Optimal parameters for the enhancement of human osteoblast-like cell proliferation in vitro via shear stress induced by high-frequency mechanical vibration
Parámetros óptimos para la mejora de la proliferación de células similares a osteoblastos humanos in vitro a través del esfuerzo cortante inducido por vibraciones mecánicas de alta frecuencia
Nahum Rosenberg, Orit Rosenberg, Jacob Halevi Politch, Haim Abramovich
Abstract
Introduction: Biomechanical stimulation of cultured human osteoblast-like cells, which is based on controlled mechanical vibration, has been previously indicated, but the exact mechanical parameters that are effective for cells' proliferation enhancement are still elusive due to the lack of direct data recordings from the stimulated cells in culture. Therefore, we developed a low friction tunable system that enables recording of a narrow range of mechanical parameters, above the infrasonic spectrum, that applied uniformly to human osteoblast-like cells in monolayer culture, aiming to identify a range of mechanical parameters that are effective to enhance osteoblast proliferation in vitro.
Methods: Human osteoblast-like cells in explant monolayer culture samples were exposed to mechanical vibration in the 10-70Hz range of frequencies for two minutes, in four 24 hours intervals. Cell numbers in culture, cellular alkaline phosphatase activity (a marker of cell maturation), and lactate dehydrogenase activity in culture media (representing cell death) were measured after the mechanical stimulation protocol application and compared statistically to the control cell cultures kept in static conditions. The cell proliferation was deduced from cell number in culture and cell death measurements.
Results: We found that 50-70 Hz of vibration frequency protocol (10-30 μm of maximal displacement amplitude, 0.03g of peak-to-peak acceleration) is optimal for enhancing cells' proliferation(p<0.05), with a parallel decrease of their maturation (p<0.01).
Discussion: We detected the optimal mechanical parameters of excitation protocol for induction of osteoblast proliferation in vitro by a mechanical platform, which can be used as a standardized method in the research of mechanotransduction in human osteoblast.
Keywords
Resumen
Introducción: La estimulación biomecánica de células similares a osteoblastos humanos cultivadas, que se basa en vibraciones mecánicas controladas se ha demostrado anteriormente, pero los parámetros mecánicos exactos que son efectivos para la mejora de la proliferación de células aún son difíciles de alcanzar debido a la falta de registros de datos directos de las células estimuladas en cultivo. Por lo tanto, desarrollamos un sistema sintonizable de baja fricción que permite el registro de un rango estrecho de parámetros mecánicos, por encima del espectro infrasónico, que se aplica de manera uniforme a células similares a osteoblastos humanos en cultivo monocapa, con el objetivo de identificar un rango de parámetros mecánicos que son efectivos para mejorar la proliferación de osteoblastos in vitro.
Métodos: Se expusieron células similares a osteoblastos humanos en muestras de cultivo de monocapa de explante a vibración mecánica en el rango de frecuencias de 10-70 Hz durante dos minutos, en cuatro intervalos de 24 horas. El número de células en cultivo, la actividad de la fosfatasa alcalina celular (un marcador de maduración celular) y la actividad de la lactato deshidrogenasa en los medios de cultivo (que representa la muerte celular) se midieron después de la aplicación del protocolo de estimulación mecánica y se compararon estadísticamente con los cultivos de células de control mantenidos en condiciones estáticas. La proliferación celular se dedujo del número de células en cultivo y mediciones de muerte celular.
Resultados: Encontramos que 50-70 Hz de protocolo de frecuencia de vibración (10-30 μm de amplitud de desplazamiento máxima, 0,03 g de aceleración de pico a pico) es óptimo para mejorar la proliferación de células (p <0,05), con una disminución paralela de su maduración (p <0.01).
Discusión: Detectamos los parámetros mecánicos óptimos del protocolo de excitación para la inducción de la proliferación de osteoblastos in vitro mediante una plataforma mecánica, que puede utilizarse como método estandarizado en la investigación de la transducción mecánica en osteoblastos humanos.
Palabras clave
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Submitted date:
03/23/2021
Reviewed date:
04/21/2021
Accepted date:
05/05/2021
Publication date:
05/10/2021
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