bluuebunny/crossref_metadata_2025 huggingface dataset:
Depletion of CD4+ T cells but not inhibition of the protective activity of IFN-gamma prevents cure of toxoplasmosis mediated by drug therapy in mice.
Authors: Araujo F G
Published: December 2022

Toxoplasmosis is a frequent opportunistic infection in patients with AIDS. In these patients the major immune deficiencies are a severe depletion of CD4+ T lymphocytes and an impaired capacity to produce IFN-gamma. A mouse model was developed and used to study the effects that depletion of CD4+ T cells and/or inhibition of the protective activity of IFN-gamma have on the effectiveness of the drug therapy for toxoplasmosis. Infection of mice with a lethal inoculum of Toxoplasma gondii cysts followed by treatment with the hydroxynaphthoquinone 566C80 or with sulfadiazine resulted in 100% survival whereas untreated controls had 100% mortality within 15 days of infection. Administration of antiserum to IFN-gamma resulted in early death of untreated mice and in 30% mortality in those treated. Administration of mAb to CD4+ T cells followed by infection with T. gondii prevented the development of both antibody and cell-mediated immune responses against the parasite. These mice resisted the acute infection while undergoing specific treatment. Discontinuation of the treatment, however, resulted in reactivation of the infection and the majority of the animals died within 17 days of suspension of the treatment. Administration of antiserum to IFN-gamma or to CD4+ T cells 24 h but not 15 days after conclusion of the treatment also resulted in mortality. These results indicate that successful treatment of toxoplasmosis depends on the status of the immune system, particularly of CD4+ T cells. Although it is speculative to compare results obtained in mice to the situation in humans, our work suggests that restoration of a competent immune response is of crucial importance for a successful treatment of toxoplasmosis in immunocompromised individuals.
IFN-gamma and IL-2 are protective in the skin but pathologic in the corneas of HSV-1-infected mice.
Authors: Hendricks R L , Tumpey T M , Finnegan A
Published: December 2022

HSV type 1 (HSV-1) infection of the mouse cornea results in a tissue-destructive inflammatory reaction in the cornea, but little or no disease in the skin surrounding the eye. Depleting T lymphocytes from mice before HSV-1 corneal infection prevents the corneal inflammation but severely exacerbates the periocular skin lesions. Studies described in this communication investigated the role of T cell cytokines in the corneal and periocular skin disease induced by HSV-1 corneal infection. Mice received weekly i.p. injections of rat mAb specific for IL-2, IL-4, or IFN-gamma beginning 1 day before (day -1) or 6 days after (day +6) corneal infection with the RE strain of HSV-1. The severity of corneal inflammation and the area of periocular skin involvement were measured. Treatment with anti-IFN-gamma or anti-IL-2 significantly reduced the incidence and severity of corneal inflammation. Treatment was equally effective when initiated on day -1 (before T cell activation) or day +6 (after T cell activation but before the initiation of corneal inflammation). Treatment with anti-IL-4 had no effect. The histologic features of corneal inflammation in mock-treated mice included neovascularization, corneal edema, and cellular infiltration. Corneas of anti-IL-2-treated mice that developed inflammation had similar but less severe histologic features. Corneas of anti-IFN-gamma-treated mice that developed inflammation had neovascularization and edema but minimal cellular infiltration. Treatment with anti-IFN-gamma or anti-IL-2 significantly exacerbated periocular skin lesions when initiated at day -1, but not when initiated at day +6. Anti-IL-4 treatment had no effect on skin lesions. Treatment with either anti-IFN-gamma or anti-IL-2, when initiated at day -1, significantly inhibited the delayed-type hypersensitivity response to HSV Ag, but when treatment was begun at day +6 only anti-IFN-gamma significantly inhibited the delayed-type hypersensitivity response. Our findings suggest that IFN-gamma and IL-2 are important elements in both an immunopathologic T-lymphocyte response to HSV-1 Ag in the cornea and a protective T lymphocyte response in the skin.
Biochemical and immunologic characterization of a major IgE-inducing filarial antigen of Brugia malayi and implications for the pathogenesis of tropical pulmonary eosinophilia.
Authors: Lobos E , Ondo A , Ottesen E A , Nutman T B
Published: December 2022

A major allergen of the human filarial parasite Brugia malayi has been identified by two-dimensional immunoblot analysis using a serum pool from patients with tropical pulmonary eosinophilia. The allergen is composed of two Ag with M(r) 23 and M(r) 25 and acidic isoelectric point (Bm23-25). Immunoblots using affinity-purified IgE antibodies to BM23-25 indicated that Bm23-25 is expressed mainly in the microfilarial stage. Digestion of the allergen with endoglycosidases indicates that it has N-linked oligosaccharide chains. Analysis of the reactivity of T cells derived from patients with lymphatic filariasis revealed that the Bm23-25 allergen was capable of stimulating T cell proliferation; Bm23-25 was also shown to induce IgE production in vitro from PBMC derived from patients with either TPE or other filarial symptoms. Bronchoalveolar lavage fluid of patients with TPE contained IgE antibodies that recognized Bm23-25 strongly, an observation suggesting that the microfilarial allergen might be involved in the pathogenesis of the TPE syndrome.
Macrophage and monocyte IL-1 beta regulation differs at multiple sites. Messenger RNA expression, translation, and post-translational processing.
Authors: Herzyk D J , Allen J N , Marsh C B , Wewers M D
Published: December 2022

