Local Oxidative Stress Expansion through Endothelial Cells – A Key Role for Gap Junction Intercellular Communication

Abstract

BackgroundMajor circulation pathologies are initiated by oxidative insult expansion from a few injured endothelial cells to distal sites; this possibly involves mechanisms that are important to understanding circulation physiology and designing therapeutic management of myocardial pathologies. We tested the hypothesis that a localized oxidative insult of endothelial cells (ECs) propagates through gap junction inter-cellular communication (GJIC).Methodology/Principal FindingsCultures comprising the bEnd.3 cell line, that have been established and recognized as suitable for examining communication among ECs, were used to study the propagation of a localized oxidative insult to remote cells. Spatially confined near infrared illumination of parental or genetically modified bEnd.3 cultures, pretreated with the photosensitizer WST11, generated O2•− and •OH radicals in the illuminated cells. Time-lapse fluorescence microscopy, utilizing various markers, and other methods, were used to monitor the response of non-illuminated bystander and remote cells. Functional GJIC among ECs was shown to be mandatory for oxidative insult propagation, comprising de-novo generation of reactive oxygen and nitrogen species (ROS and RNS, respectively), activation and nuclear translocation of c-Jun N-terminal kinase, followed by massive apoptosis in all bystander cells adjacent to the primarily injured ECs. The oxidative insult propagated through GJIC for many hours, over hundreds of microns from the primary photogeneration site. This wave is shown to be limited by intracellular ROS scavenging, chemical GJIC inhibition or genetic manipulation of connexin 43 (a key component of GJIC).Conclusion/SignificanceLocalized oxidative insults propagate through GJIC between ECs, while stimulating de-novo generation of ROS and RNS in bystander cells, thereby driving the insult's expansion.

Hydrogen peroxide as a central redox signaling molecule in physiological oxidative stress: Oxidative eustress

“…The microenvironment surrounding particular cells, notably the extracellular matrix (ECM), modifies the signal [116]. Gap junctional communication via connexin channels between cells may lead to “local oxidative stress expansion” [117].…”

Differential Effects of Indoxyl Sulfate and Inorganic Phosphate in a Murine Cerebral Endothelial Cell Line (bEnd.3)

“…This cell line retains key features of differentiated endothelium and is considered to represent an authentic model for examination of cerebral endothelial dysfunction [57]. Cells were cultured in DMEM supplemented with 10% fetal bovine serum (FBS), 2 mmol/L glutamine, 100 IU/mL penicillin, and 100 IU/mL streptomycin, and maintained at 37 °C in a humidified atmosphere containing 5% CO 2 .…”

Into rather unexplored terrain—transcellular transport across the blood–brain barrier

“…HCs in contrast provide a paracrine Ca 2+ signaling route whereby a messenger is released in the extracellular compartment and mobilizes Ca 2+ in remote cells by binding to its corresponding receptor. Intercellular Ca 2+ waves are an important communication tool for the electrically non‐excitable cells in the CNS and has also been identified in BBB endothelial cells ( in vitro ) (Feine et al, ; Park et al, ; Vandamme et al, ). Data from our lab indicate that the propagation of endothelial intercellular Ca 2+ waves is associated with high amplitude [Ca 2+ ] i changes and a large permeability increase, which is more pronounced than the permeability increase associated with BK induced Ca 2+ oscillations (De Bock et al, ).…”