STUDIES ON CON COMITANT IMMUNITY

Transcription

1 STUDIES ON CON COMITANT IMMUNITY JOHN J. BITTNER (From the Roscoe B. Jackson Memorial Laboratory, Bar Harbor, Maine) The publication of numerous communications in the Reports from the Imperial Cancer Research Fund on the production of concomitant immunity by transplantable tumors furnished the impetus for the investigation to be reported here. As stated by Woglom (1929), little advance has been made in this field since Russell's paper was published in By concomitant immunity is meant the resistance of the host, as the result of the growth of a tumor, to reinoculation of the same or different tumors. The immunity produced by one tumor may be non-specific; that is, it may not prevent the progressive growth of grafts of all tumors. Russell (1912) was of the opinion that transplantable tumors might be divided into two groups, those which produce concomitant immunity and those which do not. He drew attention to the differences observed not only between different tumors but also between different sub-strains of the same tumor. To explain some of his results, Russell stated that individual host peculiarities were sufficient to produce or prevent the immune state. In one experiment comprising 12 animals, 4 were immune; the non-immune individuals had been selected from a stock produced by random matings, and to this Russell ascribed their lack of immunity. Mouse carcinoma 63 did not produce immunity in Russell's experiment, but did in the experiments conducted by Bullock and Rohdenburg (1920b) and by Foulds (1930). Bullock and Rohdenburg (1920a) found that the variations observed in the production of induced immunity disappeared when hosts from inbred stocks were used. In their work on concomitant immunity, these authors (1920b) observed that the variations could not be explained on the basis of strain, mortality, or technic, but must be due to differences in the tumor itself. They also concluded that there was no relationship between the fluctuations in induced, concomitant, and natural immunity. An extensive experiment on the possible immunizing effects produced by mouse sarcoma 180 was conducted by Wood and Prigosen (1925). These authors inoculated grafts of x-rayed tissue into the hosts and, after a period of ten to thirty days, reinoculated the animals with normal untreated tissue. The number of animals surviving the double inoculation was 1369, of which 88 per cent were susceptible to the normal grafts of tumor 180. Wood and Prigosen concluded that the variations observed between the control and experimental animals were insignificant and that the inoculation of x-rayed grafts of tumor 180 had no immunizing effects as shown by reinoculation with untreated tissue. Andervont (1932) found that, following successful inoculation of tumor 121

2 122 JOHN J. BITTNER 180 in the tail, amputation of the tail at the end of the second week, and reinoculation of tumor 180 subcutaneously, many mice failed to grow the tumor implants, due to the concomitant immunity produced by the tail growth. Of the controls, approximately 18 per cent showed regression of the tail tumors. Andervont, in agreement with many other workers, concluded that the factors determining variations or fluctuations in the immunity were: (1) the inherent property of the tumor; (2) variations in the growth energy of the tumor; (3) differences in the animals inoculated. Mice in which tail tumors had regressed were usually negative to reinoculation, and mice negative to the caudal inoculation of tumor 180 were susceptible to groin reinoculation. Andervont also determined that hosts immune to tumor 180 were resistant to carcinoma 63, but the reverse did not hold true. The present paper discusses experiments on the possible production of concomitant immunity by mouse sarcoma 180 when inbred stocks of mice are inoculated. The technic described by Andervont was followed.' MATERIALS AND METHODS The following stocks of inbred mice were used as hosts: Stocks Generations Inbred Spontaneous Cancer Incidence Mice Secured from * X or CllA N I Bar Co,Bl C,Rr Leaden Yellow D A Z or C,1I High-Cancer High-Cancer High-Cancer Dr. C. C. Little Dr.]. M. Murray Dr. ]. 1\1. Murray Dr. C. C. Little Dr. \Y. S. Murray -----~ _. - --_._ * The writer is indebted to the investigators listed for representatives of their stocks of mice. The ancestors of the other races were given the writer, at various times, by Dr. L. C. Strong. We were able to inoculate animals from six races of mice which had been inbred brother-to-sister for 25 generations. In addition to the pure stocks of mice, various hybrid generations were also employed. In describing the hybrid generations we have used the stock letters with the filial generation represented, i.e., AXF t represents the F, generation derived by crossing mice from the A and X stocks. The relative spontaneous cancer incidence in the strains is given for future comparative study. It is evident that by the use of the above mentioned closely inbred stocks we have eliminated, as far as possible, differences in the genetic and physiological constitution of the hosts compared. The tissue inoculated was the well known Crocker Fund mouse sarcoma The author wishes to express his appreciation to Dr. J. W. Schereschewsky and Dr. H. B. Andervont for the help and cooperation given during the course of the experiment.

