Design, assembly and test of a bioreactor for the electrical stimulation of neuronal cells

• Autor(a) principal: Melo, Diogo Soares
• Data de Publicação: 2022
• Tipo de documento: Dissertação
• Idioma: eng
• Título da fonte: Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
• Texto Completo: http://hdl.handle.net/10362/147387
• topic: Tissue engineering; Neuronal regeneration; Electrical stimulation; Bioreactor; SH-SY5Y cells; Electrospinnig; Domínio/Área Científica::Engenharia e Tecnologia::Outras Engenharias e Tecnologias

Millions of people have suffered injury to their nervous system, which by its limited self- healing capacity, represents life-long complications, associated with loss of motor and sensory function. Though limited, this capacity is being extensively explored, and has been shown to increase through the use of electrical stimulation (ES). Therefore, this work was oriented towards the development of a setup for ES of neuronal cells, allowing the assessment of its potential in promoting neuronal regeneration. An ES chamber was designed using the CAD software Fusion 360TM and produced by machining of a poly(methyl methacrylate) (PMMA) block. A fixation system for con- ductive scaffolds was included, using stainless steel electrodes, which fits the description for a direct coupling method found in the literature. Connection to a power supply or a function generator is possible, allowing for delivery of both direct current (DC) and alternating current (AC) to cells. In a different design, electrical insulation of the media was attempted, defectively. The nature of this work supported the need for incorporating conductive polymers (CPs) in the scaffolds used for neuronal differentiation of cells in the stimulation chamber and so, poly(lactic acid) (PLA) aligned electruspun fibers were produced and coated with poly(3,4-ethylenedioxythiophene) (PEDOT) using vapor-phase polymerization (VPP). In this process, the polymerization takes place through the reaction of Iron(III) p-toluenesul– fonate/Fe(III)Tosylate (FeTos) included in the scaffolds with 3,4-ethylenedioxythiophene (EDOT) on vapor phase. This fibers did not exhibit cytotoxicity and electrical character- ization was attempted, using the bioreactor as a 2-point probe. Film casting using the same polymeric solutions failed, as an increase in the ratio of PLA to FeTos did not reduce film brittleness. In vitro assays were conducted both with and without stimulation. SH-SY5Y extended neurites after only 2 days of exposure to retinoic acid (RA). Cells maintained some level of differentiation and neurite directionality with time, when cultured in the produced fibers. Importantly, an electrical field of 500 mV/ cm was applied 1 h/day, for 9 days, without significant improvements on neuronal differentiation.