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The “cancer immunogenomics” paradigm has facilitated the search for tumor-specific antigens over the last 4 years by applying comprehensive cancer genomics to tumor antigen discovery. We applied this methodology to identify tumor-specific “neoantigens” in the C57BL/6-derived GL261 and VM/Dk-derived SMA-560 tumor models. Following DNA whole-exome and RNA sequencing, high-affinity candidate neoepitopes were predicted and screened for immunogenicity by ELISPOT and tetramer analyses. GL261 and SMA-560 harbored 4,932 and 2,171 nonsynonymous exome mutations, respectively, of which less than half were expressed. To establish the immunogenicities of H-2Kb and H-2Db candidate neoantigens, we assessed the ability of the epitopes predicted in silico to be the highest affinity binders to activate tumor-infiltrating T cells harvested from GL261 and SMA-560 tumors. Using IFNγ ELISPOT, we confirmed H-2Db–restricted Imp3D81N (GL261) and Odc1Q129L (SMA-560) along with H-2Kb–restricted E2f8K272R (SMA-560) as endogenous tumor-specific neoantigens that are functionally immunogenic. Furthermore, neoantigen-specific T cells to Imp3D81N and Odc1Q129L were detected within intracranial tumors as well as cervical draining lymph nodes by tetramer analysis. By establishing the immunogenicities of predicted high-affinity neoepitopes in these models, we extend the immunogenomics-based neoantigen discovery pipeline to glioblastoma models and provide a tractable system to further study the mechanism of action of T cell–activating immunotherapeutic approaches in preclinical models of glioblastoma. Cancer Immunol Res; 4(12); 1007–15. ©2016 AACR.

Multiple myeloma (MM) is the cancer of plasma cells within the bone marrow (BM) and remains incurable. Tumor-associated macrophages (TAMs) within the tumor microenvironment often display a pro-tumor phenotype and correlate with tumor proliferation, survival, and therapy resistance. Thus, TAMs have become an emerging target of interest. IL-10 is a key immunosuppressive cytokine that leads to recruitment and development of TAMs. In this study, we investigated the role of IL-10 in MM TAM development as well as the therapeutic application of IL-10/IL-10R signaling inhibition. We demonstrated that IL-10 is overexpressed in MM BM and mediates M2-like polarization of TAMs in patient BM, 3D co-cultures in vitro, and mouse models. In turn, TAMs promote MM proliferation and drug resistance, both in vitro and in vivo. Moreover, inhibition of IL-10/IL-10R pathway using a blocking IL-10R antibody prevented M2 polarization of TAMs and the consequent TAM-induced proliferation of MM, and re-sensitized MM to therapy, in vitro and in vivo. Therefore, our findings suggest that inhibition of IL-10/IL-10R axis is a novel immunotherapy strategy with monotherapy efficacy and can be further combined with current anti-MM therapy to overcome drug resistance. Future investigation is warranted to evaluate the potential of such therapy in MM patients.

PurposeProfessional program assessment is necessary in an accreditation process, in order to ensure educational quality and public accountability. One avenue of assessment is through an internship. The challenge is to determine how evidence from this indirect learning experience can aid in accreditation. The purpose of this paper is to describe the use of internship supervisor evaluation feedback within the accreditation process for a professional interior design degree program.Design/methodology/approachIn this study, internship assessment was provided by feedback from intern supervisors. Ten years of supervisor feedback were analyzed using descriptive statistics and a content analysis of supervisor comments.FindingsTwo hundred forty-seven internship supervisor evaluation documents were analyzed. Overall, supervisors positively evaluated the performance of the intern as Good to Excellent. A majority of supervisors (91%) provided comments that were positive yet vague, as most could not differentiate between the intern and the intern's performance.Practical implicationsThis study links experiential learning to its evidence that can be used in an accreditation process. The challenges for educators in developing an assessment tool useful for accreditation evidence and to be shared by multiple program degree stakeholders are also described.Originality/valueResearch on internships usually focuses on the student's viewpoint. This study is original in that it examines the use of internship supervisor's evidence in program accreditation.

Background Recent investigations of the meninges have highlighted the importance of the dura layer in central nervous system immune surveillance beyond a purely structural role. However, our understanding of the meninges largely stems from the use of pre-clinical models rather than human samples. Methods Single-cell RNA sequencing of seven non-tumor-associated human dura samples and six primary meningioma tumor samples (4 matched and 2 non-matched) was performed. Cell type identities, gene expression profiles, and T cell receptor expression were analyzed. Copy number variant (CNV) analysis was performed to identify putative tumor cells and analyze intratumoral CNV heterogeneity. Immunohistochemistry and imaging mass cytometry was performed on selected samples to validate protein expression and reveal spatial localization of select protein markers. Results In this study, we use single-cell RNA sequencing to perform the first characterization of both non-tumor-associated human dura and primary meningioma samples. First, we reveal a complex immune microenvironment in human dura that is transcriptionally distinct from that of meningioma. In addition, we characterize a functionally diverse and heterogenous landscape of non-immune cells including endothelial cells and fibroblasts. Through imaging mass cytometry, we highlight the spatial relationship among immune cell types and vasculature in non-tumor-associated dura. Utilizing T cell receptor sequencing, we show significant TCR overlap between matched dura and meningioma samples. Finally, we report copy number variant heterogeneity within our meningioma samples. Conclusions Our comprehensive investigation of both the immune and non-immune cellular landscapes of human dura and meningioma at single-cell resolution builds upon previously published data in murine models and provides new insight into previously uncharacterized roles of human dura.

