Download Lipogenesis in Sheep Adipose Tissue Maintained in Tissue Culture

BIOCHEMICAL SOCIETY TRANSACTIONS
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Fig. 2. Inactivation-re-activation cycle of microsoma/ 3-hydroxy-3-methylglutarylcoenzyme A reductase in thepresenceof 6m~-[y-~~P]ATP(specifcactivity
1.41 x lo7c.p.m.1
nmol) and 20 mM-MgC12
Twice-washed microsomal fractions were incubated at 30°C at different times with
Mg[p3’P]ATP. At 20min, a glycerol/glycerol kinase mixture was added to one series
of tubes (v), whereas to another series also 0.4ml of 105000g supernatant corresponding to 12mg of protein was added in addition (0).The 3-hydroxy-3-methylglutaryl-coenzyme A reductase activity was measured and the percentages of the initial
value (15.6nmol of mevalonate/h per mg of protein) are represented on the left ordinate
(closed symbols). The 32P bound to microsomal protein was measured and the 32P
(pmol) per mg of protein is represented on the right ordinate (open symbols).
methylglutaryl-coenzyme A interconversion (which appears independent of cyclic
AMP) still need to be answered.
Lipogenesis in Sheep Adipose Tissue Maintained in Tissue Culture :
Effects of Insulin and Growth Hormone
RICHARD G . VERNON
Department of Physiology, Hannah Research Institute, Ayr KA6 5HL, Scotland, U.K.
Early lactation in both ruminants and non-ruminants is associated with a decrease in
the rate of lipogenesis and lipoprotein lipase activity in adipose tissue (Hamosh et al.,
1970; Shirley et al., 1973;Smith, 1973;Gritchting et al., 1977), presumably to ensure
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that nutrients are preferentially utilized by the mammary gland. Prolactin is involved in
the suppression of lipoprotein lipase activity in rat adipose tissue (Zinder et al., 1974),
but other mechanisms may also operate, particularly in the ruminant, in which, in
contrast to the non-ruminant, prolactin does not appear to be necessary for normal
milk production once lactation is established (Tucker, 1974). Growth hormone, however,
is essential for lactation in ruminants (Tucker, 1974).Growth hormone is galactopoietic
in ruminants, but not in rodents (Tucker, 1974), also growth hormone concentrations
in plasma from ruminants are elevated during lactation (Forsyth & Hart, 1975). As
growth hormone has been shown to influence the metabolism of rat adipose tissue, it
seemed pertinent to investigate the effects of the hormone on the metabolism of ruminant
adipose tissue.
Sheep adipose-tissue slices were maintained in tissue culture in Medium 199
containing 25m~-Hepes,*pH 7.3, at 32°C; insulin (kindly donated by Boots Ltd.)
and sheep growth hormone (N.I.H. S11, 0.56i.u./mg, kindly donated by National
Institute of Arthritis, Metabolism and Digestive Diseases, U.S.A.) when included in
the culture medium were each at a concentration of 10pg/ml. After 1 , 2 or 3 days in culture, tissue slices were transferred to fresh Medium 199 containing insulin (lOpg/ml)
and the rates of lipogenesis and acylglycerol (glyceride) glycerol synthesis were
determined as described previously (Vernon, 1977).
As shown in Fig. 1, lipogenesis from both acetate and glucose was stimulated by
insulin during tissue culture. Growth hormone antagonized this stimulatory effect of
insulin on lipogenesis, but at least 24h in culture were required for this effect of growth
hormone to become manifest. In some experiments growth hormone by itself had an
insulin-like effect on the rate of lipogenesis. Inclusion of insulin in the culture medium
*Abbrevation used: Hepes, 4-(2-hydroxyethyl)-l-piperazine-ethanesulphonicacid.
