In vivo studies could not confirm in vitro prophylactic synergism between Moringa essential oil and Lactobacillus reuteri (MT180537)

Abstract Aerobic vaginitis (AV) is a recently defined vaginal recurring infection, which is treated with antibiotics. However, excessive and prolonged use of antibiotics disrupts healthy vaginal microflora and leads to the emergence of antibiotic resistance among pathogens. This situation has directed researchers to explore alternative antimicrobials. The current study describes in vitro and in vivo antimicrobial efficacy and pharmaceutical interactions between plant essential oils (EOs) and five lactic acid bacteria (LABs), isolated from the healthy vagina, against E. faecalis, one of the major etiological agents of AV. In vitro experiments confirm good antimicrobial activity of both plant EOs and cell free supernatant (CFS) from LABs. Based on high antimicrobial efficacy, Moringa essential oil (MO) was selected to determine its nature of interaction with CFS of five LAB strains. Synergism was recorded between MO and CFS of L. reuteri (MT180537). To validate in vitro findings, prophylactic responses of individual and synergistic application of MO and L. reuteri (MT180537) were evaluated in an E. faecalis (MW051601) induced AV murine model. The prophylactic efficacy was evidenced by a reduction in intensity of clinical symptoms, E. faecalis (MW051601) count per vaginal tissue along with a reduction in AV associated changes in histological markers of infection in animals receiving Moringa essential oil and L. reuteri (MT180537) alone or in combination. However, significant synergism between Moringa essential oil and L. reuteri (MT180537) could not be observed. Our data confirms the importance of in vivo experiments in deducing pharmacological interactions.

Resumo Vaginite aeróbica (VA) é uma infecção vaginal recorrente definida recentemente, que é tratada com antibióticos. No entanto, o uso excessivo e prolongado de antibióticos perturba a microflora vaginal saudável e leva ao surgimento de resistência aos antibióticos entre os patógenos. Esta situação levou os pesquisadores a explorar antimicrobianos alternativos. O presente estudo descreve a eficácia antimicrobiana in vitro e in vivo e as interações farmacêuticas entre óleos essenciais vegetais (OE) e cinco bactérias lácticas (BAL), isoladas de vagina sã, contra E. faecalis, um dos principais agentes etiológicos da AV. Os experimentos in vitro confirmam a boa atividade antimicrobiana de ambos os EOs de plantas e sobrenadante livre de células (CFS) de LABs. Com base na alta eficácia antimicrobiana, o óleo essencial de Moringa (MO) foi selecionado para determinar sua natureza de interação com o sobrenadante livre de células (CFS) de cinco cepas de LAB. Sinergismo foi registrado entre MO e CFS de L. reuteri (MT180537). Para validar os resultados in vitro, as respostas profiláticas da aplicação individual e sinérgica de MO e L. reuteri (MT180537) foram avaliadas em um modelo murino AV induzido por E. faecalis (MW051601). A eficácia profilática foi evidenciada por uma redução na intensidade dos sintomas clínicos, contagem de E. faecalis (MW051601) por tecido vaginal, juntamente com uma redução nas alterações associadas a AV nos marcadores histológicos de infecção em animais que receberam óleo essencial de Moringa e L. reuteri (MT180537) sozinho ou em combinação. No entanto, não foi possível observar sinergismo significativo entre o óleo essencial de Moringa e L. reuteri (MT180537). Nossos dados confirmam a importância dos experimentos in vivo na dedução de interações farmacológicas.

In vivo studies could not confirm in vitro prophylactic synergism between Moringa essential oil and Lactobacillus reuteri (MT180537) / Estudos in vivo não puderam confirmar o sinergismo profilático in vitro entre o óleo essencial de Moringa e Lactobacillus reuteri (MT180537)

Aerobic vaginitis (AV) is a recently defined vaginal recurring infection, which is treated with antibiotics. However, excessive and prolonged use of antibiotics disrupts healthy vaginal microflora and leads to the emergence of antibiotic resistance among pathogens. This situation has directed researchers to explore alternative antimicrobials. The current study describes in vitro and in vivo antimicrobial efficacy and pharmaceutical interactions between plant essential oils (EOs) and five lactic acid bacteria (LABs), isolated from the healthy vagina, against E. faecalis, one of the major etiological agents of AV. In vitro experiments confirm good antimicrobial activity of both plant EOs and cell free supernatant (CFS) from LABs. Based on high antimicrobial efficacy, Moringa essential oil (MO) was selected to determine its nature of interaction with CFS of five LAB strains. Synergism was recorded between MO and CFS of L. reuteri (MT180537). To validate in vitro findings, prophylactic responses of individual and synergistic application of MO and L. reuteri (MT180537) were evaluated in an E. faecalis (MW051601) induced AV murine model. The prophylactic efficacy was evidenced by a reduction in intensity of clinical symptoms, E. faecalis (MW051601) count per vaginal tissue along with a reduction in AV associated changes in histological markers of infection in animals receiving Moringa essential oil and L. reuteri (MT180537) alone or in combination. However, significant synergism between Moringa essential oil and L. reuteri (MT180537) could not be observed. Our data confirms the importance of in vivo experiments in deducing pharmacological interactions.

Vaginite aeróbica (VA) é uma infecção vaginal recorrente definida recentemente, que é tratada com antibióticos. No entanto, o uso excessivo e prolongado de antibióticos perturba a microflora vaginal saudável e leva ao surgimento de resistência aos antibióticos entre os patógenos. Esta situação levou os pesquisadores a explorar antimicrobianos alternativos. O presente estudo descreve a eficácia antimicrobiana in vitro e in vivo e as interações farmacêuticas entre óleos essenciais vegetais (OE) e cinco bactérias lácticas (BAL), isoladas de vagina sã, contra E. faecalis, um dos principais agentes etiológicos da AV. Os experimentos in vitro confirmam a boa atividade antimicrobiana de ambos os EOs de plantas e sobrenadante livre de células (CFS) de LABs. Com base na alta eficácia antimicrobiana, o óleo essencial de Moringa (MO) foi selecionado para determinar sua natureza de interação com o sobrenadante livre de células (CFS) de cinco cepas de LAB. Sinergismo foi registrado entre MO e CFS de L. reuteri (MT180537). Para validar os resultados in vitro, as respostas profiláticas da aplicação individual e sinérgica de MO e L. reuteri (MT180537) foram avaliadas em um modelo murino AV induzido por E. faecalis (MW051601). A eficácia profilática foi evidenciada por uma redução na intensidade dos sintomas clínicos, contagem de E. faecalis (MW051601) por tecido vaginal, juntamente com uma redução nas alterações associadas a AV nos marcadores histológicos de infecção em animais que receberam óleo essencial de Moringa e L. reuteri (MT180537) sozinho ou em combinação. No entanto, não foi possível observar sinergismo significativo entre o óleo essencial de Moringa e L. reuteri (MT180537). Nossos dados confirmam a importância dos experimentos in vivo na dedução de interações farmacológicas.

Modelos murinos para el estudio de cáncer de mama triple negativo / Murine models for the study of triple negative breast cancer

RESUMEN El estudio de diferentes variables, como patogénesis, perfil inflamatorio, identificación de blancos terapéuticos, eficacia de tratamientos en modelos murinos ha resultado uno de los más prácticos para el estudio preclínico del cáncer de mama triple negativo (CMTN), el subtipo de cáncer más agresivo, con limitada aplicación de tratamientos y baja tasa de sobrevivencia. Sin embargo, hay que reconocer que existen otros menores en los que se viene estandarizando la inducción del CMTN. En esta revisión se engloban los diferentes métodos de inducción que han permitido el desarrollo de CMTN y las aplicaciones terapéuticas más relevantes por el que se desarrollaron los modelos murinos con CMTN.

