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EXAMINING APOPTOSIS IN A SEPSIS CELL CULTURE MODEL
Author(s) -
Davis Bethany,
Del Gaizo Moore Victoria
Publication year - 2016
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.30.1_supplement.647.5
Subject(s) - apoptosis , sepsis , immune system , cytokine , immunology , programmed cell death , flow cytometry , kidney , hek 293 cells , acute kidney injury , proinflammatory cytokine , annexin , medicine , biology , inflammation , cancer research , receptor , biochemistry
Sepsis, a systemic inflammatory response to infection of the blood by pathogenic organisms, impacts millions of people around the world every year. Pro‐inflammatory cytokines are released to fight the infection but many times lead to organ damage such as Acute Kidney Injury (AKI). AKI, a disorder characterized by the abrupt deterioration in kidney function, has a broad spectrum of consequences ranging from mild kidney injury to permanent loss in renal function and kidney failure. Common amongst critically ill patients, the development of AKI is believed to be due, at lease in part, to apoptosis. Apoptosis, or programmed cell death, is controlled by a multitude of cellular mechanisms. Cytokines can initiate the extrinsic pathway of apoptosis by binding to death receptors on the cell surface. While it is definitively known that apoptosis plays a large role in immune cell decline under septic conditions, its influence in AKI is not entirely understood. The objective of this study is to determine an efficient means of examining the role of cytokines in inducing apoptosis in renal cells in order to develop greater insight into the pathogenesis of sepsis‐associated AKI. Human immune THP1 cells were treated with a synthetic LPS to stimulate cytokine secretions. Cultured human embryonic kidney (HEK) cells were treated with either these naturally secreted or recombinant cytokines. Then flow cytometry of the HEK cells stained with Annexin V‐FITC determined the amount of apoptosis in the culture sample. Flow cytometry data demonstrates that recombinant cytokines allow for quicker analysis than naturally secreted cytokines. Apoptosis amongst HEK cells treated with recombinant cytokines for 48 hours was comparable to treatment with naturally secreted cytokines for a duration of 120 hours. Treatment with recombinant cytokines results in much quicker analysis because it excludes the time necessary for THP1 stimulation and cytokine recovery as well as significantly reduces the duration of treatment. Recombinant cytokines also have the benefit of easily picking precise concentrations of TNF‐alpha and other cytokines. With the greater understanding of the apoptotic processes associated with sepsis, the biochemical mechanism at the foundation of AKI can be further elucidated and the creation of more effective treatments for this disease may be possible. Support or Funding Information Elon University Undergraduate Research Program (BD), Elon University College Fellows Program (BD), FR&D (VM)