M. E Estevez

Early Defect of Phagocytic Cell Function in Subjects at Risk for Acquired Immunodeficiency Syndrome

We studied the functions of peripheral blood monocytes and polymorphonuclear cells in 15 apparently healthy homosexual men, eight homosexual or bisexual subjects with unexplained generalized lymphadenopathies (pre‐AIDS), four homosexual men with acquired immunodeficiency syndrome (AIDS), and 15 heterosexual men. In comparison with normal controls, the homosexual groups studied presented a decreased monocyte candidacidal activity for Candida pseudotropicalis that gradually deteriorates as the clinical symptoms progress towards AIDS. The monocyte phagocytic function was retained. Although the phagocytic and candidacidal activities of the polymorphonuclear cells did not differ from those of the normal controls, the candidacidal activity in some of the cases studied was unusually enhanced, indicating that the cells were in an activated state. In addition, only two of nine sera tested from asymptomatic homosexual males were positive for antibodies to HTLV‐III/LAV, while six out of eight pre‐AIDS and both of the two AIDS patients tested had antibodies to AIDS‐associated retrovirus. We suggest that in AIDS the phagocytic system is already involved, together with B and T lymphocyte abnormalities, during the early events of the syndrome, even without the detection of AIDS‐associated retrovirus antibodies.

Lymphocyte Subpopulations and Cytokine Production in Paracoccidioidomycosis Patients

Despite the postulated role of the immune system in the control of the infection by Paracoccidioides brasiliensis, only a few studies have addressed this point in patients. The determination of total lymphocytes and their subpopulations in 6 untreated patients with the chronic form of paracoccidiodomycosis showed that half of them were lymphopenic, because of low number of CD4+ T‐lymphocytes. All patients had low CD4/CD8 ratios. On the contrary, B‐lymphocytes were normal in all patients. An additional patient, studied on treatment with ketoconazole, had normal lymphocyte counts in all subpopulations, as did one of the patients previously studied at diagnosis when he received specific antimycotic treatment. The production of interferon and tumor necrosis factor, determined by bioassay in supernatants of mononuclear blood cells of the patients, induced by interleukin 2 in vitro was significantly lower than that of normal subjects. These results show that patients with paracoccidioidomycosis have a defect in blood lymphocyte subsets as well as in the ability to produce regulatory cytokines.

Cytogenetic and immunologic phenotype findings in hodgkin's disease

There are very few chromosome studies using banding techniques of lymph nodes in Hodgkins disease (HD), and determinations of immunologic phenotypes are scarce. We have performed both cytogenetic and immunologic studies in 12 of 22 lymph node biopsies of different histologic types obtained from 20 HD patients (no mitotic cells were found in the remaining ten lymph nodes). A near-diploid modal number was obtained in 80% of the cases, and 20% showed a bimodal distribution. Clones were observed in 50% of HD lymph nodes, with chromosome markers in 60% of them. Markers 15q+, 5p-, and der(X) and a trisomy of chromosome #21 were observed in our cases. Seventy-one percent of the lymph nodes studied showed a predominance of T lymphocytes. Within the lymph nodes, where the karyotype was determined, 4/12 lymph nodes presented a predominance of B lymphocytes, and they were all included in the group with structural chromosome abnormalities.

Comparison of Candida killing activity measured by chemiluminescence and cytomorphological methods in human phagocytes

Phagocyte function can be assayed by many laboratory tests including a cytomorphological method that uses Candida cells as target. The aim of this study was to correlate this technique with the production of toxic oxygen metabolites, measured by chemiluminescence (CL). The biological function of polymorphonuclear (PMN) cells and monocytes from the blood of 24 normal subjects and 25 patients with immunodeficiency diseases were studied. CL was measured using opsonized zymosan as the stimulating agent and, for the evaluation of Candida killing activity, C. pseudotropicalis and C. albicans were used as targets. A linear correlation between CL and lytic activity was observed with both PMN and monocytes from normal subjects and patients (r = 0.563 to 0.955; P less than 0.05 to less than 0.001). Our results indicate that the production of toxic oxygen metabolites, as measured by CL is closely related to the killing of Candida by PMN and monocytes.