3 STUDIES ON CONCOMITANT IMMUNITY 123 The animals were inoculated, according to Andervont's technic, in the tail, or caudally, with an emulsion of tumor 180. The tumors were permitted to grow for two weeks, when the tails were amputated, and grafts of tumor 180 were reinoculated subcutaneously in the right axilla. Two groups of control animals were used. One group was inoculated subcutaneously with tumor 180 near the right axillary region. The other group of control mice was grafted subcutaneously on the back near the base of the tail or posteriorly. TAlILE I: Number and of Individuals Susceptible to Tumor 180 in the Experimental and Control Groups (Only the experimental animals which had tail tumors are considered) Exp. Mice with Controls Inoculated Controls Inoculated Tail Tumors in Right Axilla Posteriorly Stock +Sub X A Leaden C7Rr Cb7Rl D Z N I Yellow AXF AXF AX-ARC AX-XBC ZIF () ZIF ZNF ZI-ZBC () DA-ABC DAF, Two carcinomas of the mammary gland were also tested. Tumor A748 developed spontaneously in an individual of the A stock and tumor Z749 in the Z stock. Homologous tissue or tumor 180 was used for the caudal inoculations in testing these tumors. No experimental animals were inoculated in the posterior position following the tail grafting. RESULTS In Table I we have tabulated the results secured from the inoculation of tumor 180 in the two control and experimental groups. In the experimental class only individuals which had a tail tumor two weeks after that inoculation are included. From the table it is evident that a comparative study can not be made between the experimental and control groups of some stocks. The lack of

