OBSERVATION Architectural Organization of Filiform Papillae in Normal and Black Hairy Tongue Epithelium Dissection of Differentiation Pathways in a Complex Human Epithelium According to Their Patterns of Keratin Expression Motomu Manabe, MD, PhD; Henry W. Lim, MD; Martin Winzer, MD; Cynthia A. Loomis, MD, PhD Background: An inadequate understanding of the lying the primary papillae and between the individual pri- complex morphologic characteristics of human fili- mary papillae express esophageal-type keratins. In black form papillae has hampered the histopathological hairy tongue disease, there is a marked retention of sec- characterization of disorders affecting tongue kera- ondary papillary cells expressing hair-type keratins. tinization. To better define the 3-dimensional cyto- architecture of tongue epithelium, we performed Conclusions: Using a panel of antikeratin probes, we detailed immunohistochemical analyses of normal and define the precise topographical localization of cell black hairy tongue tissues using a panel of antikeratin populations undergoing 3 distinct differentiation pro- antibodies. grams in dorsal tongue epithelium. Comparative analy- ses of black hairy tongue specimens indicate that defec- Observations: The dome-shaped base of the human fi- tive desquamation of the cells in the central column of liform papilla (primary papilla) is surmounted by 3 to 8 filiform papillae results in the formation of highly elon- elongated structures (secondary papillae). These second- gated, cornified spines or “hairs”—the hallmark of this ary papillae are composed of a central column of epithe- disease. lial cells expressing hair-type keratins and an outer rim of cells expressing skin-type keratins. The epithelium over- Arch Dermatol. 1999;135:177-181 HE DORSAL surface of mam- cumulative data indicate that, in addition malian tongue is covered by to the ubiquitous expression of the strati- densely packed filiform pa- fied epithelial-type keratins (K5 and K14), pillae. The shape and size different regions of the tongue epithe- of these papillae vary mark- lium display distinct patterns of keratin ex- Tedly from species to species.1-8 In hu- pression. The nonkeratinized epithelium mans, the papillary architecture is more of the lateral and ventral surfaces of the complex, consisting of a central body sur- tongue, as well as the epithelium cover- rounded by several threadlike cornified ing the lateral aspects of filiform and projections, often referred to as second- fungiform papillae and the interpapillary ary papillae.5,9 Morphologically, human mucosa of the dorsal tongue, express tongue epithelium seems to be divided esophageal-type keratins (K4 and K13). In into discrete domains that undergo dis- contrast, the orthokeratinized epithe- tinct pathways of terminal differentia- lium overlying the tips of filiform papil- tion, similar to what occurs in rodent and lae makes skin-type keratins (K1 and K10), From the Departments of cow tongue epithelium. However, the mor- and the epithelium covering taste buds Dermatology, Akita University phologic compartments of the human synthesizes simple epithelial-type kerat- School of Medicine, Akita, tongue have not been as well character- ins (K8, K18, and K19). Finally, we and Japan (Dr Manabe), and Henry ized as those of the rodent because of their others14-16 have shown that tongue epithe- Ford Hospital, Detroit, Mich greater complexity. lium also produces an acidic hard-type (Dr Lim); the Keratins are a heterogeneous family keratin, characteristic of hair- or nail- Dermatohistopathology of polypeptides that form a subclass of in- type differentiation. Laboratory, Westerstede, Germany (Dr Winzer); and The termediate filaments. They serve as excel- In this article, we extend our immu- Ronald Perelman Department of lent markers for various pathways of epi- nolocalization studies on the structure and Dermatology, New York thelial differentiation and have been used compartmentalization of the human fili- University School of Medicine, to define discrete populations of keratino- form papilla and begin to explore the role NY (Dr Loomis). cytes within the tongue epithelium.10-13 The of such compartments in human disease. ARCH DERMATOL / VOL 135, FEB 1999 177 ©1999 American Medical Association. All rights reserved. Downloaded From: https://jamanetwork.com/ on 09/29/2021 RESULTS MATERIALS AND METHODS SPECIFICITY OF THE AE 20 ANTIBODY FOR HUMAN EPIDERMAL KERATIN HUMAN TISSUE In our prior study of human tongue epithelium,7 in- Normal human tongues were obtained from au- direct immunofluorescent staining was performed us- topsy within 48 hours of death. The 3 biopsy speci- ing the monoclonal antibody AE 2, which recognizes mens of BHT were obtained from former patients markers of