Recent investigation of the meninges, specifically the dura layer, has highlighted its importance in CNS immune surveillance beyond a purely structural role. However, most of our understanding of the meninges stems from the use of pre-clinical models rather than human samples. In this study, we use single cell RNA-sequencing to perform the first characterization of both non-tumor-associated human dura and meningioma samples. First, we reveal a complex immune microenvironment in human dura that is transcriptionally distinct from that of meningioma. In addition, through T cell receptor sequencing, we show significant TCR overlap between matched dura and meningioma samples. We also identify a functionally heterogeneous population of non-immune cell types and report copy-number variant heterogeneity within our meningioma samples. Our comprehensive investigation of both the immune and non-immune cell landscapes of human dura and meningioma at a single cell resolution provide new insight into previously uncharacterized roles of human dura.

We compared the performance of two 96-well multiplex immunoassay platforms in assessing plasma cytokine concentrations in patients with glioblastoma (GBM; n = 27), individuals with melanoma, breast or lung cancer metastases to the brain (n = 17), and healthy volunteers (n = 11). Assays included a bead-based fluorescence MILLIPLEX® assay/Luminex (LMX) platform and 4 planar electrochemiluminescence kits from Meso Scale Discovery (MSD). The LMX kit evaluated 21 cytokines and the 3 MSD kits evaluated 20 cytokines in total, with 19 overlapping human cytokines between platforms (GM-CSF, IFNγ, IL-1β, IL-2, IL-4, IL-5, IL-6, IL-7, IL-8, IL-10, IL-12p70, IL-13, IL-17A, IL-21, IL-23, MIP-1α, MIP-1β, MIP-3α, TNFα). The MSD platform had lower LLoQs (lower limits of quantification) than LMX for 17/19 cytokines, and higher LLoQs for IFN-γ and IL-21. The ULoQs were higher in LMX versus MSD assays for 17/19 shared analytes, but lower than MSD for IL-17A and IL-21. With LMX, all 19 shared analytes were quantifiable in each of 55 samples. Although MSD recombinant protein standard curves indicated lower LLoQs than LMX for most cytokines, MSD detected 7/19 (37%) native analytes in

Immunologic evaluation shows that most COVID-19 patients exhibit targeted immunosuppression compared to patients with influenza., We pursued a study of immune responses in coronavirus disease 2019 (COVID-19) and influenza patients. Compared to patients with influenza, patients with COVID-19 exhibited largely equivalent lymphocyte counts, fewer monocytes, and lower surface human leukocyte antigen (HLA)–class II expression on selected monocyte populations. Furthermore, decreased HLA-DR on intermediate monocytes predicted severe COVID-19 disease. In contrast to prevailing assumptions, very few (7 of 168) patients with COVID-19 exhibited cytokine profiles indicative of cytokine storm syndrome. After controlling for multiple factors including age and sample time point, patients with COVID-19 exhibited lower cytokine levels than patients with influenza. Up-regulation of IL-6, G-CSF, IL-1RA, and MCP1 predicted death in patients with COVID-19 but were not statistically higher than patients with influenza. Single-cell transcriptional profiling revealed profound suppression of interferon signaling among patients with COVID-19. When considered across the spectrum of peripheral immune profiles, patients with COVID-19 are less inflamed than patients with influenza.

Encouraging clinical results using immune checkpoint therapies to target the PD-1 axis in a variety of cancer types have paved the way for new immune therapy trials in brain tumor patients. However, the molecular mechanisms that regulate expression of the PD-1 pathway ligands, PD-L1 and PD-L2, remain poorly understood. To address this, we explored the cell-intrinsic mechanisms of constitutive PD-L1 and PD-L2 expression in brain tumors. PD-L1 and PD-L2 expression was assessed by flow cytometry and qRT-PCR in brain tumor cell lines and patient tumor-derived brain tumor-initiating cells (BTICs). Immunologic effects of PD-L2 overexpression were evaluated by IFN-γ ELISPOT. CD274 and PDCD1LG2 cis-regulatory regions were cloned from genomic DNA and assessed in full or by mutating and/or deleting regulatory elements by luciferase assays. Correlations between clinical responses and PD-L1 and PD-L2 expression status were evaluated in TCGA datasets in LGG and GBM patients. We found that a subset of brain tumor cell lines and BTICs expressed high constitutive levels of PD-L1 and PD-L2 and that PD-L2 overexpression inhibited neoantigen specific T cell IFN-γ production. Characterization of novel cis-regulatory regions in CD274 and PDCD1LG2 lead us to identify that GATA2 is sufficient to drive PD-L1 and PD-L2 expression and is necessary for PD-L2 expression. Importantly, in TCGA datasets, PD-L2 correlated with worse clinical outcomes in glioma patients.. By perturbing GATA2 biology, targeted therapies may be useful to decrease inhibitory effects of PD-L2 in the microenvironment.

