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External counting and scintiphotography in rabbits with arthritis.

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External Counting and Scintiphotography in Rabbits with Arthritis
By STEPHEN
D. SHOLKOFF,M.D., MORTONG. GLICKMAN,
M.D.,
JULIUSSCHACHTER,
PH.D., AND MALCOLMROWLAND,
PH.D.
Bedsonia arthritis in rabbits was studied
b y Anger camera scintiphotography and
external counting, using pertechnetate
99m as the label. This method of evalu-
A
MEMBER of the bedsonia group of organisms (psittacosis, lymphogranuloma
venereum, trachoma) has been identified
as a possible etiologic factor in some cases
of Reiter's syndrome.lS2 A long-term study
concerning the bedsonia group is under
way at this institution. Joints of rabbits and
monkeys have been infected by intraarticular injection of bedsonia organisms isolated from patients with classical Reiter's
syndrome? Neither clinical examination nor
conventional roentgenography of these animals has satisfactorily demonstrated the
varying intensity of arthritis. The clinical
evaluation of arthritic activity is subject to
observer error and cannot be recorded
graphically. Roentgenography does not
easily demonstrate rapidly changing i d a m matory manifestations, since structural
changes of bone and cartilage are late in
appearance and are slow to evolve,.
In seeking an objective and graphic
method for evaluating the arthritis, we
From the Department of Radiology, the G. W .
Hooper Foundation, and the Department of Pharmaceutical Chemistry, University of California,
Schools of Medicine and Pharmacy, San Francisco.
This investigation was supported b y NIH Training Grant GM 01272 frm the National Institutes
of General Medical Sciences, and Research Grant
Al 07698 from the National Institutes of Health.
STEPHEN D. SHOLKOFF, M.D.: Research
Fellow, Department o f Radiology, University of
California School of Medicine, San Francisco,
California. MORTONG. GLICKMAN,M.D.: Instructor in Residence, Department of Radiology,
220
ating the intensity of the arthritis and the
changes in the disease proved more sensitive and more accurate than did clinical and roentgenographic examinations.
studied external counting technics and
gamma scintiphotography in rabbits infected with bedsonia.
MATERIALSAND METHODS
Nine rabbits were infected with bedsonia organisms by intraarticular injection into one knee
joint. This injection consisted of 0.5 ml. of a 10
per cent yolk sac suspension of isolate 25 SM containing approximately 106 egg LD,,. The rabbits
were evaluated by clinical, roentgenographic, and
scintiphotographic examinations over a period of
3 weeks.
The scintiphotographic studies were conducted
with the Anger scintillation camera, equipped with
a highspeed automatic digital printer.* A 4000
parallel hole, low energy collimator was used. The
animals were immobilized with a recently devised
apparatus to ensure identical positioning each time
a rabbit was restudied.4 The knees were placed
under the crystal, and the trunk and lower legs
were shielded with lead (Fig. 1). Scintiphoto'Pho-Gamma I11 Scintillation Camera, Nuclear
Chicago Corporation, 333 E. Howard Ave., Des
Plaines, Ill. 60018.
University of California School of Medicine, San
Francisco, California. JULIUSSCWCHTF.R, Ph.D.:
Assistant Professor of Epidemiology and Medicine,
G. W . Hooper Foundution, University o f California
School of Medicine, Sun Francisco, California
MALCOLMROWLAND, Pn.D.: Assistant Professor,
Department o f Pharmaceutical Chemistry, University of California School of Pharmucy, San Francisco, California.
Reprint requests should be addressed to Dr.
Sholkofl, Department of Radiology, Uniljersity of
California, San Francisco Medical Center, San
Francisco, California 94122.
ARTHRITISAND RHEUMATISM,
VOL. 12, No. 3
(JUNE 1969)
221
EXTERNAL COUNTING AND SCINTIPHOTOGRAPHY
Fig. 1.-The rabbit is in immobolized position for the study with neck extended
and drawn through a notch in the boards. The camera will be positioned over the
unshielded knees. The lead extends beyond the margins of the crystal so that only the
knees are monitored.
graphic examinations were performed after intravenous injection of 500 pc of pertechnetate 99m
(99mTc0,-) at varying intervals after infection, as
shown in Table 1.
