Desarrollo e implementación de un protocolo para evaluación de la citotoxicidad in vitro en células cancerígenas humanas
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Date
2021
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Universidad Nacional de Trujillo
Abstract
Las líneas celulares y su cultivo in vitro, se utilizan ampliamente en la investigación biomédica, y es que son herramientas fundamentales utilizadas en laboratorios de todo el mundo para estudiar la fisiología y la bioquímica de las células, así como mecanismos subyacentes a la enfermedad y, efectos de fármacos y compuestos tóxicos. Para ello, es importante tener conocimiento y comprensión de los principios básicos y técnicas fundamentales que se realizan en un laboratorio de cultivo celular. Así que, el objetivo de este trabajo fue desarrollar un protocolo donde se describe los métodos utilizados para trabajar con una o más líneas celulares, que presentan una gama de fenotipos y patrones en el crecimiento con diferentes condiciones físicas y nutricionales, así mismo, las mejores prácticas para descongelar, cambiar el medio, pasar las células y crioconservar las líneas celulares, minimizando el riesgo de contaminación de microorganismos adventicios o de otras líneas celulares. Además, se describió el ensayo de reducción de bromuro de 3-(4,5-dimetiltiazol-2-ilo)-2,5-difeniltetrazol (MTT), el cual es un método que se utiliza en las pruebas de citotoxicidad in vitro, evaluando el potencial anticancerígeno de algún compuesto, mediante la determinación de la supervivencia y capacidad de proliferación de las células. En este informe se utilizaron como modelos a los compuestos 3-anilino-2-benzoil-1,4-naftoquinona (AC1D) y 3-anilino-2-(furan-2- carbonil)-1,4-naftoquinona (AC1K), sometidos in vitro contra células cancerosas humanas (próstata DU-145, vejiga T-24 y mama MCF-7), y células de riñón embrionario humano no tumoral HEK-293. Se describen los pasos para examinar los efectos citotóxicos de estos compuestos para células cancerosas, su seguridad para células sanas y el adecuado manejo de un cultivo celular, ayudando a los investigadores a familiarizarse con el uso de líneas celulares como herramientas experimentales para complementar e impulsar la investigación, sobre todo en cáncer
ABSTRACT Cell lines and their in vitro culture are widely used in biomedical research, and are fundamental tools used in laboratories around the world to study the physiology and biochemistry of cells, as well as mechanisms underlying disease and, effects of drugs and toxic compounds. To do this, it is important to have knowledge and understanding of the basic principles and fundamental techniques carried out in a cell culture laboratory. So, the objective of this work was to develop a protocol that describes the methods used to work with one or more cell lines, which present a range of phenotypes and growth patterns with different physical and nutritional conditions, as well as the best practices for thawing, changing the medium, passage of cells and cryopreservation of cell lines, minimizing the risk of contamination from adventitious microorganisms or other cell lines. In addition, the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazole bromide (MTT) reduction assay was described, which is a method used in in vitro cytotoxicity tests, evaluating the anticancer potential of a compound, by determining the survival and proliferation capacity of the cells. In this report, the compounds 3-anilino-2-benzoyl-1,4-naphthoquinone (AC1D) and 3-anilino-2-(furan-2-carbonyl)-1,4-naphthoquinone (AC1K) were used as models. subjected in vitro against human cancer cells (DU-145 prostate, T-24 bladder and MCF-7 breast), and HEK-293 non-tumor human embryonic kidney cells. The steps to examine the cytotoxic effects of these compounds for cancer cells, their safety for healthy cells, and the proper handling of a cell culture are described, helping researchers become familiar with the use of cell lines as experimental tools to complement and boost the research, especially in cancer
ABSTRACT Cell lines and their in vitro culture are widely used in biomedical research, and are fundamental tools used in laboratories around the world to study the physiology and biochemistry of cells, as well as mechanisms underlying disease and, effects of drugs and toxic compounds. To do this, it is important to have knowledge and understanding of the basic principles and fundamental techniques carried out in a cell culture laboratory. So, the objective of this work was to develop a protocol that describes the methods used to work with one or more cell lines, which present a range of phenotypes and growth patterns with different physical and nutritional conditions, as well as the best practices for thawing, changing the medium, passage of cells and cryopreservation of cell lines, minimizing the risk of contamination from adventitious microorganisms or other cell lines. In addition, the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazole bromide (MTT) reduction assay was described, which is a method used in in vitro cytotoxicity tests, evaluating the anticancer potential of a compound, by determining the survival and proliferation capacity of the cells. In this report, the compounds 3-anilino-2-benzoyl-1,4-naphthoquinone (AC1D) and 3-anilino-2-(furan-2-carbonyl)-1,4-naphthoquinone (AC1K) were used as models. subjected in vitro against human cancer cells (DU-145 prostate, T-24 bladder and MCF-7 breast), and HEK-293 non-tumor human embryonic kidney cells. The steps to examine the cytotoxic effects of these compounds for cancer cells, their safety for healthy cells, and the proper handling of a cell culture are described, helping researchers become familiar with the use of cell lines as experimental tools to complement and boost the research, especially in cancer
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Keywords
Células cancerígenas, Citotoxicidad in vitro, Laboratorio de cultivo celular