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Universidad Nacional Autónoma de México

31 ARTICLES PUBLISHED IN JoVE

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Immunology and Infection

A Simple and Efficient Method to Detect Nuclear Factor Activation in Human Neutrophils by Flow Cytometry
Erick García-García 1, Eileen Uribe-Querol 2, Carlos Rosales 3
1Department of Biological Sciences, University of Alberta, 2División de Estudios de Posgrado e Investigación, Facultad de Odontología, Universidad Nacional Autónoma de México, 3Department of Immunology, Instituto de Investigaciones Biomédicas, Universidad Nacional Autónoma de México

Neutrophils are the most abundant leukocytes in blood. Neutrophils possess transcriptionally regulated functions such as production of proinflammatory cytokines and inhibition of apoptosis. These functions can be studied with the method presented here, which allows detection and quantification of nuclear factors by flow cytometry in isolated nuclei

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Immunology and Infection

Isolation of Viral Replication Compartment-enriched Sub-nuclear Fractions from Adenovirus-infected Normal Human Cells
Paloma Hidalgo 1,2, Ramón A. Gonzalez 1
1Centro de Investigación en Dinámica Celular, Instituto de Investigación en Ciencias Básicas y Aplicadas, Universidad Autónoma del Estado de Morelos, 2Instituto de Biotecnologìa, Universidad Nacional Autónoma de México

We provide a novel strategy to isolate viral replication compartments (RC) from adenovirus (Ad)-infected human cells. This approach represents a cell-free system that can help to elucidate the molecular mechanisms regulating viral genome replication and expression as well as regulation of viral-host interactions established at the RC.

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Developmental Biology

Affinity Labeling Detection of Endogenous Receptors from Zebrafish Embryos
Tonatiuh Molina-Villa 1, Valentín Mendoza 1, Fernando López-Casillas 1
1Cell and Developmental Biology, Instituto de Fisiología Celular, Universidad Nacional Autónoma de México

A novel technique for the detection of low abundance endogenous receptors present in zebrafish embryos is described. We have named it AFLIP because it consists of affinity labeling of the receptor by its ligand linked to immunoprecipitation.

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Biochemistry

Defining Substrate Specificities for Lipase and Phospholipase Candidates
Diana X. Sahonero-Canavesi 1, Maritza Zavaleta-Pastor 1, Lourdes Martínez-Aguilar 1, Isabel M. López-Lara 1, Otto Geiger 1
1Centro de Ciencias Genómicas, Universidad Nacional Autónoma de México

Many predicted (phospho)lipases are poorly characterized with regard to their substrate specificities and physiological functions. Here we provide a protocol to optimize enzyme activities, search for natural substrates, and propose physiological functions for these enzymes.

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JoVE Journal

Fluorescence Anisotropy as a Tool to Study Protein-protein Interactions
Abril Gijsbers 1, Takuya Nishigaki 2, Nuria Sánchez-Puig 1
1Departamento de Química de Biomacromoléculas, Instituto de Química, Universidad Nacional Autónoma de México, 2Departamento de Genética del Desarrollo y Fisiología Molecular, Instituto de Biotecnología, Universidad Nacional Autónoma de México

Protein interactions are at the heart of a cell's function. Calorimetric and spectroscopic techniques are commonly used to characterize them. Here we describe fluorescence anisotropy as a tool to study the interaction between the protein mutated in the Shwachman-Diamond Syndrome (SBDS) and the Elongation factor-like 1 GTPase (EFL1).

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Behavior

fMRI Mapping of Brain Activity Associated with the Vocal Production of Consonant and Dissonant Intervals
Nadia González-García 1, Pablo L. Rendón 2
1Hospital Infantil de México "Federico Gómez", 2Centro de Ciencias Aplicadas y Desarrollo Tecnológico, Universidad Nacional Autónoma de México

The neural correlates of listening to consonant and dissonant intervals have been widely studied, but the neural mechanisms associated with production of consonant and dissonant intervals are less well known. In this article, behavioral tests and fMRI are combined with interval identification and singing tasks to describe these mechanisms.

