Introduction: Hyaluronan (HA) is a major component of the skin that exerts a variety of biological functions. Inter-α-trypsin inhibitor heavy chain (ITIH) proteins comprise a family of hyaladherins of which ITIH5 has recently been described in skin, where it plays a functional role in skin morphology and inflammatory skin diseases including allergic contact dermatitis (ACD). Objective: The current study focused on the ITIH5-HA interaction and its potential clinical and functional impact in extracellular matrix (ECM) stabilization. Methods: Studying the molecular effects of ITIH5 in skin, we established skin models comprising murine skin cells of Itih5 knockout mice and corresponding wild-type controls. In addition, human dermal fibroblasts with an ITIH5 knockdown as well as a murine recombinant Itih5 protein were established to examine the interaction between ITIH5 and HA using in vitro adhesion and HA degradation assays. To understand more precisely the role of ITIH5 in inflammatory skin diseases such as ACD, we generated ITIH5 knockout cells of the KeratinoSens® cell line. Results: Using murine skin models, ITIH5 knockdown fibroblasts, and a reactive oxygen species (ROS)-mediated HA degradation assay, we proved that ITIH5 binds to HA, thereby acting as a stabilizer of HA. Moreover, microarray profiling revealed the impact of ITIH5 on biological processes such as skin development and ECM homeostasis. Performing the in vitro KeratinoSens skin sensitization assay, we detected that ITIH5 decreases the sensitizing potential of moderate and strong contact sensitizers. Conclusion: Taken together, our experiments revealed that ITIH5 forms complexes with HA, thereby on the one hand stabilizing HA and facilitating the formation of ECM structures and on the other hand modulating inflammatory responses.

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