Array comparative genomic hybridization of keratoacanthomas and squamous cell carcinomas: different patterns of genetic aberrations suggest two distinct entities

Keratoacanthoma (KA) is a benign keratinocytic neoplasm that spontaneously regresses after 3-6 months and shares features with squamous cell carcinomas (SCCs). Furthermore, there are reports of KAs that have metastasized, invoking the question of whether KA is a variant of SCC (Hodak et al., 1993). To date, no reported criteria are sensitive enough to discriminate reliably between KA and SCC, and consequently there is a clinical need for discriminating markers. Our previous study analyzed 132 KAs and 29 SCCs and revealed significantly different regions of genomic aberrations using chromosomal comparative genomic hybridization (CGH). In the present study, we applied array CGH to investigate 98 KAs and 22 SCCs from the above samples. The result shows that all KAs and SCCs have some degree of genetic aberrations. The distribution of numbers of aberrant clones per sample differed significantly between KAs and SCCs (P<0.02), which also demonstrated recurrent aberrations that differed significantly (P<0.001), as illustrated by unsupervised cluster analysis. Classifiers for clinicopathological parameters of KAs were established based on t-test statistics and permutation tests. Tumor size, fibrosis, and inflammation, which are related to the developmental stages of KAs, showed significant (t-test, permutation test) associations with aberrations of selected genomic regions. This suggests chromosomal instability during the whole life cycle of KAs.     

周期素A在皮肤鳞状细胞癌与角化棘皮瘤中的检测

目的通过检测周期素(cyc lin)A在皮肤鳞状细胞癌(SCC)和角化棘皮瘤(KA)中的分布,探讨两者之间的关系。方法采用免疫组化技术检测12例KA和20例SCC皮损中cyc lin A蛋白。结果cyc lin A阳性细胞在KA中主要位于肿瘤的周边部,而在SCC中,呈弥漫性分布;cyc lin A在KA中的平均阳性率(13.2%)与高分化SCC(15.5%)差异无显著性(P>0.05),但与中、低分化SCC(23.4%,33.6%)差异有显著性(P<0.05)。结论cyc lin A在KA与SCC中的分布特点进一步说明了两者的异同,KA是否是SCC的分型尚需进一步探讨;cyc lin A在KA及各型SCC中的阳性率的不同说明cyc lin A可以作为评价皮肤肿瘤增殖程度的一个分子生物学指标。     

The potential role of abnormal E-cadherin and α-, β- and γ-catenin immunoreactivity in the determination of the biological behaviour of keratoacanthoma

BACKGROUND: Failure of E-cadherin and its associated proteins alpha-, beta- and gamma-catenin is believed to lead to disruption of cell-cell adhesion and to contribute to neoplasia. OBJECTIVES: To determine the pattern of E-cadherin and alpha-, beta- and gamma-catenin immunostaining in keratoacanthoma (KA) and to evaluate its potential value in routine histopathology in differentiating KA with benign from that with malignant biological behaviour. METHODS: We examined the expression of E-cadherin and alpha-, beta- and gamma-catenin in KA and correlated the histopathological features with the immunohistochemical findings. Next, we compared the immunohistochemical findings of KA with those found in malignant (squamous cell carcinoma, SCC) and benign (warts) lesions. In addition to the established histopathological criteria we used the Ki-67 index, a well-known marker of cell proliferation. Immunoperoxidase staining of E-cadherin and alpha-, beta- and gamma-catenin, and Ki-67 determination, were performed in paraffin-embedded sections of 12 KAs taken from archival material. On reviewing the histology, seven of the 12 KAs were characterized as 'classical' KA, and the rest as 'borderline' KA or KA resembling SCC. Additionally, 28 well, nine moderately and five poorly differentiated SCCs and 20 warts were examined. RESULTS: Most 'classical' KAs (79-86%) showed normal membranous immunostaining and a low Ki-67 index. The remaining 'classical' KAs showed abnormal expression, in a staining pattern resembling that of well-differentiated SCC. All 'borderline' KAs showed a high Ki-67 index (> 40%) and abnormal expression of the adhesion molecules studied, identical to that of poorly differentiated SCC. Expression of E-cadherin and alpha-, beta- and gamma-catenin was found to be more frequently abnormal in 'borderline' KA compared with that in 'classical' KA (P < 0.05). Among E-cadherin and alpha-, beta- and gamma-catenin expression and Ki-67 index, only the expression of beta-catenin was more frequently found to be abnormal in total SCC than in total KA (P < 0.05). Expression of E-cadherin and alpha-, beta- and gamma-catenin was more frequently found to be abnormal in well-differentiated SCC than in 'classical' KA (P < 0.05). In total, as well as in 'classical' or 'borderline' KA, an agreement between expression of E-cadherin and of catenins was seen. CONCLUSIONS: These findings suggest that E-cadherin and catenins may be very helpful in distinguishing between 'classical' and 'borderline' KA, as the expression of these adhesion molecules in 'classical' KA is identical to that found in normal epidermis, overlapping with well-differentiated SCC in some cases. In 'borderline' KA, expression of adhesion molecules is identical to that in poorly differentiated SCC.     