skin-type differentiation, K1 and K10.19 Un- of Manhattan Veterans Affairs Medical Center fortunately, AE 2 cross-reacts with filaggrin, a major com- in New York as well as Medical University of Lu¨ beck ponent of keratohyalin granules (data not shown), mak- in Germany. ing it difficult to localize the skin-type keratins. To obtain more definitive immunolocalization data, we generated ANTIKERATIN ANTIBODIES a monoclonal antibody, AE 20, that reacts specifically with Detailed characterizations of the monoclonal anti- the basic 67-kd human epidermal keratin, K1, as as- body AE 8, which reacts specifically with the sessed by immunoblotting (data not shown). No AE 20 esophageal-type keratin K13, and the antibody AE immunoreactivity was detected in keratin extracts from 13, which reacts with 44- to 46-kd acidic hair ker- a variety of other tissues and cultured cells tested (data atins, are described elsewhere.17,18 The AE 20 anti- not shown). body was generated against trypsinized human skin cells according to the methods described pre- SEVERAL DISTINCT DIFFERENTIATION 19 viously. The antibody to K6 was generously pro- DOMAINS OF CROWN-SHAPED vided by Dennis Roop, PhD, Baylor University, HUMAN FILIFORM PAPILLA Houston, Tex. KERATIN EXTRACTION, The human filiform papilla consists of an elevated con- GEL ELECTROPHORESIS, nective tissue core covered by a partially keratinized strati- AND IMMUNOBLOTTING fied epithelium. Small connective tissue protrusions ema- nate from the top of this central core and are surmounted Normal human epidermis and cultured epithelial by a column of epithelial cells undergoing cornification. cells from various tissues were first extracted with These elongated cornified spines are referred to as sec- 25-mmol/L Tris-hydrochloride, 0.6-mol/L po- ondary papillae and characteristically tilt toward the phar- tassium chloride, 1% Triton X-100 supplemented 20 ynx. In contrast to the papillary epithelium, the inter- with 5 protease inhibitors. The residual aqueous- papillary surface is nonkeratinized. insoluble cytoskeletal preparation, containing To facilitate the topological analysis of this com- mainly keratin proteins, was then solubilized with 2% sodium dodecyl sulfate in 25-mmol/L plex structure, we used a panel of monoclonal antibod- Tris-hydrochloride (pH 7.4) as described.20 ies recognizing keratins that are characteristic of differ- One-dimensional sodium dodecyl sulfate– ent types of stratified epithelia. The AE 8 monoclonal polyacrylamide gel electrophoresis and subsequent antibody specifically reacts with the K4 keratin, which immunoblotting were performed according to the is expressed in nonkeratinized, stratified epithelia such methods described previously.21 as in the esophagus. The AE 13 antibody recognizes acidic hair keratins, which are expressed in hair cortex, nail IMMUNOHISTOCHEMICAL STAINING plate, and mouse tongue filiform papillae.14,17 The AE 20 antibody, described above, recognizes K1, which is char- The biopsy specimens were embedded in OCT com- pound (Ty Miles Inc, Westchester, Ill), snap frozen acteristic of keratinized epithelia such as skin. The an- in liquid nitrogen, and cut into 6-µm cryostat sec- tibody to K6 recognizes a keratin that is normally ex- tions. Sections were stained by the indirect immu- pressed at a few restricted sites of the body, including nofluorescent and immunoperoxidase technique.19,22 volar skin, foreskin, oral mucosa, and outer root sheath cells of the hair follicle, as well as by all stratified epi- thelial cells undergoing hyperproliferation.23-25 Using this panel of antikeratin antibodies, we con- firmed that the epithelium covering filiform papillae is We present the characterization of a new antikeratin subdivided into at least 3 distinct cell domains (data not K1 monoclonal antibody and its use to definitively shown). As reported previously, the antiesophageal kera- localize the skin-type compartment in human tongue tin antibody, AE 8, reacted with the nonkeratinizing epi- epithelium. In addition, we provide a more detailed thelia between the filiform papillae as well as between description of the 3-dimensional organization of the the cornified hairlike projections. The AE 20 immuno- separate domains within human filiform papillae. reactivity was confined to cells at the bases of the corni- Finally, we demonstrate that the hairlike projections fied spines, indicating that the skin-type keratin K1 is re- observed in black hairy tongue disease (BHT) are due stricted to this cell population. The AE 13 antihair keratin primarily to the formation of abnormally long exten- antibody reacted only with those cells directly beneath sions of the “hair compartment” of the filiform the cornified spines, adjacent to the AE 20–positive cells. papillae.