Carcinoma-like vascular density in atypic keratoacanthoma suggests malignant progression

Abstract

Differential diagnosis between keratoacanthomas and well differentiated squamous cell carcinomas based on clinical and histomorphological data is problematic. Recent findings of cellular atypia in a large proportion of keratoacanthomas indicated that these potentially 'self-healing' cutaneous neoplasms had the potential for malignant progression. Another malignancy-associated criterion is enhanced angiogenesis with increased microvessel density. To provide further diagnostic markers for keratoacanthomas we examined microvessel density on paraffin sections of 13 keratoacanthomas in comparison with 10 normal skin biopsies and 16 late-stage skin squamous cell carcinomas by counting and by computer-assisted image analysis of CD31-immunostained vessels. A significant increase of microvessel density in 'hot spots' was observed in keratoacanthomas as compared to normal skin. Furthermore, when keratoacanthomas were subdivided into tumours with and without malignancy-associated atypic areas, only those with atypia (n=6) were significantly better vascularised than normal skin and had a mean microvessel density in the range of late-stage squamous cell carcinomas. Both keratoacanthoma subtypes revealed comparable levels of inflammatory cell infiltration, tumour cell proliferation and vascular endothelial growth factor expression (mRNA and protein). Thus, in addition to malignancy-associated cellular atypia, increased microvessel density may serve as further diagnostic parameter to discriminate keratoacanthomas with a potential to progress to malignancy.

Figure 1

Two subtypes of keratoacanthoma: (A) nKA revealing neither cellular nor nuclear atypia (H&E). (B) mKA showing prominent cellular and nuclear atypia of tumour cells (H&E). Immunohistochemical staining of microvessels using CD31-antibodies in KA. (C) nKA (counterstaining modified Masson-Goldner). (D) mKA, ‘hot spot’ in frame (t: tumour). (E) Endothelial staining in detail. VEGF expression: (F) VEGF protein localisation by immunohistochemistry in KA revealed a distribution with preference to differentiating cells. (G) Expression of VEGF-RNA revealed by radioactive in situ hybridisation in KA. VEGF anti-sense probe (dark field). (H) Corresponding sense probe control (dark field). Bar: 30 μm.

Figure 2

Linear regression of the two vascular density parameters: manual count vs % area.

Figure 3

Vessel density in normal skin and KA and the subtypes of KA: nKA, mKA compared to late-stage SCCs (# P<0.001 significant vs skin, § P<0.05 significant vs skin using all pairwise multiple comparison).