Cinética de muerte de linfocitos en diabetes mellitus tipo 1 y su posible asociación con miR-146a y miR-155 / Kinetics of cell death in type 1 diabetes mellitus and their possible association with miR-146a and miR-155

Aim: Analyze mi-146a and miR-155 expression and its correlation with the apoptosis of lymphocytes T in T1D and control patient. Patients and Methodology: 17 T1D patients (5 children between 8-14 yr and 12 adults between 19-29 yr). Activated and not activated peripheral mononuclear cells were studied were studied. Cellular activation with anti-CD3 and primary culture with interleukyne-2 by 5 days. Apoptosis assays through flow cytometry. miRNA through Taqman probes. Statistical analysis through Kruskal-Wallis and post-hoc Dunn's test. Results: Composition of virgin and memory T CD4 cells showed significant differences for stimulus response in control group (p = 0,0004). Increased memory cells count in control group activated by 7 days than basal (p = 0,0047). For early apoptosis differences were observed in days 3 and 7 with and without activation (p = 0,001). AICD apoptosis showed increases in control group after re-stimulation through TCR (p= 0,03). miR-146a expression was lower in recent-onset T1D children vs recent-onset DM1 adults (p = 0,0167). Conclusion: This study shows a differential miR-146a expression in T1D children with respect to T1D adult patients, diminished AICD mechanism in T1D and altered CD4+CD45RA-CD45R0+ memory cells generation in T1D adult patients.

Radiotherapy and immune response: the systemic effects of a local treatment

Technological developments have allowed improvements in radiotherapy delivery, with higher precision and better sparing of normal tissue. For many years, it has been well known that ionizing radiation has not only local action but also systemic effects by triggering many molecular signaling pathways. There is still a lack of knowledge of this issue. This review focuses on the current literature about the effects of ionizing radiation on the immune system, either suppressing or stimulating the host reactions against the tumor, and the factors that interact with these responses, such as the radiation dose and dose / fraction effects in the tumor microenvironment and vasculature. In addition, some implications of these effects in cancer treatment, mainly in combined strategies, are addressed from the perspective of their interactions with the more advanced technology currently available, such as heavy ion therapy and nanotechnology.

BJ-1108, a 6-Amino-2,4,5-trimethylpyridin-3-ol analogue, regulates differentiation of Th1 and Th17 cells to ameliorate experimental autoimmune encephalomyelitis

BACKGROUND: CD4+ T cells play an important role in the initiation of an immune response by providing help to other cells. Among the helper T subsets, interferon-γ (IFN-γ)-secreting T helper 1 (Th1) and IL-17-secreting T helper 17 (Th17) cells are indispensable for clearance of intracellular as well as extracellular pathogens. However, Th1 and Th17 cells are also associated with pathogenesis and contribute to the progression of multiple inflammatory conditions and autoimmune diseases. RESULTS: In the current study, we found that BJ-1108, a 6-aminopyridin-3-ol analogue, significantly inhibited Th1 and Th17 differentiation in vitro in a concentration-dependent manner, with no effect on proliferation or apoptosis of activated T cells. Moreover, BJ-1108 inhibited differentiation of Th1 and Th17 cells in ovalbumin (OVA)-specific OT II mice. A complete Freund's adjuvant (CFA)/OVA-induced inflammatory model revealed that BJ-1108 can reduce generation of proinflammatory Th1 and Th17 cells. Furthermore, in vivo studies showed that BJ-1108 delayed onset of disease and suppressed experimental autoimmune encephalomyelitis (EAE) disease progression by inhibiting differentiation of Th1 and Th17 cells. CONCLUSIONS: BJ-1108 treatment ameliorates inflammation and EAE by inhibiting Th1 and Th17 cells differentiation. Our findings suggest that BJ-1108 is a promising novel therapeutic agent for the treatment of inflammation and autoimmune disease.

Total knee replacement induces peripheral blood lymphocytes apoptosis and it is not prevented by regional anesthesia - a randomized study / Artroplastia total do joelho induz apoptose em linfócitos de sangue periférico e não é evitada por anestesia regional - Estudo randomizado