4 124 JOHN J. BITTNER mice, or of space where the mice were available, accounts for the inadequate numbers in many cases. However, a sufficient number of animals from four inbred stocks were inoculated to enable us to compare the results secured from the inoculation of all the classes. In the X stock 5.7 per cent of the individuals which had tail tumors showed progressive growth of the subcutaneous grafts. In the control groups the TABI.E II: Ratios for the Control and Experimental Groups for Eacb Stock or Generation (R.A. = Right axillary controls. Post. = Controls grafted posteriorly) X Stock Exp, 4+:66-±.29 R.A. 24+: 7- ±1.55 Post. 31+: 64- ±3.05 A Stock Exp, 41 + : 55 - ±3.23 R.A. 35+: 3- ±1.11 Post : 55 - ±3.46 Co,Or Stock Exp. 32+: 2-±0.91 R.A. 41 +: 3- ±1.1l Post. 62+: 18- ±2.49 Leaden Stock Exp, 46+: 0 R.A. 21+: 0- Post. 15+: 2-±O.89 AXF, Generation Exp, 14+:17-±1.85 R.A. 0+: 0- Post. 43+: 18- ±2.37 AXF, Generation Exp, 42+: 98- ±3.61 R.A.l01+: 3-±1.14 Post :48-±3.80 AX-ABC Generation Exp. 34+: 39- ±2.84 R.A. 22+: 0- Post. 63+: 44- ±3.39 AX-XBC Generation Exp, 11+ : 30 - ± 1.89 H..A. 15+: 2- ±0.89 Post. 34+:23--±: %± %± %± %± %± %± %± %± %± % % 88.24%± %± %± %± %± %± %± % 58.88%± %± %± %± & 2: 7t.70%±5.34 or 13.43XP.E. 2 & 3: 44.78%±5.95 or 7.53 X P.E. 1 & 3: 26.92%±3.70 or 7.28XP.E. 1 & 2 : 49.40%±4.45 or X P.E. 2 & 3: 43.04%±4.34 or 9.92XP.E. 1 & 3: 6.36%±4.65 or 1.37XP.E. 1 & 2: 0.94%±3.67 or 0.26XP.E. 2 & 3 : 15.68%±4.00 or 3.92 X P.E. 1 & 3: 16.62%±4.ll or 4.04XP.E. 1 & 2: 0.00% 2 & 3 : 1t.76%±5.23 or 2.25 X P.E. 1 & 3 : 1t.76%±5.23 or 2.25 X P.E. 1 & 3: 25.33%±6.40 or 3.96XP.E. 1 & 2: 67.12%±2.60 or 23.97XP.E. 2 & 3: 29.33%±2.77 or IO.59XP.E. 1 & 3: 37.79%±3.63 or XP.E. 1 & 2: 53.42%±3.89 or 13.73XP.E. 2 & 3: 41.12%±3.17 or 12.97XP.E. 1 & 3 : 12.30%±5.02 or 2.45 X P.E. 1 & 2: 61.41%±6.97 or 8.81 XP.E. 2 & 3: 28.60%±6.80 or 4.21 XP.E. 1&3: 32.81%±6.33 or 5.18XP.E. results were: right axilla, 77.4 per cent; posterior, 32.6 per cent susceptible. The number of mice inoculated in the respective groups were 70, 31, and 95. The mathematical differences between the various groups of experiments for the X stock were: experimental animals and controls, right axilla, X P.E.; experimental animals and controls, posterior, 7.28 X P.E.; between the control groups, 7.53 X P.E. (Table II). Ninety-six A stock animals which had tail tumors were inoculated subcutaneously with tissue from tumor 180, of which 42.7 per cent were susceptible

5 STUDIES ON CONCOMITANT IMMUNITY 125 to the latter graft. The results in the control groups were: right axilla, 38 inoculated, 92.1 per cent +; posterior, 108 grafted, 49.1 per cent +. The difference recorded between the animals having tail and subcutaneous tumors was not mathematically significant when compared with the controls inoculated posteriorly in the A stock (1.37 X P.E.). The degree of significance between the experimental and the right axilla controls was X P.E., and the two control groups 9.92 X P.E. The 46 leaden stock mice which had tail tumors all showed progressive growth of the subcutaneous implants of tumor 180. The right axillary controls were likewise all susceptible, whereas the posterior controls, 17 in number, gave only 88.2 per cent +. No significant mathematical variation was recorded when the differences for the three groups were compared. TABLE III: Experimental Animals in the Various Stocks Which Were Negative /0 Tail Inocula/ion of Tumor 180 and s Negative and Positive /0 Subcutaneous Grafts (The last column gives the percentage of the total number used which failed to grow tail tumors) Exp, Mice: Negative to Tail Inoculation Stock of Number + Sub. - Sub. Inoculated X A Leaden C. 7Br C. 7Hl D Z N I Yellow AXF AXF AX-ABC AX-XBC ZIF, Total The results for the C,.,Br stock were: experimental class, 94.1 per cent; controls, right axilla, 93.2 per cent; controls inoculated posteriorly, 77.5 per cent susceptible. The numbers of individuals inoculated in the respective groups were 34, 44, and 80. The degrees of difference recorded between the experimental group and posterior controls, and the two control classes (4.04 and 3.92 X P.E., respectively) are probably significant. Since comparisons can not be made for the C.rBl, D, Z, N and I stocks, space will not be taken to repeat the observations recorded in Table I. In the AX-ABC generation, the back-cross generation to the A stock, no mathematical difference was observed between the experimental mice and the controls inoculated posteriorly. All other comparisons for the AXF 1, AXF~. AX-XBC and AX-ABC generations gave variations which are significant.

