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Experimental Granulomatous Uveitis*
194
T. F. SCHLAEGEL, JR. AND FRED M. WILSON
the American Ophthalmological Society this year, and it will be published in the transactions of that group. However, in summary, our experiments, reported two years ago on boiled lenses, indicated that the oxygen uptake, as measured "in the Warburg ap paratus, was perhaps unreliable because it was not an index of respiratory activity of the tissue and, in the same paper, we reported that the estimation of glycolysis by the Warburg technique is a valid measurement because it is a heat-stable mechanism. It was our contention—perhaps not expressed in that paper, but expressed in the paper which was given at the American Ophthalmological Society —that the respiratory processes in the lens are
limited to the epithelium, and that the amount of respiration in the epithelium is so slight that it is overshadowed by the autoxidation of glutathione and ascorbic acid which is registered when you measure the oxygen uptake of the entire lens. In regard to the cyanide experiments, Dr. Potts, we have found that, even under the best circum stances in vitro, 20 minutes are required to estab lish equilibrium, and it is no wonder that tissue re sponse was not available. In our earlier experiments, when Dr. Robbie was still alive, we did numerous experiments in inject ing cyanide in the eye, and we were always dis appointed in not being able to get an effect.
E X P E R I M E N T A L GRANULOMATOUS U V E I T I S * STUDIES ON THE MECHANISM OF PRODUCTION OF THE CONTRALATERAL REACTION AFTER THE USE OF HORSE SERUM I N RABBITS T. F. SCHLAEGEL, JR., M.D.,
AND FRED M. WILSON,
M.D.
(With the technical assistance of Jack M. Tadman, M.S.) Indianapolis, Indiana
Several authors have made contributions to the study of experimental horse seruminduced granulomatous uveitis in rabbits. 1-12 Schlaegel showed that a granulomatous uveitis with a histopathologic picture re sembling that of sympathetic ophthalmia could be produced in both eyes of a rabbit by an injection of horse serum into one eye fol lowed in three weeks by horse serum intra^ venously.5 This procedure provides an un usual opportunity for antigen-antibody de terminations in the contralateral or "sym pathizing" eye, uncomplicated by previous manipulations. It is the purpose of this study to investigate some of the mechanisms which may be responsible for this form of contralateral granulomatous uveitis. The following major facts have been es tablished: 1. If the eye of a rabbit is injected with horse serum, a granulomatous uveitis will develop in the injected eye at about 14 days. 2. If the rabbit has been sensitized pre viously by intradermal injections of horse I
* From the Department of Ophthalmology, Indi ana University School of Medicine. This work was supported by a grant from the National Council to Combat Blindness, Inc.
serum, uveitis will develop within one day instead of 14 but the contralateral eye re mains uninvolved. 3. After the injection of horse serum into one eye of a normal rabbit, if horse serum is given intravenously one to three weeks later, uveitis will develop in the con tralateral untreated eye.4 4. The degree of contralateral uveitis is increased by use of the eye instead of the skin as a site of sensitization and also is increased by the use of large amounts of horse serum intravenously (for example, 20 ml.). 8 In the experiments being reported, two possible mechanisms were considered: (1) A direct toxic effect of horse serum on the uvea and (2) an allergic reaction involving either (a) local or (b) systemic antibodies. EXPERIMENT 1 POSSIBLE PRODUCTION OF UVEITIS BY DIRECT TOXIC ACTION OF HORSE SERUM
Method. Each of IS rabbits received 20 ml. of horse serum intravenously. The rab bits were killed at four through 21 days and their eyes enucleated. Eighteen scattered
EXPERIMENTAL GRANULOMATOUS UVEITIS
sections from each eye were cut, stained with hematoxylin and eosin, and graded as to the degree of cellular reaction. The grading of sections in this and other experiments being reported was made by one of us (T.F.S.) without knowledge of their source. These gradings were from 1 through 4, based on the degree of cellular infiltration. Results and Interpretation. Figure 1 dem onstrates that no uveitis appeared during the first week. Uveitis began to appear during the second week and, although minimal in de gree, reached a peak at 16 days. Since an antigen-antibody reaction usually begins some time after seven days and reaches a height at about 14 days, the development of uveitis within the first week would have in dicated that horse serum produces a uveitis by direct toxic action. Since the uveitis was delayed until the classic period for an anti gen-antibody reaction, the results support the antigen-antibody postulate. T H R E E EXPERIMENTS TO DIFFERENTIATE BETWEEN LOCAL AND SYSTEMIC ANTIBODY MECHANISMS EXPERIMENT 2
Method. Each of 25 rabbits received 0.1 ml. of horse serum in the right eye. Three
195
Fig. 2 (Schlaegel and Wilson). The degree φf histopathologic uveitis in the untreated left eyes at the days indicated. These rabbits had received an injection of horse serum in their right eyes followed by 20 ml. intravenously in three weeks. weeks later each rabbit was injected intra venously with 20 ml. of horse serum. Results and Interpretation. Figure 2 dem onstrates that uveitis was present within one day and reached a maximum development after five days following the intravenous in jection of horse serum. It is possible, but unlikely, that any of the horse serum in jected into the right eye would reach the left eye to stimulate the production of local anti bodies. Uveitis developed in the first week after intravenous horse serum, so the pos tulate of systemic antibodies is supported; if the uveitis had been produced by local an tibody reaction it should have been more delayed. EXPERIMENT 3
Fig. 1 (Schlaegel and Wilson). The degree of histopathologic uveitis at the days indicated after the intravenous injection of 20 ml. of horse serum.