ABSTRACT BACKGROUND: Among the many changes caused by a surgical insult one of the least studied is postoperative immunosuppression. This phenomenon is an important cause of infectious complications of surgery such as surgical site infection or hospital acquired pneumonia. One of the mechanisms leading to postoperative immunosuppression is the apoptosis of immunological cells. Anesthesia during surgery is intended to minimize harmful changes and maintain perioperative homeostasis. The aim of the study was evaluation of the effect of the anesthetic technique used for total knee replacement on postoperative peripheral blood lymphocyte apoptosis. METHODS: 34 patients undergoing primary total knee replacement were randomly assigned to two regional anesthetic protocols: spinal anesthesia and combined spinal-epidural anesthesia. 11 patients undergoing total knee replacement under general anesthesia served as control group. Before surgery, immediately after surgery, during first postoperative day and seven days after the surgery venous blood samples were taken and the immunological status of the patient was assessed with the use of flow cytometry, along with lymphocyte apoptosis using fluorescent microscopy. RESULTS: Peripheral blood lymphocyte apoptosis was seen immediately in the postoperative period and was accompanied by a decrease of the number of T cells and B cells. There were no significant differences in the number of apoptotic lymphocytes according to the anesthetic protocol. Changes in the number of T CD3/8 cells and the number of apoptotic lymphocytes were seen on the seventh day after surgery. CONCLUSION: Peripheral blood lymphocyte apoptosis is an early event in the postoperative period that lasts up to seven days and is not affected by the choice of the anesthetic technique.

RESUMO JUSTIFICATIVA E OBJETIVO: Dentre as muitas alterações causadas por uma ferida cirúrgica, uma das menos estudadas é a imunossupressão pós-operatória. Esse fenômeno é uma causa importante das complicações infecciosas relacionadas à cirurgia, como infecção do sítio cirúrgico ou pneumonia nosocomial. Um dos mecanismos que levam à imunossupressão pós-operatória é a apoptose de células imunológicas. Durante a cirurgia, a anestesia se destina a minimizar as alterações prejudiciais e manter a homeostase perioperatória. O objetivo deste estudo foi avaliar o efeito da técnica anestésica usada para artroplastia total de joelho sobre a apoptose em linfócitos de sangue periférico no pós-operatório. MÉTODOS: Trinta e quatro pacientes submetidos à artroplastia total primária de joelho foram randomicamente designados para dois protocolos de anestesia regional: raquianestesia e bloqueio combinado raqui-peridural. Onze pacientes submetidos à artroplastia total do joelho sob anestesia geral formaram o grupo controle. Antes da cirurgia, logo após a cirurgia, durante o primeiro dia de pós-operatório e sete dias após a cirurgia, amostras de sangue venoso foram colhidas e o estado imunológico do paciente foi avaliado com o uso deflow cysts 87 m, juntamente com apoptose de linfócitos com o uso de microscopia de fluorescência. RESULTADOS: Apoptose em linfócitos de sangue periférico foi observada imediatamente no pós-operatório e acompanhada por uma redução do número de células T e B. Não houve diferença significativa no número de linfócitos apoptóticos de acordo com o protocolo anestésico. Alterações no número de células T CD3/8 e no número de linfócitos apoptóticos foram observadas no sétimo dia após a cirurgia. CONCLUSÃO: Apoptose em linfócitos de sangue periférico é um evento precoce no período pós-operatório que dura até sete dias e não é afetado pela escolha da técnica anestésica.

Early surgical complications after gastric by-pass: a literature review / Complicações cirúrgicas precoces após bypass gástrico: revisão da literatura

INTRODUCTION: Gastric bypass is today the most frequently performed bariatric procedure,but, despite of it, several complications can occur with varied morbimortality. Probably all bariatric surgeons know these complications, but, as bariatric surgery continues to spread, general surgeon must be familiarized to it and its management. Gastric bypass complications can be divided into two groups: early and late complications, taking into account the two weeks period after the surgery. This paper will focus the early ones. METHOD: Literature review was carried out using Medline/PubMed, Cochrane Library, SciELO, and additional information on institutional sites of interest crossing the headings: gastric bypass AND complications; follow-up studies AND complications; postoperative complications AND anastomosis, Roux-en-Y; obesity AND postoperative complications. Search language was English. RESULTS: There were selected 26 studies that matched the headings. Early complications included: anastomotic or staple line leaks, gastrointestinal bleeding, intestinal obstruction and incorrect Roux limb reconstruction. CONCLUSION: Knowledge on strategies on how to reduce the risk and incidence of complications must be acquired, and every surgeon must be familiar with these complications in order to achieve an earlier recognition and perform the best intervention. .

