The Latex fixation test in the spinal fluid and urine of patients with rheumatoid arthritis.код для вставкиСкачать
The Latex Fixation Test in the Spinal Fluid and Urine of Patients with Rheumatoid Arthritis By WILLARD R. STARNES, ALEXANDERULLOAAND HOWARD L. HOLLEY The finding of an abnormal macroglobu- Plure gruppos de investigatores ha isolin in the serum of patients with rheu- late e characterisate un macroglobulina matoid arthritis has been isolated and anormal in le sero de patientes con characterized by several groups of in- arthritis rheumatoide. Esseva studiate vestigators. A study was undertaken to le question del presentia de iste globustudy other biological fluids as to the lina anormal in altere liquidos biologic. presence of this abnormal globulin. Liquid0 spinal e urina esseva obtenite Spinal fluids and urines were obtained ab patientes con arthritis. Le patientes from patients with arthritis. The pa- esseva dividite in tres gruppos: (1) tients were divided into three groups: Spondylitis ankylosante; (2) osteoarth(1) ankylosing spondylitis; (2) osteo- ritis; e (3) arthritis rheumatoide, active arthritis; and (3) rheumatoid arthritis, e non-tractate. Le sero ab patientes con active, untreated. The serum of patients arthritis rheumatoide esseva positive in with rheumatoid arthritis gave positive le tests de fixation a latex. Le test de latex fixation test. Latex fixation test on fixation a latex in le liquid0 spinal ab the spinal fluids from the three groups omne le tres gruppos esseva negative gave a negative latex agglutination. The pro agglutination a latex. Le urinas e urines and spinal fluids were concen- le liquidos spinal esseva concentrate a trated to a known volume with poly- un cognoscite volumine per medio de vinyl pyrrolidone (P.V.P.).There was pyrrolidona polyvinylic. Esseva trovate found an increase in the total protein un augment0 del total hexosas ligate a bound hexoses in the three latex posi- proteina in le tres concentratos urinari con positivitate a latex. tive urine concentrates. A BNORMALITIES IN THE electrophoretic pattern of proteins in patients with rheumatoid arthritis have been identified in serum, synovial fluid and spinal It has been reported that in the serum and spinal fluid of patients with active rheumatoid arthritis, there was an increase in the relative concentration of the alpha-1, alpha-2, and gamma globulins and a decrease of the albumin fraction; the tendency being for the alpha globulins to rise early in the course of the disease, followed by au elevation of the gamma globulin.* Causes for alterations in an individual protein fraction, however, are not well known, although there is evidence that thc gamma globulins are increased whenever considerable inflammation or tissue destruction is present or when the gamma globulin antibody formation is increased. The rheumatoid factor, a hemoagglutinating substance, has been isolated or partially isolated and characterized from the serum and synovial fluid of patients with rheumatoid a r t h r i t i ~By . ~ means of electrophoresis of those fluids, by the factor has been found to migrate within the gamma f r a ~ t i o n However, .~ ~ _ _ _ From the University of Akh:ir:ru Medicul Center und Veterans Administration Hospital, Birmingham, Ala. Supported in part by grant (A-3555) from the National institute of Arthritis and Metabolic Diseases, National Institutes of Health, Bethesda, Md. 364 LATEX FIXATION TEST IN RHEUMATOID ARTHRITIS 365 means of ultracentrifuge studies it has been demonstrated that the factor can be separated from the normal gamma globulin.G Since Singer and Plotz6 first applied latex particles to the hemoagglutination system test for rheumatoid arthritis, numerous other workers have confirmed the validity of this test in serum.i Others have found it positive in synovial fluid as well. It was deemed of interest to determine whether the so-called rheumatoid factor, which apparently determines the results of the agglutinaticn test