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ABSTRACT Objective to prospectively evaluate the ability of post-graduate students enrolled in a laparoscopy program of the Institute for Teaching and Research to complete single port total nephrectomies. Materials and Methods 15 post-graduate students were enrolled in the study, which was performed using the SILStm port system for single-port procedures. All participants were already proficient in total nephrectomies in animal models and performed a left followed by a right nephrectomy. Analyzed data comprised incision size, complications, and the time taken to complete each part of the procedure. Statistical significance was set at p

We propose the Augmented Grouping Approach (AugGA) and its deployment in the Augmented Grouping GO (AugGGO) scheme, for an efficient exploration of the chemical ordering (or compositional structure) of multi-component (alloyed) nanoparticles. The approach is based on a 'grouping' strategy (previously proposed for high-symmetry structures) by which the number of compositional degrees of freedom of the system is decreased by defining sets of atoms (groups, or orbits, or shells) that are constrained to be populated by the same element. Three fundamental advances are here included with respect to previous proposals: (i) groups are defined on the basis of descriptors (no point-group symmetry is assumed), (ii) bulk groups can exploit general chemical ordering patterns taken from databases, and (iii) sub-grouping is realized via a multi-descriptor strategy (here using two basic descriptors: the atomic energy and a few types of geometry patterns). The AugGGO approach is applied to two prototypical examples of binary nanoalloys: Pd-Pt and Ag-Cu, with a size between ≈500 and ≈1300 atoms, in different configurations, and the convex hull of the mixing energy as a function of composition is derived. It is shown how the three advances here proposed decisively extend the power and scope of the grouping approach: (i) making it applicable to any generic structural framework, (ii) achieving a thorough sampling of the core regions of nanoparticles, and (iii) catching exotic/unexpected chemical ordering arrangements, at a computational cost which is 1-2 orders of magnitude smaller than that of traditional Monte Carlo single-exchange techniques.

Objective: To assess the efficacy of autologous conditioned serum (ACS) for the treatment of patients with knee osteoarthritis after failure of medical treatments and platelet rich plasma (PRP) injections., Background: Knee osteoarthritis is the most common form of arthritis. Prior to prosthetic surgery these patients might benefit from medical treatments, physiotherapy, and in case of their ineffectiveness, from autologous blood component injections., Methodology: We have treated 30 patients with Kellgren-Lawrence I-III knee osteoarthritis with ACS after failure of standard medical treatments/physiotherapy and platelet rich plasma (PRP) injections for a full cycle, within the previous year from enrollment., Results: ACS administration was performed in all patients with mild side effects and produced prompt (1 month) improvements of VAS and Lequesne scales in 67% of patients and this result persisted at 6 and 12 months. No relationship between the rate of response and Kellgren-Lawrence scale at enrollment was observed whilst responders had a significantly higher amount of interleukin-1 receptor antagonist (IL1-RA) in ACS as compared to nonresponders., Conclusion: The present study confirms the efficacy of ACS in pain control and functional recovery of patients with knee osteoarthritis resistant to medical and PRP treatment. These results were obtained in a well-defined cohort of resistant patients and seem to be related with IL1-RA content in injected ACS., (© 2021 British Blood Transfusion Society.)

Background: Convolutional Neural Networks can be effectively used only when data are endowed with an intrinsic concept of neighbourhood in the input space, as is the case of pixels in images. We introduce here Ph-CNN, a novel deep learning architecture for the classification of metagenomics data based on the Convolutional Neural Networks, with the patristic distance defined on the phylogenetic tree being used as the proximity measure. The patristic distance between variables is used together with a sparsified version of MultiDimensional Scaling to embed the phylogenetic tree in a Euclidean space., Results: Ph-CNN is tested with a domain adaptation approach on synthetic data and on a metagenomics collection of gut microbiota of 38 healthy subjects and 222 Inflammatory Bowel Disease patients, divided in 6 subclasses. Classification performance is promising when compared to classical algorithms like Support Vector Machines and Random Forest and a baseline fully connected neural network, e.g. the Multi-Layer Perceptron., Conclusion: Ph-CNN represents a novel deep learning approach for the classification of metagenomics data. Operatively, the algorithm has been implemented as a custom Keras layer taking care of passing to the following convolutional layer not only the data but also the ranked list of neighbourhood of each sample, thus mimicking the case of image data, transparently to the user.