INTRODUÇÃO: O bypass gástrico é hoje o procedimento bariátrico mais realizado, mas, apesar disso, várias complicações podem ocorrer com variada morbimortalidade. Provavelmente todos os cirurgiões bariátricos conhecem essas complicações, mas como a cirurgia bariátrica continua a se espalhar, o cirurgião geral deve estar familiarizado com essas complicações e seu manuseio. As complicações do bypass gástrico podem ser divididas em dois grupos: as precoces e tardias, tendo em conta o período de duas semanas após a operação. Este artigo irá focar as precoces. MÉTODO: Foi realizada revisão da literatura utilizando as bases Medline/PubMed, Cochrane Library, SciELO, e informações adicionais sobre sites institucionais de interesse cruzando os descritores: bypass gástrico AND complicações; seguimento AND complicações; complicações pós-operatórias AND anastomose, Roux-en-Y; obesidade AND complicações pós-operatórias. A língua usada para a busca foi o inglês. RESULTADOS: Foram selecionados 26 artigos que combinavam com os descritores. As complicações imediatas foram: fístula na linha de grampeamento, sangramento gastrointestinal, obstrução intestinal e reconstrução incorreta da alça em Roux. CONCLUSÃO: O conhecimento sobre as estratégias de como reduzir o risco e incidência das complicações deve ser adquirido ao longo do tempo, e cada cirurgião deve estar familiarizado com essas complicações, a fim de reconhecê-las precocemente e realizar a melhor intervenção. .

Phenotypic and Functional Analysis of HL-60 Cells Used in Opsonophagocytic-Killing Assay for Streptococcus pneumoniae

Differentiated HL-60 is an effector cell widely used for the opsonophagocytic-killing assay (OPKA) to measure efficacy of pneumococcal vaccines. We investigated the correlation between phenotypic expression of immunoreceptors and phagocytic ability of HL-60 cells differentiated with N,N-dimethylformamide (DMF), all-trans retinoic acid (ATRA), or 1alpha, 25-dihydroxyvitamin D3 (VitD3) for 5 days. Phenotypic change was examined by flow cytometry with specific antibodies to CD11c, CD14, CD18, CD32, and CD64. Apoptosis was determined by flow cytometry using 7-aminoactinomycin D. Function was evaluated by a standard OPKA against serotype 19F and chemiluminescence-based respiratory burst assay. The expression of CD11c and CD14 gradually increased upon exposure to all three agents, while CD14 expression increased abruptly after VitD3. The expression of CD18, CD32, and CD64 increased during differentiation with all three agents. Apoptosis remained less than 10% until day 3 but increased after differentiation by DMF or ATRA. Differentiation with ATRA or VitD3 increased the respiratory burst after day 4. DMF differentiation showed a high OPKA titer at day 1 which sustained thereafter while ATRA or VitD3-differentiated cells gradually increased. Pearson analysis between the phenotypic changes and OPKA titers suggests that CD11c might be a useful differentiation marker for HL-60 cells for use in pneumococcal OPKA.

Phenotypic and Functional Analysis of HL-60 Cells Used in Opsonophagocytic-Killing Assay for Streptococcus pneumoniae

Differentiated HL-60 is an effector cell widely used for the opsonophagocytic-killing assay (OPKA) to measure efficacy of pneumococcal vaccines. We investigated the correlation between phenotypic expression of immunoreceptors and phagocytic ability of HL-60 cells differentiated with N,N-dimethylformamide (DMF), all-trans retinoic acid (ATRA), or 1alpha, 25-dihydroxyvitamin D3 (VitD3) for 5 days. Phenotypic change was examined by flow cytometry with specific antibodies to CD11c, CD14, CD18, CD32, and CD64. Apoptosis was determined by flow cytometry using 7-aminoactinomycin D. Function was evaluated by a standard OPKA against serotype 19F and chemiluminescence-based respiratory burst assay. The expression of CD11c and CD14 gradually increased upon exposure to all three agents, while CD14 expression increased abruptly after VitD3. The expression of CD18, CD32, and CD64 increased during differentiation with all three agents. Apoptosis remained less than 10% until day 3 but increased after differentiation by DMF or ATRA. Differentiation with ATRA or VitD3 increased the respiratory burst after day 4. DMF differentiation showed a high OPKA titer at day 1 which sustained thereafter while ATRA or VitD3-differentiated cells gradually increased. Pearson analysis between the phenotypic changes and OPKA titers suggests that CD11c might be a useful differentiation marker for HL-60 cells for use in pneumococcal OPKA.

Herbicide paraquat induces sex-specific variation of neuroinflammation and neurodegeneration in Drosophila melanogaster.

