THE AMERICAN JOURNAL OF CANCER · 2013-05-15 · THE AMERICAN JOURNAL OF CANCER A Continuation of The Journal of Cancer Research VOLUME XXXI DECEMBER, 1937 NUMBER 4 PAPILLOMATOSIS

THE AMERICAN JOURNAL OF CANCER

A Continuation of The Journal of Cancer Research

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VOLUME XXXI DECEMBER, 1937 NUMBER 4

PAPILLOMATOSIS I N T H E FORESTOMACH OF T H E RAT AND ITS BEARING ON T H E WORK OF FIBIGER

W. CRAMER

( Imperial Cancer Research F u n d )

This paper deals with the conditions determining the onset of an extensive benign hyperplasia of the squamous epithelium lining the forestomach of the rat, associated with an extraordinarily massive hyperkeratosis. To the naked eye this lesion appears as numerous papillomata or warty excrescences occupying a large portion of the forestomach. The existence of a moderate degree of hyperplasia of the squamous epithelium without the appearance of warts or papillomata, or with but a few isolated growths of this kind, is not included in the considerations dealt with in this paper. The author's contribution to this subject consists in two series of experiments carried out at an interval of ten years under apparently identical conditions, which gave entirely discordant results.

Papillomatosis in the forestomach of the rat, which is lined, like the oesophagus, by stratified epithelium, has been repeatedly described in the literature as having been produced experimentally. This condition is of considerable theoretical interest in view of its bearing on the problem of carcinogenesis. Fibiger appears to be the first to have described it in his well known investigations on the experimental production of cancer of the forestomach in the 'rat by Spiroptera, which were published in detail in three papers in the Zeitschrift fur Krebsforschung in 1913, 1914, and 1920. He attributed the occurrence of this papillomatosis to infection with a parasite, Gongylonema neoplasticum; but in his later papers he also emphasized the fact that the animals were kept on a diet consisting only of white bread. I t must be remembered that at that time the significance of the vitamins in nutrition was not as clearly understood as it is now, and that in particular the pathological changes induced in different organs by a diet poor or deficient

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538 W. CRAMER

in this respect was not known. Fibiger distinguished clearly between the benign hyperplasia of the papillomatosis, which in many of his experimental animals was very extensive and occurred with considerable frequency, and a hyperplastic condition of a histologically different type which he believed to be malignant; but both these lesions he attributed to the presence of Gongylonema. His papers are accompanied by numerous instructive figures, which bear out this distinction. They do not, of course, prove conclusively that the conditions described by Fibiger as malignant were true carcinomata, a contention on which doubt was thrown by Bullock and Rohdenburg in 1918, and more recently by Passey. The only conclusive evidence in support of Fibiger's contention refers not to rats but to one mouse, from which he succeeded in transplanting one of his alleged carcinomata through four gcn- erations.

The distinction between the papillomatous benign hyperplasia and the condition described by Fibiger as " carcinoma of the forestomach" is particularly emphasized in his paper published in 1921. These considerations were apparently designed to meet the criticism made in 1918 by Bullock and Rohdenburg, who produced papillary and polypoid tu~nours in the rat's forestomach, resembling the benign hyperplasia described by Fibiger, by chemical or mechanical irritants such as celluloid balls covered with spikes of pig's bristles or the introduction of Scharlach R or pine tar. No reference is made to the diet used in these experiments, but in a later paper by Pappen- heimer and Larimore, Rohdenburg is quoted as having informed these authors that the animals received bread, turnips, carrots, and greens.

In 1924, Pappenheimer and Larimore published experiments indicating that some factor concerned with the diet could produce hyperplastic and ulcerative lesions in the forestomach of the rat. A rickets-producing diet, consisting of patent flour and an excess of calcium lactate with sodiunl chloride and ferric citrate, produced ulcerative and hyperplastic lesions in no less than 86 per cent of the animals within a period of twenty-seven to thirty- five days. Neither treatment with the quartz mercury lamp nor the addition of cod-liver oil prevented the occurrence of these lesions, although rickets did not develop. In these experiments, therefore, the stomach lesions appeared even though Vitamins A and D were present. Similar lesions were found in ani~nals on a synthetic diet deficient in Vitamin A. On this diet the lesions could be prevented by the addition of cod-liver oil, but they were also prevented when the cod-liver oil added to the diet had been inactivated by heat so that xerophthalmia, but no stomach lesions, appeared. In this series, therefore, the lesions in the forestomach were preventable, although the rats were suffering from a Vitamin A deficiency.