ABSTRACT The study of different variables, such as pathogenesis, inflammatory profile, identification of therapeutic targets, efficacy of treatments in murine models has proven to be one of the most practical for the preclinical study of triple negative breast cancer (TNBC), the most aggressive subtype of cancer, with limited application of treatments and low survival rate. However, it must be recognized that there are other minors in which the induction of TNBC is being standardized. This review encompasses the different induction methods that have allowed the development of TNBC and the most relevant therapeutic applications by which murine models with TNBC were developed.

Salmonella entérica serovar Enteritidis biofilm lifestyle induces lower pathogenicity and reduces inflammatory response in a murine model compared to planktonic bacteria / Salmonella entérica serovar Enteritidis en biopelícula induce una menor patogenicidad y respuesta inflamatoria en un modelo murino que su contraparte planctónica

Abstract Salmonella enterica serovar Enteritidis (S. Enteritidis) is the most frequent serovar involved in human salmonellosis. It has been demonstrated that about 80% of infections are related to biofilm formation. There is scant information about the pathogenicity of S. Enteritidis and its relationship to biofilm production. In this regard, this study aimed to investigate the differential host response induced by S. Enteritidis biofilm and planktonic lifestyle. To this purpose, biofilm and planktonic bacteria were inoculated to BALB/c mice and epithelial cell culture. Survival studies revealed that biofilm is less virulent than planktonic cells. Reduced signs of intestinal inflammation and lower bacterial translocation were observed in animals inoculated with Salmonella biofilm compared to the planktonic group. Results showed that Salmonella biofilm was impaired for invasion of non-phagocytic cells and induces a lower inflammatory response in vivo and in vitro compared to that of planktonic bacteria. Taken together, the outcome of Salmonella-host interaction varies depending on the bacterial lifestyle.

Resumen Salmonella entérica serovar Enteritidis (S. Enteritidis) es la serovariedad más frecuentemente aislada en la salmonelosis humana. Se ha demostrado que alrededor del 80% de las infecciones están relacionadas con la formación de biopelículas. Sin embargo, la información disponible acerca de la patogenicidad de S. Enteritidis y su relación con la producción de biopelículas es escasa. Este trabajo tuvo como objetivo investigar la respuesta diferencial del huésped frente a S. Enteritidis en sus 2 estilos de vida: biopelícula y planctónico. Para ello, se inocularon bacterias en estado de biopelícula o planctónico en ratones BALB/c y cultivo de células epiteliales. Los estudios de supervivencia revelaron que Salmonella en biopelícula fue menos virulenta que su contraparte planctónica. Los animales inoculados con biopelículas presentaron una mayor conservación estructural del intestino y una menor translocación bacteriana que el grupo planctónico. Asimismo, Salmonella en biopelícula mostró una capacidad deficiente para invadir células no fagocíticas e indujo una menor respuesta inflamatoria in vivo e in vitro que las bacterias planctónicas. Se concluye que el resultado de la interacción Salmonella-huésped depende del estilo de vida bacteriano.

Prevalence of rickettsias in ticks (Acari: Ixodidae) collected in domestic animals of the northern region of Caldas department, Colombia / Prevalencia de rickettsias en garrapatas (Acari: Ixodidae) colectadas en animales domésticos del norte del departamento de Caldas, Colombia

ABSTRACT The northern region of the Caldas department in Colombia is considered an endemic area for murine typhus. Recent studies in patients with acute febrile disease demonstrated infection with the spotted fever group's rickettsiae due to an increase in the IgG titer by indirect immunofluorescence in paired sera obtained from these patients. The objective of the current research was to identify the species of ticks present in domestic animals in the northern region of Caldas and establish the presence of rickettsial genomic material in the collected ticks. Ticks were obtained from bovines, horses, and dogs in seven municipalities in the north of Caldas. Ticks were stored in 90 % ethanol until processing and were identified using taxonomic keys, DNA was extracted using commercial techniques, and the gltA gene was amplified by conventional chain reaction polymerase (PCR). Seven hundred thirteen ticks were obtained from 593 domestic animals. The highest infestation occurred in cattle, followed by canines and horses. Ticks found corresponded to the species Riphicephalus microplus, Dermacentor nitens, Amblyomma sp., and Riphicephalus sanguineus s.l. In none of the tick samples, Rickettsia-specific gltA gene DNA was found. It can be inferred that the ticks obtained are not a source of rickettsial infection for people in this department region, despite finding different species associated with the transmission of this disease.

RESUMEN La región norte del departamento de Caldas, Colombia es considerada como una zona endémica de tifo murino. Estudios recientes realizados en pacientes con enfermedad febril aguda, demostraron la infección con rickettsias, del grupo de las fiebres manchadas, debido al aumento en el título de IgG, por inmunofluorescencia indirecta (IFI), en sueros pareados, obtenidos de dichos pacientes. El objetivo de la investigación fue el de identificar las especies de garrapatas presentes en animales domésticos, de la región norte de Caldas y establecer la presencia de material genómico de rickettsias, en las garrapatas recolectadas. En siete municipios, se recolectaron garrapatas de bovinos, de equinos y de caninos. Las garrapatas, se almacenaron en etanol al 90 %, hasta su identificación taxonómica. Se extrajo el ADN, utilizando técnicas comerciales y se amplificó por reacción de cadena de la polimerasa (PCR) convencional el gen gltA. Se obtuvieron 713 garrapatas de 593 animales domésticos. La más alta infestación se presentó en bovinos, seguido de los caninos y equinos. Las garrapatas encontradas correspondieron a las especies Riphicephalus microplus, Dermacentor nitens, Amblyomma sp. y Riphicephalus sanguineus s.l. En ninguna de las muestras, se comprobó la presencia de ADN del gen gltA específico de Rickettsia. Se puede inferir que las garrapatas obtenidas no serían una fuente de infección rickettsial para las personas, en esta región del departamento; sin embargo, su presencia es un factor de riesgo para la adquisición de rickettsiosis asociadas con las fiebres manchadas.

Salmonella enterica serovar Enteritidis biofilm lifestyle induces lower pathogenicity and reduces inflammatory response in a murine model compared to planktonic bacteria.

Salmonellaenterica serovar Enteritidis (S. Enteritidis) is the most frequent serovar involved in human salmonellosis. It has been demonstrated that about 80% of infections are related to biofilm formation. There is scant information about the pathogenicity of S. Enteritidis and its relationship to biofilm production. In this regard, this study aimed to investigate the differential host response induced by S. Enteritidis biofilm and planktonic lifestyle. To this purpose, biofilm and planktonic bacteria were inoculated to BALB/c mice and epithelial cell culture. Survival studies revealed that biofilm is less virulent than planktonic cells. Reduced signs of intestinal inflammation and lower bacterial translocation were observed in animals inoculated with Salmonella biofilm compared to the planktonic group. Results showed that Salmonella biofilm was impaired for invasion of non-phagocytic cells and induces a lower inflammatory response in vivo and in vitro compared to that of planktonic bacteria. Taken together, the outcome of Salmonella-host interaction varies depending on the bacterial lifestyle.