There are several reports on herbicide paraquat (PQ)-induced Parkinsonian-like pathology in different animal models, including Drosophila melanogaster. Also, the role of some inflammatory factors, such as nitric oxide is reported in PQ-induced neuroinflammation of Drosophila. Although invertebrate model is valuable to study the conserved inflammatory pathway at the time of neurodegeneration, but neuroinflammation during PQ-mediated neurodegeneration has not been studied explicitly in Drosophila. In this study, the inflammatory response was examined in Drosophila model during PQ-induced neurodegeneration. We found that after exposure to PQ, survivability and locomotion ability were affected in both sexes of Drosophila. Behavioural symptoms indicated similar physiological features of Parkinson’s disease (PD) in different animal models, as well as in humans. Our study revealed alteration in proinflamatory factor, TNF-α and Eiger (the Drosophila homologue in TNF superfamily) was changed in PQ-treated Drosophila both at protein and mRNA level during neurodegeneration. To ensure the occurrence of neurodegeneration, tyrosine hydroxylase (TH) positive neuronal cell loss was considered as a hallmark of PD in the fly brain. Thus, our result revealed the conserved inflammatory events in terms of expression of TNF-α and Eiger present during a sublethal dose of PQ-administered neurodegeneration in male and female Drosophila with significant variation in proinflamatory factor level among both the sexes.

Efeito inibitório de quinolinas e fisalina f em células de indivíduos infectados pelo HTLV-1 / Inhibitory effect of quinolones and physalin f in cells from individuals infected with HTLV-1

A proliferação espontânea de linfócitos é um marcador da infecção pelo Vírus Linfotrópico de Célula T Humana do tipo 1 (HTL V -l)o Esta é mais elevada em pacientes com paraparesia espástica tropical/mielopatia associada ao HTL V (HAMJTSP) que em indivíduos assintomáticos. Embora o seu papel na patogênese da HAM/TSP ainda seja desconhecido, a identificação de drogas capazes de modular a proliferação espontânea pode ser relevante para o tratamento da HAM/TSP. Neste estudo nós avaliamos os efeitos dos derivados quinoIínicos BS373, Ql e Q2 e da fisalina F em culturas de células mononucleares do sangue periférico (PBMC) de indivíduos infectados pelo HTL V com HAM/TSP. Estes compostos inibiram, ex vivo, a proliferação espontânea em culturas de PBMC, conforme avaliado pela incorporação de 3H-timidina. Além disso, a produção espontânea, ex vivo, de citocinas inflamatórias foi significativamente inibida pelo composto BS373 (25 j.tM) e pela tlsahna F (10 j.tM). A expressão da proteína viral Tax foi reduzida cerca de 80% após incubação de PBMC com BS373 (25 uM). BS373 e fisalina F induziram um aumento na porcentagem de células em apoptose, como demonstrado por análise da marcação do PBMC com anexina V por citometri~ de fluxo. A análise ultraestrutural de células cultivadas na presença destes compostos mostrou vacúolos apresentando membranas de mielina, que se assemelham a compartimentos autofágicos. Em conclusão, as quinolinas e a fisalina F foram capaz de inibir a proliferação espontânea de células de indivíduos infectados pelo HTL V-I. Outros estudos são necessários para compreendermos os mecanismos pelos quais estes compostos agem no PBMC de indivíduos infectados pelo HTL V-I.

Spontaneous lymphocyte proliferation, a hallmark ofHuman- T Lymphocyte Virus Type 1 (HTL V -1) infection, is particular1y high in HTL V -associated myelopathy/tropical spastic paraparesis (HAM/TSP) patients compared to asymptomatic carriers. Although its role in the pathogenesis of HAM/TSP is stiH unknown, the identification of drugs capable of modulating the spontaneous proliferation may be reievant for the treatment ofHAM/TSP. Here we evaluated the effects ofthe quinoline derivative BS373, Q1 and Q2 and physalin F in cultures of peripheral blood mononuclear ceHs (PBMC) obtained from HTL V-infected subjects with HAM/TSP. Compounds inhibited of spontaneous proliferation in PBMC cultures, as assessed by 3H-thymidine incorporation. Additionally, the spontaneous production of inflammatory cytokines by PBMC was significantly inhibited by BS373 (25 flM) and physalin F (10 flM). The expression of the viral transcription factor Tax was reduced about 80% after incubation of PBMC with 8S373 (25 I-tM). BS373 and physalin F induced an increase in the percentage of apoptotic cells, as shown by flow cytometry analysis of annexin V-stained PBMC. Ultrastructural analysis of cultured cells in the presence of compounds showed vacuoles presented with myelin-like membranes, resembling autophagic vacuole-like compartments. In conc1usion, quinolines and physalin F was able to inhibit the spontaneous proliferation of cells from HTL V -l-infected individuaIs. Further studies are required to understand the mechanisms by which the compounds affect HTL V -1 PBMC.

