Role of B lymphocytes in cell-mediated immunity. I. Requirement for T cells or T-cell products for antigen-induced B-cell activation.

Although B lymphocytes can be triggered by B-cell mitogens and by certain other molecules to produce lymphokines, they do not produce lymphokines when stimulated with specific soluble protein antigens. We have investigated whether T-cell help would enable B cells to produce lymphokines when activated by antigens. Addition of small numbers of T cells to B-cell cultures resulted in significant production of a monocyte chemotactic factor. T cells could be replaced by supernates of antigen-stimulated T cells, demonstrating both that the chemotactic factor was B-cell-dervied and that T-cell help was mediated by a soluble factor. Although the T-cell factor was nonantigen specific, B-cell activation required the presence of both antigen and T-cell factor. Thus, it appears that although dependent upon T cells, B lymphocytes may play an important role in amplification of cell-mediated immune responses.

Identification of a specific interleukin 1 inhibitor in the urine of febrile patients.

The urine of febrile patients has been found to contain high concentrations of an inhibitor of interleukin 1 (IL-1)-induced thymocyte proliferation. The inhibitor is specific for IL-1 and does not block the effects of interleukin 2 (IL-2) or phytohemagglutin (PHA) on thymocytes, and it is not nonspecifically toxic for these cells. IL-1 inhibitor can be found in the urine of normal individuals and afebrile patients, but is present in increased concentrations in the urine of patients with fever of diverse etiologies. Preliminary physicochemical characterization indicates that the inhibitor is a 20-40-kdalton protein.

Binding of aggregated gamma-globulin to activated T lymphocytes in the guinea pig.

Heat-aggregated guinea pig gamma-globulin was shown to bind to the surface membrane of a subclass of guinea pig T lymphocytes. Cells of this subpopulation were identified as T lymphocytes because these cells did not stain for surface Ig (a B-cell marker) but did form spontaneous E-rosettes with rabbit erythrocytes (a T-cell marker). A strikingly high proportion of such aggregate-binding (Agg(+)), E-rosette-forming (E-rosette(+)), but surface Ig-negative (Ig(-)) cells were found in an inflammatory exudate. Thus purified peritoneal exudate lymphocytes (PELs) are known to consist of over 90% T cells, and 59% of these cells bound aggregates. 10% of these Agg+ Ig- E-rosette+ cells were found in draining lymph node cell populations and none in thymus cell populations. The high frequency amongst PELs suggested that these Aggregate+ Ig- E-rosette+ cells might be activated T cells as these are known to occur in high proportion in PEL populations. Confirmatory evidence for this postulate was provided by the striking increase (from 10% to 46%) of Ig- E-rosette+ cells that bound aggregates when lymph node cells were activated by antigen stimulation in vitro.

A synthetic glycolipid with B-cell mitogenic activity.

A synthetic glycolipid, N-palmitoyl-D-glucosamine, (NPG) was found to be a potent B-cell mitogen, that stimulated spleen cells from nu/nu as well as from normal mice. The proper dispersion of the water insoluble preparation is critical for the elicitation of this mitogenic effect. Limulus lysate clotting assay indicated that the NPG preparation either contains only 0.001% endotoxin contamination, or that NPG itself is 10(-5) times less active in this assay than purified endotoxic LPS. Since such low levels of endotoxin concentration are not mitogenic, it is concluded that synthetic N-palmitoyl-D-glucosamine, when properly dispersed, is itself a B-cell mitogen.

The peritoneal exudate lymphocyte. I. Differences in antigen responsiveness between peritoneal exudate and lymph node lymphocytes from immunized guinea pigs.

Peritoneal exudate lymphocytes from guinea pigs immunized with horse radish peroxidase, dinitrophenyl guinea pig albumin, or ferritin in complete Freund's adjuvant have been shown to be significantly more reactive than other lymphocytes in two in vitro assays of cellular immune function: production of macrophage migration inhibitory factor and antigen-induced lymphocyte proliferation. The enhanced reactivity of peritoneal exudate lymphocytes cannot be accounted for by artifacts introduced by column purification or by the presence of nonlymphoid accessory cells. These observations suggest that the peritoneal exudate lymphocyte pool is a highly enriched source of cellular immune effector cells with specificity directed towards those antigenic determinants to which an animal has been recently exposed.

The uropod-bearing lymphocyte of the guinea pig. Evidence for thymic origin.

In this study, the frequency of uropod formation and the type of lymphocyte bearing the uropod was investigated in various guinea pig lymphocyte populations. Without prior in vitro stimulation, almost 40% of peritoneal exudate lymphocytes (PELS) form uropods, while thymocytes and lymph node cells form far fewer. Subsequent stimulation in vitro with purified protein derivative demonstrated that there is an association between antigen reactivity and frequency of uropod formation in these populations. The ultrastructure of these uropods is identical to that described for human lymphocytes stimulated with phytohemagglutinin. In the populations studied, all the lymphocytes forming uropods lack easily detectable surface membrane immunoglobulin and are therefore most likely thymus-derived or T lymphocytes.

