Innate immunity and HPV: friends or foes

Most human papillomavirus infections are readily cleared by the host immune response. However, in some individuals, human papillomavirus can establish a persistent infection. The persistence of high-risk human papillomavirus infection is the major risk factor for cervical cancer development. These viruses have developed mechanisms to evade the host immune system, which is an important step in persistence and, ultimately, in tumor development. Several cell types, receptors, transcription factors and inflammatory mediators involved in the antiviral immune response are viral targets and contribute to tumorigenesis. These targets include antigen-presenting cells, macrophages, natural killer cells, Toll-like receptors, nuclear factor kappa B and several cytokines and chemokines, such as interleukins, interferon and tumor necrosis factor. In the present review, we address both the main innate immune response mechanisms involved in HPV infection clearance and the viral strategies that promote viral persistence and may contribute to cancer development. Finally, we discuss the possibility of exploiting this knowledge to develop effective therapeutic strategies

Oxidative stress: therapeutic approaches for cervical cancer treatment

OBJECTIVES: Oxidative stress results from an imbalance between the generation and elimination of oxidant species. This condition may result in DNA, RNA and protein damage, leading to the accumulation of genetic alterations that can favor malignant transformation. Persistent infection with high-risk human papillomavirus types is associated with inflammatory responses and reactive oxygen species production. In this context, oxidative stress, chronic inflammation and high-risk human papillomavirus can act in a synergistic manner. To counteract the harmful effects of oxidant species, protective molecules, known as antioxidant defenses, are produced by cells to maintain redox homeostasis. In recent years, the use of natural antioxidants as therapeutic strategies for cancer treatment has attracted the attention of the scientific community. This review discusses specific molecules and mechanisms that can act against or together with oxidative stress, presenting alternatives for cervical cancer prevention and treatment

An Azole-Resistant Candida parapsilosis Outbreak: Clonal Persistence in the Intensive Care Unit of a Brazilian Teaching Hospital

The incidence of candidemia by the Candida parapsilosis complex has increased considerably in recent decades, frequently related to use of indwelling intravascular catheters. The ability of this pathogen to colonize healthcare workers (HCW)' hands, and to form biofilm on medical devices has been associated with the occurrence of nosocomial outbreaks and high mortality rates. Fluconazole has been the leading antifungal drug for the treatment of invasive candidiasis in developing countries. However, azole-resistant C. parapsilosis isolates are emerging worldwide, including in Brazil. Few studies have correlated outbreak infections due to C. parapsilosis with virulence factors, such as biofilm production. We thus conducted a microbiological investigation of C. parapsilosis complex isolates from a Brazilian teaching hospital. Additionally, we identified a previously unrecognized outbreak caused by a persistent azole-resistant C. parapsilosis (sensu stricto) clone in the intensive care unit (ICU), correlating it with the main clinical data from the patients with invasive candidiasis. The molecular identification of the isolates was carried out by PCR-RFLP assay; antifungal susceptibility and biofilm formation were also evaluated. The genotyping of all C. parapsilosis (sensu stricto) was performed by microsatellite analysis and the presence of ERG11 mutations was assessed in the azole non-susceptible isolates. Fourteen C. parapsilosis (sensu stricto) isolates were recovered from patients with invasive candidiasis, eight being fluconazole and voriconazole-resistant, and two intermediate only to fluconazole (FLC). All non-susceptible isolates showed a similar pattern of biofilm formation with low biomass and metabolic activity. The A395T mutation in ERG11 was detected exclusively among the azole-resistant isolates. According to the microsatellite analysis, all azole non-susceptible isolates from the adult ICU were clustered together indicating the occurrence of an outbreak. Regarding clinical data, all patients infected by the clonal non-susceptible isolates and none of the patients infected by the susceptible isolates had been previously exposed to corticosteroids (p = 0.001), while the remaining characteristics showed no statistical significance. The current study revealed the persistence of an azole non-susceptible C. parapsilosis clone with low capacity to form biofilm over two years in the adult ICU. These results reinforce the need of epidemiological surveillance and monitoring antifungal susceptibility of C. parapsilosis isolates in hospital wards

