Journal Articles
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Item Cornified Epithelial Teeth of Jawless Vertebrates Contain Proteins Similar to Keratin-Associated Proteins of Mammalian Skin Appendages(MDPI (Basel, Switzerland), 2025-06-01) Sachslehner AP; Parry DAD; Eckhart L; Iwata J; Johnston CAKeratins and keratin-associated proteins (KRTAPs) are the main components of mammalian nails and hair. Comparative genomics and gene expression studies have revealed that keratins are conserved in all vertebrates, whereas KRTAPs exist only in mammals. Recently, we found hair keratin-like cysteine-rich keratins in jawless vertebrates with confirmed expression in the cornified epithelial teeth of the sea lamprey (Petromyzon marinus). Here, we report that KRTAP-like proteins are also present in the horny teeth of the lamprey. Mass spectrometry-based proteomics identified proteins that share features with KRTAPs, such as high contents of cysteine and tyrosine residues, which support intermolecular interactions, and abundant glycine residues, which endow the proteins with flexibility. Genes encoding KRTAP-like proteins are arranged in a cluster in P. marinus, and the presence of at least one KRTAP-like protein is conserved in phylogenetically diverse species of lamprey, including Lampetra fluviatilis, Lethenteron reissneri, Geotria australis, and Mordacia mordax. The KRTAP-like genes of lampreys contain two exons, whereas mammalian KRTAPs have only a single exon. Although KRTAPs and KRTAP-like proteins are products of independent evolution, their common expression in cornified skin appendages suggests that they fulfill similar functions.Item Culture media and format alter cellular composition and barrier integrity of porcine colonoid-derived monolayers(Taylor and Francis Group, 2024-04-02) Barnett AM; Mullaney JA; McNabb WC; Roy NCIntestinal organoid technology has revolutionized our approach to in vitro cell culture due in part to their three-dimensional structures being more like the native tissue from which they were derived with respect to cellular composition and architecture. For this reason, organoids are becoming the new gold standard for undertaking intestinal epithelial cell research. Unfortunately, their otherwise advantageous three-dimensional geometry prevents easy access to the apical epithelium, which is a major limitation when studying interactions between dietary or microbial components and host tissues. To overcome this problem, we developed porcine colonoid-derived monolayers cultured on both permeable Transwell inserts and tissue culture treated polystyrene plates. We found that seeding density and culture format altered the expression of genes encoding markers of specific cell types (stem cells, colonocytes, goblets, and enteroendocrine cells), and barrier maturation (tight junctions). Additionally, we found that changes to the formulation of the culture medium altered the cellular composition of colonoids and of monolayers derived from them, resulting in cultures with an increasingly differentiated phenotype that was similar to that of their tissue of origin.Item Porcine colonoids and enteroids keep the memory of their origin during regeneration(American Physiological Society, 2021-05-01) Barnett AM; Mullaney JA; Hendriks C; Le Borgne L; McNabb WC; Roy NCThe development of alternative in vitro culture methods has increased in the last decade as three-dimensional organoids of various tissues, including those of the small and large intestines. Due to their multicellular composition, organoids offer advantages over traditionally used immortalized or primary cell lines. However, organoids must be accurate models of their tissues of origin. This study compared gene expression profiles with respect to markers of specific cell types (stem cells, enterocytes, goblet, and enteroendocrine cells) and barrier maturation (tight junctions) of colonoid and enteroid cultures with their tissues of origin and colonoids with enteroids. Colonoids derived from three healthy pigs formed multilobed structures with a monolayer of cells similar to the crypt structures in colonic tissue. Colonoid and enteroid gene expression signatures were more similar to those found for the tissues of their origin than to each other. However, relative to their derived tissues, organoids had increased gene expression levels of stem cell markers Sox9 and Lgr5 encoding sex-determining region Y-box 9 and leucine-rich repeat-containing G protein-coupled rector 5, respectively. In contrast, expression levels of Occl and Zo1 encoding occludin and zonula occludens 1, respectively, were decreased. Expression levels of the cell lineage markers Atoh1, Cga, and Muc2 encoding atonal homolog 1, chromogranin A, and mucin 2, respectively, were decreased in colonoids, whereas Sglt1 and Apn encoding sodium-glucose transporter 1 and aminopeptidase A, respectively, were decreased in enteroids. These results indicate colonoid and enteroid cultures were predominantly comprised of undifferentiated cell types with decreased barrier maturation relative to their tissues of origin.