The RT6 T cell antigen: its role in the identification of functional subsets and in T cell activation.
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Animals↗
Biomedical subjects
D M Lubaroff
Publications and source records attributed to D M Lubaroff.
Monoclonal antibodies to RT7 and LCA antigens in the rat: cell distribution and segregation analysis.
Fluorescein-conjugated monoclonal antibodies (mAb) have been used to examine the cellular distribution and genetic relationship of the RT7 and the leukocyte common antigen (L-CA) in the rat. We demonstrate here that the RT7.1 alloantigen, recognized by the mAb BC84, is present on B lymphocytes as well as T lymphocytes, although at markedly different apparent concentrations; T cells binding approximately 4 times more BC84 than do B cells. In contrast, the polymorphic L-CA recognized by the NDS58 mAb and the non-polymorphic L-CA recognized by the OX1 mAb appear to be expressed in equal concentrations on T and B cells and differ in their tissue distribution from the RT7.1 alloantigen. We have also tested a new mAb termed 8G6.1, that is reactive with RT7b rat strains and has a cell, tissue and strain distribution profile resembling that of a polymorphic L-CA. Segregation analysis of the RT7 and polymorphic L-CA antigenic systems using a single backcross model demonstrate that the alloantigens recognized by BC84 and NDS58 cosegregate and that both are allelic with respect to 8G6.1. The L-CA antigenic determinant defined by OX1 was non-polymorphic and thus was not allelic with any of the antigenic systems tested. These results suggest that there is a close genetic relationship between the expression of RT7 and L-CA in the rat.
Alleles↗
Characterization of the heterogeneity of R3327 rat prostatic tumors derived from single-cell clones.
Prostatic adenocarcinoma is characterized by cellular diversity, which is well demonstrated in the Dunning R3327 rat prostatic adenocarcinoma. This heterogeneity may arise from epigenetic influences, ie, cellular adaptation or selection, and/or from genetic changes. To investigate the question of genetic instability, four tissue culture cell lines were derived from single cells isolated from the uncloned late (UCL) passage of the Dunning R3327H prostate cell culture. Each of these clonally derived tissue cultures was injected into castrated and intact young adult male rats for tumor production. Uncloned early (UCE) and UCL passage tissue cultures were also propagated as solid tumors. Tumors and the cultures from which they were derived were examined for evidence of phenotypic and genetic changes using morphological and cytometric methods. Transmission and scanning electron microscopy revealed only slight differences among the cell cultures. A single population of diploid cells was demonstrated in each of the cell cultures by propidium iodide staining and subsequent flow cytometric measurement of DNA content/nucleus. Tumors of unicellular as well as multicellular origin exhibited extreme heterogeneity of histological features, both among animals as well as within a single tumor. Tumors were surveyed and tissue types were characterized and cataloged. Clone 3 was generally better differentiated than the others; tumors from castrated animals were better differentiated than those from intact animals. Flow cytometry revealed multiple hyperdiploid cell populations that were variable from one sample to another. We concluded that changes in genotype as well as phenotype occurred in the tumors derived from single cells. Some of these changes may have occurred in the cells while still in culture.
Adenocarcinoma↗
Differential effects of prolactin on rat dorsolateral prostate and R3327 prostatic tumor sublines.
Many investigators have reported effects of the pituitary hormone, prolactin, on the physiology and biochemistry of the rat prostate gland, particularly the lateral or dorsolateral lobe. The Dunning R3327H is a transplantable rat prostatic adenocarcinoma derived from a spontaneous tumor of the Copenhagen rat dorsolateral prostate. This study describes and compares morphological and physiological effects of prolactin on rat dorsolateral prostate and two sublines of the Dunning tumor. Ectopic pituitary grafts were used to induce chronic hyperprolactinemia in castrated rats receiving androgen supplement to provide a relatively controlled hormonal environment in which the effects of prolactin were maximally and consistently observed. Gravimetric and biochemical analyses, as well as ultrastructural study, provided evidence of prolactin's stimulatory effect on dorsolateral prostate growth and secretory activity. Hyperprolactinemia stimulated the growth of the well-differentiated, androgen-dependent R3327/3219 tumor subline with an increase in weight, volume and the total content of DNA, protein and zinc. There were no changes in tumor morphology. In contrast, the anaplastic androgen-independent R3327/150 tumor subline did not respond to graft-induced hyperprolactinemia. This differential response of the two R3327 tumor sublines attests to the complexity of prolactin's effects on prostatic tissue and to the extent of the deterioration of endocrine control that often accompanies tumor progression. Prolactin binding in the R3327 sublines was studied using immunohistochemical staining and radioligand assay, but produced complex results which raise questions about the discrepancy between hormone binding and biological action of prolactin in prostatic tissues.