Coronavirus disease 2019 (COVID-19) is characterized by a high incidence of acute respiratory failure. The underlying immunopathology of that failure and how it compares to other causes of severe respiratory distress, such as influenza virus infection, are not fully understood. Here we addressed this by developing a prospective observational cohort of COVID-19 and influenza subjects with varying degrees of disease severity and assessing the quality and magnitude of their immune responses at the cellular and protein level. Additionally, we performed single-cell RNA transcriptional profiling of peripheral blood mononuclear cells from select subjects. The cohort consists of 79 COVID-19 subjects, 26 influenza subjects, and 15 control subjects, including 35 COVID-19 and 7 influenza subjects with acute respiratory failure. While COVID-19 subjects exhibited largely equivalent or greater activated lymphocyte counts compared to influenza subjects, they had fewer monocytes and lower surface HLA-class II expression on monocytes compared to influenza subjects and controls. At least two distinct immune profiles were observed by cytokine levels in severe COVID-19 patients: 3 of 71 patients were characterized by extreme inflammation, with greater than or equal to ~50% of the 35 cytokines measured greater than 2 standard deviations from the mean level of other severe patients (both influenza and COVID-19); the other immune profile, which characterized 68 of 71 subjects, had a mixed inflammatory signature, where 28 of 35 cytokines in COVID-19 patients had lower mean cytokine levels, though not all were statistically significant. Only 2 cytokines were higher in COVID-19 subjects compared to influenza subjects (IL-6 and IL-8). Influenza and COVID-19 patients could be distinguished statistically based on cytokine module expression, particularly after controlling for the significant effects of age on cytokine expression, but again with lower levels of most cytokines in COVID-19 subjects. Further, high circulating levels of IL-1RA and IL-6 were associated with increased odds of intubation in the combined influenza and COVID-19 cohort [OR = 3.93 and 4.30, respectively] as well as among only COVID-19 patients. Single cell transcriptional profiling of COVID-19 and influenza subjects with respiratory failure identified profound suppression in type I and type II interferon signaling in COVID-19 patients across multiple clusters. In contrast, COVID-19 cell clusters were enriched for alterations in metabolic, stress, and apoptotic pathways. These alterations were consistent with an increased glucocorticoid response in COVID-19 patients compared to influenza. When considered across the spectrum of innate and adaptive immune profiles, the immune pathologies underlying severe influenza and COVID-19 are substantially distinct. The majority of COVID-19 patients with acute respiratory failure do not have a cytokine storm phenotype but instead exhibit profound type I and type II IFN immunosuppression when compared to patients with acute influenza. Upregulation of a small number of inflammatory mediators, including IL-6, predicts acute respiratory failure in both COVID-19 and influenza patients.

Background Although clinical trials testing immunotherapies in glioblastoma (GBM) have yielded mixed results, new strategies targeting tumor-specific somatic coding mutations, termed “neoantigens,” represent promising therapeutic approaches. We characterized the microenvironment and neoantigen landscape of the aggressive CT2A GBM model in order to develop a platform to test combination checkpoint blockade and neoantigen vaccination. Methods Flow cytometric analysis was performed on intracranial CT2A and GL261 tumor-infiltrating lymphocytes (TILs). Whole-exome DNA and RNA sequencing of the CT2A murine GBM was employed to identify expressed, somatic mutations. Predicted neoantigens were identified using the pVAC-seq software suite, and top-ranking candidates were screened for reactivity by interferon-gamma enzyme linked immunospot assays. Survival analysis was performed comparing neoantigen vaccination, anti-programmed cell death ligand 1 (αPD-L1), or combination therapy. Results Compared with the GL261 model, CT2A exhibited immunologic features consistent with human GBM including reduced αPD-L1 sensitivity and hypofunctional TILs. Of the 29 CT2A neoantigens screened, we identified neoantigen-specific CD8+ T-cell responses in the intracranial TIL and draining lymph nodes to two H2-Kb restricted (Epb4H471L and Pomgnt1R497L) and one H2-Db restricted neoantigen (Plin2G332R). Survival analysis showed that therapeutic neoantigen vaccination with Epb4H471L, Pomgnt1R497L, and Plin2G332R, in combination with αPD-L1 treatment was superior to αPD-L1 alone. Conclusions We identified endogenous neoantigen specific CD8+ T cells within an αPD-L1 resistant murine GBM and show that neoantigen vaccination significantly augments survival benefit in combination with αPD-L1 treatment. These observations provide important preclinical correlates for GBM immunotherapy trials and support further investigation into the effects of multimodal immunotherapeutic interventions on antiglioma immunity. Key Points 1. Neoantigen vaccines combined with checkpoint blockade may be promising treatments. 2. CT2A tumors exhibit features of human GBM microenvironments. 3. Differential scanning fluorimetry assays may complement in silico neoantigen prediction tools.