Cellular requirements for lipopolysaccharide adjuvanticity. A role for both T lymphocytes and macrophages for in vitro responses to particulate antigens.

By employing primary cultures of purified spleen cells from lipopolysaccharide (LPS) responder (C3H/HeN or C57BL/10Sn) or nonresponder (C3H/HeJ or C57BL/10ScN) mice incubated with particulate antigen and LPS prepared by phenol-water extraction (Ph), we have presented evidence that both T cells and macrophages (MO) are required for LPS-induced adjuvanticity. First, MO derived from C3H/HeN spleen cells, when mixed with responder, C3H/HeN lymphocytes and Ph-LPS, elicited enhanced antibody responses to sheep erythrocytes (SRC) antigen, whereas lymphocytes from the nonresponder, C3H/HeJ mouse strain did not evoke this response. Similarly, purified T cells from C3H/HeN spleens, when cultured with responder, nu/nu spleen cells, and Ph-LPS yielded enhanced anti-TNP PFC responses; whereas, C3H/HeJ T cells did not potentiate immune responses when mixed with optimal concentrations of Ph-LPS. LPS prepared by butanol-water extraction elicited significant adjuvant effects with all cell combinations. Finally, purified responder T cells and MO enabled either responder or nonresponder B cells to elicit LPS potentiation. These data indicate that T cells and MO are controlling LPS-induced augmentation of B-cell responses.

Detection of a mediator derived from endotoxin-stimulated macrohpages that induces the acute phase serum amyloid A response in mice.

The mechanism by which LPS stimulates an acute phase serum amyloid A (SAA) response in C3H mice has been studied. A factor (SAA inducer) appears in the blood of C3H/HeN (lipopolysaccharide [LPS]-sensitive) mice approximately 1 h after administration of LPS, which, when passively administered, can induce C3H/HeJ mice to produce SAA although they are resistant to the LPS itself. SAA inducer has been detected in the culture medium of LPS treated C3H/HeN macrophages but not spleen cells. Thus, two stages in the induction of the acute phase SAA response are now recognized: a latent period of 2-3 h during which the SAA concentration remains at baseline values and in which SAA inducer appears, and the period of synthesis of SAA which lasts for approoximately 24 h past induction. It is proposed that a macrophage response to LPS is responsible for production of the serum mediator which induces SAA synthesis.

A human urine-derived interleukin 1 inhibitor. Homology with deoxyribonuclease I.

We have previously reported that the urine of febrile humans contained large quantities of an inhibitor of IL-1-induced murine thymocyte proliferation that was a glycoprotein between 30 and 40 kD in size. In the present study this factor has been purified to homogeneity using a sequence of eight purification steps (ammonium sulfate precipitation, ion exchange chromatography, molecular sieve chromatography, hydrophobic affinity chromatography, hydroxylapatite chromatography, fast protein liquid chromatography, and two HPLC steps). SDS-PAGE analysis indicates that the purified material is a 38-kD molecule. Evidence based on a partial amino acid sequence analysis as well as enzyme studies indicates that this inhibitor is a type of human DNase I.

Objective response of multiple myeloma to cyclosporin A.

Two patients with advanced multiple myeloma were treated with oral low-dose cyclosporin A. one without other therapy and one in conjunction with chemotherapy to which the patient had been previously unresponsive. Both patients had objective laboratory and clinical responses. In the patient treated with cyclosporin A alone, decreasing serum IL-6 and beta-2-microglobulin levels fell as the clinical response evolved. Cyclosporin A deserves further evaluation in the therapy of multiple myeloma.

Analysis of the systemic corticosteroid sensitivity of patients with primary open-angle glaucoma.

Patients with primary open-angle glaucoma have an ocular and systemic sensitivity to corticosteroids. We adapted a cellular assay that used peripheral blood lymphocytes to detect this corticosteroid sensitivity in vitro in a microtiter assay. It reduced the time, cost, and amount of blood required to examine a patient. We examined ten subjects on three separate days and demonstrated that the reliability of one 50% inhibitory concentration was about 76%. We then studied 25 patients with primary open-angle glaucoma and 25 control subjects using this in vitro assay. The patients with primary open-angle glaucoma were significantly more sensitive to corticosteroids than the control subjects (P less than .001).

Immunological comparison of plaque-resistant and plaque-susceptible inbred rat strains.

AN IMMUNOLOGICAL comparison of ODU Plaque-Resistant (RES) and Plaque-Susceptible (SUS) Rats was performed in order to determine if plaque accumulation was secondary to some immunological abnormality, and to ascertain the effects of plaque accumulation on the immune system. Plaque accumulation in SUS rats on powder diets is associated with a significant elevation in immunoglobulin levels over RES rats, especially in serum IgG and IgM. Young (less than 9 weeks) SUS rats possess fewer splenic T lymphocytes than do young RES rats. This decrease is associated with a marked decrease in the response of spleen cells in vitro to T cell mitogens, Con A and PHA. This decrease is unrelated to diet and is completely and spontaneously reversed in the adult (3 month) rats. These studies demonstrate that the accumulation of large amounts of plaque cause an elevation in immunoglobulin levels. However, plaque accumulation in SUS rats does not appear to be secondary to any demonstrable immunologic abnormality.

