Chicken Thymus Research Articles

Different myosins in myoid and entodermal reticular epithelial cells of the thymus

A sensitive immunofluorescence microscopical technique employing specific antibodies against highly purified actin and (smooth muscle type) myosin from chicken gizzard and myosin from human striated muscle was used to localize these contractile proteins in the thymus of rat, guinea-pig, cat and chicken. Myoid cells were seen to react with antibodies to striated, but not to smooth muscle type myosin, whereas reticular epithelial cells contained smooth, but not striated type myosin. Actin immunoreactivity was found in both myoid and reticular epithelial cells.

Analysis of Thymidine Kinase Activity and Glucocorticoid-Binding Capacity in the Thymuses of Healthy and Tumor-Bearing Chickens

Thymidine kinase activity in the lymphoid organs of healthy and tumor-bearing chickens was studied after hydrocortisone or dexamethasone treatment. Glucocorticoid hormones (10 mg/100 g body wt) administered twice in 48 hours resulted in an 80% decrease in thymidine kinase activity in the thymuses of healthy chickens, whereas birds with transplantable MC 29 chicken hepatoma failed to respond to the hormones. In the bursae of Fabricius of healthy or tumor-bearing chickens, thymidine kinase activity did not change considerably after administration of hydrocortisone or dexamethasone. The steroid-binding capacity in cellfree systems prepared from thymuses of tumor-bearing birds was significantly decreased. Thus the difference in steroid-binding capacity between thymuses of healthy and tumor-bearing chickens correlated well with, and could account for, the failure of hydrocortisone and dexamethasone to reduce the thymidine kinase level in tumor-bearing birds.

The Presence of Immunoregulatory Cells in Chicken Thymus: Function in B and T Cell Responses

The presence of immunoregulatory cells in chicken thymus was studied by using several different systems. Chickens injected with large numbers of syngeneic thymocytes were tested for their ability to produce antibody to heterologous red cells. Similar chickens were studied for their ability to reject allogeneic skin grafts. In separate studies, mixtures of thymocytes with spleen cells or with peripheral blood leukocytes were assayed for their ability to respond to PHA or to produce a graft-vs-host reaction in embryonic chicks. These studies indicated that immunoregulatory cells exist in chicken thymus, which displays both helper and suppressor activity. The suppressor cells were more prevalent or more easily detectable in young birds and in chickens with intact bursas. The helper function of thymocytes was seen to better advantage with cells derived from older animals and from bursectomized donors.

Cytophotometric Characterization of Cortical and Medullary Lymphocytes in the Chicken Bursa and Thymus

We have evaluated by means of cytophotometric techniques the nuclear content in DNA, in total nucleic acids and in histone proteins and the nuclear volume of the thymic and bursal lymphocytes in adult chickens. We have also made the same determinations in the developing bursal follicles before and after hatching. The results indicate that cortical differ from medullary lymphocytes for all the parameters considered both in thymic lobules and bursal follicles. Furthermore these differences appear analogous in both the organs, independently from the fact that they produce precursors of T and B lymphocytes respectively. As concerns the developing bursal follicle, the lymphocytes show the characteristics of the adult medullary lymphocytes. At the hatching changes occur in the nuclear content of total nucleic acids and histones. This is probably related to the exposure to antigenic stimulation through the cloaca.

Evaluation of the possible role of B cell receptors in the tendency of B cells to migrate into follicles in mice and chickens*

Chicken spleen and bursa cells were examined for the percentage of Fc receptor-bearing cells. Rosette formation was done with chicken 7S antibody-sensitized sheep erythrocytes and was inhibited by heat-aggregated chicken Ig. In the spleen, the percentage was found to increase with age to approximately 26% at 7 to 12 weeks. In contrast, only 3 to 5% of bursa cellss at this age demonstrated Fc receptors. Sleens from bursectomized chickens had 7--10% Fc receptor-bearing cells. In an attempt to determine a possible role of the C3 receptor on migration patterns, the effect of cobra venom factor (CVF) on the localization of transferred lymphoid cells was examined. Pretreatment of recipients with enough CVF to lower mean C3 levels to 11% of controls failed to affect follicular B cell localization in mice at either 24 or 48 h after transfer. Localization of thymus or bursa cells in chickens was similarly unaffected by CVF pretreatment. The possible roles of Fc and C3 receptors on migration of B lymphocytes into follicles and germinal centers were discussed.

