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PubMed1979

Effect of chloramphenicol on in vitro function of lymphocytes.

The effect of chloramphenicol on the in vitro function of human peripheral blood lymphocytes was studied in assays of lymphocyte transformation and lymphokine production. When lymphocytes were stimulated by phytohemagglutinin, concanavalin A, or pokeweed mitogen in the presence of various concentrations of chloramphenicol, only minimal effects on blastogenesis were noted. However, suppression by chloramphenicol of blastogenesis induced by candida antigen or streptokinase-streptodornase was greater in magnitude and was dose-dependent; blastogenesis was suppressed to 25%--30% or normal levels by concentrations of chloramphenicol of 25--50 microgram/ml. Chloramphenicol had little effect on the production of the lymphokine leukocyte migration inhibition factor by lymphocytes stimulated either by candida antigen or by concanavalin A, whereas puromycin at a concentration of 5 microgram/ml significantly suppressed this response. Thus chloramphenicol appears to suppress antigen-induced lymphocyte blastogenesis significantly but not lymphokine production by stimulated lymphocytes.

Antigens, Fungal↗

PubMed1980

Direct stimulation of lymphokine production by cephalothin.

Cephalothin significantly suppressed in vitro DNA and total protein synthesis in human peripheral blood lymphocytes stimulated by antigens or mitogens. However, similar concentrations of this antibiotic enhanced streptokinase-streptodornase-stimulated production of the lymphokine leukocyte-migration-inhibition factor (LMIF) and directly stimulated production of this lymphokine by otherwise unstimulated lymphocytes from 10 of 12 normal human subjects. Penicillin did not appear to produce these effects. Cephalothin did not interfere directly with neutrophil migration or the interaction of preformed LMIF with neutrophils. Stimulation of LMIF production by cephalothin required viable lymphocytes and was inhibited by puromycin. These results suggest that cephalothin is capable of inducing lymphokine production by human lymphocytes in a manner that appears to be nonspecific in nature. This type of effect could be the basis of some apparently immunologic reactions to this antibiotic.

Cephalothin↗

PubMed1980

Longitudinal assay of lymphocyte responsiveness in patients with major burns.

Serial blast transformation in vitro was measured in peripheral lymphocytes from 38 patients with major thermal injury. Lymphocytes were tested with the antigens streptokinase-streptodornase (SKSD), mumps and purified protein derivative (PPD), the mitogens concanavalin A (Con A) and phytohemagglutinin (PHA), and in the one-way mixed lymphocyte reaction. Statistically significant suppression by the burn injury was noticed in all measurements except response to PHA. One-time measurements were not significantly different between the patients who survived and the patients who did not survive their burn injuries. However, serial determinations of responsiveness to the three natural antigens SKSD, mumps and PPD, as well as the mixed lymphocyte reaction accurately reflected prognosis.

Adolescent↗

PubMed1982

Suppressor cell activity after major injury: indirect and direct functional assays.

In vitro responses to streptokinase-streptodornase (SKSD), mumps, and mixed lymphocyte culture (MLC) in 19 burned and 13 multitrauma patients were studied sequentially by lymphocyte tritiated thymidine incorporation and compared to responses of 28 normals. Mean responses to SKSD remained significantly depressed from normal for up to 14 to 28 days following injury: MLC responses, significantly depressed at 48 hours, recovered promptly to normal levels. Because of evidence that the proliferative capacity of the T-cell population to soluble antigens is contained within the inducer subpopulation while both inducer and suppressor subpopulations respond in MLC, the observed increase in MLC responses, coupled with a sustained depression of SKSD and mumps responses, suggests activation of a population of suppressor cells. In a direct assay of suppressor cell function, lymphocytes from three or four multitrauma patients incubated in a two-way MLC with normal lymphocytes significantly suppressed PHA responsiveness, confirming the findings of the direct assay.

Adolescent↗

PubMed1977

Application of laser cytometry to the analysis of immunologically induced in vitro lymphocyte responsiveness.

