A Versatile Bioreactor for Dynamic Suspension Cell Culture. Application to the Culture of Cancer Cell Spheroids.

Diana Massai,Giulia Cerino,Giuseppe Isu,Diego Gallo,Giuseppe Falvo D’Urso Labate,Umberto Morbiducci,Caterina Frati,Federico Quaini,Marco A Deriu,Alberto Audenino,Denise Madeddu,Angela Falco,Maurizio Pesce

doi:10.1371/journal.pone.0154610

Diana Massai, Giulia Cerino + Show 11 more

Open Access

https://doi.org/10.1371/journal.pone.0154610

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Abstract

A versatile bioreactor suitable for dynamic suspension cell culture under tunable shear stress conditions has been developed and preliminarily tested culturing cancer cell spheroids. By adopting simple technological solutions and avoiding rotating components, the bioreactor exploits the laminar hydrodynamics establishing within the culture chamber enabling dynamic cell suspension in an environment favourable to mass transport, under a wide range of tunable shear stress conditions. The design phase of the device has been supported by multiphysics modelling and has provided a comprehensive analysis of the operating principles of the bioreactor. Moreover, an explanatory example is herein presented with multiphysics simulations used to set the proper bioreactor operating conditions for preliminary in vitro biological tests on a human lung carcinoma cell line. The biological results demonstrate that the ultralow shear dynamic suspension provided by the device is beneficial for culturing cancer cell spheroids. In comparison to the static suspension control, dynamic cell suspension preserves morphological features, promotes intercellular connection, increases spheroid size (2.4-fold increase) and number of cycling cells (1.58-fold increase), and reduces double strand DNA damage (1.5-fold reduction). It is envisioned that the versatility of this bioreactor could allow investigation and expansion of different cell types in the future.

Highlights

The large scale production of cells is a mandatory step to set up economically viable in vitro experimental models for basic research, disease modelling and drug testing, and to definitely translate tissue engineering and regenerative medicine strategies to the clinical practice for therapeutic applications
We present here a versatile bioreactor suitable for tunable shear stress dynamic suspension cell culture
Cells are suspended at an average volume fraction (VF) value of approximately 0.33%, which is close to the initial VF value (0.48%), with the peak of probability density function (PDF) value equal to 2.5, corresponding to VF values between 0 and 0.5% (Fig 4A)

Summary

Introduction

The large scale production of cells is a mandatory step to set up economically viable in vitro experimental models for basic research, disease modelling and drug testing, and to definitely translate tissue engineering and regenerative medicine strategies to the clinical practice for therapeutic applications. In a scaling-up perspective and inspired by the manufacturing processes of therapeutics in biopharmaceutical industry [6,7], three-dimensional (3D) suspension culture has demonstrated to be an advantageous alternative to monolayer techniques for large-scale expansion of cells [4,5,8,9]. Suspension methods have been widely adopted: (1) for scalable and controlled expansion of stem cells [10,11,12,13,14,15] and cancer cells [16,17,18]; (2) for guiding stem cell differentiation [13,19,20,21,22]; (3) for the production of cellular spheroids and tissue-like constructs [23,24,25]. When suspension is obtained by dynamic mixing of the culture medium, (1) the formation of gradients in, e.g., temperature, pH, dissolved oxygen, nutrients/metabolites is prevented, (2) the transport of oxygen and nutrients is increased, and (3) the sedimentation of cultured cells/constructs is avoided, going beyond the intrinsic limitations of static culture systems [4,7,9,28]

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