Ayala Sharp

Aging in the T Lymphocyte Compartment

A decline in the capacity of bone marrow cells to differentiate to T lymphocytes was found when cells from young and old donors were seeded onto an alymphoid fetal thymus. A step-by-step analysis of cell-cell interactions of the lymphohemopoietic cells and the thymic stroma indicated an effect of age on a variety of cell differentiation parameters. These included a decrease in the affinity of bone marrow cells to the stroma, and in their capacity to compete with the thymic lymphoid resident cells on colonization of the thymus. There was a significant decrease in the ability of cells of old donors to replicate sequentially within the thymic microenvironment. There was a reduced capacity of bone marrow cells from aging mice to express a developmental preference after seeding onto a syngeneic fetal thymus in a mixture with cells from allogeneic donors. We addressed the question whether the aging thymus contains increased levels of immature cells that fail to differentiate in the involuted thymic microenvironment by seeding thymocytes from young and old donors onto the fetal thymic stroma. The values of T cells that developed from the old donor inoculum were lower under these conditions. Our studies suggest that at least some of the manifestations of aging in the T cell compartment are related to developmentally programmed events in the lymphohemopoietic cell compartment.

The bone marrow as an effector T cell organ in aging

The idea that the bone marrow (BM) might function as a T cell effector organ and might constitute a compensating system to the decreased T cell compartment in aging was examined by carrying out an analysis of T cell functions in this tissue. The Thy1+ cells, which were found to increase in their proportions with age in the BM, were sorted out from the BM of young and old mice by flow cytometry and their proliferative response to Concanavalin A (conA) stimulation was measured. The sorted Thy1+ cells responded to conA at levels comparable to those of splenocytes, with the old BM showing a significantly lower response than the young. To determine whether cells of the intact BM behave in a similar pattern to the sorted cells, we measured the responses induced by conA in unseparated BM cells of both age groups. The results showed that the patterns of proliferative response in the intact BM cells were different than those observed in splenocytes and in the Thy1+ sorted cells. Hence, cells of the old BM manifested initially higher levels of proliferation preceding that of the young BM, yet the response was apparent for a shorter duration. When we measured the conA induced proliferation of the intact BM cells in the presence of colchicine, the number of BM cells entering the first mitotic cycle was higher in the old. Thus, it seems that there are more effector T cells in the old BM, yet their response to stimulation is of shorter duration. This conclusion was also supported by the assessment of cytotoxic T lymphocytes precursor (CTLp) frequency and of CTL function of the BM. CTLp was higher in the old BM following 3 days of incubation and lower following 7 and 9 days of incubation. The old BM cells showed a reduced CTL response at the proliferative phase when stimulated for 4-7 days, yet their effector cell reactivity was either equal or more efficient than the young. To determine whether some of the differences between the two age groups were due to regulatory effects of the BM microenvironment, BM cells from young and old donors were admixed with young mouse splenocytes and stimulated with conA. The conA induced response was enhanced up to tenfold under these conditions yet to the same extent in both age groups. Thus, it appears that the BM has the capacity to function as a T cell effector organ.(ABSTRACT TRUNCATED AT 400 WORDS)

MHC-Linked Colonization of the Thymus and Thymocyte Development: Effects of Mature T Lymphocytes

Effects of mature T lymphocytes on thymic colonization by lymphohemopoietic cells were investigated in an in vitro experimental model, using a variety of experimental strategies. Lymphoid-depleted fetal thymus (FT) explants (C57BL/Ka, Thy1.1, H-2b) were incubated with bone marrow (BM) cells from syngeneic (C57BL/Ka; SBM) and allogeneic (BALB/c, Thy1.2, H-2d; ABM) donors. Cocultures of FT with SBM and ABM, depleted of Thy1+ or of CD3+ cells, resulted in equal proportions of lymphocytes from both BM donors. When peripheral blood lymphocytes (PBL) from synegenic or semi-allogeneic donors (F1[C57BL/Ka x C57BL/6J], Thy1.1/Thy1.2); or F1[C57BL/Ka x BALB/c], Thy1.1/Thy1.2, respectively) were added to these cultures, the total lymphocyte count per thymic lobe decreased and a developmental preference of the SBM-derived cells, as compared to the ABM-derived cells, was noted. Cells of the PBL types were also observed in the cultures. Cocultures of FT with ABM and PBL showed reduced proportions of ABM-derived cells and occurrence of cells of the PBL type. Finally, FT explants partially depleted of lymphocytes by irradiation (6 Gy), were cocultured with PBL from either syngeneic or allogeneic donors. In the presence of syngeneic PBL, the total number of cells and the proportion of double-positive (CD4+CD8+) T cells were similar to those in the FT cultured by itself, whereas in the presence of allogeneic PBL these values were reduced. The study suggests that mature T lymphocytes may play a role in the developmental processes in the thymus, and points to MHC-linked selective effects.

