Identification of cells secreting a thymostimulin-like substance and examination of some histoenzymatic pathways in aging avian primary lymphatic organs: II. Bursa of Fabricius.

The Bursa of Fabricius of 15 day, 1-, 3-, and 6 month-old adult chickens (White Leghorn strain) were studied by histological and histochemical staining, histoenzymatic reactions (LDH, SDH, a-GPDH, NAD, NADPH, Ca++-dependent ATP-ase, pH 8.5) and by anti-thymostimulin immunoreaction. Positive reactions for mucopolysaccharides and enzymatic activities were located in the epithelia of the follicles, i.e. in follicle-associated-epithelium (FAE), inter-follicle-epithelium (IFE) and in different epithelial compartments of cortical and medullary zones. Positive reaction for thymostimulin-like (TS-like) substance was restricted to FAE cells and weakly to the basal lamina of IFE. In 6-month-old chickens, the FAE cells disappeared; the phenomenon of bursal regression was evident, although not all the follicles were involved. In the few still normal follicles, the good reactivity to the enzymes tested suggests that residual physiological activity is still present, even if reduced.

Evidence for DNA damage in patients with coronary artery disease.

According to the "monoclonal hypothesis" of atherosclerosis, several studies suggest that cancer and atherosclerosis may have several fundamental biological mechanisms in common. Therefore, an increase in the mutation rate may be involved in the pathogenesis of atherosclerotic plaques. The aim of the study was to verify the presence of chromosomal damage in peripheral blood lymphocytes in patients with coronary artery disease by using micronucleus (MN) test, a reliable biomarker in genetic and cancer risk assessment. Subjects included 53 patients with documented coronary ischemic heart disease (group I); 10 patients with valvular heart disease in absence of atherosclerotic lesions of the coronary arteries (group II) and 16 healthy subjects, age- and sex-matched (group III) were studied as controls. For each subject, two separate cultures were performed and 1000 binucleated cells were scored for the evaluation of MN frequency. The mean (+/-S.E.M.) of MN frequency were 11.9+/-1.7, 5.9+/-1.2 and 3.6+/-0.7 in groups I, II and III, respectively. The MN frequency of group I was significantly higher than that of group III (P=0.02). In group I, MN frequency increased with the number of affected vessels (6.3+/-0.7, 13.9+/-1.6, 14.9+/-5.3 for one-, two-, and three-vessel disease, respectively). Scheffe's test showed that MN frequency was significantly higher in two-vessel compared with one-vessel disease (P=0.0077). Moreover, a positive relationship was found between MN levels and the severity of the disease, calculated by the Duke scoring system (R=0.28, P=0.032), as well as the systolic blood pressure (R=0.34, P=0.009). These results suggest that coronary artery disease in humans is a condition characterized by an increase of DNA damage, positively correlated with the severity of the atherosclerotic disease.

Effect of coronary bypass and cardiac valve surgery on systemic endothelial function.

Seventeen patients scheduled for a cardiac procedure necessitating cardiopulmonary bypass underwent serial perioperative assessment of brachial artery flow-mediated dilation. Patients who underwent coronary bypass surgery had a sustained systemic endothelial dysfunction in the perioperative period, whereas those undergoing cardiac valve surgery experienced transient postoperative systemic endothelial dysfunction.

HIV type 1-induced inhibition of CD45 tyrosine phosphatase activity correlates with disease progression and apoptosis, but not with anti-CD3-induced T cell proliferation.

