Hyaluronan driven by epithelial aPKC deficiency remodels the microenvironment and creates a vulnerability in mesenchymal colorectal cancer.

Mesenchymal colorectal cancer (mCRC) is microsatellite stable (MSS), highly desmoplastic, with CD8+ T cells excluded to the stromal periphery, resistant to immunotherapy, and driven by low levels of the atypical protein kinase Cs (aPKCs) in the intestinal epithelium. We show here that a salient feature of these tumors is the accumulation of hyaluronan (HA) which, along with reduced aPKC levels, predicts poor survival. HA promotes epithelial heterogeneity and the emergence of a tumor fetal metaplastic cell (TFMC) population endowed with invasive cancer features through a network of interactions with activated fibroblasts. TFMCs are sensitive to HA deposition, and their metaplastic markers have prognostic value. We demonstrate that in vivo HA degradation with a clinical dose of hyaluronidase impairs mCRC tumorigenesis and liver metastasis and enables immune checkpoint blockade therapy by promoting the recruitment of B and CD8+ T cells, including a proportion with resident memory features, and by blocking immunosuppression.

Anti-Inflammatory Effect of Tacrolimus/Hydroxypropyl-ß-Cyclodextrin Eye Drops in an Endotoxin-Induced Uveitis Model.

BACKGROUND: Uveitis is an infrequent disease which constitutes a major cause of ocular morbidity. Correct management is essential, being corticosteroids its cornerstone. In case of contraindication to corticosteroids or treatment failure, the use of topical tacrolimus (TAC) could be an alternative which has already demonstrated safety and effectiveness in other ocular pathologies. However, TAC eye drops are not marketed, thus their elaboration must be carried out in Hospital Pharmacy Departments (HPDs). METHODS: 32 Sprague-Dawley rats were divided into 4 groups of 8 rats each: (a) untreated healthy rats (Healthy); (b) untreated Endotoxin-Induced Uveitis model-rats (EIU); (c) EIU-rats treated with standard treatment of dexamethasone ophthalmic drops (DXM) and (d) EIU-rats treated with TAC-hydroxypropyl-ß-cyclodextrin eye drops previously developed by our group (TAC-HPßCD). The mRNA expression levels of IL-6, IL-8, MIP-1α and TNF-α, quantitative analysis of leucocytes in aqueous humor and histological evaluation were performed. RESULTS: TAC-HPßCD eye drops demonstrated to reduce ocular inflammation, expression of IL-6, TNF-α, MIP-1α and leukocyte infiltration in aqueous humor. CONCLUSIONS: TAC-HPßCD eye drops showed beneficial effect in EIU model in rats, positioning as an alternative for uveitis treatment in case of corticosteroids resistance or intolerance.

Tailored Hydrogels as Delivery Platforms for Conditioned Medium from Mesenchymal Stem Cells in a Model of Acute Colitis in Mice.

Inflammatory bowel disease (IBD), including Crohn's disease (CD) and ulcerative colitis (UC), is increasingly prevalent and current therapies are not completely effective. Mesenchymal stem cells are emerging as a promising therapeutic option. Here, the effect of local hydrogel application loaded with conditioned medium (CM) from human uterine cervical stem cells (hUCESC-CM) in an experimental acute colitis mice model has been evaluated. Colitis induction was carried out in C57BL/6 mice by dissolving dextran sulfate sodium (DSS) in drinking water for nine days. Ulcers were treated by rectal administration of either mesalazine (as positive control) or a mucoadhesive and thermosensitive hydrogel loaded with hUCESC-CM (H-hUCESC-CM). Body weight changes, colon length, and histopathological analysis were evaluated. In addition, pro-inflammatory TNF-α, IL-6, and IFN-γ mRNA levels were measured by qPCR. Treatment with H-hUCESC-CM inhibited body weight loss and colon shortening and induced a significant decrease in colon mucosa degeneration, as well as TNF-α, IFN-γ, and IL-6 mRNA levels. Results indicate that H-hUCESC-CM effectively alleviated DSS-induced colitis in mice, suggesting that H-hUCESC-CM may represent an attractive cell-free therapy for local treatment of IBD.

POU1F1 transcription factor induces metabolic reprogramming and breast cancer progression via LDHA regulation.