Adenocarcinoma↗
Membrane antigen phenotype of sensitized T lymphocytes mediating tuberculin-delayed hypersensitivity in rats.
The membrane antigen phenotype of immune lymph node cells (LNC) which mediate tuberculin-delayed hypersensitivity (DH) in Lewis rats was examined. The results show that the T-cell population which expresses the RT7.1 alloantigen defined by the BC 84A monoclonal antibody contained cells capable of transferring DH. Separation of the RT7.1-positive T-cell population with the monoclonal antibodies W3/25, MRC OX-8, or DS 4.23 (which defines the RT6.1 alloantigen) revealed that either the W3/25-positive or the RT6.1-negative T-cell subpopulations contained DH effector cells, whereas the corresponding MRC OX-8-positive or RT6.1-positive T-cell subpopulations did not. Moreover, when the W3/25-positive T-cell subpopulation was divided into either RT6.1-positive or RT6.1-negative T-cell subsets, only the W3/25-positive, RT6.1-negative subset transferred DH. These results indicate that the effector cells that mediate tuberculin DH are contained within the immune T-cell subset which bears both the RT7.1 and the W3/25 markers, but lacks both the MRC OX-8 and the RT6.1 markers.
Animals↗
Androgen receptor binding characteristics in the cytosol of the rat dorsolateral prostate gland and the Dunning R-3327 prostatic adenocarcinoma.
The incidence of prostatic cancer is highly correlated with advanced age, and it has been suggested that changes in androgen binding may be important in age-associated alterations in growth regulatory mechanisms of prostatic epithelial cells. In this study the effects of age on androgen binding characteristics in the dorsolateral prostate glands of young and aged Copenhagen rats were determined and the binding properties in the Dunning R3327/130 subline of rat prostatic adenocarcinoma were characterized. Tritium-labeled and nonlabeled methyltrienolone analogs (R1881) were used to study the binding properties of 5 alpha-dihydrotestosterone receptor in the cytosol of tumors and prostate glands. Binding of R1881 was low but specific for the androgen receptor as shown by competition studies in which nonlabeled R1881, 5 alpha-dihydrotestosterone, and testosterone competed successfully with 3H-R1881 for binding sites, but 17 beta-estradiol and low levels of progesterone did not. In Copenhagen dorsolateral prostate, Scatchard analysis suggested a single class of binding sites. In young animals (three to five months) the average binding capacity was 10.36 fmol/mg cytosol protein with a dissociation constant (Kd) of 2.28 nmol/L. The dorsolateral prostate of aged rats (11-16 months) showed no significant difference in specific binding characteristics as compared to the younger age group. Specific binding of 3H-R1881 in R3327/130 tumor was saturable with a single class of high-affinity binding sites having an average binding capacity of 64.77 fmol/mg cytosol protein and a Kd of 2.76 nmol/L. These data show that the tumor had approximately 6.5 times the number of binding sites as did the normal Copenhagen rat dorsolateral prostate gland. However, no age-related changes were detected through 11-16 months of age in the androgen binding characteristics of normal rat dorsolateral prostate gland that could be correlated with the higher concentration of androgen binding sites in the R3327/130 tumor subline.
Adenocarcinoma↗
Cryotherapy.
Since 1969 cryosurgical perineal destruction of the prostate has been developed at the University of Iowa for treatment of patients with prostatic cancer. The technique of this procedure has been the subject of previous reviews. This is a summary of our experience of patient survival and the effect on local prostatic cancer growth in those patients with various stages of the disease subjected to perineal cryosurgical destruction of their cancer. We also report briefly on our experience in the laboratory where the Dunning tumor model is used to study the effect of cryosurgery as well as combinations of immunological modifications.
Adult↗
Potentiation of the rat delayed-type hypersensitivity reaction by the Fc portion of human IgG1.
