Uk. Messmer et al., Suppression of apoptosis by glucocorticoids in glomerular endothelial cells: effects on proapoptotic pathways, BR J PHARM, 129(8), 2000, pp. 1673-1683
1 Tumour necrosis factor-alpha (TNF-alpha)- and lipopolysaccharide (LPS)-in
duced apoptosis of bovine glomerular endothelial cells is now recognized as
an important part in the pathogenesis of glomerulonephritis characterized
by early mitochondrial cytochrome c release, mitochondrial permeability tra
nsition, Bak protein upregulation, Bcl-X-L protein downregulation and caspa
se-3 activation.
2 Co-treatment of cells with 10 nM dexamethasone and TNF-alpha or LPS block
ed roughly 90% of apoptotic cell death in glomerular endothelial cells. The
action of glucocorticoids could be documented in that they prevented all a
poptotic markers such as DNA laddering, DNA fragmentation measured by the d
iphenylamine assay as well as morphological alterations.
3 To mechanistically elucidate the action of glucocorticoids we evaluated w
hether glucocorticoids elicit a time-dependent effect. For dexamethasone, t
o maximally inhibit DNA fragmentation a preincubation period was not requir
ed. Even if dexamethasone was supplemented 6 h following TNF-alpha or LPS w
e observed a maximal inhibitory effect.
4 Concerning its influence on TNF-alpha and LPS signal transduction, we fou
nd that dexamethasone only partially prevented cytochrome-c-release as a fi
rst sign of apoptotic cell death but efficiently blocked mitochondrial perm
eability transition. Moreover, TNF-alpha- and LPS-induced Bak upregulation,
Bcl-X-L-downregulation, and the activation of caspase-3-like proteases, me
asured fluorometrically using DEVD-AMC and PARP cleavage, were efficiently
blocked by dexamethasone.
5 We postulate that glucocorticoids exert their inhibitory action upstream
of the terminal death pathways but downstream of primary receptor mediated
signals by blocking pro-apoptotic signals pre- and/or post cytochrome c rel
ease and mitochondrial signalling.