Metabolic reprogramming is considered hallmarks of cancer. Aerobic glycolysis in tumors cells has been well-known for almost a century, but specific factors that regulate lactate generation and the effects of lactate in both cancer cells and stroma are not yet well understood. In the present study using breast cancer cell lines, human primary cultures of breast tumors, and immune deficient murine models, we demonstrate that the POU1F1 transcription factor is functionally and clinically related to both metabolic reprogramming in breast cancer cells and fibroblasts activation. Mechanistically, we demonstrate that POU1F1 transcriptionally regulates the lactate dehydrogenase A (LDHA) gene. LDHA catalyzes pyruvate into lactate instead of leading into the tricarboxylic acid cycle. Lactate increases breast cancer cell proliferation, migration, and invasion. In addition, it activates normal-associated fibroblasts (NAFs) into cancer-associated fibroblasts (CAFs). Conversely, LDHA knockdown in breast cancer cells that overexpress POU1F1 decreases tumor volume and [18F]FDG uptake in tumor xenografts of mice. Clinically, POU1F1 and LDHA expression correlate with relapse- and metastasis-free survival. Our data indicate that POU1F1 induces a metabolic reprogramming through LDHA regulation in human breast tumor cells, modifying the phenotype of both cancer cells and fibroblasts to promote cancer progression.

Detection of regulatory T cell phenotypic markers and cytokines in patients with human papillomavirus infection.

Infection with human papillomavirus (HPV) is the main cause of cervical cancer. Viral persistence is considered the main risk factor for neoplastic progression and evidence suggests that regulatory T cells (Treg) play an important role in the failure of viral elimination. The aim of this study was to detect phenotypic markers of Treg and cytokines interleukin (IL)-10 and transforming growth factor (TGF)-ß, in the cervical microenvironment of HPV-infected patients. One hundred and one samples of uterine cervix embedded in paraffin were analyzed. We used immunohistochemistry to examine the coexpression of the CD25/FOXP3 and CD4/TGF-ß markers, and the expression of GITR and IL-10 in cells present in the cervical stroma. We detected a microenvironment composed of high proportions of CD25+ FOXP3+ , CD4+ TGFß+ , IL-10+ , and GITR+ cells in samples with high viral loads and severe lesions of HPV-infected patients. The abundance of these markers, indicative of the presence of Treg cells and immunosuppressive cytokines, was significantly associated with severe lesions and elevated viral loads in the examined samples. These results suggest that Treg cells may be involved in maintaining a microenvironment favorable for viral persistence and neoplastic progression. Our findings support those of previous studies that suggested that these markers could be used to predict HPV persistence and neoplastic progression, and as potential targets for immune response modulation.

Generalidades sobre la migración de bagres amazónicos de la familia Pimelodidae y su relación con los ciclos hidrológicos / General information about the migration of Amazonian catfish from the Pimelodidae family and its relationship with hydrological cycles / Informações gerais sobre a migração dos bagres da Amazônia da família Pimelodidae e sua relação com os ciclos hidrológicos

Resumen El entendimiento de los eventos migratorios en peces dulceacuícolas es fundamental para conocer su biología y ecología, pues estos acontecimientos naturales conllevan a generar alternativas para la gestión y preservación de las pesquerías, especialmente en especies que presentan desafíos a causa de su gran capacidad de desplazamiento en toda la cuenca amazónica. Los grandes bagres de la familia Pimelodidae son caracterizados por presentar estas características migratorias, las cuales están influenciadas por el pulso de inundación, donde el incremento de las aguas por grandes precipitaciones en las cabeceras elevan el nivel de las aguas, estimulándolos a iniciar migraciones y a llevar a cabo el proceso de maduración gonadal, induciendo a los individuos a dirigirse a las áreas de desove río arriba para culminar su proceso reproductivo. El objetivo de la presente revisión, pretende abordar de forma general aspectos involucrados en la migración de peces Siluriformes y su relación con los ciclos hidrológicos.

Abstract The understanding of the migratory events in freshwater fish is fundamental to know their biology and ecology, since these natural events lead to generate alternatives for the management and preservation of fisheries, especially in species that present challenges due to their great capacity of displacement in the entire Amazon basin. The large catfish of the Pimelodidae family are characterized by presenting these migratory characteristics, which are influenced by the flood pulse, where the increase of waters due to heavy rainfall in the headwaters raises the level of the waters, stimulating them to initiate migrations and to carry performed the process of gonadal maturation, inducing individuals to go to spawning areas upstream to complete their reproductive process. The objective of the present review, aims to address in a general way to deal with aspects involved in the migration of Siluriform fishes and their relationship with hydrological cycles.