Some of the lesions observed by Pappenheimer and Larimore are figured, but these are not as extensive as the benign lesions figured by Fibiger. The authors found it difficult to interpret their results. They concluded that deficient diets favour the occurrence of the lesions, but the responsible factors could not be analysed in terms of dietary deficiencies known at that time. Since in synthetic diets, especially when they are deficient, the stomach frequently contains hair in considerable amount, they suggested that the lesions might be due to the ingestion of hair, which acts then as a mechanical

PAPILLOMATOSIS IN THE FORESTOMACH O F THE RAT 539

irritant, and they were able to produce similar lesions by mixing hair with an otherwise complete synthetic diet. They referred to the observations of Mitchell, Bradshaw and Carlson, who found hair balls in the stomachs of 90 per cent of rats kept on the synthetic standard casein paste diet of Osborne and Mendel, which was used at that time as a stock diet to ensure health and normal growth. These observers failed to find hair balls in rats kept on stock diets of mixed foods, on dry, crumbly diets, or on liquid diets. They mentioned, also, the existence of ulcerative conditions in the stomachs of rats in which hair balls were found, but without any detailed description.

In 1926, Fujimaki claimed to have produced carcinomata in the rat's forestomach by diets deficient in Vitamin A. His figures show, however, that he had produced only an extensive papillomatous hyperplasia similar to the benign hyperplasia described by Fibiger and the other workers mentioned above. No reference is made by Fujimaki to the observations published subsequently to Fibiger's earlier work. There is no clear evidence of any true malignant change, and in an editorial note added to the paper it is stated that an examination of the actual slides which had been sent by the author left in doubt the carcinomatous nature of the lesions described by him.

The statements of Fujimaki induced me, in 1927, to repeat his experiments. The results of these experiments will be given in further detail in the experimental part of this paper. Here i t is sufficient to state that at that time I had no difficulty in producing a very extensive papillomatous hyperplasia in a considerable proportion of rats kept on various diets, but that I failed to obtain any convincing evidence of a malignant change in these benign hyperplastic lesions.

In 1934, Passey published a preliminary account of experiments designed to repeat Fibiger's production of carcinoinatous changes in the rat's forestomach by infecting the animals with Gongylorzema neoplasticum. When the parasite was given to rats kept on a full, vitamin-rich diet, the forestomach showed no changes at all, not even benign hyperplasia. When the diet was changed to one of white bread, as used by Fibiger, or a synthetic diet deficient in Vitamin A, infection with Gongyloncma neoplasticum produced in the stomach areas of hyperplasia with marked keratinization but without any malignant changes supervening. But similar lesions were found when the rats were kept on these diets without being infected with the parasite. In drawing attention to this fact, that a benign hyperplasia was produced as a result of dietetic deficiencies alone, Passey stated that he was unable at the time to assess the relative importance of the parasite and of the dietetic deficiency in producing these changes; but that in his experiments the stomach lesions were more marked in those animals which had a superadded Gongylo- nema infection. As Passey's publications on this aspect of the question have been, so far, of a preliminary nature, unaccompanied by illustrations, I enquired by letter whether the benign hyperplasia observed by him was equal in degree to that figured by Fibiger. He kindly informed me that he had not been able to reproduce the gross papillomata shown in Fibiger's illustrations, and that the benign hyperplasia, although comparable to that figured by Fibiger, was not of the same degree.

540 W. CRAMER

EXPERIMENTAL OBSERVATIONS

My own experiments were begun in 1927, after the appearance of Fuji- maki's paper, to test his claim that cancer of the forestomach could be produced in rats by dietetic conditions alone. A large number of rats were kept on a diet deficient in Vitamin A. In another set of experiments, rats were kept on a diet deficient in Vitamin B ; but as it was anticipated that any effect which might be produced would require a considerable period of time, and as the rats die within about six to eight weeks on a diet completely deficient in

FIG. 1. PIZPIT.I.OMATOSIS 1X STOMACII OF RAT KEPT ON DIET DEFICIENT IN VITAMIN A FOR

ELEVEN WEEKS

Vitamin B, the experiment was arranged so that the animals were kept on the deficiency diet for four or five weeks; Vitalnin B was added during the following week, and the animals were then put back on a deficiency diet, this process being repeated several times. In other experiments which happened to be carried on during that period, and in which an abnormal diet was used, the stomach was also systematically examined.