Are keratoacanthomas variants of squamous cell carcinomas? A comparison of chromosomal aberrations by comparative genomic hybridization

Keratoacanthoma (KA) is a benign keratinocytic neoplasm that usually presents as a solitary nodule on sun-exposed areas, develops within 6-8 weeks and spontaneously regresses after 3-6 months. KAs share features such as infiltration and cytological atypia with squamous cell carcinomas (SCCs). Furthermore, there are reports of KAs that have metastasized, invoking the question of whether or not KA is a variant of SCC. To date no reported criteria are sensitive enough to discriminate reliably between KA and SCC, and consequently there is a clinical need for discriminating markers. We screened fresh frozen material from 132 KAs and 37 SCCs for gross chromosomal aberrations by using comparative genomic hybridization (CGH). Forty-nine KAs (37.1%) and 31 SCCs (83.7%) showed genomic aberrations, indicating a higher degree of chromosomal instability in SCCs. Gains of chromosomal material from 1p, 14q, 16q, 20q, and losses from 4p were seen significantly more frequently in SCCs compared with KAs (P-values 0.0033, 0.0198, 0.0301, 0.0017, and 0.0070), whereas loss from 9p was seen significantly more frequently in KAs (P-value 0.0434). The patterns of recurrent aberrations were also different in the two types of neoplasms, pointing to different genetic mechanisms involved in their developments.     

In situ characterization of lymphocytic immunophenotypes and interleukin-2 receptors in cutaneous tuberculosis and leprosy–a comparative evaluation

The granulomas of lupus vulgaris (LV) were characterized by preponderance of CD4⁺ lymphocytes and a raised CD4⁺/CD8⁺ ratio. In contrast, in scrofuloderma (SF) the CD⁸ T-lymphocyte subpopulation predominated and the CD4⁺/CD8⁺ ratio was significantly decreased. A higher percentage of lymphocytes expressed interleukin-2 receptor (IL-2R) in LV as compared with SF, indicating an activated cellular immune response in the former. Immunophenotypic changes in tuberculosis verrucosa cutis {TBVC) were intermediate between LV and SF. CD4⁺ lymphocytes were the main infiltrating T-cell type in borderline tuberculoid leprosy (BT), while CD8⁺ lymphocytes predominated in the granuloma of lepromatous lepromatous (LL). The CD4⁺/CD8⁺ ratio and percentage of lymphocytes expressing IL-2R was significantly higher in BT as compared with LL. These immunophenotypic findings suggest that in both cutaneous tuberculosis and leprosy there is a continuous spectrum with regard to cell-mediated immunity depending on the clinical presentation.     

Adhesion molecules and IL-1 costimulate T lymphocytes in the autologous MECLR in psoriasis

Membrane molecules such as CD36 (OKM5), intercellular adhesion molecule-1 (ICAM-1, CD54), gamma interferon-induced protein 10 (γ-IP10) and IL-1 are induced and/or upregulated in psoriatic epidermis. These molecules have important accessory, trafficking or signalling functions in the immune system and also play a role in the pathophysiology of psoriasis. The relevance of adhesion molecules, CD36 and epidermal IL-1 in psoriasis was studied in vitro in the autologous mixed epidermal cell-T lymphocyte reaction (MECLR). Their level of expression was quantitated in epidermal cell suspensions (ECS) from patients with psoriasis and their function was assessed by blocking with specific mAbs and antisera or by depleting CD36⁺ cells from the ECS prior to the MECLR. ECS from psoriatic lesions contained increased numbers of CD36⁺ (23±12%), ICAM-1⁺ (31±14%) and IL-1⁺ (57±21%) cells. The autologous MECLR was inhibited in saaples from all patients by mAb to CD2 (LFA-2), CD11a (LFA-1α), CD18 (LFA-1β), ICAM-1, CD58 (LFA-3) and an antiserum to IL-1β. Thus, adhesion molecules facilitate inflammation in psoriasis not only via adhesion and recruitment of T lymphocyte in psoriatic lesions, but also via activation of T cells. Furthermore CD36 molecules on psoriatic epidermal cells do not costimulate autologous T lymphocytes in psoriasis. The observed costimulatory function of IL-1β in the MECLR emphasizes its relevance in psoriasis.     

