NK cells play a key role in the control of CMV infection in mice, but the mechanism by which NK cells can recognize and kill CMV-infected cells is unclear. In this study, the modulation of NK cell ...susceptibility of human CMV (hCMV)-infected cells was examined. We used a human lung and a human foreskin fibroblast cell line infected with clinical isolates (4636, 13B, or 109B) or with laboratory strains (AD169, Towne). The results indicate that all three hCMV clinical isolates confer a strong NK resistance, whereas only marginal or variable effects in the NK recognition were found when the laboratory strains were used. The same results were obtained regardless of the conditions of infection, effector cell activation status, cell culture conditions, and/or donor-target cell combinations. The NK cell inhibition did not correlate with HLA class I expression levels on the surface of the target cell and was independent of the leukocyte Ig-like receptor-1, as evaluated in Ab blocking experiments. No relevant changes were detected in the adhesion molecules ICAM-I and LFA-3 expressed on the cell surface of cells infected with hCMV clinical and laboratory strains. We conclude that hCMV possesses other mechanisms, related neither to target cell expression of HLA-I or adhesion molecules nor to NK cell expression of leukocyte Ig-like receptor-1, that confer resistance to NK cell recognition. Such mechanisms may be lost during in vitro passage of the virus. These results emphasize the differences between clinical hCMV isolates compared with laboratory strains.
Phosphatidylinositol 3-kinase (Pl-3K) plays a key role in several cellular processes, including mitogenesis, apoptosis, actin reorganization and vesicular trafficking. The molecular events involved ...in its activation have not been fully elucidated and several reports indicate that a key event for enzyme activation is the interaction of the SH2 domains of the p85 regulatory subunit of Pl-3K with tyrosine-phosphorylated proteins. In this study, we investigated the involvement of the product of the proto-oncogene c-Cbl in the activation of Pl-3K triggered by CD16 in human NK cells and the possible mechanisms leading to Pl-3K recruitment to the plasma membrane. Our results indicate that stimulation of NK cells through CD16 results in a rapid tyrosine phosphorylation of Cbl, which is constitutively associated with Grb2 and forms an activation-dependent complex with the p85 subunit of Pl-3K. In addition, we detected the presence of the Grb2-associated tyrosine-phosphorylated p36 and Shc proteins in anti-Cbl and anti-p85 immunoprecipitates from CD16-stimulated NK cell lysates. Upon CD16 stimulation, Pl-3K activity was found associated with Cbl and to a lesser extent with Grb2 and Shc as well as with the zeta chain of the CD16 receptor complex. Overall these results suggest that the formation of a complex containing either Shc or pp36 associated with Grb2, Cbl and the p85 subunit of Pl-3K is one of the major mechanisms which might couple CD16 to the Pl-3K pathway in NK cells.
Phosphatidylinositol 3‐kinase (Pl‐3K) plays a key role in several cellular processes, including mitogenesis, apoptosis, actin reorganization and vesicular trafficking. The molecular events involved ...in its activation have not been fully elucidated and several reports indicate that a key event for enzyme activation is the interaction of the SH2 domains of the p85 regulatory subunit of Pl‐3K with tyrosine‐phosphorylated proteins. In this study, we investigated the involvement of the product of the proto‐oncogene c‐Cbl in the activation of Pl‐3K triggered by CD16 in human NK cells and the possible mechanisms leading to Pl‐3K recruitment to the plasma membrane. Our results indicate that stimulation of NK cells through CD16 results in a rapid tyrosine phosphorylation of Cbl, which is constitutively associated with Grb2 and forms an activation‐dependent complex with the p85 subunit of Pl‐3K. In addition, we detected the presence of the Grb2‐associated tyrosine‐phosphorylated p36 and Shc proteins in anti‐Cbl and anti‐p85 immunoprecipitates from CD16‐stimulated NK cell lysates. Upon CD16 stimulation, Pl‐3K activity was found associated with Cbl and to a lesser extent with Grb2 and Shc as well as with the ζ chain of the CD16 receptor complex. Overall these results suggest that the formation of a complex containing either Shc or pp36 associated with Grb2, Cbl and the p85 subunit of Pl‐3K is one of the major mechanisms which might couple CD16 to the Pl‐3K pathway in NK cells.
