Hit screening with multivariate robust outlier detection.

Hit screening, which involves the identification of compounds or targets capable of modulating disease-relevant processes, is an important step in drug discovery. Some assays, such as image-based high-content screenings, produce complex multivariate readouts. To fully exploit the richness of such data, advanced analytical methods that go beyond the conventional univariate approaches should be employed.

Highly scalable arrayed CRISPR mediated gene silencing in primary lung small airway epithelial cells.

Small airway epithelial cells (SAECs) play a central role in the pathogenesis of lung diseases and are now becoming a crucial cellular model for target identification and validation in drug discovery. However, primary cell lines such as SAECs are often difficult to transfect using traditional lipofection methods; therefore, gene editing using CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 is often carried out through ribonucleoprotein (RNP) electroporation. Here we have established a robust, scalable, and automated arrayed CRISPR nuclease (CRISPRn) screening workflow for SAECs which can be combined with a myriad of disease-specific endpoint assays.

A multi-parametric high throughput assay for detecting beta-cell proliferation in dispersed primary islets.

Identification of novel compounds to selectively induce pancreatic beta-cell proliferation has the potential to restore functional beta-cell mass and insulin secretory demand in type 2 diabetes. The rarity of islet cell clusters (comprising of only 1% of the total pancreas mass) makes such a discovery a challenge. To address this obstacle a high throughput, 384 well, plate-based multi-parametric imaging assay was developed to capture ex vivo primary islet proliferation, allowing positive identification of compounds that can selectively enhance islet beta-cell proliferation.

CGRP, adrenomedullin and adrenomedullin 2 display endogenous GPCR agonist bias in primary human cardiovascular cells.

Agonist bias occurs when different ligands produce distinct signalling outputs when acting at the same receptor. However, its physiological relevance is not always clear. Using primary human cells and gene editing techniques, we demonstrate endogenous agonist bias with physiological consequences for the calcitonin receptor-like receptor, CLR.

High Seeding Density Induces Local Hypoxia and Triggers a Proinflammatory Response in Isolated Human Islets.

Hypoxia is the main threat to morphological and functional integrity of isolated pancreatic islets. Lack of oxygen seems to be of particular importance for functionality of encapsulated islets. The present study was initiated as an experimental model for the environment experienced by human islets in a confined space present during culture, shipment, and in an implanted macrodevice.

Elevated levels of branched-chain amino acids have little effect on pancreatic islet cells, but L-arginine impairs function through activation of the endoplasmic reticulum stress response.

Recent metabolic profiling studies have identified a correlation between branched-chain amino acid levels, insulin resistance associated with prediabetes and susceptibility to type 2 diabetes. Glucose and lipids in chronic excess have been reported to induce toxic effects in pancreatic β-cells, but the effect of elevated amino acid concentrations on primary islet cell function has not been investigated to date. The aim of this study was to investigate the effect of chronic exposure to various amino acids on islet cell function in vitro.

Activation of the NLRP3 inflammasome by islet amyloid polypeptide provides a mechanism for enhanced IL-1β in type 2 diabetes.

Interleukin 1β (IL-1β) is an important inflammatory mediator of type 2 diabetes. Here we show that oligomers of islet amyloid polypeptide (IAPP), a protein that forms amyloid deposits in the pancreas during type 2 diabetes, triggered the NLRP3 inflammasome and generated mature IL-1β. One therapy for type 2 diabetes, glyburide, suppressed IAPP-mediated IL-1β production in vitro.