Claire-Michèle Farber

B cell–helper neutrophils stimulate the diversification and production of immunoglobulin in the marginal zone of the spleen

Neutrophils utilize immunoglobulins (Igs) to clear antigen, but their role in Ig production is unknown. Here we identified neutrophils around the marginal zone (MZ) of the spleen, a B cell area specialized in T-independent Ig responses to circulating antigen. Neutrophils colonized peri-MZ areas after post-natal mucosal colonization by microbes and enhanced their B-helper function upon receiving reprogramming signals from splenic sinusoidal endothelial cells, including interleukin 10 (IL-10). Splenic neutrophils induced Ig class switching, somatic hypermutation and antibody production by activating MZ B cells through a mechanism involving the cytokines BAFF, APRIL and IL-21. Neutropenic patients had fewer and hypomutated MZ B cells and less preimmune Igs to T-independent antigens, which indicates that neutrophils generate an innate layer of antimicrobial Ig defense by interacting with MZ B cells.Neutrophils use immunoglobulins to clear antigen, but their role in immunoglobulin production is unknown. Here we identified neutrophils around the marginal zone (MZ) of the spleen, a B cell area specialized in T cell–independent immunoglobulin responses to circulating antigen. Neutrophils colonized peri-MZ areas after postnatal mucosal colonization by microbes and enhanced their B cell–helper function after receiving reprogramming signals, including interleukin 10 (IL-10), from splenic sinusoidal endothelial cells. Splenic neutrophils induced immunoglobulin class switching, somatic hypermutation and antibody production by activating MZ B cells through a mechanism that involved the cytokines BAFF, APRIL and IL-21. Neutropenic patients had fewer and hypomutated MZ B cells and a lower abundance of preimmune immunoglobulins to T cell–independent antigens, which indicates that neutrophils generate an innate layer of antimicrobial immunoglobulin defense by interacting with MZ B cells.

Iron as a potential co-factor in the pathogenesis of Kaposi's sarcoma?

The role of iron in the pathogenesis of several tumours is being increasingly investigated. In particular, its involvement in the pathogenesis of Kaposis sarcoma (KS) is suggested by the distribution of the endemic form of KS corresponding to continental rifts and associated iron‐oxide‐rich volcanic clays. We investigated in vitro to what extent iron supplementation or withdrawal could affect the growth of KS‐derived cells, by analysing the effects of adding iron salts (iron chloride and ferric nitrilotriacetate) and/or reducing iron by iron chelators (desferrioxamine) on KS‐derived cell cultures. The addition of iron salts strongly stimulated the growth of KS cells, as reflected by increase in thymidine incorporation and cell number. Conversely, desferrioxamine and deferiprone inhibited cell growth. The inhibitory effect of iron chelation was more pronounced on rapidly dividing basic fibroblast‐growth‐factor‐stimulated cells. These results may point to a novel therapeutic approach to KS. Int. J. Cancer 78:720–726, 1998.

Contribution of the polymerase chain reaction to the diagnosis of tuberculous infections in children

AbstractThe purpose of the study was to evaluate the contribution of polymerase chain reaction (PCR) to the diagnosis of tuberculous infection in children. Two different PCR techniques were compared to the standard bacteriological methods for the detection ofMycobacterium tuberculosis in 157 specimens obtained from the respiratory system of 51 children. Patients were classified in three groups: 12 patients with active disease (57 specimens), 12 patients with silent tuberculous infection (23 specimens) and 27 patients without tuberculosis (77 specimens). One PCR method (PCR/Ag85) used amplification of a fragment of the genes coding for the mycobacterial antigen 85 followed by hybridization of a probe specific forM. tuberculosis on the Southern blot of amplified DNA. The other PCR technique was a nested PCR (NPCR) using double amplification of a fragment of the insertion element IS6110 only present in theM. tuberculosis genome. The sensitivities of the different techniques, compared to the clinical diagnosis, were 7.0% for acid fast staining, 22.8% for culture, 24.6% for PCR/Ag85 and 44.9% for NPCR in active disease, 4.3% for culture, 8.7% for PCR/Ag85 and 28.6% for NPCR in silent tuberculous infection. The specificities were 100% for culture, 94.8% for PCR/Ag85 and 87.9% for NPCR. Among the 12 children clinically considered as having active tuberculosis, 1 had smear positive samples, 4 had at least one positive culture, 7 at least one positive PCR/Ag85 and 9 at least one NPCR positive sample. Among the 12 children having silent tuberculous infection, none had positive smears, 1 had one positive culture, 2 had at least one positive PCR/Ag85 and 5 at least one NPCR positive sample.ConclusionOur study suggests that both PCR techniques, and especially NPCR, are able to detectM. tuberculosis DNA in specimens containing few micro-organisms. PCR methods are more sensitive than culture and the results are available more quickly. Testing multiple samples from the same individual increased the sensitivity. In view of occasional false-positive results, cultures remain the gold standard to establish definitive diagnosis of primary tuberculous infection in children.