for rheumatoid arthritis, was also present in other body fluids such as the cerebrospinal fluid and urine of patients with rheumatoid arthritis. Quantitative studies of total protein bound hexose content as well as ultracentrifuge characterization of the rheumatoid factor were also conducted. METHODS Three groups of adult male patients were studied: ( 1) Seven patients with the diamosis of ankylosing spondylitis; ( 2 ) ten patients with the diagnosis of rheumatoid arthritis, active, untreated; and (3)three patients with the diagnosis of osteoarthritis. The latex agglutination was positive only in the patients with rheumatoid arthritis. Serum and cerebrospinal fluid latex agglutinations were performed by the method of Singer and Plotzo and by the Hyland RA-Test." The urine, 250 ml., was dialyzed against running tap water for eight hours and then concentrated to 1.0 ml. with polyvinyl pyrrolidone (P.V.P.)for 24 hours at 5 C. The slide latex agglutination test was performed on 1:20 and 1:40 dilutions of the urine concentrates. All urines were tested for albumin prior to dialysis. Ten to 15 ml. of cerebrospinal fluid were dialyzed with P.V.P. to a volume of 0.1 ml. Total urine concentrate proteins were determined by the Biuret method.8 Total protein bound hexose levels were determined by the Thymol-H,SO, meihod.0 Urine electrophoretic studies were performed on a Spinco Model R electrophoretic apparatus using Verona1 buffer p H 8.6 0.05 mol. Ultracentrifuge studies were performed on a Model E Spinco ultracentrifuge. RESULTSAND DISCUSSION Table 1gives the results found in serum and cerebrospinal fluid. All cerebrospinal fluids were found to give negative agglutinations on both the latex agglutination and Hyland RA-Tests. Apparently, the rheumatoid factor is of such a high molecular weight that it does not pass the blood brain barrier. Karcher and associatesl0 suggested that in certain neurologic disease processes where there is an increase in gamma globulin, this particular globulin originates in the meningeal envelope, other globulins possibly arising from the serum. Ten specimens of urine of patients with rheumatoid arthritis were concentrated and three were found to give positive latex agglutinations (table 2). This finding indeed is interesting because, if we assume that the rheumatoid factor is a macroglobulin which apparently does not pass the choroid plexus, it is difficult to visualize how the same globulin could appear in the urine of individuals who did not have any detectable kidney disease. As shown in table 2, the three patients exhibiting the positive urine titers had relatively high serum titers. It is also of interest to note that the thirteen patients gave negative routine urine tests for albumin which were confirmed on electro"Hyland Laboratories, Los Angeles, Calif. 366 STARNES, ULLOA AND HOLLEY Table 1.-Latex Firation liter of Serum and Cerebrospinal Fluid Patients with Rheumatoid Arthritis Rheumatoid Arthritin Spondylitm Serum CSF L !-) c (-1 (-1 (-1 (-1 (-1 (-1 (--! L (-) (-1 K (-) E (-1 H (-) D (-1 Serum CSF L 1:2560 CI 1:5120 (-) (-) (-! M 1:2560 c H 1:1280 1:1280 G 1:1280 V 1:680 H 1:320 K 1:655,360 of Osteonrthritis Serum CSY - c (-1 (-) C" ( - ) (-1 L (-1 (-) (-) (-) (-) (-i (-1 (-1 F 1:20,480 ~ _ _ _( - ) _ _ - ~ Table 2.-Latex Firation Titer and Total Protein of Serum and Urine Concentrates of Patients with Rheumatoid Arthritis ~ M. K. F. Normal Serum Urine' 1:2560 1 :855,360 1:20,480 + + +- - T. P. (me. 56) 360 280 360 80 'Performed by Slide Test. phoretic studies of the same specimens. Patient M. had a total protein content of 360 mg. per cent; F., 360 mg. per cent; and K., 280 mg. per cent on the urine concentrates. As reported previously, the serum euglobulin fraction in patients with rheumatoid arthritis showed an increase in total protein bound hexose content." The present study, as shown in table 3, indicates that the total protein bound hexose content in the urine concentrates was also markedly