Histologic review of bone marrow trephine biopsies is a central component of the diagnostic and treatment response evaluation of hematologic malignancies. Well-validated antibody reagents are routinely used for immunohistochemistry of these samples to provide additional insight into abnormal antigen expression. However, current immunohistochemistry staining protocols are typically limited to only one or two markers simultaneously. Dysplastic changes in cellular morphology and dyssynchronous expression of lineage markers are common features of myelodysplastic syndromes, myeloproliferative neoplasms and secondary acute leukemias. We have integrated the use of multiple diagnostic validated antibody clones with additional antibodies for hematologic lineages and structural proteins to create a 30-marker panel for imaging mass cytometry (IMC). Antibodies included in this panel identify myeloid, lymphoid, erythroid, macrophage, vascular, megakaryocyte and stromal markers as well as markers of cellular proliferation and apoptosis. Through conjugation to elemental metal tags, the entire panel is stained simultaneously on the tissue sample, then acquired by time-of-flight mass spectrometry on a Hyperion instrument (Fluidigm). Antibody staining concentrations and antigen retrieval conditions were optimized for formalin-fixed paraffin-embedded (FFPE) bone marrow to obtain consistent staining for all markers on the panel. Redundant markers for cell populations were selected to provide further internal validation of the observed staining patterns. After data acquisition, cell segmentation algorithms using CellProfiler and ilastik were applied to quantify marker expression in single cells and Phenograph in HistoCAT was used for cell population clustering. Cluster identities for all cells are associated with the original image location in order to plot the spatial arrangement of populations. Using this highly multiplexed panel, we have imaged sets of bone marrow specimens from patients with normal bone marrow morphology and those with myeloid malignancies. We initially confirmed the staining patterns expected for each antibody patterns of co-expression of lineage markers in normal bone marrow samples. We then extended this panel to examine biopsies from patients with myelodysplastic syndrome, myelofibrosis, and secondary acute myeloid leukemia. We found a clear population of CD71+ CD235a+ erythroid cells with strong expression of the proliferative marker Ki67 located within erythroid islands in normal bone marrow samples and MDS. Cell markers of apoptosis and DNA damage are scattered at low frequency throughout the bone marrow in samples with normal bone marrow morphology, but increased clusters of the DNA damage marker phospho-H2AX are observed in selected cases of myelodysplastic syndromes. Overall, this IMC imaging approach is able to extend the current clinical immunostaining for myeloid malignancies by identifying all major bone marrow cell populations. Through highly multiplexed analysis of bone marrow cell populations, the spatial architecture of cell populations and stromal structures can be elucidated, including erythroid islands, lymphoid aggregates and changes in vascular structures with increasing severity of myelofibrosis. In ongoing studies, the development of these imaging techniques for analysis of archived FFPE bone marrow samples is being applied to translational research on hematologic diseases. Disclosures Oh: Kartos Therapeutics: Consultancy; Disc Medicine: Consultancy; PharmaEssentia: Consultancy; Constellation: Consultancy; CTI Biopharma: Consultancy; Celgene/Bristol Myers Squibb: Consultancy; Blueprint Medicines: Consultancy; Novartis: Consultancy; Gilead Sciences: Consultancy; Incyte Corporation: Consultancy.

Neoantigens represent promising targets for personalized cancer vaccine strategies. However, the feasibility of this approach in lower mutational burden tumors like glioblastoma (GBM) remains unknown. We have previously reported the use of an immunogenomics pipeline to identify candidate neoantigens in preclinical models of GBM. Here, we report the application of the same immunogenomics pipeline to identify candidate neoantigens and guide screening for neoantigen-specific T cell responses in a patient with GBM treated with a personalized synthetic long peptide vaccine following autologous tumor lysate DC vaccination. Following vaccination, reactivity to three HLA class I- and five HLA class II-restricted candidate neoantigens were detected by IFN-γ ELISPOT in peripheral blood. A similar pattern of reactivity was observed among isolated post-treatment tumor-infiltrating lymphocytes. Genomic analysis of pre- and post-treatment GBM reflected clonal remodeling. These data demonstrate the feasibility and translational potential of a therapeutic neoantigen-based vaccine approach in patients with primary CNS tumors.

The “cancer immunogenomics” paradigm has facilitated the search for tumor-specific antigens over the last 4 years by applying comprehensive cancer genomics to tumor antigen discovery. We applied this methodology to identify tumor-specific “neoantigens” in the C57BL/6-derived GL261 and VM/Dk-derived SMA-560 tumor models. Following DNA whole-exome and RNA sequencing, high-affinity candidate neoepitopes were predicted and screened for immunogenicity by ELISPOT and tetramer analyses. GL261 and SMA-560 harbored 4,932 and 2,171 nonsynonymous exome mutations, respectively, of which less than half were expressed. To establish the immunogenicities of H-2Kb and H-2Db candidate neoantigens, we assessed the ability of the epitopes predicted in silico to be the highest affinity binders to activate tumor-infiltrating T cells harvested from GL261 and SMA-560 tumors. Using IFNγ ELISPOT, we confirmed H-2Db–restricted Imp3D81N (GL261) and Odc1Q129L (SMA-560) along with H-2Kb–restricted E2f8K272R (SMA-560) as endogenous tumor-specific neoantigens that are functionally immunogenic. Furthermore, neoantigen-specific T cells to Imp3D81N and Odc1Q129L were detected within intracranial tumors as well as cervical draining lymph nodes by tetramer analysis. By establishing the immunogenicities of predicted high-affinity neoepitopes in these models, we extend the immunogenomics-based neoantigen discovery pipeline to glioblastoma models and provide a tractable system to further study the mechanism of action of T cell–activating immunotherapeutic approaches in preclinical models of glioblastoma. Cancer Immunol Res; 4(12); 1007–15. ©2016 AACR.

