KARINA INACIO LADISLAU DE CARVALHO SALMAZI
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- Tooth Tissue Engineering: The Influence of Hydrophilic Surface on Nanocrystalline Diamond Films for Human Dental Stem Cells(2013) DUAILIBI, Silvio Eduardo; DUAILIBI, Monica Talarico; FERREIRA, Lydia Masako; SALMAZI, Karina Inacio Ladislau Carvalho; SALVADORI, Maria Cecilia; TEIXEIRA, Fernanda de Sa; PASQUARELLI, Alberto; VACANTI, Joseph Phillip; YELICK, Pamela CrottyNew techniques for tissue engineering (TE) are rapidly emerging. The basic concept of autologous TE is to isolate cells from small biopsy specimens, and to expand these cells in culture for subsequent seeding onto biodegradable scaffolds. Nanocrystalline diamond films have attracted the attention of researchers from a variety of different areas in recent years, due to their unique and exceptional properties. In this approach, human dental stem cells (hDSCs) were characterized by flow cytometry and grown on diamond films with hydrogen (H)-terminated and oxygen (O)-terminated surfaces for 28 days, and then removed by lysis and washing with distilled water. Energy dispersive spectroscopy analysis was performed, showing that the regions with O-terminated surfaces contained much higher levels of deposited calcium, oxygen, and phosphorus. These results suggest that the extracellular matrix was considerably more developed in the O-terminated regions, as compared with the H-terminated regions. In addition, optical microscopy of hDSCs cultured on the diamond substrate with H- and O-terminated surfaces, before washing with distilled water, showed preferential directions of the cells arrangement, where orthogonal lines suggest that the cells appeared to be following the O-terminated regions or hydrophilic surface. These findings suggest that O-terminated diamond surfaces prepared on biodegradable scaffolds can be useful for mineralized dental tissue formation.
- CD8+T Lymphocyte Expansion, Proliferation and Activation in Dengue Fever(2015) MATOS, Andreia Manso de; CARVALHO, Karina Inacio; ROSA, Daniela Santoro; VILLAS-BOAS, Lucy Santos; SILVA, Wanessa Cardoso da; RODRIGUES, Celia Luiza de Lima; OLIVEIRA, Olimpia Massae Nakasone Peel Furtado; LEVI, Jose Eduardo; ARAUJO, Evaldo Stanislau Affonso; PANNUTI, Claudio Sergio; LUNA, Expedito Jose Albuquerque; KALLAS, Esper GeorgeDengue fever induces a robust immune response, including massive T cell activation. The level of T cell activation may, however, be associated with more severe disease. In this study, we explored the level of CD8+ T lymphocyte activation in the first six days after onset of symptoms during a DENV2 outbreak in early 2010 on the coast of Sao Paulo State, Brazil. Using flow cytometry we detected a progressive increase in the percentage of CD8+ T cells in 74 dengue fever cases. Peripheral blood mononuclear cells from 30 cases were thawed and evaluated using expanded phenotyping. The expansion of the CD8+ T cells was coupled with increased Ki67 expression. Cell activation was observed later in the course of disease, as determined by the expression of the activation markers CD38 and HLA-DR. This increased CD8+ T lymphocyte activation was observed in all memory subsets, but was more pronounced in the effector memory subset, as defined by higher CD38 expression. Our results show that most CD8+ T cell subsets are expanded during DENV2 infection and that the effector memory subset is the predominantly affected sub population.
- Variability of HIV-1 Genomes among Children and Adolescents from Sao Paulo, Brazil(2013) SANABANI, Sabri Saeed; PESSOA, Rodrigo; OLIVEIRA, Ana Carolina Soares de; MARTINEZ, Vanessa Pouza; GIRET, Maria Teresa Maidana; SUCCI, Regina Celia de Menezes; CARVALHO, Karina; TOMIYAMA, Claudia Satiko; NIXON, Douglas F.; SABINO, Ester Cerdeira; KALLAS, Esper GeorgesBackground: Genetic variability is a major feature of the human immunodeficiency virus type 1 (HIV-1) and considered the key factor to frustrating efforts to halt the virus epidemic. In this study, we aimed to investigate the genetic variability of HIV-1 strains among children and adolescents born from 1992 to 2009 in the state of Sao Paulo, Brazil. Methodology: Plasma and peripheral blood mononuclear cells (PBMC) were collected from 51 HIV-1-positive children and adolescents on ART followed between September 1992 and July 2009. After extraction, the genetic materials were used in a polymerase chain reaction (PCR) to amplify the viral near full length genomes (NFLGs) from 5 overlapped fragments. NFLGs and partial amplicons were directly sequenced and data were phylogenetically inferred. Results: Of the 51 samples studied, the NFLGs and partial fragments of HIV-1 from 42 PBMCs and 25 plasma were successfully subtyped. Results based on proviral DNA revealed that 22 (52.4%) patients were infected with subtype B, 16 (38.1%) were infected with BF1 mosaic variants and 4 (9.5%) were infected with sub-subtype F1. All the BF1 recombinants were unique and distinct from any previously identified unique or circulating recombinant forms in South America. Evidence of dual infections was detected in 3 patients coinfected with the same or distinct HIV-1 subtypes. Ten of the 31 (32.2%) and 12 of the 21 (57.1%) subjects with recovered proviral and plasma, respectively, protease sequences were infected with major mutants resistant to protease inhibitors. The V3 sequences of 14 patients with available sequences from PBMC/or plasma were predicted to be R5-tropic virus except for two patients who harbored an X4 strain. Conclusions: The high proportion of HIV-1 BF1 recombinant, coinfection rate and vertical transmission in Brazil merits urgent attention and effective measures to reduce the transmission of HIV among spouses and sex partners.
