Ronald W. Gillette

Changes in thymic estrogen receptor expression following orchidectomy

Abstract Estrogen receptor levels were measured in cytosols prepared from thymuses of age-matched normal male and female mice. Thymic estrogen receptor concentrations were significantly higher in females than in males. Castration restored the thymic estrogen receptor expression in males to levels nearly equivalent to that of females. The results suggest that androgenic hormones may suppress estrogen receptor expression in thymus and thus account at least in part for the well documented inferior immune response of males. The data further suggest a role for estrogenic hormones in thymic function and thymocyte maturation.

The effect of T lymphocyte deficiency on tumor induction and growth.

Abstract Severe reduction in the numbers of T lymphocytes as occurs in thymectomized, X-irradiated, bone marrow reconstituted mice or nude mice did not increase the animals susceptibility to chemical carcinogenesis. However, once established, many tumors grew distinctly slower in the T cell deficient animals. This was demonstrated to be due to lack of normal T cell function since tumors arising in TXB hosts progressed much faster in normal secondary hosts than in secondary TXB hosts. In most instances, established tumor cell lines from tissue culture also grew more slowly in T deficient hosts. The suggestion was made that T cells may not be required for immune surveillance while the immune mechanism(s) that promote subsequent tumor growth are T cell dependent. Alternative possibilities are discussed.

Homing of labeled lymphoid cells in athymic mice: Evidence for additional immunologic defects

51 Cr-labeled lymph node, spleen, thymus, and bone marrow cells from BALB/c-nu hybrid donors distributed in reduced percentages to the spleens, bone marrow, and small intestines of athymic nu/nu mice. Comparable cells from the lymphoid organs of athymic nude donors homed to the lymphoid organs of normal hybrid or nude recipients in similar percentages. The distribution of labeled lymphoid cells from normal donors in nude or hybrid recipients was related to cell viability since heat-killed hybrid cells were recovered in similar percentages in nude athymic or normal mice. Since labeled lymphoid cells migrated normally in thymectomized, irradiated mice reconstituted with syngeneic bone marrow, it was concluded that lack of a functioning thymus per se did not explain the results. The data suggest that additional deficiencies may exist in some lymphoid organs of the nude mouse that prevent normal migration of T lymphocytes.

Regulation by adherent cells of macromolecular synthesis in lymphocyte populations.

Abstract The role of adherent cells in the regulation of lymphocyte DNA, RNA, and protein synthesis was investigated. Splenic adherent cells suppressed DNA synthesis of spleen cells. The reverse was true of lymph node adherent cells, i.e., DNA synthesis was less in the absence of adherent cells. Histocompatibility was not required for interaction between adherent and nonadherent cells. The regulatory adherent cell in either case was not θ positive. Removal of splenic adherent cells decreased RNA and protein synthesis of nonadherent spleen cells. RNA synthesis by lymph node cells increased when adherent cells were removed, but protein synthesis was lower. Autoradiographic analysis revealed that removal of adherent spleen cells allowed a greater number of nonadherent cells to respond to the presence of mitogen. Adherent cells were observed to regulate DNA synthesis of B lymphocytes also. Lymph node adherent cells amplified DNA synthesis in both nonadherent spleen and lymph node cells, while splenic adherent cells suppressed splenic nonadherent cells but stimulated nonadherent lymph node cells. We feel the data are compatible with the idea that at least two functionally distinct adherent cell populations exist. A mechanism is suggested to explain the data.

Changes with age in the homing properties and mitogen responses of lymphocytes from normal and leukemia-prone mice.

Abstract Changes in the mitogen response and homing properties of lymphoid cells from leukemia prone AKR/J mice were compared to those of normal BALB/c mice as a function of age. During the first 35 weeks of life, BALB/c lymph node and spleen cells were significantly more responsive to phytohemagglutinin (PHA) than were AKR lymphocytes. BALB/c thymocytes exhibited a consistent low-level response to PHA that was totally missing in AKR thymus cells. Data provided by the homing studies demonstrated that the content of spleen-seeking cells rose with age in BALB/c spleens whereas the percentage recovery of spleen-seeking cells from AKR spleens decreased. The pool of lymph node-seeking cells was larger in AKR than in BALB/c prior to the development of leukemia in AKR. Radiolabeled bone marrow cells that migrate to the spleen decreased numerically with time in BALB/c animals while the opposite was observed with AKR, i.e., there was a continuous increment in the levels of spleen-seeking lymphoid cells in AKR bone marrow. The data show that the relative representation of specific subpopulations of lymphocytes is modified in AKR mice and suggest that such differences may be important in the development of AKR leukemia.