Maturation of blood monocytes into macrophages is accompanied by a number of functional changes including decreased IL-1 beta release in response to LPS. This limitation has previously been ascribed to transcriptional regulation. However, in seeming conflict with the observed depression in IL-1 beta mRNA levels, recent work demonstrates increased intracellular IL-1 beta in macrophages. Therefore, the present study sought to explain these differences by comparing IL-1 beta production from autologous alveolar macrophage and blood monocyte pairs at multiple regulatory sites, including endotoxin responsiveness, mRNA expression, protein translation, and post-translational processing. Macrophages did not differ from monocytes in endotoxin sensitivity, but when analyzed by both ELISA and Western blot, were confirmed to have limitations in IL-1 beta release. Gene expression studies demonstrated that at 4 h, macrophage IL-1 beta steady state mRNA levels were 3-fold lower than the monocyte's. However, total IL-1 beta protein production, as measured by [35S]methionine labeling with immunoprecipitation, demonstrated three- to sixfold higher amounts in macrophages at comparable time points. The enhanced protein production in the face of relatively low mRNA levels suggests that macrophages translate IL-1 beta mRNA more efficiently. Furthermore, characterization of IL-1 beta release into supernatants revealed that whereas monocyte release occurred early, represented 5 to 20% of the intracellular amounts, and contained largely processed IL-1 beta, macrophage release was delayed, represented 1 to 5% of the intracellular amounts, and contained primarily unprocessed IL-1 beta. Taken together, these data demonstrate that the limitations in alveolar macrophage IL-1 beta release occur due to slower export and conversion of 35- to 17-kDa protein and are not due to differences in sensitivity to endotoxin or to transcriptional control mechanisms.
Alpha-1-antichymotrypsin inhibits the NADPH oxidase-enzyme complex in phorbol ester-stimulated neutrophil membranes.
Authors: Kilpatrick L , McCawley L , Nachiappan V , Greer W , Majumdar S , Korchak H M , Douglas S D
Published: December 2022

The generation of superoxide anion and release of granule contents are essential to the bactericidal function of neutrophils, but may also contribute to host tissue damage during inflammation. In previous studies (J. Immunol. 146:2388), we have demonstrated that the acute phase reactant alpha-1-antichymotrypsin (ACT), a potent inhibitor of the serine protease cathepsin G, also suppresses superoxide anion generation. The inhibitory effect of ACT was not directly linked to its antiproteolytic activity and may reflect interaction at a site other than its reactive loop. To further characterize the mechanism of inhibition, we investigated the direct effects of ACT on the NADPH oxidase enzyme complex and the signaling pathways that regulate motivation of the respiratory burst. We present evidence that ACT does not intefer with agonist-stimulated calcium mobilization or translocation and activity of protein kinase C. ACT was an effective inhibitor of superoxide anion generation in membrane preparations isolated from PMA-activated cells. These results support the notion that ACT is acting on a component of the active assembled NADPH oxidase complex. Thus, ACT may have an important role in regulation of specific aspects of the inflammatory processes and the modulation of toxic oxygen-based host tissue damage.
Detection of alveolar macrophage-derived IL-1 beta in asthma. Inhibition with corticosteroids.
Authors: Borish L , Mascali J J , Dishuck J , Beam W R , Martin R J , Rosenwasser L J
Published: December 2022

This study investigated alveolar macrophage function in asthma as assessed by the presence and source of the cytokine IL-1 beta. Bronchoalveolar lavage (BAL) fluid and cell pellets were obtained at 4 a.m. from a cohort of asthmatic subjects. Normal subjects were used as controls. Asthmatic BAL fluid contained 57.0 +/- 5.9 pg/ml of IL-1 beta as determined by ELISA. No IL-1 beta could be identified in the BAL fluid of the control group. This IL-1 represented synthesis by cells of the airway as assessed by the presence of IL-1 beta-specific mRNA transcripts in the BAL cell pellets. Inasmuch as IL-1 beta transcripts are known to be present for only a short time period after activation (approximately 0.5 to 16 h), the presence of transcripts represents recent cellular activation. Using the technique of in situ hybridization, IL-1 beta transcripts were found to be located exclusively within alveolar macrophages, and a mean of 40 +/- 14% of alveolar macrophage were activated at the time of the lavage. Corticosteroids are known to be efficacious in the treatment of asthma. Administration of a single 50-mg dose of prednisone at 3 p.m. resulted in diminished IL-1 beta protein concentration in the BAL fluid (28.3 +/- 5.7 pg/ml; p < 0.01 compared to baseline) and completely abrogated IL-1 beta mRNA expression. In summary, these studies demonstrate that IL-1 beta is synthesized within the airways by alveolar macrophage and suggest that IL-1 beta may be a mediator of asthma. Inhibition of IL-1 beta may be an additional mechanism for the efficacy of corticosteroids in the treatment of asthma.
Development of murine lupus in CD4-depleted NZB/NZW mice. Sustained inhibition of residual CD4+ T cells is required to suppress autoimmunity.
Authors: Connolly K , Roubinian J R , Wofsy D
Published: December 2022