- IVIg Immune Reconstitution Treatment Alleviates the State of Persistent Immune Activation and Suppressed CD4 T Cell Counts in CVID(2013) PAQUIN-PROULX, Dominic; SANTOS, Bianca A. N.; CARVALHO, Karina I.; TOLEDO-BARROS, Myrthes; OLIVEIRA, Ana Karolina Barreto de; KOKRON, Cristina M.; KALIL, Jorge; MOLL, Markus; KALLAS, Esper G.; SANDBERG, Johan K.Common variable immunodeficiency (CVID) is characterized by defective B cell function, impaired antibody production, and increased susceptibility to bacterial infections. Here, we addressed the hypothesis that poor antibody-mediated immune control of infections may result in substantial perturbations in the T cell compartment. Newly diagnosed CVID patients were sampled before, and 6-12 months after, initiation of intravenous immunoglobulin (IVIg) therapy. Treatment-naive CVID patients displayed suppressed CD4 T cell counts and myeloid dendritic cell (mDC) levels, as well as high levels of immune activation in CD8 T cells, CD4 T cells, and invariant natural killer T (iNKT) cells. Expression of co-stimulatory receptors CD80 and CD83 was elevated in mDCs and correlated with T cell activation. Levels of both FoxP3+ T regulatory (Treg) cells and iNKT cells were low, whereas soluble CD14 (sCD14), indicative of monocyte activation, was elevated. Importantly, immune reconstitution treatment with IVIg partially restored the CD4 T cell and mDC compartments. Treatment furthermore reduced the levels of CD8 T cell activation and mDC activation, whereas levels of Treg cells and iNKT cells remained low. Thus, primary deficiency in humoral immunity with impaired control of microbial infections is associated with significant pathological changes in cell-mediated immunity. Furthermore, therapeutic enhancement of humoral immunity with IVIg infusions alleviates several of these defects, indicating a relationship between poor antibody-mediated immune control of infections and the occurrence of abnormalities in the T cell and mDC compartments. These findings help our understanding of the immunopathogenesis of primary immunodeficiency, as well as acquired immunodeficiency caused by HIV-1 infection.
- HTLV-1 Tax Specific CD8+ T Cells Express Low Levels of Tim-3 in HTLV-1 Infection: Implications for Progression to Neurological Complications(2011) NDHLOVU, Lishomwa C.; LEAL, Fabio E.; HASENKRUG, Aaron M.; JHA, Aashish R.; CARVALHO, Karina I.; ECCLES-JAMES, Ijeoma G.; BRUNO, Fernanda R.; VIEIRA, Raphaella G. S.; YORK, Vanessa A.; CHEW, Glen M.; JONES, R. Brad; TANAKA, Yuetsu; NETO, Walter K.; SANABANI, Sabri S.; OSTROWSKI, Mario A.; SEGURADO, Aluisio C.; NIXON, Douglas F.; KALLAS, Esper G.The T cell immunoglobulin mucin 3 (Tim-3) receptor is highly expressed on HIV-1-specific T cells, rendering them partially ""exhausted'' and unable to contribute to the effective immune mediated control of viral replication. To elucidate novel mechanisms contributing to the HTLV-1 neurological complex and its classic neurological presentation called HAM/TSP (HTLV-1 associated myelopathy/tropical spastic paraparesis), we investigated the expression of the Tim-3 receptor on CD8(+) T cells from a cohort of HTLV-1 seropositive asymptomatic and symptomatic patients. Patients diagnosed with HAM/TSP down-regulated Tim-3 expression on both CD8(+) and CD4(+) T cells compared to asymptomatic patients and HTLV-1 seronegative controls. HTLV-1 Tax-specific, HLA-A*02 restricted CD8(+) T cells among HAM/TSP individuals expressed markedly lower levels of Tim-3. We observed Tax expressing cells in both Tim-3(+) and Tim-3(-) fractions. Taken together, these data indicate that there is a systematic downregulation of Tim-3 levels on T cells in HTLV-1 infection, sustaining a profoundly highly active population of potentially pathogenic T cells that may allow for the development of HTLV-1 complications.
