Am. Rodriguez-lopez et al., Increased apoptosis susceptibility in mesangial cells from spontaneously hypertensive rats, MICROVASC R, 59(1), 2000, pp. 80-87
We have examined the susceptibility to apoptosis in mesangial cells from sp
ontaneously hypertensive rats (SHR) or from normotensive rats (WKY) and the
possible involvement of nitric oxide in this process. Mesangial cells mono
layers from either SHR or normal rats were incubated for 12 h in medium wit
h or without fetal calf serum (FCS) and with or without thapsigargin (Tg, 1
0(-6) M). A series of cultures from rats of both groups was treated with N-
G-nitro-l-arginine methyl ester (L-NAME, 10(-4) M). We assessed apoptosis b
y propidium iodide staining, by TUNEL nitrite production (Griess reaction),
by inducible nitric oxide synthase (iNOS) and Bcl-2 and Bas by Western blo
t. Incubated with a FCS-free medium, cells from SHR showed a significantly
higher apoptotic rate (10.7 +/- 2.0) than with 10% FCS (10% FCS, 4.7 +/- 0.
3), while WKY cells did not show this increment (10% FCS, 4.7 +/- 0.3; 0% F
CS, 5.9 +/- 0.3). Apoptosis in cells from WKY increased when incubated with
thapsigargin in FCS-free medium (0% FCS+ Tg, 17.7 +/- 2.9%) and increased
even more in SHR cells (0% FCS+ Tg, 19.7 +/- 2.9%). Treatment with L-NAME d
ecreased thapsigargin-induced apoptosis in both SHR (8.2 +/- 2.4%) and WKY
cells (9.3 +/- 2.4%). An increase in nitrite production and iNOS expression
was detected in groups in which the apoptosis rate was elevated. A high ra
te of apoptosis was also associated with a decrease in the Bcl-2/Bax ratio.
Our results indicate that in SHR cells, short-term serum deprivation and t
he increase in intracellular free calcium concentration with thapsigargin a
re able to enhance the apoptosis rate in primary cultures and that the expr
ession of iNOS, and hence NO production, is involved in this effect. (C) 20
00 Academic Press.