Chronic administration of anti-CD4 mAb prevents autoimmune disease in NZB/NZW F1 (B/W) mice. This may be due either to CD4 cell depletion or to inhibition of CD4 cell function. To evaluate the relative importance of these mechanisms, we devised a system in which the consequences of cell depletion could be analyzed independent of the inhibitory effects of chronic mAb therapy. This was accomplished by performing adult thymectomy before mAb administration. Specifically, female B/W mice underwent thymectomy or sham thymectomy at age 6 wk, followed at age 3 mo by a short course of either anti-CD4 (2 mg/wk for 3 wk) or saline. Treatment with anti-CD4 depleted 90% of circulating CD4 cells, but a small subpopulation (10%) of CD4 cells was refractory to depletion. In non-thymectomized mice, the CD4 population gradually reconstituted after cessation of therapy. In contrast, in thymectomized mice, recovery of CD4 cells was prevented by the absence of the thymus. Despite the striking reduction in CD4 cells in thymectomized mice, severe autoimmune disease developed, with autoantibody levels, proteinuria, and mortality comparable with non-thymectomized, nondepleted controls. The unexpected development of lupus nephritis in thymectomized, CD4-depleted B/W mice suggested that the thymus might be required to achieve the benefits of therapy with anti-CD4. To exclude this possibility, we demonstrated that chronic therapy with anti-CD4 prevents autoimmunity in thymectomized B/W mice. These findings imply that: 1) substantial depletion of CD4 T cells is not sufficient to suppress autoimmunity; 2) suppression of autoimmunity requires sustained functional inhibition of CD4 T cells; and 3) a small subpopulation of CD4 cells that is refractory to depletion by anti-CD4 is sufficient to promote the full expression of murine lupus in B/W mice.
Structure and binding properties of a monoclonal anti-idiotypic autoantibody to anti-DNA with epibody activity.
Authors: Fischel R , Eilat D
Published: December 2022

We report the isolation and characterization of a mouse autoanti-idiotypic mAb (D7.4 IgG2a), which is directed against a major public Id (A52 IgG2b) in the murine and human autoimmune response to DNA. The natural anti-Id mAb has been produced in the course of the SLE-like disease in a female NZB/NZW F1 mouse and showed a dual specificity (epibody activity) for the public Id (A52) and for the autoantigen (DNA). The two binding activities were shown to reside in the Fab portion of the epibody and were highly specific for their respective Ag. A complete nucleotide sequence analysis of the D7.4 H and L chain V-region genes combined with computer comparisons to available Ig sequences may suggest a charge interaction between the H chain CDR3 segments of the Id and anti-Id antibodies. The D7.4 epibody may be a component of the self-binding, idiotypically connected network of natural antibodies. Alternatively, it could be elicited against the potentially pathogenic, DNA-containing immune complexes in order to facilitate their removal from the circulation of diseased individuals.
Novel deletion and a new missense mutation (Glu 217 Lys) at the catalytic site in two adenosine deaminase alleles of a patient with neonatal onset adenosine deaminase- severe combined immunodeficiency.
Authors: Hirschhorn R , Nicknam M N , Eng F , Yang D R , Borkowsky W
Published: December 2022

Mutations at the adenosine deaminase (ADA) locus result in a spectrum of disorders, encompassing a fulminant neonatal onset severe combined immunodeficiency (SCID) and childhood onset immunodeficiency, as well as apparently normal immune function. The extent of accumulation of the toxic metabolite, deoxyATP, correlates directly with severity of disease. We have now determined the mutations on both alleles of a child with fulminant, neonatal onset ADA- SCID and accumulation of extremely high concentrations of deoxyATP. The genotype was consistent with the severely affected phenotype. One allele carried a large deletion that arose by non-homologous recombination and included the first five exons and promoter region. The second allele carried a missense mutation (G649A) resulting in replacement of Glu217, an amino acid involved in the catalytic site, by Lys and predicting a major alteration in charge. Expression of the mutant cDNA in Cos cells confirmed that the mutation abolished enzyme activity. We have previously reported that a missense mutation at the preceding codon is similarly associated with neonatal onset ADA- SCID and accumulation of extremely high deoxyATP. These findings suggest that genotype-phenotype correlations may be apparent for ADA- SCID, despite the role that random variation in exposure to environmental pathogens may play in the initial phenotype. Such genotype-phenotype correlations may be important to consider in evaluating results of ongoing trials of "gene" and enzyme replacement therapy.
Detection of a vigorous HIV-1-specific cytotoxic T lymphocyte response in cerebrospinal fluid from infected persons with AIDS dementia complex.
Authors: Jassoy C , Johnson R P , Navia B A , Worth J , Walker B D
Published: December 2022

AIDS dementia complex is a common neurologic disorder in later stages of HIV-1 infection. Because virus-specific CTL have been shown to contribute to neurologic disease in certain viral illnesses, we examined the cerebrospinal fluid of HIV-1-infected persons with various stages of AIDS dementia complex for the presence of HIV-1-specific CTL. In five of six subjects studied, HIV-1-specific CTL were identified in the cerebrospinal fluid. These CTL were directed at epitopes within the gag, reverse transcriptase, envelope, and nef proteins and restricted by HLA class I Ag. In four of these subjects, virus-specific CTL were detected in higher numbers in the cerebrospinal fluid compared to the peripheral blood, suggesting a specific recruitment to or local induction within the nervous system. These studies demonstrate the presence of a vigorous and broadly directed CTL response to HIV-1 in the central nervous system of infected persons with AIDS dementia complex, and provide immunologic evidence of localized intrathecal infection. Although HIV-1-specific CTL may serve to inhibit viral replication in the central nervous system, the presence of a persistent CTL response in the central nervous system may also contribute to the neurologic disorders characteristic of HIV-1 infection.
Further Studies on the Influence of Heparin on Anaphylactic Shock in the Guinea Pig
Authors: Van de Carr F. Rene , Williams O. B.
Published: December 2022

