Measurement of gene expression in human retinal microvessels by solution hybridization

E. Cagliero, M. B. Grant, M. Lorenzi

Research output: Contribution to journalArticle

13 Scopus citations

Abstract

Changes in gene expression could play a central role in the phenotypic abnormalities of the retinal vascular cells observed in diabetic retinopathy and other retinal diseases. To measure gene expression in human retinal microvessels, a RNA-probe excess solution hybridization assay was used. Retinal microvessels were isolated from eyes obtained within 36 hr of death, and intact RNA was extracted by the guanidine method. Hybridization of poly(A)+ RNA northern blots revealed only the cytoskeletal β-actin message; by using the more sensitive solution hybridization assay, the plasminogen activator-inhibitor 1 (PAI-1) and von Willebrand factor (vWF) mRNAs were quantified. The prevalence of these transcripts in the retinal microvessels was 0.04 x 106 copies/ng RNA for PAI-1 and 0.14 x 106 copies/ng RNA for vWF, much less than the prevalence in human umbilical vein endothelial cells (1.93 x 106 and 3.90 x 106, respectively). The PAI-1 mRNA levels in retinal microvessels isolated from five type II diabetic patients were significantly higher than those in vessels isolated from ten age-matched controls (0.06 x 106 versus 0.04 x 106 copies/ng RNA, P < 0.05). The solution hybridization assay accurately measured low-abundance mRNAs in human retinal microvessels; determination of gene expression in these cells could aid in understanding the pathogenesis of important ophthalmologic diseases such as diabetic retinopathy.

Original languageEnglish (US)
Pages (from-to)1439-1445
Number of pages7
JournalInvestigative Ophthalmology and Visual Science
Volume32
Issue number5
StatePublished - Jan 1 1991

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ASJC Scopus subject areas

  • Ophthalmology
  • Sensory Systems
  • Cellular and Molecular Neuroscience

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