The animals were killed when still apparently well, so that the stomach was, as a rule, distended with food. The oesophagus and the duodenum were tied off, and the whole stomach fixed in formalin-saline for twenty-four hours. I t was then cut open and examined. In this way one avoids the collapse of the sto~nach which occurs when it is opened before being fixed, and which leads to a folding of the stomach wall. In the rats kept on a synthetic diet deficient in Vitamin A, the forestomach showed papillomatosis, varying greatly in extent, however, in different animals. Sometimes a few papillomata only were present: in other animals the whole forestomach was filled with papillomata, and all degrees between these two extremes were observed. The phenomenon, however, did not occur in every animal, nor was it simply a question of the length of time during which the animals had been subjected to the deficient diet. Further, the effect was not specific for Vitamin A deficiency, but was found also in rats which had been kept for several months on a diet deficient in Vitamin B, as described above. I t was also found occasionally in animals kept on a synthetic diet containing an abnormal salt mixture. Lastly 8 rats, aged a year or more, from our laboratory stock were examined. They had never been kept on any vitamin-deficient diet, but had

PAPILLOMATOSIS IN THE FORESTOMACH OF THE RAT 54 1

received our ordinary laboratory diet of boiled maize, rice and bread, supplemented twice a week with cod-liver oil and Marmite, the latter a yeast extract rich in the Vitamin B, and B:! complex. In 3 of these 8 rats the forestomach showed numerous papillomata, and in one of the 3 this condition had developed to a very extreme degree. Findlay, who was at that time working in these laboratories carrying out experiments on rats kept on a synthetic diet deficient in Vitamin B:! only, also noted the occurrence of these hyperplastic lesions of the forestolnach in his animals. Altogether about 40 rats exhibiting an extensive hyperplasia in the epithelium of the forestomach clearly visible

FIGS. 2 AXD 3 . PIIOTOMICROC:RAPIIS O r LCSIONS IN S T O ~ ~ A T I I SHOWN I N FIG. 1 Above is a section through 2 papillomata ( X 8 ) . Below is a high-power view ( X 50) of a

section through the papilloma outlined in Fig. 2 .

to the naked eye'came under observation; but not one of them had developed an undoubtedly malignant tumour of the forestomach. The condition observed by us closely resembled the extensive benign papillomatosis described in the papers on Spiroptera cancer by Fibiger, who, as mentioned in the introduction, differentiated this condition clearly from a malignant condition also found by him, but much less frequently.

Histologically, there was in our material a hyperplasia of the squamous epithelium, varying greatly in degree with hyperkeratosis; but even when this hyperplasia was very massive, it never invaded the muscular wall of the stomach. These papillomata of the forestomach resembled in their histological appearance the large warts often obtained in the skin of mice by

542 W. CRAMER

painting with tar, which, while increasing fairly rapidly in width, maintain a smooth base and fail to show infiltrative growth for a long period, although a malignant change eventually supervenes in their immediate neighbourhood. An occasional occurrence of hair in the stomach of rats kept on this diet which had been noted by Pappenheimer and Larimore, and also by Carlson and his colleagues, was confirmed.

Since both the naked-eye appearance and the histological features of this condition have already been illustrated in great detail by Fibiger, and also

FIG. 4. PAPILLOM.\TOSIS I N STOMACII OF RAT KEPT O N DIET DEFICIENT lX V I T \ ~ I I ? ~ A FOR

EI.EVEN WEI~KS

by Bullock and Rohdenburg, and by Pappenheimer and Larimore, only a few figures are given to illustrate the results obtained by us. Figs. 1-3 show the lesions in the forestomach of a rat kept on a diet deficient in Vitamin A for eleven weeks. Fig. 1 shows an extensive papilloillatosis affecting a large portion of the forestomach. Fig. 2 , a low-power illicroscopic view, represents a section through two of these papillomata, while Fig. 3, a high-power view through the smaller of these two papillomata, illustrates the existence of a considerable degree of epithelial hyperplasia without any evidence of infiltrative growth into the muscu~ar wall of the stomach. Figs. 4-6 illustrate the

PAPILLOMATOSIS IN THE FORESTOMACH O F THE RAT 543

condition of the forestomach of two rats kept on an A-deficient diet for eleven weeks, Fig. 5 giving the histological appearance of the condition represented in Fig. 4 ; Fig. 7 that of a rat kept on a B-deficient diet for one month. Figs. 8 and 9 represent the stomach of a rat kept for thirteen months on a natural diet of bread, biscuits, maize and rice, to which cod-liver oil and Marmite had been added.