Sensitization with Eustrongylides sp. (Nematoda: Dioctophymatidae) antigens induce production of specific IgG and IgE in murine model / Sensibilização com antígenos de Eustrongylides sp. (Nematoda: Dioctophymatidae) induzindo a produção de IgG e IgE em modelo murino

Abstract Fish consumption plays an important role in human diet. Hoplias malabaricus, commonly known as traíra, is a freshwater fish widely appreciated in several Brazilian states and frequently infected by Eustrongylides sp. fourth-instar larvae (L4). The aim of the present study was to evaluate allergenic potential of Eustrongylides sp. L4 crude extract (CEE). BALB/c mice were immunized intraperitoneally (IP) by 10 μg CEE with 2 mg of aluminum hydroxide on days 0 and 35. Specific IgG and IgE antibody levels were determined after immunization and cellular immunity was evaluated by assessing intradermal reaction in ear pavilion. Epicutaneous sensitization was performed in dorsal region by antigen exposure using a Finn-type chamber containing 50 μg of CEE or saline solution, followed by evaluation of specific antibody levels. IP immunization resulted in a gradual increase in IgG antibody levels and transitory IgE production. Significant increase in ear thickness was observed in cellular hypersensitivity reaction. In case of antigen exposure by epicutaneous route, CEE was able to induce meaningfully increased levels of specific IgG and IgE antibodies as well as heightened cellular immunity. Both intraperitoneal immunization and epicutaneous contact with Eustrongylides sp. larval antigens were observed for first time to be capable of inducing immunological sensitization in mice.

Resumo Consumir peixe constitui papel importante na dieta humana. Hoplias malabaricus, comumente chamado de traíra, peixe de água doce largamente apreciado no Brasil, é frequentemente infectado com larvas de quarto estágio (L4) de Eustrongylides sp. O presente estudo objetivou avaliar o potencial alergênico do Extrato Bruto de L4 de Eustrongylides sp. (EBE). Camundongos BALB/c foram imunizados intraperitonealmente (IP) por 10 μg de EBE com 2 mg de hidróxido de alumínio nos dias 0 e 35. Após imunização, determinaram-se níveis específicos de anticorpos IgG e IgE e avaliou-se a imunidade celular pela reação intradérmica no pavilhão auricular. Realizou-se sensibilização epicutânea na região dorsal pela exposição ao antígeno, utilizando-se câmara tipo Finn, contendo 50 μg de CEE ou solução salina. Após exposições, foram avaliados níveis específicos de anticorpos. Na imunização via IP, houve aumento gradual nos níveis de anticorpos IgG e produção de IgE transitória. Foi observado aumento significativo na espessura do pavilhão auricular na reação de hipersensibilidade celular. Na exposição ao antígeno pela via epicutânea, o EBE induziu aumento significante nos níveis de anticorpos IgG e IgE específicos e induziu imunidade celular. Pela primeira vez, observou-se que a imunização intraperitoneal e contato epicutâneo com antígenos larvares de Eustrongylides sp. são capazes de induzir sensibilização imunológica em camundongos.

Avaliação da técnica de vitrificação de ovários de camundongos da linhagem B6D2F1, pertencentes ao Instituto de Ciência e Tecnologia em Biomodelos - ICTB/Fiocruz-RJ / Evaluation of ovary vitrification technique from B6D2F1 strain belonging to the Institute of Biomodels Science and Technology ICTB / Fiocruz-RJ

Objetivou-se testar a vitrificação de ovários de camundongos do ICTB/Fiocruz. Inicialmente, fez-se coleta e maturação in vitro dos oócitos de ovários a fresco e vitrificados, bem como avaliação de estruturas no cultivo embrionário, pós-fertilização in vitro. Fêmeas B6D2F1 foram eutanasiadas para remoção dos ovários (n=60) e divididas em três grupos: grupo 1 (n=30 animais) - oócito de ovários vitrificados, maturados e fertilizados in vitro (120 fragmentos); grupo 2 (n=15) (controle 1) - oócitos coletados a fresco, maturados e fertilizados in vitro; e grupo 3 (n=15) (controle 2) - oócitos maturados in vivo e fertilizados in vitro. A técnica foi verificada no desenvolvimento embrionário in vitro, que foi avaliado pelo teste de qui-quadrado (BioStat 5.0). Recuperaram-se 123, 224 e 328 oócitos nos G1, G2 e G3, respectivamente. Observaram-se diferenças significativas nas taxas de clivagem às 24 horas (embriões ≥ 2 células) entre G1 (8%) e G2 (32%) (P<0,1) e G1 e G3 (49%) (P<0,05), mas não entre G2 e G3 (P>0,05). Para blastocistos, às 96 horas, os grupos G1, G2 e G3 apresentaram, respectivamente, 6%, 11% e 46%, diferindo significativamente entre eles (P<0,05). A vitrificação de ovários, a maturação oocitária e a fertilização in vitro são alternativas para a produção de embriões de camundongos in vitro.(AU)

This work aimed test ovarian vitrification of hybrid mouse from ICTB/Fiocruz. Protocol collection and oocyte in vitro maturation from fresh and vitrified ovaries was established and embryos were evaluated after fertilization. B6D2F1 females were euthanized for ovarian removal (n= 60) and divided into 3 groups: G1 (n= 30) - ovaries fragmented (n= 120), vitrified, matured and fertilized; G2 (n= 15) - in vitro fertilization of oocytes matured in vitro from fresh ovaries; G3 (n= 15) - ampulla region oocytes in vitro fertilizated. Viability was verified by thawing, oocyte in vitro maturation and fertilization. In vitro embryo development of each group was evaluated by Chi-square test (BioStat 5.0). 123, 224 and 328 oocytes were recovered from G1, G2 and G3, respectively. Significant differences were observed in cleavage rates at 24 hours (embryos with 2 cells or more) between G1 (8%) and G2 (32%) (P< 0.1) and G1 and G3 (49%) (P< 0.05) but not between G2 and G3 (P> 0.05). Blastocysts at 96 hours presented 6%, 11% and 46%, respectively for G1, G2 and G3, differing significantly (P< 0.05). Ovary vitrification, oocyte in vitro maturation and in vitro fertilization were available for the production of in vitro mouse embryos.(AU)

Should we expect neurological symptoms in the SARS-CoV-2 epidemic? / ¿Es esperable que haya cuadros neurológicos por la pandemia por SARS-CoV-2?

INTRODUCTION: There is growing evidence that SARS-CoV-2 can gain access to the central nervous system (CNS). We revise the literature on coronavirus infection of the CNS associated with neurological diseases. DEVELOPMENT: Neurological symptoms were rarely reported in the SARS-CoV and MERS-CoV epidemics, although isolated cases were described. There are also reports of cases of neurological symptoms associated with CoV-OC43 and CoV-229E infection. The presence of neurological lesions, especially demyelinating lesions in the mouse hepatitis virus model, may explain the mechanisms by which coronaviruses enter the CNS, particularly those related with the immune response. This may explain the presence of coronavirus in patients with multiple sclerosis. We review the specific characteristics of SARS-CoV-2 and address the question of whether the high number of cases may be associated with greater CNS involvement. CONCLUSION: Although neurological symptoms are not frequent in coronavirus epidemics, the high number of patients with SARS-CoV-2 infection may explain the presence of the virus in the CNS and increase the likelihood of early- or delayed-onset neurological symptoms. Follow-up of patients affected by the SARS-CoV-2 epidemic should include careful assessment of the CNS.

