Search

Checkpoint inhibitors have significantly changed the prognosis of patients with relapsing refractory classical Hodgkin's lymphoma (cHL), demonstrating excellent results in heavily pretreated patients. However, there is still limited data on the real-world experience with PD-1 inhibitors in cHL. Within the context of the Apulian hematological network (Rete Ematologica Pugliese, REP), we performed a retrospective, multicenter analysis of 66 patients with relapsing refractory cHL who had received PD-1 inhibitors in the non-trial setting. Forty-three patients (65%) were treated with nivolumab and 23 (35%) with pembrolizumab. Thirty-one (47%) and 8 (12%) patients underwent autologous or allogeneic stem cell transplantation prior to checkpoint inhibitor therapy, respectively. The median number of lines of treatment attempted prior to PD-1 inhibitor therapy was 4 (range, 3 to 7). All patients had received brentuximab vedotin prior to checkpoint inhibitor therapy. The overall response rate to PD-1 inhibitors therapy was 70% (47% complete remission (CR) and 23% partial remission (PR)). Twenty-four immune-related adverse events (19 (80%) grades 1-2; 5 (20%) grades 3-4) were documented (4 gastrointestinal, 4 hepatic, 6 fever, 4 hematological, 3 dermatological, 3 allergic rhinitis). Toxicity resolved in all patients, and there were no deaths attributed to checkpoint inhibitor therapy. After a median follow-up of 26 months (range 3-72 months), 54 patients (82%) are alive, and 12 (18%) died. The cause of death was attributed to disease progression in 9 patients and sepsis in 3 patients. After PD-1 inhibitor therapy, 22 patients (33%) relapsed or progressed. The overall survival and progression-free survival at 5 years were 65% and 54%, respectively. This study confirms the efficacy and tolerability of PD-1 inhibitor therapy in relapsed refractory cHL in a real-world setting, demonstrating similar clinical outcomes and toxicity profiles compared to clinical studies., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Aims: Frailty is common in patients with atrial fibrillation (AF), with possible impact on therapies and outcomes. However, definitions of frailty are variable, and may not overlap with frailty perception among physicians. We evaluated the prevalence of frailty as perceived by enrolling physicians in the Edoxaban Treatment in Routine Clinical Practice for Patients With Non-Valvular AF (ETNA-AF)-Europe registry (NCT02944019), and compared it with an objective frailty assessment., Methods and Results: ETNA-AF-Europe is a prospective, multi-centre, post-authorization, observational study. There we assessed the presence of frailty according to (i) a binary subjective investigators' judgement and (ii) an objective measure, the Modified Frailty Index. Baseline data on frailty were available in 13 621/13 980 patients. Prevalence of perceived frailty was 10.6%, with high variability among participating countries and healthcare settings (range 5.9-19.6%). Conversely, only 5.0% of patients had objective frailty, with minimal variability (range 4.5-6.7%); and only <1% of patients were identified as frail by both approaches. Compared with non-frailty-perceived, perceived frail patients were older, more frequently female, and with lower body weight; conversely, objectively frail patients had more comorbidities. Non-recommended edoxaban dose regimens were more frequently prescribed in both frail patient categories., Conclusions: Physicians' perception of frailty in AF patients is variable, mainly driven by age, sex, and weight, and quite different compared with the results of an objective frailty assessment. Whatever the approach, frailty appears to be associated with non-recommended anticoagulant dosages. Whether this apparent inappropriateness influences hard outcomes remains to be assessed., Competing Interests: Conflict of interest: I.D. reports having received speaker fees from Boehringer Ingelheim, Bayer, Bristol-Myers Squibb/Pfizer, Daiichi-Sankyo, Medtronic, Boston Scientific, and Biotronik. A.B. reports being involved in research sponsored by and is a member of advisory panels and speakers’ bureau for Daiichi Sankyo, Pfizer, BMS, Bayer, and Boehringer Ingelheim. P.-E.R., M.C.M., and J.A.G.d.S. are employees of Daiichi Sankyo Europe GmbH. L.P. has received fees and honoraria from Daiichi-Sankyo, SOTIO Biotech, and Beckman-Coulter. P.K. receives research support from European Union, British Heart Foundation, Leducq Foundation, Medical Research Council (UK), and German Centre for Cardiovascular Research, from several drug and device companies active in atrial fibrillation, and has received honoraria from several such companies. He is listed as inventor on two patents held by the University of Birmingham (Atrial Fibrillation Therapy WO 2015140571, Markers for Atrial Fibrillation WO 2016012783). He is co-principal investigator of ETNA-AF-Europe. R.D.C. co-authored ESC Guidelines on Atrial Fibrillation 2010–2012; acted as a Steering Committee member and National Coordinator for Italy, and co-authored manuscripts published on APPRAISE-2, ARISTOTLE, AVERROES, ENGAGE AF-TIMI 48, and Re-DUAL PCI. R.D.C. has received fees, honoraria, and research funding from Sanofi-Aventis, Boehringer Ingelheim, Bayer, Bristol-Myers Squibb/Pfizer, Daiichi-Sankyo, Novartis, Portola, Roche, and Merck. He is co-principal investigator of ETNA-AF-Europe. All remaining authors have declared no conflicts of interest., (© The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology.)