CIK cells are a subset of effector lymphocytes endowed with a non-MHC restricted anti-tumor activity making them an appealing and promising cell population for adoptive immunotherapy. CIK are usually generated ex-vivo by initial priming with Interferon-γ (IFN-γ) and monoclonal antibody against CD3 (anti-CD3), followed by culture in medium containing Interleukin-2 (IL-2). Interleukin-15 (IL-15) shares with IL-2 similar biological functions and recently it has been reported to induce CIK with increased anti-leukemic potential. The aim of the study was to compare the killing efficacy of CIK generated by IL-2 alone or IL-2 and IL-15 toward tumor targets of different origins, leukemic cells and malignant cells from epithelial solid tumors. CIK bulk cultures were examined for cell proliferation, surface phenotype and cytotoxic potential against tumor cell lines K562, HL60, HeLa and MCF-7. The results showed that IL-15 is able to induce a selective expansion of CIK cells, but it is less effective in sustaining CIK cell proliferation compared to IL-2. Conversely, our data confirm and reinforce the feature of IL-15 to induce CIK cells with a potent cytotoxic activity mostly against tumor cells from epithelial solid malignancies via NKG2D-mediated mechanism., (Copyright © 2016 American Society for Histocompatibility and Immunogenetics. Published by Elsevier Inc. All rights reserved.)

Background: Mesenchymal stromal cells (MSCs) have been largely investigated, in the past decade, as potential therapeutic strategies for various acute and chronic pathological conditions. MSCs isolated from different sources, such as bone marrow (BM), umbilical cord tissue (UCT) and adipose tissue (AT), share many biological features, although they may show some differences on cumulative yield, proliferative ability and differentiation potential. The standardization of MSCs growth and their functional amplification is a mandatory objective of cell therapies. The aim of this study was to evaluate the cumulative yield and the ex vivo amplification potential of MSCs obtained from various sources and different subjects, using defined culture conditions with a standardized platelet lysate (PL) as growth stimulus., Methods: MSCs isolated from BM, UCT and AT and expanded in human PL were compared in terms of cumulative yield and growth potential per gram of starting tissue. MSCs morphology, phenotype, differentiation potential, and immunomodulatory properties were also investigated to evaluate their biological characteristics., Results: The use of standardized PL-based culture conditions resulted in a very low variability of MSC growth. Our data showed that AT has the greater capacity to generate MSC per gram of initial tissue, compared to BM and UCT. However, UCT-MSCs replicated faster than AT-MSCs and BM-MSCs, revealing a greater proliferation capacity of this source irrespective of its lower MSC yield. All MSCs exhibited the typical MSC phenotype and the ability to differentiate into all mesodermal lineages, while BM-MSCs showed the most prominent immunosuppressive effect in vitro., Conclusions: The adoption of standardized culture conditions may help researchers and clinicians to reveal particular characteristics and inter-individual variability of MSCs sourced from different tissues. These data will be beneficial to set the standards for tissue collection and MSCs clinical-scale expansion both for cell banking and for cell-based therapy settings.

Alternatives to the use of fetal bovine serum (FBS) have been investigated to ensure xeno-free growth condition. In this study we evaluated the efficacy of human platelet lysate (PL) as a substitute of FBS for the in vitro culture of some human cell lines. PL was obtained by pools of pathogen inactivated human donor platelet (PLT) concentrates. Human leukemia cell lines (KG-1, K562, JURKAT, HL-60) and epithelial tumor cell lines (HeLa and MCF-7) were cultured with either FBS or PL. Changes in cell proliferation, viability, morphology, surface markers and cell cycle were evaluated for each cell line. Functional characteristics were analysed by drug sensitivity test and cytotoxicity assay. Our results demonstrated that PL can support growth and expansion of all cell lines, although the cells cultured in presence of PL experienced a less massive proliferation compared to those grown with FBS. We found a comparable percentage of viable specific marker-expressing cells in both conditions, confirming lineage fidelity in all cultures. Functionality assays showed that cells in both FBS- and PL-supported cultures maintained their normal responsiveness to adriamycin and NK cell-mediated lysis. Our findings indicate that PL is a feasible serum substitute for supporting growth and propagation of haematopoietic and epithelial cell lines with many advantages from a perspective of process standardization, ethicality and product safety.