Evaluación de formulaciones inmunoterapéuticas experimentales en el modelo hibrido murino de Reaccion del Injerto contra el Hospedador / Evaluation of experimental immunotherapeutic formulations in the hybrid murine model of Graft-versus-Host Reaction

Introducción: La Enfermedad del Injerto Contra el Hospedador es la complicación más frecuente de los Trasplantes de Células Madre Hematopoyéticas y de todos los trasplantes que contengan células inmunocompetentes alogénicas, el 100 por ciento la padecen y cerca del 30 por ciento mueren por su causa; una proporción alta de casos son esteroide-refractarios, asimismo otras medidas inmunosupresoras modernas fracasan. En los campos de la Inmunoterapia y la Vaccinología también existe una escasez preocupante de inmunomoduladores de origen biológico potentes, efectivos, seguros y de amplio espectro. Existe un modelo híbrido murino de gran utilidad metodológica para estudios experimentales. Objetivo: Evaluar dos formulaciones novedosas de origen biotecnológico, una de ellas inmunopotenciadora y otra inmunosupresora, desarrolladas como cocleatos. Material y Métodos: Mediante Microscopia Electrónica y RT-PCR se caracterizaron las formulaciones como nanopartículas y su capacidad de regular la expresión del ARNm de linfoquinas definitorias de sus perfiles, respectivamente. Empleando el modelo de Enfermedad del Injerto Contra el Hospedador en ratón híbrido F1 (CBAxC57BL), se evaluó su carácter inmunomodulador in vivo . Resultados: Partiendo de los proteoliposomas de Neisseria meningitidis, se obtuvieron dos formulaciones en forma de cocleatos, ambas con diámetros de partícula inferior a 100nm. La Formulación 1mostró un perfil proinflamatorio con potente capacidad de aumentar el IFNγ y el TNFα y potenció el Índice de Bazo hasta 2,05 en el modelo EICH con p=0,0002. La Formulación 2 mostró un perfil supresor-regulatorio con potente capacidad de aumentar la IL-10 y el TGFβ y además de suprimir la producción de TNFα. En el modelo usado, esta formulación, suprimió el Índice de Bazo de manera dosis dependiente y con alta significación estadística. Se corroboró el conocido perfil de seguridad y ausencia de reactogenicidad de ambas formulaciones. Conclusiones: Ambas formulaciones tienen potencial aplicación en los campos de la terapia de Enfermedad del Injerto Contra el Hospedador en otras patologías y en Vaccinología. Los resultados obtenidos en el presente trabajo fundamentan la conveniencia de continuar el desarrollo farmacéutico y completar la preclínica de ambas formulaciones(AU)

Introduction: Graft-versus-host disease is the most frequent complication of Hematopoietic Stem Cell Transplants and all transplants containing allogeneic immunocompetent cells; 100 percent of patients suffer from this complication and about 30 percent die for this particular cause. A high proportion of cases are steroid-refractory; likewise, other modern immunosuppressive measures fail. In the fields of Immunotherapy and Vaccinology, there is also a worrying shortage of powerful, effective, safe and broad spectrum immunomodulators of biological origin. There is a hybrid murine model of great methodological utility for experimental studies. Objective: To evaluate two novel formulations of biotechnological origin: an immunopotentiator formulation and an immunosuppressive one, which were developed as cochleates. Material and Methods: The formulations assayed by Electron Microscopy and RT-PCR were characterized as nanoparticles and for their capacity to regulate lymphokine mRNA expression profile, respectively. The immunomodulatory character was evaluated in vivo using Graft-versus-host disease in (CBAxC57BL) F1 hybrid mice. Results: Starting from the proteoliposomes derived from Neisseria meningitides, two cochleate formulations were obtained, both with particle diameters below 100 nm. Formulation 1 showed a proinflammatory profile with potent capacity to increase IFNγ and TNFα, and potentiated the Spleen Index up to 2.05 in the GVDH model with p = 0.0002. Formulation 2 showed a suppressor/regulatory profile with potent capacity to increase IL-10 and TGFβ and suppress the production of TNFα. In the model used, this formulation suppressed the Spleen Index in a dose-dependent manner with high statistical significance. The known safety profile and absence of reactogenicity of both formulations was corroborated. Conclusions: Both formulations have potential application in the fields of GVHD therapy and other pathologies as well as in Vaccinology. The results obtained in the present work suggest the usefulness to continue with the pharmaceutical development and complete the preclinical studies of both formulations(AU)

Avaliação dos efeitos da Suplementação do Ácido Graxo ômega 3 em modelo de tumor de mama murino e mucosite induzida por 5-Fluouracil / Evaluation of the effects of Omega 3 Fatty Acid Supplementation in a model of murine breast tumor and mucositis induced by 5-Fluouracil

O ômega 3 é um ácido graxo poliinsaturado utilizado como agente terapêutico no tratamento de diversas doenças. Evidências sugerem efeito anti-inflamatório com sua utilização. A escolha do tratamento eficiente que minimize os efeitos adversos do tratamento oncológico tem sido desafiadora para a prática clínica. Dessa forma, ele seria interessante na prevenção e / ou tratamento da mucosite, uma inflamação que acomete as mucosas da boca ao ânus, decorrente da utilização de quimioterápico e radioterápico. As consequências deste agravo vão desde a perda de peso, anorexia e odinofagia, ao aumento do risco de translocação bacteriana, em virtude do aumento da permeabilidade intestinal (PI). Tais complicações podem levar à interrupção do tratamento quimioterápico até que o paciente se restabeleça. Trabalhos anteriores do nosso grupos de pesquisa demonstraram que o ômega 3 foi capaz de atenuar a perda de peso e reduzir a PI pela regulação da apoptose de células intestinais. Entretanto, possíveis efeitos da suplementação do ômega 3 no tumor não foram investigados. O presente estudo avaliou o efeito deste composto na mucosite induzida por quimioterápico em modelo de tumor de mama murino. Para tanto, camundongos Balb/c foram divididos nos grupos controle (CTL), controle tumor (CTLTU), tumor ômega 3 (TUW3), tumor 5-fluouracil (TU5FU) e tumor 5-fluouracil ômega 3 (TU5FUW3). Nos animais dos grupos com tumor foi realizada a inoculação das células tumorais (dia 1) e, uma vez que os tumores já se encontravam palpáveis (dia10), os grupos TUW3 e TU5FUW3 iniciaram a suplementação com ômega 3 pela ração por 10 dias, enquanto os demais animais receberam ração controle. Ao final desse período, os animais dos grupos TU5FU e TU5FUW3 receberam injeção intraperitoneal de 5-FU (300mg/kg) para indução da mucosite, enquanto os demais animais receberam a injeção de solução salina. Após 72 horas, os animais foram eutanasiados para coleta de dados. A suplementação com ômega 3 não foi capaz de prevenir a perda de peso em decorrência da mucosite. Entretanto, os animais do grupo TU5FUW3 apresentaram redução da PI por meio da recuperação das junções firmes ZO-1 e ocludina, além da preservação da arquitetura dos vilos. Os animais do grupo CTLTU também apresentaram aumento da PI e redução da expressão das proteínas de junção, provavelmente pela liberação de citocinas TNF-α e IL-1ß, conforme dosagem destas no baço. Nos animais suplementados com ômega 3 foi observado redução do tamanho do tumor e do número de metástases tumorais, além de redução da proliferação de células tumorais e aumento da afinidade das células tumorais ao quimioterápico, levando à apoptose destas. Este processo de citotoxicidade seletiva foi observado pela redução da toxicidade hepática. Diante desses resultados, pode-se concluir que a suplementação de ômega 3, por período de dez dias, foi capaz de atenuar os efeitos da mucosite induzida por quimioterápico, sem causar prejuízo à sua atividade antitumoral.