Resumo O entendimento dos eventos migratórios em peixes de água doce é fundamental para conhecer os aspectos biológicos e ecológicos das espécies, este tipo de fenômenos naturais podem nos orientar a gerar alternativas para a gestão e preservação dos recursos pesqueiros, especialmente daquelas espécies que apresentam a capacidade de se deslocarem por toda a bacia amazônica. Os grandes bagres da família Pimelodidae são caracterizados por apresentar este tipo migração, onde ao estarem diretamente influenciados pelo pulso de inundação, especificamente quando ocorre aumento do nível das águas pelas precipitações na região andina, são estimulados a iniciarem migrações reprodutivas, induzindo-os a se dirigirem rio acima nas cabeceiras para culminar seu processo reprodutivo. O objetivo desta revisão foi abordar de forma geral os aspectos envolvidos na migração de peixes da ordem Siluriformes e sua relação com os períodos hidrométricos

Human papillomavirus and coinfections with Chlamydia trachomatis, Gardnerella vaginalis, and Trichomonas vaginalis in self-collected samples from female sex workers in the Central-Western region of Brazil / Papilomavírus humano e coinfecção por Chlamydia trachomatis, Gardnerella vaginalis e Trichomonas vaginalis em amostras autocoletadas de mulheres profissionais do sexo da região Centro-Oeste do Brasil

ABSTRACT Introduction: Human papillomavirus (HPV) is intimately associated with cervical cancer, and the presence of coinfections, such as with Chlamydia trachomatis, Gardnerella vaginalis and Trichomonas vaginalis, may potentiate or facilitate HPV infection. Female sex workers are considered vulnerable to the acquisition of these infections due to exposure to risk factors. Objective: To determine HPV infection, viral types and coinfections in self-collected samples from female sex workers. Methods: Self-collected samples from female sex workers, of vaginal canal and uterine cervix, were subjected to HPV-deoxyribonucleic acid (DNA) detection, viral genotyping by type-specific polymerase chain reaction (PCR), restriction fragment length polymorphism (RFLP), and the detection of coinfection. Results: HPV-DNA was detected in 19.4% of the samples, and HPV 31, 6, and 53 were the most frequently detected types. There was a predominance of high-risk oncogenic HPV (HR-HPV) and a strong presence of simultaneous infections with multiple HPV types (84.6%). Coinfections with both HPV and C. trachomatis, and HPV and G. vaginalis were detected. The variables that were statistically associated with HPV infection and the presence of multiple infections were non-use of condoms and non-compliance with regular cervical cytology screening. Conclusion: The results highlight the importance of more comprehensive studies among vulnerable populations, aiming to establish measures to raise awareness about the risks of contracting sexually transmitted infections, as well as to support future studies for introducing HPV vaccines with wider coverage of viral types.

RESUMO Introdução: O papilomavírus humano (HPV) está intimamente associado ao câncer cervical, e a presença de coinfecções, como por Chlamydia trachomatis, Gardnerella vaginalis e Trichomonas vaginalis, pode potencializar ou facilitar a infecção por HPV. As mulheres profissionais do sexo são consideradas vulneráveis à aquisição dessas infecções devido à exposição aos fatores de risco. Objetivo: Determinar a infecção por HPV, os tipos virais e as coinfecções em amostras autocoletadas de mulheres profissionais do sexo. Métodos: Amostras autocoletadas de mulheres profissionais do sexo, do canal vaginal e da cérvice uterina, foram submetidas a detecção do HPV-ácido desoxirribonucleico (DNA), genotipagem viral por reação em cadeia da polimerase (PCR) tipo específica e restriction fragment length polymorphism (RFLP) e detecção de coinfecção. Resultados: O HPV-DNA foi detectado em 19,4% das amostras, sendo os tipos HPV 31, 6 e 53 os mais frequentes. Houve predominância de HPV de alto risco (HR-HPV) e elevada presença de infecções múltiplas (84,6%). A presença de coinfecções foi observada tanto para HPV e C. trachomatis quanto para HPV e G. vaginalis. Observou-se também que mulheres profissionais do sexo que não fazem uso de preservativos e aquelas que não realizam o exame citológico rotineiramente estão predispostas à aquisição da infecção causada pelo HPV. Conclusão: Os resultados obtidos ressaltam a importância de estudos mais abrangentes entre as populações vulneráveis, objetivando estabelecer medidas para a conscientização sobre os riscos de aquisição das infecções sexualmente transmitidas, bem como auxiliar estudos futuros para introdução de vacinas contra o HPV com maior cobertura de tipos virais.