This study defines an assay (laser analysis) that is a significant advance in our ability to quantitate and analyze immunologically induced in vitro lymphocyte responsiveness. Laser analysis is demonstrated to parallel radionucleotide incorporation (3H-thymidine) in terms of kinetic pattern, dose response characteristics, and statistical accuracy while exeeding radionucleotide incorporation in sensitivity. Direct quantitation of lymphocyte responsiveness, in terms of cellular proliferation, disclosed that substantial numbers of small lymphocytes were produced during in vitro stimulation with mitogen (concanavalin A) or antigen (streptokinase-streptodornase) in addition to the expected increase in lymphoblasts. The magnitude of this "total cellular response" (lymphocytes plus lymphoblasts) was found to be similar for antigen and mitogen stimulation, a finding not suggested by routine radionucleotide incorporation or morphological assays.

Concanavalin A↗

PubMed1980

Cimetidine-induced augmentation of human lymphocyte blastogenesis by mitogen, bacterial antigen, and alloantigen.

The effect of Cimetidine, a histamine-type 2 receptor antagonist, was evaluated on the in vitro proliferative response of normal human peripheral blood lymphocytes (PBLs). Cimetidine (10(-3) to 10(-8) M) increased mitogen-induced blastogenesis by 22% (phytohemagglutinin (PHA) and by 27% (pokeweed) over nondrug-treated control values (P less than 005 for PHA and pokeweed). Preincubation of PBLs with Cimetidine further augmented blastogenesis as much as 2- to 3-fold (P less than 0.005 for both mitogens). Multiple testing of the same normal subject demonstrated consistent reproducibility of increased proliferation by Cimetidine. Similar statistically significant amplifications of the proliferative res-ponse were observed when bacterial antigen (streptokinase-streptodornase) or alloantigen was used to induce blastogenesis. Optimally effective concentrations of Cimetidine ranged from 10(-5) to 10(-7) M, which corresponds to expected clinical serum levels. These observations suggest that a histamine-type 2 receptor antagonist is capable of modulating the proliferative response of PBLs in the absence of exogenously added histamine. The immunoregulatory implication of this Cimetidine-induced proliferative augmentation is discussed in relation to clinical transplantation and cancer immunotherapy.

Antigens, Bacterial↗

PubMed1979

Comparison of lymphocyte function after isolation by ficoll-hypaque flotation or elutriation.

Mononuclear leukocytes were separated from whole blood by ficoll-hypaque flotation and by elutriation (counterflow centrifugation). Lymphocytes isolated from 6 control subjects by elutriation showed a 30% greater response to stimulation with phytohemagglutinin and 130% greater response to streptokinase-streptodornase stimulation than did autologous lymphocytes obtained by ficoll-hypaque separation. Cell yields of major mononuclear cell subpopulations and cell viability were comparable after separation of leukocytes by both techniques. These results indicate that ficoll-hypaque flotation may diminish lymphocyte responses, and that elutriation offers a useful alternative to ficoll-hypaque separation. In addition, elutriation may be the preferable method for evaluation of lymphocytes from patients with suspected immunologic dysfunction and may be valuable in the isolation of mononuclear cells from infiltrated skin lesions.

Cell Separation↗

PubMed1978

Cell-mediated immune reactions in asthma.

Abnormally high levels of IgE are produced in various diseases, including atopic eczema, and a functional decrease in the T-lymphocyte population occurs at the same time. T-cell function has been studied in a group of asthma patients with high, normal and low IgE levels and in a control group. T-cell function appears to be reduced in asthmatic patients with high concentrations of IgE when skin tests are performed with Candida antigen (P less than 0.05). There was also a decline in the lymphocyte transformation tests with candida antigen in autologous serum (P less than 0.05), although not in foetal calf serum. The responses to in vivo tests with streptokinase-streptodornase were normal, as was the response of the lymphocytes to PHA.

Adolescent↗

PubMed1992

Cellular immunity in tuberculous pleural effusions: evidence of spontaneous lymphocyte proliferation and antigen-specific accelerated responses to purified protein derivative (PPD).

