells / Q000201:enzymology*; D007703:Peptidyl-Dipeptidase A / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D011918:Rats, Inbred SHR; D017208:Rats, Wistar; D017385:Sequence Homology; D013347:Subcellular Fractions / Q000201:enzymology / Q000378:metabolism*
D012333:RNA, Messenger; D000804:Angiotensin II; D007703:Peptidyl-Dipeptidase A