The Effect of Fever Artificially Induced on the Skin Sensitivity of Guinea Pigs to Turpentine

THE EFFECT OF FEVER ARTIFICIALLY INDUCED ON THE SKIN SENSITIVITY OF GUINEA PIGS TO TURPENTINE LEON GOLDMAN, M.D.' (Received for publication May 16, 1939)

Of course, the most important phase of the study of contact dermatitis at the present time is the determination of the background for the development of this form of dermatitis. This is much more significant than the collection of proved contact agents and adding those to a list which will always be limitless. Modern developments in the study of this background include the use of animals such as the guinea pig and the monkey (1—7),

as good test subjects for the production of contact dermatitis, the influence of diet with special reference to vitamin C requirements (8—12), the use of varied antigen mixtures (13) instead of the simple antigen alone, and the preparation of the skin site for

the application of the antigen. This report aims at the study in addition to the above of the influence of artificial fever. In the course of some work on guinea pig sensitizations to fractional distillates of turpentine it was decided to study the effect of fever on the induction of sensitization. To narrow the problem, whole turpentine mixture was used instead of the fractional distillates. Turpentine was chosen because this is an important and practical eczematogenic agent. The dermatological aspects of what is called turpentine have been reviewed thoroughly by Bonnevie (14) in his recent book. Burckhardt (9) has sensitized guinea pig skin to turpentine and produced an eczematoid derma1 From the Kettering Laboratory of Applied Physiology of the College of Medicine of the University of Cincinnati and the Department of Fever Therapy Research of the Cincinnati General Hospital. Read before the Second Annual Meeting of The Society for Investigative Dermatalogy, Inc., St. Louis, May 16, 1939.

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titis. In some of the animals the sensitivity lasted for several years. To eliminate the factor of primary irritation as much as possible, the hair of the animal was removed by close clippings (not shaving or sulphide epilation) a concentration of only 50 per cent turpentine in olive oil was used, and the animals were kept in separate cages to avoid the complications of lacerations from fighting. Males and females were used. Approximately the same set of color combinations was employed for each of the groups. The diet was kept constant for all animals, and included lettuce, carrots, hay, oats and water. With the 50 per cent turpentine in oil mixture, the control groups, of only 5 pigs, in the first part of the experiment developed sensitivity after the 4—5th

application. Following this, the skin became infiltrated, crusted and scaling. The appearance was similar to Mayer's animal with ursol hypersensitivity. Microscopic sections of the skin in the acute phase showed the usual picture of the

eczematous reaction. Section of the indurated skin showed marked acanthosis with pyknotic nuclei and considerable increase in melanin throughout the epidermis; there were scattered foci of lymphocytic infiltrate in the derma; the appendages were sparse. In this small control series the animals reacted also to d-pinen, l.pinen and also diluted fractional distillates of 145—150°, 150—154°, 154—

155°, 155—156°C. It was not possible to make out any differences in intensity of reaction with these various fractions. If full strength turpentine was used on the

skin, the animal became very excited, twisted and turned about, attempted to rub and scratch the skin. Some hours later, redness, infiltration or even necrosis would be evident. No microscopic studies of this irritant reaction was made. The turpentine used was steam distilled wood turpentine (American). One year later, another series of 5 guinea pigs were used as controls, and very similar results were obtained. Here, too, the same batch (older) of turpentine was used. One pig showed no reactions to repeated applications of the 50 per cent turpentine

mixture but the next application of concentrated turpentine produced a violent reaction. To study the effect of artificial fever, the guinea pigs were put in cages, directly

in the Kettering-Simpson-Sittler Hypertherm. For the first batch of 5 pigs studied, this air conditioned box had the temperature of the dry bulb at 120—140°F.

with a humidity of 30—45 per cent. For a recent group of 5 guinea pigs, the dry bulb temperature was held to 110—112°F. with a humidity of 80—90 per cent. The

initial rectal temperature of the guinea pigs in the series varied between 101— 106°F. Skin temperature readings were not done. Under the conditions of the experiment fever was produced in the guinea pigs by conduction and by the interference with heat loss by evaporation of moisture from the pulmonary system. With the higher box temperature and the lower humidity approximately 13—15 minutes was the maximum period the animals could remain in the hypertherm. The rectal temperature would be between 108—112°F., the skin felt hot. The animals were weak and listless and scarcely moved. When they did move, they crawled slowly and dragged their hind legs as if they were paralized. The respiratory and pulse rates were increased. This 'heat stroke" could be fatal with the temperature remaining elevated even with the death of the animal. With these animals, the gross visceral findings were essentially negative save for

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marked engorgement of the blood vessels. No microscopic sections were done.