Omega 3 is a polyunsaturated fatty acid used as a therapeutic agent in the treatment of various diseases. It is widely accepted that its use can exert an anti-inflammatory effect. Thus, it would be interesting in the prevention and treatment of mucositis, which is described as an inflammation that affects mucosal membranes from the mouth to the anus, due to the use of chemotherapy and radiotherapy. Patients who suffer from this outcome, usually presents weight loss, anorexia and odynophagia,also there is an increased risk of bacterial translocation, due to the increase of the intestinal permeability (IP). Such complications can lead to the interruption of chemotherapy treatment until the patient is recovered. Previous work has shown that omega 3 was able to reduce weight loss and reduce IP by reducing intestinal cell apoptosis. However, the effects of omega 3 supplementation from the diet on the tumor remains unknown. This study aimed to investigate the effect of this compound on chemotherapy-induced mucositis in the murine breast tumor model. For that, BALB/ c mice were divided into the control (CTL), tumor control (CTLTU), omega 3 tumor (TUW3), 5-fluouracil tumor (TU5FU) and 5-fluouracil omega 3 tumor (TU5FUW3) groups. Tumor cells were inoculated (day 1) and, since they were already palpable (day 10), groups TUW3 and TU5FUW3 started receiving the chow supplemented with omega 3 for 10 days, while the other animals received control chow. At the end of this period, animals from groups TU5FU and TU5FUW3 received an intraperitoneal injection of 5-FU (300mg / kg) to induce mucositis, while the other animals received an injection of saline. After 72 hours, all animals were euthanized for data collection. Omega 3 supplementation was not able to prevent weight loss due to mucositis. However, animals from TU5FUW3 group showed a reduction in IP by recovering the tight-junctions ZO-1 and occludin, and by preventing the mucosal damage. Animals from CTLTU group also showed increased PI and reduced expression of tight-junction proteins, probably due to the release of cytokines TNF-α and IL-1ß, according to their dosage in the spleen. In addition, our results have demonstrated that animals supplemented with omega 3, have shown a reduction in the size of the tumor and the number of metastasis in lungs. We have found that the reduction was mediated through a reduction in the proliferation of tumor cells and also by an increased affinity of tumor cells to the chemotherapeutic drug, leading to their apoptosis. This selective cytotoxicity process leads to the reduction of liver toxicity. Given these results, we can conclude that omega 3 supplementation, for ten days, was able to attenuate the effects of chemotherapy-induced mucositis, without causing damage to its antitumor activity.

Evaluación de la capacidad patogénica de Fusarium spp. en modelos vegetal y murino / Evaluation of Fusarium spp. pathogenicity in plant and murine models

El género Fusarium es ampliamente conocido por su capacidad fitopatógena, típicamente asociada al marchitamiento vascular. Sin embargo, se ha reportado como un patógeno oportunista en pacientes inmunocompetentes e inmunocomprometidos, por lo que puede ser considerado como un microorganismo de interés en estudios de patogenicidad en diferentes hospederos. Este trabajo evaluó la capacidad patogénica de aislamientos de Fusarium spp. de diferentes orígenes en hospederos vegetales y en un hospedero animal (modelo murino). Doce aislamientos de Fusarium spp. de origen vegetal, animal superficial, humano superficial y humano sistémico fueron inoculados en plantas de tomate, gulupa y clavel, y en ratones BALB/c, inmunocompetentes e inmunosuprimidos. Las pruebas de patogenicidad en plantas no mostraron todos los síntomas asociados al marchitamiento vascular en los tres modelos vegetales, pero la colonización y la necrosis de los haces vasculares observada en todos los casos, independientemente de la especie de Fusarium y el origen del aislamiento, demostró el potencial infeccioso de Fusarium spp. en las diferentes especies de plantas. Por otro lado, las pruebas de patogenicidad en el modelo murino evidenciaron alteraciones del comportamiento. Asimismo, se observó en el modelo murino que todos los aislamientos infectaron y colonizaron diferentes órganos, independientemente de su origen, de la especie o del estado inmunitario del hospedero, pero solamente cinco (de diferente origen y correspondientes a diferentes especies) generaron mortalidad. En contraste, la prueba de inoculación superficial no evidenció lesiones ni colonización. Los resultados observados indican el potencial papel patogénico de los aislamientos de Fusarium spp. en los diferentes tipos de hospederos. Sin embargo, es necesario profundizar en estudios de factores de patogenicidad que expliquen la capacidad de este género para colonizar múltiples hospederos.

The genus Fusarium is widely recognized for its phytopathogenic capacity. However, it has been reported as an opportunistic pathogen in immunocompetent and immunocompromised patients. Thus, it can be considered a microorganism of interest in pathogenicity studies on different hosts. Therefore, this work evaluated the pathogenicity of Fusarium spp. isolates from different origins in plants and animals (murine hosts). Twelve isolates of Fusarium spp. from plants, animal superficial mycoses, and human superficial and systemic mycoses were inoculated in tomato, passion fruit and carnation plants, and in immunocompetent and immunosuppressed BALB/c mice. Pathogenicity tests in plants did not show all the symptoms associated with vascular wilt in the three plant models; however, colonization and necrosis of the vascular bundles, regardless of the species and origin of the isolates, showed the infective potential of Fusarium spp. in different plant species. Moreover, the pathogenicity tests in the murine model revealed behavioral changes. It was noteworthy that only five isolates (different origin and species) caused mortality. Additionally, it was observed that all isolates infected and colonized different organs, regardless of the species and origin of the isolates or host immune status. In contrast, the superficial inoculation test showed no evidence of epidermal injury or colonization. The observed results in plant and murine models suggest the pathogenic potential of Fusarium spp. isolates in different types of hosts. However, further studies on pathogenicity are needed to confirm the multihost capacity of this genus.

[Evaluation of Fusarium spp. pathogenicity in plant and murine models]. / Evaluación de la capacidad patogénica de Fusarium spp. en modelos vegetal y murino.

The genus Fusarium is widely recognized for its phytopathogenic capacity. However, it has been reported as an opportunistic pathogen in immunocompetent and immunocompromised patients. Thus, it can be considered a microorganism of interest in pathogenicity studies on different hosts. Therefore, this work evaluated the pathogenicity of Fusarium spp. isolates from different origins in plants and animals (murine hosts). Twelve isolates of Fusarium spp. from plants, animal superficial mycoses, and human superficial and systemic mycoses were inoculated in tomato, passion fruit and carnation plants, and in immunocompetent and immunosuppressed BALB/c mice. Pathogenicity tests in plants did not show all the symptoms associated with vascular wilt in the three plant models; however, colonization and necrosis of the vascular bundles, regardless of the species and origin of the isolates, showed the infective potential of Fusarium spp. in different plant species. Moreover, the pathogenicity tests in the murine model revealed behavioral changes. It was noteworthy that only five isolates (different origin and species) caused mortality. Additionally, it was observed that all isolates infected and colonized different organs, regardless of the species and origin of the isolates or host immune status. In contrast, the superficial inoculation test showed no evidence of epidermal injury or colonization. The observed results in plant and murine models suggest the pathogenic potential of Fusarium spp. isolates in different types of hosts. However, further studies on pathogenicity are needed to confirm the multihost capacity of this genus.

