A novel organotypic 3D sweat gland model with physiological functionality

Abstract

Dysregulated human eccrine sweat glands can negatively impact the quality-of-life of people suffering from disorders like hyperhidrosis. Inability of sweating can even result in serious health effects in humans affected by anhidrosis. The underlying mechanisms must be elucidated and a reliable in vitro test system for drug screening must be developed. Here we describe a novel organotypic three-dimensional (3D) sweat gland model made of primary human eccrine sweat gland cells. Initial experiments revealed that eccrine sweat gland cells in a two-dimensional (2D) culture lose typical physiological markers. To resemble the in vivo situation as close as possible, we applied the hanging drop cultivation technology regaining most of the markers when cultured in its natural spherical environment. To compare the organotypic 3D sweat gland model versus human sweat glands in vivo, we compared markers relevant for the eccrine sweat gland using transcriptomic and proteomic analysis. Comparing the marker profile, a high in vitro-in vivo correlation was shown. Carcinoembryonic antigen-related cell adhesion molecule 5 (CEACAM5), muscarinic acetylcholine receptor M3 (CHRM3), Na+-K+-Cl- cotransporter 1 (NKCC1), calcium-activated chloride channel anoctamin-1 (ANO1/TMEM16A), and aquaporin-5 (AQP5) are found at significant expression levels in the 3D model. Moreover, cholinergic stimulation with acetylcholine or pilocarpine leads to calcium influx monitored in a calcium flux assay. Cholinergic stimulation cannot be achieved with the sweat gland cell line NCL-SG3 used as a sweat gland model system. Our results show clear benefits of the organotypic 3D sweat gland model versus 2D cultures in terms of the expression of essential eccrine sweat gland key regulators and in the physiological response to stimulation. Taken together, this novel organotypic 3D sweat gland model shows a good in vitro-in vivo correlation and is an appropriate alternative for screening of potential bioactives regulating the sweat mechanism.

Hyaluronan Regulates Eyelid and Meibomian Gland Morphogenesis

“…In general, when using the hanging drop assay, factors that promote the formation of spheroids can be considered positive regulators of the developmental process. 53 , 54 Thus, HMWHA could be considered to be a positive cue for development of the MG. Interestingly, phalloidin staining revealed that in the center of the spheroids, the MG epithelial cells started to form acini ( Fig. 14 E).…”

An efficient method for eccrine gland isolation from human scalp

“…Isolation of human EGs ex vivo that maintain their morphological integrity and function can also be an invaluable source for the cosmetic industry interested in studying the sweat response to pharmacological products in healthy as well as in pathologic disorders (eg anhidrosis, hypo‐ and hyperhidrosis). As a further fascinating challenge, the possibility exists of investigating the clinical outcome after transplanting exclusively EGs in special clinical situations, for example to extensive deep‐burn survivors whose scarring skin covered with skin grafts is devoid of sweat glands and suffers from heat intolerance …”

Detection of fluid secretion of three‐dimensional reconstructed eccrine sweat glands by magnetic resonance imaging

“…Cholinergic stimulation leads to intracellular Ca 2+ ‐uptake of secretory cells, followed by the opening of K + and Cl − channels, thus activating the NKCC1 cotransporter . An in vitro study by Klaka et al., using a calcium flux assay, showed that cholinergic stimulation with acetylcholine or pilocarpine gives rise to calcium influx in a 3D hanging drop culture model of eccrine sweat glands . Another study using confocal laser scanning microscopy showed that when two‐dimensional (2D) cultured sweat gland cells were stimulated by acetylcholine, an increase in intracellular calcium concentration occurs, similar to cells in vivo .…”

“…[1,3,20] An in vitro study by Klaka et al, using a calcium flux assay, showed that cholinergic stimulation with acetylcholine or pilocarpine gives rise to calcium influx in a 3D hanging drop culture model of eccrine sweat glands. [21] Another study using confocal laser scanning microscopy showed that when two-dimensional (2D) cultured sweat gland cells were stimulated by acetylcholine, an increase in intracellular calcium concentration occurs, similar to cells in vivo. [22] However, there are no effective methods to detect fluid secretion of regenerated or reconstructed eccrine sweat glands.…”