Using the differential display PCR method, we have isolated an mRNA downregulated in doxorubicin resistant human cell lines. The full length cDNA clone was identified as the human homologue of yeast ...RPB11 subunit of RNA polymerase II. Northern blot analysis of normal tissues detected a particularly high expression of RPB11 mRNA in heart and skeletal muscle. Reduction of this mRNA expression was observed in all the cell lines tested after drug treatment and was paralleled by a similar decrease of the protein levels. These findings suggest that doxorubicin may exert in vivo specific inhibitory effects on a major component of the transcription machinery.
Increasing experimental evidence demonstrates that malignant transformation is associated with changes in the repertoire of expression of the integrin family of molecules, which mediate cell-matrix ...and cell-cell interactions. We have analyzed immunohistochemically and immunochemically the expression of VLA-3 integrin and its known ligands, namely, laminin (LM), fibronectin (FN), collagen type IV (Coll IV), nicein (NIC), and entactin/nidogen (ENT), in lung tumors of various histological types. alpha 3 beta 1 was detectable in normal bronchial epithelium and along basement membranes of alveolar walls. In non-small cell lung carcinomas (NSCLC) the integrin was expressed in 82% of the cases, independently of histological type and degree of differentiation of the tumors. On the other hand, only 13% of the small cell lung carcinomas (SCLC) displayed a weak and heterogeneous distribution of the alpha 3 beta 1 complex. Our findings were confirmed immunochemically using long-term tumor cell lines. While the expression of both alpha 3 beta 1 and ligands LM, FN, Coll IV, and Ent correlated in NSCLC with the presence of basement membranes, FN was the only ligand detectable in the stroma of SCLCs. A selective loss of nicein in basement membranes was demonstrated in NSCLC indicating an impairment of expression of this glycoprotein following malignant transformation.
The ability of tumor cells to interact with the extracellular matrix (ECM) is functionally mediated by a variety of receptor molecules among which the integrins are a well-characterized family of ...mediators. In this study we have investigated immunohistochemically the in vivo expression of the alpha 3/beta 1 promiscuous receptor for ECM constituents in a variety of human solid malignancies. Although the receptor appears to undergo changes in distribution patterns, its expression is maintained in a high percentage of primary (76%) and metastatic (82%) tumors. Furthermore, the comparative immunohistochemical evaluation of alpha 3/beta 1 and its ligands, in a selected number of tumors of different histotypes, demonstrated that the expression of this integrin correlates with the presence of at least one ligand, either around nests of neoplastic cells or at the epithelial-stromal interface. The highly conserved expression of alpha 3/beta 1 shown in this study suggests that this receptor may play a role in tumor growth at the primary as well as at the metastatic site.
Natural Killer (NK) cells are lymphoid cells that participate in innate immunity and in early defense against infections and tumors. NK cells express cell surface activating and inhibitory receptors ...that allow them to recognize and kill infected or tumor cells and rapidly produce cytokines and chemokines. Increasing evidence asserts an important immunomodulatory role of NK cells. We will discuss different mechanisms used by NK cells to regulate T cell-mediated immune responses.
Drowning is quick, silent and recognized as a serious public health problem worldwide, but it is neglected. Children and adolescents receive insufficient education about drowning prevention. The ...objective of this study was to monitor changes in the Drowning Prevention Knowledge Level (DPKL) in elementary school children and adolescents at CAp-UERJ, Rio de Janeiro, Brazil. The methodology was a longitudinal study carried out from 2022 to 2024 with students from the (5th, 6th, 7th, 8th and 9th years) of CAp-UERJ. 12 classes were monitored until 2024, totaling 336 students, four of which were in the 7th year, four in the 8th year and four in the 9th year, with a total of 112 students per year of schooling. A structured questionnaire divided into three parts containing 20 items about DPKL was answered at the school. In the 1st part, students correlated the universal figures used on signs to prevent drowning with the texts that signify those images (7 questions), in the 2nd part they had to relate the colors of the green, yellow and red flags and their meaning in regarding bathing conditions (3 questions) and in the last part, the student marked yes or no on statements related to the correct behavior to be adopted in the aquatic environment (10 questions). Each class received 3 interventions based on the individual DPKL result. When all years of schooling were analyzed together in three moments (2022, 2023 and 2024), the result showed that there was an improvement in knowledge about the 7 prevention signs, to the point that 99.1% of students now knew the meanings of the signs, and in two of them, sign no. 2, which refers to the emergency telephone and in no. 3, pushing prohibited, there was 100% accuracy. It was also possible to verify that after the intervention, almost all (98%) of the students correctly understood the meaning of the colors of the green, yellow and red flags. Regarding the behavior of playing nearby or putting their hand in the drain that sucks water from the pool, the result showed that the youngest were the ones who improved the most after intervention, going from 77.2% to 99.1% of those who responded correctly in 2024; about entering the pool, diving with a “somersault”, in a dangerous and inappropriate way, it was found that the older they were, the more aware of the danger caused by headfirst dives. Regarding the DPKL categorization, it was noted that 99% of students were classified as DPKL excellent in 2024. Those in the 8th and 9th years obtained 100% in the classification as DPKL excellent. It can be concluded that the school was a good place to carry out preventive interventions on drowning using an approach focused on behavior change, conceptual identification and student awareness in order to improve DPKL. The diagnostic use of DPKL to monitor school-age students can help identify safety values and concepts in certain regions of the country or specific groups that are not familiar with aquatic environments and thus help to formulate preventive interventions if necessary.
