Graduation Year

2020

Document Type

Dissertation

Degree

Ph.D.

Degree Name

Doctor of Philosophy (Ph.D.)

Degree Granting Department

Medical Sciences

Major Professor

Jerome Breslin, Ph.D.

Committee Member

Javier Cuevas, Ph.D.

Committee Member

Joshua Scallan, Ph.D.

Committee Member

Patricia Kruk, Ph.D.

Keywords

Endothelial Barrier, eNOS, Glycolysis, Sigma-1R

Abstract

The sigma-1 receptor (σ1) is a single 25 kD polypeptide that acts as a chaperone protein residing primarily in the endoplasmic reticulum Its interaction with mitochondria at the mitochondria-associated ER membrane domain is well-documented. Following activation, σ1 binds to the inositol trisphosphate receptor (Hayashi & Su), and modulates intracellular calcium homeostasis. Also, the activated σ1 modulates plasma membrane receptors and ion channel functions, and may regulate cellular excitability. Further, σ1 affects trafficking of lipids and proteins essential for neurotransmission, cell growth and motility. Activation of σ1 provides neuroprotection and cardio-protection in various models. Examples of neuroprotection include but not limited to its therapeutic use in neuro-psychiatric disorders, cognitive disorders, neurodegenerative diseases, pain and stroke. In regards to cardio-protection, Bhuiyan et al have shown that cardiac σ1 receptors have a role in ameliorating cardiac hypertrophy in pressure overload induced heart failure model. In addition to neuro- and cardio-protection, some studies have shown different actions of σ1 in endothelium such as enhancement of nitric oxide production or modulation of calcium homeostasis. The recent finding by Dan-Yang Liu et al that σ1 activation alleviates blood brain barrier dysfunction in vascular dementia mice suggests its role in maintaining endothelial junctional integrity, which has not previously been studied in detail.

In the current project we investigated the potential involvement of σ1 receptors in enhancing endothelial barrier function and modulating arteriolar diameter pursuing both genetic and pharmacological approaches. We assessed the extent to which σ1 activation can protect endothelial cells from barrier disrupting agents such as the inflammatory cytokine IL-1β or the mitochondrial ionophore Carbonyl cyanide m-chlorophenyl hydrazine (Ganote & Armstrong). σ1 are known to maintain mitochondrial integrity, improve cell survival and stress response via mitochondria and regulate oxidative stress derived from mitochondria based on their localization at mitochondria associated ER membrane in neurological models. However, there is lack of studies about its effects in regard to endothelial mitochondria or bioenergetics particularly glycolysis. The present project will outline its role in maintaining endothelial barrier integrity with respect to endothelial junctional proteins and endothelial bio-energetics.

Several reports suggest that σ1 activation improves outcomes following stroke injury. Previously identified mechanisms include preserving calcium homeostasis and glial cell function, lowering infarct volume. Our recent study shows that σ1 agonism by afobazole relaxes rat mesenteric lymphatic vessels. The response was attenuated by the NO synthase (Hernandez-Resendiz et al.) inhibitor L-NAME. Furthermore, afobazole was shown to elicit elevated NO production in cultured lymphatic endothelial cells (Trujillo et al., 2017). Other studies have also suggested the potential link between σ1 an eNOS. Bhuiyan et al have demonstrated that σ1 agonism by DHEA treatment induces stimulation of Akt-eNOS signaling pathways in rat thoracic aorta (Bhuiyan et al., 2011). They have also shown that σ1 activation protects against hypertension-induced kidney injury by stimulation of Akt-eNOS signaling pathway (Bhuiyan et al., 2011b). Thus, in the current project in addition to studying intracellular mechanisms, we also investigated the ex vivo role of σ1 activation in brain arteriolar diameter using and also human arteriolar diameter modulation.

In conclusion, we elucidated several previously unreported roles of σ1 in endothelium. We found that σ1 is critical and important in maintain basal levels of endothelial barrier maintenance. We also highlighted the role of σ1 activation in eNOS activation and NO production. We reported that the effect of σ1 activation on endothelial barrier is tightly linked to shifting metabolic pathways towards enhancing glycolysis and glycolytic ATP production which is important in maintaining barrier function. Future studies are needed to examine more pathways involving σ1 signaling in endothelial cells and validate these pathways in animal models.

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