Studies of the T-lymphocytes resident in the spleens of thymectomized, irradiated, bone marrow-reconstituted (TXB) mice☆

Abstract The T-lymphocytes resident in the spleens of thymectomized, lethally irradiated mice that had been reconstituted with syngeneic bone marrow (TXB) were characterized. Both recently reconstituted N-TXB, (approximately 3 weeks after bone marrow injection) and aged (>6 months after reconstitution) A-TXB animals were studied. The T-lymphocytes from spleens of recently reconstituted N-TXB donors did not respond to PHA but did react significantly to Concanavalin A (Con A). The lack of PHA sensitivity was not due to dilution of reactive cells by other cell types. Removal of adherent cells, likewise, did not restore N-TXB spleen cell PHA responsiveness. N-TXB splenic T-cells were cortisone resistant. N-TXB spleen cells by themselves did not cause a graft vs host response. However, N-TXB spleen cells amplified the graft vs host response of normal lymph node cells but not N-TXB lymph node cells. Addition of cyclic GMP enhanced [ 3 H]thymidine uptake of N-TXB spleen cells caused by Con A. N-TXB spleen cells were exclusively spleen seeking. The Con A reactive cell within N-TXB spleens was demonstrated to be of donor origin. Fetal liver as well as syngeneic bone marrow contained cells capable of reconstituting the Con A response. Spleen cells from aged. (>6 months) A-TXB were found to be PHA sensitive. Competitive inhibition assays measuring θ expression in A-TXB spleen cells indicate a significant increase in the θ positive lymphocyte population occurred with time. The data indicate that considerable reconstitution of θ positive cells had occurred in A-TXB donors. The results also suggest that the T-lymphocyte population of the TXB spleen may be a unique subpopulation of T-lymphocytes that resides exclusively in spleen and bone marrow.

A Study of the Functional Capacities of Spleen Cells After Maintenance in vitro

Summary Spleen cells were cultured for varying lengths of time in vitro. The cells were then injected into lethally irradiated mice to ascertain their ability to protect the animals against irradiation death. Complete loss of ability to protect irradiated animals was found to occur after 9 days in vitro despite the injection of adequate numbers of viable spleen cells. The significance of these results is discussed.

Further evidence for the existence of B-lymphocyte subpopulations☆

Abstract We have studied the homing properties of B lymphocytes by using 51 Cr-labeled lymphoid cells obtained from athymic, nu/nu mice, and animals made T-lymphocyte deficient by thymectomy and lethal irradiation followed by reconstitution with syngeneic bone marrow. Comparison was made to the patterns of distribution observed when cell preparations containing normal numbers of T and B lymphocytes were migrated. A small but significant percentage of labeled lymphocytes from lymph nodes, spleen, Peyers Patches, and bone marrow of T-cell-deficient animals was shown to be lymph node seeking. Secondary transfers of lymph node cells from primary recipients caused enrichment of this lymph node-seeking population. Treatment of T-lymphocyte-deficient lymphoid cell preparations with neuraminidase reduced the percentages of cells homing to the lymph nodes. The data showed that B lymphocytes exhibit unique homing properties when injected into normal recipients. In addition, direct comparison of the homing patterns of B lymphocytes prepared from spleen and lymph nodes of athymic mice revealed differences suggesting that these lymphoid organs contained unique mixtures of at least two different kinds of B cell. The evidence supports the notion that the B-lymphocyte populations contain at least two subpopulations, one of which possesses the ability to home to lymph nodes.

Variation in the homing properties of 51Cr-labeled thy antigen-positive lymphoma

Abstract The homing properties of five thy antigen-positive murine lymphoma cell lines in normal syngeneic mice were compared to those of thymus, lymph node, bone marrow, and spleen cells. YAC lymphoma cells migrated in significant numbers to the spleen but in somewhat smaller percentages than normal lymphoid cells. An immunoselected subline of YAC was recovered from recipient spleens in about double the amounts observed with the parental lymphoma line. The migration patterns produced by lymphoma YC8, AKR, and LSTRA more closely resemble those of the selected subline of YAC. None of the lymphoma used in this study homed to recipient lymph nodes. That the selective distribution of labeled lymphoma cells was a function of cell viability was demonstrated by the fact that heat-killed lymphoma cells were recovered almost exclusively from the liver. Neuraminidase treatment, prior to 51 Cr labeling, changed the migration in vivo of all the lymphoma studied except YAC. In contrast, trypsinization of the lymphoma did not influence their distribution in vivo . Binding of concanavalin A (Con A) to the cell membrane modified the homing of normal lymph node cells but did not change the migration of lymphoma cells. The conclusion was drawn that many receptors normally present on T lymphocytes are missing or modified after transformation and that the lymphoma in question more closely resembles the sessile splenic T1 lymphocyte.

Kinetic studies of macrophages: II. The effect of specific antisera on radiolabeled peritoneal cells

Abstract The distribution of intraperitoneally injected 51Cr-labeled peritoneal cells in syngeneic recipient mice was markedly altered by treatment with antimacrophage serum. The spleen-seeking segment of the peritoneal population was virtually eliminated by treatment of the cells in vitro with antimacrophage serum. Pretreatment of recipient mice with the same serum resulted in increased accumulation of labeled cells in the spleen and reduced migration of cells to the bone marrow. Injection of the recipients with antimacrophage serum at the time of cell introduction caused decreased recovery of labeled peritoneal cells from the spleen. Treatment of donors with antimacrophage serum resulted in nearly normal distribution patterns. Antithymocyte serum and normal rabbit serum did not effect the distribution of intraperitoneally injected labeled peritoneal cells. AMS was shown to be ineffective in promoting the survival of skin allografts when ALS activity was removed by absorption with lymphocytes.