Acta histochem. (lena) 93, 256-263 (1992) Gustav Fischer Verlag lena' Stuttgart· New York
Departments of Dermatology!) and Plastic Surgery2), and Centre of Animal Research 3 ), Friedrich-Schiller-University, lena, Germany
Histochemistry of the porcine pilosebaceous unit By UWE WOLLINA'), UWE BERGER2), CHRISTINE STOLLE l ), HEIKO STOLLE l ), HARALD SCHUBERT3) and CHRISTINA HIPLER l ) With 2 Figures (Received February 22, 1991; revised lune 28, 1991; accepted November 28, 1991) Key words: Porcine skin, Pilosebaceous unit, Human skin, Lectin histochemistry, Immunohistochemistry
Abstract The present study describes lectin and immunoreactivilY in the pilosebaceous unit of porcine skin. Complex carbohydrates of mucin and biantennary Man/Gluc types were distributed among hair follicle epithelia (hair root sheaths, cuticula, shaft, and shaft matrix). Sebaceous glands expressed biantennary Man/Gluc carbohydrates and GalNAc residues. The expression of simple-type and epidermis-like keratins was confmned by immunohistochemistry with monoclonal antibodies. Filaggrin-positive cells were found in the keratinizing zone of HENLE'S layer in anagen follicles. The innermost layer of the outer hair root sheath was stained with antibodies against the epidermal growth factor-receptor, keratin 10 and Ki67 antigen. The differences to humans were remarkably small.
1. Introduction The domestic pig has become an accepted model of medical research. In particular, porcine skin resembles human skin in some important qualities making it reliable for experimental dermatology, surgery, toxicology, pharmacology, and radiology (ARCHAMBEAU and BENNETT 1984; MARACRIAN and CALHOUN 1966; MONTAGNA and YUN 1964; MEYER et al. 1986; TSUKISE and MEYER 1983; WOLLINA et al. 1991 a, b). In contrast to human skin, pigs show a trio clustering of hair follicles. Hair growth in pigs but not in humans discloses a remarkable annual cyclic activity (WATSON and MOORE 1990). In most domestic mammals, sebaceous glands are paired. In pigs they are larger than in densely haired species (MARCARIAN and CALHOUN 1966; MONTAGNA and YUN 1964; MEYER et al. 1978; MEYER and NEURAND 1987). Therefore, it seems questionable, that porcine skin provides a reliable model in biomedical hair research. Since histochemical data on the porcine hair follicle and sebaceous gland are extremely sparse, the present study was initiated in order to elucidate the differences of porcine and human skin.
2. Material and methods 2.1. Tissue samples were obtained from the hairy skin ofthe trunk of 6 domestic pigs of both sexes (age 10 to 15 month; Institute of Animal Contagion Research, Academy of Veterinary Sciences, lena) and of 15 minipigs [MINILEWEj (age 2 to 2.5 a; 12 female and 3 castrated males; Centre of Animal Research, University of lena) under anesthesia. Skin specimens were snap frozen in liquid nitrogen and cut at 5 ~m.
Histochemistry of the porcine pilosebaceous unit
Table I. Lectins used. Abbreviation
Major sugar specificity*
Con A PSA UEA I WGA
Canava/ia ensiformis Pi"lIIJ1 sativum Ulex europaeus I Triticulll vulgare
ty-D-Man/ty-D-Gluc D-Man, D-Gluc ty-L-Fuc GlcNAc131 -> 4 sequences
* According to GALLAGHER (1984) and OSAWA and TSUJI (1987).
2.2. Leetins (Table I) were labelled with f1uorescein-isothiocyanate and applied on unfixed frozen sections in the direct fluorescence technique [(DIF); (WOLLINA et al. 1990a)] with a modification of JOHNSON and DE NOGUEIRA ARANJO (1981) to reduce fading. PSA was purchased from Sigma. PNA from SIFIN (BerlinWeiBensee/FRG), ConA from the Academy of Agricultural Sciences (Gatersleben/FRG), WGA from the Department of Biotechnology. University of Leipzig, and UEA from Boehringer. Mannheim/FRG. Table 2. Antibodies lCK. cytokeratin(s)]. Specificity
CK 8, 18. 19
CK 13, 16
IgG I (mouse)
IgG I (mouse)
Vim 9 (I)
vimentin type 9
IgG I (mouse)
IgG I (mouse)
Paesel & Lorei
Dr. WENZ, JenalFRG
collagen type IV
IgG 1 (mouse)
19G 1 (mouse)
2.3. Po/y- and monoclonal antibodies against tissue antigens are listed in Table 2. Secondary antibodies for peroxidase technique (POX) were purchased from Dako (Hamburg/FRG). For immunostaining we followed the protocol of STERNBERGER et al. (1970) using acetone-fixed tissue sections. POX was developed with 3-amino-9ethylcarbazole (EGA-Chemie. Steinheim). 2.4. Staining CO/1/ro/s included inhibition of lectins by monosaccharides (WOLLINA et al. 1990a) and substitution of the primary antibodies by buffer solution. Examination was done with either the lenalumar fluorescence microscope or the Histova/light microscope (Carl Zeiss lena, FRG).
3. Results The distribution of lectin binding sites and tissue antigens in porcine and human epithelia of the pilosebaceous unit is summarized in Table 3. Domestic pigs and minipigs provided identical histochemical results. 17
Acta histochem .. Bd. 93. t
U. WOLLINA et al.
Table 3. Comparison of histochemical reactivity in the pilosebaceous units of porcine (P) and human skin (H). Tissue
PSA, PNA, ConA ACAM (supraglandular) K 8.12 anti-filaggrin supraglandular zone keratinizing zone
++ + ++
+ ++ + + +
Hair follicle outer hair root sheath
innermost layer of the outer root sheath inner hair root sheath
29.11 Ki67, RKSE 60 ConA ACAM Cam 5.2 (infraglandular) RKSE 60
shaft and shaft matrix
RKSE 60 PNA PSAlUEA I ACAM Cam 5.2 PSA
dermal hair follicle sheath
anti-collagen anti-vimentin, Vim9(l)
Mm. arrectores pili
++ ++ ++ +* +*
+ + ++ ++ ++ ++ +* +*
+* ++ ++ ++ + +
+* +* ++ ++ + + +
++ ++ + ++
++ ++ + +
Sebaceous gland peripheral sebocytes
ConA, WGA ACAM
sebaceous duct cells
ACAM, K 8.12
* Single cells or mosaic-like pattern. §
The shaft was PSA-negative.
3. 1. Sebaceous glands Sebocytes: No distinct patterns were observed between porcine and human specimens. In general, the basal (immature) sebocytes showed a stronger reactivity with lectins as well as with antibodies. These cells expressed