Nicotinamide mononucleotide adenylyltransferase, a nonhistone chromatin protein: Immunochemical comparative studies

Rabbit antibodies to partially purified nicotinamide mononucleotide adenylyltransferase precipitated the enzyme, which remained fully active in the insoluble complexes. Precipitation of antigen-excess soluble complexes with sheep anti-rabbit γ globulin increased the sensitivity of the immunoassay. With this double-antibody assay, the enzymes from chicken erythrocytes, liver, kidney, and thymus showed nearly identical reactivity. Goose, pheasant, and turkey enzymes were highly cross-reactive with the chicken form; pigeon liver enzyme was markedly less reactive. There was no cross-reactivity with fish, amphibian, or mammalian enzymes. The specificity of the antiserum was increased by absorption of antibodies to nonenzyme proteins. The absorbed serum still precipitated the enzyme; in complement fixation assays, it reacted with an antigen that behaved like the enzyme. This antigen was detectable in whole chromatin and in the proteins extracted from chromatin by high salt or urea concentrations. Its immunological reactivity survived exposure to 0.5 m urea, but was reduced by exposure to 6.0 m urea plus 0.4 m guanidine. The enzyme was present as an inactive, partially denatured protein in nonhistone chromatin proteins prepared with these reagents.

Development of the embryonic chicken thymus: I. Characteristic synchronous morphogenesis of lymphocytes accompanied by the appearance of an embryonic thymus-specific antigen

Differentiation of the lymphoid cells of embryonic chicken thymus was studied morphologically and serologically from the tenth day of incubation to hatching. Lymphoid cells became markedly smaller in size at two stages, from the eleventh to the thirteenth and from the fifteenth to the sixteenth days of incubation. During the first stage, lymphoid cells changed notably in other criteria as well. The mean generation time was prolonged from approximately 12 to 36 hr on Day 13. The proportion of those cells that possessed embryonic thymus-specific antigen synchronously increased from 4–7 to 60–75% during the 24-hr period between the twelfth and the thirteenth days and reached a plateau level on Day 14. Morphologically, cytoplasmic processes became conspicuous in lymphoid cells on Days 11 and 12. These processes probably separated from the main body of the actively transforming lymphoid cells. In this phenomenon, a septum composed of microfilaments and small vesicles was formed at the neck of the process. Between the twelfth and the thirteenth days, the majority of lymphoid cells took the features typical of differentiated lymphocytes, i.e., small round shapes with scanty cytoplasm and distinct heterochromatin in their nuclei.

Development of the embryonic chicken thymus: II. Differentiation of the epithelial cells studied by electron microscopy

The differentiation of thymic epithelial cells in embryonic chickens was studied ultrastructurally from the tenth day of incubation to hatching. Reticuloepithelial cells first displayed functionally differentiated features between the twelfth and thirteenth days of incubation, whereas medullary epithelial cells with cysts and/or numerous granules did not display such features until the fifteenth day of incubation or later. Between the twelfth and thirteenth days, reticuloepithelial cells came to possess characteristic electron-lucent vacuoles and dense bodies. Furthermore, some of the vacuoles, probably as a result of fusion, seemed to contain the electron-dense bodies. The density of the material of such inclusion vacuoles tended to decrease after its entrance into the vacuoles. These findings suggested that the material in the inclusion vacuoles may be released into cytoplasm and, thereafter, may be secreted to exterior of the cells. Since a series of distinct morphological changes of reticuloepithelial cells took place simultaneously with an abrupt increase of lymphoctes bearing the embryonic thymus-specific antigen (T antigen), it is likely that the material supposedly released from the inclusion vacuoles of reticuloepithelial cells could be responsible for differentiation of lymphoid cells into embryonic T-antigen-bearing lymphocytes.

A Nonproducer T Lymphoblastoid Cell Line from Marek's Disease Transplantable Tumor (JMV)

A continuous lymphoblastoid cell line was established from a JMV tumor transplant related to Marek's disease (MD). It is designated RPL1 (JMV) lymphoblastoid cell line. This cell line contains DNA sequences complementary to MD virus DNA and has an antigen similar to MD-tumor-associated surface antigen (MATSA). However, it lacks any MD virus (MDV) rescuable in vivo or in vitro. The cell line has surface antigens typical of chicken thymus cells (T cells) and histocompatability antigens different from those of the host chicken.

The antigen-inexperienced thymic suppressor cells: a class of lymphocytes in the young chicken thymus that inhibits antibody production and cell-mediated immune responses.

Transfer of thymus cells from young chickens in combination with a light whole body irradiation (360 R) was found to suppress the rejection of skin grafts across strong histocompatibility (B) differences. On the average, the suppressed animals also showed decreased serum hemagglutinin titers against erythrocytes of the skin donor strain and a decreased graft-versus-host (GvH) reactivity against embryos of this strain. The thymic suppressor cells can be obtained from animals that have not experienced the antigen under test. However, after transfer and contact to the antigen (skin graft) they can lead to the formation of specific ("activated") suppressor cells and can mediate in the long run a specific inhibition of the response to this antigen. The suppressive activity is associated with a bursa-dependent cellular subpopulation in the thymus that is different from B lymphocytes, B precursor cells or GvH-reactive T cells. The bursa dependency of the thymic suppressor cell suggests that functionally different lineages of thymic and thymus-derived lymphocytes are derived from different sources of prethymic stem cells. The suppressor cells are predominantly found in the young chicken thymus and already detectable in the 16-day-old embryo, while poor suppressive activity is found in the adult thymus. The suppressive effect can be obtained with thymus cells from either syngeneic or allogeneic donors. Embryonic allogeneic donors provide suppressive cell preparations free of GvH reactivity. The possibility that the thymus suppressor cells mediate self tolerance and "neonatal tolerance" is discussed.