In an earlier paper (1) we confirmed the observations of Kyes and Strauser (2) with regard to the inhibitory effect of heparin on the anaphylactic syndrome. In our studies guinea pigs hypersensitive to horse serum, instead of pigeons hypersensitive to sheep serum, as used by Kyes and Strauser, were employed. In view of the failure of Hanzlik, Butt and Stockton (3), of Hyde (4), and of Reed and Lamson (5) to support our findings, we present additional data which substantiate our previous findings, using as a criterion of shock the classical symptoms of anaphylaxis. We have checked the reaction by the Dale and by the bronchospasm method of Koessler and Lewis (6). In a limited number of animals we controlled the coagulation time of the blood by means of the capillary tube method prior to the injection of the shocking dose of the antigen and also at the height of shock.
The Effect of Bacteriophage upon the Phenomena of Leukocytosis and Phagocytosis
Authors: Nelson A. R.
Published: December 2022

Summary and Conclusions

The intravenous injection of anti-staphylococcus bacteriophage exerts a marked influence on the type of leukocytic response elicited by susceptible staphylococci, but has only a slight effect upon the type of response due to resistant organisms. This influence is characterized by the fact that a single injection of the bacteriophage together with susceptible organisms causes a decrease in the degree and time of appearance of the leukocytosis as compared with that due to susceptible organisms alone. Repeated injections of the bacteriophage increase the period of leukopenia and decrease the degree of leukocytosis as compared with that due to susceptible organisms alone. The bacteriophage does not alter the type of response due to resistant organisms in any way comparable to the effect of the bacteriophage on the type of leukocytic response due to susceptible organisms. When leukocytes and organisms are brought into contact with the anti-staphylococcus bacteriophage in vitro, either in the presence or absence of serum, the degree of phagocytosis in the case of susceptible organisms is markedly increased, whereas in the case of resistant staphylococci the degree of phagocytosis is practically unaffected. When anti-staphylococcus bacteriophage is injected intravenously there is an almost immediate increase in the phagocytic power of the leukocytes for susceptible organisms, whereas for resistant organisms the phagocytic power remains practically unaltered. The opsonic power is not altered appreciably in either case.
Anaphylaxis
Authors: Sherwood Noble P.
Published: December 2022

Since the work of Gay and Southard (1) in 1907 a number of papers have appeared describing histological and physiological changes in intact and isolated hearts of the guinea pig, dog, cat, rabbit and frog. Since the completion of this work Gahringer (2) reported on clinical anaphylaxis in pigeons and more recently Hanzlik (3) has reported on the “crop reaction” in pigeons. So far as I know there has been no work reported on sensitization phenomena in the embryonic chick. This would seem desirable because it would afford an opportunity to study sensitization phenomena in an intact living organism before the heart had connections with the central nervous system and before even the anlage of the liver and other organs were in evidence, providing sensitization could be established. It would also permit of the study of such phenomena in an exceedingly simple isolated heart whose functional disturbances as recently pointed out by Lewis (4) are quite comparable to those of the adult mammalian heart.
Further Observations Concerning the Sensitization of Tuberculous Guinea Pigs
Authors: Dienes L.
Published: December 2022

In a former paper (1) we stated that in the study of sensitization of tuberculous guinea pigs we met with the difficulty that the procedure which in the majority of animals gave strong sensitization from August until November 1926, in the following months only rarely gave positive results. With the timothy pollen we were unable to reproduce the strong sensitization which we observed in one guinea pig in November. For awhile we concentrated our efforts on the timothy pollen. After unsuccessful trials with small and large doses injected subcutaneously over long periods we have been led to try to inject the pollen and the tubercle bacillus together, and then to inject the pollen in the tuberculous lesions. One out of three guinea pigs treated in the last mentioned way developed strong sensitiveness.
Studies on the Antigenic Properties of the Ultraviruses
Authors: Schultz E. W. , Bullock L. T. , Lawrence F.
Published: December 2022

No evidence of specific complement fixation could be demonstrated when immune sera, produced in rabbits with pure vaccinia brain-virus, were tested against antigens prepared from the lesions of cutaneous vaccinia or against brain-virus antigens. No specific complement fixation occurred when immune sera, produced with cutaneous vaccinia virus, were tested against antigens prepared from brain-virus. Although the latter sera fixed complement in the presence of antigens prepared from cutaneous vaccinia pulp, an almost equivalent degree of complement fixation was observed in the presence of antigens prepared from bacteria cultured from this pulp. Antisera produced against bacteria, harvested from the pulp of cutaneous vaccinia lesions, fixed complement, in the presence of antigens prepared from cutaneous vaccinia lesions. No evidence for the presence of specific precipitins in the two kinds of vaccinia immune sera could be elicited. Precipitates, which did occur in certain cases, were shown to be due to unspecific reactions. Virulicidal antibodies were easily demonstrable in all the immune sera examined for this property.
Susceptibility of Eskimos to the Common Cold and a Study of Their Natural Immunity to Diphtheria, Scarlet Fever and Bacterial Filtrates
Authors: Heinbecker Peter , Irvine-Jones Edith I. M.
Published: December 2022

Eskimos are very susceptible to upper respiratory infections on contact with the outside world. Ordinary bacterial infections rarely occur. Diphtheria and scarlet fever are unknown clinically and in a group of about fifty subjects all were negative to the Dick test and all the adults were also Schick negative. Children up to the age of twelve years were invariably Schick positive.

