James Clagett

The distribution of rapidly and slowly renewed T, B, and “null” lymphocytes in mouse bone marrow, thymus, lymph nodes, and spleen

Abstract Combined radioautography and immunofluorescence were employed to discern the proportions of rapidly renewed (RR) and slowly renewed (SR) T, B, and null cells in mouse lymph nodes (LN), spleen (Spl), thymus (Thy), and the lymphocyte-rich fraction of bone marrow (BML). Thy and BML were found to consist predominantly of cells with a rapid turnover rate (95.4 and 76.9%, respectively) in accord with their roles as primary lymphoid organs. In contrast, the secondary lymphoid organs were primarily composed of SR cells (LN, 73.4%; Spl, 67.8%) and contained a more even admixture of the six lymphocyte subsets. Most cells in Thy were RR T cells (93.3%), whereas the predominant lymphocyte subpopulation in both LN and Spl consisted of SR T cells (56 and 50%, respectively), with RR T cells being much less frequent (14.6% LN; 6.8% Spl). BML contained both RR and SR cells which stained with the T-cell reagent (7.8 and 7.1%). These percentages are probably overestimates, however, since this reagent stained some nonlymphoid cells in BML. RR B cells were most plentiful in BML (31.4%) and Spl (23.3%), less common in LN (11.2%), and rare in Thy (0.4%). SR B cells were equally numerous in Spl (16%), LN (15.4%), and BML (15.3%). RR null cells were the most prevalent cell type in BML (37.7%), but were infrequent in other tissues (1.6% Thy, 2.1% LN, 2.1% Spl). SR null cells were rare in all tissues (0% Thy, 2% LN, 2.1% Spl, 0.7% BML). These experiments represent the first comprehensive investigation of the composition of the lymphomyeloid organs in terms of RR and SR T, B, and null cells. They conclusively demonstrate RR and SR lymphocytes in all three functional categories (T, B, and null) and show characteristic tissue distributions of the six lymphocyte subsets.

A cat model for the evaluation of mechanisms of bone resorption: induction of bone loss by simulated immune complexes and inhibition by indomethacin.

SummaryWhen simulated immune complexes (SIC) (heat-aggregated IgG) possessing many of the properties of true antigen-antibody complexes were injected via the root canal into the periapical tissues of cat maxillary cuspids, radiographically and histologically evident bone resorption occurred at these sites within 7 days. Bone loss was accompanied in all cases by inflammation of the surrounding collagenous connective tissues and was characterized by the presence of osteoclasts. Bone resorption, but not the accumulation of inflammatory cells, was blocked by the systemic administration of indomethacin, an inhibitor of prostaglandin synthetase. The most likely explanation is that SIC-activated mechanisms such as the complement cascade, prostaglandin synthesis, and neutrophil degranulation were responsible for the bone loss. The minor inflammation and bone loss that followed the repeated injections of BSA and of monomeric IgG can best be explained as a response to trauma. The data presented establish that the cat maxillary cuspid is a useful model in which to explore the mechanism underlying pathological bone resorption.

Mitogen-induced hyperproliferation response of peripheral blood mononuclear cells from patients with severe generalized periodontitis: Lack of correlation with proportions of T cells and T-cell subsets☆

Severe generalized periodontitis (SGP) is a localized inflammatory disease which differs clinically from common periodontitis in that it leads to remarkable extensive alveolar bone loss in relatively young adults. There is evidence that B-cell responses to bacterial substances may play a major role in the pathogenesis of this disease. In the present report, we show that a B-cell mitogen from Actinomyces viscosus (AVIS) bacteria provokes a hyperproliferation response of peripheral blood mononuclear cells (PBMNC) from these patients. In addition, AVIS-stimulated PBMNC from SGP patients proliferate for longer periods in culture than do PBMNC from control subjects. There were, however, no differences between patients and controls in the numbers of immunoglobulin-secreting cells in these cultures as determined by an indirect plaque-forming cell assay. The possibility that differences in numerical proportions of regulatory T-cell subsets may play a role in the mitogen-induced hyperproliferation phenomenon is examined. PBMNC were stained with fluorescein isothiocyanate-conjugated monoclonal antibodies OKT3, OKT4, and OKT8 in order to identify, respectively, total T cells, helper/inducer, and suppressor/cytotoxic subsets. Flow cytometric analysis of such specifically stained cell preparations from 14 control subjects and 14 SGP patients did not reveal any significant differences between the proportions of total T cells or T-cell subsets of the two groups. Furthermore, there were no statistically significant correlations between the magnitude of proliferation responses and the proportions of total T cells or either of the T-cell subsets.