The tyrosine phosphatase CD45 is a key positive element in multiple lymphocyte signaling pathways. To understand the contribution of CD45 to HIV-1-induced T cell hyporesponsiveness and apoptosis we evaluated the CD45-associated tyrosine phosphatase activity of lymphocytes from patients with different stages of HIV-1 disease and compared it with CD45 expression, spontaneous and Fas-induced apoptosis, anti-CD3-induced T cell proliferation, distribution of CCR5 delta32/wt, and cytokine production. The proliferative response to anti-CD3 as well as the CD45-associated phosphatase activity were significantly reduced in progressors. In long-term nonprogressors (LTNPs) the proliferative response to anti-CD3 was also diminished, although to a lesser extent, while the tyrosine phosphatase activity was not significantly impaired. One-third of LTNPs were found positive for the 32-bp deletion of the CCR5 gene. This mutation had no effects on anti-CD3 proliferative response or CD45 phosphatase activity. A significant reduction in IL-2 and IFN-gamma was observed in both LTNPs and in normal progressors, whereas IL-4 production was significantly decreased only in progressors. Last, we observed a significant correlation between CD45 phosphatase activity and apoptosis. We therefore conclude that the impairment of CD45 tyrosine phosphatase activity correlates with disease progression and the level of T cell apoptosis, but not with anti-CD3-induced T cell proliferation. Moreover, we suggest that evaluation of CD45 tyrosine phosphatase activity may represent an additional tool with which to assess disease progression.

Chemokines, sTNF-Rs and sCD30 serum levels in healthy aged people and centenarians.

Several lines of evidence point to a profound remodelling of the cytokine network in healthy elderly subjects, with decreased type-1 cytokine production (IL 2) and a shift to type 0 and 2. We have also observed an increase of proinflammatory cytokines (IL-1, IL-6, TNF-alpha) in vitro, and an increase of circulating stem cell factor in vivo. In this setting, we studied changes of chemokines (MCP-1 and RANTES) with aging, as well as other molecules, namely, sTNF-RI and sTNF-RII, and the soluble form of the CD30 molecule (sCD30), involved in the pro- and antiinflammatory cytokine balance. The subjects enrolled in the study belonged to three different selected healthy groups of young, aged and centenarians. The presence of rheumatoid factor (RF) and antinuclear antibodies (ANA) was simultaneously assessed. The results show that MCP-1 serum levels were higher in the healthy aged and lowest in the young, while RANTES increased exclusively in centenarians. Only centenarians had autoantibodies (ANA and RF). sTNF-RI and sTNF-RII were significantly elevated in healthy old subjects compared to the young, and even higher in selected centenarians compared to the other age groups. sCD30 serum levels were significantly raised in centenarians compared to the young, despite absence of circulating CD30+ cells in the peripheral blood of the whole study population. No relationship among serum values of these different members of the TNF-R family was found, despite a strong correlation for sTNF-RI and sTNF-RII in all groups. We hypothesize that the increased chemokine levels in aged people, and raised sCD30 levels in centenarians, may reflect a general shift towards type 0/2 cytokines in normal aging, which may be responsible, at least in part, for the appearance of circulating autoantibodies without definite clinical consequences at advanced age.

The role of oxidative imbalance in progression to AIDS: effect of the thiol supplier N-acetylcysteine.

In this study we investigate the redox profile of HIV+ patients at different stages of disease with regard to immunological parameters, i.e., the number of circulating CD4+ and CD8+ lymphocytes. For this purpose, peripheral blood mononuclear cells (PBMCs) obtained from healthy donors, HIV+ patients in the asymptomatic phase, long-term nonProgressors (LTNPs), and AIDS patients have been considered. Cells have been exposed in vitro to the prooxidizing agent menadione, which is able to induce superoxide anion formation, and the susceptibility of the cells to the induced oxidative stress was estimated. Moreover, the possibility that the susceptibility of the cells to oxidative stress might be reduced by preexposing them to the antioxidizing agent N-acetylcysteine (NAC) has also been analyzed. The results obtained can be summarized as follows: (1) treatment with the prooxidant agent is capable of inducing massive morphological alterations in PBMCs. In particular, a significant correlation was found between the decrease in number of CD4+ lymphocytes in patients at different stages of disease and the susceptibility of their PBMCs to oxidative stress; (2) preincubation with NAC was able to preserve partially the ultrastructural characteristics of PBMCs isolated from HIV+ patients. In particular, a direct relationship was found between the efficacy of NAC protection and CD4 counts; (3) evaluation of the plasma index of peroxidation and the number of circulating CD4 lymphocytes indicates the existence of a positive correlation between "systemic" oxidative imbalance and stage of the disease; and (4) cells from LTNPs display either oxidative susceptibility or oxidative markers similar to those of healthy donor cells. Our study suggests that the redox profile of patients may be considered a predictive marker of AIDS progression and that the acute infection and the asymptomatic phase of the disease may represent a useful period in which the combined use of antiretroviral and antioxidant drugs may be beneficial.