Three serums were found to contain antitoxin both for diphtheria and scarlet fever. It is therefore concluded that the immunity to the disease and the negative skin tests depend on the presence of antitoxin. This is interpreted as being due to a natural hereditary immunity dependent upon some non-specific antitoxic mechanism. Skin reactions with filtrates of streptococci isolated from cases of rheumatic fever were mildly positive in a small percentage of cases. Neutralizing antitoxin was demonstrated in all three sera but it was not invariably present for all three toxins. The Eskimos showed a high percentage of positive reactions when tested with a staphylococcus aureus filtrate.
Studies on the Antigenic Properties of the Ultraviruses
Authors: Schultz E. W. , Hoyt J.
Published: December 2022

No evidence of specific complement fixing antibodies against the virus of herpes was found in the serum of rabbits which had been immunized by different routes with virus-brain suspensions. These immune sera also failed to present any evidence of specific precipitating antibodies. Virulicidal antibodies were demonstrable in all of the immune sera tested for this property.
Methods of Obtaining and Preserving Antibodies
Authors: Darányi J.
Published: December 2022

Whereas in the treatment of animals with bacteria or toxin-antigens a certain poisoning results from the first injection, the danger of the treatment with protein-antigens (e.g., blood corpuscles and heterologous sera) lies in the anaphylactic shock, which usually occurs after the second or third injection. It is well known how difficult it is to produce efficient antibodies against protein-antigens on account of the hypersensitiveness. The danger of anaphylaxis increases especially when we endeavor to immunize to a high degree and in this way collect antibodies in the animal blood. The best method of antibody production is the intravenous application of the antigens; however, this form of immunizing causes the greatest number of anaphylactic deaths. Subcutaneous or intraperitoneal injections do not give sufficient stimulation for antibody formation. The acknowledged best antibody producers are rabbits, in the serum of which we can collect hemolysins, agglutinins, precipitins, etc., much better than in the larger animals.
Quantitative Studies on the Action of Compound Hemolysins
Authors: Gahl Rudolf
Published: December 2022

A method of plotting hemolytic data was developed and it was shown that curves representing the same percentage of hemolysis are parallel, as long as the concentration of blood cells in the mixture is not changed, and can be made to coincide by a shift of one or both coördinate axes. The shape of the curves is, therefore, independent of the concentration or titer of amboceptor and complement serum. This conclusion was derived under certain assumptions, the principal one being that the components are individual substances and not mixtures of two or three. A comparison of the shape of such “characteristics” should, therefore, permit conclusions as to the purity of the components used for the tests.
Cell Antigens and Individual Specificity
Authors: Landsteiner Karl
Published: December 2022

The general aspects of immunological specificity have been discussed before this society in a masterly manner by authorities like Hektoen and Wells, on former similar occasions. If the following review concerns itself again with related matters, this may be justified by the broadening of the subject of serological specificity which has branched off from medical immunology as a somewhat independent line of research.

The interest in the particular topic of blood antigens has been revived of late by investigations centering round several distinct issues, such as the principles of species specificity, the chemical nature of cell antigens and the human blood groups. Beginning with the blood groups, one of the questions which has arisen is whether the differences of human blood are limited to the four groups or whether there are other variations.
Cytokine synthesis by intestinal intraepithelial lymphocytes. Both gamma/delta T cell receptor-positive and alpha/beta T cell receptor-positive T cells in the G1 phase of cell cycle produce IFN-gamma and IL-5.
Authors: Yamamoto M , Fujihashi K , Beagley K W , McGhee J R , Kiyono H
Published: December 2022

Murine intestinal intraepithelial lymphocytes (IEL) have been shown to contain subsets of alpha/beta TCR+ and gamma/delta TCR+ T cells that spontaneously produce cytokines such as IFN-gamma and IL-5. We have now determined the nature and cell cycle stage of these cytokine-producing T lymphocytes in EIL by using IFN-gamma- and IL-5-specific ELISPOT assay, cytokine-specific mRNA-cDNA dot-blot hybridization and polymerase chain reaction, and flow cytometry (FACS) for DNA analysis. When CD3+ T cells from IEL of normal C3H/HeN mice were separated into low and high density fractions by discontinuous Percoll gradients, IFN-gamma and IL-5 spot-forming cells were only found in the former population. Analysis of mRNA for these cytokines by both IFN-gamma- and IL-5-specific dot-blot hybridization and polymerase chain reaction revealed that higher levels of message for IFN-gamma and IL-5 were also seen in the low density fraction. However, cell cycle analysis of these two fractions by FACS using propidium iodide showed a similar pattern of cell cycle stages in both low and high density populations (G0 + G1 approximately 96 to 98% and S/G2 + M approximately 2 to 4%). Finally, mRNA from gamma/delta TCR+ and alpha/beta TCR+ T cells in both low and high density fractions of IEL were analyzed for IFN-gamma and IL-5 message by polymerase chain reaction. After 35 cycles of amplification, both gamma/delta TCR+ and alpha/beta TCR+ T cells in the low density fraction expressed higher levels of message for these two cytokines when compared with the high density population. These results have now shown that both gamma/delta and alpha/beta TCR+ IEL can be separated into low and high density subsets and both fractions possess a similar stage of cell cycle. However, only the low density cells (in G1 phase) of both gamma/delta and alpha/beta TCR types possess increased cytokine-specific mRNA and produce the cytokines IFN-gamma and IL-5. Our results suggest that alpha/beta TCR+ and gamma/delta TCR+ IEL can produce cytokines without cell proliferation.
Expression pattern of the most JH-proximal human VH gene segment (VH6) in the B cell and antibody repertoire suggests a role of VH6-encoded IgM antibodies in early ontogeny.
Authors: Van Es J H , Raaphorst F M , van Tol M J , Meyling F H , Logtenberg T
Published: December 2022

