Re-activation of Human Endogenous Retrovirus-K in Neuroinflammatory Disease

A thesis submitted to the Faculty of Graduate Studies in partial fulfillment of the requirements for the Master of Science degree. Department of Biology, Master of Science in Bioscience, Technology, and Public Policy program, The University of Winnipeg.A new appreciation of the microbiome is changing the way we perceive human health and disease. The holobiontic nature of humans is even etched into our DNA in the form of viral symbionts. Empirical evidence for the presence of endogenous retroviruses (ERVs) in the human genome and their activity in homeostatic and pathological states has accumulated; however, no causal relationship with human disease has been established to date. In this review, we will focus on the role of endogenous retrovirus-K in neurological disease. Specifically, we will attempt to reconcile the pathological contribution of ERVK in disparate neurological diseases by providing evidence as to inter-individual differences in ERVK genotypes, addressing the molecular regulation of ERVK, and providing detailed examples of ERVK-mediated processes in nervous system diseases.ALS Association, Canada Foundation for Innovation, Manitoba Medical Service Foundation, Manitoba Health Research Council, University of Winnipeg, CIHR Frederick Banting and Charles Best Canada Graduate Scholarship, and Manitoba Graduate ScholarshipMaster of Science in Bioscience, Technology, and Public Polic

Human cytomegalovirus vaccination: progress and perspectives of recombinant gB

A vaccine for Human cytomegalovirus (HCMV) remains a high priority as complications following infection are observed in immunocompromised individuals and in congenitally infected neonates. Numerous preclinical and clinical studies have investigated vaccine strategies ranging from live attenuated preparations, nucleic acid-based approaches and recombinant delivery systems to subunit vaccines. These have defined the importance of both cell-mediated and humoral immunity to viral gB in the control of HCMV infection. This review will cover clinical trials investigating vaccine approaches that have incorporated gB and discuss the future perspectives of the recombinant gB subunit vaccine for HCMV

Development of a Fluorescent Assay to Search New Drugs Using Stable tdTomato-Leishmania, and the Selection of Galangin as a Candidate With Anti-Leishmanial Activity

Antimonials continue to be considered the first-line treatment for leishmaniases, but its use entails a wide range of side effects and serious reactions. The search of new drugs requires the development of methods more sensitive and faster than the conventional ones. We developed and validated a fluorescence assay based in the expression of tdTomato protein by Leishmania, and we applied this method to evaluate the activity in vitro of flavonoids and reference drugs. The pIR1SAT/tdTomato was constructed and integrated into the genome of Leishmania (Leishmania) amazonensis. Parasites were selected with nourseothricin (NTC). The relation of L. amaz/tc3 fluorescence and the number of parasites was determined; then the growth in vitro and infectivity in BALB/c mice was characterized. To validate the fluorescence assay, the efficacy of miltefosine and meglumine antimoniate was compared with the conventional methods. After that, the method was used to assess in vitro the activity of flavonoids; and the mechanism of action of the most active compound was evaluated by transmission electron microscopy and ELISA. A linear correlation was observed between the emission of fluorescence of L. amaz/tc3 and the number of parasites (r2 = 0.98), and the fluorescence was stable in the absence of NTC. No differences were observed in terms of infectivity between L. amaz/tc3 and wild strain. The efficacy of miltefosine and meglumine antimoniate determined by the fluorescence assay and the microscopic test showed no differences, however, in vivo the fluorescence assay was more sensitive than limiting dilution assay. Screening assay revealed that the flavonoid galangin (GAL) was the most active compound with IC50 values of 53.09 µM and 20.59 µM in promastigotes and intracellular amastigotes, respectively. Furthermore, GAL induced mitochondrial swelling, lipid inclusion bodies and vacuolization in promastigotes; and up-modulated the production of IL-12 p70 in infected macrophages. The fluorescence assay is a useful tool to assess the anti-leishmanial activity of new compounds. However, the assay has some limitations in the macrophage-amastigote model that might be related with an interfere of flavanol aglycones with the fluorescence readout of tdTomato. Finally, GAL is a promising candidate for the development of new treatment against the leishmaniasisFil: Garcia Bustos, Maria Fernanda. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; ArgentinaFil: Moya Alvarez, Agustin. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; ArgentinaFil: Pérez Brandan, Cecilia María. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; ArgentinaFil: Parodi Ramoneda, Cecilia María. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; ArgentinaFil: Sosa, Andrea Mabel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; ArgentinaFil: Buttazzoni Zuñiga, Valeria Carolina. Facultad de Ciencias Agrarias y Veterinarias ; Universidad Catolica de Salta;Fil: Pastrana, Oscar Marcelo. Facultad de Ciencias Agrarias y Veterinarias ; Universidad Catolica de Salta;Fil: Manghera, Paula Mariana. Facultad de Ciencias Agrarias y Veterinarias ; Universidad Catolica de Salta;Fil: Peñalva, Pablo Alejandro. Facultad de Ciencias Agrarias y Veterinarias ; Universidad Catolica de Salta;Fil: Marco, Jorge Diego. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; ArgentinaFil: Barroso, Paola Andrea. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentin

Inception of early-life allergen–induced airway hyperresponsiveness is reliant on IL-13+CD4+ T cells