Background: Mesenchymal stromal cells (MSCs) isolated from human umbilical cord tissue (UCT) can be considered the perfect candidates for cell-based therapies and regenerative medicine. UCT-derived MSCs can be cryogenically stored in cell banks and expanded as needed for therapeutic uses., Study Design and Methods: We developed a new method for UCT-MSC isolation, cryopreservation, and expansion, following all criteria required by a stem cell bank. UCT-MSCs were isolated either by manual dissociation (MM) or by a semiautomatic dissociation system (SAM). In both protocols UCTs were treated enzymatically using Type IV collagenase good manufacturing practices (GMP) graded and hyaluronidase (medicinal product). Isolated UCT-MSCs were cryopreserved and analyzed after thawing for phenotype; for proliferation rate; and for their osteogenic, adipogenic, and chondrogenic differentiation capabilities., Results: We found that SAM reduced the time of tissue enzyme exposure and enabled us to obtain a homogeneous single-cell suspension deprived of tissue fragments. The isolated cells in both groups showed high expression of MSC markers CD105, CD73, and CD90 and similar differentiation capabilities, phenotype, and proliferation potential. Moreover, the final yield of MSCs was comparable between the two techniques., Conclusion: In this study, we have established a reliable and standardized protocol to isolate UCT-MSCs from UCT for cell banking purposes. Processing the whole umbilical tissue with GMP-graded enzymes using a semiautomatic dissociator allowed us to obtain a single-cell suspension product with a known number of isolated cells that can be cryopreserved right after isolation and thawed as needed for expansion and clinical use., (© 2015 AABB.)

Recently, it was shown that the spectrum of anomalous dimensions and other important observables in planar N=4 supersymmetric Yang-Mills theory are encoded into a simple nonlinear Riemann-Hilbert problem: the Pμ system or quantum spectral curve. In this Letter, we extend this formulation to the N=6 supersymmetric Chern-Simons theory introduced by Aharony, Bergman, Jafferis, and Maldacena. This may be an important step towards the exact determination of the interpolating function h(λ) characterizing the integrability of this model. We also discuss a surprising relation between the quantum spectral curves for the N=4 supersymmetric Yang-Mills theory and the N=6 supersymmetric Chern-Simons theory considered here.

Background: Supplements to support clinical-grade cultures of mesenchymal stem cells (MSC) are required to promote growth and expansion of these cells. Platelet lysate (PL) is a human blood component which may replace animal serum in MSC cultures being rich in various growth factors. Here, we describe a plasma poor pathogen-free platelet lysate obtained by pooling 12 platelet (PLT) units, to produce a standardized and safe supplement for clinical-grade expansion of MSC., Methods: PL lots were obtained by combining 2 6-unit PLT pools in additive solution (AS) following a transfusional-based procedure including pathogen inactivation (PI) by Intercept technology and 3 cycles of freezing/thawing, followed by membrane removal. Three PI-PL and 3 control PL lots were produced to compare their ability to sustain bone marrow derived MSC selection and expansion. Moreover, two further PL, subjected to PI or not, were also produced starting from the same initial PLT pools to evaluate the impact of PI on growth factor concentration and capacity to sustain cell growth. Additional PI-PL lots were used for comparison with fetal bovine serum (FBS) on MSC expansion. Immunoregulatory properties of PI-PL-generated MSC were documented in vitro by mixed lymphocyte culture (MLC) and peripheral blood mononuclear cells (PBMC) mitogen induced proliferation., Results: PI-PL and PL control lots had similar concentrations of 4 well-described growth factors endowed with MSC stimulating ability. Initial growth and MSC expansion by PI-PL and PL controls were comparable either using different MSC populations or in head to head experiments. Moreover, PI-PL and PL control sustained similar MSC growth of frozen/thawed MSC. Multilineage differentiation of PI-derived and PI-PL-derived MSC were maintained in any MSC cultures as well as their immunoregulatory properties. Finally, no direct impact of PI on growth factor concentration and MSC growth support was observed, whereas the capacity of FBS to sustain MSC expansion in basic medium was irrelevant as compared to PL and PI-PL., Conclusion: The replacement of animal additives with human supplements is a basic issue in MSC ex vivo production. PI-PL represents a standardized, plasma-poor, human preparation which appears as a safe and good candidate to stimulate MSC growth in clinical-scale cultures.

