αB-Crystallin Peptide Fused with Elastin-like Polypeptide: Intracellular Activity in Retinal Pigment Epithelial Cells Challenged with Oxidative Stress

• Prif Awduron: Sara Aly Attia (Awdur), Anh Tan Truong (Awdur), Alvin Phan (Awdur), Shin-Jae Lee (Awdur), Manal Abanmai (Awdur), Marinella Markanovic (Awdur), Hugo Avila (Awdur), Haozhong Luo (Awdur), Atham Ali (Awdur), Parameswaran G. Sreekumar (Awdur), Ram Kannan (Awdur), J. Andrew MacKay (Awdur)
• Fformat: Llyfr
• Cyhoeddwyd: MDPI AG, 2023-09-01T00:00:00Z.
• Pynciau: article
• Mynediad Ar-lein: Connect to this object online.

Crynodeb

Background: Oxidative stress-induced retinal degeneration is among the main contributing factors of serious ocular pathologies that can lead to irreversible blindness. αB-crystallin (cry) is an abundant component of the visual pathway in the vitreous humor, which modulates protein and cellular homeostasis. Within this protein exists a 20 amino acid fragment (mini-cry) with both chaperone and antiapoptotic activity. This study fuses this mini-cry peptide to two temperature-sensitive elastin-like polypeptides (ELP) with the goal of prolonging its activity in the retina. Methods: The biophysical properties and chaperone activity of cry-ELPs were confirmed by mass spectrometry, cloud-point determination, and dynamic light scattering 'DLS'. For the first time, this work compares a simpler ELP architecture, cry-V96, with a previously reported ELP diblock copolymer, cry-SI. Their relative mechanisms of cellular uptake and antiapoptotic potential were tested using retinal pigment epithelial cells (ARPE-19). Oxidative stress was induced with H₂O₂ and comparative internalization of both cry-ELPs was made using 2D and 3D culture models. We also explored the role of lysosomal membrane permeabilization by confocal microscopy. Results: The results indicated successful ELP fusion, cellular association with both 2D and 3D cultures, which were enhanced by oxidative stress. Both constructs suppressed apoptotic signaling (cleaved caspase-3); however, cry-V96 exhibited greater lysosomal escape. Conclusions: ELP architecture is a critical factor to optimize delivery of therapeutic peptides, such as the anti-apoptotic mini-cry peptide; furthermore, the protection of mini-cry via ELPs is enhanced by lysosomal membrane permeabilization.

Disgrifiad o'r Eitem

10.3390/antiox12101817
2076-3921