In this experiment the titer of antigen and antibody was measured in both blood serum and aqueous humor. - Method. The right eyes of 30 rabbits were injected with 0.1 ml. of horse serum. At three weeks, 20 ml. of horse serum was given intravenously. Blood and aqueous samples were collected at intervals before and after this intravenous injection of horse serum. Six rabbits were used for each
T. F. SCHLAEGEL, JR. AND FRED M. WILSON
196
time interval and the blood and aqueous samples tested for antigen and antibody. Since secondary aqueous is richer in anti bodies, a rabbit was used only twice and then only after a rest of at least two weeks. Because fat in the blood may give false positive reactions, the rabbits were fasted before the samples were drawn. Determination of antihorse-serum antibodies A modification of the technique described by Hanks 13 was adopted. The horse serum was diluted with distilled water and then incorporated in the gelatin base to give the desired final concentrations. The gelatin base is composed as follows: Gelatin (Difco) NaCl Phenol Glycerol Distilled water
S.O gm. 1.0 gm. 0.5 gm. 20.0 ml. 80.0 ml.
After the ingredients are dissolved the pH is adjusted to 7.0 with N/10 NaOH. The gelatinized antigen is liquefied in a 37°C. water bath and pipetted into the bot tom of 6.0 by 50 mm. precipitin tubes, after which it is chilled in the refrigerator until solid. The aqueous humor is then layered over the antigen. The tests are incubated for 48 hours at 20°C. At the end of this period the tubes are examined for a precipitate at the interphase of antigen and aqueous humor. Besides the titrations on the aqueous hu mor, serum from the rabbits used in these experiments was also titrated for the pres ence of antihorse-serum antibody. The gelatinized antigen test also was used for these tests.
cipitin tubes. The diluted aqueous humor, or test serum, was then stratified over the antiserum and the tubes incubated at room temperature for four hours. The reactions were noted by holding the tubes before a source of obliquely transmitted light. Results (Table 1) and Interpretation. 1. After the injection of 0.1 ml. of horse serum into the right eyes, none was detected in the blood or aqueous of the contralateral eye at the days tested, presumably because of the small amount injected. 2. The injection into the right eyes pro duced an appreciable antibody titer in the blood; some of these antibodies entered the contralateral eyes after some lag. 3. Three days after the intravenous in jection, the antibody titer in the blood had fallen drastically and antibodies in the aque ous were no longer detectable. The intra venous antigen apparently removed the anti bodies by combining with them. Experiment 3 supports the postulate of systemic antibodies. EXPERIMENT 4
Method. Fifteen rabbits received 10 ml. of rabbit serum intravenously; this serum TABLE 1 EXPERIMENT 3
(The right eyes of 30 rabbits were injected with 0.1 ml. of horse, serum. An intravenous injection of 20 ml. was given at 21 days. At the days indicated the left aqueous and blood of six rabbits were tested for antigen and antibody. The mean of these values appears in this table.) No. of Days Blood Serum from Injec tion into Right Antigen Anti Eyes body
Determination of horse serum in aqueous humor The ring test was used to detect the pres ence of horse serum in the aqueous humor. Two-tenths ml. of pooled antihorse serum (rabbit serum, ring test titer—1:6,400) was placed in the bottoms of 6.0 by SO mm. pre
IV
7 10 14 17 91 24 27 35
0* 0 0 0
1/132 1/164 1/4800 1/4660
1/800 1/203 1/7.5
1/25 1/3466 1/9280
Aqueous Humor (Left eyes) Antigen
Anti body
0t 0
1/1.66 1/3.33 1/6.66 1/11.6
ot 0
1/4.5 0
ot
* Two of five rabbits had a 1:2 titer. t One of six rabbits had a 1:2 titer.