The existence of papillomatosis of the forestonlach can often be detected only if the stomach is opened. But when the condition is very pronounced,

it is sonletimes visible from the outside, as shown in Fig 10. The outside of this stomach of a rat kept on a B-deficient diet for four months in the manner described above, gives the impression to the naked eye of the presence of multiple carcinomata in the forestomach. Histological examination revealed, however, only an extensive hyperplasia of benign type without infiltrative growth into the muscular wall of the stomach.

Two other types of hyperplasia of the squamous epithelium of the rat's forestomach have been observed. One, represented in Fig 11, appears to the naked eye as a few isolated crater-like warts. They have been described in the literature as ulcerative lesions. Microscopically there is a marked hyperplasia of the squamous epithelium with hyperkeratosis. These lesions resemble most of those described by Pappenheimer and Larimore. In our

544 W. CRAMER

first series of experiments they were found not infrequently in rats kept on a variety of diets, and they occurred, also, though much less frequently, in our last series of experiments. The stomach shown in Fig. 11 is from a rat in our recent series kept on a diet of white flour, calciuln phosphate, and calcium carbonate.

A very striking but unusual example of epithelial hyperplasia was found by us once under conditions in which it was least expected to occur. The

FIG. 8. PAPII.I.OMATOSIS IN STOXIACII OF RAT KEPT FOR TIIIRTEFN RIOSTHS ON A N I i T l r ~ , ~ ~ DILT SUPPLEMENTED WITII COD-I.IVER OIL AND MAKMITE

material was obtained from a different laboratory, in which rats were kept on a stock diet of kitchen scraps and bread. From the point of view of growth and the general health of the animals, this diet was found to be a very satisfactory one. When these rats died or were killed the stomachs were kept at our request in forinol saline. In one of these animals, which died after being kept on this diet for thirteen months, the stonlach was filled with a mass consisting mainly of hair and of sawdust, used for the bedding of the cages. When this was removed the forestomach was found to contain a large egg-shaped tunlour (Fig. 1 2 ) which, when cut in half, proved to con-

PAPILLOMATOSIS IN THE FORESTOMACH OF THE RAT 545

sist mainly of a very large mushroom-shaped papilloma, attached to the wall of the forestomach by a stalk (Fig. 13). The histological appearance of this tumour, which was unique in our experience, is represented in Figs. 14-16. The low-power view given in Fig. 14 shows that the tumour consists of a large sessile papilloma made up of a mass of connective tissue lined by hyperplastic epithelium. A high-power view of a part of this papilloma is repro- duced in Fig. 15. Bound up with the papilloma by the hyperkeratotic covering

are two large cysts filled with keratin and a small wart. A high-power view of the wart is given in Fig. 16. Although isolated fields of either the large papilloma or the small wart might lead to the impression that malignancy has developed, the low-power view of the whole mass (Fig. 14) gives no evidence of true infiltrative malignant growth in any part.

For comparison, the warty growths produced in the mouse's skin by carcinogenic substances, which have been referred to above as growing to a considerable size while retaining their benign character, but which eventually became associated with the developnlent of malignancy in their immediate neighbourhood, are illustrated in Figs. 17-20. Fig. 17 is a large papilloma with hyperkeratosis and a smooth base giving no evidence of infiltrating malignant growth in the immediate neighbourhood. Fig. 18 shows two such papillomata: in the one to the left infiltrative growth is just beginning to develop a t the left-hand corner; the papilloma to the right is still without evidence of infiltration. Fig. 19 represents a large papilloma with massive

546 W. CRAMER

hyperkeratosis and a smooth, even base; infiltrative malignant growth, however, is beginning from a centre outside the papilloma, but in its immediate neighbourhood. Fig. 20 represents a large papilloma with infiltrative malignant growth arising from its base.

These 4 examples of papillon~ata of the mouse's skin have been selected from a large number of similar preparations in our possession as affording a contrast between the frequency with which malignancy develops in connection with large papillomata of the skin, and the rarity if not complete absence of

such a development in the much more extensive papillomatosis of the rat's forestomach.