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JoVE Journal

CAPRRESI: Chimera Assembly by Plasmid Recovery and Restriction Enzyme Site Insertion
Orlando Santillán 1, Miguel A. Ramírez-Romero 2, Guillermo Dávila 3
1Centro de Ciencias Genómicas, Universidad Nacional Autónoma de México, 2Oncomedic Incorporation, Ciudad de México, 3Laboratorio Internacional de Investigación sobre el Genoma Humano, Universidad Nacional Autónoma de México

Here, we present chimera assembly by plasmid recovery and restriction enzyme site insertion (CAPRRESI), a protocol based on the insertion of restriction enzyme sites into synonym DNA sequences and functional plasmid recovery. This protocol is a fast and low-cost method for fusing protein-coding genes.

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JoVE Journal

Desensitization and Recovery of Crayfish Photoreceptors Upon Delivery of a Light Stimulus
Carolina Barriga-Montoya 1, Araceli de la O-Martínez 1, Arturo Picones 2, Arturo Hernández-Cruz 2, Beatriz Fuentes-Pardo 1, Froylán Gómez-Lagunas 1
1Departamento de Fisiología, Facultad de Medicina, Universidad Nacional Autónoma de México, 2Laboratorio Nacional de Canalopatías, Instituto de Fisiología Celular, Universidad Nacional Autónoma de México

A protocol for the study of desensitization and sensitivity recovery of crayfish photoreceptors as a function of circadian time is presented.

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Cancer Research

Analyzing the Communication Between Monocytes and Primary Breast Cancer Cells in an Extracellular Matrix Extract (ECME)-based Three-dimensional System
Nancy Adriana Espinoza-Sánchez *1,2, Gloria Karina Chimal-Ramírez *1, Ezequiel Moisés Fuentes-Pananá 1
1Unidad de Investigación en Virología y Cáncer, Hospital Infantil de México Federico Gómez, 2Programa de Doctorado en Ciencias Biomédicas, Facultad de Medicina, Universidad Nacional Autónoma de México

Here, we describe a three-dimensional culture method to analyze the morphology of primary breast cancer cells, as well as to study their direct/indirect interactions with monocytes and the outcomes such as collagen degradation, immune cell recruitment, cell invasion, and promotion of cancer-related inflammation.

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Biochemistry

Lipid Index Determination by Liquid Fluorescence Recovery in the Fungal Pathogen Ustilago Maydis
Lucero Romero-Aguilar *1,2, Mónica Montero-Lomeli *3, Juan Pablo Pardo 2, Guadalupe Guerra- Sánchez 1
1Escuela Nacional de Ciencias Biológicas, Instituto Politécnico Nacional, 2Facultad de Medicina, Universidad Nacional Autónoma de México, 3Instituto de Bioquímica Médica Leopoldo de Meis, Universidade Federal do Rio de Janeiro

Here, we describe a protocol to obtain the lipid droplet index (LD index) to study the dynamics of triacylglycerols in cells cultured in high-throughput experiments. The LD index assay is an easy and reliable method that uses BODIPY 493/503. This assay does not need dispendious lipid extraction or microscopy analysis.

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Genetics

Promoter Capture Hi-C: High-resolution, Genome-wide Profiling of Promoter Interactions
Stefan Schoenfelder *1, Biola-Maria Javierre *1,2, Mayra Furlan-Magaril 1,3, Steven W. Wingett 1,4, Peter Fraser 1,5
1Nuclear Dynamics Programme, The Babraham Institute, Babraham Research Campus, 2IJC Building, Campus ICO-Germans Trias i Pujol, Josep Carreras Leukemia Research Institute, 3Departamento de Genética Molecular, Instituto de Fisiología Celular, Universidad Nacional Autónoma de México, 4Bioinformatics Group, The Babraham Institute, Babraham Research Campus, 5Department of Biological Science, Florida State University

DNA regulatory elements, such as enhancers, control gene expression by physically contacting target gene promoters, often through long-range chromosomal interactions spanning large genomic distances. Promoter Capture Hi-C (PCHi-C) identifies significant interactions between promoters and distal regions, enabling the assignment of potential regulatory sequences to their target genes.

