A Versatile Model of Microfluidic Perifusion System for the Evaluation of C-Peptide Secretion Profiles: Comparison Between Human Pancreatic Islets and Hlsc-Derived Islet-Like Structures  
[PERSON]  
1 Unicyte AG, Molecular Biotechnology Center (MBC), University of Turin, Via Nizza, 52, 10126 Turin, Italy; [EMAIL_ADDRESS]
22i3T Societa per la Gestione Dell'incubatore di Imprese e per il Trasferimento Tecnologico Scarl, University of Turin, 10126 Turin, Italy; [EMAIL_ADDRESS]
3 Unicyte AG, Unicyte srl, Via Lugaro, 15, 10100 Turin, Italy; [EMAIL_ADDRESS]
4 Department of Medical Sciences, University of Turin, Corso Doglioti 14, 10126 Turin, Italy; [EMAIL_ADDRESS]
5 Correspondence: [EMAIL_ADDRESS]; Tel. : +39-011-6709588  
used to study insulin secretion in vitro [4]. The static glucose-stimulated insulin secretion (SGSIS) method consists of exposing insulin-producing cells to different glucose concentrations (or other secretagogues) under static conditions during specific time intervals (in general of 1-2 h each), and quantifying the release of insulin/C-peptide. Although it is a fairly simple method and does not require specialized devices, it has two essential limitations: (1) the static batch-type environment substantially differs from the in vivo milieu, and (2) the accumulation of the secreted insulin/C-peptide (or different hormones) may act in an autocrine manner influencing the final result [4]. On the other hand, dynamic glucose-stimulated insulin secretion (DGSIS) protocols were developed to recapitulate the in vivo environment of insulin-producing cells. In addition, dynamic protocols for assessing insulin secretion were present before the static system, just not developed fully due to technical limitations [5,6]. For instance, technical issues related to the use of the whole pancreas (hemolysis and self-digestion of the organ by exocrine enzymes) delayed a broad adoption of the dynamic perifusion methodologies. Later, in the early 1970s, the possibility to enzymatically isolate rat PIs together with the introduction of synthetic materials (plastic chambers and polyethylene tubing) allowed the modernization of the dynamic system which led to milestone discoveries such as the biphasic profile of insulin release [7]. During that period, a major limitation of the improved dynamic system was obtaining flow rates lower than 900 \(\upmu\)L/min--as a result the samples were very diluted therefore requiring highly sensitive analytical methods to quantify insulin content. More than two decades ago, the so-called \"lab-on-a-chip revolution\" was launched in the field of analytical chemistry through the introduction of the microfluidic analysis concept. This allowed the possibility of achieving lesser and more accurate flow rates through smaller devices [8,9] made of materials suitable for cell cultures [10]. On the basis of such technological advances, several groups investigated the functional secretome signature of insulin-producing cells by combining microfluidic systems with electrophoretic immunoassay assays, which allowed real-time detection of both insulin and calcium oscillations [11, 12, 13]. More recently, Raman microscopy has been incorporated to visualize molecular structures such as lipids, mitochondria, and nuclei during glucose stimulation [14]. However, since most of these devices are: commercially unavailable, complex, and expensive, their widespread distribution and use are limited. Although efforts have been devoted in developing user-friendly microfluidic devices [15,16], the diabetes research community still needs a flexible, standardized, and less expensive microfluidic perfusion device. Herein, we describe and validate a novel microfluidic perifusion system (MPS) based entirely on commercially available components that can easily be adopted for islet research.