Apoptosis of non-parasitised red blood cells in Plasmodium yoelii malaria

Recently, while studying erythrocytic apoptosis during Plasmodium yoelii infection, we observed an increase in the levels of non-parasitised red blood cell (nRBC) apoptosis, which could be related to malarial anaemia. Therefore, in the present study, we attempted to investigate whether nRBC apoptosis is associated with the peripheral RBC count, parasite load or immune response. To this end, BALB/c mice were infected with P. yoelii 17XL and nRBC apoptosis, number of peripheral RBCs, parasitaemia and plasmatic levels of cytokines, nitric oxide and anti-RBC antibodies were evaluated at the early and late stages of anaemia. The apoptosis of nRBCs increased at the late stage and was associated with parasitaemia, but not with the intensity of the immune response. The increased percentage of nRBC apoptosis that was observed when anaemia was accentuated was not related to a reduction in peripheral RBCs. We conclude that nRBC apoptosis in P. yoelii malaria appears to be induced in response to a high parasite load. Further studies on malaria models in which acute anaemia develops during low parasitaemia are needed to identify the potential pathogenic role of nRBC apoptosis.

Resveratrol diminishes platelet aggregation and increases susceptibility of K562 tumor cells to natural killer cells.

Platelet aggregation around migrating cancer cells protects them against the activity of natural killer cells (NKCs). The inability of immune system to response results in the progression of malignant diseases. This study was designed to evaluate the effects of resveratrol (3, 4', 5-trihydroxystilbene) on platelet aggregation and NKCs activity. Experiments were designed to evaluate the platelet aggregation, production of thromboxane B2 (TXB2), estimation of expression of the platelet receptor GpIIb/IIIa (major biological markers for platelet aggregation) and functional activity of the NKCs against the K562 cancer cell line after incubation with various concentrations of reveratrol. Resveratrol at a concentration of 3 × 10-3Μ completely inhibited platelet aggregation (p<0.05), decreased TXB2 levels (p<0.05) and inhibited the expression of receptor GpIIb/IIIa in non-stimulated platelets (p<0.05). At the same concentration, it increased the NKCs cytotoxic activity at an average rate of 319 ± 34, 450 ± 34 and 62 ± 2.4% (p<0.05) in the NKC/targets cells ratios of 12.5:1, 25:1 and 50:1, respectively. Thus, resveratrol not only completely inhibited platelet aggregation and reduced TXB2 levels and expression of receptor GpIIb/IIIa, but also increased the cytotoxic activity of NKCs in vitro and thus increased the susceptibility of tumor cells to NKCs. Thus, resveratrol can be used as an additional supplement to modulate the immune system and to inhibit platelet aggregation in thromboembolic episodes. Further clinical investigation in vivo could lead to specific concentrations that may maximize the beneficial effect of resveratrol.

Antiviral immunity in the red swamp crayfish, Procambarus Clarkii: hemocyte production, proliferation and apoptosis

In the present study, the pathogenic mechanism of white spot syndrome [WSSV] in crayfish, Procambarus clarkii by investigating activities of immune cells related to innate immune function during infection was explored. White spot disease caused by WSSV leads to devastating losses in crayfish farming. Examination by transmission electron microscopy revealed abundant WSSV particles and significant changes in the different lymphoid organs of infected crayfish. WSSV infection caused parts of the gill epithelium and microvilli to be reduced in number and size or damaged, meanwhile, the mitochondria, morphology changed, with parts of the cristae diminished leaving large vacuoles. Moreover, electron dense deposits appeared and hetero-chromatinized nuclei could be seen in blood cells with ruptured nuclear membranes and outflow of nucleoplasm. Also, evident were very densely basophilic inclusions were found in interstitial hepato-pancreatic tissue, connective tissue underlying the mid gut, cardiac tissue, gill tissue and hematopoietic tissue. Transmission electron microscopy revealed the presence of previously undescribed rod-shaped, enveloped versions in the cytoplasm adjacent to the nuclei of cells from various tissues

Estado funcional dos linfócitos T CD4+ e CD8+ e seu papel na progressão lenta da infecção por HIV em pacientes pediátricos / Functional state of CD4+ and CD8+ T lymphocytes and their role in the slow progression of HIV infection in pediatric patients