Competing Interests: The authors declare no conflicts of interest., (Copyright © 2019 the Author(s). Published by Wolters Kluwer Health, Inc. on behalf of the European Hematology Association.)

Introduction: Non-familial ascending thoracic aorta dilation and aneurysms (TAAs) are silent diseases in elderly patients. Histopathology revealed that functionally polarized infiltrating CD4+ T-cells play a key role in aortic wall weakening., Objective: To evaluate the possible associations between phenotype and cytokine production of circulating CD4+ T-lymphocytes and the presence of TAA in patients with aortic valve disease (AVD)., Methods: We studied blood samples from 10 patients with TAA and 10 patients with AVD. Flow cytometry was used to quantify: a) CD4+ T-lymphocytes surface expression of CD25, CD28, and chemokine receptors (CCR5, CXCR3, CX3CR1); b) fractions of in vitro stimulated CD4+ T-cells producing cytokines (interferon gamma [IFN-γ], interleukin [IL]-17A, IL-21, IL-10); c) CD4+CD25highFoxP3+ regulatory T-cells (Treg) fraction. Enzyme-linked immunosorbent assays (ELISA) were performed for cytokines (IFN-γ, IL-6, IL-10, IL-17A, IL-23, transforming growth factor beta [TGF-β]) and chemokines (RANTES, CX3CL1)., Results: The total CD4+CD28±CD4+/CX3CR1+ T-cells fraction was higher (P=0.0323) in AVD (20.452±4.673) than in TAA patients (8.633±2.030). The frequency ratio of CD4+ T-lymphocytes producing IFN-γ vs. IL-17A+IL-21 cytokine-producing CD4+ T-cells was higher (P=0.0239) in AVD (2.102±0.272) than in TAA (1.365±0.123) patients. The sum of CD4+CD28±CD4+/CX3CR1+ T-cells correlated positively with values of the previous cytokine ratio (P=0.0002, R=0.732). The ratio of CD4+CD28±CD4+/CX3CR1+ T-cells vs. Treg was higher (P=0.0008) in AVD (20.859±3.393) than in TAA (6.367±1.277) patients., Conclusion: Our results show that the presence of TAA in subjects with AVD is associated with imbalance between phenotypic and cytokine-producing subsets of circulating CD4+ T-lymphocytes, prevalently oriented towards a pro-fibrotic and IFN-γ counteracting effect to functional polarization.