The crystal was split electrically so that each
knee was counted separately. The counts were
collected continuously for 3 to 13 hours after each
injection, and the number collected was printed
automatically at 1min. intervals during each study
period. Photographs were exposed by collecting
2000 counts.
Control studies were performed prior to the
bedsonia infection. Three animals had intravenous
99mTc04- studies beginning at 8 and 48 hours
after injection of 1.0 ml. saline into a knee joint.
This was done to ensure that needle introduction
did not produce early detectable change in the
labeling characteristics of the joints. Subsequent
intravenous studies 1 to 12 weeks later were per-
formed on these rabbits to c o & m that the previous intraarticular injections had been innocuous.
RESULTS
Neither
scintiphotography, external
counting, roentgenography, nor physical
examination demonstrated any abnormalities in the control studies.
At frequent intervals after infection, all
the joints were examined for heat, swelling,
and passive range of motion. During the
first 12 hours after infection, no abnormality was discernible. At 48 hours, severe inflammatory response was demonstrated in
all the inoculated joints. By 3 weeks, the
range of motion in all rabbits had de-
SHOLKOFF ET AL.
Fig. 2.-Roentgenogram 3 days after infection of the left knee. Although not clearly
reproduced, minimal soft tissue swelling is present over the left knee on the original
film. No other abnormalities are seen.
creased and in 5 of these soft tissue thickening was detectable. By that time, however, neither warmth over the joint nor effusion was evident.
The roentgenograms demonstrated minimal soft tissue swelling as early as 24 hours
after inoculation (Fig. 2). This finding remained constant, but neither bone changes
nor evidence of cartilage destruction was
present at any time during the 3 weeks of
study.
As early as 4 hours after infection, the
intravenous pertechnetate studies showed
increased radioactivity in the affected knee
relative both to the contralateral knee and
to the knees as they had appeared prior to
infection (Figs. 3A and B, 4A and B). This
relationship persisted throughout the
study, as shown by the scintiphotographs
in Figs. 5 and 6.
The results can be expressed numerically
as the ratio, Ci/C,, where C , = maximum
counts per minute from the infected knee,
and C, = counts at that time from the
contralateral (normal) knee (Table 1).For
those animals studied prior to infection, the
counts over the knees were maximum between 10 and 50 min. (mean = 33) after
injection of pertechnetate. In the infected
animals, the maximum count rate was
reached at 20 to 58 min. (mean = 42).
The range of counts collected per minute
at the maximum for each study was 3200
to 10,000. The room background was 200
counts per minute. It was measured before
and after each study, using the same detector.
The varying intensity of the arthritis is
easily appreciated by comparing the ratios
at different times. The use of ratios rather
than absolute numbers eliminated inaccuracies caused by differences in animal
weight, symmetry of the extremities, response to infection, rate of excretion of
pertechnetate, and other natural variables.
Histologic examination of the infected
joints of 2 rabbits at the end of the study
confirmed the presence of chronic inflam-
EXTERNAL COUNTING AND SCINTIPHOTOGRAPHY
223
Fig. 3.-(A) Rabbit 1: Intravenous 9SmTcOp- study performed one week after intraarticular injection of 1.0 ml. saline into the right knee, and prior to infection shows no
detectable abnormality. (B) Rabbit 1: Increased labeling of the left knee is present as
early as 6 hours after infection with bedsonia.
SHOLKOFP ET AL.
Fig. 4.-(A) Rabbit 2: Normal intravenous 9grnTcO4- study before infection. (B)
Rabbit 2: Twenty-fourhours after infection of the left knee.
.TERNAL COUNTING AND SCINTIPHOTOGRAPHY
Fig. 5.-Rabbit
1: Intravenous study at 3 weeks.
Fig. 6 . R a b b i t 2: Intravenous study at 3 weeks.
226
SHOLKOFF ET AL.