We have developed a mAb (JE-6) that recognizes an Id encoded by the most JH-proximal human VH gene segment (VH6) in or near germ-line configuration. This mAb was used to determine the frequency of Id JE6+ B cells in large collections of monoclonal EBV-transformed and short term B cell lines derived from fetal, neonatal, and adult lymphoid tissues. Moreover, we investigated the presence of Id JE-6+ Ig in sera from neonates and adults and determined the (auto)antigen binding properties of VH6-encoded IgM mAb. We detected a fivefold overrepresentation of VH6-expressing IgM producing B cells in fetal tissues, cord blood, and adult bone marrow relative to adult blood. In cord blood, but not in adult blood sera, germ-line VH6-encoded IgM molecules were readily detectable. IgM secreted by VH6-expressing B cell clones displayed highly conserved and virtually identical autoantigen binding properties, independent of the length and composition of the IgH chain CDR3 region and L chain isotype. Collectively, these results suggest that the VH6 gene and the antibodies it encodes play an important role in early human ontogeny.
Allergen-specific modulation of cytokine synthesis patterns and IgE responses in vivo with chemically modified allergen.
Authors: Gieni R S , Yang X , HayGlass K T
Published: December 2022

Hypersensitivity and IgE synthesis are highly dependent on the balance in which production of IL-4 and IFN-gamma is induced. An immunologic approach that alters the dominant pattern of cytokine synthesis and antibody production that is elicited after exposure to native allergen is described. High M(r), glutaraldehyde-polymerized OVA administered (i.p.) before or after immunization with unmodified OVA induces > or = 95% inhibition of specific IgE synthesis concomitant with 300- to 800-fold increases in IgG2a production in C57BL/6 mice. These changes result from a genetically controlled shift in the pattern of cytokine production within the allergen-specific T cell repertoire as demonstrated by i) susceptibility of the changes induced upon administration of modified allergen to in vivo treatment with anti-IFN-gamma mAb and ii) a 5- to 7-fold increase in the ratio of IFN-gamma:IL-4 synthesis after overnight culture directly ex vivo. This system should prove useful in identification of the factors which are influential in the commitment of T cells to Th1- or Th2-like patterns of cytokine synthesis. Moreover, as defective induction of IFN-gamma by allergen-specific T cells appears to play a role in elevated IgE synthesis and human allergy, this approach may have therapeutic potential.
Fresh and cultured Langerhans cells display differential capacities to activate hapten-specific T cells.
Authors: Dai R , Grammer S F , Streilein J W
Published: December 2022

Langerhans cells (LC) that have been cultured for 3 days acquire potent T cell-activating properties when compared to freshly prepared, uncultured LC. By contrast, fresh LC are superior to cultured LC in the ability to process native protein Ag. To define further the disparate functional properties of these epidermally derived APC, freshly isolated and cultured epidermal cells (EC) enriched for LC were prepared from BALB/c mice. Highly purified T cells from naive mice, and from mice sensitized epicutaneously with dinitrofluorobenzene, have been examined for their capacity to respond to fresh and cultured EC; 1) in the presence of staphylococcal enterotoxin B; and 2) after the EC had been derivatized with dinitrofluorobenzene. Both fresh and cultured EC activated syngeneic T cells in the presence of staphylococcal enterotoxin B, and fresh and cultured DNP-derivatized EC induced proliferation among DNP-specific T cells. Only cultured, hapten-derivatized EC were able to activate unprimed syngeneic T cells in vitro, and these cells responded as though "primed" when re-exposed to DNP-derivatized spleen cells in secondary cultures. In addition, naive lymphocytes that were activated by cultured DNP-EC were able to evoke local contact hypersensitivity reactions when injected into the pinnae of naive mice that were then painted with dinitrofluorobenzene. By contrast, naive syngeneic T cells exposed to fresh DNP-EC neither proliferated nor differentiated into effector cells. We conclude that fresh LC can constitutively activate primed, but not unprimed, hapten-specific T cells, whereas cultured LC readily both primed and unprimed T cells. The capacity of hapten-derivatized cultured EC to convert naive, hapten-specific T cells into cells that mediate contact hypersensitivity supports the proposal that cultured LC are the functional equivalents of epidermal LC that have migrated to draining lymph nodes. The ability of hapten-derivatized fresh LC to activate primed, hapten-specific T cells is consistent with the view that fresh LC are functionally equivalent to LC within the epidermis.
Delimitation of the proliferative stages in the human thymus indicates that cell expansion occurs before the expression of CD3 (T cell receptor).
Authors: Alvarez-Vallina L , González A , Gambón F , Kreisler M , Díaz-Espada F
Published: December 2022