T cells migrate to tumour sites after extracorporeal interleukin 2 stimulation and reinfusion in a patient with metastatic melanoma

Peripheral blood mononuclear cells (PBMC) were taken by leukapheresis from a patient with melanoma skin metastases and stimulated in vitro using 1000 IU recombinant interleukin 2 (IL-2)/ml to generate lymphokine-activated killer cells (LAK cells). Two-colour immunofluorescence analysis demonstrated an IL-2-induced up-regulation of CD25 on natural killer cells (CD56⁺) as well as on T lymphocytes (CD3⁺). After radiolabelling with indium-111, the cells were reinfuse. Gamma-camera imaging revealed an enrichment at the tumour sites. Immunostaining of tumour tissue taken before and after scintigraphy demonstrated CD25⁺ Tlymphocytes (CD2⁺, CD3⁺), but no natural killer cells (CD16⁺, CD56⁺) infiltrating the metastases. LAK cell enrichment at melanoma metastases in vivo did not involve natural killer cells, but was characterized by increased numbers of activated T lymphocytes in this patient.     

Comparison of mitotic cyclins and cyclin-dependent kinase expression in keratoacanthoma and squamous cell carcinoma

Disruption of the cell-cycle regulation through over-expression or mutation of cyclins and cyclin-dependent kinases has been implicated in carcinogenesis. In order to determine whether keratoacanthoma (KA) is unique or a variant of squamous cell carcinoma (SCC) and whether expression of mitosis-related antigens are associated with KAs' tendency to regress, we compared the immunohistochemical expression of mitotic cyclins (cyclins A and B) and their cyclin-dependent kinase p34(cdc2) in 21 KAs, 8 regressing KAs, and 28 conventional squamous cell carcinomas. KAs showed both overlap and significant differences in expression of these mitosis-related antigens compared to SCCs. Basal and parabasal pattern of expression of cyclins A and B significantly predominated in KAs in contrast to SCCs which exhibited diffuse pattern (cyclin A 86%/cyclin B 64% vs. 25%/36%, p < 0.01). However, no differences in the highest mean level of expression in 'hot spot' loci of cyclins A and B were identified comparing KAs to SCCs (19%/12% vs. 25%/13%, p > 0.05). For the cyclin-dependent kinase p34(cdc2), no differences in pattern, distribution or mean levels of expression were found. For cyclins A and B, regressing KA showed significantly more regional tumor labeling (88%/88% vs. 57%/33%, p = 0.03) and a lower mean level of immunoreactivity (5%/4% vs. 19%/12%, p = 0.001) compared to mature KAs. These findings indicate a role for mitotic cyclins in the evolution of both SCC and KA. The overlapping patterns of expression for these mitosis-related antigens suggest that KAs represent a variant of SCC that exhibit an overwhelming but not absolute tendency to involute.     

角化棘皮瘤皮损CD4 CD8和细胞凋亡的检测

目的探讨角化棘皮瘤(keratoacanthoma,KA)皮损中T淋巴细胞与细胞凋亡及其意义。方法应用免疫组化技术检测皮损部位CD4,CD8,CD56,S100蛋白;用末端脱氧核糖核酸转移酶(TdT)介导的dUTP生物素缺口末端标记技术(TUNEL),原位检测28例角化棘皮瘤皮损中凋亡细胞。结果28例KA中,CD4,CD8,CD56,S100蛋白的阳性率分别为85.71%,92.86%,82.14%和67.86%。CD4阳性率低于CD8,但无显著性差异(t=1.54,P>0.05)。46.43%KA(13/28)瘤中心出现凋亡细胞。KA凋亡率(21.89%)与CD8阳性率呈显著正相关(r=0.904,P<0.001)。结论淋巴细胞介导的细胞凋亡可能在KA的自行消退中发挥重要作用。     

Analysis of microsatellite instability and loss of heterozygosity in keratoacanthoma

