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A. Krishnakumar, D. P. Dash, D. A. Simpson, A. W. Stitt, C. E. Willoughby; Alternative Splicing of the Erythropoietin Receptor (EPOR) in the Normal Retina. Invest. Ophthalmol. Vis. Sci. 2010;51(13):5997.
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© ARVO (1962-2015); The Authors (2016-present)
Alternative splicing of the erythropoietin receptor (EPOR) can generate protein isoforms which could impinge on the therapeutic ability of recombinant erythropoietin (EPO) or EPO analogues in the treatment of retinal vascular disease and in retinal neuroprotection.The aim of this study was to determine the pattern of alternative splicing pattern of EPOR in the normal murine and human retinae.
RNA was extracted from pooled, normal adult murine (C57) and human retinae, spontaneously immortalized human muller glial (MIO-M1) (Limb, et al 2002) and RPE-19 cell lines. Variant specific PCR of cDNA was performed for known EPOR splice variants: EPOR-F (full length transcript); EPOR-S (soluble variant) and EPOR-T (truncated variant). PCR products were separated on 1% lithium borate agarose gels, gel extracted and sequenced to confirm transcript identity. Isoform specific primers and RT-qPCR was used to quantify EPOR splice variants in the normal murine and human retinae.
In the normal murine and human adult retina three known alternative transcripts were identified by RT-PCR (1.7 kb for EPOR-F, 300 bp for EPOR-T and 200 bp for EPOR-S) and confirmed by sequencing following gel extraction. RT-qPCR demonstrated that EPOR-F shows maximal expression followed by EPOR-S and EPOR-T which show almost negligible expression in the normal murine and human retinae.
The identification of EPOR-S and EPOR-T has not previously been reported in the murine or human retina. Alternative splicing in retinal EPOR could potentially result in unexpected deleterious, functional impact and could have significant therapeutic consequences when using EPO analogues in retinal vascular disease and neuro-protection.
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