Expression levels of Fibulin- 2 (Fbln2) and Fibulin-5 (Fbln5) on subretinal fluid in human primary rhegmatogenous retinal detachment.

Abstract

Purpose : To compare the expression levels of Fibulin- 2 (Fbln2) and Fibulin-5 (Fbln5) on subretinal fluid samples in human primary rhegmatogenous retinal detachment with aqueous humor from eyes that had senile cataract surgery.

Methods : For WB analysis the monoclonal anti- Fbln2 and anti-Fbln5 antibodies were either obtained from a commercial source (Santacruz Biotechnology). Protein extracts from six patients were prepared and estimated from subretinal fluid of rhegmatogenous retinal detachment. Then, 50ug/ul were electrophoretically transferred onto nitrocelulose and immunoblotted for both fibulins. Aqueous humor samples from normal patients were used as control.
Fbln2 and Fbln5 mRNA were analyzed in these samples by qRT-PCR. The RNA extract were prepared using the Directzol RNA isolation kit. Contaminating DNA was then removed by a DNase I solution. One microgram of total RNA was used for reverse transcription into the complementary DNA (cDNA), using the kit Brilliant (from Agilent Technologies). The polymerase chain reaction (qPCR) was performed on transcribed cDNA, using Fbln2 and Fbln5 specific primers using the qPCR kit. The mixtures were amplified in a thermal cycler Rotorgene (QIAGEN). The present study received approval from the Local Ethic Committee. All patients gave their full written consent for participation.

Results : The levels of Fbln 2 and Fbln5 mRNA in the cellular fraction of subretinal fluid in retinal detachment were significantly higher as compared with that in normal samples (aqueous humor) (P<0.05). In the same way, the protein levels of Fbln2 and Fbln in these samples shown a similar pattern when they were analyzed by immunoblotting.

Conclusions : In this report we show the presence and the up-regulation of Fbln 2 and Fbln5 in the cellular fraction of subretinal fluid of patients in primary rhegmatogenous retinal detachment.

This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.