Efeitos à distância de feridas cirúrgicas em mucosa oral sobre populações granulocíticas na medula óssea em modelo murino / Remote effects of surgical oral mucosa wounds on populations granulocytes in the bone marrow in a murine model

Neste trabalho procuramos determinar os efeitos à distância de um ferimento cirúrgico único na cavidade bucal (localizado no palato duro, de extensão 3 mm) sobre a medula óssea em camundongos C57BL/6. Para isto, quantificamos células totais, eosinófilos, neutrófilos e fagócitos mononucleares, tanto na medula óssea recém-coletada, como na medula óssea cultivada em presença de diferentes citocinas. As citocinas foram utilizadas para revelar defeitos sutis que não são detectáveis no exame da medula óssea recém-coletada. Na medula óssea recém-coletada, foi encontrado um aumento modesto, mas significativo, nos números de neutrófilos. Em culturas incubadas com Interleucina (IL)-3, foi observada uma redução significativa dos números de células totais, neutrófilos e fagócitos mononucleares. Em culturas incubadas com Fator Estimulante de Colônias de Granulócitos (G-CSF), foram observados efeitos negativos semelhantes aos encontrados com IL-3, com supressão de células totais, neutrófilos e fagócitos mononucleares. Em culturas incubadas com Fator Estimulante de Colônias de Granulócitos e Macrófagos (GM-CSF), foi observada redução significativa somente sobre o número de neutrófilos. O mesmo ocorreu na presença de Fator Estimulante de Colônias de Macrófagos (M-CSF). Utilizamos também rhG-CSF (Filgrastim®), que tem atividade em cultura de células murinas, sendo observado efeito negativo apenas sobre neutrófilos, em contraste com o observado na presença de G-CSF murino. O efeito positivo sobre o número de neutrófilos na medula óssea recém-coletada poderia ser explicado pela exposição a microrganismos da microbiota bucal no local da lesão, causando bacteremia. Para testar esta hipótese, adotamos uma estratégia de desinfecção local (Clorhexidina a 0,2% e 0,04%) antes da realização do corte, e avaliamos a repercussão deste tratamento sobre as diferentes respostas anteriormente caracterizadas. O tratamento com Clorhexidina a 0,2% eliminou os efeitos observados anteriormente sobre os números de células totais, neutrófilos e fagócitos mononucleares. Em contrapartida, após o tratamento com Clorhexidina a 0,04% alguns dos efeitos anteriormente encontrados, quanto aos números de neutrófilos e de fagócitos mononucleares, persistiram. Procuramos também avaliar se o bloqueio da resposta de glicocorticóides endógenos ao trauma cirúrgico, pelo uso de Mifepristone (RU486), antagonista do receptor de glicocorticóides, poderia modificar os efeitos da ferida na cavidade bucal sobre a medula óssea. Não houve evidência de bloqueio específico dos efeitos do trauma com o uso de RU486, comparativamente aos controles que receberam veículo. Em experimentos-piloto, avaliamos os efeitos do trauma cirúrgico, induzido em camundongos BP-2 segundo os protocolos utilizados anteriormente em camundongos C57BL/6, obtendo evidência de respostas importantes entre cepas no padrão de resposta da medula óssea. Dentro das limitações do presente estudo, podemos concluir que o trauma cirúrgico bucal atinge a medula óssea à distância, demonstrando efeitos positivos 24h após o trauma e negativos quando utilizamos citocinas. Além disso, a presença de bactérias bucais é responsável pela depressão da resposta da medula. (AU)

In this study, we aimed to identify, using C57BL/6 mice, long-distance effects of a single, surgical oral wound of 3 mm extension in the hard palate, on the bone marrow. To do so, we enumerated total cells as well as eosinophil, neutrophil and mononuclear phagocytes, from recently harvested bone marrow, and from cultured bone marrow following different cytokine stimuli. Cytokines were used to reveal subtle effects on the recently harvest bone marrow. In recently harvested bone marrow, a small but significant increase was found in neutrophil numbers. In cultures exposed to interleukin (IL)-3 there were decreased total cell, neutrophil and mononuclear phagocyte numbers. In cultures exposed to granulocyte-colony stimulating factor (G-CSF), reduced total cell, neutrophil and mononuclear phagocyte numbers were observed resembling those in IL-3-exposed cultures. In cultures exposed to granulocytemacrophage-colony stimulating factor (GM-CSF), significant reduction was fund only in neutrophil numbers. The occurred when macrophage-colony stimulating factor (M-CSF) was used. We also used recombinant human G-CSF (rhG-CSF, Filgrastim®), which acts on murine cells, observing reduction in neutrophil numbers, unlike observations of murine G-CSF. The increase in neutrophil numbers in the fresh bone marrow might be induced by exposure of wounded surfaces to oral microorganisms and possibly by bacteremia. To test this hypothesis, the hard palate was disinfected with clorhexidine (CHX) 0.2% or 0.04%, prior to wounding. The treatment with CHX 0.2% abolished the cytokine effects in the total cell, neutrophil and mononuclear phagocyte numbers. By contrast, after treatment with CHX 0.04%, cytokine effects on the neutrophil and mononuclear phagocyte numbers persisted. We also sought to evaluate whether the effect of oral wounding on bone marrow was dependent on endogenous glucocorticoids. Blockade of the glucocorticoid antagonist receptor with mefipristone (RU486) provided results comparable to those of control group given vehicle only, thus ruling out this hypothesis. In pilot experiments, we also evaluated the effects of surgical trauma in BP-2 mice, using the protocol previously used for C57BL/6 mice, and found significant differences in the bone marrow responses between these two strains. Briefly, our limited experimental evidence shows that oral surgical trauma can influence the bone marrow at distance over a 24h postoperative period, and inducing positive or negative effects on various leukocyte lineages, before and after culture with cytokines. Available evidence points to oral bacteria as involved in these long-distance effects on bone-marrow. (AU)

Resposta imunológica a antígenos de Hysterothylacium deardorffoverstreetorum de peixes teleósteos / Imunne response against Hysterothylacium deardorffoverestreetorum from teleost fish

Anisaquidose é uma doença provocada por parasitos da família Anisakidae e se caracteriza por manifestações gastrointestinais e alérgicas. O Anisakis simplex é o parasito mais patogênico ao homem e altamente alergênico. Porém, outros anisaquídeos também são danosos aos humanos, mas é desconhecida a imunogenicidade dessas larvas. O objetivo deste trabalho foi avaliar o potencial imunogênico do parasito Hysterothylacium deardorffoverestreetorum (HD) em modelo murino. Camundongos da linhagem BALB/c foram divididos em três grupos experimentais e receberam as preparações antigênicas obtidas de larvas de HD: extrato bruto de larvas (EBH), extrato secretado/ excretado de larvas (ESH) e extrato bruto de larvas após excreção/secreção (EEH). Amostras séricas foram obtidas em diferentes dias após imunização para determinação dos níveis de anticorpos específicos pelo ensaio imunoenzimático (ELISA). Os resultados demonstram aumento na produção de imunoglobulina (Ig) G após a segunda imunização, com aumento progressivo após a terceira imunização. Já em relação à IgE, a reatividade foi mais tardia, demonstrando aumento progressivo após a terceira imunização. Foi avaliada a imunidade celular por meio da intradermorreação, como resultado estatisticamente significativo em relação ao controle utilizado. Este experimento é a primeira descrição da potencialidade patogênica desse parasito em mamíferos e representa um avanço no diagnóstico da anisaquidose humana.(AU)

Anisaquidosis is a disease caused by parasites of Anisakidae family and is characterized by gastrointestinal and allergic reactions. The Anisakis simplex is a more pathogenic Anisakidae to humans and is highly allergenic. However, other species of this family also have characteristics that are harmful to humans, but little is known about the immunogenicity this parasites. The objective of this study was to experimentally assess the immunogenic potential of the parasite Hysterothylacium deardorffoverestreetorum (H.D) in mice. Mice of inbred BALB/c strain were divided into three groups and received three immunizations of the following antigenic preparations obtained from L3 larvae H.D: Crude larval extract of H.D (CEH) Extract secreted / excreted larvae H.D. (ESH) and crude extract of larvae after excretion / secretion (EEH). Serum samples were obtained on different days after immunization to determine the levels of circulating specific antibodies by enzyme-linked immunosorbent assay (ELISA). The results show increased production of immunoglobulin (Ig) G after the second immunization with a gradual increase after the third immunization. Regarding IgE reactivity, this occurred later, demonstrating a progressive increase only after the third immunization. Cellular immunity was evaluated by intradermal, and showed statistically significant result compared to the control used. This experiment is the first description of the pathogenic potential of this parasite in mammals and represents a breakthrough in the diagnosis of human Anisakidosis.(AU)