Lymphocyte activation gene-3 (LAG-3) expression and IFN-gamma production are variably coregulated in different human T lymphocyte subpopulations.

We evaluated the relationship between cytokine profile and the expression of the lymphocyte activation gene-3 (LAG-3) in both T cell clones and polyclonal T cell lines; LAG-3 is a CD4-like protein whose expression is reportedly restricted to Th1/0 cells and dependent upon IFN-gamma. We found that, while LAG-3 was expressed only by CD4+ T cell clones producing IFN-gamma, most CD8+ clones producing IL-4 but not IFN-gamma (i.e., with a T cytotoxic-2-like profile) were LAG-3+. The intensity of LAG-3 expression by CD8+ clones correlated with the amount of released IFN-gamma, suggesting that this cytokine is not required for expression but rather for the up-regulation of LAG-3. Flow cytometric analyses of polyclonal T cell lines confirmed that LAG-3 could be expressed by both CD4+ and CD8+ cells that did not contain cytoplasmic IFN-gamma. In these cell lines, large proportions of CD4+ and CD8+ cells coexpressed LAG-3 and CD30, a putative marker of Th2-like cells. Overall, our data do not support the earlier suggestion that LAG-3 and CD30 are selective markers of T cells with type-1 and type-2 cytokine profiles, respectively.

HIV-specific lymphoproliferative responses in asymptomatic HIV-infected individuals.

In vitro lymphoproliferative responses to HIV-1 recombinant antigens (gp160, p24, and Rev protein) were studied in 83 patients with asymptomatic HIV-1 infection (CDC groups II and III) and circulating CD4 lymphocyte numbers > 400/mm3. Significant response to at least one of the three antigens was detected in 52.4% of the subjects, but the responses were weak, and concordance of the response to the three antigens was rare, the frequency of individuals responding to each antigen not exceeding 22.4%. Increasing frequencies of response were observed when recall antigens (tetanus toxoid and Candida albicans glycomannoprotein) (65.5%) and anti-CD3 MoAb (76.6%) were used as stimuli. Although a significant association between lymphocyte response to p24, but not gp160, and steadiness of CD4 lymphocyte numbers before the assay was observed, no predictive value for lack of CD4 cell decrease was confirmed for either antigen, and fluctuation of the responses to HIV antigens was seen during subsequent follow up. The panel of T cell assays used could be regarded as appropriate for monitoring both HIV-specific responses and T lymphocyte function during immunotherapy with soluble HIV antigens.

Identification of cells secreting a thymostimulin-like substance and examination of some histoenzymatic pathways in aging avian primary lymphatic organs: I. thymus.

Thymi of adult chickens (White Leghorn strain), ages of 6, 18, 45, days and 1, 3, 6, months, were studied by histological and histochemical stainings, histoenzymatic reactions (LDH, SDH, alpha-GPDH, NADH, NADPH, Ca++ dependent ATPase, pH 8.5) and by anti-thymostimulin immunoreaction. Positive reactions for thymostimulin-like substance, mucopolysaccharides and enzymatic activities were located in different epithelial compartments of the cortical and medullary zones. In 3 and 6 month-old chickens, there was a progressive reduction of the cortical area and of the number of cortical and medullary epithelial cells, and, in the thymi of the 6 month-old chickens, a partial decline in the immunological reactivity to anti-TS, but the intensity of the histochemical and enzymatic reactivities, in comparison with the thymi of the youngest chickens, was not decreased. This may suggest that the cortical and medullary epithelial cells still present in these thymi are capable of secreting and expressing activities and, thus, may maintain their functions.