The kinetics of in vitro cellular proliferation against a PPD of Mycobacterium tuberculosis or streptococcal antigen (streptokinase-streptodornase) was evaluated in pleural fluid and peripheral blood mononuclear cells (PBMC) from patients with tuberculous and non-tuberculous pleuritis. The peak proliferative response to PPD by mononuclear cells from pleural fluid occurred earlier (day 3) in 65% of patients with tuberculosis, a finding not seen in non-tuberculous effusions. Spontaneous lymphocyte proliferation of both peripheral blood lymphocytes and pleural effusion lymphocytes was frequently observed, irrespective of etiology. However, 20 of 21 tuberculous patients manifesting spontaneous lymphocyte proliferation had accelerated kinetics of proliferation to PPD, which was antigen-specific. These results suggest that spontaneous lymphocyte proliferation occurs as a response to antigen stimulation at the site of disease, and is not a non-specific response to inflammation. Furthermore, enhanced reactivity against mycobacterial antigen, manifested by accelerated kinetics of proliferation, has diagnostic potential in patients with pleural effusions.

Humans↗

PubMed1985

Delayed hypersensitivity skin testing of 150 volunteers. Retesting results and antibody measurements.

We studied skin test reactivity to five commonly used antigens by testing 150 healthy adults. The delayed hypersensitivity (DH) skin test is widely used to assess the immune status of patients. The battery of antigens suitable for use may vary in different countries, but the reactivity to the antigens in our population did not differ remarkably from reports of other authors. The reactivity rates were: candida 32.7%, mumps 86%, streptokinase-streptodornase (SK-SD) 70%, trichophyton 0% and tuberculin 58.7%. Sixteen of the subjects were retested after 2 weeks. Only eight of them showed unchanged reactions to all five antigens. Specific IgG antibody concentrations measured by enzyme-linked immunosorbent assay (ELISA) against each of the five antigens in the serum of 42 subjects before and after testing showed great inter-individual variation. The antibody concentration did not correlate with the DH skin test results, but the testing itself increased the production of anti-mumps- and anti-SK-SD-antibodies.

Adolescent↗

PubMed1982

Augmentation of delayed-type hypersensitivity in antigen-primed guinea pigs by human dialyzable leukocyte extract. Chromatographic and enzymatic characterization of the active principle.

Column chromatographic purification and sensitivity towards enzymatic treatments of dialyzable transfer factor (TFd), the immunologically specific component of dialyzable leukocyte extract (DLE), have previously been used in its biochemical characterisation. In the present work we studied the effect of enzymes and the Sephadex G-10 chromatographic separation of the components of DLE augmenting delayed-type hypersensitivity. Skin reactivities to streptokinase-streptodornase (SK-SD) and tuberculin PPD were significantly augmented by injecting DLE into antigen-primed guinea pigs. The augmentation caused by DLE treatment correlated to the pre-existing level of immunity in the recipients. Most of the augmentory activity resided in 2 adjacent fractions, eluting early from a Sephadex G-10 column. This augmentation was destroyed by alkaline hydrolysis, by treatment with pronase, proteinase K, ribonuclease, and nuclease P1, but not by alkaline phosphatase or phosphodiesterase II. The observed sensitivities towards these enzymes, except that for ribonuclease, were closely similar to those described for the specific TFd component of DLE. These results are compatible with the idea that either the nonspecific augmenting and the specific TFd molecules are principally similar, or that the TFd molecules, in addition to their capacity to transfer specific immunity, also have an augmenting effect, which needs in its manifestation a sub-threshold dose of immunogen.

Adjuvants, Immunologic↗

PubMed1976

Ontogeny of human cell-mediated immunity: age-related variation of in vitro infantile lymphocyte transformation.