Immunologic and virologic response after tetanus toxoid booster among HIV-1- and HIV-2-infected Senegalese individuals

Twelve HIV-1-infected, nine HIV-2-infected patients and eight HIV-negative subjects were given a 40IU booster dose of tetanus toxoid (TT). Blood was collected on days 0, 7 and 30 after immunization. Changes in HIV-1 or HIV-2 RNA load were evaluated by nested PCR. TT-IgG antibody levels were quantified by ELISA. CD4 cell counts as well as activation, memory and maturation markers of T lymphocyte subsets were determined by flow cytometry. The induction of apoptosis was investigated using 7-aminoactinomycin D (AAD) and propidium iodide (PI) staining. Proliferative responses to TT and pokeweed mitogen (PWM) were determined by the level of [(3)H] thymidine incorporation. Seven and 30 days after immunization, there was no detectable increase in HIV-1 or HIV-2 plasma load. There were also no changes in CD4 cell counts, CD69, HLA-DR and memory CD45RO or naive CD45RA antigens. Immunization did not increase the spontaneous apoptosis of peripheral blood mononuclear cells (PBMCs), CD4+ and CD8+ T cells subsets neither in controls nor in HIV-infected patients. Similarly, apoptosis induced in vitro by PWM or by the specific TT recall antigen did not vary during the study period. The proliferative response to PWM and to the TT recall antigen was decreased both in HIV-1- and HIV-2-infected patients compared to HIV-negative controls. Immunization significantly increased the TT-IgG levels in healthy controls and in HIV-infected patients. However, the anti-TT-IgG response, as measured by the fold-increase index between days 0 and 30, was significantly higher in healthy controls than in HIV-1- (P=0.036) and HIV-2-infected patients (P=0.003). In conclusion, we found no deleterious immunologic or virologic effect was detected in healthy HIV-1- and HIV-2-infected individuals after antigenic challenge with a TT booster. However, the response to TT vaccination was lower in HIV-1- and in HIV-2-infected individuals than in healthy HIV-negative controls.

A multidot immunobinding assay for the serodiagnosis of tuberculosis: Comparison with an enzyme-linked immunosorbent assay

A simple dot immunobinding (dot blot) assay procedure has been developed for the detection of antibodies directed against a soluble mycobacterial antigen preparation. This technique was compared with the widely used ELISA, in a study of samples from tuberculous patients. Dot blots were read on a densitometer. The correlation between both assays was excellent (r = 0.91; P less than 0.001); 90% of sera from tuberculous patients were detected using both techniques and a serial two-fold dilution method. Assessments of the end-points of titration curves by reflectometry and simple visual interpretation gave similar results. The dot blot assay is easier to perform and appears to be a practical alternative to ELISA for the detection of anti-mycobacterial antibodies in tuberculous patients.

Local anti-P32 humoral response in tuberculous meningitis

We report five cases of severe pulmonary tuberculosis admitted to hospital with a suspicion of meningeal involvement. The diagnosis of tuberculous meningitis was confirmed by standard bacteriological techniques in two of the five patients. Specific IgG class antibodies directed against the recently purified BCG antigen P32 were detected by a dot immunoblotting technique in the serum and in the cerebrospinal fluid of each patient; however, a higher anti-P32 immunoglobulins/total immunoglobulins ratio was observed in the cerebrospinal fluid of patients with tuberculous meningitis than in their serum while the reverse situation was observed in the other patients.

Assay of specific antibody response to mycobacterial antigen for the diagnosis of a pleural effusion in a patient with aids

A diagnosis of mycobacterial infection was supported by a serological assay in a patient with AIDS. Specific antibody levels were not above the threshold of positivity determined in non-immunodeficient patients, but sera obtained previously were available, and a significant rise in titre was observed.

T Cell Reactivity against Mycolyl Transferase Antigen 85 of M. tuberculosis in HIV-TB Coinfected Subjects and in AIDS Patients Suffering from Tuberculosis and Nontuberculous Mycobacterial Infections

The mycolyl transferase antigen 85 complex is a major secreted protein family from mycobacterial culture filtrate, demonstrating powerful T cell stimulatory properties in most HIV-negative, tuberculin-positive volunteers with latent M.tuberculosis infection and only weak responses in HIV-negative tuberculosis patients. Here, we have analyzed T cell reactivity against PPD and Ag85 in HIV-infected individuals, without or with clinical symptoms of tuberculosis, and in AIDS patients with disease caused by nontuberculous mycobacteria. Whereas responses to PPD were not significantly different in HIV-negative and HIV-positive tuberculin-positive volunteers, responses to Ag85 were significantly decreased in the HIV-positive (CDC-A and CDC-B) group. Tuberculosis patients demonstrated low T cell reactivity against Ag85, irrespective of HIV infection, and finally AIDS patients suffering from NTM infections were completely nonreactive to Ag85. A one-year follow-up of twelve HIV-positive tuberculin-positive individuals indicated a decreased reactivity against Ag85 in patients developing clinical tuberculosis, highlighting the protective potential of this antigen.