0 1/1.66 1/3.3
197
EXPERIMENTAL GRANULΦMATOUS UVEITIS TABLE 2 EXPERIMENT 5
(Fifteen rabbits of a skin group were sensitized to horse serum by an injection into the skin of the back. Another 15 rabbits, comprising an eye group, were sensitized by an injection into the right eye. At two weeks, left eye and skin sensitivity tests were per formed on both groups and the left eyes later Sec tioned and graded for degree of uveitis. There is no significant difference between the skin and eye groups.) r™ ^roup Skin Left Eye Fig. 3 (Schlaegel and Wilson). The degree of histopathologic uvetitis at the days indicated. These rabbits had received antihorse serum antibodies intravenously followed on the next day by an injec tion of the antigen (0.1 ml. of horse serum) into the vitreous.
was from rabbits made highly immune to horse serum. On the next day both eyes were injected with 0.1 ml. of horse serum. Results and Interpretation. Figure 3 dem onstrates that uveitis was detected at one day and reached a peak on the eighth day. Ac cording to the systemic antibody postulate, antibodies reach the contralateral eye first and react with subsequently appearing horse serum. In this experiment we simulated these conditions by injecting antibodies intravenously and by supplying the antigen by injection into the eyes on the next day. Since the uveitis which was produced was severe and in every way comparable to that previously produced, 1 ' 3 the postulate of systemic antibodies is supported. ELECTIVE SENSITIZATION EXPERIMENT 5
Method. One group of 15 rabbits was sen sitized to horse serum by an injection into their ritfht eyes and another group of IS rab bits by an injection into the skin of the back. Skin and left-eye sensitivity tests were done on both groups after two weeks. The
Degree of Clinical Reaction Left Eye 2.7 3.1
Skin 2.2 2.6
Degree of Histopathologic Uveitis in the Left Eyes 2.49 2.65
skin and left-eye reactions were graded on a 0 through 4 scale. Then the left eyes were enucleated, studied for the degree of uveitis, and graded on a 0 through 4 scale. Results and Interpretation. Table 2 dem onstrates no significant difference between the skin and eye groups after calculation of T. It is possible that an injection into the right eye of a rabbit could in some way espe cially predispose it to an ocular reaction on the contralateral side simply because the sen sitizing injection was made into an eye and not elsewhere. If elective sensitization had occurred, the eyes of the eye group and the skin of the skin group should have shown the greater reactions. There is a tendency for the eye group to give higher readings in all three categories. These findings are against elective sensitization and evidence only of the possibility that eye injections will produce a greater general sensitivity than will injections into the skin of the back. COMMENT
The body of evidence now accumulated strongly supports the role of systemic anti bodies in the production of uveitis from horse serum although it does not rule out the possibility that local antibodies also play a part. The evidence does not prove that the combination of antigen and antibody in it self stimulates the granulomatous response.
198
T. F. SCHLAEGEL, JR. AND FRED M. WILSON
Neither does it mean that histamine which might be liberated by the antigen-antibody reaction is the agent, since histamine has been. proved incapable of inciting cellular infiltration.1* What substance stimulates the granulomatous response remains unknown. SUMMARY
Five experiments were performed on 100 rabbits in an attempt to elucidate the mech-
anism of production of contralateral granulomatous uveitis from the use of horse serum in rabbits. These experiments, as well as past ones, support the postulate of systemic antibodies. Antibodies, formed as a result of injection of horse serum into rabbits, enter the eyes and react with sub sequently appearing horse serum to incite a granulomatous uveitis. 1100 West Michigan Street (7).
