Use of stem cells from mismatched related donors

Mismatched haploidentical bone marrow transplantations from a related donor have been the topic of clinical and laboratory research for more than 20 years. During that time, new treatment strategies have been designed based on animal experiments, and, since our group introduced the megadose inoculum which combines T-cell-depleted bone marrow cells with a large number of granulocyte colony-stimulating factor-mobilized peripheral blood stem cells and a more intensive conditioning regimen, have done much to overcome the problems of graft-versus-host disease and graft rejection. As most patients have a full haplotype mismatched relative available, this technique means that a far greater number of patients with hematologic malignancies can be offered a T-cell-depleted transplantation as curative therapy.     

Direct Methods Optimised for Solving Crystal Structure by Powder Diffraction Data: Limits,Strategies, and Prospects

The ab-initio crystal structure solution by powder diffraction data requires great efforts because of the collapse of the experimental information onto the one dimensional 2θ axis of the pattern. Different strategies will be described aiming at improving the process of extraction of the integrated intensities from the experimental pattern in order to make more straightforward the structure solution process by direct methods. Particular attention will be devoted to the EXPO program. Some of its performance will be analysed and results will be shown.     

The X-linked lymphoproliferative-disease gene product SAP regulates signals induced through the co-receptor SLAM

In addition to triggering the activation of B- or T-cell antigen receptors, the binding of a ligand to its receptor at the cell surface can sometimes determine the physiological outcome of interactions between antigen-presenting cells, T and B lymphocytes. The protein SLAM (also known as CDw150), which is present on the surface of B and T cells, forms such a receptor-ligand pair as it is a self-ligand. We now show that a T-cell-specific, SLAM-associated protein (SAP), which contains an SH2 domain and a short tall, acts as an inhibitor by blocking recruitment of the SH2-domain-containing signal-transduction molecule SHP-2 to a docking site in the SLAM cytoplasmic region. The gene encoding SAP maps to the same area of the X chromosome as the locus for X-linked lymphoproliferative disease (XLP) and we found mutations in the SAP gene in three XLP patients. Absence of the inhibitor SAP in XLP patients affects T/B-cell interactions induced by SLAM, leading to an inability to control B-cell proliferation caused by Epstein-Barr virus infections.     

Successful engraftment of T-cell-depleted haploidentical "three-loci" incompatible transplants in leukemia patients by addition of recombinant human granulocyte colony-stimulating factor-mobilized peripheral blood progenitor cells to bone marrow inoculum

Patients who undergo transplantation with haploidentical "three-loci" mismatched T-cell-depleted bone marrow (BM) are at high risk for graft failure. To overcome the host-versus-graft barrier, we increased the size of the graft inoculum, which has been shown to be a major factor in controlling both immune rejection and stem cell competition in murine models. Seventeen patients (mean age, 23.2 years; range, 6 to 51 years) with end-stage chemoresistant leukemia were received transplants of a combination of BM with recombinant human granulocyte colony-stimulating factor-mobilized peripheral blood progenitor cells from HLA-haploidentical "three-loci" incompatible family members. The average concentration of colony-forming unit-granulocyte-macrophage in the final inoculum was sevenfold to 10-fold greater than that found in BM alone. The sole graft-versus-host disease (GVHD) prophylaxis consisted of T-cell depletion of the graft by the soybean agglutination and E-rosetting technique. The conditioning regimen included total body irradiation in a single fraction at a fast dose rate, antithymocyte globulin, cyclophosphamide and thiotepa to provide both immunosuppression and myeloablation. One patient rejected the graft and the other 16 had early and sustained full donor-type engraftment. One patient who received a much greater quantity of T lymphocytes than any other patient died from grade IV acute GVHD. There were no other cases of GVHD > or = grade II. Nine patients died from transplant-related toxicity, 2 relapsed, and 6 patients are alive and event-free at a median follow-up of 230 days (range, 100 to 485 days). Our results show that a highly immunosuppressive and myeloablative conditioning followed by transplantation of a large number of stem cells depleted of T lymphocytes by soybean agglutination and E-rosetting technique has made transplantation of three HLA-antigen disparate grafts possible, with only rare cases of GVHD.     