OBJETIVO: Avaliar o estado funcional dos linfócitos T CD4+ e CD8+ de pacientes pediátricos venezuelanos infectados pelo HIV-1. MÉTODOS: As crianças foram agrupadas como progressoras rápidas (PRs) ou progressoras lentas (PLs), com base no quadro clínico. Para determinar a funcionalidade dos linfócitos T CD4+ e CD8+, foram utilizadas técnicas de citometria de fluxo e caracterizados parâmetros de funcionalidade dessas células por meio de testes ex vivo como expressão de CD95/Fas e de CD127 e frequência de apoptose. Além disso, determinamos, em células mononucleares de sangue periférico, a proliferação do HIV e a produção de interleucina-10 (IL-10), do fator de necrose tumoral alfa (TNF-α) e de interferon gama (IFN-γ), e também estimamos o IFN-γ intracelular em células T CD4+. RESULTADOS: Nossos resultados indicam que vários mecanismos moleculares e celulares dos linfócitos T CD4+ e CD8+ tiveram piora nos PRs em comparação com PLs e controles. Ambos os tipos de linfócitos T dos PRs apresentaram aumento na expressão de CD95/Fas (p < 0,01), redução na expressão de CD127 (p < 0,01) e elevação na frequência de apoptose (p < 0,01). Além disso, as células T desses pacientes apresentaram diminuição na capacidade de proliferação mitogênica (p < 0,05), redução na porcentagem de linfócitos T CD4+ produtores de IFN-γ (p < 0,05) e menor capacidade de produção de IL-10, TNF-α e IFN-γ (p < 0,01) em comparação com PLs e controles. CONCLUSÃO: Nossos resultados indicam que o declínio das respostas moleculares e celulares dos linfócitos T está relacionado a uma rápida progressão e à diminuição na resistência à infecção pelo HIV-1 em crianças.

OBJECTIVE: To evaluate simultaneously the functional state of CD4+ and CD8+ T lymphocytes from Venezuelan HIV-1-infected pediatric patients. METHODS: Children were assigned to subgroups of rapid progressors (RPs) and slow progressors (SPs), based on clinical features. To determine the degree of CD4+ and CD8+ T-lymphocyte functionality, flow cytometry techniques were used, and diverse parameters of the functionality of these cells were characterized by ex vivo tests, such as expression of CD95/Fas and CD127, and frequency of apoptosis. In addition, we determined, in cultured peripheral blood mononuclear cells, HIV-specific proliferation and the production of interleukin-10 (IL-10), tumor necrosis factor alpha (TNF-α) and interferon gamma (IFN-γ), besides measuring intracellular IFN-γ in CD4+ T cells. RESULTS: Our results indicate that several molecular and cellular mechanisms of CD4+ and CD8+ T lymphocytes are deteriorated in RPs in comparison with SPs and controls. Indeed, both types of T lymphocytes from RPs exhibited an increased expression of CD95/Fas (p < 0.01), a significantly reduced expression of CD127 (p < 0.01), and an augmented frequency of apoptosis (p < 0.01). Furthermore, T cells from these patients displayed a diminished capacity of mitogen proliferation (p < 0.05), a reduced percentage of IFN-γ producing CD4+ T lymphocytes (p < 0.05) and a smaller capacity of IL-10, TNF-α and IFN-γ production (p < 0.01) in comparison with SP and control patients. CONCLUSION: Our findings indicate that the decline of the normal T lymphocyte molecular and cellular responses is related to a rapid progression and a decreased resistance to HIV-1 infection in children.

Construction of a recombinant adenovirus co-expressing truncated human prostate-specific membrane antigen and mouse 4-1BBL genes and its effect on dendritic cells

Our aim was to construct a recombinant adenovirus co-expressing truncated human prostate-specific membrane antigen (tPSMA) and mouse 4-1BBL genes and to determine its effect on dendritic cells (DCs) generated from bone marrow suspensions harvested from C57BL/6 mice for which the effect of 4-1BBL on DCs is not clear, especially during DCs processing tumor-associated antigen. Replication deficient adenovirus AdMaxTM Expression System was used to construct recombinant adenovirus Ad-tPSMA-internal ribosome entry site-mouse 4-1BBL (Ad-tPSMA-IRES-m4-1BBL) and Ad-enhanced green fluorescent protein. Day 7 proliferating DC aggregates generated from C57BL/6 mice were collected as immature DCs and further mature DCs were obtained by lipopolysaccharide activated immature DCs. After DCs were exposed to the recombinant adenovirus with 250 multiplicity of infection, the expression of tPSMA and m4-1BBL proteins were detected by Western blot, and the apoptosis and phenotype of DCs were analyzed by flow cytometry. Cytokines (IL-6 and IL-12) in the supernatant were detected by enzyme-linked immunosorbent assay (ELISA). Proliferation of T cells was detected by allogeneic mixed lymphocyte reactions. The tPSMA and m4-1BBL proteins were expressed correctly. The apoptosis rate of DCs transfected with Ad-tPSMA-IRES-m4-1BBL was 14.6 percent, lower than that of control DCs. The expression of co-stimulatory molecules [CD80 (81.6 ± 5.4 percent) and CD86 (80.13 ± 2.81 percent)] up-regulated in Ad-tPSMA-IRES-m4-1BBL-pulsed DCs, and the level of IL-6 (3960.2 ± 50.54 pg/mL) and IL-12 (249.57 ± 12.51 pg/mL) production in Ad-tPSMA-IRES-m4-1BBL-transduced DCs were significantly higher (P < 0.05) than those in control DCs. Ad-tPSMA-IRES-m4-1BBL induced higher T-cell proliferation (OD450 = 0.614 ± 0.018), indicating that this recombinant adenovirus can effectively enhance the activity of DCs.