REFERENCES
1. Sattler, H. C, zur Nedden, Krusius, F. F., and Kόmmel, R.: Quoted by Foss. 2. Foss, B.: Experimental anaphylactic iridocyclitis. Acta Path. & Microbiol. Scand. Suppl., 81, 1949. 3. Schlaegel, T. F., Jr., and Davis, J. B.: The reaction of the rabbit eye to horse serum; sensitization by intradermal injection. Am. J. Ophth., 26:78S, 1943. 4. Schlaegel, T. F., Jr.: A comparison of the ocular reactions of pigmented and albino rabbits to normal horse serum; intraocular followed by intravenous injection. Am. J. Ophth., 27:137, 1944. 5. : Bilateral granulomatous uveitis from the use of horse serum in rabbits. Am. J. Ophth., 30:1225,1947. 6. Biegel, A. C.: Effect of cortisone in horse serum uveitis in rabbits. Arch. Ophth., 45 :2S8 (March) 195.1. 7. Bick, M. W., and Wood, R. M.: Heparin and uveitis; an experimental study. Am. J. Ophth., 33: 1878, 1950. 8. Burky, E. L.: Relation of ocular sensitivity to the Arthus phenomenon in the rabbit. Arch. Ophth., 10:368, 1933. 9. Seegal, D., and Seegal, B. C.: Local organ hypersensitiveness: III. Further observations on the experimental production in the rabbit eye. J. Exper. Med., 54:249, 1931. 10. Riehm, W.: Experimentelle Uebertragung eines lokalen Ueberempfindlichkeitszustandes von einem paarig angelegtem Organ das andere zugleich ein Beitrag zur Genese der sympathischen Ophthalmie. Deutsch med. Woch., 55 :907, 1929. 11. Guy, L.: Passage of horse serum from the blood stream into the aqueous humor of normal and immunized animals. Am. J. Ophth., 23:1209, 1940. 12. Collins, R. C.: Experimental studies on sympathetic ophthalmia. Am. J. Ophth., 32:1687, 1949. 13. Hanks, J. H.: A ring precipitin test for estimating the concentration of antibody in small amounts of immune serum. J. Immunol., 28:95,1935. 14. Schlaegel, T. F., Jr.: Histamine and uveal infiltration. Am. J. Ophth., 32:1331, 1949.
T. F. SCHLAEGEL, JR. AND FRED M. WILSON
the American Ophthalmological Society this year, and it will be published in the transactions of that group. However, in summary, our experiments, reported two years ago on boiled lenses, indicated that the oxygen uptake, as measured "in the Warburg ap paratus, was perhaps unreliable because it was not an index of respiratory activity of the tissue and, in the same paper, we reported that the estimation of glycolysis by the Warburg technique is a valid measurement because it is a heat-stable mechanism. It was our contention—perhaps not expressed in that paper, but expressed in the paper which was given at the American Ophthalmological Society —that the respiratory processes in the lens are
limited to the epithelium, and that the amount of respiration in the epithelium is so slight that it is overshadowed by the autoxidation of glutathione and ascorbic acid which is registered when you measure the oxygen uptake of the entire lens. In regard to the cyanide experiments, Dr. Potts, we have found that, even under the best circum stances in vitro, 20 minutes are required to estab lish equilibrium, and it is no wonder that tissue re sponse was not available. In our earlier experiments, when Dr. Robbie was still alive, we did numerous experiments in inject ing cyanide in the eye, and we were always dis appointed in not being able to get an effect.
E X P E R I M E N T A L GRANULOMATOUS U V E I T I S * STUDIES ON THE MECHANISM OF PRODUCTION OF THE CONTRALATERAL REACTION AFTER THE USE OF HORSE SERUM I N RABBITS T. F. SCHLAEGEL, JR., M.D.,
AND FRED M. WILSON,
M.D.
