J Bacteriol Virol.  2007 Dec;37(4):241-248. 10.4167/jbv.2007.37.4.241.

The Expression of TLR3 and Cytokines Induced by Poly I:C in Human Retinal Pigment Epithelial Cells

• Choi SJ
• Kim J
• Park SJ
• Park HS
• Lee KH
• Kim SK
• Park JY

• KMID: 1513585
• DOI: http://doi.org/10.4167/jbv.2007.37.4.241

Abstract

In this study, we examined the expression of Toll-like receptor3 (TLR3) by human retinal pigment epithelial cells (RPE) and determined whether exposure to the TLR3 agonist polyinosinic-polycytidylic acid (poly I:C) would induced the expression of cytokines in these cells. RT-PCR revealed that TLR3 was constitutively expressed in human RPE, and its expression was increased by treatment with poly I:C. After treatment with poly I:C, we determined the expression levels of pro-inflammatory cytokines in human RPE using RT-PCR and ELISA. We demonstrated that poly I:C treatment increased the production of TNF-alpha, IL-6, and IL-8 in human RPE. Upon exposure to poly I:C, human RPE initiated antiviral response resulting in the induction of IFN-beta mRNA expression and 2',5'-oligoadenylate synthetase mRNA expression. These results suggest that human RPE may participate in ocular defense mechanism against viral infection through TLR3.

Keyword

Toll-like receptor3; Poly I:C; Viral infection; Retinal pigment epithelium; Interferon

MeSH Terms

2',5'-Oligoadenylate Synthetase
Cytokines*
Enzyme-Linked Immunosorbent Assay
Epithelial Cells*
Humans*
Interferons
Interleukin-6
Interleukin-8
Poly I-C
Retinal Pigment Epithelium
Retinaldehyde*
RNA, Messenger
Tumor Necrosis Factor-alpha
2',5'-Oligoadenylate Synthetase
Cytokines
Interferons
Interleukin-6
Interleukin-8
Poly I-C
RNA, Messenger
Retinaldehyde
Tumor Necrosis Factor-alpha

Figure

• 

  Figure 1. The effect of poly I:C on TLR3 expression in human fetal RPE. (A) After treatment with 50 μg/ml of poly I:C, total RNA was isolated and RT-PCR was performed. (B) After poly I:C treatment for 7h with various concentrations, real time PCR was performed using primer sets of GAPDH, and TLR3. Comparative expression levels were calculated.

• 

  Figure 2. The effect of poly I:C on TNF-α and IL-8 expression. The human fetal RPE was treated with 50 μg/ml of poly I:C. At different time intervals, total RNAs were isolated. Then, mRNA levels of TNF-α (A), and IL-8 (B) were analyzed using RT-PCR.

• 

  Figure 3. The effect of poly I:C on IL-6 production. (A) The human fetal RPE was treated with various doses of poly I:C for 7h. Total RNAs were isolated and mRNA levels of IL-6 was analyzed using RT-PCR. (B) The human fetal RPE was incubated with poly I:C for 24h and culture supernatants were then harvested. The levels of IL-6 in the culture supernatants were measured using ELISA.

• 

  Figure 4. The effect of poly I:C on IFN-β expression in human fetal RPE. (A) After poly I:C treatment for 7h at variable dosages, total RNA was isolated and RT-PCR was performed. (B) After treatment with 50 μg/ml of poly I:C, total RNA was isolated and RT-PCR was performed.

• 

  Figure 5. The effect of poly I:C on 2′,5′-OAS expression in human fetal RPE. (A) After poly I:C treatment for 7h with various concentrations, total RNA was isolated and RT-PCR was performed. (B) After treatment with 10 μg/ml of poly I:C, total RNA was isolated and RT-PCR was performed. (C) After treatment with various concentrations of LPS, total RNA was isolated and RT-PCR was performed.

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