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International audience; Neural crest cells constitute a unique population of progenitor cells with extensive stem cell capacities able to navigate throughout various environments in the embryo and are a source of multiple cell types, including neurons, glia, melanocytes, smooth muscles, endocrine cells, cardiac cells, and also skeletal and supportive tissues in the head. Neural crest cells are not restricted to the embryo but persist as well in adult tissues where they provide a reservoir of stem cells with great therapeutic promise. Many fundamental questions in cell, developmental, and stem cell biology can be addressed using this system. During the last decades there has been an increased availability of elaborated techniques, animal models, and molecular tools to tackle neural crest cell development. However, these approaches are often very challenging and difficult to establish and they are not adapted for rapid functional investigations of mechanisms driving cell migration and differentiation. In addition, they are not adequate for collecting pure populations of neural crest cells usable in large scale analyses and for stem cell studies. Transferring and adapting the neural crest system in tissue culture may then represent an attractive alternative, opening up numerous prospects. Here we describe a simple method for establishing primary cultures of neural crest cells derived from trunk neural tubes using the avian embryo as a source of cells. This protocol is suited for producing pure populations of neural crest cells that can be processed for cytological, cellular, and functional approaches aimed at characterizing their phenotype, behavior, and potential. © 2020 Wiley Periodicals LLC. Basic Protocol: Primary cultures of avian trunk neural crest cells Support Protocol: Adaptations for immunofluorescence labeling and videomicroscopy.

Candidiasis is characterized by susceptibility to recurrent or persistent infections caused by Candida spp., typically Candida albicans, of cutaneous and mucosal surfaces. In this report, function and frequency of Th17 cells as well as genetics of patients susceptible to mucocutaneous candidiasis were studied. For patients, T-cell proliferation tests in response to Candida antigen, Th17 cell proportions, and STAT1 phosphorylation were evaluated through flow cytometry. Expression of IL17A, IL17F and IL22 genes were measured by real-time quantitative PCR. At the same time, whole exome sequencing was performed for all patients. We identified two heterozygous substitutions, one: c.821G > A (p. R274Q) was found in a multiplex family with three individuals affected, the second one: c.812A > C (p. Q271P) was found in a sporadic case. Both mutations are located in the coiled-coil domain (CCD) of STAT1. The frequency of Th17 cells, IL17A, IL17F, and IL22 gene expression in patients’ peripheral blood mononuclear cells (PBMCs), and T-cell proliferation to Candida antigens were significantly reduced in the patients as compared to healthy controls. An increased STAT1 phosphorylation was observed in patients’ PBMCs upon interferon (IFN)-γ stimulation as compared to healthy controls. We report two different but neighboring heterozygous mutations, located in exon 10 of the STAT1 gene, in four Iranian patients with CMC, one of whom also had hypothyroidism. These mutations were associated with impaired T cell proliferation to Candida antigen, low Th17 cell proportions, and increased STAT1 phosphorylation upon IFN-γ. We suggest that interfering with STAT1 phosphorylation might be a promising way for potential therapeutic measurements for such patients.