(With the technical assistance of Jack M. Tadman, M.S.) Indianapolis, Indiana
Several authors have made contributions to the study of experimental horse seruminduced granulomatous uveitis in rabbits. 1-12 Schlaegel showed that a granulomatous uveitis with a histopathologic picture re sembling that of sympathetic ophthalmia could be produced in both eyes of a rabbit by an injection of horse serum into one eye fol lowed in three weeks by horse serum intra^ venously.5 This procedure provides an un usual opportunity for antigen-antibody de terminations in the contralateral or "sym pathizing" eye, uncomplicated by previous manipulations. It is the purpose of this study to investigate some of the mechanisms which may be responsible for this form of contralateral granulomatous uveitis. The following major facts have been es tablished: 1. If the eye of a rabbit is injected with horse serum, a granulomatous uveitis will develop in the injected eye at about 14 days. 2. If the rabbit has been sensitized pre viously by intradermal injections of horse I
* From the Department of Ophthalmology, Indi ana University School of Medicine. This work was supported by a grant from the National Council to Combat Blindness, Inc.
serum, uveitis will develop within one day instead of 14 but the contralateral eye re mains uninvolved. 3. After the injection of horse serum into one eye of a normal rabbit, if horse serum is given intravenously one to three weeks later, uveitis will develop in the con tralateral untreated eye.4 4. The degree of contralateral uveitis is increased by use of the eye instead of the skin as a site of sensitization and also is increased by the use of large amounts of horse serum intravenously (for example, 20 ml.). 8 In the experiments being reported, two possible mechanisms were considered: (1) A direct toxic effect of horse serum on the uvea and (2) an allergic reaction involving either (a) local or (b) systemic antibodies. EXPERIMENT 1 POSSIBLE PRODUCTION OF UVEITIS BY DIRECT TOXIC ACTION OF HORSE SERUM
Method. Each of IS rabbits received 20 ml. of horse serum intravenously. The rab bits were killed at four through 21 days and their eyes enucleated. Eighteen scattered
EXPERIMENTAL GRANULOMATOUS UVEITIS
sections from each eye were cut, stained with hematoxylin and eosin, and graded as to the degree of cellular reaction. The grading of sections in this and other experiments being reported was made by one of us (T.F.S.) without knowledge of their source. These gradings were from 1 through 4, based on the degree of cellular infiltration. Results and Interpretation. Figure 1 dem onstrates that no uveitis appeared during the first week. Uveitis began to appear during the second week and, although minimal in de gree, reached a peak at 16 days. Since an antigen-antibody reaction usually begins some time after seven days and reaches a height at about 14 days, the development of uveitis within the first week would have in dicated that horse serum produces a uveitis by direct toxic action. Since the uveitis was delayed until the classic period for an anti gen-antibody reaction, the results support the antigen-antibody postulate. T H R E E EXPERIMENTS TO DIFFERENTIATE BETWEEN LOCAL AND SYSTEMIC ANTIBODY MECHANISMS EXPERIMENT 2
Method. Each of 25 rabbits received 0.1 ml. of horse serum in the right eye. Three
195
Fig. 2 (Schlaegel and Wilson). The degree φf histopathologic uveitis in the untreated left eyes at the days indicated. These rabbits had received an injection of horse serum in their right eyes followed by 20 ml. intravenously in three weeks. weeks later each rabbit was injected intra venously with 20 ml. of horse serum. Results and Interpretation. Figure 2 dem onstrates that uveitis was present within one day and reached a maximum development after five days following the intravenous in jection of horse serum. It is possible, but unlikely, that any of the horse serum in jected into the right eye would reach the left eye to stimulate the production of local anti bodies. Uveitis developed in the first week after intravenous horse serum, so the pos tulate of systemic antibodies is supported; if the uveitis had been produced by local an tibody reaction it should have been more delayed. EXPERIMENT 3
Fig. 1 (Schlaegel and Wilson). The degree of histopathologic uveitis at the days indicated after the intravenous injection of 20 ml. of horse serum.
In this experiment the titer of antigen and antibody was measured in both blood serum and aqueous humor. - Method. The right eyes of 30 rabbits were injected with 0.1 ml. of horse serum. At three weeks, 20 ml. of horse serum was given intravenously. Blood and aqueous samples were collected at intervals before and after this intravenous injection of horse serum. Six rabbits were used for each
T. F. SCHLAEGEL, JR. AND FRED M. WILSON
196
time interval and the blood and aqueous samples tested for antigen and antibody. Since secondary aqueous is richer in anti bodies, a rabbit was used only twice and then only after a rest of at least two weeks. Because fat in the blood may give false positive reactions, the rabbits were fasted before the samples were drawn. Determination of antihorse-serum antibodies A modification of the technique described by Hanks 13 was adopted. The horse serum was diluted with distilled water and then incorporated in the gelatin base to give the desired final concentrations. The gelatin base is composed as follows: Gelatin (Difco) NaCl Phenol Glycerol Distilled water
S.O gm. 1.0 gm. 0.5 gm. 20.0 ml. 80.0 ml.