Thymidine uptake of unstimulated and of phytohemagglutinin (PHA)-, streptokinase-streptodornase (SK-SD)- and candida extract (candida)-stimulated lymphocytes of normal infants less than 20 months old was evaluated. Thymidine uptake of unstimulated and PHA-stimulated young infants' lymphocytes was avid, resembling that of newborn cells. Over periods of weeks to months, infantile lymphocytes demonstrated transition in unstimulated and PHA-induced thymidine uptake from response patterns like cord blood cells to ones more characteristic of adult lymphocytes. Candida- and SK-SD-induced thymidine uptake of lymphocytes from very young infants was likewise apparently quite avid, resembling cord blood cells. However, factoring out high unstimulated thymidine uptake by conversion of data to stimulation indexes clarified differing paces of acquisition of transformation responsiveness of the two naturally acquired infectious antigens. Specific lymphocytes reactivity to SK-SD was quite low in all groups of infants compared with candida-induced transformation, which in some infants acquired adult proportions.

Aging↗

PubMed1978

Cell-mediated immunity in diabetes mellitus.

Cell-mediated immunity was evaluated in patients with diabetes mellitus by delayed hypersensitivity skin tests and in vitro lymphocyte transformations. Only 44% of diabetic patients had skin test reactivity to Candida antigen, compared with 88% of normal controls (P < 0.001). Insulin-dependent diabetic (IDD) patients had abnormally low lymphocyte transformation responses to phytohemagglutinin, concanavalin A, and streptokinase-streptodornase (P < 0.05). This defect was not corrected by culturing the cells in nondiabetic plasma. IDD patients with persistent hyperglycemia (fasting serum glucose level, >200 mg/dl) had lower levels of transformation than did IDD patients with fasting serum glucose levels less than 150 mg/dl. Lymphocytes from two IDD patients with poor lymphocyte transformation responses had marked improvement in response to phytohemagglutinin when the lymphocytes were cultured after a preincubation period designed to deplete cultures of suppressor activity. Seven IDD patients were studied serially over 12 months. Lymphocyte transformation responses in four of these patients improved coincidentally with a change in the level of fasting hyperglycemia from >200 to <150 mg/dl. The other three IDD patients with consistent fasting serum glucose levels of >200 mg/dl had poor lymphocyte transformation responses. Diabetic patients have demonstrable defects in lymphocyte function which improved in a small number of patients with reduction in the level of fasting hyperglycemia.

Adolescent↗

PubMed1980

Modulation of immunoreactivity to periodontal disease-associated microorganisms during pregnancy.

The lymphocyte blastogenic response to a panel of antigens and mitogens was assessed in a group of 20 women throughout their pregnancy. In addition, a group of five nonpregnant women was monitored simultaneously to identify variations in response to the same stimulants. The stimulants included orally associated bacterial antigens (Streptococcus sanguis, Actinomyces viscosus, Bacteroides asaccharolyticus, Bacteroides melaninogenicus subsp. intermedius, Bacteroides [Capnocytophaga] ochraceus, and Fusobacterium nucleatum) and non-orally associated-stimulants (streptokinase-streptodornase, tetanus toxoid, concanavalin A, phytohemagglutinin, and pokeweed mitogen). Intrinsic (cells cultured in male AB plasma) suppression of the lymphocyte response to these stimulants was observed to occur by the second trmester of pregnancy and was resolved after parturition. Additionally, an extrinsic (cells cultured in autologous plasma) suppression was also suggested to occur in a similar manner. There was no detectable enhancement of the blastogenic response to oral bacteria associated with elevated gingivitis, which is generally reported to occur during nonpregnancy gingivitis. We propose that concomitant immunosuppression occurs during the second trimester, which masks such enhancement.

Actinomyces↗

PubMed1973

In vitro correlates of delayed hypersensitivity in man: ambiguity of polymorphonuclear neutrophils as indicator cells in leukocyte migration test.