Table 1.-Ratio
Rabbit
Number
Initial
Ratio
1.08
033
0.91
1.03
0.96
1.04
0.93
N.D. $
1
2
3
4
5
6
7
8
9
N.D.$
Ci/C, for Each Intravenous Pertechnetate Study
4 to 6 hr..
ratio
24 hr.,
ratio
Time After Infection*
48 hr..
ratio
98 hr.,
rPti0
1.18
2.34t
2.18
0.93 **
1.15
3 wk.
IUtiO’
1.21
1.40
1.36
1.20
1.24
1.26
1.44
1.72
1.90
* The observations at the above times were compared to the initial values and found to be signifi0.01.The ratios obtained in the studies after infection have each been divided by their
cant, P
corresponding initial value so that they will be comparable with one another.
t Clinically the most severely involved, the only animal in which severe decreased range of motion
was evident during the h s t week after infection.
$ N.D. =Not done. The mean of the other studies was 0.98. The theoretically expected value
was 1.00.
** Confirmed negative study by scintiphotography. All the others were obviously positive by sdntiphotography.
<
gressed. Structural bone abnormalities and
osteoporosis were not manifest during the
study and would not be expected to respond to changes in the disease.
Scintiphotographic technics demonDISCUSSION
strated evidence of inflammation before
Organisms of the bedsonia group of bac- such inflammation was seen clinically. No
teria produce a chronic inflammatory ar- false positive or false negative results were
thritis in rabbits. Regardless of whether this obtained. The intensity of local involveagent is related to Reiter’s syndrome, acute ment by inflammation could be graded oband chronic arthritis develops in rabbits in- jectively by external counting and scintifected with bedsonia. These animals there- photographs. On serial studies, scintifore are ideal models for the study of meth- photographs and ratios of counts collected
ods to evaluate arthritis.
from the knees correlated well with each
In our experimental animals, clinical ex- other and with the changing clinical state.
amination failed to detect the disease in its
We believe that this method of examinaearly stages and was consistently accurate tion is the most valid, simple method of
only in the most florid stage. Because the evaluating early stages of arthritis and its
results were not quantifiable, an accurate changing state. It is useful in animal reestimate of improvement, progression, or search, where objectivity is essential and a
stability of the disease was impossible on clinical history is not possible. Its potential
usefulness in clinical practice will be the
subsequent examinations.
The roentgenograms demonstrated soft subject of a later report.
Pertechnetate scintiphotography is not
tissue swelling early, but the degree of
swelling did not change as the disease pro- specific for detection of arthritis. Any local-
matory changes. Bedsonia organisms were
reisolated from the synovial membranes.
The noninjected knee of each rabbit was
normal.
227
EXTJ3RNAL COUNTING AND SCINTIPHOTOGRAPHY
ized inflammatory process might produce
similar abnormal scintiphotos and count
ratios and could therefore be studied by
this method.
SUMMARIO
IN INTERLINGUA
Arthritis a Bedsonia in conilios esseva studiate per scintiphotographia a camera
Anger e contation externe, con le us0 de pertechnetato-99m como marca. Iste methodo
d e evalutar le intensitate del arthritis e le alterationes occurrente in le curso del
tempore in le morbo se provava plus sensibile e plus accurate que examines clinic e
roentgenographic.
REFERENCES
1. Schachter, J., Barnes, M. G., Jones, J. P., Jr.,
Engleman, E. P., and Meyer, K. F.: Isolation of
bedsoniae from the joints of patients with Reiter's
syndrome. Proc. SOC. Exp. Biol. Med. 122:283,
1966.
2. Amor, B., Coste, F., and Delbarre, F.: Mise
en Lvidence, par culture d'agents du genre BedSonia dans le liquide articulaire en cas de rheu-
matisme de Fiessinger-Leroy-Reiter (F. L. R. ). C.
R. Acad. Sci. [D] 264:1365, 1967.
3. Smith, D. E., James, P., Schachter, J., EngIeman, E. P., and Meyer, K. F.: Experimental bedSonia arthritis in the rabbit. In preparation.
4. Sholkoff, S. D., Glickman, M. G., and Powell,
M. R.: Restraint of small animals for radiopharmaceutical studies. In preparation.
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