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Behavior

A Two-interval Forced-choice Task for Multisensory Comparisons
Fabiola Duarte 1, Tonatiuh Figueroa 1, Luis Lemus 1
1Neurociencia Cognitiva, Instituto de Fisiología Celular, Universidad Nacional Autónoma de México

Psychophysics is essential for studying perception phenomena through sensory information. Here we present a protocol to perform a two-interval forced-choice task as implemented in a previous report on human psychophysics where participants estimated the duration of visual, auditory, or audiovisual intervals of aperiodic trains of pulses.

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Chemistry

A Facile Synthetic Method to Obtain Bismuth Oxyiodide Microspheres Highly Functional for the Photocatalytic Processes of Water Depuration
Juan C. Durán-Álvarez 1, Carolina Martínez 1, Adriana C. Mera 2, Raquel Del Angel 1, Nini J. Gutiérrez-Moreno 1, Rodolfo Zanella 1
1Instituto de Ciencias Aplicadas y Tecnología, Universidad Nacional Autónoma de México, 2Instituto de Investigación Multidisciplinario en Ciencia y Tecnología, Universidad de La Serena

This article describes a synthetic method to obtain bismuth oxyiodide microspheres, which are highly functional to perform the photocatalytic removal of organic pollutants, such as ciprofloxacin, in water under UV-A/visible light irradiation.

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Developmental Biology

In Vitro Culture of Epithelial Cells from Different Anatomical Regions of the Human Amniotic Membrane
Daniela Avila-González 1, Guadalupe García-López 1, Néstor E. Díaz-Martínez 2, Héctor Flores-Herrera 3, Anayansi Molina-Hernández 1, Wendy Portillo 4, Néstor F. Díaz 1
1Departamento de Fisiología y Desarrollo Celular, Instituto Nacional de Perinatología, 2Biotecnología Médica y Farmacéutica CONACYT, Centro de Investigación y Asistencia en Tecnología y Diseño del Estado de Jalisco (CIATEJ), 3Departamento de Inmunobioquímica, Instituto Nacional de Perinatología, 4Instituto de Neurobiología, UNAM

This protocol describes the isolation of epithelial cells from different anatomical regions of the human amniotic membrane to determine their heterogeneity and functional properties for possible application in clinical and physiopathological models.

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Neuroscience

Culture of Neurospheres Derived from the Neurogenic Niches in Adult Prairie Voles
Daniela Ávila-González 1, Larry J. Young 2, Francisco Camacho 1, Raúl G. Paredes 1,3, Néstor F. Díaz 4, Wendy Portillo 1
1Departamento de Neurobiología Conductual y Cognitiva, Instituto de Neurobiología, Universidad Nacional Autónoma de México, 2Silvio O. Conte Center for Oxytocin and Social Cognition, Center for Translational Social Neuroscience, Department of Psychiatry and Behavioral Sciences, Yerkes National Primate Research Center, Emory University, 3Escuela Nacional de Estudios Superiores Juriquilla, Universidad Nacional Autónoma de México, 4Departamento de Fisiología y Desarrollo Celular, Instituto Nacional de Perinatología

We established the conditions to culture neural progenitor cells from the subventricular zone and dentate gyrus of the adult brain of prairie voles, as a complementary in vitro study, to analyze the sex-dependent differences between neurogenic niches that could be part of functional plastic changes associated with social behaviors.

