The influence of thymectomy on Rous sarcoma regression.

The incidence of regression of wing-web tumors induced by Rous sarcoma virus was shown to be dependent on the quantity of thymus tissue remaining after neonatal thymectomy in chickens of inbred line 6. Frequency of metastasis was associated negatively with the amount of thymus tissue present. Tumor regression and metastasis restriction both appeared dependent on the quantity of thymic tissue present.

Causes of mortality in chickens that regressed a Rous sarcoma virus-induced tumor.

Rous sarcomas were induced in 6-week-old chickens of several genetically different stocks: inbred lines C, 6(1), 6(3), and 7(2); crosses of inbred lines (6(3) X 7(2)) F4 and (6(1) X 15(1)) F2 X 6(1); and reciprocal crosses (15(1) X 100) F1 X 15(1) and (15(1) X 100) F1 X 100. The resulting tumors were scored for size six times during a 10-week period. Females that had completely regressed their sarcomas were placed in individual laying cages and examined weekly for reappearance of a tumor. After death, the probable cause was determined by necropsy. The major causes of death in the pooled sample of 49 females were fatty liver hemorrhagic syndrome (24.6%), reproductive disorder (14.2%), Marek's disease (12.2%), and lymphoid leukosis (6.1%). Elapsed time between tumor regression and death from any cause ranged from 21 days to 1930 days (5.3 years). One tumor recurred, this in a bird which eventually died with a massive sarcoma in the left wingweb and Rous metastasis in liver tissue. These data provide evidence of specific resistance to neoplastic disease.

Effect of regression of Rous sarcoma tumors upon egg production in an inbred line of White Leghorns.

Reserach was conducted to determine whether development and subsequent regression of a Rous sarcoma virus (RSV) induced wing-web tumor influenced egg production. Fifty-seven six-week old pullet chicks of inbred line 6 of the United States Department of Agriculture, Regional Poultry Research Laboratory, East Lansing, Michigan, were inoculated subcutaneously in the left wing-web with 0.1 ml. of a 10-minus 3 dilution of a pseudotype of Bryan high titer RSV designated BH-RSV (RAV-1). Thirty chicks were left uninoculated. Each chick was examined for tumor growth at regular intervals to 10 weeks post-inoculation. A tumor was considered regressed if it disappeared completely. Ninteen regressor and 22 uninoculated females were placed in laying cages at 4.5 months of age and egg production data obtained over seven 28-day periods. The difference in hen-day egg production between regressors and uninoculated controls favored regressors by 2.7 eggs/bird and was statistically significant. Physiological stimulus from cellular immunity, linkage and pleiotropy are discussed as possible caused of the higher egg production in regressors.

Metastasis of Rous sarcoma tumors in chickens is influenced by the major histocompatibility (B) complex and sex.

Six-week-old second generation progeny from the cross of inbred Lines 6(1) and 15(1), segregating into three major histocompatibility (B) complex groups (B2/B2, B2/B5, and B5/B5), were inoculated subcutaneously in the wingweb with one of three pseudotypes of Rous sarcoma virus. Chickens that died during a 10-week period after inoculation were necropsied and scrutinized for gross metastasis and histological sections of at least one lesion per affected organ examined for Rous sarcoma-transformed cells. By definition, a metastatic tumor was one located in an organ or tissue other than the primary inoculation site and having the histological appearance of a Rous sarcoma. Sarcomas developed in 1144 chickens, 390 of which died with tumor. For B2/B2, compared to B5/B5 hosts, mortality was 8 vs. 93%, median days to death were 45 vs. 31, and metastatic frequency was significantly lower, 32 vs. 58%. Disseminated lesions were significantly less frequent in females than males and grew preferentially in the heart and pericardial sac. Because the frequency of metastasis was significantly lower in B2/B2 than in B5/B5 chickens, a gene(s) within, or closely linked to, the B complex sharply retards the spread of Rous sarcoma virus-induced tumors.

Detection of cellular immunity to rous tumors of chickens by the leukocyte migration inhibition reaction.

Peripheral blood leukocytes obtained from Rous sarcoma-bearing chickens were tested for their ability to be inhibited from migrating in vitro by soluble tumor extract. It was found that the inhibition of leukocytes from chickens with regressing Rous tumors was significantly greater than that of chickens with progressing tumors or non-tumor bearing controls.

Genetic nature of regression of Rous sarcoma virus-induced tumors in crosses of Regional Poultry Research Laboratory lines 6 and 7(2).

Regional Poultry Research Laboratory (RPRL) inbred lines 6(1), 6(3), and 7(2) and F1, F2, and reciprocal backcross progenies of these lines were used to investigate host-gene effects upon the regression of Rous sarcoma virus (RSV)-induced tumors. Lines 6(1) and 6(3) were susceptible to subgroup A and C lymphoid leukosis (LL) viruses and line 7(2) was resistant to subgroup A but was segregating for susceptibility to subgroup C LL virus. Viruses of RSV subgroups A and C were used. Lines 6(1), 6(3), and 7(2) were homozygous for shared blood group alleles B2, C5, L1 and r. The incidence of tumor regression was higher in line 6(3) than in 7(2), and in reciprocal F1 crosses of these lines was of the same order of magnitude as in line 6(3). Progeny from F1 generation parents mated to line 6(3) had a higher frequency of regression than offspring from F1 generation parents mated to line 7(2). The frequency of regression in F2 generation offspring was intermediate between the two backcrosses. The data suggest that either a locus (or loci) other than L and B has a role in Rous tumor regression in this species or the immune response region of the B blood group-major histocompatibility complex differs in the two lines even though the serological and/or graft vs. host regions have not been shown to differ.

