The term nonmelanoma skin cancer (NMSC) includes squamous cell carcinoma (SCC) and basal cell carcinoma (BCC) 1. Many factors contribute toward the increased incidence of NMSC, and these are mainly linked to activation of proto-oncogenes and inhibition of tumor-suppressor genes 2.
Programmed cell death protein 4 (PDCD4) is a tumor-suppressor gene that was identified to be upregulated in apoptosis 3,4. It influences transcription, translation, and signal transduction pathways in a way that suppresses tumorous growth 5. Experimental studies showed that PDCD4 could suppress the transformation of mouse epidermal cells exposed to different carcinogenesis promoters 6,7. Accordingly, its high expression in tumors was associated with less aggressiveness and a better prognosis 8.
As limited work has been carried out on the expression of PDCD4 in NMSC, the current study aimed to study its expression in BCC and SCC in comparison with psoriasis (benign proliferative skin disease) and normal skin.
Patients and methods
The study was approved by the Dermatology Department Research Ethical Committee. Deparaffinized sections of 3 µm cut from routinely fixed and paraffin-embedded specimens of BCC (nine cases), SCC (five cases, moderately differentiated), psoriasis vulgaris (10 cases), and normal skin (six biopsies) were stained immunohistochemically with rabbit antihuman PDCD4 monoclonal antibody (EPR3431) (Novus Biologicals, Littleton, Colorado, USA) using the avidin–biotin complex peroxidase staining technique 9. The intensity and pattern of staining were assessed. In addition, a morphometric analysis was carried out using the Leica Qwin 500 Image Analyzer (LEICA Imaging Systems Ltd, Cambridge, UK), which consists of a Leica DM-LB microscope with a JVC color video camera attached to a computer system. Marker color was first detected, followed by the formation of a binary image for the area stained by the marker. The mean area percent of PDCD4 staining was estimated for each studied case.
Photomicrographs depicting the various findings were obtained using a Canon Power Shot G15 digital camera (Canon, New York, New York, USA), connected to a Primo Star Zeiss Microscope (Zeiss, Oberkochen, Germany). All photomicrographs presented are according to their original magnification.
Apart from strong focal nuclear positivity in the basaloid cells in three (33.3%) BCC cases (Fig. 1), all BCC cases showed negative PDCD4 expression in the tumor islands (Fig. 2). Well-differentiated areas of SCC showed moderately positive (mainly cytoplasmic) staining of PDCD4 (Fig. 3), whereas undifferentiated areas were negative (Fig. 4). In psoriasis, moderate predominantly suprabasal PDCD4 cytoplasmic positivity was detected in eight (80%) cases (Fig. 5). This was diffuse in three cases, patchy in three cases, and focal in two cases. Negative expression was detected in two (20%) psoriatic cases. All normal skin biopsies showed epidermal nuclear staining of PDCD4 mainly in suprabasal layers (Fig. 6). Staining was also strongly detected in sebaceous lobules, ductal, and secretory portions of sweat glands. In all specimens, staining of inflammatory cells, fibroblasts, sebaceous lobules, and sweat gland structures was considered a positive internal control.
Table 1 shows the results of morphometric analysis of PDCD4 staining in all studied specimens. No significant difference was found in PDCD4 staining in BCC, SCC, and psoriasis compared with normal skin (P=0.1, 0.4, and 0.9, respectively). Also, no significant difference was found on comparing BCC and SCC with psoriasis (P=0.25 and 0.4, respectively).
In agreement with Matsuhashi et al.10, almost no PDCD4 expression was detected in BCC cases. Focal nuclear positivity was detected only in three cases. Interestingly, this focal nuclear positivity was detected in areas of the tumor showing scattered apoptotic bodies on hematoxylin and eosin examination, providing more evidence of the role of PDCD4 in promoting apoptosis of epithelial cells.
To date, PDCD4 expression in cutaneous SCC has not been studied previously. However, its expression has been studied in laryngeal SCC 11, esophageal SCC 12, oral SCC 13, and in lung cancer including SCC 8. Similar to our findings, these studies reported loss of PDCD4 expression in high-grade undifferentiated areas (cases) of SCC. Because PDCD4 contributes toward the differentiation of keratinocytes by inhibiting AP-1 activities 7,14 and activating caspase-mediated pathways 10,15, well-differentiated areas of SCC showed evident PDCD4 expression.
Suprabasal cytoplasmic expression of PDCD4 was detected in 80% of psoriasis cases. It is speculated that this cytoplasmic upregulation of PDCD4 in psoriatic skin represents an additional mechanism, together with P16-induced senescence of keratinocytes 16, in preventing neoplastic transformation in this highly proliferative skin disorder.
Suprabasal nuclear expression of PDCD4 in all normal skin biopsies in the current study supports previous results that reported its expression in differentiating (suprabasal) layers of the skin and hair follicles (suprabulbar) and its absence or reduced expression in the proliferating basal cell layer and in the hair bulb 10.
Although normal skin showed nuclear staining of PDCD4 in differentiating suprabasal layers, well-differentiated areas of SCC and psoriatic epidermis showed cytoplasmic staining. In most studies on cancer cell lines, cytoplasmic expression of PDCD4 was detected 17. This may reflect a reaction of the keratinocytes to changes in its microenvironment both in psoriasis and in SCC, with subsequent increased activity of differentiating pathways and altered expression patterns.
This study provides preliminary support for a potential role of PDCD4 in tumor suppression in NMSC. Therapeutic approaches that ultimately lead to upregulation of PDCD4 expression in tumor cells with subsequent promotion of tumor cell apoptosis may represent a new approach in NMSC therapy. Further studies on larger number of cases are still needed to validate these results.
The authors would like to thank all residents in the Dermatology and Surgery Departments at Cairo University and National Research Centre for helping us with the recruitment of cases.
Conflicts of interest
There are no conflicts of interest.
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