IGA surface determinants on cells from the chicken embryo.

Reports on chicken IgA were published almost simultaneously in 1972 by three different groups of investigators (1–3). Antisera to chicken I have been produced using serum, bile or oviduct washings as antigen source in rabbits (1–3) or pheasants (4). IgA is present in chicken serum and secretions in a variety of polymer molecules (5). The distribution of cells with I surface determinants has been studied in different organs of the chicken (6,7) Recently it has been suggested that I appears very early in phylogenesis (8). In continuation of work on the distrubtion of lyphoid cells bearing immunoglobulin surface determinants in hatched chickens (9), cells from the yolk sac, bursa of Fabricius, thymus and spleen of embryonal chickens have been tested in an indirect immunofluorescence test for surface determinants reacting with anti chicken IgA sera.KeywordsEvans BlueThymus CellEmbryonal ChickenEmbryonal LifeChicken SerumThese keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Analysis of time dependent changes of lymphopoietic activity in organ cultures of embryonic chicken thymus.

The time dependent development of lymphocytes in organ cultures of the thymus obtained from 10-day-old chick embryos was characterized by an initial phase of exponential increase of the number of lymphocytes per thymus followed by a plateau phase with no further increase in cell number. The proportion of cells in DNA synthesis dropped rapidly during the first 10 days of culture. Simultaneously the lymphocytes turned progressively smaller, as evidenced by both cell diameter and dry mass and constituted a homogeneous population of small cells at the end of the culture period. Thymic anlagen partially depleted of lymphoid precursor cells by a short hot pulse with 3H-TdR showed a prolonged exponential phase and reached normal plateau cell numbers 2-4 days later than usual. Furthermore, at least in the first part of the plateau phase, a reduction in the number of lymphoid cells per thymus resulted in a recovery in terms of the cell number which was associateed with increased DNA synthesis. These results are compatible with the regulation of thymic lymphopoiesis in organ culture through a mechanism operating via cell density.

Immunoglobulin-bearing lymphocytes of the chicken. I. Heavy chain immunoglobulin commitment and organ distribution.

When purified anti-immunoglobulin light chain antibodies were used in indirect immunofluorescence or labeled with 125I for autoradiographic staining, a similar percentage of Ig-bearing lymphocytes were detected by both techniques in lymphoid cell suspensions from the thymus or blood of 8-14-week-old chickens. However, a larger proportion of Ig positive lymphocytes were detected in suspensions of bursal cells by the more sensitive autoradiographic method, suggesting a lower surface density of Ig: perhaps on newly differentiated stem cells. In thymus and spleen suspensions, the proportions of Ig positive lymphocytes carrying mu and gamma-chains were roughly equal, whereas in the B cell populations of the bursa and blood, cells carrying surface gamma-chains predominated. IgA-bearing lymphocytes were only a minor population (< 5%) in lymphocyte suspensions prepared from the thymus, bursa, blood and spleen of adult chickens, but formed almost 50% of the Ig-bearing lymphocytes in the caecal tonsils.

Lymphocyte subpopulations in the thymus.

The thymus of mice and chickens contains at least four discrete populations of lymphoid cells: Two distinct cortical populations of small lymphocytes (early and late population), a hydrocortisone resistant and presumably medullary population of small lymphocytes, and a population of medium sized lymphocytes (prolymphocytes) (see Table I and Figure 3). These four cell types were identified with preparative cell separation techniques (e.g. cell electrophoresis, BSA-density gradient centrifugation, and velocity sedimentation) in combination with size distribution analysis. The combination of these techniques provides two-dimensional distribution patterns ('fingerprints') with high power of resolution. At present the two cortical populations of small lymphocytes cannot be identified as distinct populations by any other method. The physical parameters also provide useful markers for the identification and comparison of cellular subpopulations in animals of different ages, different strains, and to a certain degree even of different species. It is believed that each of these subpopulations is in itself heterogeneous in respect to antigen specificity, and it is proposed to call lymphocytes with different antigen specificity but identical physical characteristics 'isotypic lymphocytes'. The medium and large thymocytes serve as progenitors of the small thymic lymphocytes, as shown by different investigators. Small and larger lymphocytes are thus believed to represent different stages on developmental pathways (vertical heterogeneity). The different populations of small thymocytes, on the other hand, are believed to represent different independent pathways (horizontal heterogeneity). There is clearly the possibility that functionally distinct sublines of peripheral T-cells are generated by separate developmental pathways in the thymus, and the functional properties of single thymic cell types (e.g. of the thymic suppressor cells) may accordingly correspond to the functional properties of their peripheral progeny.

