Staining of human skin with RGB trichrome unveils a proteoglycan-enriched zone in the hair dermal sheath

Abstract

The skin is the largest organ in the body and plays several essential functions acting as a barrier that protects us from physical and chemical insults, prevents the entrance of pathogens and the loss of water, besides playing an esential role in the regulation of body temperature. The skin displays a high regenerative capacity, evidenced by its self-renewing activity and the process of wound healing, driven by the existence of several resident stem cell populations. Due to the high prevalence of skin pathology, and their anatomical accessibility, skin biopsies and their subsequent microscopic observation constitute a powerfull tool for the study of primary skin diseases, as well as cutaneous manifestations of systemic diseases. This gave rise to dermatopathology as a specific discipline that unifies dermatology and pathology. In this setting, staining with hematoxylin and eosin constitutes the gold standard method for microscopic observation and diagnosis. Yet, several additional stains are used for specific purposes, such as trichrome stains for the staining of collagens in the extracellular matrix. We have applied a recently developed stain (RGB trichrome, acronym for picrosirius Red, fast Green and alcian Blue) to human samples to assess the staining outcomes in normal skin tissues. RGB staining provides a high contrasted interface between epidermis and dermis, and a comprehensible staining of the different dermal structures such as blood vessels, nerves, and sweat and sebaceous glands. The specific staining of collagens by picrosirius red can be useful for the objective quantification of these proteins under polarized light microscopy. In hair follicles, RGB staining resulted in specific staining of the epidermal sheaths and the encasing connective tissue (i.e., dermal sheath). Interestingly, the dermal sheath show two domains in which blue predominates over red staining, thus indicating that proteoglycans prevail in these areas. These two zones are the dermal papilla and an uncharacterized zone located at the upper isthmus, that we have denominated as proteoglycan-enriched belt (PEB). While the role of the papilla in the induction and regulation of hair growth is clearly established, the possible role of PEB remains to be determined and merits additional investigation.