BEAM (carmustine [bis-chloroethylnitrosourea (BCNU)]-etoposide-cytarabine-melphalan) chemotherapy is the standard conditioning regimen for autologous stem cell transplantation (ASCT) in lymphomas. Owing to BCNU shortages, many centers switched to fotemustine-substituted BEAM (FEAM), lacking proof of equivalence. We conducted a retrospective cohort study in 18 Italian centers to compare the safety and efficacy of BEAM and FEAM regimens for ASCT in lymphomas performed from 2008 to 2015. We enrolled 1038 patients (BEAM = 607, FEAM = 431), of which 27% had Hodgkin lymphoma (HL), 14% indolent non-Hodgkin lymphoma (NHL), and 59% aggressive NHL. Baseline characteristics including age, sex, stage, B-symptoms, extranodal involvement, previous treatments, response before ASCT, and overall conditioning intensity were well balanced between BEAM and FEAM; notable exceptions were median ASCT year (BEAM = 2011 versus FEAM = 2013, P < .001), Sorror score ≥3 (BEAM = 15% versus FEAM = 10%, P = .017), and radiotherapy use (BEAM = 18% versus FEAM = 10%, P < .001). FEAM conditioning resulted in higher rates of gastrointestinal and infectious toxicities, including severe oral mucositis grade ≥3 (BEAM = 31% versus FEAM = 44%, P < .001), and sepsis from Gram-negative bacteria (mean isolates/patient: BEAM = .1 versus FEAM = .19, P < .001). Response status at day 100 post-ASCT (overall response: BEAM = 91% versus FEAM = 88%, P = .42), 2-year overall survival (83.9%; 95% confidence interval [CI], 81.5% to 86.1%) and progression-free survival (70.3%; 95% CI, 67.4% to 73.1%) were not different in the two groups. Mortality from infection was higher in the FEAM group (subhazard ratio, 1.99; 95% CI, 1.02 to 3.88; P = .04). BEAM and FEAM do not appear different in terms of survival and disease control. However, due to concerns of higher toxicity, fotemustine substitution in BEAM does not seem justified, if not for easier supply., (Copyright © 2018 The American Society for Blood and Marrow Transplantation. Published by Elsevier Inc. All rights reserved.)

Objective: The angiotensin-converting enzyme (ACE) is highly expressed in the aneurysmal vascular wall, in both animal models and human disease. Genetic variations in ACE could be crucial in determining the risk of thoracic aortic aneurysm (TAA). The aim of the present study was to examine the role of ACE insertion/deletion polymorphism on the risk of TAA in patients with bicuspid aortic valves or tricuspid aortic valves., Methods: We enrolled 216 patients (158 men; age, 58.9±14.9 years) with TAA, associated with bicuspid aortic valves (n=105) and tricuspid aortic valves (n=111) compared with 312 patients (252 men; age, 54.6±11.0 years) with angiographically proven coronary artery disease and 300 healthy controls (91 men; age, 40.4±10.5 years)., Results: The genotype distribution of ACE insertion/deletion was significantly different between the patients with TAA compared with both the control group (P=.0005) and the coronary artery disease group (P=.03). The genotypes were not different between the control group and the coronary artery disease group (P=.3). Compared with the controls, both the bicuspid aortic valve patients (P=.0008) and tricuspid aortic valve patients (P<.0001) had a greater frequency of allele D. The aortic diameters were significantly different among the three genotypes (48.3±6.6, 45.3±8.9, 39.9±8.7 for the DD, DI, and II genotypes, respectively; P=.0002). A synergistic effect between the ACE D allele and hypertension was found for both an increased aortic diameter (P=.003) and the risk of TAA (P<.001). On multivariate logistic regression analysis, D allele (odds ratio, 3.0; 95% confidence interval, 1.1-8.1; P=.03) was a significant predictor of TAA., Conclusions: ACE insertion/deletion polymorphism represents a genetic biomarker for TAA. These findings could have a significant effect on both the early detection and effective pharmacologic treatment of aortic disease., (Copyright © 2012 The American Association for Thoracic Surgery. Published by Mosby, Inc. All rights reserved.)

Objective: Hormone replacement therapy (HRT) is acknowledged as the gold standard for the alleviation of climacteric vasomotor symptoms. Prothrombotic genetic variants have been suggested to increase thrombotic risk among HRT users. The aim of the study was to determine whether a positive family history may identify a genetic predisposition for thrombosis in women before prescribing HRT., Methods: From January 2005 to May 2009, we consecutively enrolled 145 asymptomatic women (mean age 51.2 ± 5.4 years) without previous episodes of venous and/or arterial thrombosis referred to our Genetics Research Unit before starting HRT. A detailed family history was reconstructed and we identified 48 women (33.1%) with a positive family history, defined as venous thromboembolism and/or stroke or heart attack, in first-degree relatives before 60 years for men and 65 years for women. A group of 121 women (mean age 54.0 ± 9.1 years) with an episode of venous and/or arterial thrombosis was also included. Genetic screening for factor V Leiden, prothrombin G20210A and methylenetetrahydrofolate reductase C677T polymorphisms was performed., Results: The frequency of factor V Leiden or prothrombin G20210A mutations was significantly higher both in asymptomatic women with a positive family history (16.7% vs. 2.1%, p = 0.001) and in patients with thrombosis (12.4% vs. 2.1%; p = 0.005) compared with asymptomatic women without a family history. Multivariate regression analysis showed a synergic effect between the presence of one prothrombotic mutation and family history on the risk of thrombosis (odds ratio 3.7, 95% confidence interval 1.9-7.2)., Conclusions: A positive family history of thrombosis is a sensitive indicator for selected genetic testing in high-risk women before starting HRT.