After the ingredients are dissolved the pH is adjusted to 7.0 with N/10 NaOH. The gelatinized antigen is liquefied in a 37°C. water bath and pipetted into the bot tom of 6.0 by 50 mm. precipitin tubes, after which it is chilled in the refrigerator until solid. The aqueous humor is then layered over the antigen. The tests are incubated for 48 hours at 20°C. At the end of this period the tubes are examined for a precipitate at the interphase of antigen and aqueous humor. Besides the titrations on the aqueous hu mor, serum from the rabbits used in these experiments was also titrated for the pres ence of antihorse-serum antibody. The gelatinized antigen test also was used for these tests.
cipitin tubes. The diluted aqueous humor, or test serum, was then stratified over the antiserum and the tubes incubated at room temperature for four hours. The reactions were noted by holding the tubes before a source of obliquely transmitted light. Results (Table 1) and Interpretation. 1. After the injection of 0.1 ml. of horse serum into the right eyes, none was detected in the blood or aqueous of the contralateral eye at the days tested, presumably because of the small amount injected. 2. The injection into the right eyes pro duced an appreciable antibody titer in the blood; some of these antibodies entered the contralateral eyes after some lag. 3. Three days after the intravenous in jection, the antibody titer in the blood had fallen drastically and antibodies in the aque ous were no longer detectable. The intra venous antigen apparently removed the anti bodies by combining with them. Experiment 3 supports the postulate of systemic antibodies. EXPERIMENT 4
Method. Fifteen rabbits received 10 ml. of rabbit serum intravenously; this serum TABLE 1 EXPERIMENT 3
(The right eyes of 30 rabbits were injected with 0.1 ml. of horse, serum. An intravenous injection of 20 ml. was given at 21 days. At the days indicated the left aqueous and blood of six rabbits were tested for antigen and antibody. The mean of these values appears in this table.) No. of Days Blood Serum from Injec tion into Right Antigen Anti Eyes body
Determination of horse serum in aqueous humor The ring test was used to detect the pres ence of horse serum in the aqueous humor. Two-tenths ml. of pooled antihorse serum (rabbit serum, ring test titer—1:6,400) was placed in the bottoms of 6.0 by SO mm. pre
IV
7 10 14 17 91 24 27 35
0* 0 0 0
1/132 1/164 1/4800 1/4660
1/800 1/203 1/7.5
1/25 1/3466 1/9280
Aqueous Humor (Left eyes) Antigen
Anti body
0t 0
1/1.66 1/3.33 1/6.66 1/11.6
ot 0
1/4.5 0
ot
* Two of five rabbits had a 1:2 titer. t One of six rabbits had a 1:2 titer.
0 1/1.66 1/3.3
197
EXPERIMENTAL GRANULΦMATOUS UVEITIS TABLE 2 EXPERIMENT 5
(Fifteen rabbits of a skin group were sensitized to horse serum by an injection into the skin of the back. Another 15 rabbits, comprising an eye group, were sensitized by an injection into the right eye. At two weeks, left eye and skin sensitivity tests were per formed on both groups and the left eyes later Sec tioned and graded for degree of uveitis. There is no significant difference between the skin and eye groups.) r™ ^roup Skin Left Eye Fig. 3 (Schlaegel and Wilson). The degree of histopathologic uvetitis at the days indicated. These rabbits had received antihorse serum antibodies intravenously followed on the next day by an injec tion of the antigen (0.1 ml. of horse serum) into the vitreous.
