4p
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peroxisomes, catalase; nuclei, DNA. Good resolution of the
various organelles was obtained and the localization of the
various organelle markers was confirmed by the use of selective
membrane perturbants.
The equilibrium densities and enzymic activities have been
compared in lymphocytes from control subjects and patients
with adult-onset hypogamrna-globulinaernia. In the patients'
lymphocytes, there was a striking decrease in activity of
5'-nucleotidase but other plasma membrane marker enzymes
and the density distribution of this organelle were unaltered.
There was no change in the activity or distribution of
peroxisomal or lysosomal enzymes but the endoplasmic
reticulum showed a decrease in enzyme activity and in the
proportion of dense membranes. There was a small but
significant decrease in lactate and malate dehydrogenases but
the density distributions of the mitochondria were normal.
These results indicate that there are significant abnormalities
of the lymphocyte organelles in immunodeficiency disorders
but their relationship to the pathogenesis of the impaired
y-globulin synthesis remains to be determined.
11. MONOCYTE
FUNCTION
IN
RHEUMATOID
ARTHRITIS (RA) WITH VASCULITIS AND SYSTEMIC
LUPUS ERYTHEMATOSUS (SLE): EVIDENCE FOR A
DEFECT IN IMMUNE PHAGOCYTOSIS
N. HURST AND G. NUKI
Rheumatic Diseases Unit and Department ofMedicine (WGH),
University ofEdinburgh. Scotland, U.K.
Evidence is accumulating which suggests that cells of the
monocyte/macrophage system may play a key role in the
pathogenesis of RA. As part of a series of investigations of
monocyte function in patients with rheumatic diseases a method
has been developed for studying monocyte phagocytosis in cells
separated from to ml of venous blood. Pre-opsonized heat-killed
Candida alb/cans are mixed in suspension in Teflon wells and
numbers of the extracellular yeast remaining are counted at
timed intervals. Kinetic studies show that the rate of phagocytosis is first order with respect to both yeast and monocyte
concentrations so that the efficiency of phagocytosis can be
expressed by a rate constant.
Fourteen patients with definite or classical RA and 14 healthy
volunteers matched for age and sex have been compared. The
phagocytic rate constant was not different in patients with
rheumatoid factor (RF)-positive RA compared with controls but
was significantly reduced in all five RA patients with active
vasculitis. Depression of the phagocytic rate constant was not
correlated with clinical disease activity, erythrocyte sedimentation rate, titres of RF or antinuclear factors.
To determine whether this phagocytic defect is specific for
either the Fe or C, receptor on monocytes, the phagocytic rate
constant for both IgG-coated C. alb/cans and C,-coated
Saccharomyces cereuisiae has been measured in patients with
rheumatoid vasculitis and SLE. Preliminary results suggest that
there may be a selective C, receptor defect in rheumatoid
vasculitis, and in SLE the Fe receptor, the C, receptor or both
may be involved. Results of parallel measurements of total
complement activity (CH,o), complement components (C" C.'
C,) breakdown products) and C ,Q binding will be presented.
13. CHARACTERIZATION OF THE HEPATOCYTE
PLASMA MEMBRANE TRANSPORT MECHANISM FOR
LACTATE
J. P. MONSON, J. A. SMITH. R. D. COHEN AND R. A. ILES
Metabolic and Endocrinology Unit. The London Hospital.
London EIIBB
Preliminary evidence has been presented for a stereoselective
transport mechanism for lactate in hepatocyte plasma membranes (Monson et al., 1979, Clinical Science, 57, 30p-3Ip).
The present work extends the findings by examining the kinetics
of initial lactate uptake into isolated rat hepatocytes and the
effects of potential inhibitors of anion transport, changes in
extracellular pH and change in temperature on initial uptake.
The cellular uptake of L(+ )-lactate in IS s involves two
components; one is saturable but the other fails to saturate with
extracellular lactate concentrations up to 30 mmol/l, suggesting
passive diffusion as its likely basis. The passive diffusion
component may be largely an artifact of the isolated cell
preparation in which an abnormally large surface area is
presented for diffusion. The saturable component, which is
stereoselective, conforms to Michaelis-Menten kinetics, yielding
values for K m of 1·25 mmol/l and a minimum V max. of 8·3 pmol
min- 1 g-l wet WI. Lactate entry is not rate-limiting for overall
lactate removal under these conditions but could be in vivo
under some circumstances.
