Dielectric spectroscopy for monitoring human pancreatic islet differentiation within cell-seeded scaffolds in a perfusion bioreactor system

Abstract

The long-term in vitro culture and differentiation of human pancreatic islets is still hindered by the inability to emulate a suitable microenvironment mimicking physiological extracellular matrix (ECM) support and nutrient/oxygen perfusion. This is further amplified by the current lack of a non-invasive and rapid monitoring system to readily evaluate cellular processes. In this study, we realized a viable method for non-invasively monitoring isolated human pancreatic islets in vitro. Islets are induced to dedifferentiate into proliferative duct-like structures (DLS) in preparation for potential and subsequent re-differentiation into functional islet-like structures (ILS) in a process reminiscent of islet regeneration strategies. This long-term in vitro process is conducted within a three-dimensional microenvironment involving islets embedded in an optimized ECM gel supported by microfabricated three-dimensional scaffolds. The islet-scaffold is then housed and continuously perfused within chambers of a bioreactor platform. The process in its entirety is monitored through dielectric spectroscopy measurements, yielding an accurate representation of cellular morphology, functionality, and volume fraction. This non-invasive and real-time monitoring tool can be further manipulated to elucidate important information about the optimized cellular microenvironment required for maintaining long-term culture and achieve efficient differentiation for islet regeneration.

Elaboration of a finite element model of pancreatic islet dielectric response to gap junction expression and insulin release

“…Accordingly, the elaborated FEA model contains gap junctions between cells and adjoining membranes representing adherens and tight junctions. The shape of the model dielectric spectra was comparable with the experimental islet dielectric spectra obtained in our previous work (7,18). Gap junction density between the model islet cells was varied to evaluate the effect of this parameter on the dielectric spectra.…”

Impedance-based cellular assays for regenerative medicine

“…Cell proliferation and differentiation have been investigated with EIS with a dielectric probe that limited the use of the low frequency field [84 -86] but was able to monitor large-scale samples. The sensitivity was increased by Daoud et al [87] by using ad hoc macrochambers lined up with parallel-plate platinum electrodes; they enabled impedance measurements in the beta-dispersion frequency range. The epithelial differentiation processes of Madin-Darby canine kidney cells to hollow cyst-like structures were captured with this technique.…”

The emerging field of pancreatic tissue engineering: A systematic review and evidence map of scaffold materials and scaffolding techniques for insulin-secreting cells

“…59 The lack of retrievability of the microcapsules is a disadvantage of this technique, so different in vivo monitoring methods have been investigated for non-invasive tracking of the microcapsules. 59,69,81,82 The application of new imaging techniques can also provide information about cell morphology, spatial distribution, and oxygen supply within a scaffold or hydrogel encapsulation. 83,84…”

“…112 It can be concluded that the alginate/gelatin composite hydrogel that was plotted is not the ideal biomaterial for the purpose of engineering an insulin-secreting construct with this technique. A study by Daoud et al 82,113 used PLGA to 3D-print a scaffold matrix. A cell–gel composite containing human islet cells, collagen I, collagen IV, and fibronectin was then added.…”