Effects of prostaglandin on the antigen- and mitogen-driven responses of peripheral blood lymphocytes from patients with adult and juvenile periodontitis☆

Abstract Peripheral blood lymphoid cells were obtained from individuals with juvenile periodontitis or the adult form of the disease and from normal control subjects and were cultured with and without bacterial substances or polyclonal activators. Blastogenic response and cytotoxic factor production were measured and the effectiveness of prostaglandin- E 2 in inhibiting these cell functions was evaluated. The data show that cells obtained from individuals with juvenile periodontitis in general respond to stimulation in vitro more vigorously and are suppressed by prostaglandin E 2 less effectively than cells from individuals with the adult form of the disease or cells from normal control individuals. Measurement of activated lymphoid cell responses in the presence and absence of PGE 2 may have diagnostic usefulness in distinguishing individuals with the juvenile form from those with adult forms of the disease.

Effects of cell concentration and exogenous prostaglandin on the interaction and responsiveness of human peripheral blood leukocytes.

Abstract The role of cell-to-cell interactions and the effects of endogenous and exogenous prostaglandin on the mitogenic and antigenic responsiveness of peripheral blood leukocytes from five adult human subjects were evaluated. Cells were activated by phytohemagglutinin (PHA) or a homogenate of Actinomyces viscosus (AVIS), a gram-positive suspected pathogen, in the presence or absence of prostaglandin E2 (PGE2) or indomethacin. A cell concentration range from 0.1 to 10 × 105 was studied using flat-bottom microtest wells. Cells from all five donors, who ranged in age from 33 to 46 years and who were healthy and clinically and radiographically free of inflammatory periodontal disease, could be induced to respond to AVIS blastogenically by increasing the cell concentration. PGE2 at 10 μM suppressed responsiveness to AVIS at all cell concentrations, while the PHA response was suppressed at some concentrations and enhanced at others. Regression analysis of the log-log transformation of the data indicated that a single cell type may be responsible for the PHA response. In contrast, the responses to AVIS appeared to be more complex. In order to obtain a significant DNA synthetic response, 5- to 10-fold more cells were required than for PHA, and two cell types participated.

Immunoregulation in severe generalized periodontitis.

Severe generalized periodontitis (SGP) is an inflammatory disease which leads to extensive alveolar bone loss in young adults. Peripheral blood lymphocytes from SGP patients have been previously reported to exhibit an in vitro hyperproliferative response when exposed to B cell mitogens derived from Staphylococcus aureus and Actinomyces viscosus. Therefore hyperresponsiveness to B-cell mitogens could be an important pathogenic factor in the susceptibility to and progression of SGP. We have tested whether the hyperproliferative response of lymphocytes from SGP patients was due to (i) a functional deficiency of suppressor T cells, or (ii) to numerical alterations of lymphocytes. Supernatant fluids from concanavalin A-stimulated T cells from 14 SGP patients and 14 normal subjects were compared for their ability to suppress the IgM synthesis of B-cell mitogen-stimulated mouse splenocytes. No significant differences were noted in suppressor T-cell function between control subjects and SGP patients. However, SGP patients had significantly higher lymphocyte counts than control subjects, and there was a positive correlation between high lymphocyte counts and high mitogen-stimulated proliferation. SGP patients also had higher lymphocyte:monocyte ratios than control subjects, suggesting that a defect in macrophage-mediated suppression might be involved in the hyperproliferation phenomenon. Our data do not support the hypothesis that a suppressor T-cell defect is the cause of mitogen-induced hyperproliferative responsiveness of peripheral blood lymphocytes from SGP patients. Rather, hyperproliferation may be due to an expansion of the lymphocyte pool which responds to mitogens, or/and a regulatory disturbance which arises because of altered lymphocyte:macrophage ratios.