The use of parent report to assess the quality of care in primary care visits among children with asthma.

OBJECTIVE: To determine the accuracy of parent report and the accuracy of the medical record in documenting physician performance of elements of pediatric asthma care in the primary care setting. METHODS: A convenience sample of 79 English-speaking parents of 4--12-year old children with asthma presenting to medical center--affiliated inner-city primary care pediatric clinics in the Bronx, Dallas, and Chicago was enrolled, and the office visit was audiotaped. Parents were interviewed 1--16 days after the visit by telephone. OUTCOME MEASURES: Accuracy of parent report was the primary outcome. The "reference standard" was an independent evaluation of the audiotaped record of the primary care visit. The National Asthma Education and Prevention Program was used as a guide to select data elements to assess quality of pediatric asthma care during primary care visits. RESULTS: Sufficient documentation was significantly (P <.001) less likely to be present in the medical record than in the follow-up interview for each element of care. When these elements were combined into a cumulative score, 71% of parent interviews but only 37% of medical records scored > or = 5 (out of a possible 6), with 29% of medical records scoring < 3. Parents were able to accurately report (concordance of parent data with audiotape reference standard) whether or not the visit had included performance of 5 of the 6 elements of care. CONCLUSIONS: Our study suggests that parent telephone interview within 2 weeks after the visit is more accurate than the medical record for documentation of the quality of asthma care in pediatric primary care visits. The medical record was not sufficient to assess the quality of primary care related to asthma, primarily because of missing data. Therefore, our data suggest that assessing quality of care using the medical record will not only bias the findings in the direction of more deficient care but will also make improvement in care more difficult. Further validation of our strategy for using parent report to assess the quality of care in primary care visits will require its application in a variety of other primary care settings.

Evaluation of delayed hypersensitivity: from PPD to poison ivy.

Intact cell mediated immunity (CMI) requires a complex interaction between lymphocytes, macrophages, and cytokines. The presence of functional CMI can be easily tested in vivo by delayed hypersensitivity (DTH) skin testing. Delayed hypersensitivity skin testing has been commonly used in three ways; anergy screening, testing for infection with intracellular pathogens, and testing for sensitivity to contact allergens. Accurate anergy screening requires intradermal testing with four or more common recall antigens such as Candida albicans, tetanus toxoid, mumps, and purified protein derivative. Problems of antigen potency and availability can be overcome by using a multiple time test-like device that contains seven different recall antigens (Multitest CMI). Although testing for infection was once common, it is now only recommended for tuberculosis screening because most other antigens are either not predictive of infection or not commercially available. Accurate testing for contact allergy requires careful attention to technique, and limitation of allergens to the 20 antigens that are approved by the American Academy of Dermatology.

Effect of inoculation of sarcoid tissue into athymic (nude) mice.

Previous reports have suggested that the inoculation of sterile homogenates of human sarcoidosis tissue produce pathological changes or death in T cell deficient mice, while other reports have challenged these results. Because of the importance of this point, the potential infectiousness of human sarcoid tissue was investigated using athymic (nude) mice. Eight tissue specimens were obtained from patients with sarcoidosis and inoculated once into a total of sixty-two nude mice. These experiments were conducted over a period of two years. The tissue specimens inoculated included biopsies of lung, lymph node and muscle. A non-tissue buffer was used as a sham control and inoculated into ten nude mice. Of the eight tissue specimens, six were found to contain non-caseating granulomas consistent with sarcoidosis. Only one of these six specimens induced significant pathological changes when injected into nude mice: two mice developed reticulum cell sarcoma and one developed membranous glomerulonephritis. Five inoculations of sarcoid tissue into a new athymic mouse colony housed in resterilized cages produced no detectable pathology. Two different tissue specimens not containing sarcoid granulomas were inoculated into two groups of nude mice. Both of these specimens induced pathological changes. One mouse developed a reticulum cell sarcoma, one developed lymphoma and one developed interstitial nephritis. A saline control inoculation produced no pathology in any mice. Mortality rates were similar in sarcoid inoculated, normal tissue inoculated and saline inoculated mice. These findings suggest that if sarcoidosis has an infectious etiology, the responsible agent is not one that easily infects athymic mice.(ABSTRACT TRUNCATED AT 250 WORDS)

Discrimination between urticaria-prone and other allergic patients by intradermal skin testing with codeine.

To study the ability of cutaneous mast cells to degranulate in urticaria-prone patients, subjects were skin tested with the known mast cell degranulator, codeine sulfate. Sensitivity to codeine as determined by the concentrations of codeine necessary to cause a net wheal of 5 mm was compared between urticaria-prone subjects, allergic subjects, and normal control subjects. Urticaria-prone subjects were more sensitive to codeine at every concentration tested and exhibited a mean reactivity to codeine that was almost 100 times that of the other allergic individuals and normal control subjects. This difference could not be explained by an increased sensitivity to histamine in 71% of urticaria-prone patients nor by any dermatographic tendencies or increased relative allergic reactivity. These findings suggest that codeine skin testing can be used to identify a distinct population of patients with urticaria.