Initial entry of lactate on the transporter is inhibited by
d-cyano-3-hydroxycinnamate and by the thiol reagent p-chloromercuriphenylsulphonate, but not by the inhibitor of the general
anion
transporter 4-acetamido-4-isothiocyanostilbene-2,2'disulphonic acid (SITS).
Lactate entry exhibits a high temperature dependence up to
25°C but apparent activation energy falls at higher temperatures in keeping with the behaviour of several aniontransport mechanisms.
Initial rate of entry is enhanced by lowering extracellular pH
but the relationship of the lactate distribution ratio to that of H+
indicates that facilitated non-ionic entry is not the sole mechanism of 'carrier'-assisted transport. This finding could be explained
by either a hydrogen ion symport or hydroxyl ion antiport
mechanism with additional exchange of lactate for another
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intracellular anion. This mechanism is consistent with the known
intracellular alkalinizing effect of lactate metabolism in liver.
14. MEASUREMENT OF INTESTINAL PERMEABILITY
IN HUMAN NEONATES
scans and mean faecal isotope excretion was 0·4 ± SO 0·4%
(range 0·1-1·0%).
Indium-labelled leucocyte scanning with measurement of
faecal recovery may allow regional localization of Crohn's
disease and provide an objective measurement of leucocyte loss
into the intestine.
E. A. FAGAN, R. MURPHY, D. BENTLEY AND V. S. CHADWICK
Departments of Medicine and Paediatrics. Royal Postgraduate
Medical School, London WI20HS
A new high-pressure liquid chromatography (HPLC) technique
for measuring polyethyleneglycol (PEG) polymer molecules in
urine has been developed and applied to the study of intestinal
permeability in human neonates. A mixture of PEG polymer
molecules of different sizes (mol. wt, 300-4000) was administered by mouth and the differential absorption from the gut
assessed by the free excretion into urine over a 6 h period. A plot
of percentage absorption versus molecular weight, based on the
urinary excretion profile, was obtained from tests performed in
15 healthy full-term neonates (five aged I day; five aged 2-3
days and five aged 5-9 days) and compared with results in six
normal healthy adults. Day I babies showed markedly increased
gut permeability to PEG polymers of all sizes compared with
day 5-9 babies, whose excretion profile resembled that of adults.
The profile from day 2-3 babies was intermediate between those
of day I and days 5-9. No selectivity over the molecular weight
range 326-638 was observed in day I or 2 babies but a highly
selective profile (decreasing absorption with increasing
molecular weight) was achieved by day 6, in fact, in some day 6
babies the permeability profile was more selective than in adults.
Delayed acquisition of the adult-type selective profile was
observed in one day 9 baby.
Marked changes in gut permeability to inert polymer
molecules were observed in the first few days of life in human
neonates. These changes in man may represent the process of
intestinal 'closure' described in a variety of animal species.
16. CHOLESTERYL
CHOLECYSTITIS
ESTERS
IN
BILE
IN
E. KOUROUMALIS, P. E. Ross, D. HOPWOOD AND I. A. D.
BOUCHIER
Departments of Medicine and Pathology, Ninewells Hospital
and Medical School, Dundee. Scotland. U.K.
Gall-bladder bile samples from 77 patients with cholecystitis
were analysed by g.l.c. to determine cholesteryl ester concentrations. A few samples were identified by GC-MS.
In 20 samples biliary cholesterol (>4%) was esterified, mainly
with oleate or palmitate, while in eight white bile samples the
small amount of cholesterol present was mainly esterified (up to
90%). Cholesteryl ester profiles of gall-bladder mucosa and bile
differed, and it is therefore improbable that mucosa is the sole
source of biliary cholesteryl esters. This supposition was
supported by demonstration of cholesteryl esters in T-tube bile.
Sequential T-tube samples showed cholesteryl esters up to 6 h
after cholecystectomy but concentrations fell to negligible levels
within 24 h, suggesting that hepatic secretion of cholesteryl
esters may be regulated by the intact enterohepatic circulation.
