Expression of serine protease inhibitor 3 in ocular tissues in endotoxin-induced uveitis in rat.
OBJECTIVE
To ascribe the serine protease inhibitor 3 (SPI-3) as an ocular acute inflammatory molecule and to clarify its producing cells in an endotoxin-induced uveitis (EIU) model.


METHODS
Male Wistar rats were injected with lipopolysaccharide (LPS), and the expression of SPI-3 mRNA in the ocular tissues was examined by in situ hybridization (ISH) and Northern blot analysis. A combination of ISH and immunohistochemistry (IHC) were performed to prove the colocalization of SPI-3 mRNA and either glial fibrillary acidic protein (GFAP) or OX-42. The expression of phosphorylated STAT3 (pSTAT3) was demonstrated by IHC and Western blot after LPS injection. The colocalization of SPI-3 mRNA and pSTAT3 was finally examined by the double labeling of ISH and IHC.


RESULTS
After LPS injection, the expression of SPI-3 mRNA in ocular tissues was quickly upregulated and reached a peak between 12 and 24 hours after injection. An intense mRNA signal was observed in epithelial cells of the iris and ciliary body and the innermost retinal layer. In the retina, SPI-3 mRNA was colocalized with GFAP, demonstrating that the cells expressing SPI-3 mRNA were astrocytes. After LPS treatment, SPI-3 mRNA and pSTAT3 were colocalized in retinal astrocytes, and pSTAT3 expression appeared slightly earlier than that of SPI-3 mRNA.


CONCLUSIONS
Ocular inflammation induced the transient expression of SPI-3 mRNA in retinal astrocytes and epithelial cells in the iris and ciliary body, particularly during early phase of the inflammation. Simultaneously, the activation of STAT3 (phosphorylation of STAT3) occurred slightly earlier in astrocytes. This supports the previous in vitro results that SPI-3 expression is induced in a STAT3-mediated manner. SPI-3 may have some crucial roles in preventing some degenerative proteolysis, which is induced by inflammatory stimuli.
Takamiya
A
A
Department of Anatomy, Asahikawa Medical College, Asahikawa, Japan.
Takeda
M
M
Yoshida
A
A
Kiyama
H
H
D000209:Acute-Phase Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism; D000818:Animals; D015703:Antigens, CD; D000951:Antigens, Neoplasm; D000954:Antigens, Surface; D001253:Astrocytes / Q000201:enzymology; D030161:Avian Proteins; D051926:Basigin; D001798:Blood Proteins; D015152:Blotting, Northern; D002924:Ciliary Body / Q000201:enzymology; D004268:DNA-Binding Proteins / Q000378:metabolism; D004847:Epithelial Cells / Q000201:enzymology; D005123:Eye / Q000201:enzymology* / Q000473:pathology; D005136:Eye Proteins / Q000378:metabolism; D005904:Glial Fibrillary Acidic Protein / Q000378:metabolism; D007124:Immunoenzyme Techniques; D017403:In Situ Hybridization; D007498:Iris / Q000201:enzymology; D008070:Lipopolysaccharides; D008297:Male; D008562:Membrane Glycoproteins / Q000378:metabolism; D012333:RNA, Messenger / Q000096:biosynthesis; D051381:Rats; D017208:Rats, Wistar; D012160:Retina / Q000201:enzymology; D050796:STAT3 Transcription Factor; D012486:Salmonella typhimurium; D015842:Serine Proteinase Inhibitors / Q000096:biosynthesis / Q000235:genetics*; D015843:Serpins; D015534:Trans-Activators / Q000378:metabolism; D014605:Uveitis / Q000201:enzymology* / Q000473:pathology
D000209:Acute-Phase Proteins; D015703:Antigens, CD; D000951:Antigens, Neoplasm; D000954:Antigens, Surface; D030161:Avian Proteins; D001798:Blood Proteins; C102624:Bsg protein, Gallus gallus; C079432:Bsg protein, rat; D004268:DNA-Binding Proteins; D005136:Eye Proteins; D005904:Glial Fibrillary Acidic Protein; D008070:Lipopolysaccharides; D008562:Membrane Glycoproteins; D012333:RNA, Messenger; D050796:STAT3 Transcription Factor; D015842:Serine Proteinase Inhibitors; C403317:Serpina3n protein, rat; D015843:Serpins; C494088:Stat3 protein, rat; D015534:Trans-Activators; D051926:Basigin