Background Aims: We have recently shown that thymoglobulin (TG) efficiently expands cytokine-induced killer (CIK) cells in combination with interferon (IFN)-γ and interleukin (IL)-2 (ITG2 protocol). It is presently unknown whether the infusion of autologous immune effector cells generated by TG, IFN-γ and IL-2 is feasible and safe., Methods: Five patients with advanced and/or refractory solid tumors were enrolled in the present phase I/II study. Peripheral blood mononuclear cells (PBMC) collected by leukapheresis were stimulated under good manufacturing practice (GMP)-conditions with IFN-γ, followed by TG and IL-2. After 2-3 weeks in culture, a median of 4.65 × 10(6) immune effector cells per kilogram of recipient's body weight was obtained and infused intravenously. The median time from enrollment into the study to infusion of the expanded CIK cells was 30 days., Results: ITG2 efficiently expanded immune effector cells that comprised both conventional natural killer (NK) cells and CD3(+) CD16(+) CD56(+) CIK cells. One patient with advanced melanoma died because of disease progression before the infusion of CIK cells. The target dose of at least 2.5 × 10(6) CIK cells/kg of recipient's body weight was reached in four out of five evaluable patients. CIK cells were administered intravenously without any measurable toxicity. In vitro, CIK cells exerted lytic activity against cervical cancer cells. The median survival was 4.5 months (range 1-13) from the first infusion of CIK cells., Conclusions: This study has highlighted the feasibility and safety of the administration of CIK cells generated with the ITG2 protocol. Whether CIK cells can help control disease burden in patients with advanced malignancies will be determined in future clinical trials.

Introduction: Accurate white blood cell counting (WBC) and differential count by blood analyzers could allow a more informative characterization of granulocyte colony-stimulating factor (G-CSF) mobilized blood (MB), leukapheresis products (LP), and cord blood (CB). However, reliable counting by a blood cell analyzer in this setting is a major challenge owing to quali-quantitative abnormalities of blood cells., Methods: We evaluated the performances of the analyzer Pentra DX 120 by Horiba ABX working with dedicated cell-gating profiles, which generate three-part differential counts in samples obtained from donors' MB, LP, and CB. The results of the analyzer were compared to counts obtained by flow cytometry and manual counts, the latter performed for reference validation and in the case of discrepant results between study and reference counts., Results: Pentra DX 120 generated highly correlated counts (R > 0.91 in all cases) to those obtained by flow cytometry in all samples (MB, LP, and CB) with high degree of count accuracy in most cases and referred to WBC absolute count and differential count including lymphocytes (LYM) %, monocytes (MON) %, and polymorphonuclear leukocytes (PMN) %. Accuracy, judged by the difference between study and reference counts and expressed as percentage of reference count, ranged from 0.8% to 8.6%, and sporadic loss of accuracy occurred for MON % only in no more than 10% of CB samples., Conclusion: The ABX Pentra DX 120 provided accurate WBC count and differential count during MB, LP, and CB analyses and allowed a better characterization of donors' hematologic status and graft composition., (© 2011 Blackwell Publishing Ltd.)

Background: The aim of this study was to evaluate the quality of red blood cell concentrates obtained from donated whole blood, selected for transfusion therapy of thalassaemic patients, by measuring the following parameters: haemoglobin, haematocrit, percentage haemolysis, residual leucocyte count and residual protein content., Materials and Methods: Overall 345 red cell concentrates were evaluated, of which 205 had been filtered in-line pre-storage and washed and 140 were buffy coat-depleted and used within 2 days of collection. Of the buffy coat-depleted concentrates, 62 were leucodepleted and 78 washed and leucodepleted post-storage all within 2 days of collection. The off-line filters used for the leucodepletion were gamma-irradiated polyester with a pore size of 200 μm. The washing procedure was automated (Haemonetics ACP 215, Braintree, MA, USA). The haematological parameters were evaluated by a blood cell counter (Coulter, Ramsey, IL, USA) and the white blood cell count by cytofluorimetry (FACScan)., Results: Ninety-five percent (194/205) of the red cell concentrates that had been filtered pre-storage and washed, 92% (57/62) of the red cell concentrates that had been leucodepleted post-storage and 94% (73/78) of the those subjected to both treatments had normal values of haemoglobin (>40 g/unit), haematocrit (between 50-70%), percentage haemolysis (<0.8/unit), white cell count (<1×10(6)) and residual protein content (<0.5 g/L). Five percent (11/205) of the red cell concentrates that had been filtered pre-storage and washed, 8% (5/62) of those leucodepleted post-storage after 2 days and 6% (5/78) of those that underwent both procedures had a haemoglobin content <40 g/unit and a haematocrit <50%., Conclusions: The preparation procedures had been carried out satisfactorily; nevertheless, transfusion therapy with some "low dose" normal units could be less effective and might, therefore, result in greater transfusion requirements in patients receiving such units.