Immunosuppressive effect of syngeneic thymus cells on allograft rejection.

Transfer of thymus cells from young chickens to syngeneic recipients suppresses the allograft rejection between strains differing at the major histocompatibility (B) locus. Thymus cell transfer in combination with a light whole body irradiation (360 R) prolongs significantly the mean rejection time of skin allografts and leads in a proportion of recipients to long-lasting graft survival (greater than 200 days). Three weeks after the cell transfer, the suppression appears to be antigen specific, as judged by the normal reactivity against third-party skin grafts. From the types of thymus cells preparations that are effective in these experiments, it is inferred that the suppressor cell is a bursa-dependent lymphocyte, which is predominantly found in the young chicken thymus and which is different from B-lymphocytes, B-precursor cells, or graft-versus-host-reactive T-cells.

Migratory patterns of B lymphocytes: II. Fate of cells from central lymphoid organs in the chicken

Studies were made of the fate of 3H-nucleoside-labeled chicken bursa, thymus and bone marrow cells after intravenous injection into 8-day-old, irradiated chickens. A higher rate of proliferation of bursa cells was indicated by the fact that bursa cells incorporated more 3H-thymidine and 3H-adenosine than thymus cells, while 3H-uridine incorporation was similar for these two cell types. Homing patterns in recipient spleen tissue were similar for all three nucleoside-labeled bursa cells. Bursa cells localized in follicular and periellipsoidal areas, whereas thymus cells preferentially homed to periarteriolar sheaths. Bone marrow cells were found in all these locations. Bursa cells from chickens aged 4 weeks and older showed a greater tendency to home to the spleen than did bursa cells from 8-day-old donor chickens. Homing to other tissues appeared independent of donor age.

Shared antigenicity between chicken brain and bursa of fabricius.

Antisera to chicken brain, thymus and bursa of Fabricius were prepared in rabbits and rendered organ-specific by appropriate absorptions. Brain was shown by immunofluorescence and immune adherence to have common antigenicity with the bursa of Fabricius but not with the thymus.

Detection of T-cell surface antigens in a Marek's disease lymphoblastoid cell line.

Indirect membrane immunofluorescence was applied in the study of a lymphoblastoid cell line derived from Marek's disease lymphoma. The cell line tacked surface immunoglobulins and reacted specifically with antisera prepared against chicken thymus cells. Cells transformed by Marek's disease virus were of thymus origin.

Studies of the activity in chicken serum that stimulates the lymphoid development of the embryonic chicken thymus in organ culture.

Some properties of the thymus stimulating activity in chicken serum (TSAS) which is essential for the lymphoid development of the embryonic chicken thymus in organ culture, have been studied and a partial purification has been performed. TSAS resisted heating for 30 min at 60° C but not higher temperatures. During dialysis and ultrafiltration of serum a loss of TSAS down to a constant level of about 50 per cent of the original activity was obtained. TSAS was sensitive to treatment at pH 5 or lower but resisted alkaline pH up to at least pH 10. When serum was fractionated with ammonium sulphate little TSAS precipitated at 2.43 M ammonium sulphate or lower. At higher concentrations TSAS was found in the precipitates, but even at 4.05 M significant activity remained in the supernatant. TSAS was recovered from a fraction dominated by the serum albumins after gel filtration on Sephadex G‐200 and G‐100. Further fractionation by ion exchange chromatography on DEAE‐Sephadex separated the TSAS from serum albumin. These results suggest that part of the TSAS is associated with one or more specific factors with a molecular weight of approximately 70,000. The effect of the partly purified TSAS on the growth and morphologic development of the embryonic thymus in organ culture was studied.

Isolation of surface immunoglobulins from lymphocytes from chicken thymus and bursa.

ALTHOUGH the chicken is more closely related to dinosaurs and crocodiles than to the mouse1, the clear-cut demarcation between thymus-derived (T) and bursa-derived (B) lymphocytes in this bird2,3 offers an excellent system for study of the two lymphocyte types. B cells have been demonstrated to possess surface immunoglobulin3–5, while T cells have not, although it has been established that light chains at least are expressed on specifically activated chicken T cells6. So far, however, intact polypeptide chains have not been demonstrated on T and B cells. We now describe the isolation and partial characterisation of surface immunoglobulins of chicken B and T lymphocytes. We have evidence that both lymphocyte types have surface immunoglobulins consisting of light chains, μ chains and a heavy chain of molecular weight 40,000.