Efeito do estresse pré-natal na regulação da inflamação alérgica pulmonar no modelo murino de asma experimental

Devido ao seu rápido crescimento, o feto é particularmente vulnerável a insultos e modificações no millieu hormonal. Este fato sugere que situações adversas experimentadas pela mãe grávida podem alterar o desenvolvimento e a saúde da prole, explicado principalmente pela permeabilidade da barreira placentária a diversos hormônios e substâncias. O objetivo deste trabalho foi estudar o efeito do estresse pré-natal na regulação da inflamação alérgica pulmonar, empregando o modelo murino de asma experimental. Para este propósito foram utilizadas camundongas virgens da linhagem Swiss, com 50 dias de idade. Foi empregado o modelo de choque nas patas para promover o estresse pré-natal e o modelo do "metrô de Nova Efeito do estresse pré-natal na regulação da inflamação alérgica pulmonar no modelo murino de asma experimental 65 Iorque" para o estresse pós-natal. As fêmeas foram distribuídas em 4 grupos experimentais: CC: fêmeas não estressadas; CE: fêmeas estressadas pós-natalmente aos 60 dias de idade (PND60), EC: fêmeas nascidas de mães estressadas entre o dia 15 (GD15) e 18 de gestação (GD18); EE: fêmeas nascidas de mães estressadas entre o GD15 e GD18 e estressadas pós-natalmente aos PND60. A indução da inflamação alérgica pulmonar foi realizada através da sensibilização dos animais com solução de ovalbumina (OVA) 0,1 mg.Kg-1 sc para avaliação do leucograma, lavado broncoalveolar (BAL), celularidade hematopoiética medular e neuroquímica. Os experimentos foram realizados 24h após a última sessão de nebulização. O número de células do BAL foi significantemente maior nos animais do grupo EE, em relação àqueles dos grupos CC (P0.05) para os linfócitos, neutrófilos, eosinófilos e monócitos; porém, observou-se diferenças significativas (P<0.05) entre o número de bastonetes dos grupos, sendo maior nos animais do grupo CC em relação àqueles do grupo EC. O número de células hematopoiéticas da medula óssea foi significantemente (P<0.05) menor nos animais do grupo EE, em relação àqueles do grupo CC. No córtex pré-frontal, há diferenças significantes na relação Ácido Homovanílico/ Dopamina (HVA/DA) (P<0.05), sendo maior nos animais do grupo EC, em relação àqueles do grupo CE. Em conclusão, o estresse pré-natal levou a modulação de células do sistema imune (SI) dos neonatos, evidenciado após a exposição a estresse agudo pós-natal, amplificando a resposta alérgica pulmonar. Sugere-se que a maior susceptibilidade dos animais do grupo EE seja consequência de alterações induzidas pelo estresse pré-natal no eixo hipotálamo-pituitária-adrenal (HPA).

Debido a su rápido crecimiento el feto es particularmente vulnerable a los cambios en el ambiente hormonal. Esto sugiere que situaciones adversas de la madre durante la gestación pueden alterar el desarrollo y la salud de la descendencia, principalmente debido a la permeabilidad de la barrera placentaria a diversas hormonas y sustancias. El objetivo del presente trabajo fue estudiar el efecto del estrés prenatal sobre la regulación de la inflamación alérgica pulmonar, empleando el modelo murino de asma experimental. Para este propósito fueron utilizadas ratonas vírgenes de linaje suizo de 50 d de edad. Fue empleado el modelo de descargas eléctricas en las patas (Footshock) para inducir el estrés prenatal y el modelo de estrés denominado "metro de Nueva York" para el estrés posnatal. Las hembras fueron divididas en 4 grupos experimentales: CC: hembras no estresadas; CE: hembras estresadas posnatalmente a los 60 d de edad (PND60); EC: hembras nacidas de madres estresadas entre el día 15 (GD15) y 18 de gestación (GD18); EE: hembras nacidas de madres estresadas entre el GD15 y GD18 y estresadas posnatalmente al PND60. La inducción de la inflamación alérgica pulmonar fue realizada a través de la sensibilización de los animales con solución de ovoalbúmina (OVA) 0,1 mg.Kg-1 sc. para posteriormente evaluar leucograma, lavado broncoalveolar (BAL), celularidad hematopoyética medular y neuroquímica. Los experimentos fueron realizados 24 horas después de la última sesión de nebulización. El número de células del BAL fue significativamente mayor en los animales del grupo EE, en comparación con los del grupo CC (P0.05) para los linfocitos, neutrófilos, eosinófilos y monocitos; sin embargo, se observaron diferencias significativas (P<0.05) entre los grupos en el número de bastonetes, siendo mayor en los animales del grupo CC en relación al grupo EC. El número de células hematopoyéticas de la médula ósea fue significativamente menor (P<0.05) en los animales del Grupo EE, en comparación con los del grupo de CC. En la corteza prefrontal, hubo diferencias significativas en la relación Ácido Homovanílico/ Dopamina (HVA/DA) (P<0.05), siendo mayor en los animales del grupo EC, en comparación con los del grupo CE. En conclusión, el estrés prenatal produjo modulación de las células del sistema inmune (SI) de los neonatos, evidenciado después de la exposición a un estrés agudo posnatal, por la amplificación de la respuesta alérgica pulmonar. Se sugiere que la mayor susceptibilidad de los animales del grupo EE sea resultado de los cambios inducidos por el estrés prenatal en el eje hipotálamo-pituitaria-adrenal (HPA).

Due to the rapid growth of the fetus it is particularly vulnerable to insults and changes in hormonal milieu. Therefore, is suggested that adverse situations experienced by the pregnant mother can alter the development and health of offspring, mainly due to the permeability of the placental barrier to various hormones and substances. The aim of the present investigation was to study the effects of prenatal stress in the regulation of pulmonary allergic inflammation, employing the murine model of experimental asthma. For this purpose, were used virgin female mice, Swiss lineage, of 50 days old. The models used were foot shock to induce prenatally stress, and "New York subway" stress to induce postnatally stress. Females were divided into 4 groups: CC group: not stressed females; CE group: postnatally stressed females (PND60); EC: females born from stressed mothers (GD15 to GD18); EE Group: females born from stressed mothers (GD15 to GD18) (footshock) and postnatally stressed (PND60). The induction of allergic pulmonary inflammation was done through sensitization of animals with 0,1 mg.Kg-1 sc of ovalbumin (OVA) solution, to further evaluate leukogram, bronchoalveolar lavage (BAL) hematopoietic marrow cellularity and neurochemistry. The experiments were performed 24 hours after the last session of nebulization. The number of BAL cells was significantly higher in EE group animals compared with the CC group (P0.05) for lymphocytes, neutrophils, eosinophils and monocytes; however, there were significant differences (P<0.05) observed in the number of rods cells between groups, being higher in animals the CC group compared to EC group. The number of hematopoietic cells of the bone marrow was significantly lower (P<0.05) in animals of Group EE, compared with CC group. In the prefrontal cortex, there were significant differences in homovanillic acid /dopamine (HVA/DA) (P<0.05) rate, being higher in the EC group, compared to EC group. In conclusion, prenatal stress modulated the immune system (SI) cells of neonates, evidenced after exposure to a post-natal acute stress by amplification of pulmonary allergic response. It is suggested that the increased susceptibility of animals EE group is a result of changes induced by prenatal stress on hypothalamus pituitary-adrenal (HPA) axis.