Impaired Phagolysosomal Fusion of Peripheral Blood Monocytes from HIV‐Infected Subjects

We evaluated phagolysosomal fusion in peripheral blood monocytes from 20 HIV‐infected individuals and 40 normal controls, using a fluorescence assay with acridine orange as marker. The percentages of phagolysosomal fusion of monocytes from HIV‐infected subjects, after 30 and 60 min of yeast ingestion, (mean ± standard deviation) 57·2 ± 17 and 63·2 ± 18·6, respectively, when compared to normal controls (72·4 ± 7·8 and 77 ± 8·1), did not differ significantly. However, there was a direct linear association between the percentages of phagolysosomal fusion and CD4+ lymphocytes (P<0·001) or CD4/CD8 T‐cell ratio (P<0·01). These results suggest that phagolysosomal dysfunction becomes evident at late stages of HIV infection and progresses as CD4+ T‐lymphocyte count and CD4/CD8 T‐cell ratio decrease. On the other hand, recombinant gp120 inhibited significantly normal phagolysosomal fusion at concentrations ranging between 1 and 1000 ng/ml. Taking together the results obtained, we can conclude that gp120 could be responsible for monocyte phagolysosomal dysfunction observed in HIV infected patients.

Natural Killer Suppressor Factors in Sera from HIV‐Infected Subjects

We investigated the presence of NK suppressor factors in HIV+ sera. We further investigated if gp120 could be one of the substances responsible for the impairment of NKC regulation found in HIV+ asymptomatic patients. Our results indicate that HIV+ sera inhibit significantly normal NKC in a dose‐dependent way, even at concentrations as low as 1%. The inhibitory effect of HIV+ sera decreased, but was not completely removed, by adsorptions of IgG or by treatment with a MoAb against human FcIgG. Pretreatment of normal effector cells with anti‐CD 16 MoAb slightly reduced their cytotoxic capability, but did not modify the suppressor effect of HIV+ sera. The preincubation of normal PBMC with recombinant gp120 had also a suppressor effect even at 10ng/ml. Pretreatment of HIV+ sera with anti‐gp120 or anti‐FcIgG MoAb reduced, but not completely, their inhibitory effect. In conclusion, HIV+ serum has a dose‐dependent inhibitory effect on normal NKC. Most of this inhibition is caused by IgG, but other substances, such as gp120, can also contribute to it. Since the removal of IgG and further treatments of HIV+ sera were not able to abrogate completely the NK suppression, other serum factors still undetermined (TNF‐α, other cytokines), should be considered.

Dysfunction of monocytes in Hodgkin's disease by excessive production of PGE-2 in long-term remission patients

The candidacidal activity and the production of oxygen radicals by monocytes were investigated in untreated and long‐term remission patients with Hodgkins disease (HD). Both groups showed a decreased candidacidal function of monocytes with a chemiluminescence (CL) response significantly lower and delayed with respect to normal controls. Indomethacin at 1 μg/ml corrected the monocyte deficiency increasing the CL response to normal values and normalizing the kinetics in the untreated patients. However, in patients in remission, the peak was delayed and followed by a significant increase in the production of oxygen radicals compared with untreated patients. A direct linear correlation was found between the percentages of lysed Candida and maximum CL peak of stimulated monocytes. When prostaglandin E2 (PGE‐2) levels, measured in supernatants of cultured mononuclear cells, were plotted against the percentages of killed Candida, an inverse linear correlation was found. Therefore, monocytes from HD patients have a dysfunction in the generation of oxygen radicals and a decreased candidacidal activity associated with excessive production of PGE‐2. Indomethacin can correct the oxidative metabolism in the untreated patients while in apparently “cured” patients the disorder persists.