The relative importance of the bone marrow and spleen in the production and dissemination of B lymphocytes.

Abstract The relative importance of the bone marrow and spleen in the production of B lymphocytes was investigated in guinea pigs by the combined use of [ 3 H]TdR radio-autography and fluorescent microscopy after the staining of B cells by FITC-F(ab′) 2 -goat-anti-guinea pig Ig. Large and small lymphoid cells possess sIg in the marrow and spleen but B cell turnover in the marrow exceeds that in the spleen. That newly generated bone marrow B cells are not derived from an extramyeloid bursa equivalent was demonstrated by the absence of [ 3 H]TdR labeled B cells in tibial marrow 72 hr after [ 3 H]TdR was administered systemically, while the circulation to the hind limbs was occluded. Pulse and chase studies with [ 3 H]TdR showed that large marrow B cells are derived from sIg-negative, proliferating precursors resident in the bone marrow and not from the enlargement of activated small B lymphocytes. The acquisition of [ 3 H]TdR by splenic B cells lagged behind that observed in the marrow. Three days after topical labeling of tibial and femoral bone marrow with [ 3 H]TdR, a substantial proportion of splenic B cells were replaced by cells that had seeded there from the labeled marrow. The studies unequivocally identify the bone marrow as the organ of primary importance in B cell generation and indicate that in the guinea pig rapidly renewed B lymphocytes of the spleen are replaced by lymphocytes recently generated in bone marrow. The rate of replacement of B lymphocytes in the lymph node by cells newly generated in the bone marrow takes place at a slower tempo than in the spleen.

Chymopapain C, an Immunosuppressive Protease: I. Partial Purification and Characterization

Summary This study has demonstrated the presence of a proteolytic enzyme, chymopapain C, in the extract of Carica papaya. This enzyme differs significantly from papain in enzymatic activity, thermal half-life and action of human IgG. However, this enzyme is similar to commercial chymopapain in its activity and in its digestion of human IgG, but differs in molecular weight, thermal half-life and relative electrophoretic mobility.

THE BEIGE (bg) GENE INFLUENCES THE DEVELOPMENT OF AUTOIMMUNE DISEASE IN SB/Le MALE MICE

Publisher Summary This chapter focuses on recessive mutation beige ( bg ) that produces a number of pleiotropic effects in mice. This includes increased susceptibility to pneumonitis and cytomegalovirus infections, defective granulocyte chemotaxis and lysosomal enzyme secretion, and giant lysosomal granules in granule-containing cells. The frequency of natural killer (NK) cells in beige mice is normal, and after exposure to interferon (IFN) or IFN inducers, NK cell activity in beige mice can be augmented to almost normal levels. A progressive fatal autoimmune disease develops spontaneously that is markedly accelerated in male animals. Like the BXSB mice derived from them, male SB mice develop a fatal lupus-like syndrome and that the bg locus has a profound effect on the development of this autoimmune disease. In a study described in the chapter, cell subsets were quantified on ortho-fluorescence-activated cell sorter (FACS) using fluorescein-conjugated rat monoclonal antibodies specific for mouse differentiation antigens. The bg mutation does not selectively impair NK lymphoid cells and should not be evoked simply as a murine model of NK deficiency.

Mitogenic activity of Actinomyces viscosus. II. Induction of DNA and immunoglobulin synthesis in rabbit B lymphocytes.

Abstract A cell wall extract from Actinomyces viscosus (AVIS) induces blastogenesis in a large proportion of lymphocytes from non-immunized rabbits. The response appears to be a polyclonal B cell response, since purified T cells are unresponsive, whereas B cell-enriched cultures exhibit 3-fold higher DNA synthesis than conventional splenocyte cultures. Furthermore, AVIS triggers immunoglobulin synthesis and plasma cell formation in vitro . The rabbit response is similar to that of the mouse, and we suggest that non-specific polyclonal B cell responsiveness to Actinomyces bacteria may be an important host defense mechanism common to these, and probably other, mammalian species.