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Neuroscience

Unraveling the Role of Discrete Areas of the Rat Brain in the Regulation of Ovulation through Reversible Inactivation by Tetrodotoxin Microinjections
Carlos-Camilo Silva *1,2, Montserrat Bolaños-Hurtado *1, Cinthia Juárez-Tapia 4, Angélica Flores 1,2, Isabel Arrieta-Cruz 3,5, María-Esther Cruz 3, Roberto Domínguez 1,2,3
1Chronobiology of Reproduction Research Lab, Biology of Reproduction Research Unit, Facultad de Estudios Superiores Zaragoza, Universidad Nacional Autónoma de México, 2Developmental Biology Lab, Biology of Reproduction Research Unit, Facultad de Estudios Superiores Zaragoza, Universidad Nacional Autónoma de México, 3Neuroendocrinology Lab, Biology of Reproduction Research Unit, Facultad de Estudios Superiores Zaragoza, Universidad Nacional Autónoma de México, 4Facultad de Ciencias, Universidad Nacional Autónoma de México, 5Department of Basic Research, Instituto Nacional de Geriatria, Ministry of Health

This protocol describes the construction of a low-cost microinjection system, its stereotaxic implantation into deep-brain structures and the procedure for timed microinjections of tetrodotoxin in awake and unrestrained rats. The goal is to reveal the participation of hypothalamic structures in the regulation of ovulation by inhibiting their neural activity.

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Environment

Using Tree-Rings to Reconstruct Fire History Information from Forested Areas
Julián Cerano-Paredes 1, Jose M. Iniguez 2, José Villanueva-Díaz 1, Rosalinda Cervantes-Martínez 1, Víctor H. Cambrón-Sandoval 3, Josué R. Estrada-Arellano 4, Gerardo Esquivel-Arriaga 1, Osvaldo Franco-Ramos 5, Lorenzo Vázquez-Selem 5, Gabriel F. Cardoza-Martínez 4
1CENID-RASPA, INIFAP, 2Rocky Mountain Research Station, USDA Forest Service, 3Facultad de Ciencias Naturales, Universidad Autónoma de Querétaro, 4Facultad de Ciencias Biológicas, Universidad Juárez del Estado de Durango, 5Instituto de Geografía, Universidad Nacional Autónoma de México

This work explains the most appropriate techniques and methods for conducting a fire history study from beginning site selection to final analysis of fire-climate relationship.

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Genetics

In-Nucleus Hi-C in Drosophila Cells
Ayerim Esquivel-López 1, Rodrigo Arzate-Mejía 1, Rosario Pérez-Molina 1, Mayra Furlan-Magaril 1
1Departamento de Genética Molecular, Instituto de Fisiología Celular, Universidad Nacional Autónoma de México

The genome is organized in the nuclear space into different structures that can be revealed through chromosome conformation capture technologies. The in-nucleus Hi-C method provides a genome-wide collection of chromatin interactions in Drosophila cell lines, which generates contact maps that can be explored at megabase resolution at restriction fragment level.

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Biology

Immunofluorescent Staining for Visualization of Heterochromatin Associated Proteins in Drosophila Salivary Glands
Silvia Meyer-Nava 1, Mario Zurita 1, Viviana Valadez-Graham 1
1Instituto de Biotecnología, Departamento de Genética del Desarrollo y Fisiología Molecular, Universidad Nacional Autónoma de México

This protocol aims to visualize heterochromatin aggregates in Drosophila polytene cells.

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Medicine

Isolated Lung Perfusion System in the Rabbit Model
Alejandro Pacheco-Baltazar 2, José Luis Arreola-Ramírez 1, Jesús Alquicira-Mireles 1, Patricia Segura-Medina 1
1Departamento de Investigación en Hiperreactividad Bronquial, Instituto Nacional de Enfermedades Respiratorias, 2Facultad de Medicina, Universidad Nacional Autónoma de México

The isolated rabbit lung preparation is a gold standard tool in pulmonary research. This publication aims to describe the technique as developed for the study of physiological and pathological mechanisms involved in airway reactivity, lung preservation, and preclinical research in lung transplantation and pulmonary edema.