Precursor of T lymphocytes undergo proliferation and maturation under the influence of the thymic microenvironment. In our study, we have attempted to determine the distribution of human postnatal thymocytes in division according to their stage of differentiation. Our data show that about 11.5% of all thymic cells are in S/G2/M phases, and that a subset of the cortical and precortical subpopulations contains most of the dividing cells. Rate of cell division is maintained at high levels from the prethymocyte precursor along the successive stages of differentiation represented by CD1+CD3-CD4-CD8- and CD1+CD3-CD4+CD8- cells. The percentage of dividing cells is maximal in an intermediate subset of CD1+CD3-CD4+CD8-CD45RO+ cells defined by the distinct expression of class I HLAdim/high molecules, which could contain cells in transit from prethymocytes to double-positive cortical cells. The CD3- fraction of the double-positive cortical cells contains most of the dividing thymocytes, although the rate of division within this subset is much less than that of the precursor CD1+CD3-CD4+CD8- cells. In a linear scheme of differentiation, cell division stops at or near the point of initiation of CD3 expression. These results suggest that in human thymus cell expansion takes place before the initiation of the positive selection process. According to this view the stringency of the selection process would require the previous generation of a large number of precursors to permit the production of sufficient numbers of mature T cells.
Disulfide linkage between C3b and tetanus toxin on tetanus toxin-specific EBV-transformed B cells.
Authors: Jacquier M R , Gabert F M , Villiers C L , Colomb M G
Published: December 2022

EBV-transformed B cells specific for tetanus toxin were found to bind C3b in excess over the expected figures based on the number of complement receptors CR1. This was confirmed by analysis of cell extracts by SDS-PAGE giving evidence for C3b-membrane protein complexes that were disrupted upon reduction. Alkylation of C3b-free cysteine abolished formation of these complexes and only a noncovalent binding of C3b to CR1 was observed, which could be inhibited by mAb to CR1. When C3b was incubated with the same cells coated with tetanus toxin bound to their specific membrane Ig, preferential formation of disulfide-bonded complexes between tetanus toxin and C3b was observed. These observations correspond to a novel capacity of C3b to interact covalently through its cysteine 1010 with free SH groups of protein acceptors. One hypothesis is that the disulfide bond formation is catalyzed by a thioredoxin-like protein secreted and expressed on the membrane of EBV-transformed B cells. In the context of Ag processing and presentation by B cells, disulfide binding of chaperone C3b to Ag is likely to persist during transcytosis and to play a significant role in the modulation of the processing.
Bone marrow-derived dendritic epidermal T cells express T cell receptor-alpha beta/CD3 and CD8. Evidence for their extrathymic maturation.
Authors: Shiohara T , Moriya N , Hayakawa J , Arahari K , Yagita H , Nagashima M , Ishikawa H
Published: December 2022

The majority of murine Thy-1+ dendritic epidermal T cells (DETC) express virtually identical TCR encoded by V gamma 5 and V delta 1 genes and are derived from early fetal thymic precursors. However, not consistent with this notion is an early finding that DETC arise continuously from bone marrow (BM) precursors by a thymus-independent mechanism. To address this issue, donor-type DETC were characterized in lethally irradiated mice that were reconstituted by Thy-1-disparate BM cells with or without a thymus. The BM-derived DETC, unlike their normal TCR-gamma delta counterparts, were found to express the TCR-alpha beta/CD3 complex and CD8, and their migration to the epidermis dermis occurred independently of the thymus. The numbers of the BM-derived DETC increased with time and reached a plateau 6 mo after BM transfer, at which time the TCR-alpha beta/CD3 complex was expressed on a small fraction of the DETC in athymic BM chimeras. Although no further increase in the number was observed at later times, at 1 yr after transfer most of the BM-derived DETC came to express the TCR-alpha beta/CD3 complex in the absence of thymic influence. By contrast, most of BM-derived T cells in other lymphoid organs from athymic BM chimeras still failed to express the TCR-alpha beta/CD3 complex even at 1 yr after transfer. These results suggest that extrathymic differentiation of BM-derived DETC could occur with the epidermal microenvironment.
Role of CD8 in staphylococcal enterotoxin B-mediated lysis by cytotoxic T lymphocytes.
Authors: Hoo W S , Kranz D M
Published: December 2022

Recent evidence has suggested that recognition of superantigens such as the staphylococcal enterotoxins by CTL occurs independently of the accessory molecule CD8. These conclusions are based on the observation that antibodies to CD8 do not appear to be effective inhibitors of T cell lysis that is mediated by enterotoxin. This is in contrast to the well-known inhibitory effects of anti-CD8 antibodies on T cell activation by most peptide/class I complexes. In this study, we show that lysis of staphylococcus enterotoxin B (SEB)-bearing target cells by the mouse alloreactive CTL clone 2C is inhibited by anti-CD8 antibodies. SEB-mediated lysis by a polyclonal population of mouse CTL was also inhibited by anti-CD8 antibodies, but only under conditions where the SEB concentration is low. Inhibition occurs even when class I negative Daudi cells are used as targets. Thus, the observed inhibition does not appear to be due to the prevention of intercellular interactions between CD8 and class I molecules but is probably a consequence of preventing the intracellular association of CD8 and TCR. At the high ligand densities used in most previous studies, very few of the CD8/TCR complexes may be required for activation. Under these conditions, lysis may appear to be CD8 "independent" because 1) there are a sufficient number of preexisting CD8/TCR complexes for activation; or 2) prohibitively high concentrations of antibody would be needed to saturate unassociated CD8.
Ca2+ ionophore A23187-dependent stabilization of granulocyte-macrophage colony-stimulating factor messenger RNA in murine thymoma EL-4 cells is mediated through two distinct regions in the 3'-untranslated region.
Authors: Iwai Y , Akahane K , Pluznik D H , Cohen R B
Published: December 2022