Decreased phagolysosomal fusion of peripheral blood monocytes from patients with thalassemia major.

We evaluated the phagolysosomal fusion of peripheral blood monocytes from 15 patients with thalassemia major and 10 thalassemia major carriers using a cytomorphological method with acridine orange as fusion marker. The monocyte phagolysosomal fusion of thalassemic patients was decreased (49.6 +/- 8.6%, mean +/- SD) and differed significantly (p < 0.05) from those of carriers and normal controls (65.7 +/- 11.4% and 74.6 +/- 5.7%, respectively). In vitro deferoxamine partially improved monocyte phagolysosomal fusion of patients with thalassemia major, and did not affect monocyte function in carriers and healthy subjects. Furthermore, in vitro addition of ferrous sulfate decreased normal phagolysosomal fusion. We conclude that the monocyte phagolysosomal fusion dysfunction of thalassemic patients could be related to iron overload.

Effect of Indomethacin and Thymostimulin on Natural Killer Activity of Thalassemic Patients and Normal Subjects

To evaluate whether indomethacin (IM) and thymostimulin (TP-1) could revert the alterations of natural killer cytotoxicity (NKC) observed in patients and carriers of Thalassemia Major (TM), 14 TM patients, 10 TM carriers, 16 normal controls, and 4 polytransfused patients were studied. In vitro, IM induced an increase in NKC of normal controls and TM carriers (Δ CI: + 5.98±2.49 and + 9.77±6.82 respectively), but not in TM patients (-0.32±1.45) or other polytransfused recipients. The addition of IM to Concanavalin A (Con A) also resulted in an increase of NKC in normal controls and TM carriers, similar to that induced by each substance separately in normal controls, or by IM alone in TM carriers. Similarly, TP-1 (50 μg/ml) induced increases of NKC in normal subjects (+ 4.66±3.62), but not in TM patients (-1.1±2.43). The impairment in NKC observed in TM and polytransfused patients, and the absence of response to Con A in both, TM patients and carriers, do not seem to be mediated by an excessive production of prostaglandins. The lack of response not only to Con A, but also to TP-1 could indicate a quantitative or qualitative defect in T cells, that affects NKC regulation, or a deficient response of NK cells to modulators produced by T lymphocytes.

Acute lymphoblastic leukemia in Argentina. Relationship between surface markers and prognosis.

Leukemic blasts from patients with acute lymphoblastic leukemia (ALL) were tested initially for the following surface markers: sheep erythrocyte receptors at 4 degrees C and 37 degrees C; receptor for the C3 fraction of complement; mouse erythrocyte receptor; and surface immunoglobulins. We found that 73% of the ALL were devoid of these markers, 9% had C3 receptor only, 2% were B-ALL, and 15% were T-ALL. Only one of 147 cases tested expressed receptors for mouse erythrocyte receptor. C3-ALL predominated in girls and the prognosis was similar to that of the ALL devoid of markers. T-ALL was associated with some high-risk factors, such as high initial leukocyte counts, and a predominance in boys older than 5 years of age. Further studies were done in which cytoplasmic immunoglobulins were evaluated and surface antigens were identified by the following monoclonal antibodies: OKT1, OKT3, OKT4, OKT6, OKT8, OKT9, OKT10, OKT11a, OKIa1, and J5 (anti-CALLA). Less than 1% were B-All, 15% were T-ALL, and approximately 85% were non-T, non-B ALL. Of the latter group, 69% were common-ALL, 16% were pre-B ALL, and 15% were null-ALL. Within the T-ALL, the expression of the T antigenic mosaic was heterogeneous. Results by a multivariate analysis demonstrated that sex, age, initial leukocyte count, and T-cell phenotype were independent variables with a negative prognostic value (p less than 0.01), and the only combination with significant interaction was between T-cell phenotype and leukocyte counts.