Even in virtual environments women shop and men build: A social role perspective on Second Life

The present study examined whether traditional gender role expectations (Eagly, 1987) influence behaviors in non-traditional contexts such as online virtual environments. Participants were 352 Second Life users who reported their activities and experiences in Second Life. Results indicated that men and women differed in the types of activities they engaged in a manner predicted by social role theory. Specifically, as compared to women, men were more likely to report building things (e.g. objects), to own and work on their own virtual property, and were less likely to change their avatar’s appearance. Women, as compared to men, were more likely to meet people, shop, regularly change their avatar’s appearance, and buy clothes/objects for their avatar. The present study adds to our understanding of how traditional gender role expectations may carry over to online virtual worlds and influence online behavior.     

Social evaluations of embodied agents and avatars

The purpose of this study was to examine social evaluations (i.e., perceptions of empathy and positivity) following peoples’ interactions with digital human representations. Female research participants engaged in a 3-min interaction while immersed in a 3-D immersive virtual environment with a “peer counselor.” Participants were led to believe that the peer counselor was either an embodied agent (i.e., computer algorithm) or an avatar (i.e., another person). During the interaction, the peer counselor either smiled or not. As predicted, a digitally-rendered smile was found to affect participants’ social evaluations. However, these effects were moderated by participants’ beliefs about their interaction partner. Specifically, smiles enhanced social evaluations of embodied agents but degraded them for avatars. Although these results are consistent with other findings concerning the communicative realism of embodied agents and avatars they uniquely demonstrate that people’s beliefs alone, rather than actual differences in virtual representations, can impact social evaluations.     

Serratiopeptidase Affects the Physiology of Pseudomonas aeruginosa Isolates from Cystic Fibrosis Patients

Pseudomonas aeruginosa is frequently involved in cystic fibrosis (CF) airway infections. Biofilm, motility, production of toxins and the invasion of host cells are different factors that increase P. aeruginosa’s virulence. The sessile phenotype offers protection to bacterial cells and resistance to antimicrobials and host immune attacks. Motility also contributes to bacterial colonization of surfaces and, consequently, to biofilm formation. Furthermore, the ability to adhere is the prelude for the internalization into lung cells, a common immune evasion mechanism used by most intracellular bacteria, such as P. aeruginosa. In previous studies we evaluated the activity of metalloprotease serratiopeptidase (SPEP) in impairing virulence-related properties in Gram-positive bacteria. This work aimed to investigate SPEP’s effects on different physiological aspects related to the virulence of P. aeruginosa isolated from CF patients, such as biofilm production, pyoverdine and pyocyanin production and invasion in alveolar epithelial cells. Obtained results showed that SPEP was able to impair the attachment to inert surfaces as well as adhesion/invasion of eukaryotic cells. Conversely, SPEP’s effect on pyocyanin and pyoverdine production was strongly strain-dependent, with an increase and/or a decrease of their production. Moreover, SPEP seemed to increase swarming motility and staphylolytic protease production. Our results suggest that a large number of clinical strains should be studied in-depth before drawing definitive conclusions. Why different strains sometimes react in opposing ways to a specific treatment is of great interest and will be the object of future studies. Therefore, SPEP affects P. aeruginosa’s physiology by differently acting on several bacterial factors related to its virulence.     

Occurrence of non-lactic acid bacteria populations involved in protein hydrolysis of cold-stored high moisture Mozzarella cheese

The aim of this study was to analyse non-lactic acid bacteria populations (NLABPs) and evaluate their role in proteolysis of cold-stored high moisture (HM) Mozzarella cheese. NLABPs reached values close to 8 log cfu mL⁻¹ after seven days of cold storage. Sequencing of 16 rDNA and rpoB genes and molecular biotyping allowed to identify 66 bacterial strains belonging to 25 species from 15 genera, mainly represented by Pseudomonas, Acinetobacter, and Rahnella. Fifteen strains showed proteolytic activity values higher than 1000.00 μg Gly mL⁻¹ after 24 h of growth in skimmed milk. Moreover, as shown by Urea-PAGE, 11 proteolytic strains caused partial or total disappearance of at least one of the caseins. Their proteolytic behaviour was assessed even when they grew inside the governing liquid together with HM Mozzarella cheese at 4 °C for 12 days. This is the first report that throws light on the complexity of NLABPs in HM Mozzarella cheese, demonstrating that some strains caused the partial hydrolysis of α, β, and γ caseins on its outer surface where a concomitant wrinkling and successive exfoliation became visible without significant changes in texture characteristics.