Analysis of interleukin-1beta-modulated mRNA gene transcription in human gingival keratinocytes by epithelia-specific cDNA microarrays.
OBJECTIVE
Proinflammatory cytokines such as interleukin-1beta are known to be synthesized in oral gingivitis and periodontitis and lead to the activation of the transcription factor nuclear factor-kappaB (NF-kappaB). Although numerous effects of interleukin-1beta on mesenchymal cells are known, e.g. up-regulation of intercellular adhesion molecule-1 in endothelial cells, little is known of the effects of interleukin-1beta on oral keratinocytes. The purpose of the present study was to seek interleukin-1beta-mediated alterations in mRNA gene transcription and a putative activation of NF-kappaB in oral gingival keratinocytes.


METHODS
As an in vitro model for gingivitis and periodontitis, immortalized human gingival keratinocytes (IHGK) were stimulated with the proinflammatory cytokine interleukin-1beta. An epithelia-specific cDNA microarray was used to analyze mRNA expression profiles from IHGK cells treated with 200 units interleukin-1beta/ml for 3, 6, 9, 12, and 24 h. Indirect immunofluorescence was carried out to detect NF-kappaB in IHGK following interleukin-1beta treatment.


RESULTS
Detailed analysis revealed distinct patterns of time-dependent changes, including genes induced or repressed early (3-6 h) or late (12-24 h) after interleukin-1beta treatment. Differentially expressed genes were involved in (i) cell stress, (ii) DNA repair, (iii) cell cycle and proliferation, (iv) anti-pathogen response, (v) extracellular matrix turnover, and (vi) angiogenesis. A large number of genes were responsive to NF-kappaB and induction was concomitant with nuclear translocation of the p65 RelA subunit of NF-kappaB. Interestingly, many of these genes contain multiple NF-kappaB binding sites in their promoters.


CONCLUSIONS
Analysis of altered gene expression allows identification of gene networks associated with inflammatory responses. In addition to a number of well-known genes involved in gingivitis and periodontitis, we identified novel candidates that might be associated with the onset and maintenance of an inflammatory disease.
Steinberg
T
T
Department of Orthodontics and Dentofacial Orthopedics, Dental School, University of Heidelberg, Im Neueheimer Feld 400, 69129 Heidelberg, Germany. thorsten_steinberg@med.uni-heidelberg.de
Dannewitz
B
B
Tomakidi
P
P
Hoheisel
J D
JD
Müssig
E
E
Kohl
A
A
Nees
M
M
D018797:Cell Cycle Proteins / Q000096:biosynthesis; D002461:Cell Line, Transformed; D016207:Cytokines / Q000096:biosynthesis; D045643:DNA Repair Enzymes / Q000096:biosynthesis; D016326:Extracellular Matrix Proteins / Q000096:biosynthesis; D019084:Fluorescent Antibody Technique, Indirect; D020869:Gene Expression Profiling; D053263:Gene Regulatory Networks; D005881:Gingiva / Q000166:cytology / Q000378:metabolism*; D005891:Gingivitis / Q000235:genetics* / Q000378:metabolism; D006360:Heat-Shock Proteins / Q000096:biosynthesis; D006801:Humans; D053583:Interleukin-1beta / Q000502:physiology*; D015603:Keratinocytes / Q000378:metabolism*; D020782:Matrix Metalloproteinases / Q000096:biosynthesis; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism*; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods; D011401:Promoter Regions, Genetic; D021381:Protein Transport; D012333:RNA, Messenger / Q000096:biosynthesis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015533:Transcriptional Activation / Q000502:physiology*; D042461:Vascular Endothelial Growth Factor A / Q000096:biosynthesis
D018797:Cell Cycle Proteins; D016207:Cytokines; D016326:Extracellular Matrix Proteins; D006360:Heat-Shock Proteins; D053583:Interleukin-1beta; D016328:NF-kappa B; D012333:RNA, Messenger; D042461:Vascular Endothelial Growth Factor A; D020782:Matrix Metalloproteinases; D045643:DNA Repair Enzymes