Medicinal plants have been used for centuries as an alternative treatment for health problems. Schinus terebinthifolius Raddi is a median tree that belongs to the Anacardiaceae family. The antibacterial effect of leaves extract of this plant has already been demonstrated. The objective of this study was to test a substance derived from this plant to be used in pre- and post-milking teat antisepsis. The hydroalcoholic extract of Brazilian pepper tree was used in opposite quarters for twelve consecutive weeks, and commercial iodine was used as control. None of the indices analyzed - black background mug, CMT, intramammary infections, skin health, and teat health - showed a statistical difference between the treatments, suggesting that the plant extract can be used in pre- and post-milking teat antisepsis, as a substitute for conventional products for herds in an agroecological production system. Resumo As plantas medicinais têm sido usadas há séculos como alternativa de tratamento para problemas de saúde. Schinus terebinthifolius Raddi ou aroeira é uma árvore mediana pertencente à família Anacardiaceae. O efeito antibacteriano de extratos de folhas dessa planta já foi demonstrado. O objetivo deste estudo foi testar o extrato de Schinus terebinthifolius Raddi na antissepsia de tetos pré e pós-ordenha. Foi utilizado extrato hidroalcoólico da aroeira, tendo como controle iodo comercial, em quartos opostos, durante doze semanas consecutivas. Nenhum dos índices analisados - caneca de fundo escuro, CMT, novas infecções intramamárias, "saúde da pele e do esfíncter do teto" - demonstraram diferenças estatísticas entre os tratamentos usados, sugerindo que o extrato da planta pode ser usado na pré e pós-ordenha para desinfecção de tetos, como substituto dos produtos convencionais, para rebanhos em sistema de produção orgânica.

Submitted by Ubirajara Cruz (ubirajara.cruz@gmail.com) on 2017-03-27T16:35:17Z No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Tese_Diane_Schiavon.pdf: 775620 bytes, checksum: ef7ee3eff59d2a06e02224b7b6522877 (MD5) Approved for entry into archive by Aline Batista (alinehb.ufpel@gmail.com) on 2017-03-27T20:55:16Z (GMT) No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Tese_Diane_Schiavon.pdf: 775620 bytes, checksum: ef7ee3eff59d2a06e02224b7b6522877 (MD5) Made available in DSpace on 2017-03-27T20:55:16Z (GMT). No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Tese_Diane_Schiavon.pdf: 775620 bytes, checksum: ef7ee3eff59d2a06e02224b7b6522877 (MD5) Previous issue date: 2015-09-23 Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES A utilização dos vegetais é uma prática usual, principalmente para as comunidades rurais. Uma das formas de se obter esses dados sobre as plantas medicinais é através de levantamentos etnobotânicos. É através da etnobotânica que se busca o conhecimento e o resgate do saber tradicional particularmente relacionado ao uso dos recursos da flora. O trabalho tem como objetivo resgatar o conhecimento etnobotânico de informantes sobre plantas medicinais do município de Pelotas e Capão do Leão e validar in vitro a ação antibacteriana das plantas que possuem este fim, frente às principais bactérias isoladas do úbere bovino com mastite. Foi realizado um resgate etnobotânico com três informantes. Foram identificadas 83 espécies vegetais distribuídas em 40 famílias botânicas, sendo as famílias Asteraceae e Lamiaceae as de maior número de espécie sendo a folha a parte da planta mais utilizada. Das 83 plantas identificadas, 26 foram citadas como antimicrobianos e validadas através da Técnica de Microdiluição em Caldo. Foram 16 as espécies que apresentam alguma ação antibacteriana frente às bactérias testadas. Podemos concluir que as informantes utilizam uma grande diversidade de plantas medicinais tanto na cura como na prevenção das doenças e que as espécies pariparoba (Piper regnellii (Miq.) C. DC.) e teta de cadela (Zanthoxylum astrigerum (R.S.Cowan) P.G. Waterman) são as mais promissoras dentre as testadas, podendo ser posteriormente realizados estudos in vivo com a finalidade de determinar sua ação na prevenção da mastite bovina. The use of plant is a common practice, especially for rural communities. One way to get this data on medicinal plants is through ethnobotanical surveys. It is through ethnobotany that seeks knowledge and the rescue of traditional knowledge particularly related to the use of flora resources. The study aims to rescue the ethnobotanical knowledge of informants on medicinal plants of Pelotas and Capão do Leão and validate the in vitro antibacterial activity of the plants that hold this end, against the major bacteria isolated from bovine udder with mastitis. An ethnobotanical rescue with three informants was conducted. We identified 83 plant species belonging to 40 botanical families, with the Asteraceae and Lamiaceae families the most representative in number of species and the sheet of the most used plant. Of the 83 plants identified, 26 were cited as antimicrobial and validated through the microdilution technique in broth. There were 16 plants that have some antibacterial action in the face of bacteria tested. We can conclude that the informants use a wide variety of medicinal plants both in healing and in disease prevention and the pariparoba species (Piper reginelli (Miq.) C. DC.) And bitch theta (Zanthoxylum astrigerum (RSCowan) PG Waterman ) are the most promising among the different and may be performed later in vivo studies with the purpose of determining its effect on the prevention of bovine mastitis.

