Immunohistochemical demonstration of keratins in the epidermal layers of the Malayan pangolin (Manis javanica), with remarks on the evolution of the integumental scale armour

Abstract

Using immunohistochemistry, the study demonstrates the distribution of keratins (pan-keratin with CK1-8, 10, 14-16, 19; keratins CK1, 5, 6, 9, 10; hair keratins AE13, AE14) in the epidermis of the Malayan pangolin (Manis javanica). A varying reaction spectrum was observed for pan-keratin, with body region-dependent negative to very strong reaction intensities. The dorsolateral epidermis exhibited positive reactions only in its vital layers, whereas the abdominal epidermis showed strong positive reactions in the soft two outer strata. The single acidic and basic-to-neutral (cyto)keratins produced clear variations compared to the pan-keratin tinging. E.g., CK1 appeared in all epidermal layers of both body regions, except for the ventral stratum corneum, whereas CK5, 6, 9, 10 were restricted to the soft ventral epidermis. Here, distinctly positive reactions were confined to the stratum granulosum, except for CK6 that appeared in the soft stratum corneum. A different staining pattern was obvious for the hair keratins, i.e., positive reactions of AE13 concentrated only in the granular layer of the dorsal epidermis. In the abdominal epidermis, remarkable tinging for AE14 was visible in the stratum basale, decreasing toward the corneal layer, but was also found in the outer root sheath cells of the hair follicles in the ventral body part. Our findings are discussed related to the evolution of the horny dorsal scales of the pangolin, which may have started from the tail root, projecting forward to the head.

Figure 1.

View of the dorsolateral body region (a) with its hard horny scales showing a polished surface, and of the abdominal body region (b) with a dense coat of fine hairs, shown near the anogenital region of the Malayan pangolin (Manis javanica).

Figure 2.

Demonstration of a standard spectrum of epithelial (cyto)keratins in the epidermis of the Malayan pangolin; reactions of pan-keratin (a) in the dorsolateral scale region and (b) in the scale-free ventral body region; reactions of CK 1 are obvious (c) in the scale hinge part of the dorsal body region, and (d) in the ventral body region; strong staining is best visible in the vital epidermis; DAB visualization. SB, stratum basale; SS, stratum spinosum; SG, stratum granulosum; oSC, outer stratum corneum.).

Figure 3.

Demonstration of four specific epithelial (cyto)keratins in the soft abdominal epidermis of the Malayan pangolin; CK5 in (a), CK6 in (b), CK9 in (c), and CK10 in (d); it is shown that in the epidermis layers of this body part the reactions are rather weak to medium; DAB visualization. SB, stratum basale; SS, stratum spinosum; SG, stratum granulosum; SC, stratum corneum).

Figure 4.

Demonstration of hard hair keratins in the epidermis and dermis of the Malayan pangolin; distinctly positive reactions for AE13 are found in (a) the layers of the vital epidermis of a scale hinge region, particularly the stratum granulosum, for AE14 in (b) the ventral body region, but somewhat restricted to the stratum basale of the ventral body region; moreover, positive staining for AE14 also concentrated (c) in the outer root sheath of hair follicles; DAB visualization. SB, stratum basale; SS, stratum spinosum; SG, stratum granulosum; SC, stratum corneum; oSC, outer stratum corneum; oRS, outer root sheath; iRS, inner root sheath; D, dermis.