Real-time cellular impedance monitoring and imaging of biological barriers in a dual-flow membrane bioreactor

Abstract

The generation of physiologically relevant in-vitro models of biological barriers can play a key role in understanding human diseases and in the development of more predictive methods for assessing toxicity and drug or nutrient absorption. Here, we present an advanced cell culture system able to mimic the dynamic environment of biological barriers while monitoring cell behaviour through real-time impedance measurements and imaging. It consists of a fluidic device with an apical and a basal flow compartment separated by a semi-permeable membrane. The main features of the device are the integration of a transepithelial electrical impedance (TEEI) meter and transparent windows for optical monitoring within a dual flow system. Caco-2 cells were cultured in the TEEI bioreactor under both flow and static conditions. Although no differences in the expression of peripheral actin and occludin were visible, the cells in dynamic conditions developed higher impedance values at low frequencies, indicative of a higher paracellular electrical impedance and thus suggesting accelerated barrier and tight junction formation with respect to the static cultures. TEEI measurements at high frequency also enabled monitoring the evolution of transcellular impedance during culture. The cells subject to flow showed a typical RC behaviour, while the controls showed minimal capacitive behaviour, again highlighting the differences between flow and static conditions.

A dynamic in-vitro model of the intestinal epithelium for the investigation of P-glycoprotein activity

“…At higher frequency, the capacitor becomes progressively more conductive and, in the presence of a compact cell layer with well-established tight junctions, the current is able to ow across the cells (transcellular current). The higher TEER values in the bioreactors with respect to transwells suggest that the uidic conditions improve the barrier's integrity and tightness, as reported in Cacopardo et al 2019 (16).…”

“…In order to understand how the presence of ow could affect the modulation of P-gp, we studied the inhibition of intestinal P-gp by culturing Caco-2 cells in bioreactors. The systems we used combine a twocompartment cell culture device with apical and basal media ow and have been described by Giusti (Giusti et al 2014) and Cacopardo (16). Both studies demonstrate that they are capable of better mimicking physiological barriers as they recapitulate the dynamic environment of the intestine.…”

“…Usually, TEER is measured applying a low-frequency (f<5kHz) current stimulus across the cellular barrier and recording the resulting voltage (16,20). In this study, it was monitored at 40 Hz with an integrated cellular impedance-meter (16,21), in the bioreactors, and at 12.5 Hz with an Epithelial VoltOhmMeter (EVOM, World Precision Instruments, Florida USA) provided with chopsticks electrodes, in the Transwells. As reported in Cacopardo et al (16), the measurements performed by the two instruments are equivalent.…”

Promoting Cardiac Regeneration and Repair Using Acellular Biomaterials

“…These models are currently being developed to simulate ischemic injury to enable detailed investigation into the mechanisms of ischemic heart damage and the effects of therapeutic biomaterials at a mechanistic level (Chen and Vunjak-Novakovic, 2018). Larger scale milli-fluidic bioreactors use a larger scale device in comparison to microfluidic devices in order to facilitate live imaging of an artificially perfused environment while also using impedence-and resistancebased techniques to examine integrity of a layer of cells (Cacopardo et al, 2019).…”

A dynamic in-vitro model of the intestinal epithelium for the investigation of P-glycoprotein activity

“…In order to understand how the presence of flow could affect the modulation of P-gp, we studied the inhibition of intestinal P-gp by culturing Caco-2 cells in bioreactors. The systems we used combine a two-compartment cell culture device with apical and basal media flow and have been described by Giusti (Giusti et al 2014) and Cacopardo (16). Both studies demonstrate that they are capable of better mimicking physiological barriers as they recapitulate the dynamic environment of the intestine.…”