Free fatty acid concentration showed an inverse relationship
with cholesteryl ester concentration during this time.
In chronic cholecystitis a negative correlation (r = -0·55, P
< 0·001) between esterified cholesterol concentration and bile
dry weight indicates absorption of cholesteryl esters by
gall-bladder mucosa in contrast with cholesterolosis, where
esters are apparently not absorbed.
We conclude that cholesteryl esters are found in hepatic
gall-bladder bile and that their presence depends upon an intact
enterohepatic circulation and functioning gall-bladder mucosa.
15. INDIUM-LABELLED LEUCOCYTE SCANNING IN
CROHN'S DISEASE
S. H. SAVERYMUITU, A. P. PETERS, J. P. LAVENDER, H. J.
HODGSON AND V. S. CHADWICK
Gastroenterology Unit. Royal Postgraduate Medical School.
London WI20HS
The distribution and fate of autologous indium-labelled leucocytes, after intravenous injection, was investigated in eight
patients with Crohn's disease, as an aid to assessment of disease
localization and severity. Control studies were performed in
eight patients with the irritable bowel syndrome, one patient with
an unexplained leucocytosis, but no intestinal disease. and one
patient with eosinophilic gastroenteritis. Blood (50 ml) was
taken and the leucocytes were labelled with '''indium (100-300
/lCi). After intravenous injection of the labelled cells, scans were
obtained, with a G.E.C. wide-angle gamma camera, at 3 hand
24 h. Stools were collected for 4 days and total indium content
measured to determine the per cent of administered dose
excreted. The proportions of faecal radioactivity in the particulate phase and supernatant were determined after high-speed
centrifugation of homogenized stool.
All patients studied showed hepatic and splenic uptake of
labelled cells. Intra-abdominal localization (graded + to + + + +)
was found only in the patients with Crohn's disease. Early scans
demonstrated the site of intestinal involvement and were
compared with radiological changes; later scans demonstrated a
column of labelled cells in transit through the intestine, distal to
the site of inflammation. Faecal radioactivity counting showed
mean recovery of 9·25 ± so 10·5% (range 1·9-34%) in the
patients. the most strongly positive scans producing the greatest
faecal recoveries of isotope. which was present mainly in the
particulate phase of stool. All the control subjects had negative
17. NO SUPPORT FROM COMPLEMENT PROFILES
INCLUDING IN VIVO C3 ACTIVATION FOR IMMUNE
COMPLEX AETIOLOGY IN THE PATHOGENESIS OF
PRIMARY BILIARY CIRRHOSIS
A. F. MACKLON. P. BIRD, G. BIRD AND O. JAMES
Medical Unit No.1. Freeman Hospital. Newcastle upon Tyne,
U.K.
Some evidence supporting the hypothesis that primary biliary
cirrhosis (PBC) is aetiologically associated with immune
complex formation and activation of complement was recently
challenged (Goldberg et al., 1980, Gastroenterology, 78, 1306).
We examined circulating complement profiles in 71 patients with
histologically and immunologically proven PBC: group I, 12
without symptoms and serum alkaline phosphatase (SAP) <
100 Lu./I; group II, 22 asymptomatic and SAP> 100 Lu./I;
Group III, 37 symptomatic patients (19 had received penicillamine, IIIP, 18 had not, IIINP). Results were compared with
23 female age-matched normal controls.
Total haemolytic complement (CH,o), factor B, total alternative pathway haemolytic activity (A PH A), C3 and C4
concentrations were measured in all. Products of C3 and factor
B activation were sought in 20 patients and 10 controls in serum
(calcium lactate buffer) and in EDTA plasma (EDTA buffer).
CH,o was increased in groups I, II and IIINP (P < 0·01) and
IIIP (P < 0·001). Factor B was higher than in controls in all
groups (P < 0·001). APHA was not significantly different from
controls in any group. C3 concentrations were increased in
groups II (P < 0·05), IIINP (P < 0·00 I) and IIIP (P < 0·0 I).
C4 concentrations were decreased only in group IIIP (P <
0·05).