O afogamento é rápido, silencioso e reconhecido como um grave problema de saúde pública em todo o mundo, mas é negligenciado. As crianças e adolescentes recebem educação insuficiente sobre prevenção de afogamento. O objetivo deste estudo foi acompanhar as mudanças do Nível de Conhecimento Preventivo de Afogamento (NCPA) em crianças e adolescentes escolares do ensino fundamental do CAp-UERJ, Rio de Janeiro, Brasil. A metodologia foi de estudo longitudinal realizado de 2022 até 2024 com estudantes do (5º, 6º, 7º, 8º e 9º anos) do CAp-UERJ. Foram acompanhadas 12 turmas até 2024, totalizando 336 alunos, sendo quatro turmas que cursavam o 7° ano, quatro do 8º ano e quatro do 9º ano, com total de 112 alunos por ano de escolaridade. Um questionário estruturado dividido em três partes contendo 20 itens sobre NCPA foi respondido na escola. Na 1ª parte os alunos correlacionavam as figuras universais usadas nas placas para prevenção do afogamento aos textos que significam aquelas imagens (7 questões), na 2ª parte tinham que fazer a relação entre as cores das bandeiras verde, amarela e vermelha e o seu significado em relação às condições de banho (3 questões) e na última parte, o escolar marcava sim ou não em afirmações relacionadas ao comportamento correto a ser adotada no ambiente aquático (10 questões). Cada turma recebeu 3 intervenções com base no resultado individual do NCPA. Quando analisados todos os anos de escolaridade juntos em três momentos (2022, 2023 e 2024) o resultado mostrou que houve melhora no conhecimento sobre as 7 placas de prevenção, a ponto de, 99,1% dos alunos passaram a conhecer os significados das placas, e em duas delas, placa n.º 2, que se refere ao telefone de emergência e na de n.º 3, proibido empurrar, houve 100% de acerto. Foi possível verificar também que após a intervenção, quase a totalidade (98%) dos escolares acertaram o significado das cores das bandeiras verde, amarelo e vermelha. Em relação ao comportamento de brincar próximo ou colocar a mão no dreno que aspira a água da piscina o resultado mostrou que os mais novos foram os que mais melhoraram após intervenção, passando de 77,2% para 99,1% os que responderam corretamente em 2024; sobre à entrada na piscina mergulhando com “salto mortal”, de forma perigosa e inadequada, verificou-se que quanto mais velho, mais noção do perigo, causado por mergulhos de cabeça eles tinham. Em relação a categorização do NCPA, notou-se que 99% dos alunos foram classificados como NCPA excelente em 2024. Sendo que os que cursavam o 8º e 9º ano obtiveram 100% na classificação como NCPA excelente. Pode-se concluir que a escola foi um bom local para se realizar intervenções preventivas sobre afogamentos usando abordagem focada na mudança de comportamento, identificação conceitual e conscientização dos alunos a fim de melhorar o NCPA. A utilização diagnóstica do NCPA para acompanhamento dos alunos em idade escolar pode ajudar a identificar valores e conceitos de segurança em determinadas regiões do país ou grupos específicos que não estão familiarizados com ambientes aquáticos e assim ajudar a formular intervenções preventivas caso necessário.
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