Predominant overexpression of CD25/FOXP3, IFN-γ, and suppressive cytokines in high-grade lesion samples infected with human papillomavirus / PPredominância de superexpressão de CD25/FOXP3, IFN-γ e citocinas supressoras em amostras de lesão de alto grau infectadas pelo papilomavírus humano

ABSTRACT Introduction: Human papillomavirus (HPV) persistent infection is the leading cause of cervical cancer and its precursor lesions, and the inappropriate immune response is among the factors that contribute to viral persistence. This may be influenced by regulatory T (Treg) cells and the production of immunosuppressive cytokines, such as transforming growth factor beta (TGF-β) and interleukin-10 (IL-10). Objective: We established the profile of the predominant response, Th1 or immunosuppressive response, in the tissue microenvironment, by detecting interferon-gamma (IFN-γ), TGF-β, and IL-10, as well as the co-expression of IL-2 receptor alpha (CD25) and forkhead box P3 (FOXP3). Methods: Seventy-four samples from uterine cervix biopsies that underwent HPV deoxyribonucleic acid (DNA) detection and histopathology analysis were immunostained to detect CD25/FOXP3, IFN-γ and suppressive cytokines in lymphocytes. Results: The microenvironment of high-grade squamous intraepithelial lesion (HSIL) samples with high numbers of viral particles (≥ 10,000 copies/ml) contained high numbers of CD25/FOXP3+, TGF-β+, IL-10+, and IFN-γ+ cells. Conclusion: The co-expression of CD25/FOXP3 and the expression of TGF-β, and IL-10 in HSIL samples suggest the existence of Treg cells in these locations, although IFN-γ expression was observed in several cells in these samples. Our data suggest that this cytokine could be related to immunosuppressed microenvironment maintenance, favoring the persistent HPV infection and the progression to carcinoma.

RESUMO Introdução: A infecção persistente por papilomavírus humano (HPV) é a principal causa do câncer cervical e suas lesões precursoras, e a resposta imune inadequada está entre os fatores que contribuem para a persistência viral. Isso pode ser influenciado por células T regulatórias (Treg) e pela produção de citocinas imunossupressoras, como o fator de transformação de crescimento beta (TGF-β) e a interleucina 10 (IL-10). Objetivo: Estabelecemos o perfil de resposta predominante, resposta Th1 ou imunossupressora, no microambiente tecidual, pela detecção de interferon gama (IFN-γ), TGF-β, e IL-10, bem como a coexpressão do receptor da cadeia alfa da IL-2 (CD25) e do forkhead box P3 (FOXP3). Método: Setenta e quatro amostras de biópsias de cérvice uterina, submetidas à detecção do ácido desoxirribonucleico (DNA) de HPV e à análise histopatológica, foram utilizadas nas reações de imuno-histoquímica para detectar IFN-γ, TGF-β, IL-10 e CD25/FOXP3 em linfócitos. Resultados: O microambiente das amostras de lesões intraepiteliais escamosas de alto grau (HSIL) com elevado números de partículas virais (≥ 10.000 cópias/ml) continha elevado número de células CD25/FOXP3+, TGF-β+, IL-10+ e IFN-γ+. Conclusão: A coexpressão de CD25/FOXP3 e a expressão de TGF-β nas amostras HSIL sugerem a existência de células Treg nesses locais, embora a expressão de IFN-γ tenha sido observada em várias células. Nossos dados sugerem que essa citocina pode estar relacionada com a manutenção do microambiente imunossuprimido, favorecendo a infecção persistente por HPV e a progressão para carcinoma.

Productive and Economic Responses in Grazing Dairy Cows to Grain Supplementation on Family Farms in the South of Brazil.