Abstract —Older oral contraceptive (OC) formulations containing high doses of potent synthetic estrogens and progestins are associated with increased risk of thrombosis. To examine the effects of current low-dose OC and hormone replacement therapy (HRT) regimens on arterial thrombosis, premenopausal and surgically postmenopausal cynomolgus monkeys were divided into four treatment groups. Premenopausal monkeys were given either no OCs or ethinyl estradiol and levonorgestrel as an OC at a dose equivalent to that currently given to women. Postmenopausal monkeys were given either no HRT or conjugated equine estrogens and medroxyprogesterone as an HRT at a dose equivalent to that currently given to women. The monkeys were fed an atherogenic diet containing these treatments for 27 to 30 months. At the end of this time, arterial thrombosis was evaluated with a standardized stenosis/injury procedure in the left carotid artery. Blood flow velocity was monitored for cyclic or permanent occlusive thrombosis. The current OC and HRT regimens did not increase the susceptibility of the artery wall to develop an occlusive thrombus following injury and stenosis. In fact, there was a reduction in the incidence of thrombosis in the OC animals compared with untreated controls. Increased amounts of atherosclerosis were associated with an increased incidence of occlusive arterial thrombosis. Several selected coagulation parameters [von Willebrand factor, protein C, lipoprotein(a), and platelet aggregation] did not appear to be associated with either the amount of atherosclerosis or incidence of arterial thrombosis.

The peptidoglycan-polysaccharide (PGPS) model using inbred rats closely mimics Crohn's disease. Our aim was to identify mouse strains that develop ileocolitis in response to bowel wall injection with PGPS. Mouse strains studied included NOD2 knockout animals, RICK/RIP2 knockout animals, and genetically inbred strains that are susceptible to inflammation. Mice underwent laparotomy with intramural injection of PGPS or human serum albumin in the terminal ileum, ileal Peyer's patches, and cecum. Gross abdominal score, cecal histologic score, and levels of pro-fibrotic factor mRNAs were determined 20 to 32 days after laparotomy. PGPS-injected wild-type and knockout mice with mutations in the NOD2 pathway had higher abdominal scores than human serum albumin-injected mice. The RICK knockout animals tended to have higher mean abdominal scores than the NOD2 knockout animals, but the differences were not significant. CBA/J mice were shown to have the most robust response to PGPS, demonstrating consistently higher abdominal scores than other strains. Animals killed on day 26 had an average gross abdominal score of 6.1 ± 1.5, compared with those on day 20 (3.0 ± 0.0) or day 32 (2.8 ± 0.9). PGPS-injected CBA/J mice studied 26 days after laparotomy developed the most robust inflammation and most closely mimicked the PGPS rat model and human Crohn's disease.

Tagetes minuta L. é uma planta aromática vulgarmente conhecida como chinchilho, cravo-de-defunto, chinchila, picão-do-reino entre outros. O objetivo do trabalho foi executar uma revisão sistemática sobre Tagetes minuta com a finalidade de identificar os principais componentes químicos, usos populares e atividades biológicas. Como estratégia de busca da revisão sistemática, a palavra-chave “Tagetes minuta” foi utilizada no Scielo, Pubmed e Google acadêmico. Na pesquisa inicial, foram identificadas 429 publicações, sendo 384 no Google acadêmico, 9 no Scielo e 36 no Pubmed. Após a leitura do título e do resumo, foram avaliados 111 trabalhos por se enquadrarem dentro dos critérios de inclusão. Na segunda etapa, foi realizada a leitura da publicação e se estas se enquadravam nos objetivos do trabalho, a publicação era selecionada. Com este procedimento, 51 publicações foram escolhidas. A revisão sistemática mostrou que a planta Tagetes minuta, além de ser popularmente conhecida por indicações medicinais, foi cientificamente testada e apresentou atividade antibacteriana, antifúngica, larvicida, inseticida, antiparasitária, nematicida, anti-hiperglicêmica, antitumoral, biocida e antioxidante.