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also increased the rate of acylglycerolglycerolsynthesis from glucose and therate of lactate
release into the culture medium, indicating that insulin was stimulating a general
increase in the rate of glucose uptake and utilization. Growth hormone, however,
did not antagonize the effect of insulin on acylglycerol glycerol synthesis or lactate production. Also, growth hormone did not affect the rate of glycerol release into the culture
medium, indicating that it was not affecting the rate of lipolysis.
Growth hormone, in the presence of insulin, thus had a relatively specific effect on
the rate of lipogenesis, which could not be attributed to a decreased rate of glucose
uptake or an increased rate of lipolysis.
The growth hormone used in the present study contained small amounts of other
pituitary hormones, and possible effects of these contaminants on sheep adipose-tissue
metabolism have yet to be assessed. However, the results to date suggest that growth
hormone has a direct effect on sheep adipose-tissue metabolism and that the change in
plasma growth hormone/insulin ratio observed during lactation in ruminants may at
least be partly responsible for the decreased rate of lipogenesis in ruminant adipose tissue
during early lactation.
I thank Miss S . M. Park and Mr. E. Finley for technical assistance.
Forsyth, I. A. &Hart, I. C. (1975) in Growth Hormone and Relatedpeptides (Pecile, A. & Muller,
E. E., eds.), pp. 422-432, Elsevier, New York
Gritchting, G., Baldwin, R. L. & Smith, N. E. (1977) J. Dairy Sci. 60,Suppl. 1, 197a
Hamosh, M., Clary, T. R., Chernik, S. S. & Scow, R. 0. (1970) Biochim. Biophys. Acta 210,
473-482
Shirley, J. E., Emery, R. S., Convey, E. M. & Oxender, W. D. (1973) J . Dairy Sci. 56,569-574
Smith, R. W. (1973) J. Dairy Res. 40,353-360
Tucker, H. A. (1974) in Lactation (Larson, B. L. & Smith, U. R., eds.), vol. 1, pp. 277-326,
Academic Press, New York and London
Vernon, R. G. (1977) Int. J. Biochem. 8 , 517-523
Zinder, O., Hamosh, M., Fleck, T. R. C. &Scow, R. 0. (1974) Ant. J . Physiol. 226,744-748
The Urea Cycle in Different Types of Macrophages
FRIEDRICH HOFMANN, JURGEN KREUSCH, KLAUS-PETER MAIER,*
PAUL GERHARD MUNDERt and KARL DECKER
Biochemisches Institut an der Medizinischen Fakultat der Universitat,
*Medizinische Universitatsklinik and ?Max-Planck-Institut fur Immunobiologie,
07800 Freiburg im Breisgau, West Germany
Since Krebs & Henseleit (1932) described the urea cycle, urea formation from
ammonium and bicarbonate ions has been considered as a specific function of
mammalian liver. In the course of studies on the localization of urea-cycle enzymes
we found urea formation from various amino acids (Wagle et al., 1976) by a complete
urea cycle (Kreusch et al., 1977) not only in hepatocytes but also in the sinusoidal fraction
of liver cells.
As Kupffer cells (the major part of this fraction) are considered as a part of the
macrophage system (van Furth, 1975), it seemed advisable also to study urea formation
and the urea-cycle enzymes in various types of macrophages.
Mice bone marrow (Metcalf, 1977) and rabbit alveolar macrophages (Myrvik et al.,
1962) were isolated by routine methods. Rat peritoneal macrophages were obtained by
the lysolecithine method (Munder & Modolell, 1974). Rat hepatocytes were prepared as
described by Hofmann &Decker (1975). Viability of cells was always measured by determination of enzyme leakage into the incubation medium. This medium consisted of
Krebs-Henseleit (1932) buffer, pH 7.40, supplemented with 5.5m~-~-glucose.
Incubations were performed in stoppered plastic vials containing 10-20mg of cells/ml at
100 oscillations/min in a metabolic incubator (37°C) under an atmosphere of 0 2 / C 0 2
(19 :I).
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