Optimised generation of iPSC-derived macrophages and dendritic cells that are functionally and transcriptionally similar to their primary counterparts.

Induced pluripotent stem cells (iPSC) offer the possibility to generate diverse disease-relevant cell types, from any genetic background with the use of cellular reprogramming and directed differentiation. This provides a powerful platform for disease modeling, drug screening and cell therapeutics. The critical question is how the differentiated iPSC-derived cells translate to their primary counterparts. Our refinement of a published differentiation protocol produces a CD14+ monocytic lineage at a higher yield, in a smaller format and at a lower cost. These iPSC-derived monocytes can be further differentiated into macrophages or dendritic cells (DC), both with similar morphological and functional profiles as compared to their primary counterparts. Transcriptomic analysis of iPSC-derived cells at different stages of differentiation as well as comparison to their blood-derived counterparts demonstrates a complete switch of iPSCs to cells expressing a monocyte, macrophage or DC specific gene profile. iPSC-derived macrophages respond to LPS treatment by inducing expression of classic macrophage pro-inflammatory response markers. Interestingly, though iPSC-derived DC show similarities to monocyte derived DC, they are more similar transcriptionally to a newly described subpopulation of AXL+ DC. Thus, our study provides a detailed and accurate profile of iPSC-derived monocytic lineage cells.

Application of cryopreserved cells to HERG screening using a non-radioactive Rb+ efflux assay.

The aim of the present study was to test the use of cryopreserved cells for human ether-ago- go (hERG) channel screening. A stable Chinese hamster ovary-K1 cell line expressing hERG channels was used in a non-radioactive Rb+ efflux assay for screening hERG channel activity. Cells prepared from normal cultures and following cryopreservation were compared with regard to time course of Rb+ efflux response at different concentrations of KCl, overall cell morphology, signal-to noise assay window, assay robustness, and 50% inhibitory concentration (IC50) of the hERG blocker dofetilide. Our results showed that cryopreserved hERG-expressing cells at 1, 2, and 3 days following plating at different cell densities had no obvious morphological changes in comparison to the growing cells. The assay window, Z' factor, and IC50 of dofetilide were also very similar in all series of cells tested. It was concluded that the cryopreserved hERG cells prepared using the protocol described are as effective as growing hERG cells in the non-radioactive Rb+ efflux assay.

TRPV1 signaling: mechanistic understanding and therapeutic potential.

Transient receptor potential vanilloid 1 (TRPV1) is a non-selective cation channel gated by noxious heat, vanilloids and extracellular protons. TRPV1 is acting as an important signal integrator in sensory nociceptors under physiological and pathological conditions including inflammation and neuropathy. Because of its integrative signaling properties in response to inflammatory stimuli, TRPV1 agonists and antagonists are predicted to inhibit the sensation of ongoing or burning pain that is reported by patients suffering from chronic pain, therefore offering an unprecedented advantage in selectively inhibiting painful signaling from where it is initiated. In this article, we firstly summarize recent advances in the understanding of the role of TRPV1 in pain signaling, including a overview of clinical pharmacological trials using TRPV1 agonists and antagonists. Finally, we also present an update on the mechanistic understanding and controlling of hyperthermia caused by TRPU1 antagonists, and provide perspective for future study.