Pervasive gaps in Amazonian ecological research

Biodiversity loss is one of the main challenges of our time,1,2 and attempts to address it require a clear un derstanding of how ecological communities respond to environmental change across time and space.3,4 While the increasing availability of global databases on ecological communities has advanced our knowledge of biodiversity sensitivity to environmental changes,5–7 vast areas of the tropics remain understudied.8–11 In the American tropics, Amazonia stands out as the world’s most diverse rainforest and the primary source of Neotropical biodiversity,12 but it remains among the least known forests in America and is often underrepre sented in biodiversity databases.13–15 To worsen this situation, human-induced modifications16,17 may elim inate pieces of the Amazon’s biodiversity puzzle before we can use them to understand how ecological com munities are responding. To increase generalization and applicability of biodiversity knowledge,18,19 it is thus crucial to reduce biases in ecological research, particularly in regions projected to face the most pronounced environmental changes. We integrate ecological community metadata of 7,694 sampling sites for multiple or ganism groups in a machine learning model framework to map the research probability across the Brazilian Amazonia, while identifying the region’s vulnerability to environmental change. 15%–18% of the most ne glected areas in ecological research are expected to experience severe climate or land use changes by 2050. This means that unless we take immediate action, we will not be able to establish their current status, much less monitor how it is changing and what is being lostinfo:eu-repo/semantics/publishedVersio

Pervasive gaps in Amazonian ecological research

Biodiversity loss is one of the main challenges of our time,1,2 and attempts to address it require a clear understanding of how ecological communities respond to environmental change across time and space.3,4 While the increasing availability of global databases on ecological communities has advanced our knowledge of biodiversity sensitivity to environmental changes,5,6,7 vast areas of the tropics remain understudied.8,9,10,11 In the American tropics, Amazonia stands out as the world's most diverse rainforest and the primary source of Neotropical biodiversity,12 but it remains among the least known forests in America and is often underrepresented in biodiversity databases.13,14,15 To worsen this situation, human-induced modifications16,17 may eliminate pieces of the Amazon's biodiversity puzzle before we can use them to understand how ecological communities are responding. To increase generalization and applicability of biodiversity knowledge,18,19 it is thus crucial to reduce biases in ecological research, particularly in regions projected to face the most pronounced environmental changes. We integrate ecological community metadata of 7,694 sampling sites for multiple organism groups in a machine learning model framework to map the research probability across the Brazilian Amazonia, while identifying the region's vulnerability to environmental change. 15%–18% of the most neglected areas in ecological research are expected to experience severe climate or land use changes by 2050. This means that unless we take immediate action, we will not be able to establish their current status, much less monitor how it is changing and what is being lost

Functional evaluation of the interleukin-1 pathway in keratinocytes that express HPV oncogenes