was from rabbits made highly immune to horse serum. On the next day both eyes were injected with 0.1 ml. of horse serum. Results and Interpretation. Figure 3 dem onstrates that uveitis was detected at one day and reached a peak on the eighth day. Ac cording to the systemic antibody postulate, antibodies reach the contralateral eye first and react with subsequently appearing horse serum. In this experiment we simulated these conditions by injecting antibodies intravenously and by supplying the antigen by injection into the eyes on the next day. Since the uveitis which was produced was severe and in every way comparable to that previously produced, 1 ' 3 the postulate of systemic antibodies is supported. ELECTIVE SENSITIZATION EXPERIMENT 5
Method. One group of 15 rabbits was sen sitized to horse serum by an injection into their ritfht eyes and another group of IS rab bits by an injection into the skin of the back. Skin and left-eye sensitivity tests were done on both groups after two weeks. The
Degree of Clinical Reaction Left Eye 2.7 3.1
Skin 2.2 2.6
Degree of Histopathologic Uveitis in the Left Eyes 2.49 2.65
skin and left-eye reactions were graded on a 0 through 4 scale. Then the left eyes were enucleated, studied for the degree of uveitis, and graded on a 0 through 4 scale. Results and Interpretation. Table 2 dem onstrates no significant difference between the skin and eye groups after calculation of T. It is possible that an injection into the right eye of a rabbit could in some way espe cially predispose it to an ocular reaction on the contralateral side simply because the sen sitizing injection was made into an eye and not elsewhere. If elective sensitization had occurred, the eyes of the eye group and the skin of the skin group should have shown the greater reactions. There is a tendency for the eye group to give higher readings in all three categories. These findings are against elective sensitization and evidence only of the possibility that eye injections will produce a greater general sensitivity than will injections into the skin of the back. COMMENT
The body of evidence now accumulated strongly supports the role of systemic anti bodies in the production of uveitis from horse serum although it does not rule out the possibility that local antibodies also play a part. The evidence does not prove that the combination of antigen and antibody in it self stimulates the granulomatous response.
198
T. F. SCHLAEGEL, JR. AND FRED M. WILSON
Neither does it mean that histamine which might be liberated by the antigen-antibody reaction is the agent, since histamine has been. proved incapable of inciting cellular infiltration.1* What substance stimulates the granulomatous response remains unknown. SUMMARY
Five experiments were performed on 100 rabbits in an attempt to elucidate the mech-
anism of production of contralateral granulomatous uveitis from the use of horse serum in rabbits. These experiments, as well as past ones, support the postulate of systemic antibodies. Antibodies, formed as a result of injection of horse serum into rabbits, enter the eyes and react with sub sequently appearing horse serum to incite a granulomatous uveitis. 1100 West Michigan Street (7).
REFERENCES
1. Sattler, H. C, zur Nedden, Krusius, F. F., and Kόmmel, R.: Quoted by Foss. 2. Foss, B.: Experimental anaphylactic iridocyclitis. Acta Path. & Microbiol. Scand. Suppl., 81, 1949. 3. Schlaegel, T. F., Jr., and Davis, J. B.: The reaction of the rabbit eye to horse serum; sensitization by intradermal injection. Am. J. Ophth., 26:78S, 1943. 4. Schlaegel, T. F., Jr.: A comparison of the ocular reactions of pigmented and albino rabbits to normal horse serum; intraocular followed by intravenous injection. Am. J. Ophth., 27:137, 1944. 5. : Bilateral granulomatous uveitis from the use of horse serum in rabbits. Am. J. Ophth., 30:1225,1947. 6. Biegel, A. C.: Effect of cortisone in horse serum uveitis in rabbits. Arch. Ophth., 45 :2S8 (March) 195.1. 7. Bick, M. W., and Wood, R. M.: Heparin and uveitis; an experimental study. Am. J. Ophth., 33: 1878, 1950. 8. Burky, E. L.: Relation of ocular sensitivity to the Arthus phenomenon in the rabbit. Arch. Ophth., 10:368, 1933. 9. Seegal, D., and Seegal, B. C.: Local organ hypersensitiveness: III. Further observations on the experimental production in the rabbit eye. J. Exper. Med., 54:249, 1931. 10. Riehm, W.: Experimentelle Uebertragung eines lokalen Ueberempfindlichkeitszustandes von einem paarig angelegtem Organ das andere zugleich ein Beitrag zur Genese der sympathischen Ophthalmie. Deutsch med. Woch., 55 :907, 1929. 11. Guy, L.: Passage of horse serum from the blood stream into the aqueous humor of normal and immunized animals. Am. J. Ophth., 23:1209, 1940. 12. Collins, R. C.: Experimental studies on sympathetic ophthalmia. Am. J. Ophth., 32:1687, 1949. 13. Hanks, J. H.: A ring precipitin test for estimating the concentration of antibody in small amounts of immune serum. J. Immunol., 28:95,1935. 14. Schlaegel, T. F., Jr.: Histamine and uveal infiltration. Am. J. Ophth., 32:1331, 1949.