Made available in DSpace on 2014-08-20T14:38:03Z (GMT). No. of bitstreams: 1 dissertacao_diane_bender_schiavon.pdf: 500386 bytes, checksum: 48fa2cd250b94f5f938cb28a34fe2107 (MD5) Previous issue date: 2011-02-24 Bovine Mastitis is a disease that causes high damage to milk production. Phytotherapy is the basis of Tagetes minuta L. can be used for disease control agroecological production units. The study aimed to evaluate application of antiseptics from medicinal plant from the postmilking teat disinfection. The treatments consisted of commercial iodine with treatment 1, used as teat pos-dipping on the right side, and hydro alcoholic extract from leaves of Tagetes minuta and macerated seeds Linum usitatissimum L., both at 10%, used on teats left as pos-dipping, cows in a commercial production unit that had, in average of 60 milking animals during the experiment that lasted 12 weeks. The prevalence of subclinical mastitis was evaluated using the CMT test once a week. Test mug of dark background was done at all milking. Sample of milk from all quarters was collected weekly for the isolation and characterization of present microorganisms . The incidence of new intramammary infections was calculated using the number of infection-free days by quarters, counted by weekly analysis of milk from all quarters in blood agar culture, as the denominator and the number of new cases as the numerator, defined as culture positive by Staphylococcus spp. or Streptococcus spp. in Sampling of milk per room. The index was corrected to 1000 / 4 days. Exclusion criteria: infection with Streptococcus spp or Staphylococcus spp in early labor and up to two weeks after a collection microbiologically positive. Animals with two successive collections negative after a positive were considered cured and reintroduced in the counting of days without infection. The chi-square test was used to compare the frequency of events between treatments. The CMT's weekly prevalence ranged in group 1 between 29.5% and 17.1%, and group 2 from 29.7% to 19.6%, not significantly different in any of weeks. The incidence of positive cultures for Staphylococcus / Streptococcus was 3.93 and 6.96 / 1000 / 4 / day for Groups 1 and 2 respectively, p = 0.057. There were four cases of mastitis clinic during the experiment, two in each treatment. We concluded the use of plant extracts for the disinfection of teats pos-dipping can be useful systems for milk production agro ecological. Mastite bovina é a doença que causa os maiores prejuízos na produção leiteira. A fitoterapia a base de Tagetes minuta pode ser utilizada para controle da enfermidade em unidades de produção agroecológicas. O trabalho objetivou avaliar a aplicação de anti-sépticos obtidos de planta medicinal na desinfecção de tetos pós-ordenha. Os tratamentos consistiram em iodo comercial como tratamento 1, utilizado como pós-dipping nos tetos do lado direito, e extrato hidroalcoolico de folhas de Tagetes minuta L. e macerado de sementes de Linum usitatissimum L., ambos à 10%, utilizados nos tetos esquerdos como pos-dipping de vacas em uma unidade de produção comercial que possuía, em média, 60 animais em ordenha durante o experimento que teve duração de 12 semanas. A prevalência de mastite sub-clínica foi avaliada utilizando o teste de CMT uma vez por semana. Teste da caneca de fundo escuro foi realizado em todas as ordenhas. Amostra de leite de todos os quartos foi coletada semanalmente para isolamento e caracterização dos microrganismos presentes. A incidência de novas infecções intra-mamárias foi calculada utilizando o número de dias livres de infecção por quarto, contados pela análise semanal de leite de todos os quartos em cultura de Agar sangue, como denominador, e o número de casos novos como numerador, definido como cultura positiva para Staphylococcus spp. ou Streptococcus spp. na amostragem do leite por quarto. O índice foi corrigido para 1000 quartos dia. Foram critérios de exclusão: infecção por Streptococcus spp ou Staphylococcus spp no início do trabalho e até duas semanas após uma coleta microbiologicamente positiva. Animais com duas coletas sucessivas negativas após uma positiva foram considerados curados e reintroduzidos na contagem de dias sem infecção. O teste do qui-quadrado foi utilizado para comparação das frequências dos eventos entre os tratamentos. A prevalência semanal do CMT variou no grupo 1 entre 29,5% e 17,1%, e no grupo 2 de 29,7% a 19,6%, não diferindo significativamente em nenhuma das semanas. A incidência de cultura positiva para Staphylococcus/Streptococcus foi de 3,93 e 6,96/1000 quartos/dia para os grupos 1 e 2 respectivamente, com p=0,057. Houve quatro casos de mastite clínica durante o experimento, dois em cada tratamento. Concluímos que o uso de extratos de plantas na desinfecção de tetos pós-ordenha pode ser útil aos sistemas de produção de leite agroecológico.

Introduction: bovine mastitis is a major problem in dairy production, mainly caused by Gram-positive bacteria. The search for active ingredients that act upon these microorganisms is growing due to occurrence of bacterial multiresistance. Objectives: to assess the minimum bactericidal concentration (MBC) of Origanum vulgare L. (oregano) essential oil against bacteria isolated from mastitic milk. Methods: the antimicrobial activity of O. vulgare essential oil was measured against 71 bacteria from the genera Streptococcus, Staphylococcus and Corynebacterium isolated from the bovine milk and against three pattern strains, that is, Pseudomonas aeruginosa, Staphylococcus aureus and Escherichia coli. The technique was dilution in microplate. Results: the mean minimal bactericidal concentration (MBC) ranged from 0.23 % to 2 % against the bacteria isolated from bovine milk, with the lowest concentration for Streptococcus genus and the highest for coagulase-negative Staphylococcus. Regarding the pattern strains, the MBC was 3.17 % and 0.35 % for S. aureus and Escherichia coli respectively and showed no effect for Pseudomonas aeruginosa. Conclusions: the results confirmed the in vitro activity of Origanum vulgare L. oil against the bovine mastitis-related bateria. Introducción: la mastitis bovina es el mayor problema en la producción lechera, causada principalmente por bacterias grampositivas. La búsqueda de principios activos que actúen en esos microrganismos es creciente, sobre todo por la ocurrencia de multirresistencia bacteriana. Objetivos: evaluar la concentración bactericida mínima del aceite esencial de Origanum vulgare L. (orégano) frente a bacterias aisladas de leche mastítica. Métodos: se evaluó la actividad antimicrobiana del aceite esencial de O. vulgare frente a 71 bacterias aisladas de leche bovina, de los géneros Streptococcus, Staphylococcus y Corynebacterium; y 3 cepas patrón de Pseudomonas aeruginosa, Staphylococcus aureus y Escherichia coli. La técnica utilizada fue de dilución en microplaca. Resultados: la concentración bactericida mínima media varió de 0,23 a 2 % frente a las bacterias aisladas de leche bovina, con la menor concentración para el género Streptococcus y la mayor para Staphylococcus coagulasa negativa. En cuanto a las cepas patrones la concentración bactericida mínima fue de 3,17 y 0,35 % para S. aureus y Escherichia coli, respectivamente; no presentó efecto para Pseudomonas aeruginosa. Conclusiones: en los resultados se comprobó la actividad in vitro del aceite de orégano frente a las bacterias relacionadas con la mastitis bovina.