We analyze the role of the Ca2+ ionophore A23187 in the induction of GM-CSF mRNA expression in EL-4 thymoma cells. Northern analysis shows that A23187 increases the half-life of GM-CSF mRNA. To identify potential Ca2+ response elements in the GM-CSF mRNA, we produced stable transfectants containing pRSV-CAT (EL-4cat) or hybrid constructs in which most of the GM-CSF 3'-untranslated region (EL-4gm) or the adenosine-uridine boxes alone (EL-4au) were placed in a downstream position from the CAT coding region. A23187 induces a 4.4-fold increase in CAT activity in EL-4cat cells and a 210-fold and 48-fold increase in CAT activity in EL-4gm and EL-4au cells, respectively. Actinomycin D chase experiments in transfected cells demonstrate that A23187 increases the half-life of CAT mRNA from 15 min to 3 h in EL-4au cells and more than 3 h in EL-4gm cells, suggesting that the effect of Ca2+ is mediated predominantly by the adenosine-uridine boxes with a smaller contribution from upstream regions. To map these upstream regions, we transfected cells with constructs containing mutations of the 3'-untranslated region. With two of these mutations, corresponding to a region located about 160 bases upstream of the adenosine-uridine boxes, CAT activity was induced only 50-fold compared to 200-fold in EL-4gm cells. These data indicate that two regions within the GM-CSF 3'-untranslated region interact to modulate Ca2+ effects on GM-CSF mRNA half-life.
Different effects of substitutions at residues 224 and 228 of MHC class I on the recognition of CD8.
Authors: Sekimata M , Tanabe M , Sarai A , Yamamoto J , Kariyone A , Nakauchi H , Egawa K , Takiguchi M
Published: December 2022

Previous studies indicated that weak xenoresponse to HLA class I by mouse T cells is due to the inefficient interaction of mouse CD8 with the alpha 3 domain of HLA class I. The present study using chimeric H-2Kb molecules with recombinant alpha 3 domain between H-2Kb and HLA-B7 as well as single amino acid mutants of H-2Kb demonstrated that each substitution at residues 224 and 228 affects recognition of CD8-dependent mouse CTL clones. On the other hand, reactivity of IL-2-producing H-2Kb-specific T cell hybridoma transfected with mouse CD8 alpha was abrogated by substitution at residue 224 but not by that at residue 228. This indicates that the substitution at residue 228 affects recognition of CD8-dependent CTL but does not critically affect binding of CD8 to MHC class I molecules, although residue 224 abrogates binding of CD8. The model structure of the alpha 3 domain of H-2Kb suggests that the substitution at residue 224 induces conformational change of CD8 binding loop, whereas minimum structure change by the substitution at residue 228 is expected. It is therefore speculated that minimum structure change of CD8 binding loop by substitution at residue 228 may influence binding affinity of CD8, which abrogates recognition of CD8-dependent CTL but not IL-2 production of the CD8-dependent T cell hybridoma.
Human monoclonal antibody HA-1A binds to endotoxin via an epitope in the lipid A domain of lipopolysaccharide.
Authors: Bogard W C , Siegel S A , Leone A O , Damiano E , Shealy D J , Ely T M , Frederick B , Mascelli M A , Siegel R C , Machielse B
Published: December 2022

HA-1A, a human IgM mAb, has been shown to significantly reduce mortality in septic patients with Gram-negative bacteremia, especially those with septic shock, in a controlled clinical trial. To confirm the reported specificity of this antibody for the lipid A domain of endotoxin, several assay systems were developed. These assay systems included an ELISA, which measured the binding of HA-1A to lipid A adsorbed to a solid phase; a rate nephelometry assay, which measured the ability of HA-1A to bind and aggregate lipid A in solution; and a dot-blot immunoassay, which measured the ability of HA-1A to interact with lipid A adsorbed to Immobilon-P. In all three assay systems, HA-1A bound in a dose-dependent manner to lipid A prepared from Salmonella minnesota R595 LPS, whereas negative control human IgM mAb or polyclonal antibodies did not. Several experimental approaches were employed to demonstrate the specificity of HA-1A in these assay systems. Both polymyxin B and murine IgG mAb (8A1) with a specificity for lipid A were able to competitively inhibit HA-1A reactivity with lipid A in a dose-dependent manner. Furthermore, a murine IgG anti-Id mAb (9B5.5) developed against HA-1A was also able to block the binding of HA-1A to lipid A in these assay formats. HA-1A reactivity with synthetic lipid A confirmed that HA-1A binding to the natural lipid A was not the result of contaminants in the latter. Finally, the reactivity of HA-1A against a variety of glucosamine-containing and fatty acid-containing compounds was assessed. Some weak interaction was seen with cardiolipin and chitin, but not with serum proteins, lipoteichoic acid, or DNA. Collectively, these results conclusively establish that HA-1A binds to the lipid A region of LPS by an interaction with the V region of the antibody.