Fc fragments derived from a human IgG1 myeloma protein potentiate the rat delayed-type hypersensitivity (DTH) reaction to antigen challenge. Lewis rats immunized with heat-killed tubercle bacilli give augmented DTH reactions to the purified protein derivative of tuberculin when Fc fragments are included in the challenge dose. Similar potentiation of DTH by pFc' fragments indicates that the active site is located in the CH3 domain of IgG1. Histologic evaluation of the augmented reaction sites revealed predominantly mononuclear cell infiltrates characteristic of DTH reactions. Skin tests of tubercle bacilli-sensitized rats with an unrelated antigen and/or Fc fragments fail to elicit significant reactions. Augmentation of the DTH reaction to purified protein derivative is restricted to the Fc or pFc' region fragments since intact monomeric IgG1, Fab fragments, and bovine serum albumin were all shown not to be active potentiators. The DTH reaction of ovalbumin-sensitized rats was similarly augmented when Fc fragments were included with a challenge dose of ovalbumin, thus supporting the general nature of the phenomenon. These results support the concept of Ig molecules as multifunctional proteins that can not only serve effector functions but also participate in the regulation of immune responses.
Animals↗
Identification of thymocyte progenitors in hemopoietic tissues of the rat. I. A quantitative assay system for thymocyte regeneration.
A quantitative adoptive transfer system is described which can be used to precisely monitor the generation of thymocytes and peripheral T cell subsets by precursor cells in hemopoietic tissues of rats. This assay utilizes the rat pan-T cell alloantigens, A.R.T.-1a and A.R.T.-1b, and the fluorescence-activated cell sorter (FACS) to directly enumerate donor- and host-origin thymocytes and T cells in irradiated, histo-compatible recipients. The assay has the advantage over related systems in that it detects more than 95% of total newly-formed thymocytes and T cells at all stages of differentiation; that it is highly sensitive (as few as 1% of the appropriate A.R.T.-1 bearing cells can be detected in cell mixtures); and that it permits selective recovery of donor-and/or host-origin cells for further characterization. The results show that, after a lag period of 10-12 days, the regenerative kinetics of donor-origin thymocytes are linear with respect to time and cell dose, over a range of 2.5 to 50 X 10(6) bone marrow cells. Above a threshold of approximately 400R, the regenerative kinetics of donor-origin thymocytes are independent of irradiation dose, but are inversely related to the age of the recipient. On a per cell basis, bone marrow cells are more efficient than spleen cells at regenerating the thymus. By 4 months after bone marrow cell transfer, permanent chimeras are established in which the proportion of donor-origin T cells approximates that of donor-origin thymocytes, and in which the ratio of presumptive helper (W3/25+) and suppressor/cytotoxic (OX8+) donor-origin T cells is normal. The assay therefore appears to be suitable for following both the purification of prothymocytes from rat hemopoietic tissues and the proliferation and differentiation of their progeny in thymus and peripheral lymphoid tissues.
Animals↗
Characterization of monoclonal antibodies that define rat T cell alloantigens.
A panel of monoclonal antibodies with specificity for rat T cell alloantigens was prepared by somatic cell hybridization with the use of spleen cells from either BH (ART-1b, 2b) or NBR (ART-1b, 2a) rats immunized with LEW (ART-1a, 2a) thymus, spleen, or lymph node cells. Comparison of the antibodies resulted in three distinct patterns of complement-mediated cytotoxicity. Further comparison of four representative hybridoma antibodies with antisera having specificity for ART antigens by indirect immunofluorescence demonstrated that two of the three patterns corresponded to anti-ART-1a and anti-ART-2a specificities. The patterns of binding of the anti-ART hybridomas were shown to be different from those produced by mouse monoclonal antibodies having anti-leukocyte-common (L-C) or anti-Thy-1.1 activities as well as antibodies produced by the monoclonal W3/25, which detect a subpopulation of T lymphocytes. Functional studies performed in vitro with cells fractionated by treatment with the monoclonal anti-ART antibodies and complement showed the ART-1 determinant recognized by the hybridoma products BC 84A or BY 12.3 is expressed by mature cytolytic T lymphocytes (CTL) and by cells involved in the generation of CTL during MLR, but not plaque-forming cells. Together, the data suggest the antibodies BC 84A and BY 12.3 can be used to define rat T cell populations. To date, the functional significance of cell populations recognized by monoclonal antibodies having anti-ART-2a or anti-ART-3-like activities has not been determined.
Animals↗
Genetic linkage and cell distribution analysis of T cell alloantigens in the rat.
