Læknablaðið : fylgirit - 01.09.1993, Blaðsíða 34

32
LÆKNABLAÐIÐ/FYLGIRIT 24
Stimulation or Inhibition of Protein Kinase C
V 2 Blocks VLDL Secretion of Primary Hepatocytes
Cultures. 'O.G. Bjornsson, C. Bourgeois, G.F.
Gibbons. Metabolic Research Laboratory,
Radcliffe Infirmary, Oxford, U.K.
We have demonstrated that the calcium mob-
ilizing hormones angiotensin II, vasopressin,
and endothelin-1, suppress VLDL secretion of
primary hepatocytes cultures, and. even when
applied at picomolar concs. Partial depletion
of calcium from the endoplasmic reticulum (ER)
is a likely cause [1], but since these horm-
ones in addition to producing IP^-induced Ca^+
release from ER, also stimulate diacylglycer-
ol production and activation of protein kinase
C, this latter process could not be excluded.
In the present work we have applied the pro-
tein kinase C activator phorbol 12-myristate
13-acetate (PMA) (10~8 - 10”6 M), or the pro-
tein kinase C blocker chelerythrine (5 x 10"7
- 5 x ÍO"7^ M) to a 24 h rat hepatocytes cult-
ure. Both compounds suppressed VLDL triacyl-
glycerol secretion in a conc. dependent manner
(max. suppression of triacylglycerol to < 15%
of normal; 47 ± 10 pg/mg vs. control 410 ± 38
pg/mg, SEM, n = 4, P < 0.01). The 4 oC-phorbol
12-myristate 13-acetate, which has no effect
on protein kinase C, had minimal effect on
VLDL secretion. None of the abovenamed hormon-
es or compounds affected ketogenesis, indicat-
ing that the reduction in VLDL secretion was
not due to enhanced fatty acid oxidation of
the cells. On the contrary, PMA and cheleryth-
Regulation of Hepatic VLDL Metabolism by cAMP
and Ca2+ Mediated Signal Transduction Mechan-
isms. ^Ö.G. Bjornsson, G.F. Gibbons. Metabolic
Research Laboratory, Radcliffe Infirmary,
Oxford, U.K.
Previously we have demonstrated that com-
pounds which raise cellular levels of cAMP
(incl. glucagon, forskolin), suppress VLDL
secretion, and that the specific protein kin-
as A blocker, RpcAMPS, inhibits this mechan-
ism [1,2]. We have also shown that compounds
which affect cellular [Ca2+]i (e.g verapamil,
A23187) are able to suppress VLDL secretion
to almost undetectable levels [3]. The pre-
sent studies show that the initial transient
rise in cellular cAMP levels induced by gluca-
gon (10"7 M) is sufficient to maintain a long-
term (24 h) inhibitory effect on the VLDL
metabolism. Forskolin (10"4 M) has a similar
effect on VLDL metabolism as glucagon and
also causes a transient increase in cellular
conc. of cAMP, while the forskolin analogue
1,9-dideoxyforskolin suppressed VLDL secret-
ion without having any effect on cAMP. VLDL
secretion could also be varied by varying the
conc. of Ca2+ of the culture medium, and VLDL
secretion was blocked by thapsigargin, the
inhibitor of the Ca2+/ATPase pump of the
endoplasmic reticulum. The calcium mobilizing
hormones angiotensin II, vasopressin and endo-
thelin-1 (10"12 - 10"7 M), or phenylephrine
(10"9 - 10"4 M), all suppressed VLDL secret-
ion in a conc. dependent manner in cells cult-
rine caused a moderate increase in cellular
levels of triacylglycerol (max. increase to
163% of control) and cholesterol (max. 150%),
suggesting that reduced VLDL secretion, ob-
served in the presence of these compounds,
led to accumulation of lipids within the
cells.- These data are consistent with the
hypothesis that hormones which stimulate
polyphosphoinositide hydrolysis may regulate
hepatic VLDL metabolism by mechanism which
involves protein phosphorylation induced by
protein kinase C. - [1] Lodish, H.F., N. Kong
J. Biol. Chem. 265:10893-10899, 1990.
ured in normal medium (calcium conc. 0.8 mM);
however, by raising the Ca2+ conc. of the
medium to 2.4 mM, the suppressive effect of
the hormones on VLDL secretion was abolished.
The present data support further the idea that
cAMP and Ca2+ regulate VLDL metabolism. The
data also indicate that hepatic VLDL secretion
can be regulated by mechanism which is inde-
pendent of these two intracellular mediators.
- [1] Biochem. J. 281:381-386, 1992; [2] Eur.
J. Clin. Invest., in press; [3] J. Lipid Res.
33:1017-1027, 1992.