6 126 JOHN J. BITTNER The fluctuation in the percentage of " takes" is very noticeable when comparisons are made between successive experiments for the same stocks, especially in the posterior control and experimental classes. In Table III are listed the mice of the various stocks and generations which were negative to the caudal inoculation of tumor 180. The total number negative was 171, of which 14 or 8.2 per cent grew the subcutaneous transplants of tumor 180 progressively. Thirteen of the susceptible animals were members of the leaden, C,;Br, C,;Bl and D stocks, races which gave a high percentage of subcutaneous " takes" following successful tail inoculation of tumor 180. Approximately 20 per cent of all the experimental animals inoculated were negative caudally. TABLE IV: Results of the Subcutaneous Inoculation of Tumor Z7.JQ Following Tail Inoculation with Both Tumors Z7-/9 and 180 Experimental Controls Stock Tail Tumor + Sub Z ZIF IO Z lJ ZIF t lJ ZI-ZBC Z ZI-ZBC ZNF t Z ZNF t 180 II T,\lII.E V: Results of Tail Inoculation with Tumor 180 Followed by Subcutaneous Inoculation with Tumor A7.J8 Stock Tail Tumor Experimental Controls + Sub. + A DA-ABC The results secured from the inoculation of tumor Z749, a carcinoma of the mammary gland, are tabulated in Table IV. Tumor Z749 was inoculated caudally into the 90 mice of the Z stock, ZIFt, ZI-ZBC and ZNF t generations. All were susceptible to subcutaneous implants of tumor Z749. The number of control individuals inoculated with tumor Z749 was 215 of the various generations (Table IV). In all the transplants grew progressively. Twelve of the ZIFt hybrid animals were inoculated in the posterior dorsal subcutaneous position. Tumor A748, a carcinoma of the mammary gland from an A stock mouse, was also tested following the tail inoculation of tumor 180. The number of

7 STUDIES ON CONCOMITANT IMMUNITY 127 animals inoculated was 22 of the A stock and 29 representing the DA-ABC hybrid generation (Table V). No negative subcutaneous observations were recorded. The 94 mice from the A stock, used as controls, all gave progressive growth of tumor A748. SUMMARY Some inbred stocks of mice showed significant differences between results observed following subcutaneous transplantation of sarcoma 180 at different inoculation sites. In other strains the percentage of mice susceptible to subcutaneous grafts was significantly lower following successful tail inoculation, than in control groups. One control group of one inbred stock gave results which differed from the other control group but not from the experimental class of the same stock. Stocks which gave a high percentage of " takes" in both control groups gave similar results in the experimental class. Mice negative to caudal inoculations of tumor 180 were usually negative to subcutaneous reinoculation with the same tumor. No concomitant immunity was produced to tumors which arose spontaneously in individuals of pure stocks following the caudal inoculation of tumor 180 or homologous tissue. LITERATURE CITED ANDERVONT, H. B.: Public Health Reports 47: 1859, BULLOCK, F. D., AND ROHDENBURG, G. L.: J. Cancer Research 5: 119, 1920 (a). BULLOCK, F. D" AND ROHDENBURG, G. L.: J. Cancer Research 5: 129, 1920 (b). FOULDS, L.: Ninth Sci. Rept., Imperial Cancer Research Fund, London, 1930, P: 93. RUSSELL, B. R. G.: Fifth Sci. Rept., Imperial Cancer Research Fund, London, 1912, p. 1. WOGLOM, W. H.: Cancer Review 4: 129, WOOD, F. C., AND PRIGOSEN, R. E.: J. Cancer Research 9: 287, 1925.