September 2016
Volume 57, Issue 12
Open Access
ARVO Annual Meeting Abstract  |   September 2016
Exosomes Released From Human Retinal Endothelial Cells Upon 15-HETE Treatment or ER Stress Induction: Potential Role of Nanoparticles in Diabetic Retinopathy
Author Affiliations & Notes
  • Khaled Elmasry
    Cellular Biology and Anatomy, Medical College of Georgia at Augusta University, Augusta, Georgia, United States
    Human Anatomy and Embryology, Faculty of Medicine, Mansoura University, Mansoura, Egypt
  • Khaled Hussein
    Oral Biology, Dental college of Georgia at Augusta University, Augusta, Georgia, United States
    Surgery and Medicine, Oral and Dental Research Division, National Research Center, Cairo, Egypt
  • Heba Mohamed Saleh
    Oral Biology, Dental college of Georgia at Augusta University, Augusta, Georgia, United States
    Culver Vision Discovery Institute and Ophthalmology, Augusta, Georgia, United States
  • Ahmed S Ibrahim
    Oral Biology, Dental college of Georgia at Augusta University, Augusta, Georgia, United States
    Clinical Biochemistry, Faculty of Pharmacy, Mansoura University, Mansoura, Egypt
  • Nehal M El-Sherbiny
    Oral Biology, Dental college of Georgia at Augusta University, Augusta, Georgia, United States
    Clinical Biochemistry, Faculty of Pharmacy, Mansoura University, Mansoura, Egypt
  • Inas Helwa
    Cellular Biology and Anatomy, Medical College of Georgia at Augusta University, Augusta, Georgia, United States
  • Yutao Liu
    Cellular Biology and Anatomy, Medical College of Georgia at Augusta University, Augusta, Georgia, United States
    Culver Vision Discovery Institute and Ophthalmology, Augusta, Georgia, United States
  • Mohamed Al-Sayed Al-Shabrawey
    Oral Biology, Dental college of Georgia at Augusta University, Augusta, Georgia, United States
    Culver Vision Discovery Institute and Ophthalmology, Augusta, Georgia, United States
  • Footnotes
    Commercial Relationships   Khaled Elmasry, None; Khaled Hussein, None; Heba Saleh, None; Ahmed Ibrahim, None; Nehal El-Sherbiny, None; Inas Helwa, None; Yutao Liu, None; Mohamed Al-Shabrawey, None
  • Footnotes
    Support  NIH (1R01EY023315) 
Investigative Ophthalmology & Visual Science September 2016, Vol.57, 3230. doi:
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      Khaled Elmasry, Khaled Hussein, Heba Mohamed Saleh, Ahmed S Ibrahim, Nehal M El-Sherbiny, Inas Helwa, Yutao Liu, Mohamed Al-Sayed Al-Shabrawey; Exosomes Released From Human Retinal Endothelial Cells Upon 15-HETE Treatment or ER Stress Induction: Potential Role of Nanoparticles in Diabetic Retinopathy. Invest. Ophthalmol. Vis. Sci. 2016;57(12):3230.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose : Exosomes are specialized nanoparticles containing lipids, proteins and miRNAs. They have been described as biomarkers in several pathological conditions. We recently showed that 12/15 lipoxygenase (12/15-LOX) and its lipid metabolites 12- and 15- Hydroxyicosatetraenoic acid (12- and 15-HETE) are implicated in retinal microvascular dysfunction during diabetic retinopathy (DR). The goal of the current study was to investigate the potential contribution of ER stress and exosomes in retinal microvascular dysfunction induced by 12/15-LOX-derived lipid metabolites.

Methods : Primary human retinal microvascular endothelial cells (HRMECs) were grown till ~90% confluence, then we shift to serum free media 12 hours before the treatment. The cells then treated with 15-HETE (0.1 µM), tunicamycin (5µg/ml) or their vehicles (n=3) for 30 minutes or 4 hours. Cell lysate was used to examine the effect of 15-HETE on ER stress markers; XBP-1s, p-PERK and PDI using Western blot. HRMECs conditional media were collected and exosomes were isolated using Invitrogen Total Exosome Isolation Reagent (from cell culture media) according to the manufacturer’s protocol. The size and quantity of isolated exosomes were characterized using nanoparticle tracking analysis with ZetaView.

Results : 15-HETE induced ER stress in HRMECs with elevated expression of XBP-1s (P value < 0.0001), p-PERK (P value= 0.0280) and PDI (P value=0.0098). The quantity of exosomes secreted by HRMECs is significantly increased by 15-HETE treatment (P value =0.0347) and ER stress induction by tunicamycin (P value =0.0132) for 30 minutes in comparison to the vehicle treated groups. Furthermore, 15-HETE significantly increased the quantity of secreted exosomes after 4 hours (P value =0.0108), while the further increase by tunicamycin was not significant (P value =0.1705) compared to the vehicle treated group. However, there was no significant difference in the quantity of exosomes released under the same treatment at different time points.

Conclusions : This study represents the first report to examine the potential role of HRMECs-derived exosomes in DR. Further investigations will be done to examine the lipid, protein and RNA content of the released endothelial exosomes and whether they play a role in retinal intercellular communication.

This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.

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