We analyzed microsatellite instability (MSI) and loss of heterozygosity (LOH) at 17 microsatellite markers located on chromosomes 2p, 3p, 5q, 6q, 9p, 9q, 17p and 18q in 19 randomly selected keratoacantomas (KAs), in one cutaneous lesion that histologically could not unequivocally be differentiated from squamous cell carcinoma, and in one patient with multiple KAs of longstanding duration. The goals of our study were to determine whether, in a similar manner to some visceral carcinomas, genomic instability could be detected in KAs and to clarify whether molecular analysis might be useful to further characterize KA. MSI was observed in 2 of 21 cases (9.5%) at 5 of 17 loci examined. In one patient with a solitary KA, the presence of MSI and a family history of visceral malignant tumours suggested that the patient might have belonged to a family with Muir-Torre syndrome. In one other MSI ⁺ KA, a definite differential diagnosis in relation to squamous cell carcinoma could not be established. In addition, one sample displayed LOH at 2 of 17 loci analysed whereas in the patient with multiple KAs, LOH at one locus was the only alteration found. In conclusion, the low frequency of MSI and LOH detected in our study suggests that these genetic events are uncommon in KA unless it is associated with a familial disease (e.g. Muir-Torre syndrome) or it has more aggressive histological features. Received: 12 August 1996     

Phenotypic determination of T-lymphocytes responding to chemotactic stimulation from fMLP,IL-8, human IL-10, and epidermal lymphocyte chemotactic factor

Summary Human T lymphocytes were collected after they had migrated towards N-formyl-methionyl-leukyl-phenylalanine (fMLP), rIL-8, human IL-10 (hIL-10), and epidermal lymphocyte chemotactic factor (ELCF). They were stained for determination of their phenotype by FACS analysis using anti-CD4, -CD8, -CD18, -CD45R0 and OPD4 antibodies. Human IL-10 increased the percentage of CD8⁺ T lymphocytes in the migrating cell population by 152% compared with cells migrating towards the medium and decreased the number of CD4⁺ T lymphocytes by 79%. ELCF increased the number of CD4⁺ T lymphocytes by 18%, and the number of CD45R0⁺ T lymphocytes by 52%, while the number of CD8⁺ T lymphocytes was decreased by 20%. rIL-8 increased the number of CD4⁺ T lymphocytes and decreased the CD8⁺ T lymphocytes. The distribution of the different subpopulations of T lymphocytes was not changed significantly by fMLP. The observed changes in the phenotypes did not occur when incubating T lymphocytes with the chemotaxins. Our observations demonstrate that individual chemotactic factors will attract specific subsets of T lymphocytes. They may help to explain the predominance of memory T lymphocytes (CD4R0⁺, CD4⁺) in allergic contact dermatitis and certain other skin diseases. They also confirm the results of a recent study, that showed hIL-10 to be selectively chemotactic for CD8⁺ T lymphocytes.     

Humanized anti-CD2 monoclonal antibody treatment of plaque psoriasis: efficacy and pharmacodynamic results of two randomized,double-blind,placebo-controlled studies of intravenous and subcutaneous siplizumab

New biologic therapies focused primarily on cytokine pathways, some targeting T cell-mediated immune responses, are being developed for the treatment of psoriasis. Siplizumab is a humanized anti-CD2 monoclonal antibody that interferes with costimulation necessary for T cell activation and proliferation. We assessed the biological activity, serum concentrations, and pharmacodynamic effects of siplizumab in patients with plaque psoriasis. Two multicenter, phase II randomized, double-blind, placebo-controlled studies were conducted: one study randomized 124 patients to one of two intravenous (IV) doses (0.012 and 0.04 mg/kg) of siplizumab, given every 2 weeks × 8 doses; the other study randomized 420 patients to one of three subcutaneous (SC) dose regimens of siplizumab given weekly (5 mg for 12 weeks, 5 mg for 6 weeks, and 7 mg for 4 weeks) or placebo for 12 weeks. Adults with plaque psoriasis involving ≥10% of the body surface area and who were not receiving psoriasis therapy were eligible. Treatment with siplizumab resulted in reductions in psoriasis severity, but most of the effects were not statistically significant compared with placebo. Statistically significant differences among IV siplizumab-treated and placebo groups were observed at study day 28, with greater psoriasis area and severity index (PASI) score reductions from baseline in the siplizumab groups. The difference in PASI50 response rates between the 0.04 mg/kg siplizumab and placebo groups was also statistically significant at day 28. A trend toward clinical improvement was observed in SC siplizumab-treated groups. Significant reductions in circulating absolute lymphocyte counts and CD2⁺ (CD3⁺, CD8⁺, and CD16⁺/56⁺), but not CD2⁻ (CD19⁺ and CD14⁺), lymphocyte populations were observed. These changes were not accompanied by concomitant reductions in infiltrating CD3⁺ lymphocytes in psoriatic lesions, epidermal thickness, or keratin 16 (K16) and intercellular adhesion molecule (ICAM) expression. The effect of siplizumab did not differentially affect CD45RO⁺ and CD45RA⁺ lymphocytes. Low or undetectable mean trough serum concentrations of siplizumab following IV or SC treatment were observed. Pharmacokinetic data coupled with higher-than-expected placebo clinical response rates may partly explain siplizumab’s marginal clinical activity. Higher doses of siplizumab may be required to detect significant improvements in psoriasis; however, further development of this agent was not planned.     