One brain, mutilated in the removal, was examined by Dr. Charles D. Aring of the

Laboratory of Neuropathology who reported "disintegration of the cytoplasm of the nerve cells of the cerebral cortex." The nerve cells in the basal ganglia and brain stem showed no abvious abnormality. When the animals lived, the temperature returned to normal in 20--SO minutes. Several animals were found dead in their cages the next day even after the temperature had returned to normal. For this group of experiments it was decided to use the "maximum safe" temperature period, in other words, a fairly severe temperature reaction. At this time, to the previously clipped skin over the flank, the 50 per cent turpentine in olive oil was rubbed in gently with a cotton applicator. In the first group of only 5 animals no skin reactions were noted after repeated applications of the turpentine to the skin at the height of the fever reaction. Two of these animals died, however, after the 6th treatment. The three others received 13 treatments, however; unfortunately, no additional sensitization experiments were done with these surviving animals. With the newer regulating conditions of the hypertherm, namely the lower box temperature and the higher humidity, another series of 5 pigs was studied. With three of this group, higher fevers were used. With two of this group, lower temperatures of 105.6—108.2° were used. None of these animals have developed any skin reactions following the turpentine paintings from July through December, and in two of the pigs through March. There was also a control group treated by fever. These pigs were given fever with the turpentine group but had only olive oil rubbed on their skin at the height of the fever. No skin reactions were observed. Two of this group died also with the 6th fever

treatment. In 1935, three pigs were given fever and at the height of the fever, 10 per cent formaldehyde was rubbed on the skin. No reactions were noted after five treatment periods. No controls were done with this group. In a few animals, fever was induced by electro-magnetic induction with short wave. The pig was wrapped in a towel and fixed securely within the coil. Rectal temperatures were taken at various periods. This work has just started and only a few such treatments have been given.

With the small number of animals used it was not possible to

induce sensitivity to turpentine when 50 per cent steam dis-. tilled wood turpentine in olive oil was applied to the skin, at high levels of rectal temperature produced artificially. Because of the small number of animals and the short series of experiments it

would be hazardous to draw any definite conclusions. Fever has not been used in this connection before but there has been much work on the effect of artificial fever on animals, chiefly

rabbits. Warren (15) claims that in rabbits, dogs, guinea pigs, and calves the brain temperature is 0.5—0.7°C. above the rectal temperature. With dogs, it was found that the serum potassium roe from 30 to 50 per cent above the control level with the in-

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duction of fever. In three of the four animals the rise in potassium tended roughly to parallel the degree of fever. In three of the four dogs the sodium and chloride level remained unchanged. The sodium potassium ratio showed a fall with development of fever. In two dogs examined, the serum calcium fell slightly as

fever developed (16). Hartman (17) believes that anoxemia is the "underlying cause" of many visceral lesions described in patients and experimental animals after fever death. The chief visceral changes have been described in the brain, adrenals, and then in lungs and kidneys. Loseke and Gunderson (20) mention significant decrease in liver glycogen in rabbits following fever treatment. With regard to the use of short electric wave, Jorns (18) believes that with rabbits the short wave currents have no influence on the growth of hair or the healing of wounds. Krusen and Elkins (19) report no demonstrable changes in nerve tissue in laboratory animals whose temperatures were elevated 111.5 to 111.9 for from 33 to 75 minutes. Confusing reports have been given of the effect of fever on opsonins, agglutinins and the complement.