The regulation of neutrophil functions by Type I cGMP-dependent protein kinase (cGKI) was investigated in wild-type (WT) and cGKI-deficient (cGKI−/−) mice. We demonstrate that murine neutrophils expressed cGKIα. Similar to the regulation of Ca2+ by cGKI in other cells, there was a cGMP-dependent decrease in Ca2+ transients in response to C5a in WT, but not cGKI−/− bone marrow neutrophils. In vitro chemotaxis of bone marrow neutrophils to C5a or IL-8 was significantly greater in cGKI−/− than in WT. Enhanced chemotaxis was also observed with cGKI−/− peritoneal exudate neutrophils (PE-N). In vivo chemotaxis with an arachidonic acid-induced inflammatory ear model revealed an increase in both ear weight and myeloperoxidase (MPO) activity in ear punches of cGKI−/− vs WT mice. These changes were attributable to enhanced vascular permeability and increased neutrophil infiltration. The total extractable content of MPO, but not lysozyme, was significantly greater in cGKI−/− than in WT PE-N. Furthermore, the percentage of MPO released in response to fMLP from cGKI−/− (69%) was greater than that from WT PE-N (36%). PMA failed to induce MPO release from PE-N of either genotype. In contrast, fMLP and PMA released equivalent amounts of lysozyme from PE-N. However, the percentage released was less in cGKI−/− (∼60%) than in WT (∼90%) PE-N. Superoxide release (maximum velocity) revealed no genotype differences in responses to PMA or fMLP stimulation. In summary, these results show that cGKIα down-regulates Ca2+ transients and chemotaxis in murine neutrophils. The regulatory influences of cGKIα on the secretagogue responses are complex, depending on the granule subtype.

A specific plasma kallikrein inhibitor, Bz-Pro-Phe-boroArg (P8720), was used to define the relationship between the kallikrein-kinin (K-K) system and acute intestinal inflammation induced by bacterial peptidoglycan-polysaccharide (PG-APS) in Lewis rats. Group I received human serum albumin (HSA) intramurally in the intestine and was treated with HSA. Group II received PG-APS and was treated with P8720. Group III received PG-APS and was treated with HSA. P8720 attenuated the decrease of high-molecular-weight kininogen and factor XI activity (group II vs group III,P

Aberrant functions of T cells and B cells may impair B cell memory and are thought to allow development of autoantibodies. Rarely, B cell memory defects and/or autoimmunity can be traced to specific mutations; however, in most cases which properties of T cells and B cells cause deficient B cell responses and/or autoimmunity are unclear. To answer these questions we studied child recipients of cardiac transplantation during infancy who, owing to removal of the thymus and T cell depletion at the time of surgery, have a greatly reduced T cell receptor repertoire diversity. We found these children to have impaired memory IgG antibody responses to polypeptide vaccines compared to controls, even though they appeared to mount IgM responses. These results suggest that the defective T cell compartment in recipients of cardiac transplantation in infancy impairs B cell memory and/or isotype class- switching. Remarkably, despite these defects and despite ongoing treatment with immunosuppressive drugs, 6 of the 9 subjects studied produced anti-ssDNA and rheumatoid factor antibodies as early as of 2 years of age. These findings suggest that the diversity of T cells and/or the availability of recent thymic emigrants are needed to generate B cell memory and prevent autoimmunity.

Background: Insulinlike growth factor I (IGF-I) is mitogenic for fibroblasts and smooth muscle cells and stimulates collagen synthesis. The present study tested the hypothesis that IGF-I is important in the development of granulomatous inflammation and fibrosis. Methods: IGF-I messenger RNA (mRNA) was measured in bowel and liver of rats with peptidoglycan-polysaccharide-induced chronic granulomatous enterocolitis and hepatitis using RNase protection. Cellular sites of IGF-I mRNA and IGF-I peptide precursor were localized by in situ hybridization and immunohistochemistry, respectively. Sites of IGF-I synthesis were compared with sites of interleukin 1β mRNA expression. Results: IGF-I mRNA was increased 3.7-fold in cecal tissue from peptidoglycan-polysaccharide-injected rats compared with controls. IGF-I mRNA was up-regulated in fibroblastlike cells in the intensely fibrotic periphery of cecal and hepatic granulomas. This region also expressed IGF-I peptide precursor. Interleukin 1 mRNA localized to macrophage-like cells in the center of granulomas. Conclusions: IGF-I may be important in the development of fibrosis in this model of Crohn's disease. The localization of IGF-I and interleukin 1 mRNAs to distinct but adjacent sites is consistent with a paracrine interaction between cells expressing IGF-I and interleukin 1.