Interactions of bone marrow cells from young and old mice with syngeneic and allogeneic thymic tissue

Age-related changes manifested in MHC-linked recognition of bone marrow (BM) cells by the thymic stroma were studied in vitro model of thymus-BM chimeras. Fetal thymuses (FT) depleted of self-lymphocytes were colonized with BM cells from syngeneic and allogeneic donor mice. When cells from young (3-month-old) or old (24-month-old) donors syngeneic to the stroma were seeded in a mixture with cells of allogeneic young origins (C57BL/6J-Thy1.2 and ARK/J-Thy1.1 seeded onto C57BL/6J FT), the syngeneic cells showed an age-related developmental advantage. Accordingly, cells from the old syngeneic mice manifested a significantly reduced capacity to compete with allogeneic cells when compared with the young syngeneic cells. When allogeneic BM cells from young or old mice were seeded onto the thymic stroma in a mixture with BM cells from young donors syngeneic to that stroma (BALB/c-Thy1.2 mixed with C57BL/Ka-Thy1.1 seeded onto C57BL/6J or C57BL/Ka FT), the Thy1+ cells which developed were mainly of syngeneic origin. The age of the allogeneic cells had no significant effect on the results. However, when old allogeneic cells were mixed with old syngeneic cells, the developmental advantage of the syngeneic cells was not manifested. When seeding of allogeneic cells was followed 1 day later by seeding of syngeneic cells, the syngeneic advantage was eliminated, suggesting that the MHC-linked competition began during the first 24 hr of contact with the thymic tissue. When BM-derived thmocytes grown in FT explants were transferred onto second FT recipient explants of the same genotype as the first ones, the syngeneic advantage was abolished, suggesting either that the thymic microenvironment was modified as a result of colonization or that it induced a change in the BM cells. In this respect, the young allogeneic BM-derived thymocytes showed a significant advantage when compared with the old cells. Thus, the MHC-linked syngeneic preference in the early development of BM cells is also manifested in aging mice, yet at a level that is significantly reduced compared with that seen in the young mice.

MHC-linked syngeneic developmental preference in thymic lobes colonized with bone marrow cells: a mathematical model.

Reconstitution of the T-cell compartment after bone marrow transplantation depends on successful colonization of the thymus by bone-marrow-derived progenitor cells. Recent studies compared the development of syngeneic and allogeneic bone-marrow-derived cells in cocultures with lymphoid-depleted fetal thymus explants, leading to the discovery of MHC-linked syngeneic developmental preference (SDP) in the thymus. To determine the nature of cell interactions among the bone marrow and thymic elements that might underlie SDP, we analyzed this phenomenon by mathematical modeling. The results indicate that syngeneic mature T cells, responsible for inducing this preference, probably interfere both with the seeding of allogeneic bone-marrow-derived thymocyte progenitors in the thymic stroma and with their subsequent proliferation. In addition, the possibility of augmented death among the developing allogeneic thymocytes cannot be ruled out.

An anti-acetylcholine receptor monoclonal antibody cross-reacts with phosvitin

Rabbit and mouse anti‐Torpedo acetylcholine receptor antibodies cross‐reacted partially with the highly phosphorylated protein, phosvitin. We have selected an anti‐Torpedo acetylcholine receptor monoclonal antibody which binds specifically to phosvitin; this binding is inhibited by acetylcholine receptor. These findings suggest that a phosphorylated amino acid residue may be a part of the determinant on the acetylcholine receptor recognized by this monoclonal antibody.