Expression of keratin K2e in cutaneous and oral lesions: association with keratinocyte activation, proliferation, and keratinization

Abstract

The cytoskeleton in keratinocytes is a complex of highly homologous structural proteins derived from two families of type I and type II polypeptides. Keratin K2e is a type II polypeptide that is expressed in epidermis late in differentiation. Here we report the influence of keratinocyte activation, proliferation, and keratinization on K2e expression in samples of cutaneous and oral lesions. The normal expression of K2e in the upper spinous and granular layers of interfollicular epidermis is increased in keloid scars but showed distinct down-regulation in psoriasis and hypertrophic scars where keratinocytes are known to undergo activation. Unlike normal and psoriatic skin, K2e expression in hypertrophic and keloid scars began in the deepest suprabasal layer. In cutaneous basal and squamous cell carcinomas, K2e was absent in most tumor islands but the overlying epidermis showed strong expression. No significant K2e expression in nonkeratinized or keratinized oral epithelia, including buccal mucosa, lateral border of tongue and gingiva was detected. In oral lichen planus K2e expression was undetectable, but in benign keratoses of lingual mucosa induction of K2e along with K1 and K10 was observed. In mild-to-moderate oral dysplasia with orthokeratinization, K2e was highly expressed compared with parakeratinized areas but in severe dysplasia as well as in oral squamous cell carcinoma, K2e expression was undetectable. Taken together, the data suggest that K2e expression in skin is sensitive to keratinocyte activation but its up-regulation in oral lesions is a reflection of the degree of orthokeratinization.

Figure 1.

Immunoreactivity of keratin polypeptides with keratin monoclonal antibodies. Human epidermis from breast skin was removed either by treatment with dispase (lane 1) or keratinocytes isolated by treatment with trypsin (lane 2) and cultured until they became confluent (lane 3). Keratin polypeptides were isolated from these samples as described in the Materials and Methods section and analyzed on SDS polyacrylamide gels. The protein profile as determined by Coomassie brilliant blue staining is shown in the CBB panel. The polypeptides were transferred onto nitrocellulose membranes and probed for keratins K14 (LLOO1), K6 (LLO25), K2e (LHK2e), K10 (RKSE60), and K1 (LHK1).

Figure 2.

Expression of keratin polypeptides and Ki67 in normal epidermis from different body sites. Histological appearance of normal abdominal epidermis (A) and a few Ki67-positive cells restricted to the normal basal layer in an adjacent section (B). In the same specimen, protein for K1 (C) and K10 (D) are homogeneously expressed throughout the suprabasal compartment whereas expression of K2e protein is restricted to the upper suprabasal layers (E). However, in a sample of facial skin K2e expression appears to begin in the deepest suprabasal layer (F). In penile epidermis K2e shows sporadic and patchy expression (G) whereas in foot arch epidermis K2e is restricted to layers immediately beneath the stratum corneum (H). Original magnifications: ×115 (A–F); ×57 (G, H).

Figure 3.

Expression of keratin polypeptides in cutaneous hyperproliferative lesions. Sections from frozen psoriatic, HTS, and keloid scar epidermis were stained with different keratin and Ki67 antibodies. In psoriatic epidermis, K1 (A) and K16 (B) show homogeneous distribution in the suprabasal compartment whereas K6 is heterogeneously expressed (C) and K2e is either down-regulated (D) or heterogeneously expressed with little or no expression above dermal papillae (E). Increased proliferation compared to normal epidermis was demonstrated by Ki67 staining. F: In HTS epidermis the expression of K1 (G) and K2e (H) was uniformly distributed throughout the suprabasal compartment, K16 is heterogeneously expressed (I), and Ki67 was localized in basal and epibasal layers (J). In keloid scar epidermis keratins K1 (K) and K2e (L) were primarily homogeneously expressed in the suprabasal compartment. Original magnifications: ×78 (A–E); ×155 (F–L).

Figure 4.

Expression of keratins K1, K10, and K2e in normal oral epithelia. Histological appearance of essentially normal gingiva showing parakeratinization (A). In adjacent sections, expression of K1 (B) and K10 (C) is restricted to the upper suprabasal compartment whereas K2e is absent (D). In the ventral surface of tongue, transcript for K2e (E) shows a much wider suprabasal distribution than its protein, which is localized to isolated cells (F). Proteins for K1 (G) and K10 (H) are present in columns of keratinocytes over the connective tissue papillae, although K1 is more widely distributed. Original magnifications, ×50.

Figure 5.

Expression of keratins K1, K10, and K2e in hyperkeratotic lesions and oral lichen planus. In hyperkeratotic lesions, K1 is homogeneously distributed throughout the suprabasal layers (A) whereas expression of K10 is restricted to cells over connective tissue papillae (B) and mRNA for K2e is uniformly expressed in the basal and lower suprabasal layers (C), K2e protein was expressed mostly in cells underneath the orthokeratinized layer (D). In an active area (lymphocytes in epithelium, apoptosis, atrophy; visible only at higher magnification) of oral lichen planus, K1 protein is sporadically expressed in the suprabasal compartment (E) whereas K2e was absent (F). In inactive areas (minimal lymphocytic infiltration and apoptosis, epithelium normal or acanthotic), K1 shows strong but heterogeneous suprabasal distribution (G) whereas K10 is uniformly expressed in the same compartment (H). Original magnifications: ×25 (A, B); ×50 (C–H).

Figure 6.

Expression of keratins K1, K10, and K2e in oral dysplasia. In mild oral dysplasia, K1 is homogeneously expressed in suprabasal layers (A) whereas K10 shows patchy expression mostly as columns of keratinocytes located over connective tissue papillae (B), K2e mRNA is uniformly expressed in the basal and lower suprabasal layers (C) in the presence of little or none of its protein (D). In moderate dysplasia, protein expression of K1 was weak and heterogeneous in parakeratinized region (E) but in the adjacent orthokeratinized region K1 expression was strong and homogeneous (F). K2e mRNA expression was detected in the para- and orthokeratinized regions (G) but protein expression was weaker in the parakeratinized region (H) compared with the orthokeratinized (I). In moderate/severe dysplasia, K1 was mostly localized in the upper suprabasal layers (J) and K2e mRNA was detectable in basal and suprabasal layers (K) but the protein was only expressed focally in the upper suprabasal keratinocytes (L). Original magnifications: ×160 (A–I); ×80 (J–L).

Figure 7.

Expression of keratin polypeptides in oral SCCs. In a well-differentiated oral SCC protein expression for K1 (A) and K10 (B, C) was detected within tumor islands. The K2e expression in these samples was weak or undetectable (D, E) although mRNA was still transcribed in most cells of the tumor islands (F). Original magnifications, ×50.