Introdução: A resposta imune desempenha um papel importante na eliminação das infecções causadas pelo HPV. Como estratégia de evasão do sistema imunológico, o vírus pode modular a produção e secreção de citocinas pelas células infectadas. Além disso, a resposta imune pode contribuir para a criação de um quadro de inflamação crônica, que favorece a progressão das lesões relacionadas ao vírus e o desenvolvimento do câncer. Objetivos: O objetivo deste estudo foi avaliar o papel da via da interleucina-1, mais especificamente IL-1alfa, IL-1beta e IL-18, importantes citocinas pró-inflamatórias, no estabelecimento e progressão dos tumores cervicais. Material e métodos: Queratinócitos normais e transduzidos com os oncogenes E6 e/ou E7 de HPV16, além das linhagens tumorais de origem cervical C33, SiHa e HeLa, foram avaliados quanto à expressão (transcritos e proteínas) de IL-1alfa, IL-1beta, IL-18, seus receptores e antagonistas naturais. O gene codificador de IL-18 foi editado por meio do sistema CRISPR-Cas9, e as linhagens celulares silenciadas foram avaliadas quanto à capacidade de invasão, migração, proliferação, viabilidade e indução da angiogênese. Além disso, 62 biópsias provenientes de pacientes diagnosticadas com câncer cervical avançado foram analisadas por imuno-histoquímica visando à avaliação da expressão de IL-18 nos tecidos tumorais e sua relação com o prognóstico. Resultados: Foi verificado que, embora presente nos queratinócitos normais e transduzidos com os oncogenes do HPV, a expressão de IL-1alfa e IL-1beta encontra-se drasticamente reduzida nas linhagens tumorais de origem cervical, ao contrário de IL-18, que permanece sendo expressa de forma significativa. O silenciamento de IL-18 por CRISPR-Cas9 reduziu a capacidade proliferativa e de formação de colônias das linhagens tumorais cervicais HPV positivas, mas associou-se à tendência de maior capacidade invasiva e de indução da angiogênese. A análise imuno-histoquímica das biópsias mostrou que a expressão de IL-18 na borda dos tumores está associada à resposta completa ao tratamento e menor risco de recorrência linfática (p=0.027 e p=0.034, respectivamente). Embora não se tenha atingido significância estatística, observou-se clara tendência a maiores taxas de sobrevida global e sobrevida livre de doença nos tumores em cuja localização do infiltrado inflamatório era peritumoral (p=0.056 e p=0.165, respectivamente). Da mesma forma, a maior expressão de IL-18 no núcleo e no citoplasma das células tumorais foi associada à tendência de melhor prognóstico. Conclusões: Considerando-se os resultados acima expostos, verificou-se que, embora presentes nos queratinócitos normais e transduzidos com os oncogenes de HPV16, IL-1alfa, IL-1beta, seu antagonista natural e seus receptores são modulados negativamente de forma importante nas linhagens tumorais de origem cervical C33, SiHa e HeLa. Ao contrário, IL-18, seu antagonista natural e seus receptores continuam sendo expressos nas mesmas células. A análise conjunta dos resultados envolvendo as linhagens silenciadas para IL-18 e dos dados obtidos por imunohistoquímica sugere um papel protetor de IL-18, que deve ser confirmado por meio da ampliação da amostra estudada. Deve ser considerada a interação entre as células tumorais e o microambiente, mais especificamente, neste caso, o microambiente imune tumoral, no qual IL-18 constitui um importante mediador. Pretende-se que esta seja a próxima etapa deste estudoIntroduction: The immune response plays an important role in clearing HPV infections. As a strategy to evade the immune system, the virus can modulate cytokine production and secretion by infected cells. In addition, the immune response can contribute to create a chronic inflammatory condition, which can favor the progression of virus related lesions and cancer development. Objectives: The objective of this study was to evaluate the role of the interleukin-1 pathway, more specifically IL-1alpha, IL-1beta and IL-18, important pro-inflammatory cytokines, in the establishment and progression of cervical tumors. Material and methods: Normal keratinocytes and keratinocytes transduced with HPV16 E6 and/or E7 oncogenes, in addition to the cervical tumor cell lines C33, SiHa and HeLa, were evaluated for the expression (transcripts and proteins) of IL-1alpha, IL-1beta, IL-18, their natural antagonists and receptors. The gene encoding IL-18 was edited using CRISPRCas9 system, and silenced cell lines were evaluated for invasiveness, migration, proliferation, colony-forming capacity and angiogenesis induction. In addition, 62 biopsies from patients diagnosed with advanced cervical cancer were assessed by immunohistochemistry to assess IL-18 expression in tumor tissues and its relationship with the patients\' prognosis. Results: It was found that, although present in normal and transduced keratinocytes, the expression of IL-1alpha and IL-1beta is drastically reduced in cervical tumor cell lines, in contrast to IL-18, which remains expressed in a meaningful way. IL-18 silencing by CRISPR-Cas9 reduced the proliferative and colony-forming capacity of HPV-positive cervical tumor cell lines, but was associated with the tendency towards greater invasive capacity and angiogenesis induction. The biopsies immunohistochemical analysis showed that IL-18 expression at the edge of the tumor is associated with a complete response to the treatment and a lower risk of lymphatic recurrence (p=0.027 and p=0.034, respectively). Although no statistical significance was reached, there is a clear tendency towards higher rates of overall survival and disease free survival in tumors showing peritumoral location of the inflammatory infiltrate (p=0.056 and p=0.165, respectively). Likewise, the increased IL-18 expression in tumor cells nucleus and cytoplasm was associated with a trend towards better prognosis. Conclusions: Considering the above results, it was found that, although present in normal and transduced keratinocytes, IL-1alpha, IL-1beta, their natural antagonist and receptors are significantly negatively modulated in cervical tumor cell lines C33, SiHa and HeLa. In contrast, IL-18, its natural antagonist and receptors continue to be expressed in the same cells. Together, IL-18 silenced cells analysis and immunohistochemistry data suggest a protective role for IL-18, which should be confirmed by expanding the studied sample. The interaction between tumor cells and the microenvironment must be considered, more specifically, in this case, the tumor immune microenvironment, in which IL-18 is an important mediator. This is intended to be the next stage of this stud

Molecular techniques application for the complementary laboratory diagnosis of viral infections of the central nervous system, at the University Hospital of USP.