Engelhardt and Mahler (21) have mentioned that patients showing an elevation of temperature (method ?) and sweating were more sensitive to irritants applied in patch tests. With increased sweating without temperature elevation the effect was not noticeable. In relation to this, several patients who had had positive patch tests had these tests re-applied shortly before the

fever treatment. The treatment with the Hypertherm did not seem to change the intensity of the patch test reaction. Also, on several patients the histamin wheal was studied at various periods during the fever treatment. The erythema was, of course, difficult to evaluate because of the erythema from the fever box. The wheal measurements may have shown a slight decrease in area as compared to the wheal produced before the treatment. The differences, however, were not at all significant. On a few patients simultaneous intradermal histamin injections were done on the forearm covered with the electromagnetic coil from a short wave apparatus and the untreated forearm. There were no significant differences between the two forearms.

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At the suggestion of Dr. I. A. Mirsky and together with Dr. P. Wasserman preliminary experiments were done to determine the effect of the fever produced by the Hypertherm on anaphylaxis in

guinea pigs. With the first series there was rather definite inhibition of anaphylaxis. More details will be reported on this later. SUMMARY

In small groups of guinea pigs and with a small series of experiments, severe artificial fever reactions, as produced by the Hypertherm, prevented the development of skin hypersensitivity to turpentine. REFERENCES (1) MAysia, R. L.: Die Ursolidiosynkrasie des Meerschweinchens. Arch. 1. Dermat. u. Syph., 163: 223, 1931. (2) BLOCH, B., UND STEINER-WOUELJSCH: Die Sensibilisierung des Meerschwein-

chens gegen Primein. Arch. f. Dermat. u. Syph., 152: 283,

1926.

(3) JADASSOHN, WERNER: Sensibilisierung der Haut des Meerschweinchens auf

Phenyihydrazin. Kim. Wschr., 9: 551, 1930. (4) FREI, W.: Ueber wilikuerliche Sensibilisierung gegen chemisch definierte

Substanzen II Mitteilung. Untersuchungen mit Neosalvarsan am Tier. Kim. Wochenschr., 7:1026, 1928. (5) SULZBERGER, M. B.: Hypersensitiveness to arsphenamine in guinea pigs;

experiments in prevention and in desensitization. Arch. of Derm. & Syph., 20: 669 (Nov.) 1929.

(6) KILE, Roy L.: Sensitization of animals to plant oils. Arch. Derm. & Syph., 34: 90 (July) 1936.

(7) SiszoN, F. A.: Observations on poison ivy hypersensitiveness in guinea pigs. J. Immunol., 30: 275 (April) 1935. (8) C0RMIA, F. A.: Experimental arsphenamine dermatitis I. Canadian Med. Ass. Jour., 34: 272, 1936. (9) BURCKHARDT, W.: Experimental sensitization of guinea pigs to oil of turpen-

tine. Acta dermat-venereol, 19: 359 (Aug.) 1938. Abst. Arch. Derm. & Syph., 39: 718 (April) 1939. (10) SULZBERGER, M. B., AND OsEE, B. L.: Influence of ascorbic acid of diet on

sensitization of guinea pigs to neoarsphenamine. Proc. Soc. Exper. Biol. and Med., 32: 716 (Feb.) 1936.

(11) KuE, Roy L., AND PEPPLE A. W.: Further investigations of poison ivy hypersensitiveness in guinea pigs. Jour. Invest. Derm., 1: 59 (Feb.) 1938.

(12) Mu. Jui, Wu.: Experimenteller Beitrag zur Therapie allergischer Erkrankungen. Arch. f. Derm. u. Syph., 165: 27, 1932. (13) HAXTHAUSEN, H.: Experimentelle cutane Uberempflndlichkeit gegen Sublirnat. Arch. f. Dermat. u. Syph., 170: 378, 1934.

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(14) BONNEVIE, P0UL: Aetiologie und Pathogenese der Ekzemkrankheiten, Nyt, Nordisk Forlag. Arnold Busck, Kopenhagen, 1939. (15) WARREN, STAFFORD L.: First International Conference on Fever Therapy.

p. 45, Paul B. Hoefer, Inc., N. Y., 1937. (16) BISGARD, J. DEWEY, MCINTYRE, A. R., AND ASHEROFF, W.: Studies of so-

dium, potassium and chlorides of blood serum in experimental traumatic shock, shock of induced hyperpyrexia, high intestinal obstruction

and duodenal fistulas. Surgery, 4: 528, 1938. Abst. Am. J. Syph. Gonorr. & Yen. Dis., 23: 132 (Jan.) 1939. (17) HARTMAN, FRANK W.: Pathogenesis of lesions following artificial fever.

