Comprehensive analysis of gene expression in the junctional epithelium by laser microdissection and microarray analysis.
OBJECTIVE
The junctional epithelium attaches to the tooth enamel at the dentogingival junction. The attachment mechanisms of the junctional epithelium have been studied histologically, but the molecular functions of the junctional epithelium have not been elucidated. The aim of this study was to perform a comprehensive analysis of gene expression in the junctional epithelium and to search for specific genetic markers of the junctional epithelium.


METHODS
A comprehensive analysis of genes expressed in the mouse junctional epithelium and oral gingival epithelium was performed using laser microdissection and microarray analysis. To extract high-quality RNA from these tissues, we made frozen sections using a modified film method. Confirmation of the differential expression of selected genes was performed by quantitative real-time PCR and immunohistochemistry.


RESULTS
The modified method produced RNA of sufficient quality for microarray analysis. The result of microarray analysis showed that 841 genes were up-regulated in the junctional epithelium compared with the oral gingival epithelium, and five were increased more than 50-fold in the junctional epithelium. These five genes were secretory leukocyte protease inhibitor (Slpi), keratin 17 (Krt17), annexin A1 (Anxa1), myosin light peptide 6 (Myl6) and endoplasmic reticulum protein 29 (Erp29). In particular, Slpi expression in the junctional epithelium was approximately 100-fold higher than in the oral gingival epithelium by real-time PCR. Additionally, immunohistochemistry indicated that the Slpi protein is highly expressed in the junctional epithelium.


CONCLUSIONS
We developed a method for generating fresh-frozen tissue sections suitable for extraction of good-quality RNA. We determined that Slpi is characteristically expressed in the junctional epithelium. Our results provide a substantial advance in the analysis of gene expression in the junctional epithelium.
Hayashi
Y
Y
Department of Periodontology, Showa University School of Dentistry, Ohta-ku, Tokyo, Japan.
Matsunaga
T
T
Yamamoto
G
G
Nishii
K
K
Usui
M
M
Yamamoto
M
M
Tachikawa
T
T
D000818:Animals; D017305:Annexin A1 / Q000096:biosynthesis / Q000235:genetics; D004721:Endoplasmic Reticulum; D004846:Epithelial Attachment / Q000201:enzymology / Q000378:metabolism*; D005629:Frozen Sections; D020869:Gene Expression Profiling / Q000379:methods*; D005881:Gingiva / Q000378:metabolism; D006360:Heat-Shock Proteins / Q000096:biosynthesis / Q000235:genetics; D053537:Keratin-17 / Q000096:biosynthesis / Q000235:genetics; D054020:Lasers, Gas; D051379:Mice; D042282:Microdissection / Q000379:methods; D018994:Myosin Light Chains / Q000096:biosynthesis / Q000235:genetics; D020411:Oligonucleotide Array Sequence Analysis; D053481:Secretory Leukocyte Peptidase Inhibitor / Q000096:biosynthesis* / Q000235:genetics
D017305:Annexin A1; C491737:Erp29 protein, mouse; D006360:Heat-Shock Proteins; D053537:Keratin-17; D018994:Myosin Light Chains; D053481:Secretory Leukocyte Peptidase Inhibitor; C506815:Slpi protein, mouse