Robert H. Rice, Marjorie A. Phillips, John P. Sundberg

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Localization of Hair Shaft Protein VSIG8 in the Hair Follicle, Nail Unit, and Oral Cavity Robert H. Rice, Marjorie A. Phillips, John P. Sundberg Journal of Investigative Dermatology Volume 131, Issue 9, Pages 1936-1938 (September 2011) DOI: 10.1038/jid.2011.133 Copyright © 2011 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 1 Immunoreactivity of VSIG8 antiserum in BALB/cByJ+/+ albino mouse hair follicles. (a) Immunoreactivity was limited to cuticle and cortex layers of the hair shaft, whereas the medulla was not labeled. (b) Immunostaining was evident intracellularly and at the periphery of cuticle cells. (c) In the transmission electron micrograph of hairs from FVB/NJ mice, note the prominent projections from the cortex into the medulla (white arrows). (d) In immunohistochemical analysis of BALB/cByJ hair fibers (late anagen follicles), VSIG8 antiserum labels these cortical projections intensely (white arrows). (e–k) Immunoreactivity is shown through the hair cycle in representative hair follicle vertical sections from C3H/HeJ+/+ mice. Skin samples were collected for analysis at 3-day intervals after synchronization by wax stripping. Black melanin pigment is easily differentiated from the brown diaminobenzidine label. The cuticle of the hair shaft and cortex are labeled. Samples at days 0 (not shown) and 3 revealed no immunoreactivity, those at days 6–18 were immunopositive (late anagen and early catagen), and those at days 21 and 24 (not shown) were immunonegative. VSIG8 expression in the superficial layers of nail matrix is shown (l) in a newborn C57BL/6J mouse, (m) enlarged below. (n) VSIG8 immunoreactivity in the oral cavity was evident in superficial layers of the interpapillary epithelium of the dorsal tongue, (o) at the base of the tongue, and (p) in the gingival epithelium adjacent to the tooth. For these studies, wild-type mice of six strains were used (nine C57BL/6J, three NON/ShiLtJ, one DBA/1LacJ, one C3H/HeJ, and one CBA/J); in addition, two female C3H/HeJ mice at each time point beginning at 70 days of age (24 mice) were used for wax stripping. No variation in expression patterns was observed. (a, b, e–k, m, o, p) Bars=2μm and (l, n) bars=10μm. Journal of Investigative Dermatology 2011 131, 1936-1938DOI: (10.1038/jid.2011.133) Copyright © 2011 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 2 Immunoblotting of tissue extracts and transfected coding region. Extracts of rat esophagus (lane 1), tongue (lane 2), mouse tongue (lane 3), and esophagus (lane 4) yielded single bands of ≈45kDa with mobility matching that was expressed by the human VSIG8 coding sequence (lanes 5 and 7) upon transient transfection of HeLa (or human embryonic kidney 293, not shown) cultures. Lanes 6 and 8 show no bands in parallel mock-transfected cultures. Lanes 5 and 6 were blotted with absorbed antiserum, whereas lanes 7 and 8 were blotted with unabsorbed antiserum. Journal of Investigative Dermatology 2011 131, 1936-1938DOI: (10.1038/jid.2011.133) Copyright © 2011 The Society for Investigative Dermatology, Inc Terms and Conditions