Background: Cytokine-induced killer (CIK) cells are typically differentiated in vitro with interferon (IFN)-γ and αCD3 monoclonal antibodies (mAb), followed by the repeated provision of interleukin (IL)-2. It is presently unknown whether thymoglobulin (TG), a preparation of polyclonal rabbit γ immunoglobulins directed against human thymocytes, can improve the generation efficiency of CIK cells compared with αCD3 mAb in a clinical-grade culture protocol., Methods: Peripheral blood mononuclear cells (PBMC) from 10 healthy donors and 4 patients with solid cancer were primed with IFN-γ on day 0 and low (50 ng/ml), intermediate (250 ng/ml) and high (500 ng/ml) concentrations of either αCD3 mAb or TG on day 1, and were fed with IL-2 every 3 days for 21 days. Aliquots of cells were harvested weekly to monitor the expression of representative members of the killer-like immunoglobulin receptor (KIR), NK inhibitory receptor, NK activating receptor and NK triggering receptor families. We also quantified the frequency of bona fide regulatory T cells (Treg), a T-cell subset implicated in the down-regulation of anti-tumor immunity, and tested the in vitro cytotoxic activity of CIK cells against NK-sensitive, chronic myeloid leukaemia K562 cells., Results: CIK cells expanded more vigorously in cultures supplemented with intermediate and high concentrations of TG compared with 50 ng/ml αCD3 mAb. TG-driven CIK cells expressed a constellation of NK activating/inhibitory receptors, such as CD158a and CD158b, NKp46, NKG2D and NKG2A/CD94, released high quantities of IL-12p40 and efficiently lysed K562 target cells. Of interest, the frequency of Treg cells was lower at any time-point compared with PBMC cultures nurtured with αCD3 mAb. Cancer patient-derived CIK cells were also expanded after priming with TG, but they expressed lower levels of the NKp46 triggering receptor and NKG2D activating receptor, thus manifesting a reduced ability to lyse K562 cells., Conclusions: TG fosters the generation of functional CIK cells with no concomitant expansion of tumor-suppressive Treg cells. The culture conditions described herein should be applicable to cancer-bearing individuals, although the differentiation of fully functional CIK cells may be hindered in patients with advanced malignancies.

Defects in chromosome segregation play a critical role in producing genomic instability and aneuploidy, which are associated with congenital diseases and carcinogenesis. We recently provided evidence from immunofluorescence and electron microscopy studies that merotelic kinetochore orientation is a major mechanism for lagging chromosomes during mitosis in PtK1 cells. Here we investigate whether human primary fibroblasts exhibit similar errors in chromosome segregation and if at least part of lagging chromosomes may arise in cells entering anaphase in the presence of mono-oriented chromosomes. By using in situ hybridization with alphoid probes to chromosome 7 and 11 we showed that loss of a single sister is much more frequent than loss of both sisters from the same chromosome in anatelophases from human primary fibroblasts released from a nocodazole-induced mitotic arrest, as predicted from merotelic orientation of single kinetochores. Furthermore, the lagging of pairs of separated sisters was higher than expected from random chance indicating that merotelic orientation of one sister may promote merotelic orientation of the other. Kinetochores of lagging chromosomes in anaphase human cells were found to be devoid of the mitotic checkpoint phosphoepitopes recognized by the 3F3/2 antibody, suggesting that they attached kinetochore microtubules prior to anaphase onset. Live cell imaging of H2B histone-GFP-transfected cells showed that cells with mono-oriented chromosomes never enter anaphase and that lagging chromosomes appear during anaphase after chromosome alignment occurs during metaphase. Thus, our results demonstrate that the mitotic checkpoint efficiently prevents the possible aneuploid burden due to mono-oriented chromosomes and that merotelic kinetochore orientation is a major limitation for accurate chromosome segregation and a potentially important mechanism of aneuploidy in human cells.

The scaling form of the free energy near a critical point allows for the definition of various thermodynamical amplitudes and the determination of their dependence on the microscopic nonuniversal scales. Universal quantities can be obtained by considering special combinations of the amplitudes. Together with the critical exponents they characterize the universality classes and may be useful quantities for their experimental identification. We compute the universal amplitude ratios for the tricritical Ising model in two dimensions by using several theoretical methods from perturbed conformal field theory and scattering integrable quantum field theory. The theoretical approaches are further supported and integrated by results coming from a numerical determination of the energy eigenvalues and eigenvectors of off-critical systems in an infinite cylinder.