The T cell alloantigens A.R.T.-1, A.R.T.-2, Pta, Ag-F and RT-Ly-2 were examined for linkage to albinism and the haemoglobin variant Hbb, and for cell expression similarities in the rat. The A.R.T.-1 alloantigen was not linked to either A.R.T.-2, albinism or Hbb. The A.R.T.-2 alloantigen was demonstrated to be closely associated to albinism and Hbb with a recombination frequency of approximately 3% with albinism and 1% with Hbb. The four recombinants (out of 104 animals examined) were subsequently analysed with alloantisera to Ag-F, Pta and RT-Ly-2. These alloantisera detected an antigenic system(s) which appeared to co-segregate with A.R.T.-2 in the recombinants studied Characterization of the specific T cell populations expressing each alloantigen demonstrated that removal of the cells bearing any one of the alloantigens removed all the cells expressing any of the other alloantigens. These results demonstrate that the four alloantigenic systems A.R.T.-2, Pta, Ag-F and RT-Ly-2 are located in the same genetic region of linkage group I, and appear to be expressed on the same peripheral T cell subpopulation(s) in the rat.
Animals↗
Maturation of functional T-lymphocyte subpopulations in the rat.
The T-lymphocyte differentiation alloantigens, A.R.T.-1 and A.R.T.-2 were used to follow the functional development of these cells in the rat. Delayed hypersensitivity and lymphocytes cytotoxic for allogeneic tumor cells were present in adoptively transferred recipients by 3 wk postirradiation and fetal liver reconstitution. However, these animals did not regain appreciable GvH potential until 7.5 wk postreconstitution. Normal percentages of A.R.T-1 lymphocytes were observed at all times in the spleen and lymph nodes of these animals, whereas, below normal numbers of A.R.T-2 lymphocytes were observed up to 7.5 wk postreconstitution. Depletion experiments using specific anti-A.R.T. sera demonstrated that the proliferative response to PHA and the cytotoxic lymphocyte response to alloantigens were reduced after treatment with anti-A.R.T.-1, but not with anti-A.R.T.-2. In contrast, the GvH potential of lymph node cells was reduced by treatment with either anti-A.R.T.-1 or anti-A.R.T.-2 serum plus complement. The use of these A.R.T. alloantisera in the rat has greatly facilitated the establishment of differentiation patterns for functional T lymphocytes and has resulted in the association of specific functional T-cell subpopulations with characteristic surface alloantigens.
Animals↗
Immunologic aspects of the prostate.
Our experiments involving the use of the Dunning R3327 adenocarcinoma as an animal model of prostatic cancer as well as clinical studies on the immunocompetence of prostatic cancer patients are described. Utilizing the Dunning tumor, we have demonstrated that this transplantable adenocarcinoma of the rat prostate was similar to human prostatic cancer with respect to its macroscopic and microscopic appearances, growth rate, growth differential in male and female recipients, and some of its metastatic potential. Cryosurgery was capable of destroying the primary tumor as it can in man. Both antibody and cellular immune responses could be produced against antigens associated with the tumor cells. Tumor-bearing rats treated by cryosurgery in combination with BCG were capable of producing an antitumor immunity that protected them from rechallenge. Clinical studies of prostatic cancer patients showed a diminished in vitro immunity, but the responses of the cancer patients were not significantly different from those of patients with benign prostatic disease.
Adenocarcinoma↗
A quantitative assay system for thymocyte regeneration in the rat.
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Animals↗
Separation of antibody specificities in anti-A.R.T.T-cell sera.
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Absorption↗
The Dunning tumors.
We have described the Dunning tumor, R3327, as a most useful model system in studies of human prostatic cancer. The tumor resembles the human disease in its histologic picture, ultrastructure, and most of its metastatic capabilities. The one exception to the latter is the apparent absence of metastatic lesions in the bone. Like the variability seen in human cancer of the prostate, the R3327 has been shown to develop multiple sublines, each with different histologic pictures, growth rate, and androgen dependence. Thus far the literature is replete with reports of the Dunning tumor as a model in hormonal, chemotherapeutic, and immunologic experiments. When we look at the results obtained in the five years since the Dunning tumor has been resurrected, we can be encouraged with what the future of prostatic cancer research holds with the use of this system.
Adenocarcinoma↗
Studies on a weakly immunogenic squamous cell carcinoma of the rat prostate.
Different methods were tested to elicit and assess transplantation immunity against the weakly immunogenic 11095A squamous cell carcinoma, a methylcholanthrene-induced prostatic tumor of the inbred F344 rat strain. The results described emphasize the difficulty with which effective immune responses are induced by very weak transplantation antigens. The importance of using poorly immunogenic tumors as research models is discussed.
Animals↗
Comparative studies of T-lymphocyte antigens in the rat: are ART, Ly, Pta, and Ag-F the same?
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Animals↗