Effect of pituitary adenylate cyclase-activating polypeptide (PACAP) on IL-6, IL-8, and MCP-1 expression in human retinal pigment epithelial cell line.
OBJECTIVE
To examine pituitary adenylate cyclase-activating polypeptide (PACAP) receptors (PAC1, VPAC1, and VPAC2) mRNA and the effect of PACAP on interleukin-6 (IL-6), interleukin-8 (IL-8), and monocyte chemotactic protein-1 (MCP-1) expression in human retinal pigment epithelial cell line (ARPE-19) stimulated with interleukin-1beta (IL-1beta).


METHODS
Expression of PACAP receptor mRNA was examined by reverse transcription polymerase chain reaction (RT-PCR). PACAP and IL-1beta were added to serum-free medium. IL-6, IL-8, and MCP-1 mRNA were measured by real-time PCR. IL-6, IL-8, and MCP-1 protein concentrations were measured using enzyme-linked immunosorbent assay. Nuclear factor kappaB (NF-kappaB) translocation was examined by immunofluorescence.


RESULTS
PAC1 and VCAP1 receptors mRNA were expressed in unstimulated cells. VCAP2 mRNA was expressed in cells stimulated with IL-1beta. IL-1beta stimulated IL-6, IL-8, and MCP-1 mRNA expression and protein levels. PACAP (10(- 7) to 10(- 6) M) inhibited IL-1beta -stimulated IL-6, IL-8, and MCP-1 mRNA and protein levels. Immunofluorescence of NF-kappaB in the nucleus was dense 30 min after stimulation with IL-1beta, and it was decreased by PACAP.


CONCLUSIONS
ARPE-19 cells had PACAP receptors mRNA. PACAP inhibited IL-6, IL-8, and MCP-1 expression and protein secretion. Possibly, the effect on cytokines may be via suppression of NF-kappaB translocation.
Zhang
Xue-Yun
XY
Department of Ophthalmology, Toyama Medical and Pharmaceutical University, Toyama, Japan. ophthal@ms.toyama-mpu.ac.jp
Hayasaka
Seiji
S
Chi
Zai-Long
ZL
Cui
Hu-Shan
HS
Hayasaka
Yoriko
Y
D002460:Cell Line; D018932:Chemokine CCL2 / Q000235:genetics* / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D019084:Fluorescent Antibody Technique, Indirect; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D007375:Interleukin-1 / Q000494:pharmacology; D015850:Interleukin-6 / Q000235:genetics* / Q000378:metabolism; D016209:Interleukin-8 / Q000235:genetics* / Q000378:metabolism; D018377:Neurotransmitter Agents / Q000494:pharmacology*; D010857:Pigment Epithelium of Eye / Q000187:drug effects* / Q000378:metabolism; D051219:Pituitary Adenylate Cyclase-Activating Polypeptide / Q000494:pharmacology*; D012333:RNA, Messenger / Q000378:metabolism; D051236:Receptors, Pituitary Adenylate Cyclase-Activating Polypeptide / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D051996:Transcription Factor RelA / Q000378:metabolism
C111835:CCL2 protein, human; D018932:Chemokine CCL2; D007375:Interleukin-1; D015850:Interleukin-6; D016209:Interleukin-8; D018377:Neurotransmitter Agents; D051219:Pituitary Adenylate Cyclase-Activating Polypeptide; D012333:RNA, Messenger; D051236:Receptors, Pituitary Adenylate Cyclase-Activating Polypeptide; D051996:Transcription Factor RelA