Distribution of the Human Intracellular Serpin Protease Inhibitor 8 in Human Tissues

Abstract

S U M M A R YOvalbumin-like serine protease inhibitors are mainly localized intracellularly and their in vivo functions are largely unknown. To elucidate their physiological role(s), we studied the expression of one of these inhibitors, protease inhibitor 8 (PI-8), in normal human tissues by immunohistochemistry using a PI-8-specific monoclonal antibody. PI-8 was strongly expressed in the nuclei of squamous epithelium of mouth, pharynx, esophagus, and epidermis, and by the epithelial layer of skin appendages, particularly by more differentiated epithelial cells. PI-8 was also expressed by monocytes and by neuroendocrine cells in the pituitary gland, pancreas, and digestive tract. Monocytes showed nuclear and cytoplasmic localization of PI-8, whereas neuroendocrine cells showed only cytoplasmic staining. In vitro nuclear localization of PI-8 was confirmed by confocal analysis using serpin-transfected HeLa cells. Furthermore, mutation of the P 1 residue did not affect the subcellular distribution pattern of PI-8, indicating that its nuclear localization is independent of the interaction with its target protease. We conclude that PI-8 has a unique distribution pattern in human tissues compared to the distribution patterns of other intracellular serpins. Additional studies must be performed to elucidate its physiological role.

Intrahepatic Lymphocyte Expression of Dipeptidyl Peptidase I-Processed Granzyme B and Perforin Induces Hepatocyte Expression of Serine Proteinase Inhibitor 6 (Serpinb9/SPI-6)

“…mRNA levels of two additional serpinB9 homologs, serpin9g and serpin9e, remained unaltered even in recipients of very high doses of IFN-␣ (data not shown). mRNA expression of constitutively expressed genes, such as cyclophilin and SPI-8, a serpin previously observed to be expressed in normal liver (21,32), also remained unaltered over a broad range of IFN-␣ dosing.…”

Intracellular serpin SERPINB6 (PI6) is abundantly expressed by human mast cells and forms complexes with β-tryptase monomers

“…The human mast cell line-1 (HMC-1; a kind gift from Dr J. Butterfield, Mayo Clinic, Rochester, MN) was cultured as originally described. 15 Cell lysates were prepared as described previously 16 and were stored at Ϫ80°C until use. Protein concentration was measured using the Bradford analysis (Bio-Rad Laboratories, Munchen, Germany) according to the instructions of the manufacturer.…”

“…16,17,19 For detection of SERPINB6, mast cell tryptase, or CD117, mAb PI6-18 (IgG1, 5g/mL), mAb AA1 (IgG1, 1:200), or pAb anti-CD117 (c-kit, 1:50; DAKO) were used, respectively. Tissue sections were subjected to antigen retrieval by boiling in 10 mM sodium citrate buffer, pH 6, for 10 minutes in a microwave oven.…”

“…Of all antibodies tested, mAb PI6-25 displayed optimal reactivity with SERPINB6 during Western blotting and did not cross-react with SERPINB8 or SERPINB9, expressed by transformed Jurkat cells ( Figure 1C) or with recombinant proteins (results not shown). Expression of SERPINB8 or SERPINB9 in the Jurkat cells was assessed with specific antibodies against these proteins, as previously published 16,17 (results not shown).…”

Regulation of vitamin D receptor function in MEN1-related parathyroid adenomas

“…All tissues were sampled from surgical specimens within 2 hours after resection. Immunohistochemical staining was essentially performed as previously described using rabbit polyclonal antibodies against menin (dilution 1:100), against H3K4me3 (1:7,500) and against histone H3 (1:15,000) (Strik et al, 2002). The optimal dilution for each antibody was determined by end-point titrations on tissue slides of human tonsil.…”