The results of these experiments, therefore, confirm those of Rohdenburg and Bullock, and of Pappenhein~er and Larimore, in showing that an extensive benign hyperplasia comparable to the benign hyperplastic lesions first described by Fibiger can be produced without the intervention of Gongylonema neoplasticum. They also show that it is difficult to associate the occurrence of these lesions with any specific vitamin deficiency, although a synthetic diet deficient in Vitamin A appeared to be followed most frequently by their development. Another difficulty in explaining their occurrence lies in the fact


that their incidence in a group of rats kept on the same diet was very irregular. In some animals the papillomatous hyperplasia was very pronounced after a short interval of time; while in others kept on the same diet over a longer period it was much less marked, or even absent. Lastly the lesion was found also in rats kept on a natural diet supplemented with cod-liver oil and Mar- mite. Since the occurrence of these lesions could not be identified with the presence or absence of any known factor, dietetic or otherwise, only a brief account of these experiments was published in the 26th Annual Report of the

Imperial Cancer Research Fund, recording the failure to confirm the claim made by Fujimaki that it was possible to induce a malignant change in the forestomach of the rat by dietetic measures.

Three years ago these experiments were renewed with the object of determining whether a malignant change could be induced by superimposing upon the benign papillomatosis produced by dietetic means the action of a carcinogenic substance. I t was surprising to find that the same synthetic diets which previously had produced a papillomatosis, that is to say synthetic diets deficient in Vitamin A or in Vitamin B, now failed to induce this condition. Very occasionally a few, small, isolated, crater-like thickenings were obtained, but in the great majority of animals the epithelium of the forestomach remained normal, or showed only a very slight degree of hyperplasia not visible to the naked eye. The same negative results were obtained with the diet employed by Pappenheimer and Larimore, consisting of patent flour and a salt mixture rich in calcium lactate. With this diet Pappenheimer and Larimore obtained lesions in 86 per cent of their rats after a very short lapse of time, twenty- seven to thirty-five days. In our experiments with this diet these lesions did not appear.

Fibiger repeatedly states in his papers that his animals were kept on a diet composed exclusively of white bread and water. Since the papillomatous condition of the forestomach, as distinct from the malignant condition described by Fibiger, has been induced by several observers by dietetic measures

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only, the question arises whether the diet of white bread and water is capable of inducing papillomatosis in the rat's forestornach. White bread, i.e. bread made of white flour and yeast, is completely deficient in Vitamin A. I t is also very poor in Vitamin B, for white patent flour is practically free from this vitamin and the amount of yeast added to the flour and water in making the bread is so small (about 2 per cent) that the quantity of this vitamin intro- duced into the dough is too insignificant to be detectable by the ordinary biological assay.

Here a difficulty arises. In our experience, it is not possible to keep rats alive on a diet of white bread and water for longer than a period of two months. This is due partly to the vitamin deficiencies just mentioned and partly to other factors to which reference will be made presently. But many of Fibiger's rats lived for periods of six to nine months, and Passey and his co-workers refer in a recent paper to a rat kept on white bread and water for over a year (409 days). The only explanation which offers itself is that the bread used in these experiments was not simple " white bread," i.e. bread made of white flour, water, and yeast, but " milk bread," i.e. white bread made from white flour, to which a small amount of milk has been added, and yeast. Such milk bread contains among other things small amounts of Vitamin A and Vitamin B, and rats can be kept on it over long periods.

Whatever the explanation may be of the long survival in Fibiger's and in Passey's experiments, it is clear that the deficiency of white bread in Vitamins A and B cannot be the essential factor determining the incidence of papillomatosis in the rat's forestomach. We have already referred to the evidence from the experiments of Pappenheimer and Larimore, and in our second experimental series synthetic diets completely deficient in either Vitamin A or Vitamin B failed to produce this condition. I t seemed possible, therefore, that some other factor in the white bread diet was responsible: either a defect such as the low biological nature of the proteins or the presence of a substance acting as an irritant. In order to test for this possibility, a diet was prepared in which the deficiency of white bread in vitamins, especially in Vitamin B, was corrected without the addition of any other food- stuff. This can be done by adding to white flour wheat germ in the proportion of 1 part of germ flour to 3 parts of white flour. Wheat germ is rich in both Vitamin B and Vitamin E; it also contains a small amount of Vitamin A. Even on this diet it is not possible to maintain rats for more than a few months. The disproportion in the mineral elements Ca and P, which is an important defect of both white flour and germ flour, was corrected by the addition of a calcium salt such as calcium lactate, so as to bring the Ca/P quotient to the approximate value of 1. On such a diet rats can be kept alive over long periods, a year or longer. Many of the rats grow and live without any signs of ill health, even after a year; but some, when kept for such a long period on this diet, develop chronic septic infections, especially in the lungs, indicating that the Vitamin A content is not sufficiently high. Such a diet would show if the papillomatosis of the rat's stomach were due to the low biological value of the proteins of wheaten flour, or alternatively to the presence of some substance acting as a mild chronic irritant. Out of 28 rats kept on this diet for periods varying from nine to fifteen months, the forestomach

550 W. CRAMER

showed no lesion visible to the naked eye in 27 animals, and in the one exception the lesion was limited to a small number of warty excrescences.