Differentiation of progressive versus regressive Rous virus-induced avian sarcomas according to tumor and infiltrating lymphocyte fine structure.

Electron microscope examination of progressing and regressing Rous virus-induced sarcomas in an inbred line of White Leghorns revealed that regressing tumors contained moderate to marked lymphocyte infiltration, frequent contact between lymphocytes and tumor cells, and extensive areas of necrosis. Lymphocytes infiltrating regressing tumors exhibited a polar accumulation of organelles at the point of contact between lymphocyte and target cell. On the other hand, progressing tumors contained low to moderate numbers of lymphocytes, infrequent lymphocyte-tumor cell interaction and less evidence of tumor cell degeneration. Lymphocytes from progressing tumors lacked the polar organization of organelles. This experimental system is offered as a means of studying the role of lymphocytes in tumor regression.

Lymphocytotoxicity of chickens bearing Rous sarcomas.

A 51Cr uptake microcytotoxicity assay was used to determine the cytotoxicity of lymphocytes from chickens bearing Rous sarcomas. Differences in lymphocyte cytotoxicity among individual chickens were statistically significant. Lymphocytes from chickens regressing (or having regressed) their tumors were significantly more cytotoxic than lymphocytes from hosts with progressing tumors. Lymphocytes from chickens with progressing tumors tended to enhance tumor cell growth in vitro.

A simplified technique for obtaining purified peripheral lymphocytes from chickens.

A simplified technique for isolating purified avian peripheral lymphocytes using a Ficoll-diatrizoate density is described. The procedure requires only 2 ml. of blood, can be completed in 5 min., and is suitable for the repeated testing of relatively large numbers of birds.

Major histocompatibility complex (B): effect on the response of chickens to a second challenge with rous sarcoma virus.

The influence of B genotype on the formation of a second RSV-induced tumor was studied. Line 6(3) (B2/B2) and (6(1) x 15(1))F3 and F4 B2/B2 and B5/B5 chickens were challenged with RSV in the left wingweb at 6 weeks of age. Tumor-bearing animals then were rechallenged with RSV in the right wingweb at either 10, 11, or 12 days post primary inoculation. Most B2/B2 hosts resisted development of a second tumor compared to none of the B5/B5 hosts. Failure of a second tumor to develop in B2/B2 hosts was not explained by humoral neutralization of RSV in vivo, since sera from 83% of B2/B2 and 100% of B5/B5 tumor bearing chickens failed to neutralize virus prior to 12 days post primary inoculation. Humoral antiviral immunity appeared later in B5/B5 than in B2/B2 hosts. Humoral antiviral immunity appears to play little if any role in resistance to early RSV-induced tumor development, but likely both humoral and cell-mediated antitumor immunity do.

Rous sarcoma regression in seven highly inbred lines of White Leghorns.

Response to Rous sarcoma virus (RSV)-induced tumors was studied in Regional Poultry Research Laboratory (RPRL) lines 61, 63, 72, 100, 151, and 15I5 and in Reaseheath line C, all highly inbred White Leghorn stocks. Virus inoculations were made in chickens at 6 weeks of age. Tumors were scored subjectively for size on a regular basis and in some instances a tumor profile index (TPI) was assigned which characterized tumor development over a 10 week period for each chicken (TPI 1 = complete regression in 28 days; TPI 5 = terminal tumor). The frequency of tumor regression, terminal tumors, and metastases and mean TPI was examined. The incidence of tumor regression ranged from 92% in line 61 to 0 % in lines 151 and 15I5. The frequency of terminal tumors varied from 100% in line 151 to 2% in line61, while metastasis in chickens with terminal tumors differed from 92% in line 15I5 to 0% in line 61. Mean TPI ranged from 2.0 in line 61 to 4.6 in line 15I5 and 4.7 in line C. The erythrocyte alloantigen genotype at the B blood group locus, (part of the B complex, MHC) and 11 additional blood group loci were known for each of the lines. The data indicate that genetic differences in tumor regression may be pronounced between inbred lines which share similar, if not identical, B locus erythrocyte alloantigens and that other unknown genes are also involved.

MHC and non-MHC genetic influences on Rous sarcoma metastasis in chickens.

The B5/B5 genotype, in Leghorns, was associated with a high degree of metastasis of Rous sarcoma virus-induced tumors, but in combination with a Leghorn-New Hampshire background markedly less metastasis occurred. Initially, four mating types were used: B5/B5 X B5/B5 chickens from the F5 generation of the cross of Leghorn lines 6(1) and 15(1), B24/B24 X B24/B24 chickens from line UNH 105 (New Hampshires), and reciprocal crosses of B5/B5 X B24/B24 chickens. Subsequently, F2 generation progeny of the cross of B5/B5 and B24/B24 breeders, as well as B24/B24 line UNH 105 and B5/B5 (6(1) X 15(1))F2 chickens, were used. Six-week-old chickens were inoculated in the wingweb with Rous sarcoma virus. Chickens dying during a 10-week period after inoculation were necropsied and suspect metastatic lesions examined histologically. Among 234 terminal chickens from the initial four mating types the incidence of metastasis associated with B5/B5 Leghorns (66%) was substantially higher than for B24/B24 New Hampshires (12%) and B5/B24 progeny of reciprocal Leghorn-New Hampshire crosses (19 and 24%). Subsequently, among 524 terminal hosts in the Leghorn-New Hampshire F2 population, B genotype significantly influenced tumor dissemination. However, among 52 concurrently challenged B5/B5 hosts from the (6(1) X 15(1))F2 population the incidence of metastasis (60%) was significantly higher than among 122 B5/B5 hosts from the Leghorn-New Hampshire F2 population (31%), indicating a non-major histocompatibility complex genetic effect on metastasis.