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J.–H. Sohn, Y. Wang, H.J. Kaplan, N.S. Bora; Complement Regulatory Proteins Are Expressed on Human Retinoblastoma Cells . Invest. Ophthalmol. Vis. Sci. 2005;46(13):1010.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: We have previously observed that various complement regulatory protein (CRPs) – namely MCP, DAF and CD59 – are differentially expressed in the normal human eye. This study was performed to investigate the expression of MCP, DAF and CD59 on human retinoblastoma cells. Methods: Y79 human retinoblastoma cells (ATCC HTB 18) were obtained from the American Type Culture Collection and grown under standard conditions using RPMI–1640 medium supplemented with 15% fetal calf serum, 0.02% glutamine and 50 ug/ml gentamycin. The expression of CRPs was examined by flow cytometry and semi–quantative RT–PCR analysis. Total RNA was extracted from Y79 cells using RNeasy Kit (Promega). Reverse transcription polymerase chain reaction (RT–PCR) was performed using a RNA–PCR kit obtained from Applied Biosystems (Foster City, CA) according to the manufacturer’s recommendation. The sense and anti–sense oligonucleotide primers were synthesized at Integrated DNA technologies (Coralville, IA). RT–PCR products were analyzed on 2% agarose gel and visualized under UV light. Bands corresponding to the sequences of interest were scanned and visualized by Bio–Rad imaging densitometer. Monoclonal antibodies against human MCP, DAF and CD59 were used in flow cytometric analysis. Results: Our results of flow cytometry and RT–PCR analysis demonstrate that MCP, DAF and CD59 transcripts as well as proteins are abundantly expressed on retinoblastoma cells. Conclusions: Identification of CRPs on human retinoblastoma cells may be a major advance toward the understanding of the mechanism(s) involved in the immune evasion by retinoblastoma. These studies may have therapeutic potential.
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