Airway hyperresponsiveness (AHR) is a critical feature of wheezing and asthma in children, but the initiating immune mechanisms remain unconfirmed. We demonstrate that both recombinant interleukin-33 (rIL-33) and allergen [house dust mite (HDM) or Alternaria alternata] exposure from day 3 of life resulted in significantly increased pulmonary IL-13+CD4+ T cells, which were indispensable for the development of AHR. In contrast, adult mice had a predominance of pulmonary LinnegCD45+CD90+IL-13+ type 2 innate lymphoid cells (ILC2s) after administration of rIL-33. HDM exposure of neonatal IL-33 knockout (KO) mice still resulted in AHR. However, neonatal CD4creIL-13 KO mice (lacking IL-13+CD4+ T cells) exposed to allergen from day 3 of life were protected from AHR despite persistent pulmonary eosinophilia, elevated IL-33 levels, and IL-13+ ILCs. Moreover, neonatal mice were protected from AHR when inhaled Acinetobacter lwoffii (an environmental bacterial isolate found in cattle farms, which is known to protect from childhood asthma) was administered concurrent with HDM. A. lwoffii blocked the expansion of pulmonary IL-13+CD4+ T cells, whereas IL-13+ ILCs and IL-33 remained elevated. Administration of A. lwoffii mirrored the findings from the CD4creIL-13 KO mice, providing a translational approach for disease protection in early life. These data demonstrate that IL-13+CD4+ T cells, rather than IL-13+ ILCs or IL-33, are critical for inception of allergic AHR in early life

Transcriptional profiling of HERV-K(HML-2) in amyotrophic lateral sclerosis and potential implications for expression of HML-2 proteins

Abstract Background Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disorder. About 90% of ALS cases are without a known genetic cause. The human endogenous retrovirus multi-copy HERV-K(HML-2) group was recently reported to potentially contribute to neurodegeneration and disease pathogenesis in ALS because of transcriptional upregulation and toxic effects of HML-2 Envelope (Env) protein. Env and other proteins are encoded by some transcriptionally active HML-2 loci. However, more detailed information is required regarding which HML-2 loci are transcribed in ALS, which of their proteins are expressed, and differences between the disease and non-disease states. Methods For brain and spinal cord tissue samples from ALS patients and controls, we identified transcribed HML-2 loci by generating and mapping HML-2-specific cDNA sequences. We predicted expression of HML-2 env gene-derived proteins based on the observed cDNA sequences. Furthermore, we determined overall HML-2 transcript levels by RT-qPCR and investigated presence of HML-2 Env protein in ALS and control tissue samples by Western blotting. Results We identified 24 different transcribed HML-2 loci. Some of those loci are transcribed at relatively high levels. However, significant differences in HML-2 loci transcriptional activities were not seen when comparing ALS and controls. Likewise, overall HML-2 transcript levels, as determined by RT-qPCR, were not significantly different between ALS and controls. Indeed, we were unable to detect full-length HML-2 Env protein in ALS and control tissue samples despite reasonable sensitivity. Rather our analyses suggest that a number of HML-2 protein variants other than full-length Env may potentially be expressed in ALS patients. Conclusions Our results expand and refine recent publications on HERV-K(HML-2) and ALS. Some of our results are in conflict with recent findings and call for further specific analyses. Our profiling of HML-2 transcription in ALS opens up the possibility that HML-2 proteins other than canonical full-length Env may have to be considered when studying the role of HML-2 in ALS disease

Electrocardiogram pearl: ST-T changes in patient with chest pain – Ischemia or infarction?

Most common electrocardiogram (ECG) findings of myocardial ischemia are ST segment deviations & T wave (ST-T) alterations. However, multiple other conditions can cause ST-T changes mimicking ischemia including ventricular hypertrophy, bundle branch block, electrolyte imbalance, drugs, channelopathies, etc. Uncommonly, incorrect placement of limb leads can also produce ST-T changes leading to diagnostic dilemma. We report a case of erroneous limb-lead placement in a 45 years male mimicking ischemic ECG changes

El uso de la prescripción en el proceso de enseñanza y aprendizaje de Farmacología veterinaria

p. 99-101En el presente trabajo mencionamos nuestra experiencia con el uso de recetas, vademécum y formas farmacéuticas en el desarrollo de las clases prácticas del cursado de Farmacología. El objetivo de esta presentación es promover el uso de estas herramientas para las clases prácticas de Farmacología y como soporte en la interacción grupal de los alumnos, fomentando al mismo tiempo la autonomía con proyección al auto aprendizaje. Se describen algunas tareas prácticas desarrolladas durante el cursado (resolución de casos, cálculo de dosis, elaboración de recetas, uso de vademécum), para mejorar la futura prescripción de medicamentos Obtuvimos resultados favorables, por lo que consideramos que el docente debe acompañar a sus alumnos, en la enseñanza de la prescripción de medicamentos, con técnicas pedagógicas motivadoras, para conseguir transferir conocimientos en la clase práctica, y promover la formación de profesionales racionales.Fil: Martinis Mercado, Daniela. Universidad Católica de Salta. Facultad de Ciencias Agrarias y Veterinarias; Argentina.Fil: Fernández, José Rodrigo. Universidad Católica de Salta; Argentina.Fil: Manghera, Paula Mariana. Universidad Católica de Salta; Argentina