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Developmental Biology

Chicken Recombinant Limbs Assay to Understand Morphogenesis, Patterning, and Early Steps in Cell Differentiation
Jessica Cristina Marín-Llera 1, Montse Fernández-Calderón 2, Jesús Chimal-Monroy 1
1Departamento de Medicina Genómica y Toxicología Ambiental, Instituto de Investigaciones Biomédicas, Universidad Nacional Autónoma de México, 2Departamento de Anatomía y Biología Celular and IDIVAL, Facultad de Medicina, Universidad de Cantabria

Recombinant limbs are a powerful experimental model that allows for studying the process of cell differentiation and the generation of patterns under the influence of embryonic signals. This protocol presents a detailed method for generating recombinant limbs with chicken limb-mesodermal cells, adaptable to other cell types obtained from different organisms.

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Developmental Biology

Analysis of Cell Differentiation, Morphogenesis, and Patterning During Chicken Embryogenesis Using the Soaked-Bead Assay
Jessica Cristina Marín-Llera 1, Jesús Chimal-Monroy 1
1Departamento de Medicina Genómica y Toxicología Ambiental, Instituto de Investigaciones Biomédicas, Universidad Nacional Autónoma de México

The soaked bead assay involves targeted delivery of test reagent at any developmental time point to study the regulation of cell differentiation and morphogenesis. A detailed protocol, applicable to any experimental animal model, for preparing three different types of soaked beads and implanting these in the interdigit of a chicken embryo is presented.

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Environment

How to Extract Climate Variability from Tree-Rings
Julián Cerano-Paredes 1, Paul Szejner 2, Genaro Gutiérrez-García 2, Rosalinda Cervantes-Martínez 1, Víctor H. Cambrón-Sandoval 3, José Villanueva-Díaz 1, Josué R. Estrada-Arellano 4, Osvaldo Franco-Ramos 5, Lorenzo Vázquez-Selem 5, Luis U. Castruita-Esparza 6
1INIFAP, CENID-RASPA, 2Instituto de Geología, Universidad Nacional Autónoma de México, 3Facultad de Ciencias Naturales, Universidad Autónoma de Querétaro, 4Facultad de Ciencias Biológicas, Universidad Juárez del Estado de Durango, 5Instituto de Geografía, Universidad Nacional Autónoma de México, 6Facultad de Ciencias Agrícolas y Forestales, Universidad Autónoma de Chihuahua

Tree-ring climate reconstructions can be helpful to better understand past climate variability beyond instrumental records. This protocol shows how to reconstruct past climate using tree rings and meteorological instrumental records.

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Biochemistry

Identification of Hemolytic and Phospholipase Activity in Crude Extracts from Sea Anemones by Straightforward Bioassays
Santos Ramírez-Carreto 1, Sandra I. Salazar-García 1, Giovanni Macías Martínez 1, Claudia Rodríguez-Almazán 1,2
1Instituto de Biotecnología, Universidad Nacional Autónoma de México, 2Departamento de Micro y Nanotecnologías, Instituto de Ciencias Aplicadas y Tecnología, Universidad Nacional Autónoma de México

Here, we describe a protocol to obtain crude venom extract from sea anemone and detect its hemolytic and phospholipase activity.

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Immunology and Infection

A Strategy for the Study of IL-9-Producing Lymphoid Cells in the Nippostrongylus brasiliensis Infection Model
Ofelia Muñoz-Paleta *1, Enrique Olguín-Martínez *1, Blanca Estela Ruiz-Medina *1, Alejandra Alonso-Quintana 1, Ma Cleofas Marcial-Medina 1, Paula Licona-Limón 1
1Departamento de Biología Celular y del Desarrollo, Instituto de Fisiología Celular, Universidad Nacional Autónoma de México

IL-9-expressing T and ILC2 cells are induced during N. brasiliensis infection, yet their characterization has been largely overlooked in the infected intestine due to their low frequency and differential kinetics. This protocol describes the isolation of these cells from different target organs and confirmation of their identity via flow cytometry at different infection stages.