Activation of human vascular endothelium in melanoma metastases induces ICAM‐1 and E‐selectin expression and results in increased infiltration with effector lymphocytes

Lymphocytic infiltration into melanoma tissue is an important prerequisite for effective antitumoral immunity. However, analysis of human metastatic melanoma has shown that leucocyte adhesion receptor expression on melanoma blood vessels is very low or absent, thereby impairing the entry of cytotoxic lymphocytes into tumor tissue. We hypothesized that adhesion molecules can be induced on melanoma vasculature allowing better infiltration of cytotoxic lymphocytes. Quantitative real‐time PCR and immunofluorescence staining indicated that the adhesion molecules ICAM‐1 (CD54) and E‐selectin (CD62E) can be significantly induced by intralesional application of TNF alpha in tissue from human melanoma metastases either in vitro or in vivo when grafted onto immunodeficient NSG (NOD.Cg‐PrkdcscidIl2rgtm1Wjl/SzJ) mice that preserved human vessels. Furthermore, activated human autologous CD3⁺ lymphocytes were injected intravenously into mice bearing melanoma xenografts treated with TNF‐α or PBS in addition to the leucocyte chemoattractant TARC (CCL17). Significantly increased numbers of CD8⁺ cells were detected in TNF‐α–treated melanoma metastases compared with PBS‐treated controls. In addition, tumor cell apoptosis was enhanced and melanoma cell proliferation reduced as shown by TUNEL assay and KI‐67 staining. We conclude that adhesion molecules can be induced on human melanoma vasculature resulting in significantly improved homing of activated autologous cytotoxic T cells to melanoma tissue and inhibition of melanoma cell proliferation. These observations should be considered when designing protocols for immunotherapy of malignant melanoma.     

Keratinocyte and dermal vascular endothelial cell capacities remain unimpaired in the margin of chronic venous ulcer

The role of endogenously produced cytokines and growth factors in the impaired healing of chronic leg ulcers remains uncertain. The aim of this study was to determine the functional capacity of skin cells in ulcer bed tissue compared to those in the edge of ulcers and skin distal to ulcers. Biopsies from leg ulcers of ten randomly selected patients were examined immunohistochemically for cytokines and growth factors produced by keratinocytes (KC) and vascular endothelial cells (EC). The phenotype of leukocytes infiltrating venous ulcers and the expression of vascular adhesion molecules responsible for extravasation were also studied. The expression of cytokines and growth factors by KC was similar in areas adjacent and remote from an ulcer. In the dermis adjacent to an ulcer, the expression of IL-1, IL-1, IL-1Ra, EGF and PDGFa by EC was higher than the levels of expression in EC from the distant dermis. The expression of IL-6, TNF and GM-CSF was comparable to that in cells from intact dermis. For all these factors staining was cytoplasmic, suggesting production in these areas. Ulcer bed tissue contained few fibroblasts and blood capillaries showing a high staining intensity for CD62E and CD106 EC adhesion molecules but no FGF2 expression (P<0.05). The intensity of staining for scavenging CD15⁺elastase⁺ granulocytes and CD35⁺ (C3bR) activated macrophages in the ulcer bed was comparable to that in the margin but higher than that in the distant dermis (P<0.05), whereas staining for CD68⁺, HLA DR⁺, TGF⁺ and CD54⁺ dermal macrophages was similar in all areas. There was reduced staining for CD4⁺ and CD8⁺ cells in the ulcer bed (P<0.05). There were no CD1a⁺ Langerhans cells in the epidermis encroaching upon the granulation tissue and there was reduced CD1a staining in the adjacent epidermis (P<0.05). In conclusion, there is chronic accumulation of scavenging cells with lack of remodeling of the granulation tissue and, at the same time, preserved cytokine and growth factor secretory potential of KC and dermal EC in non-healing venous leg ulcers.     