Technical Aspects and Benefits of Experimental Mouse Lung Transplantation. / Aspectos técnicos y utilidades del trasplante pulmonar experimental murino.

In recent years, the number of lung transplantations performed as the last option for many respiratory diseases has grown considerably, both in adults and children. However, the causes for the relatively short survival of lungs compared to other organ transplants still need to be studied. Techniques have improved since the 1950s when experimental lung transplantation began, and the different animal species used now include rodents. The advantage of using these small species is that the surgical model has been expanded and standardized, and different respiratory problems can be studied. In this review we examine the different technical strategies used in experimental transplantation in rats and mice, focusing on surgical techniques and anesthesia and monitoring methods, and highlighting the major contributions of mouse lung transplantation to the field.

Leishmaniasis in Turkey: first clinical isolation of Leishmania major from 18 autochthonous cases of cutaneous leishmaniasis in four geographical regions.

OBJECTIVE: To report isolation of Leishmania major strains obtained from 18 Turkish autochthonous cutaneous leishmaniasis (CL) patients infected with L. major between 2011 and 2014. METHODS: Initial diagnosis relied on microscopy and culture in enriched medium, prepared by adding specific amounts of liver extract, protein and lipid sources to NNN medium. Promastigotes were then transferred to RPMI medium including 10% of foetal calf serum for mass culture. Species-specific real-time PCR targeting ITS1 region of Leishmania spp. was performed using both lesion aspiration samples and cultured promastigotes. Two of 18 isolates were identified by isoenzyme analysis in the Leishmaniasis Reference Center in Montpellier, France. Each isolate was inoculated into the footpads of six mice to observe the pathogenicity of L. major. Developing lesions were observed, and the thickening of footpads was measured weekly. RESULTS: Melting curve analyses of 18 isolates showed a peak concordant with L. major, and two of them were confirmed by isoenzyme analyses as L. major zymodeme MON103. In the mouse model, acute lesions seen on day 21 were accepted as an indication of heavy infection. Severe impairments were observed on all mouse footpads over 3 weeks, which even progressed to extremity amputation. CONCLUSION: Cutaneous leishmaniasis-causing L. major was recently identified in Adana province in southern Turkey, with PCR. Our study shows that such CL cases are not limited to Adana but currently present from western to Southeastern Anatolia, and along the Mediterranean coast. The role of small mammals, the main reservoirs of L. major in Anatolia, needs to be elucidated, as do the underlying factors that cause severe clinical manifestations in L. major infections in Turkey, contrary to the infections in neighbouring countries.

Avances en las inmunoterapias para alergias alimentarias / New insights in immunotherapies for food allergies / Novas perspectivas em imunoterapias para alergias alimentares

Las enfermedades alérgicas son las inmunopatologías que con mayor prevalencia se presentan en el mundo. Pueden o no estar mediadas por anticuerpos IgE, sin embargo estas últimas son las que más intensamente se han estudiado por el riesgo que presentan para la vida del paciente. Si bien el único tratamiento que logra revertir estos mecanismos es la no exposición al alergeno, esto no siempre es posible. Por esta razón, y a partir del mayor conocimiento alcanzado del sistema inmune de mucosas junto al desarrollo de modelos animales de alergia, existe un marcado interés en la especialidad para el desarrollo de inmunoterapias que controlen y reviertan el estado de alergia. A partir de los ensayos pre-clínicos en animales y la aplicación de protocolos terapéuticos en ensayos clínicos, se han desarrollado terapias mucosales que logran inducir mecanismos de tolerancia específicos del alergeno, los cuales son capaces de revertir la sensibilización alérgica. Dado que el principal escollo siguen siendo las reacciones adversas inducidas durante el tratamiento, se requiere profundizar los estudios para desarrollar protocolos terapéuticos más seguros. En este punto la medicina traslacional encuentra un campo próspero para fortalecer las interacciones entre la ciencia básica, la aplicada y la clínica.

Allergic diseases are the most prevalent immunopathologies worldwide. Although different mechanisms -IgE-independent or IgE-dependent- can be involved in the immunopathogenesis, the latter are the most studied reactions since they can be life-threatening. Nowadays, allergen avoidance is the unique effective treatment for allergic patients. However, this is rather difficult to implement. For this reason, and based on the new insights into the mucosal immune system and the development of animal models of allergy, there is an increasing interest in developing novel therapies to control or reverse allergic disorders. Pre-clinical studies and clinical trials have been successful to prove that immunotherapies may accomplish mucosal mechanisms of allergen-specific tolerance, which are able to revoke the allergic sensitization. Since the main obstacle in these therapies still has adverse reactions induced during treatment, further studies are required to explore safe and effective therapeutic protocols. At this point, translational medicine is a flourishing field in the areas of basic science, applied science, and clinical research.

As doenças alérgicas são as imunopatologias mais prevalentes em todo o mundo. Embora possam estar mediadas ou não por anticorpos IgE, estas últimas são as reacções mais intensamente estudadas, devido ao risco que apresentam para a vida do paciente. Ainda que o único tratamento eficaz para reverter este mecanismos seja a não exposição dos pacientes ao alergeno, isto nem sempre é possível. Por este motivo, e com base nas novas perspectivas sobre o sistema imune de mucosas, junto com o desenvolvimento de modelos e para o animais de alergia, existe um interesse crescente na especialidade para o desenvolvimento de imunoterapias que controlem e revertam o estado de alergia. A partir de estudos pré-clínicos em animais e a aplicação de protocolos terapêuticos em ensaios clínico, foram desenvolvidas terapias mucosas que conseguem induzir mecanismos de tolerância específicos do alergeno, que são capazes de reverter a sensibilização alérgica. Devido a que o principal obstáculo nestas terapias continuam sendo as reações adversas induzidas durante o tratamento, é necessário realizar mais estudos para desenvolver protocolos terapêuticos mais seguros. Neste ponto, medicina translacional é um campo próspero para fortalecer as interações entre a ciência básica, a aplicada e a clínica.

Eficiency of different doses of rituximab in rheumatoid arthritis. / Eficiencia de diferentes dosis de rituximab en la artritis reumatoide.

OBJECTIVE: Evaluate the effectiveness, cost and safety of rituximab in patients with rheumatoid arthritis (RA) depending on the dose used. MATERIAL AND METHODS: Retrospective observational study conducted on 52 patients with RA treated with at least one dose of rituximab for 135.3 patient-years were included. Three treatment groups were obtained: (G1) First course and following two 1g infusions separated by 15 days; (G2) First course 2 infusions of 1g followed by 2 infusions of 500mg; (G3) First course and followed by 2 infusions of 500mg separated by 15 days. Re-treatments were administered on-demand according to the clinical activity. The retention time (Log-Rank), retreats and adverse events rates (incidence rate ratio) and treatment costs per patient-month of rituximab were analysed by groups. RESULTS: Group 2 showed a better cost-effectiveness ratio than group 1, as it was associated with a longer retention of rituximab (mean [95% CI] 65.7 [60.8 to 70.7] months vs 33.5 [22.7 to 44.3]; P<.001) and a lower rate of severe adverse events with only a slight increase in the rate of retreatment (courses/patient-year [95% CI] 1.66 [1.39 to 1.93] vs. 1.01 [0.69 to 1.34]; P=.005), and in the costs (median/patient-month, €484.89 vs. €473.45). Although group 3 was €41.20/patient-month cheaper than group 2, it was associated with a higher rate of re-treatments and shorter retention of rituximab (P<.001). CONCLUSIONS: The use of full-dose rituximab at onset, followed by reduced doses in successive courses administered on-demand retreatment may be the most cost-effective option.