Delayed cutaneous hypersensitivity (DCH) of 12 normal adult subjects to purified protein derivative (PPD) of Mycobacterium tuberculosis, streptococcal streptokinase-streptodornase (SK-SD), and Candida albicans Dermatophytin O (DO) was assayed in vivo by skin testing and compared with such in vitro correlates of cellular immunity as lymphocyte transformation (LT) and inhibition of leukocyte migration (ILM) from microcapillary tubes or in agarose gel. LT was shown to be the best in vitro correlate of specific lymphocyte sensitization with all antigens. In the ILM assays, PPD showed good correlation with in vivo DCH and in vitro LT; SK-SD showed partial correlation; DO showed no correlation, not being active in any of the ILM tests. Cell distribution and morphology of stained migration patterns, ILM tests performed on separated populations of lymphocytes and polymorphonuclear leukocytes (PMN), as well as the ability of test antigens to stimulate PMN cells to reduce nitroblue-tetrazolium dye, indicated that in ILM tests mononuclear cells were not inhibited in their migration, whereas migration of PMN cells appeared to depend on their direct reaction with the test antigens.

Antigens, Bacterial↗

PubMed1976

Cellular requirements for simulation by the purified streptococcal antigen DSR.

Delayed skin reactor (DSR) was prepared from streptokinase-streptodornase. Peripheral blood mononuclear cells, were collected from a Ficoll-Hypaque gradient and subsequently subdivided into T cell-, B cell-, or monocyte-depleted populations. Mononuclear cells with complement receptors (B cells) were removed by a nylon-wool column adsorption. Sheep erythrocyte rosette forming cells (T cells) were removed by centrifugation on a second gradient. Populations of cells which did not adhere to plastic and/or ingest iron filings were monocyte-depleted. While response to DSR as measured by incorporation of tritiated thymidine into DNA was not affected by removing B lymphocytes, it was significantly diminished by removing T lymphocytes. Monocyte-depleted cells incorporated less tritiated thymidine into DNA in both antigen stimulated and control cells, but the stimulation index was not significantly altered.

Antigens↗

PubMed1977

Biologic activity of transfer factor tested in vitro with cord blood lymphocytes.

Cord blood lymphocytes, generally believed to be immunologically uncommitted, were used for investigating the possible conversion by transfer factor (TF) of non-immune to immune lymphocytes in vitro. TF was prepared from pooled buffy coats of normal blood bank donors by ultrafiltration and lyophilization, and it ability to influence lymphocyte transformation was assessed by measuring increases in 3H-thymidine uptake into DNA after stimulation with purified protein derivative of tuberculin and/or streptokinase-streptodornase in 23 experiments with cord blood and 15 adult controls. TF stimulated cord blood lymphocytes nonspecifically when added to the medium alone. In the presence of antigen, TF acted in a synergistic, antigen-dependent way with either amplifying or inhibitory effects in adult good responders. These effects were negligible in adult low responders and in cord blood. From our observations no specific conversion of lymphocytes by TF in vitro can be deduced.

Adult↗

PubMed1992

Alveolar macrophage accessory cell function in bronchial asthma.

The capacity of peripheral blood monocytes and alveolar macrophages (AM) obtained by bronchoalveolar lavage (BAL) to present recall antigens, namely, tuberculin purified protein derivative (PPD) or streptokinase-streptodornase (SKSD), to highly purified autologous T-cells has been studied in 11 asthmatic and 11 healthy, nonatopic normal subjects. In the asthmatic group, AM accessory cell function was variable, and most subjects were unable to present either recall antigen as effectively as blood monocytes, although one asthmatic subject demonstrated larger proliferative responses than blood monocytes for both antigens. AM accessory cell activity was not antigen-specific, and there was a correlation between accessory cell efficacy for the two antigens (r = 0.92; confidence interval, 0.53 to 0.98). Furthermore, a correlation existed between the percentage lymphocyte count in the BAL fluid and the ratio of macrophage to monocyte antigen-presenting capability for both PPD (r = 0.92; 95% confidence interval, 0.83 to 0.99) and SKSD (r = 0.90; 95% confidence interval, 0.45 to 0.98). In the normal subjects, AM were also unable to act effectively as accessory cells for the presentation of PPD and SKSD in the majority of subjects. No correlation existed between the percentage lymphocytes in BAL fluid and the ratio of AM to monocyte accessory cell function. These results suggest an association between AM accessory function and the presence of BAL lymphocytes in bronchial asthma.

Adult↗