- Proteins of Leishmania (Viannia) shawi confer protection associated with Th1 immune response and memory generation(2012) PASSERO, Luiz Felipe D.; CARVALHO, Ana Kely; BORDON, Maria L. A. C.; BONFIM-MELO, Alexis; CARVALHO, Karina; KALLAS, Esper G.; SANTOS, Bianca B. A.; TOYAMA, Marcos H.; PAES-LEME, Adriana; CORBETT, Carlos E. P.; LAURENTI, Marcia D.Background: Leishmania (Viannia) shawi parasite was first characterized in 1989. Recently the protective effects of soluble leishmanial antigen (SLA) from L. (V.) shawi promastigotes were demonstrated using BALB/c mice, the susceptibility model for this parasite. In order to identify protective fractions, SLA was fractionated by reverse phase HPLC and five antigenic fractions were obtained. Methods: F1 fraction was purified from L. (V.) shawi parasite extract by reverse phase HPLC. BALB/c mice were immunized once a week for two consecutive weeks by subcutaneous routes in the rump, using 25 mu g of F1. After 1 and 16 weeks of last immunization, groups were challenged in the footpad with L. (V.) shawi promastigotes. After 2 months, those same mice were sacrificed and parasite burden, cellular and humoral immune responses were evaluated. Results: The F1 fraction induced a high degree of protection associated with an increase in IFN-gamma, a decrease in IL-4, increased cell proliferation and activation of CD8(+)T lymphocytes. Long-term protection was acquired in F1-immunized mice, associated with increased CD4(+) central memory T lymphocytes and activation of both CD4+ and CD8(+) T cells. In addition, F1-immunized groups showed an increase in IgG2a levels. Conclusions: The inductor capability of antigens to generate memory lymphocytes that can proliferate and secrete beneficial cytokines upon infection could be an important factor in the development of vaccine candidates against American Tegumentary Leishmaniasis.
- Subsets of Memory CD4(+) T Cell and Bactericidal Antibody Response to Neisseria meningitidis Serogroup C after Immunization of HIV-Infected Children and Adolescents(2014) MILAGRES, Lucimar G.; COSTA, Priscilla R.; SILVA, Giselle P.; CARVALHO, Karina I.; PEREIRA-MANFRO, Wania F.; FERREIRA, Bianca; BARRETO, Daniella M.; FROTA, Ana Cristina C.; HOFER, Cristina B.; KALLAS, Esper G.Meningococcal disease is endemic in Brazil, with periodic outbreaks and case fatality rates reach as high as 18 to 20% of cases. Conjugate vaccines against meningococci are immunogenic in healthy children. However, we have previously shown a poor bactericidal antibody response to a Men C conjugate vaccine in Brazilian HIV-infected children and adolescents after a single vaccine administration. The goal of the present work was to investigate associations between bactericidal antibody response induced by MenC vaccine and the frequency and activation profile (expression of CD38, HLA-DR and CCR5 molecules) of total CD4(+) memory T cell sub-populations in HIV-1-infected children and adolescents. Responders to vaccination against MenC had a predominance (about 44%) of CD4(+) T-INTERMEDIATE subset followed by T-TRANSITIONAL memory subset (23 to 26%). Importantly, CD4(+) T-INT frequency was positively associated with bactericidal antibody response induced by vaccination. The positive correlation persisted despite the observation that the frequency T-INT CD38(+)HLA-DR+ was higher in responders. In contrast, CD4(+) T-CENTRAL MEMORY (T-CM) subset negatively correlated with bactericidal antibodies. In conclusion, these data indicate that less differentiated CD+ T cells, like TCM may be constantly differentiating into intermediate and later differentiated CD4(+) T cell subsets. These include CD4 T-INT subset which showed a positive association with bactericidal antibodies.
- Imbalanced expression of functional surface molecules in regulatory and effector T cells in systemic lupus erythematosus(2014) MESQUITA JUNIOR, D.; CRUVINEL, W. M.; ARAUJO, J. A. P.; SALMAZI, K. C.; KALLAS, E. G.; ANDRADE, L. E. C.Regulatory T (TREG) cells play an important role in maintaining immune tolerance and avoiding autoimmunity. We analyzed the expression of membrane molecules in TREG and effector T cells in systemic lupus erythematosus (SLE). TREG and effector T cells were analyzed for the expression of CTLA-4, PD1, CD28, CD95, GITR, HLA-DR, OX40, CD40L, and CD45RO in 26 patients with active disease, 31 with inactive disease, and 26 healthy controls. TREG cells were defined as CD25(+/high)CD127 (circle divide/low)FoxP3(+), and effector T cells were defined as CD25(+) CD127(+) FoxP3(circle divide). The ratio of TREG to effector T cells expressing GITR, PD1, HLA-DR, OX40, CD40L, and CD45RO was determined in the three groups. The frequency of TREG cells was similar in patients with SLE and controls. However, SLE patients had a decreased frequency of CTLA-4(+)TREG and CD28(+) TREG cells and an increased frequency of CD40L(+)TREG cells. There was a decrease in the TREG/effector-T ratio for GITR(+), HLA-DR+, OX40(+), and CD45RO(+) cells, and an increased ratio of TREG/ effector-T CD40L(+) cells in patients with SLE. In addition, CD40L(+)TREG cell frequency correlated with the SLE disease activity index (P=0.0163). In conclusion, our findings showed several abnormalities in the expression of functionally critical surface molecules in TREG and effector T cells in SLE that may be relevant to the pathogenesis of this disease.