Medical Research Society
52 P
gens. More recent studies have shown associations with
the DQ subregion. The DP subregion lies centromeric to
DQ, and although DP-typing can be performed by primed
lymphocyte testing, this technique is difficult and not
widely available. The aim of this study was to examine
the DP-alpha gene with regard to susceptibility to CD,
using recombinant DNA techniques.
DNA from 58 CD patients and 80 healthy controls was
studied by Southern blotting, using a Bgl I1 digest and
a genomic DP-alpha probe (kindly provided by J. Trowsdale,
ICRF, London). This detects a polymorphism characterised
by allelic fragments sized 3.5 and 2.2 kb. The distribution of genotypes in controls and patients is shown in
the table:
genotype:
controls n=80
CD
n=58
3.5/3.5
3.5/2.2
2.2/2.2
3.75%
32.5%
74.1%
63.75%
25.9 %
0
%
p(O.0001
Thus CD is strongly associated with possession of the 3.5
kb allele; the relative risk (calculated by the method
of Woolf) is 5.0 (95% confidence limits 2.2 - 7.9).
We conclude that genes important in determining susceptibility to CD are located centromeric to the HLA-DQ
subregion and closer to the HLA-DP subregion.
THE APOPROTEIN B
GENE I N EXTRACORONARY ATHEXOSCLEROSIS
185 A POLYMORPHISM OF
G.
O'Connor, J .
Stocks, and D.J. Galton.
Diabetes
and
Lipid
Bartholomew's Lospital,
Laboratory,
LONDON EC1 7 B E .
St.
Athrrosclerosis
has
a
strong
familial
com?onent and
reconbinant DNA techno lo:,^ can
be used to identify the genes responsible for
this.
The apoprotein B gene is a useful
candidate,
since it's product i s the major
of
low density
prorein
component
lipoproteins.
We
studied a group of 4 6
patients
with
angio~raphically
proven
si t e s
at
e 1: t ra c o r ona r y
at hero s c 1e K O s i s
(carotid and aortofemoral)
to determine the
frequencies of restriction fragment
length
polymorphisms.
These
were
identified by
hybridization of a cloned human apoprotein E
gene to patient DNA which v7as digested with
the restriction endonuclease
Xba-1.
The
results were compared with those f r o m a group
o f 4 7 healthy controls selected from a health
screen in^ clinic.
from
whole
blood
and
DNA was isolated
dizested vith Xba-1.
The resultant fragments
were s ~ p a r a t e d according
to size using
agaruse &el electrophoresis, and
transferred
Southern blotting.
to a nylon nembrane b y
T h e position of tiLe fraL;ments was revealed by
hybridization to a labelled apo-B gene probe
and autoradioGraphy.
The fragments obtained wPre 8.6 kilobase
5.0 kilobase pairs ( X 2
pairs ( X I allele) and
allele)
in length. The X 1 allele frequency
0.5 in patients.
was 0.415 in cor.tro1.s and
The X 2 allele frequencies w e r ~0 . 5 6 5 and 0.5
respectively (chi square = 4 . 4 3 ,
p < 0.05).
The genotype frrquencies did
not
differ
between the groups.
186
SPECIFIC INHIBITION OF MACROPHAGE PHAGOCYTOSIS OF
SENESCENT HUMAN NEUTROPHILS BY MONOSACCHARIDES
J. SAVILL AND C. HASLETT. Department of Medicine, Royal
Postgraduate Medical School, DU Cane Road, London W12 OHS.
Macrophage (Md) phagocytosis of neutrophils (PMN) occurs
in vivo. At inflammatory sites, specific recognition and
"disposal" of intact effete PMN by Md may limit tissue
injury by preventing release of phloqistic contents from
PMN otherwise dooned to disintegration. In vitro evidence
suggests that aging PMN may develop specific surface
changes recognizable to Md: human PMN isolated from normal
peripheral blood and "aged" in culture for 24 hours are
phagocytosed by monocyte derived Md; freshly isolated PMN
are not recognized. HLA compatibility and serum are not
required (1). We have developed a 30 minutes serum free
assay of M+aged PMN interaction, quantified by microscopy
and myeloperoxidase staining. At 37OC and pH 7.4. 46.1
+2.3% (mean fSE, n=159) of Md ingested aged PMN. (Only
3.4 +0.3%, n=36, of Md recognize freshly isolated PMN).