In patients with dilated cardiomyopathy (DCM) abnormal myocardial blood flow (MBF) has been associated with coronary microvascular dysfunction. The aim of this study was to test the hypothesis that osteopontin (OPN) plasma levels could be associated with the activation of the renin-aldosterone system (RAS) in these patients and be involved in mediating myocardial and coronary damage. In 66 patients with idiopathic left ventricular dysfunction of variable severity the plasma levels of OPN were correlated with biomarkers of systemic metabolism, RAS activation, myocardial dysfunction and with clinical indexes of left ventricle (LV) function and perfusion obtained by 2D-echocardiography and PET. As compared to controls, patients showed a significant increase of inflammatory markers (OPN: 508+/-30.8ng/ml vs. 426.9+/-16.4, p<0.05 and interleukin (IL)-6: 1.71+/-0.29pg/ml vs. 0.38+/-0.03pg/ml, p<0.001) and of indexes of cardiac damage. OPN levels were significantly correlated with the extent of microvascular dysfunction (MBF at rest: p=0.01; during dipyridamole: p=0.0003) and with plasma renin activity (PRA) (r=0.26, p=0.04). Both in patients with milder or more severe LV dysfunction lower MBF values were associated with higher OPN levels and PRA. These results suggest a interdependent role of RAS and vascular inflammation in cardiomyopathy., (2009 Elsevier Ltd. All rights reserved.)

Rheumatoid arthritis (RA) patients are characterized by increased arterial stiffness, an independent predictor of cardiovascular risk. It has been suggested that osteopontin (OPN), a cytokine involved in RA pathogenesis, might have vascular effects. To study a possible relationship between OPN and arterial stiffness, aortic pulse wave velocity (PWV) was measured by tonometry in 69 patients (41 with RA, 28 with systemic sclerosis [SSc]) and 18 healthy controls. Plasma OPN levels, oxidative stress markers, and endothelin 1 (ET-1) were assessed. OPN levels were significantly (P < 0.05) higher in RA (median 9.93, range 4.36-47.80 ng/mL) than in SSc (4.3, 2.1-19.7 ng/mL) or controls (5.2, 4.1-9.4 ng/mL). In RA patients, log-OPN was related to log-C-reactive protein (log-CRP) (r = 0.30, P < 0.05), age (r = 0.38, P < 0.01), Health Assessment Questionnaire (HAQ) (r = 0.58, P < 0.0001), and inversely related to total cholesterol (r = -0.33, P < 0.05) and apolipoprotein A (apoA) (r = -0.58, P < 0.001), but not to oxidative stress markers and ET-1. PWV was similar in RA (median 8.1, range 4.7-16.4 m/s) and SSc (median 8.7, range 7.1-13.1 m/s), but significantly greater (P < 0.01) than controls (median 7.5, range 4.1-10.4 m/s). Aortic PWV was related to log-OPN (r = 0.40, P < 0.01) only in RA patients. It also was related to age (r = 0.34, P < 0.05), mean blood pressure (r = 0.44, P < 0.001), and HAQ (r = 0.48, P < 0.001). In multiple regression analysis (r(2) = 0.36), including confounders, log-OPN remained a significant predictor (P < 0.05) of PWV in RA. Elevated plasma OPN levels are associated with increased arterial stiffness in RA patients, suggesting that this protein might represent a bridge protein between inflammation and the consequent joint damage and cardiovascular risk in RA patients.

Electron-density modification (EDM) procedures are the classical tool for driving model phases closer to those of the target structure. They are often combined with automated model-building programs to provide a correct protein model. The task is not always performed, mostly because of the large initial phase error. A recently proposed procedure combined EDM with DEDM (difference electron-density modification); the method was applied to the refinement of phases obtained by molecular replacement, ab initio or SAD phasing [Caliandro, Carrozzini, Cascarano, Giacovazzo, Mazzone & Siliqi (2009), Acta Cryst. D65, 249-256] and was more effective in improving phases than EDM alone. In this paper, a novel fully automated protocol for protein structure refinement based on the iterative application of automated model-building programs combined with the additional power derived from the EDM-DEDM algorithm is presented. The cyclic procedure was successfully tested on challenging cases for which all other approaches had failed.

