The effects of proinflammatory cytokines on cytokine-chemokine gene expression profiles in the human corneal endothelium.
OBJECTIVE
To determine the effects of proinflammatory cytokines on differential gene expression profiles in the human corneal endothelium (HCE), by using a cDNA expression array.


METHODS
A human cDNA expression array technology was used to study the simultaneous expression of HCE incubated with interleukin (IL)-1alpha and tumor necrosis factor-(TNF)-alpha. Gene-specific semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA) were performed to examine the gene and protein expression patterns revealed by the cDNA expression array, in the presence and absence of proinflammatory cytokines. Moreover, the expression of these genes was studied in ex vivo HCE of donor cornea by RT-PCR.


RESULTS
IL-1alpha and TNF-alpha upregulated the expression of 46 of 268 genes for cytokines, chemokines, and their receptors in stimulated HCE. The most upregulated genes in the cDNA expression array, those of monocyte chemotactic protein (MCP)-1 (CCL2), IL-8 (CXCL8), IL-6, and growth-related beta (GRObeta, CXCL2), were studied. Semiquantitative RT-PCR and ELISA analyses revealed the proinflammatory cytokine-mediated changes in the respective gene transcription and protein expression levels. The mRNAs were detected in ex vivo HCE of donor cornea stimulated with proinflammatory cytokines.


CONCLUSIONS
HCE can abundantly express cytokines and chemokines through the stimulation of proinflammatory cytokines. The detected genes, those of CCL2, CXCL8, IL-6, and CXCL2, in HCE could facilitate understanding of the inflammatory responses, including the production of keratic precipitates and the correlation between CE and an inflamed cornea or aqueous humor.
Yamagami
Hiroko
H
Department of Ophthalmology, Omiya Medical Center, Jichi Medical School, Saitama, Japan.
Yamagami
Satoru
S
Inoki
Taeko
T
Amano
Shiro
S
Miyata
Kazunori
K
D002478:Cells, Cultured; D018925:Chemokines / Q000096:biosynthesis / Q000235:genetics*; D016207:Cytokines / Q000096:biosynthesis / Q000235:genetics*; D018076:DNA, Complementary / Q000032:analysis; D004728:Endothelium, Corneal / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D015870:Gene Expression; D020869:Gene Expression Profiling; D006801:Humans; D007375:Interleukin-1 / Q000494:pharmacology*; D020411:Oligonucleotide Array Sequence Analysis; D012333:RNA, Messenger / Q000096:biosynthesis; D019707:Receptors, Chemokine / Q000235:genetics; D018121:Receptors, Cytokine / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*; D015854:Up-Regulation
D018925:Chemokines; D016207:Cytokines; D018076:DNA, Complementary; D007375:Interleukin-1; D012333:RNA, Messenger; D019707:Receptors, Chemokine; D018121:Receptors, Cytokine; D014409:Tumor Necrosis Factor-alpha