Enterovírus (HEV), herpesvírus 1 e 2 (HHV-1 e HHV-2) e adenovírus (HAdV) são importantes agentes de infecções do SNC. Neste trabalho, técnicas moleculares foram aplicadas para a detecção destes vírus em quadros de infecção do SNC. Amostras de líquor foram colhidas de pacientes atendidos no HU-USP entre agosto e novembro/2010 e fevereiro/2012 a janeiro/2013. Através da Nested-PCR HEV foram detectados em 9,8% das amostras, HAdV em 2,5% e HHV-1 e 2 em 1,1%, além de 3 casos de coinfecção, 2 entre HEV e HHV, e 1 entre HEV e HAdV. O material genético viral foi extraído através dos métodos Qiaamp DNA Blood (Qiagen®) e MagMAXTM Viral RNA Isolation (Ambiom), e este último pareceu mais adequado à aplicação na rotina clínica. A análise quimiocitológica do líquor mostrou-se importante no direcionamento da conduta clínica, mas a detecção do vírus é fundamental para a conclusão do diagnóstico. A PCR em tempo real, cuja padronização foi iniciada neste trabalho, consiste em importante ferramenta para a utilização futura no diagnóstico complementar das infecções virais do SNC.Enteroviruses (HEV), herpesviruses 1 and 2 (HHV-1 and HHV-2) and adenoviruses (HAdV) are important causative agents of infections of the CNS. In this study, molecular techniques were applied to the detection of these viruses. CSF samples were collected from patients treated at the University Hospital of USP, between August and November, 2010, and February 2012 and January 2013. By the Nested-PCR reaction, HEV were detected in 9.8% of the samples, HAdV in 2.5% and HHV-1 and 2 in 1.1%. There were 3 cases of coinfection: 2 with HEV and HHV and other with HEV and HAdV. The viral genetic materials were extracted by QIAamp DNA Blood kit (Qiagen®) and MagMAXTM Viral RNA Isolation (Ambiom), and the second one showed to be more suitable for the application in clinical diagnosis. The CSF chemocytologic analysis proved to be important in directing the clinical conduct, but the detection of viruses is essential for the diagnosis. The real time PCR, which standardization was initiated in this work, will be an important tool for complementary diagnosis of viral infections of the CNS

Innate immunity and HPV: friends or foes

Most human papillomavirus infections are readily cleared by the host immune response. However, in some individuals, human papillomavirus can establish a persistent infection. The persistence of high-risk human papillomavirus infection is the major risk factor for cervical cancer development. These viruses have developed mechanisms to evade the host immune system, which is an important step in persistence and, ultimately, in tumor development. Several cell types, receptors, transcription factors and inflammatory mediators involved in the antiviral immune response are viral targets and contribute to tumorigenesis. These targets include antigen-presenting cells, macrophages, natural killer cells, Toll-like receptors, nuclear factor kappa B and several cytokines and chemokines, such as interleukins, interferon and tumor necrosis factor. In the present review, we address both the main innate immune response mechanisms involved in HPV infection clearance and the viral strategies that promote viral persistence and may contribute to cancer development. Finally, we discuss the possibility of exploiting this knowledge to develop effective therapeutic strategies

Oxidative stress: therapeutic approaches for cervical cancer treatment

OBJECTIVES: Oxidative stress results from an imbalance between the generation and elimination of oxidant species. This condition may result in DNA, RNA and protein damage, leading to the accumulation of genetic alterations that can favor malignant transformation. Persistent infection with high-risk human papillomavirus types is associated with inflammatory responses and reactive oxygen species production. In this context, oxidative stress, chronic inflammation and high-risk human papillomavirus can act in a synergistic manner. To counteract the harmful effects of oxidant species, protective molecules, known as antioxidant defenses, are produced by cells to maintain redox homeostasis. In recent years, the use of natural antioxidants as therapeutic strategies for cancer treatment has attracted the attention of the scientific community. This review discusses specific molecules and mechanisms that can act against or together with oxidative stress, presenting alternatives for cervical cancer prevention and treatment