A specific sugar-lectin recognition system in Md-aged PMN
interaction was sought by inclusion of monosaccharides at
lOmMolar (ionic strength 0.145) and 5 0 mMolar (ionic
strength 0.125) in the assay at pH 7.4; the proportion of
Md ingesting aged PMN was expressed as % of control (mean
tSE; *indicates pCO.01 by Wilcoxon rank sum).
Glucose (10 mM:95.9 f2.1%, n=12 - 50 mM:98.1 +4.6%, n=5),
Fucose (lOmM:99.6 +2.6%, n=12 - 50mM:lOO.l ?3.0%, n=8),
Mannose (lOmM:99.1 +3.0%, n=10 - 50mM:100.5 ?3.9%, n=6),
Galactose (10mM:100.2 +3.1%, n=10 - 50mM:gg.l fl.l%, n=6),
N-acetylglucosamine (lOmM:97.5 +1.6%, n=15 - 50mM:100.3
+2.7%, n=8) and N-acetylgalactosamine (10mM:gg.g +2.4%,
n=10 - 50mM:100.2 +2.2%, n=8) had no effect on Md phagocytosis of aged PMN. However, the amino-sugars glucosamine
(1OmM:39.7 ?1.6%*, n=32 - 50mM:23.5 ?3.1%*, n=10) and
galactosamine (lOmM:49.5 f2.3%*, n=15 - 50mM:27.4 ?0.9%,
n=5) specifically inhibited Md phagocytosis of aged PMN phaqocytosia of opsonised ox erythrocytes was not affected
(>99%) nor was aged PMN viability (>97% by trypan blue
exclusion). In summary, glucosamine and galactosamine
specifically inhibit Md phaqocytosis of aged PMN, suggesting that a lectin-like cell surface structure may mediate
this interaction.
(1) Newman et al, J Exp Med 1982: 156
:
430.
187
REDUCED PIdDTEIN SYNTHESIS A3 A CAUSE OF AKOHOLF
INDUCED SKELETIU. MUSCLE MYVPAPAI
V.R. PREEDY AND T.J. PETERS
Division of Clinical Cell Biology,
Centre, Harrow, Middlesex
MRC
Clinical Research
Alcoholic patients frequently exhibit skeletal muscle
weakness and wasting. Histological investigations have
revealed a selective reduction in the diameter of Type
11 fibres. Type I fibres are relatively unaffected.
These changes may be the direct result of ethanol
toxicity on protein synthesis in skeletal muscle. To
test this hypothesis we measured fractional rates of
protein synthesis in 1009 male rats (n-7-9) given an
i.p. injection of ethanol (75nmol/kg BW). Controls were
injected with an isovolumetric amount of 0.15mol/l NaC1.
At the e d of 140 min rats were intravenously injected
with [ 4 'Hlphenylalanine ( 1 5 O ~ l / l O O gBw) to measure
fractional rates of protein synthesis. Results, based
on the assumption that the precursor phenylalanine waa
derived from the intraoellular pool, shared that the
synthesis of myofibrillar proteins in gastrocnemius
muscle fell by 26% (p < 0.01). A similar decline was
observed in sarcoplasmic ( 2 6 % fall, p < 0.01) and
stromal fraction8 (29% fall, p < 0.01). Rates of
protein synthesis w e r e also examined in Type I
(plantaris) and Type I1 (soleus) fibre-rich muscles.
The decline in the Type I muscle (22% fall, p < 0.01)
was less than the decline in Type I1 muscles (30% fall,
p < 0.001). There was a significant change in the Type
I/Type I1 ratio of synthesia rates (11% increase, p <
0 . 0 5 ) . Similar fractional synthesis rates were obtained
when the data was calculated from the assumption that
the precursor phenylalanine was derived from the
extracellular pool. In conclusion, muscle protein
synthesis is reduced in direct response to ethanol. The
relative sensitivity of protein synthesis in Type I1
fibre-rich muscles may be related to the selective
reduction in the diameter of Type I1 fibres of alcoholic
patients.