CHEMOTHERAPY OF ACUTE MYELOBLASTIC LEUKEMIA IN AN HIV CARRIER

To the Editor: Myelodysplasia is known to occur with an increased frequency in HIV + patients; lymphoproliferative neoplasms mostly of the B-cell lineage have also been frequently described (1). However, myeloblastic leukemias are not frequently described in HIVinfected patients we found 2 cases in the literature (2, 3), neither addressing the problem of treatment of those malignancies in this particular setting. Our patient is a 53-yr-old white male homosexual, known to have anti-HIV antibodies since November 1989. Regular checkups since were unremarkable, except for a CD4 count consistently above lOOO/ mm3, the presence of HIV antibodies on Western blot; clinical status was perfect. On 8/10/92 he still had no complaints, physical examination showed a consistently obese abdomen; spleen was not palpated. White blood cell count was 80000/mm3 with 3000 platelets/mm3 and 3% neutrophils. The 5 1 % blasts were myeloperoxidaseand Sudan black-positive. Positive findings included a 15 cm x 12 cm x 6 m spleen on echography and 7 decayed teeth. Serum lysozyme was 16 pg/mm3 (upper limit of normal 13 pg/mm3). The patient was well and apyretic. Bone marrow examination showed numerous cells with 5 1.2% myeloblasts (peroxidaseand Sudan black-positive, CD34 + , CD 13 + and CD 15 + ). There were 35 % eosinophils. Megakaryocytes were absent. Karyotype the blast cells showed was 46 XY with inv (16) (p13q22) typical of FAB ANLL M4eo (3). Chemotherapy was instituted consisting of idarubicin 12 mg/m2 on d 1 to 3, and cytarabine 200 mg/ m’ on d 1 to 7; blasts were undetectable in peripheral blood by d 5 ; aplasia also occurred. The patient was discharged after 3 wk; 1 wk later, consolidation was given (idarubicin 12mg/m2 d 1, cytarabine 200 mg/m2 d 1 to 7). Bone marrow control showed no blasts 3 wk later (on 17/12/92). No further chemotherapy was administered, monthly follow-up visits were satisfactory; CD4 levels were 1000/mm3 on 13/4/93. To our knowledge this is the first report of successful treatment of myeloblastic leukemia in a carrier of HIV. One patient with a hybrid leukemia (B-cell ALL and FAB M4 ANLL) has been described who died before any attempt at treatment (2). We believe the FAB M4 ANLL presented by our patient has no direct connection with his HIV status. Of interest is the fact that p24 was never isolated in the patient’s serum, nor were infectious viral particles detected in coculture assays (patient’s mononuclear cells cultured with blood donor lymphocytes). Chemotherapy thus did not seem to enhance HIV infection; since idarubicin as well as cytarabine (which is a pyrimidine analogue) inhibit DNA synthesis, we cannot exclude that viral replication was suppressed as well.

Surveillance case definition for AIDS in resource-poor countries

A reply is offered to an article by De Cock and colleagues proposing a new AIDS surveillance case definition which requires serological HIV for use in resource-poor countries. The proposed definition was evaluated on 423 consecutive HIV seropositive patients at 3 HIV treatment centers in Belgium using a standard questionnaire. All patients were examined and CD4 lymphocyte counts were done. The study population consisted of 329 males and 86 females. 192 patients acquired HIV infection through homosexual contact 187 through heterosexual contact 8 through IV drug use 6 through blood transfusion and 22 through unknown routes. 114 had WHO clinical stage I disease 119 stage II 93 stage III and 89 stage IV. Opportunistic infections included: Pneumocystis carinii pneumonia (21 patients) Kaposis sarcoma (18) candida esophagitis (13) Mycobacterium tuberculosis infection (15) cryptococcal meningitis (9) cerebral toxoplasmosis (9) chronic Herpes simplex infection (6) M avium complex infection (3) and cytomegalovirus infection (1). A CD4 lymphocyte count under 200/mcl was used for advanced HIV disease. The sensitivity of the proposed AIDS case definition was slightly lower than that of the WHO/CDC 1987 AIDS case definition but much higher than that of the clinical WHO AIDS case definition. M tuberculosis infection (included in the newly proposed AIDS case definition) is much more prevalent in resource-poor countries than in Europe thus the sensitivity of this definition may be higher in those countries. The proposed AIDS case definition will have a much higher specificity and positive predictive value than the clinical WHO AIDS case definition when evaluating HIV seropositive and seronegative patients. In resource-poor countries the proposed AIDS case definition should be preferred when an HIV test result is available but AIDS case surveillance with a clinical AIDS case definition should continue.