On the Actiological Factors Concerned in the Papillomutosis

I t is clearly difficult a t present to give an explanation of the factors which determine the onset of the papillornatosis, for an adequate explanation must account not only for its occurrence but also for its non-appearance under apparently identical conditions. The following considerations are therefore necessarily of a purely provisional nature.

Provisionally, then, it seems advisable to consider the aetiology of the papillo~natosis apart from that of the ulcerative lesions, of which one exanlple has been given. I t is not difficult to admit that the ulcerative lesions may be due to mechanical irritants, such as the ingestion of hair or sawdust from the bedding, as suggested by the experiments of Bullock and Rohdenburg, and there is evidence that rats kept on synthetic diets, especially unbalanced synthetic diets, do ingest such materials. On the other hand, if that is the correct explanation, it is difficult to understand why in our recent series of experiments these ulcerative lesions were present so rarely, even when unbalanced synthetic diets were used.

As regards the papillomatosis, the outstanding fact is that it has not been possible in our recent series of experiments to produce this condition in the rat's forestomach by any of the dietetic measures which in our first series had apparently been followed by the development of this condition. As far as one can judge from the published figures of Fibiger, the papillomatosis observed by us ten years ago was as extensive as that found by him. The positive results obtained by us then show clearly that the occurrence of even a very extensive benign hyperplasia is not dependent upon the presence of Gongylonema ncoplasticum, as Fibiger believed.

On the other hand, failure to reproduce this condition in our recent experiments indicates that this benign hyperplasia is due to some factor other than diet, that has not yet been recognised. One possibility is that in some strains of rats the forestomach may be more susceptible to dietetic defects than in others. This possibility is suggested because the strain of rats used now in our laboratories-the Wistar strain-is different from the strain used when our earlier experiments were carried out. There is the further fact, pointed out many years ago (Cramer, 1922), that rats from different stocks vary greatly in their response to vitamin deficiencies, especially to a deficiency in Vitamin A. The apparently endemic occurrence of the papillomatosis suggests as another possibility a cage infection, possibly by a virus similar to the virus responsible for the infective papilloma of the skin of rabbits. In this case it would be conceivable that the dietetic measures play a contributory part either by allowing the survival of this agent in the forestomach, or by inducing slight defects in the epithelium, which by themselves are not sufficient to induce the formation of papillomatous masses. The correctness or incor- rectness of this explanation can be tested when it has become possible to produce again this massive papillomatosis.

Fig. 17. Simple large papilloma; no evidence of infiltrative growth. Fig. 18. T w o largc papillomata. infiltrative growth beginning at left edge of left papilloma. Fig. 19. Large papilloma with smooth base, infiltrative malignant growth beginning on its right side, outside the papilloma but in its immediate neighbourhood. Fig. 20. Large papilloma, infiltrative malignant growth developing a t its base.

552 W. CRAMER

On the Interpretation of Fibigcr's Observations

The results of our recent experimental series have an immediate bearing on the interpretation of Fibiger's observations. These have been subjected in recent years to very serious criticisms by Passey and his co-workers. Fibiger took the precaution of documenting his publications by numerous clear illustrations, which leave no doubt that he had produced in the forestomach of rats kept on white bread and water and infected with Gongylo- nema, two types of hyperplastic conditions: a papillomatosis, which he described as a benign hyperplasia, and which was sometinzes so extensive as to fill the forestomach, and secondly a hyperplasia of a different histological type showing a more infiltrative type of growth which Fibiger believed to be malignant. In further support of the interpretation of this second type of hyperplasia as a malignant one, Fibiger claimed to have found metastatic deposits in the lungs. The development of both types of hyperplasia he attributed to the presence of Gongylonema ~zeoplasticztm, enhanced, perhaps, by the concomitant action of the diet of bread and water. From what has been said in the preceding section, it is clear that the development of the benign papillonzatosis must be attributed not to the presence of the parasite, but to some factor which has not yet been identified. This factor probably requires for its action an unbalanced diet, but, as our second series of experiments shows, such diets alone are not in themselves sufficient to produce the papillomatosis.

