July 2019
Volume 60, Issue 9
Open Access
ARVO Annual Meeting Abstract  |   July 2019
Alpha-1 antitrypsin avoids epithelial to mesenchymal transition in ARPE-19 cells in a model of Diabetic Retinopathy
Author Affiliations & Notes
    Nanomedicine & Vision, IIMT-Austral-CONICET, Pilar, Buenos Aires, Argentina
  • Juan Pablo Salica
    Nanomedicine & Vision, IIMT-Austral-CONICET, Pilar, Buenos Aires, Argentina
  • Micaela Petrigliano
    Nanomedicine & Vision, IIMT-Austral-CONICET, Pilar, Buenos Aires, Argentina
  • Eduardo Chuluyan
    Universidad Austral, Pilar, BUENOS AIRES, Argentina
    Farmacología, UBA, Buenos Aires, Buenos Aires, Argentina
  • Juan E Gallo
    Nanomedicine & Vision, IIMT-Austral-CONICET, Pilar, Buenos Aires, Argentina
    Oftalmología, Hospital Universitario Austral, Pilar, Buenos Aires, Argentina
  • Footnotes
    Commercial Relationships   MARÍA CONSTANZA POTILINSKI, None; Juan Pablo Salica, None; Micaela Petrigliano, None; Eduardo Chuluyan, None; Juan Gallo, None
  • Footnotes
    Support  PICTO-Austral
Investigative Ophthalmology & Visual Science July 2019, Vol.60, 5364. doi:
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      MARÍA CONSTANZA POTILINSKI, Juan Pablo Salica, Micaela Petrigliano, Eduardo Chuluyan, Juan E Gallo; Alpha-1 antitrypsin avoids epithelial to mesenchymal transition in ARPE-19 cells in a model of Diabetic Retinopathy. Invest. Ophthalmol. Vis. Sci. 2019;60(9):5364.

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

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Purpose : Diabetic retinopathy (DR) is a leading cause of blindness in working-age population and is regarded as a microvascular complication due to the breakdown of the endothelial barrier.
The retinal pigment epithelium (RPE), which is the major component of the outer blood-retinal barrier between the choroid and neurosensory retina, is also the key to maintain the integrity of retinal tissues. In DR, as the disease progresses, cell-cell junction complexes between RPE cells are to disassemble losing their polarization and the leakage can be found through paracellular space. Dissociation of cultured RPE cells leads to dedifferentiation of the cells into fibroblast-like cells through epithelial to mesenchymal transition (EMT). To maintain the integrity of epithelial cells, adherens junctions are critical by forming cell-cell contacts as protein complexes consisting of cadherin homodimers. Nuclear factor-kappa-B (NFκB) is a multifunctional transcription factor participating in the transcriptional regulation of a variety of inflammatory mediators, enzymes and adhesion molecules. Tranglutaminase2 (TGM2) is a protein involved in many processes, such as angiogenesis, cell adhesion, migration, survival and EMT.
In this work, we explore the expression of NFκB, TGM2, and other proteins involved in integrin mediated signaling pathway as HIF1α and E-Cadherin on ARPE-19 cells exposed to high glycemia and Alpha-1 antitrypsin (A1AT) treatment.

Methods : ARPE-19 cells (ATCC® CRL-2302TM, Manassas, Virginia, USA) were maintained in DMEM/F12 (Invitrogen, Carlsbad, California, USA) containing 2μM L-glutamine, 100U/ml penicillin, 100μg/ml streptomycin, and 10% fetal bovine serum. ARPE-19 cells (passages 9 to 12) were incubated 16h with DMEM 5,5mM glucose (Control), DMEM 5,5mM glucose + 4.5mg/ml A1AT (Control+A1AT), DMEM 30mM glucose (Diabetic), DMEM 30mM glucose + 4.5mg/ml A1AT (Diabetic+A1AT). Cells were harvested with RIPA buffer for Western blot assay or fixed for immunohistochemistry.

Results :
Protein expressions of NFκB p65, TGM2, AKT, pAKT and HIF1α were diminished with A1AT treatment, while E-Cadherin was increased.

Conclusions : Outcomes support the hypothesis that A1AT increases E-Cadherin expression through decreasing integrin mediated signaling avoiding EMT. These results might help to understand mechanisms involved in inflammatory processes in DR and make A1AT a suitable molecule to DR treatment.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.


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