Advisor(s)

Mark Olah, PhD
Ohio Northern University
Pharmaceutical & Biomedical Sciences
m-olah@onu.edu

Document Type

Conference Proceedings

Location

ONU McIntosh Center; Wishing Well

Start Date

22-4-2022 10:00 AM

End Date

22-4-2022 11:00 AM

Abstract

Inflammatory states such as sepsis are associated with elevated tumor necrosis factor alpha (TNF-⍺), which facilitates inflammation, increases the production of reactive oxygen species (ROS) and oxidative stress, and contributes to death of endothelial cells (EC). Exchange Factor Directly Activated by cAMP (EPAC1) is an intracellular cyclic-AMP (cAMP) sensor involved in EC responses such as barrier function, angiogenesis and proliferation. Previous studies in our lab demonstrated activation of EPAC1 protected EC from TNF⍺-induced cell death. Specifically, microvascular EC were pretreated for 45 minutes with 100 mM 8-pCPT-2′-O-Me-cAMP (8CPT, specific EPAC1 activator) or 10 mM forskolin (adenylyl cyclase activator that increases cAMP), before a 24-hour exposure to 5 ng/ml TNF-⍺. MTT assays showed that relative to control, elevation of cAMP and specific EPAC1 activation attenuated TNF-⍺-induced EC death (98.7±3.3% and 92.4±4.8% viability in pretreated groups, respectively, compared to 76.3±2.3% in TNF-⍺-alone). This Honors Capstone Enhancement is a mock grant proposal to examine the hypothesis that EPAC1 activation inhibits the pro-apoptotic and ROS-elevating effects of TNF-⍺. The mechanisms of EPAC1 protection from TNF-⍺-induced EC apoptosis will be investigated by measuring the rate of apoptosis, apoptotic markers expression, ROS levels, and changes in mitochondrial membrane potential.

Notes

This presentation is part of the Honors Capstone Enhancement Presentation series.

Open Access

Available to all.

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Apr 22nd, 10:00 AM Apr 22nd, 11:00 AM

Study proposal for additional investigation of EPAC1 in endothelial apoptosis

ONU McIntosh Center; Wishing Well

Inflammatory states such as sepsis are associated with elevated tumor necrosis factor alpha (TNF-⍺), which facilitates inflammation, increases the production of reactive oxygen species (ROS) and oxidative stress, and contributes to death of endothelial cells (EC). Exchange Factor Directly Activated by cAMP (EPAC1) is an intracellular cyclic-AMP (cAMP) sensor involved in EC responses such as barrier function, angiogenesis and proliferation. Previous studies in our lab demonstrated activation of EPAC1 protected EC from TNF⍺-induced cell death. Specifically, microvascular EC were pretreated for 45 minutes with 100 mM 8-pCPT-2′-O-Me-cAMP (8CPT, specific EPAC1 activator) or 10 mM forskolin (adenylyl cyclase activator that increases cAMP), before a 24-hour exposure to 5 ng/ml TNF-⍺. MTT assays showed that relative to control, elevation of cAMP and specific EPAC1 activation attenuated TNF-⍺-induced EC death (98.7±3.3% and 92.4±4.8% viability in pretreated groups, respectively, compared to 76.3±2.3% in TNF-⍺-alone). This Honors Capstone Enhancement is a mock grant proposal to examine the hypothesis that EPAC1 activation inhibits the pro-apoptotic and ROS-elevating effects of TNF-⍺. The mechanisms of EPAC1 protection from TNF-⍺-induced EC apoptosis will be investigated by measuring the rate of apoptosis, apoptotic markers expression, ROS levels, and changes in mitochondrial membrane potential.