Early detection of hepatocellular carcinoma on top of liver cirrhosis: the fas receptor and ligand system

Hepatocyte aberrations, accumulation of chromosomal damage and possibly initiation of hepatic carcinogenesis is thought to be caused by the continued viral replication and the persistent attempt by a less than optimal immune response to eliminate hepatitis C virus [HCV] infected cells. The identification of the "death factors" including Fas and its Ligand [Fas-L] as a major regulator of both apoptosis and immune function has provided insight into an attractive mechanism of tumor escape from immune clearance. To assess the hepatic expression of Fas/Fas-L, the Fas receptor [Fas-R] expression on lymphocyte, and serum soluble Fas [sFas] in an attempt to analyze the role of Fas receptor/ligand system in the multistep process of fibrosis/carcinogenesis and the possible use of the serum marker as possible candidate biomarkers for an early detection of hepatocellular carcinoma [HCC]. The current study included 100 samples from cases at Theodor Bilharz Research Institute and Kasr Al Aini Hospital in Egypt. There were 90 cases of chronic hepatitis C [CHC] infection [and negative hepatitis B virus infection]. There were 30 cases without liver cirrhosis, 30 cases with liver cirrhosis and 30 cases with HCC. 10 liver biopsies were taken from healthy livers as normal controls. Histopathologic study and immunohistochemistry for detection of hepatic Fas and Fas-L expression were determined for all cases. Electron microscopy [EM] and immunoelectron microscopy [IEM] examination for detection of Fas-R expression on lymphocytes were also done. sFas, liver function tests, serologic markers for viral hepatitis, and serum alpha-fetoprotein level [alpha-FP] were done. The sFas in both HCC and CHC with cirrhosis patients were significantly higher than those of normal controls and CHC without cirrhosis [P<0.01], but there was no significant difference between the cirrhosis and HCC patients. Positive hepatic expression of both Fas and Fas-L were significantly increased in the diseased groups [p<0. 01] compared to the control specimens. A progressive Fas and FasL increase from CHC without cirrhosis to CHC with cirrhosis followed by a decline from the latter to HCC. Apoptotic Fas and Fas-L proteins expression was significantly increased with the necroinflammatory activity and the advancement of fibrosis. There was a non-significant negative correlation between sFas and hepatic Fas. In addition a significant over expression of Fas-R on separated lymphocytes was associated with a higher frequency of apoptotic cell death as detected by EM examination. Conclusion: The Fas receptor/ligand system was significantly involved in the process of liver cirrhosis converting into HCC. Down-regulation of Fas expression, up-regulation of Fas-L expression in hepatocytes and elevation of serum sFas level were important in tumor evasion from immune surveillance and in hepatic carcinogenesis

IL-10 is Predominantly Produced by CD19(low)CD5(+) Regulatory B Cell Subpopulation: Characterisation of CD19 (high) and CD19(low) Subpopulations of CD5(+) B cells

IL-10 production by CD19(+)CD5(+) B cells was investigated, by determining the expression levels of CD19, a classical B cell marker. Peripheral mononuclear cells were stained with fluorescence-conjugated anti-CD5, anti-CD19, anti-IL-10, and Annexin V. Interestingly, IL-10-producing B cells were found to be localised within the CD19(low)CD5(+) B cell subset. Apoptotic changes were also observed mainly in CD19(low) cells among B cells. Thus, CD5(+) B cells should be classified as CD19(high) and CD19(low) cells, and the immunological significance of CD19 for the IL-10 production by CD5(+) B cells requires further studies.