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Cancer Research

Antisense Oligonucleotides as a Tool for Prolonged Knockdown of Nuclear lncRNAs in Human Cell Lines
Rogelio Montiel-Manriquez 1, Clementina Castro-Hernández 1, Cristian Arriaga-Canon 1, Luis A. Herrera 1,2
1Unidad de Investigación Biomédica en Cáncer, Instituto Nacional de Cancerología-Instituto de Investigaciones Biomédicas, Universidad Nacional Autónoma de México, 2Escuela de Medicina y Ciencias de la Salud, Tecnológico de Monterrey

To analyze the function of lncRNAs in time-dependent processes such as chromosomal instability, a prolonged knockdown effect must be achieved. To that purpose, presented here is a protocol that uses modified antisense oligonucleotides to achieve effective knockdown in cell lines for 21 days.

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Biochemistry

Detection and Quantification of Mono-Rhamnolipids and Di-Rhamnolipids Produced by Pseudomonas aeruginosa
Abigail González-Valdez 1, Jessica Hernández-Pineda 2, Gloria Soberón-Chávez 1
1Departamento de Biología Molecular y Biotecnología, Instituto de Investigaciones Biomédicas, Universidad Nacional Autónoma de México, 2Departamento de Infectología e Inmunología, Instituto Nacional de Perinatología

Pseudomonas aeruginosa produces the rhamnolipid biosurfactants. Thin-layer chromatography detects and determines the proportion of mono- and di-rhamnolipids produced by each strain. Quantification of total rhamnolipids involves assessing rhamnose equivalents present in these biosurfactants extracted from the culture supernatants using the orcinol method.

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Environment

Biogas Purification through the use of a Microalgae-Bacterial System in Semi-Industrial High Rate Algal Ponds
Mariana Vega Blanes 1, Isaac Jhonnatan Pérez-Hermosillo 2, Arnold Ramírez Rueda 1, Armando González Sánchez 1
1Instituto de Ingeniería, Ciudad Universitaria, Universidad Nacional Autónoma de México, 2Granos y Servicios Integrales

Air pollution impacts the quality of life of all organisms. Here, we describe the use of microalgae biotechnology for the treatment of biogas (simultaneous removal of carbon dioxide and hydrogen sulfide) and the production of biomethane through semi-industrial open high-rate algal ponds and subsequent analysis of treatment efficiency, pH, dissolved oxygen, and microalgae growth.

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Biochemistry

Estimation of Structural Sensitivity of Intrinsically Disordered Regions in Response to Hyperosmotic Stress in Living Cells Using FRET
Constanza Enriquez-Toledo *1, Cesar A. Ponce-Diego *1, Cesar L. Cuevas-Velazquez 1
1Departamento de Bioquímica, Facultad de Química, Universidad Nacional Autónoma de México

Intrinsically disordered regions (IDRs) are flexible protein domains that modify their conformation in response to environmental changes. Ensemble fluorescence resonance energy transfer (FRET) can estimate protein dimensions under different conditions. We present a FRET approach to assess IDR structural sensitivity in living Saccharomyces cerevisiae cells under hyperosmotic stress.

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Chemistry

Visualizing and Analyzing the Chemical Space of Natural Product Databases for Drug Discovery
Haruna Luz Barazorda-Ccahuana 1, K. Eurídice Juárez-Mercado 2, José L. Medina-Franco 2, Miguel Angel Chavez-Fumagalli 1
1Computational Biology and Chemistry Research Group, Vicerrectorado de Investigación, Universidad Católica de Santa María, 2DIFACQUIM Research Group, Department of Pharmacy, School of Chemistry, Universidad Nacional Autónoma de México

Here, we provide a methodology that uses different molecular representations to display and analyze the chemical space of natural compound data sets, with a focus on applications related to drug discovery.

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Genetics

Mitochondrial Transformation in Baker's Yeast to Study Translation and Respiratory Complex Assembly
Yolanda Camacho-Villasana *1, Ulrik Pedroza-Dávila *1, Xochitl Perez-Martinez 1
1Departamento de Genética Molecular, Instituto de Fisiología Celular, Universidad Nacional Autónoma de México

This work explains how to transform yeast mitochondria using a biolistic method. We also show how to select and purify the transformants and how to introduce the desired mutation in the target position within the mitochondrial genome.

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