Refined Immunochemical Characterization in Healthy Dog Skin of the Epidermal Cornification Proteins, Filaggrin, and Corneodesmosin

Abstract

Filaggrin (FLG) and corneodesmosin (CDSN) are two key proteins of the human epidermis. FLG loss-of-function mutations are the strongest genetic risk factors for human atopic dermatitis. Studies of the epidermal distribution of canine FLG and CDSN are limited. Our aim was to better characterize the distribution of FLG and CDSN in canine skin. Using immunohistochemistry on beagle skin, we screened a series of monoclonal antibodies (mAbs) specific for human FLG and CDSN. The cross-reactive mAbs were further used using immunoelectron microscopy and Western blotting. The structure of canine CDSN and FLG was determined using publicly available databases. In the epidermis, four anti-FLG mAbs stained keratohyalin granules in the granular keratinocytes and corneocyte matrix of the lower cornified layer. In urea-extracts of dog epidermis, several bands corresponding to proFLG and FLG monomers were detected. One anti-CDSN mAb stained the cytoplasm of granular keratinocytes and cells of both the inner root sheath and medulla of hair follicles. Dog CDSN was located in lamellar bodies, in the extracellular parts of desmosomes and in corneodesmosomes. A protein of 52 kDa was immunodetected. Genomic DNA analysis revealed that the amino acid sequence and structure of canine and human CDSN were highly similar.

Update on canine filaggrin: a review

“…This distribution is similar to what has been described in human skin (Pellerin et al 2013). This localization pattern was first evidenced using immunohistochemistry (IHC) (Kanda et al 2013;Pin et al 2019) and subsequently confirmed by post-embedding immunoelectron microscopy (Pin et al 2019). One study, using IHC , reported that patterns of Flg expression were similar in different breeds (poodles, golden retrievers, shiz tzus, pugs and labradors) and body locations (16 different regions).…”

“…In western-blot assays, two bands, of 59 and 54 kDa, corresponding to the two different predicted dog Flg monomers, were found (Kanda et al 2013;Pin et al 2019). Additionally, a band of 250 kDa that probably corresponded to canine proFlg, and some intermediate bands suggestive of proFlg processing were also observed (Pin et al 2019).…”

“…In healthy dogs, proFlg is located in keratohyalin granules of the stratum granulosum cells, and Flg in the matrix of the lower SC (Kanda et al 2013;Pin et al 2019). No labelling is observed in the upper SC, probably because of the total degradation of Flg and the formation of NMF.…”

“…However, there are major structural differences between the precursors in the two species. The number of FLG repeats varies from 10 to 12 in humans (Brown and McLean 2012) and is only 4 in dogs (Kanda et al 2013;Pin et al 2019). The number of amino acids composing the monomers and the Cand N-terminal domains are also substantially different between the species.…”

“…Canine and human proFLG sequences display little amino acid similarity (33% as shown using the Basic Local Alignment Search Tool (BLAST); the authors' unpublished data) except at the level of the S100 homologous part of the N-terminus (75%). Additionally, whereas the sequences of human FLG repeats display 40% amino acid sequence variability, there are only rare changes in the sequences of canine Flg monomers (Kanda et al 2013;Pin et al 2019).…”