Apoptose e modelos in vivo para estudo das moléculas relacionadas a este fenômeno / Apoptosis and in vivo models to study the molecules related to this phenomenon

Apoptosis or programmed cell death is a physiological process, essential for eliminating cells in excess or that are no longer necessary to the organism, acting on tissue homeostasis, although the phenomenon is also involved in pathological conditions. Apoptosis promotes activation of biochemical pathways inside cells called caspase pathway, of the proteins responsible for the cleavage of several cell substrates, leading to cell death. Antiapoptotic members of the Bcl-2 family (B cell CLL/lymphoma 2), that belong to the intrinsic route of the activation of caspases, such as Bcl-xL (extra-large B-cell lymphoma) and Bcl-w (Bcl-2-like 2), act predominantly to prevent that pro-apoptotic members, such as Bax (Bcl-2-associated X protein) and Bak (Bcl-2 relative bak) lead to cell death. Antiapoptotic molecules are considered potentially oncogenic. Murine models are known to be valuable systems for the experimental analysis of oncogenes in vivo, and for the identification of pharmacological targets for cancer and to assess antitumor therapies. Given the importance of tumorigenesis studies on the immune responses to cancer and the possibility of investigating the participation of antiapoptotic molecules in tumor progression in vivo, the development of new models may be platforms for studies on tumorigenesis, immune antitumor responses, investigation of the ectopic expression of antiapoptotic molecules and immunotherapies for tumors.

A apoptose, ou morte celular programada, é um processo fisiológico essencial para a eliminação de células em excesso ou que não são mais necessárias ao organismo, atuando na homeostase dos tecidos; entretanto, esse fenômeno também está envolvido em condições patológicas. A apoptose promove a ativação de vias bioquímicas dentro das células, denominada via das caspases, proteínas responsáveis pela clivagem de diversos substratos celulares, levando as células à morte. Membros antiapoptóticos da família Bcl-2 (B cell CLL/lymphoma 2), pertencentes à via intrínseca de ativação das caspases, como Bcl-xL (B-cell lymphoma-extra large) e Bcl-w (Bcl-2-like 2) atuam predominantemente prevenindo que os membros pró-apoptóticos, como Bax (Bcl-2-associated X protein) e Bak (Bcl-2 relative bak) ocasionem a morte celular. Moléculas antiapoptóticas são consideradas potencialmente oncogênicas. Sabe-se que os modelos murinos são sistemas valiosos para a análise experimental de oncogenes in vivo, bem como para a identificação de alvos farmacológicos do câncer e para avaliar terapias antitumorais. Em vista da importância dos estudos de tumorigênese e respostas imunes contra o câncer e da possibilidade de investigar a participação de moléculas antiapoptóticas na progressão tumoral in vivo, o desenvolvimento de novos modelos poderá servir como plataforma para estudos de tumorigênese, respostas imunes antitumorais, investigação de expressão ectópica de moléculas antiapoptóticas e imunoterapias contra tumores.

Os leucócitos e a resposta inflamatória na lesão de isquemia-reperfusão / Leukocytes and the inflammatory response in ischemia-reperfusion injury

Os eventos de isquemia-reperfusão desencadeiam uma resposta inflamatória sistêmica que pode levar a lesões celulares e até falência de órgãos. Tais repercussões são notadas no pós-operatório de cirurgias, em especial, com o uso de circulação extracorpórea. Sabe-se, atualmente, que os leucócitos exercem importante papel neste processo. Assim, este estudo aborda o papel dos leucócitos na fisiopatologia das lesões de isquemia-reperfusão e a ativação das cascatas inflamatórias por esse processo e procura auxiliar na compreensão destes mecanismos assim como trazer contribuições acerca das abordagens terapêuticas que possam atenuá-los. Esta revisão bibliográfica retrospectiva foi realizada a partir de documentos científicos publicados nos últimos dez anos, em português e inglês, indexados em bases de dados internacionais Medline e SciELO e de textos clássicos relacionados. Os descritores pesquisados foram: isquemia-reperfusão, leucócitos, resposta inflamatória, circulação extracorpórea, efeitos adversos e apoptose.

The events of ischemia-reperfusion injury triggers a systemic inflammatory response and can lead to cellular injury and organ failure. Such effects are noted in the postoperative recovery, especially with the use of cardiopulmonary bypass. Nowadays, it is known that leukocytes play an important role in this process. Therefore, this study addresses the role of leukocytes in the physiopathology of ischemia-reperfusion injuries and activation of inflammatory cascades through this process and seek to help in the understanding of these mechanisms as well as to bring contributions on the therapeutic approaches that can mitigate them. This retrospective review was performed from indexed scientific papers published over the last ten years in Portuguese and English in international databases MEDLINE and SciELO and related classic texts. The descriptors investigated were: ischemia-reperfusion, leukocytes, inflammatory response, cardiopulmonary bypass, adverse effects and apoptosis.