Abstract 42 Allergy and Immunology Platform, Monday, 5/3

Squamous cell carcinoma antigens 1 and 2 (SCCA1 and SCCA2) are serine proteinase inhibitors (serpins) that are 95% identical. However, they inhibit distinct classes of proteinases: SCCA1 inhibits papain-like lysosomal cysteine proteinases (cathepsins L, S, and K), whereas SCCA2 inhibits myeloid granule proteinases (cathepsin G and mast cell chymase). To define the tissue-specific and intracellular distribution of these serpins, we first generated a panel of discriminatory monoclonal antibodies. An immunohistochemical survey of normal human tissues showed that SCCA1 and SCCA2 were co-expressed in the suprabasal layers of the stratified squamous epithelia lining mucosal surfaces, the pseudostratified epithelia lining the conducting airways, the infundibulum of hair follicles, and Hassall's corpuscles of the thymus. Since SCCA1 and SCCA2 inhibit proteinases involved in inflammatory reactions, we hypothesized that these serpins are upregulated in response to this type of stimulus. We observed increased SCCA1 and SCCA2 expression in the suprabasal epidermis of benign and malignant inflammatory skin disorders (e.g., psoriasis, eczematous dermatitis, lichen planus, squamous cell carcinoma, inflamed seborrheic keratosis, and bacterial, viral, and fungal infectious diseases), but not in non-inflammatory epidermal disorders (non-inflamed seborrheic keratosis, harlequin ichthyosis, and epidermolysis bullosa simplex).

To identify the subcellular localization of SCCA1 and SCCA2 we examined epithelial-like cells using immunofluorescence, SCCA1/2-GFP fusion proteins, transient transfections, and confocal microscopy. SCCA1 and SCCA2 co-localized to the cytosolic compartment and were not associated with nuclei, mitochondria, lysosomes, microtubules, actin, or the Golgi. This finding was confirmed by subcellular fractionation studies using differential centrifugation. Pulse-chase studies showed that even in the presence of PMA, SCCA1 and SCCA2 were retained within the cytosol and were not secreted appreciably into the medium.

To determine if an inflammatory cytokine could increase SCCA1 and SCCA2 expression directly, epithelial cells were cultured in the presence of TNF-α. TNF-α stimulation increased SCCA1 expression both at the protein and mRNA levels.

Collectively, these data suggest that SCCA1 and SCCA2 are intracellular, cytosolic proteins whose expression is upregulated during inflammation. We hypothesize that these serpins may protect epithelial cell surfaces from exogenous or endogenous proteinase-mediated injury.