We consider the universality class of the two-dimensional tricritical Ising model. The scaling form of the free energy leads to the definition of universal ratios of critical amplitudes which may have experimental relevance. We compute these universal ratios by a combined use of results coming from perturbed conformal field theory, integrable quantum field theory, and numerical methods.

In this work we have used the inhibitor of F-actin polymerisation cytochalasin B (Cyt B) to test the hypothesis that the contractile ring and the central spindle are mutually interdependent structures in mammalian mitotic cells. Double fluorescence staining of alpha-tubulin and F-actin was employed to analyse anaphase and telophase figures from asynchronously growing cultures and prometaphase-synchronised cells. Testing for the presence of the central spindle and contractile ring in human primary fibroblasts, human hepatoma cells and Chinese hamster cells after Cyt B treatment showed that both structures were simultaneously absent in over 60% of treated anaphases and 80% of telophases. Experiments on resumption of cytokinesis in cleavage-arrested cells further showed that Cyt B-treated human fibroblasts proceeded to cleavage within minutes after removal of the drug from the medium, concomitant with the re-formation of both cellular structures in cleaving cells. These data suggest that the presence of a correctly assembled contractile ring is essential for the formation and persistence of the central spindle during ana-telophase and provide further support for the idea of a strong co-operative interaction between these two structures during cytokinesis.

The activation and differentiation of resting B cells into Ig secreting cells are regulated by T cells, macrophages and their secreted factors. The present study evaluated the effect of cyclosporin A (CsA) on this process. Peripheral blood lymphomonocytes (PBMC) drawn from healthy donors were stimulated with protein A (PA) or with lipopolysaccharides plus pokeweed (LPS+PWM) in either the presence or the absence of CsA. Phenotypic B cell changes and immunoglobulin production was then analyzed. The data revealed that CsA decreased the expression of B cell surface receptors of the activation phase, and enhanced the resting phase receptors. Different effects of CsA were found on B cell differentiation, depending on its induction by PA or LPS+PWM. In the first system, CsA decreased the expression of differentiation phase receptors and the secretion of free Ig. In cultures stimulated with LPS+PWM, CsA increased the differentiated phase receptors and Ig secretion. Thus, CsA seemed to act as a blocking agent of the activation phase and as a modulator of the differentiation phase and of IgG secretion, depending upon the antigen used for stimulation.

The blastogenic response to mitogens of peripheral blood mononuclear cells (PBMC) obtained from healthy volunteers and patients with chronic or acute tuberculosis (TB) was evaluated. Cells derived from TB patients showed a reduced proliferative capacity compared to that of healthy individuals. Three possible causes of such an impairment were investigated, namely: 1) a change in the proportion of lymphocyte subpopulations; 2) an altered ratio between monocytes and lymphocytes and 3) a reduction in the state of monocyte-macrophage activation, with an impaired production of interleukin-1 (IL-1). We observed no significant modification of lymphocyte subsets from TB patients and normal individuals. However, the relative number of monocytes in the patients was always higher than the controls. Furthermore, circulating monocytes from the patients with TB exhibited a decreased phagocytosis of latex beads, a normal expression of DR antigens, and an increased spontaneous production of IL-1. The possibility that the hyperactivation of macrophages may be responsible for the observed low blastogenic response is discussed.

The proliferation and development of cytotoxic T cells was investigated in human peripheral blood mononuclear cell (PBMC) cultures stimulated with an antigenic extract from Candida albicans (MPPS), or with the purified protein derivative from Mycobacterium tuberculosis (PPD), or with human recombinant interleukin 2 (rIL-2). Microbial antigen- and rIL-2-induced cytotoxic T cells were able to lyse both natural killer (NK) sensitive and resistant targets. No correlation was observed between the development of T cell cytotoxicity and interferon (IFN) production in vitro. The addition of anti-class II monoclonal antibodies at the beginning of MPPS/PPD-stimulated cultures inhibited the cell proliferation, IFN production and T cell cytotoxicity, while all these cellular activities were not inhibited by anti-class II antibodies in rIL-2-stimulated cultures. Finally, antibodies to class I determinants inhibit T cell cytotoxicity, suggesting a role of such determinants in the development of the non-adaptive immunity to microbial infections.