Two questions, therefore, arise: ( I ) Is Gongylonema neoplasticum responsible for the development of

the lesion believed by Fibiger to be malignant? ( 2 ) Does this lesion represent a true carcinoma of the forestomach? In reply to the first question, it can be argued that Gongyloncma is not re-

sponsible for this type of hyperplasia, since Passey and his collaborators failed to obtain this lesion in rats kept on a diet of bread and water and infected with Gongylonema. But they failed, also, to obtain as extensive a degree of the benign hyperplasia as that figured by Fibiger; and it may be possible that Gongylone?na can elicit the type of hyperplasia believed by Fibiger to be malignant only in a forestomach in which a very extensive benign hyperplasia exists, although Fibiger himself did not hold this view. The answer to this question must, therefore, remain in abeyance until it has become possible to reproduce a t will the same degree of the benign hyperplasia observed by Fibiger and found also in our first series of experiments.

As to the second question, the malignant nature of this lesion is open to serious doubt. In so far as it is based upon metastatic spread in rats it must be discarded in view of the criticism by Passey, 1,eese and Knox, at least for the present; for it is now well known that in lungs of rats kept on a diet deficient in Vitamin A, such as the diet of white bread certainly is, a condition of septic bronchiectasis develops. The lesions which can be seen in such lungs closely resemble the figures of the alleged lung metastases published by Fibiger.

The only conclusive evidence in Fibiger's work that the infection with Gongylonenza on a diet of bread and water produces a carcinoizla in the

PAPILLOMATOSIS I N THE FORESTOMACH OF THE RAT 553

forestomach is now reduced to his experiments with mice, in which 3 out of 100 animals developed in the forestomach a histologically malignant type of hyperplasia, with peritoneal metastases in 1 instance. The peritoneal metastases in this last-mentioned animal could be transplanted successfully through four generations. These experiments on mice have apparently never been repeated.

At present we must conclude that diet and parasite are in themselves not sufficient to account for the lesions observed by Fibiger, and that there must be yet another factor involved which has not yet been identified. In this connection it is interesting to note that Fibiger himself considered the question whether it was Congylo~zema itself which was responsible for these lesions, or whether its effect might not be produced by virtue of acting as a host to a virus in the sense first suggested by Borrel.

While it has become necessary to readjust the interpretation of Fibiger's observations in the light of our present more complete knowledge, it would be a mistake not to accept his actual observations as valid facts. Our recent experience-that is to say, our inability to reproduce the extensive papillomatosis previously observed by us, although working under apparently identical conditions-is so vivid that but for the existence of photographic records taken ten years ago I myself would now doubt that this extensive papillomatosis had ever been obtained in our experiments. Fibiger's detailed docu- mentation of his observations can leave no doubt that he obtained a very extensive benign hyperplasia and another type which he, as an experienced morbid histologist, believed to be a malignant one. Passey and his co-workers failed to obtain this malignant type of hyperplasia, and they give good reasons for doubting whether it was truly malignant. But they also failed to obtain the same extensive degree of benign hyperplasia. The problem which faces us immediately is, therefore, not so much whether the questionably malignant type of hyperplasia was, or was not, truly malignant, but what are the factors determining the onset of the extensive benign papilloinatosis of the forestomach. When these factors have been identified and the papillomatosis can be produced experimentally at will, it may be possible to determine the true nature of the lesion which Fibiger claimed to be a carcinoma of the forestomach. The fact that considerable doubt exists concerning the true nature of this lesion is interesting from the point of view of the general relationship of the benign hyperplasia of squamous epithelium to the development of cancer, and requires a brief consideration.

On the Relationship between a Benign Hyperplasia and the Subsequent Development o j filalignancy

I t has been stated frequently, although by no means generally accepted, that there is no sharp dividing line between the aetiology of benign and of malignant tumours: the two are said to differ from each other only in degree. The phenomenon of papillomatosis of the rat's forestomach shows that an extensive benign hyperplasia of the squamous epithelium can exist to an extent equalling, and even greatly exceeding, that observed in the squamous epithelium of the mouse's skin treated with carcinogenic substances. But