increased. Comparatively, the percentage of total protein bound hexose in the urine total protein of these three patients was increased over the average of ten normal subjects. The ultracentrifuge studies of the urine protein concentrates revealed a globulin with a mobility of O.QS, but not the presence of the characteristic 19s globulin usually associated with the rheumatoid factor. This finding, however, may be related to the low concentration of the macroglobulins in urine or other agglutinating substances which would give a positive test. SUMMARY 1. Seven patients with ankylosing spondylitis, nine patients with rheumatoid arthritis, active, and three patients with osteoarthritis gave negative latex fixation test on the cerebrospinal fluid. 2. Of the thirteen patients examined with rheumatoid arthritis and high serum titers, three gave positive latex fixation tests on urine protein concentrates. 3. All urine concentrates were examined electrophoretically and found to be within normal limits. There was an increase in the total protein bound hexose content of the urine protein concentrates of the patients with rheumatoid arthritis. 367 LATEX FIXATION TEST IN RHEUMATOID ARTHRITIS ___ Table 3.-Carbohydrate and Protein Content of Urine Concentrates Name M. F. Total Protein mg. % T.P.R.H.* mg. 90' Percent T.P.U.H.' of T.P.t 360 360 280 28.8 70.8 44.4 8% 20% 16% 8.8 11% K. Normal ( Av. 10 samples) 80 'Total Protein bound hexose (T.P.B.H.). tTotal protein (T.P.). 4. Ultracentrifuge studies on the latex positive urine concentrates revealed a component with a mobility of 0.9s. REFERENCES 1. Ropes, M. W., Perlmann, G. E., Kaufman, D., and Bauer, W.: The electrophoretic distribution of proteins in plasma in rheumatoid arthritis. J. Clin. Invest. 33~311,1954. 2. Perlmann, G.E., Ropes, M. W., Kaufman, D., and Bauer, W.: The elertrophoretic patterns of proteins in synovial fluid and serum in rheumatoid arthritis. J. Clin. Invest. 33:319, 1954. 3. Starnes, W.K.,Hammack, W. J., Ulloa, A., Holley, H. L., and Hill, S. R.: Studies on serum and spinal fluid protein patterns in patients with rheumatoid arthritis spondylitis. Clin. Res. 6:1W, 1958. 4. Svartz, N., Carlson, A., Schlosamann, K., and Ehrenberg, A.: Isolation of the rheumatoid factor. Acta med. scandinav. 160:87, 1958. 5. Kranklin, E. C., Holman, H. R., MullerEberhard, H. I., and Kunkel, H. J.: An unusual protein component of high molecular weight in the serum of certain patients with rheumatoid arthritis. J. Exper. Med. 105425, 1937. 6. Singer, J. M., and Plotz, C. M.: The latex fixation test. I. Application to the serologic diagnosis of rheumatoid arthritis. Ani. 1. Med. 21:888, 1956. 7. Holley, H. L., Ulloa, A,, Henry, M., and Johnson, M. L.: A comparison of the latex fixation-whole serum, latex fixation-euglobulin fraction, and bentonite flocculation test in the laboratory diagnosis of rheumatoid arthritis. Am. J. hf. Sc. 237345, 1959. 8. Fister, H. J.: Manual of standardized procedures for spectrophotometric chemistry. New York, Standard Scientific Supply Corp., 1960, p. MA. 9. Shetlar, M. R., and Masters, Y. F.: Use of thymol-sulfuric acid reaction for determination of carbohydrates in biological materials. Ann. Chern. 29: 402, 1957. 10. Karcher, D.,and Lowenthal, A.: L'origine des proteines de liquide cephalorachidien. Clin. Chim. Acta. 3:78, 1958. 11. Starnes, W. R., Ulloa, .4., and Holley, H. L.: Glycoprotein studies on the rheumatoid factor. Am. J. Med. 27: 326, 1959. Willurd R. Starnes, M.S., C,hief Biochemist, Metabolic Laboratories, Veterans Administration Hospital, Birmingham, .4h. Alexander Ulloa, LM.D., lmtructor in Medicine, University of Alabama Medical Cater, Birmingham, Ah., and Trainee of the National Imtitute of Arthritis and Metabolic Diseases, National lnstitutes of Health, Bethesdu, M d . Howard L. Holley, M.D.,Professor of Medicine and Director of the Section of Arthritis and Rheumatism, University of Alubamu Medical Center and Veterans Administration Hospital, Birmingham, A h .