First International Conf. on Fever Therapy, p. 43, Paul B. Hoeber, Inc., N. Y. 1937. (18) JORNS: quoted by WILLIAM BIERMAN—The Medical Applications of the

Short Wave Current. P. 133, Wm. Wood & Co., Baltimore, 1938. (19) KRUSEN, FRANK H., AND ELKINS, EARL C.: Fever therapy by physical means. J. A. M. A., 112: 1689 (April 29) 1939. (20) LOSEKE, LUCILLE L., AND GUNDRRSON, MILLARD F.: Depletion of liver

glycogen under experimental fever induction in rabbits. 7th Annual Fever Conf., St. Louis, Nov. 11—12, 1938. (21) ENGELHARDT, W., AND MAHLER, W.: Is there a connection between the re-

sults of patch tests of Jadassohn and Bloch and fever and sweating? Dermat. Wchnschr., 99: 1205 (Sept. 15) 1934. Abst. Arch. Derm. & Syph., 32: 298 (Aug.) 1935.

DISCUSSION Da. I. ARTHUR MIRSKY, Cincinnati: Our purpose in continuing this study was

to attempt to find some definite mechanism in explanation of Dr. Goldman's observations. Together with Dr. P. Wasserman, we studied a series of guinea pigs who were sensitized to horse serum and subsequently upon being treated with a shock dose of this serum, developed a typical anaphylactic reaction. A similarly sensitized series were at the same time put into a fever box for 12 to 15 minutes, by which time their body temperature reached 109 to 1100. In only one instance out of 12 sensitized guinea pigs receiving the shock dose after fever treatment did death occur. However, in no instance did any animal develop the typical anaphylactic reaction, nor did any of the symptoms of anaphylaxis become obvious, and upon autopsy no animal revealed distended lungs. This experiment suggests that raising the body temperature can prevent the anaphylactic reaction. In the attempt to find whether histamine might play a role in this phenomenon, we studied the reaction of a series of guinea pigs to histamine, and found that no difference occurred so far as the mortality was concerned, whether the animals' temperatures were normal or high. It is of interest to note that in three sensitized guinea pigs, 24 hours after being placed in the fever box, at which time body temperature was normal, the administration of a shock dose of horse serum did not produce any of the signs or symptoms of the anaphylactic reaction. This suggests that fever therapy has an influence on either anti-

bodies or on the reaction between antibodies and antigen.

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DR. CLARENCE BERNSTEIN, Chicago: This work is very interesting. It is well

known that anaphylactic responses may be entirely prevented by many procedures, such as intravenous injection of vitamin C, or of any of several substances that produce a physiologic shock to the organism. Certain changes in the plasma protein, sedimentation rate, etc., occur. These studies have been made in the human after subjection to fever therapy. I do not know whether any chemical studies have been done in the work under discussion. Changes in this respect may have played a large role in doing away with the anaphylactic response. It is of immense value to know whether fever therapy has been tried in human cases of skin sensitization and what results have been found. Today reports are conflicting, and require better control observations. Further work of this nature would supply a scientific basis for fever therapy in this field, and therefore a sounder basis for its use. Da. J. GARDNER HOPKINS, New York: In generalized urticaria due to heat the

wheals will often not appear in the area heated, apparently because dilated blood

vessels carry away the H-substance so rapidly that any reaction is prevented. This might explain some failure of reactions in the heated animals, but I do not see how it can apply to the anaphylactic part of these experiments. For clarification, I wonder if Dr. Goldman would again tell us about the effect

of the hypertherm exposure on sensitization to turpentine. I understood from his abstract that there was no marked difference between the heated and the unheated animals, but I understand from his remarks just now that this was not the case, that there was a difference. DR. LEON GOLDMAN, Cincinnati: In the series of cases which will be reported tomorrow, we found that there was no benefit derived from fever therapy in atopic

dermatitis, and likewise in the cases of contact dermatitis. From this brief report it is rather interesting that artificial fever had a definite effect on two entirely different mechanisms in the guinea pig, an eczematous skin reaction and an anaphylactic phenomenon.

Apparently there was a misprint in the prepared abstract. The corrected abstract should read that there was a marked difference between the heated and unheated animals.

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