Pasture-based dairy production has been a major source of income for most family farms in the south of Brazil. Increasing milk prices have spurred an increase in grain supplementation, which has been poorly implemented, resulting in low levels of efficiency. To evaluate the consequences of supplementation on milk production and composition, grazing behavior and economic return, the widely used grain management system (CC-commercial concentrate, containing 21% CP, offered at 1 kg per 3.7 L of milk) was compared with an energy supplement (GC-ground corn, with 9.5% CP, offered at 0.4% of live weight). Ten Holstein cows were paired into two groups, and subjected to the two treatments in a crossover design. The cows remained in the same grazing group, and the grain supplement was offered individually at milking time and consumed completely. Each experimental period lasted 14 days, with 10 days for diet adaptation and four days for data collection; individual milk production and samples were collected to determine levels of fat, protein, lactose, carotenoids, vitamin A and N-urea. Grazing behavior was observed (scans every 5 min) in the first 4 h after the morning milking, and chemical composition of hand plucked samples of forage were measured. The cost of the supplement and profitability per treatment were calculated. Cows supplemented with GC consumed herbage with higher crude protein (CP: 16.23 vs. 14.62%; p < 0.05), had higher biting rate (44.21 vs. 39.54 bites/min; p < 0.03) and grazing time (22.20 vs. 20.55 scans; p < 0.05) than when receiving CC. There were no differences in milk composition between treatments (p > 0.05). However, higher concentrations of ß-carotene and total carotenoids were detected in the milk of cows at 70-164 days of lactation, compared to <70 days of lactation (p < 0.05). Milk production was higher (13.19 vs. 11.59 kg/day; p < 0.05) when cows consumed CC, but resulted in lower profitability compared to GC (US$ 4.39 vs. US$ 4.83/cow per day). Our results show that higher productivity does not necessarily improve profitability. Cows receiving supplement with lower levels of protein were able to adjust their grazing behavior to meet their protein needs and this level of diet modification did not alter milk composition.

Análise das soluções de hipoclorito de sódioutilizadas por endodontistas / Evaluation of sodium hypochlorite solutionsused by endodontists

Introdução: O sucesso da terapia endodôntica está condicionado adiversos fatores, entre os quais o uso de substâncias químicas deboa qualidade. Objetivo: Neste trabalho investigaram-se algumascaracterísticas das soluções de hipoclorito de sódio utilizadas emclínicas de atendimento, tais como concentração, pH e armazenagem.Material e métodos: Para isso, distribuiu-se formulário com perguntasrelativas à solução empregada a 100 endodontistas, além de pedir-lhes100 ml do produto. As amostras foram levadas ao laboratório paraanálise por titulometria da concentração de cloro residual livre, bemcomo do pH das soluções, com o auxílio de um peagômetro. Resultados:Apenas 1% dos profissionais conhece o valor do pH das soluções. Dasamostras recolhidas, 36% apresentaram pH 9, 22% pH 10, 16% compH 13, 14% pH 12, 11% pH 11 e 1% pH 8. Quanto aos profissionais,37% mantinham seus frascos sob refrigeração e 67% em temperatura ambiente. Conclusão: Os endodontistas usam soluções de hipocloritode sódio diferentes daquelas que desejam por desconhecerem ascondições necessárias para a manutenção da estabilidade química.

Introduction: The success of endodontic therapy is conditioned byseveral variables, being the use of good quality chemical agents one ofthem. Objective: In this study, some characteristics of commonly usedsodium hypochlorite irrigants were investigated, such as: concentration,pH, and storage conditions. Material and methods: For this purpose, aquestionnaire comprising questions regarding the employed irrigant wassent to 100 endodontists. Besides that, a 100mL sample of the sodiumhypochlorite irrigant used by them was collected. Free residual chlorinedetermination was laboratorially carried out through titration method.Additionally, pH was measured, through peagameter reading. Results:Only 1% of the professionals did know the pH value of the solutionsused by them. Concerning to pH value, 36% of the solutions presentedpH 9; 22%, pH 10; 16%, pH 13; 14%, pH 12; 11%, pH 11; and 1%, pH8. 37% of the endodontists kept the solution flasks under refrigeration,and 67% kept them at environmental temperature. Conclusion: It can beconcluded that the endodontists are using altered hypochlorite solutionsbecause they fail to observe the necessary conditions for maintainingthe solution?s chemical stability.