Expression of Neuropsin in the Keratinizing Epithelial Tissue–Immunohistochemical Analysis of Wild-Type and Nude Mice Naoko Inoue, Katsuki Kuwae, Akemi.

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Expression of Neuropsin in the Keratinizing Epithelial Tissue–Immunohistochemical Analysis of Wild-Type and Nude Mice Naoko Inoue, Katsuki Kuwae, Akemi Ishida-Yamamoto, Hajime Iizuka, Masao Shibata, Shigetaka Yoshida, Keiko Kato, Sadao Shiosaka Journal of Investigative Dermatology Volume 110, Issue 6, Pages 923-931 (June 1998) DOI: 10.1046/j.1523-1747.1998.00212.x Copyright © 1998 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 1 Confocal laser microscopic images of NPi in mouse skin. Cytoplasmic and lamellar-like peripheral distribution of NPi in the stratum spinosum (arrowheads) and in the stratum granulosum, respectively (a). Palm skin of mice at postnatal day 8.5. (a, b) NPi of the hair cortex and inner root sheath (c, d) [higher magnification of boxed area in (c) is shown in (d)]. Longitudinal section of back skin (e, f). NPi of the hair cortex and the Huxlay layer of the vibrissa (g, i). Transverse section of upper lip skin (g, i). Note the eosinophilic portion of the vibrissa showed strong immunoreactivity although the noneosinophilic portion lacked immunoreactivity [ arrow in (g)]. Higher magnification of boxed area in (g) is shown in (i). NPi was seen on the outer membrane of the trichohyalin granules [ arrowheads in (i) and (j)]. Parts (a), (d), and (i) are double staining images of fluorescein isothiocyanate-immunostaining of NP (green fluorescence) and nuclear staining with propidium iodide (red fluorescence). Hematoxylin and eosin staining of the same fields are presented on the right for precise identification of the tissue organization (b, e, f, h, j). Parts (a), (c), (d), (g), and (i) are at the same magnifications as (b), (e), (f), (h), and (j), respectively. bc, basal cell; h, hair; he, Henle layer; hu, Huxlay layer; ors, outer root sheath; sc, stratum corneum; sg, stratum granulosum; ss, stratum spinosum. Scale bars, (b, f, j) 50 μm; (e, h) 100 μm. Journal of Investigative Dermatology 1998 110, 923-931DOI: (10.1046/j.1523-1747.1998.00212.x) Copyright © 1998 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 2 Developmental changes in NPi in the mouse back skin. No NPi was seen in the epidermis at E14.5 (a), but NPi appeared at E15.5 (c). The granular cells appeared at this stage as shown by arrows. NPi increased at E16.5 (e), E17.5 (g), and P0.5 (i), but was slightly decreased in adult mice (6 W; k). At E17.5 (g) and P0.5 (i), lamellar-like NPi structures become prominent. Hematoxylin and eosin staining of the same fields are presented on the right (b, d, f, h, j, i). All photographs are at the same magnification. kg, keratohyalin granule; pd, periderm; pil, pilus; sb, stratum basale; sc, stratum corneum; sg, stratum granulosum; ss, stratum spinosum. Scale bar, 50 μm. Journal of Investigative Dermatology 1998 110, 923-931DOI: (10.1046/j.1523-1747.1998.00212.x) Copyright © 1998 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 3 Dynamic changes in NPi expression during morphogenetic event at the opening of the eyelids. NPi in the skin showed increased intensity and became invaginated in eyelids fuse to the direction of conjunctivae at P0.5 (a), P3.5 (b), and P9.5 (d). The photograph in (c) shows hematoxylin and eosin staining of the section shown in (b). Note, no NPi was observed in the epithelium cornea [ arrows in (a) and (d)]. Scale bars, (c, d) 50 μm; (a) and (b) are the same magnification as (c). Journal of Investigative Dermatology 1998 110, 923-931DOI: (10.1046/j.1523-1747.1998.00212.x) Copyright © 1998 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 4 Developmental changes of NPi in the tongue and palatum. NPi structures in the tongue and palatum at E15.5 (a), E16.5 (c), E17.5 (e), and P0.5 (g). In (a), NPi was first detected on the surface of the tongue. Note the saw-like NPi structure in the developing tongue at E16.5 before morphogenesis (c). The photographs in (b), (d), (f), and (h) show hematoxylin and eosin staining of the same sections as (a), (c), (e), and (g), respectively. bc, basal cell; pal, palatum; ton, tongue. Scale bar, 50 μm. Journal of Investigative Dermatology 1998 110, 923-931DOI: (10.1046/j.1523-1747.1998.00212.x) Copyright © 1998 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 5 Early appearance of NPi in the nasal pit and nasal cavity. (a) NPi structure at E14.5 and (c) higher magnification of the boxed area. (e) NPi structure at E15.5. The photographs in (b), (d), and (f) show hematoxylin and eosin staining of the sections in (a), (c), and (e), respectively. bc, basal cell; sc, stratum corneum. Scale bars, (b, f) 100 μm; (d) 50 μm. Journal of Investigative Dermatology 1998 110, 923-931DOI: (10.1046/j.1523-1747.1998.00212.x) Copyright © 1998 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 6 NPi structures in mouse stomach. NPi at P4.5 (a), P9.5 (c), and adult (6 W; e), and (g) higher magnification of the boxed area in (e). The photographs in (b), (d), (f), and (h) show hematoxylin and eosin staining of the sections in (a), (c), (e), and (g), respectively. The boxed area in (d) is enlarged in the insert. Note the clear keratohyalin granules. Dashed lines in (b), (d), and (h) represent the boundaries between the stratified squamous (sse) and the simple columnar (sce) epithelia. Arrows in (a) represent the dim NPi at P4.5 in the primordial forestomach. Asterisks show self-fluorescent luminal (lu) contents in (e) and (g). bc, basal cell. Scale bars, (b, d, h) 50 μm; (f) 100 μm. Journal of Investigative Dermatology 1998 110, 923-931DOI: (10.1046/j.1523-1747.1998.00212.x) Copyright © 1998 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 7 Comparison of epithelial NPi of nude and control mice. NPi of BALB/ C+/+ (a–d) and NPi of BALB/C nude mouse (nu/nu; e–h) in the upper lip skin. Photographs (a)–(h) were randomly taken by separate immunostained tissue sections from BALB/C+/+ (a–d) and BALB/C nude mice (e–h). (a–e) Three week old mice; (f–h) 4 wk old mouse. Note the broader and more diffuse immunostaining of NPi in nu/nu mouse skins (e–h) in the cytoplasm of the stratum granullosum compared with that in BALB/C+/+ mouse skins (a–d). All photographs are at the same magnification. Scale bar, 50 μm. Journal of Investigative Dermatology 1998 110, 923-931DOI: (10.1046/j.1523-1747.1998.00212.x) Copyright © 1998 The Society for Investigative Dermatology, Inc Terms and Conditions