554 W. CRAMER

there is this significant difference: that in the squalllous epithelium of the mouse's skin the benign hyperplasia is almost always followed by the development in its immediate neighbourhood of a malignant type of hyperplasia, while in the squamous epithelium of the rat's forestomach the subsequent development of a carcinoma has been completely absent in our experiments and in those of all other observers since Fibiger. This fact is the more striking as the histological picture of a papilloma in the rat's forestomach closely resembles that of certain types of papillomata found in the mouse's skin after the application of carcinogenic substances. I t is evident, therefore, that the existence of a benign hyperplasia in the squamous epithelium, however extensive, is not in itself sufficient to induce malignancy. From a study of experimental carcinogenesis in the mouse's skin, we had been led to conclude that the development of malignancy in the skin epithelium mani- festing a benign hyperplasia is a new and additional process, and not merely a continuation of the benign hyperplastic process (Cramer 1929). The existence in the rat's forestomach of an extensive benign papillomatosis without any evidence of a malignant change confirms this conclusion in a particularly striking manner. The association in the forestonlach of both a benign and a malignant type of hyperplasia has been observed so far only by Fibiger, and he, as stated in the Introduction, draws a sharp distinction between the two. He emphasizes the fact that the malignant hyperplasia can develop in a stomach in which the papillonlatous benign hyperplasia is co~npletely absent, and he concludes that the benign hyperplastic conditions of the stomach must not be regarded as a necessary preliminary step in the development of a carcinoma, or as a precancerous condition, even although they were in his experiments sometimes con~plicated by the development of a carcinoma.

A very extensive benign hyperplasia and hyperkeratosis of the squalllous epithelium of the rat's forestomach, consisting in the development of numerous papillolnata and warty excrescences, and occupying a large portion of the forestomach, were observed by Fibiger in rats kept on a diet of bread and water and infected with Gongylonema ncoplasticum. He also observed in these rats a histologically different type of hyperplasia which he believed to be a carcinoma of the squamous epithelium. Both these lesions he attributed to the presence of Congyloncma. Since a benign hyperplasia equal in extent to that found by Fibiger, but not associated with a carcinoma, has since been observed in rats not infected with Gongyloncma, this nematode cannot be held responsible for the benign hyperplasia. The factors determining the onset of this benign papillomatosis have not yet been identified. Unbalanced diets, especially diets deficient in Vitamin A, may play a contributory part, but they are not the chief determining factor, for, although this extensive papillomatosis was frequently found in rats kept on such diets ten years ago, it failed to appear in experiments carried out recently under apparently identical conditions. Possible explanations for this discrepancy are suggested.

The question whether another histologically different type of hyperplasia in the rat's forestomach which Fibiger believed to be a carcinoma of the


squanious epithelium was a true malignant hyperplasia, and if so, whether it was produced by Go~zgylonema neoplasticzrm cannot be decided until all the factors responsible for the development of the extensive benign papillomatosis have been identified, so that this latter lesion can be made to develop experimentally a t will. On the other hand, it is possible to conclude from the experiments of Passey and his collaborators that infection with Gongy- loncma, even when combined with an unbalanced diet, is not in itself sufficient to produce a carcinoma of the forestomach.

The general question of the relationship between the existence of a benign hyperplasia of the squamous epithelium and a subsequent development of malignancy is briefly discussed.

Lastly, the, a t present, erratic development in the rat's forestomach of an extensive papillomatosis which may simulate superficially the appearance of malignant new growths must be taken into account in experiments designed to test the carcinogenic action of substances, especially of substances present in food or drink, if grave errors are to be avoided. In this connection it is well to bear in mind that the lesions, when they occur, are restricted to the forestomach, which is lined with squamous epithelium and which has no counterpart in the human stomach.

BULLOCK, F. D., AND ROHDENBURG, G. L.: J. Cancer Research 3: 227, 1918. CRAMER, W.: 26th Annual Report of the Imperial Cancer Research Fund, 1927-28, p. 8. CRAMER, W.: Brit. J. Exper. Path. 10: 335, 1929; 3: 298, 1922. FIBIGER, J . : Ztschr. f . Krebsforsch. 13: 217, 1913; 14: 295, 1914; 17: 1, 1920. FISDLAY, G. hf . : J. Path. & Bact. 31: 353, 1928. FUJIMAKI, Y.: J. Cancer Research, 10: 469, 1926. PAPPEKHEIMER, A. M.. AND LARIAIORE, L. G.: J. Exp. Med. 40: 719, 1924. PASSEY, R. D.: Annual Report of the Yorkshire Council of Brit. Emp. Cancer Campaign,

1934, p. 9 ; 1935, p. 8. PASSEY, R. D., LEESE, A., AND KNOX, J. C.: J. Path. & Bact. 42: 425, 1936.

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THE AMERICAN JOURNAL OF CANCER · 2013-05-15 · THE AMERICAN JOURNAL OF CANCER A Continuation of The Journal of Cancer Research VOLUME XXXI DECEMBER, 1937 NUMBER 4 PAPILLOMATOSIS