Spontaneous regression of keratoacanthoma can be promoted by topical treatment with imiquimod cream

Imiquimod, the first member of a new class of immune response modifiers, is approved for the treatment of external genital and perianal warts. Recently, many clinical trials highlighted the potential of imiquimod as a treatment for other viral infections and cutaneous neoplasms. We report two cases of facial keratoacanthomas (KA) treated with topical 5% imiquimod cream. Patients were successfully cleared of KAs after treatment for 8 weeks. No recurrence occurred after a 1-year follow-up. Despite the fact that KAs are characterized by the potential for spontaneous regression, it is possible that a faster activation of CD4+ lymphocytes, via interferon release and cytokine secretion takes place after imiquimod application leading to KA regression.     

Neutrophils infiltrating ultraviolet B-irradiated normal human skin display high IL-10 expression

Exposure to an erythemal dose of ultraviolet B (UVB) is known to induce interleukin (IL-10) expression in human skin. It is generally believed that this IL-10 is predominantly expressed by CD11b⁺HLA-DR⁺ macrophages that infiltrate the UVB-exposed skin. This cytokine is presumed to contribute to the immunosuppressive effects of UVB by inhibiting cell-mediated immune responses. We recently demonstrated that neutrophils, which also invade UVB-irradiated skin, express CD11b and HLA-DR as well. In addition, we showed that the presence of these neutrophils affects T-cell responses in primary T-cell cultures derived from UVB-exposed skin. Since neutrophils invade UVB-exposed skin and, like macrophages, express CD11b and HLA-DR, we sought to determine whether neutrophils represent another source of IL-10. Skin biopsies were obtained from four healthy volunteers before and 2 days after exposure to four minimal erythema doses of UVB. A series of immunohistochemical double-staining procedures using the following markers was performed: IL-10, CD11b, HLA-DR, CD36, neutrophil elastase, and CD66b. As expected IL-10 could be detected in CD11b⁺HLA-DR⁺CD36⁺ macrophages in the epidermis and dermis of UVB-exposed skin. Surprisingly, the majority of the abundant IL-10 expression was found in CD11b⁺HLA-DR⁺elastase⁺CD66b⁺ neutrophils. Cytospin preparations from dermal cell suspensions confirmed the IL-10 expression by neutrophils displaying characteristic multilobular nuclei. Thus, neutrophils in UVB-exposed skin express IL-10 and should be recognized as active coplayers in the creation of the UVB-induced immunosuppressive microenvironment.     

Expression of class II beta-tubulin in non-melanoma cutaneous tumors

BACKGROUND: Different combinations of beta-tubulin isotypes contribute to the diverse functions of microtubules (MTs). Class II beta-tubulin (class II tubulin) is up-regulated in differentiated keratinocytes. In contrast, the expression of class II tubulin in follicular differentiation and cutaneous tumors has not been studied. METHODS: The immunohistochemical expression of class II tubulin was investigated in 117 cutaneous tumors: 30 squamous cell carcinomas (SCCs), seven keratoacanthomas (KAs), 57 basal cell carcinomas (BCCs), 23 trichoepitheliomas (TEs), and in the adjacent non-neoplastic skin. RESULTS: Class II tubulin was expressed in the keratinocytes of the granular layer, melanocytes, hair cortical and cuticular cells, inner root sheath (IRS), companion layer (CL) of the outer root sheath (ORS), and mesenchymal cells with Schwannian or myogenic differentiation. Moreover, class II tubulin expression was increased in the areas of squamous or follicular differentiation in cutaneous tumors. On grading the follicular differentiation or myofibroblastic response with anti-class II tubulin, TE showed follicular differentiation more frequently (p < 0.001) with less of a myofibroblastic response (p = 0.001) than BCC. CONCLUSIONS: Class II tubulin expression is closely related to squamous or follicular differentiation and may be helpful in distinguishing most SCCs from KAs and BCC from TE. However, it does not reliably distinguish well-differentiated, crateriform SCC from KA.