The DEDM (difference electron-density modification) algorithm has been described in a recent paper [Caliandro et al. (2008), Acta Cryst. A64, 519-528]: it breaks down the collinearity between model structure phases and difference structure phase estimates. The new difference electron-density produced by DEDM, summed to the calculated Fourier maps, is expected to provide a representation of the full structure that is more accurate than that obtained by the observed Fourier synthesis. In the same paper, the DEDM algorithm was combined with the EDM (electron-density modification) approach to give the EDM-DEDM procedure which, when applied to practical molecular-replacement cases, was able to improve the model structures. In this paper, it is shown that EDM-DEDM suffers from some critical points that did not allow cyclical application of the procedure. These points are identified and modifications are made to allow iteration of the procedure. The applications indicate that EDM-DEDM may become a fundamental tool in protein crystallography.

Bouveret's syndrome is a rare condition usually caused by a single large stone impacted in the duodenum. This is a cause of gastric outlet. Even if endoscopy is the mainstay of diagnosis, the radiographic examinations are also important too. Generally, the stones are too large to be removed endoscopically. Conservative endoscopic treatment should be attempted initially, and if it fails, surgical approach should be performed.

Aortic Valve Disease, includes a range of disorder severity from mild leaflet thickening without valve obstruction, "aortic sclerosis", to severe calcified aortic stenosis. It is a slowly progressive active process of valve modification similar atherosclerosis for cardiovascular risk factors, lipoprotein deposition, chronic inflammation, and calcification. Systemic signs of inflammation, as wall and serum CRP, similar to those found in atherosclerosis, are present in patients with degenerative aortic valve stenosis and may be expression of a common disease, useful in monitoring of stenosis progression.

In patients with primary Ab deficiencies, hematological and immunological abnormalities are frequently observed. A regenerative failure of hemopoietic stem/progenitor cells has been hypothesized. We evaluated in the bone marrow (BM) of 11 patients with common variable immunodeficiency, the phenotype of BM progenitors and their in vitro growth by colony-forming cell (CFC) and long-term culture (LTC) assays. A significant decrease in erythroid and mixed CFC and, to a greater extent, in primitive LTC-CFC progenitors was observed in patients compared with healthy controls. The frequency of BM pre-B and pro-B cells correlated directly with the absolute number of CD19+ lymphocytes. BM cells cultured in vitro produced spontaneously lower amounts of IL-2 and elevated levels of TNF-alpha compared with controls, indicating a skewing toward a proapoptotic cytokine pattern. In addition, stromal cells generated after BM LTC secreted less IL-7 and displayed by immunohistochemistry an altered phenotype. These findings were associated with a significant decrease in naive Th cells coexpressing CD31 in the peripheral blood. These results indicate an impaired growth and differentiation capacity of progenitor cells in patients with common variable immunodeficiency.

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.

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.

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.

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 purpose of this study is to offer a view of ion beam thinning at atomistic level. Therefore, a computer simulation of the type molecular dynamics is used to study the evolution of a stepped silicon surface undergoing ion bombardment. According to the methods used in radiation damage studies, the effect of the impinging beam is described by constructing a large ensemble of trajectories of displaced silicon atoms. The effects of the beam parameters, such as energy and angle, as well as the ones arising from the surface topography, are obtained from the properties of this ensemble. The results of the simulations are discussed in the light of the mesoscopic models of surface evolution.

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.

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.

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.

To investigate whether defective costimulatory signals could be involved in the loss of T lymphocyte functions during HIV-1 infection, we tested the effect of CD28 costimulation on both T cell receptor/CD3 and HIV-1 antigen-induced proliferative responses. Although CD3-mediated responses significantly decreased with more advanced stages of HIV-1 infection, the ability of potentiating the responses through CD28 costimulation was maintained at all stages and did not differ from that of HIV-1- subjects. When CD28 costimulation was studied in lymphocyte cultures stimulated with HIV-1 gp160 or p24, potentiation was seen only when a significant response was present without additional CD28 triggering, namely in subjects receiving active immunization with recombinant gp160. These results confirm the integrity of the CD28 pathway of costimulation during HIV-1 infection, and suggest that lymphocytes responding to soluble HIV-1 antigen are not deleted in HIV-1-infected patients, but do not receive significant priming during the natural course of the infection.