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Investigation of Gene Expressions Related to Oxidative Stress and Apoptosis in Human Thoracic Aortic Aneurysms
Thoracic aortic aneurysm (TAA) is a relatively rare disease with a high risk of mortality. It has been shown that reactive oxygen species (ROS) play an important role in the pathogenesis of this disease. Increased oxidative stress induces apoptosis in smooth muscle cells within the aorta. Loss of these cells is one of the prominent factors lead to development of TAA.
The aim of this study is to contribute to current knowledge on death pathways in TAA by comparing the expression levels of some genes related to oxidative stress and apoptosis in smooth muscle cells from aneurysmal (experimental group) and non-aneurysmal (control group) tissues.
Both aneurysmal and non-aneurysmal tissues were collected during the surgical operation by expert medical doctors at Kartal Kosuyolu Training and Research Hospital with the approval of ethics council. Primary cell cultures were established using the tissue samples. Oxysterols (7-ketocholesterol and 25-hydroxycholesterol) were used to induce oxidative stress in smooth muscle cells. Intracellular ROS levels were determined after the treatment with these agents. The cell death in response to ROS was confirmed via treatment with antioxidant agents, resveratrol and N-acetylcysteine. The mean of cell death via apoptosis was determined by live cell imaging microscopy, DNA condensation/fragmentation analyses and caspase 3 activity measurements.
The role of a selected set of genes (Bax, p53, caspase 3, Akt1 and Akt2) in the apoptosis of TAA cells was investigated using gene silencing technology. The silencing efficiency was verified using real-time polymerase chain reaction (RT-PCR). In addition, expression levels of SOD and Nox4 genes involving oxidative stress response were analyzed.
It was observed that oxysterols caused apoptosis through inducing oxidative stress, and increased caspase 3 activity in both experimental and control cells. However, expression level of apoptotic genes, Bax, p53 and caspase 3, and oxidative stress response genes, SOD and Nox4 did not change significantly in these cells.
With these findings, it was concluded that 1) the oxidative stress response has triggered cell death via apoptotic pathways in TAAs; 2) SOD and Nox4 expression levels have not changed; 3) cells could undergo apoptosis independent of Bax, p53 and caspase 3; 4) cell death has not been enhanced by silencing anti-apoptotic genes, Akt1 or Akt2; and 5) all these processes in TAAs have occurred similar to non-aneurysmal tissues.
For a better understanding of whole mechanism of TAA all related genes might be studied on protein level and other death pathways should also be investigated.
  

YÖRÜK Emre
Danışman : Doç. Dr. Gülruh ALBAYRAK

Anabilim Dalı : Moleküler Biyoloji ve Genetik

Programı : -

Mezuniyet Yılı : 2014

Tez Savunma Jürisi : Doç. Dr. Gülruh ALBAYRAK

Prof. Dr. Şule ARI

Prof. Dr. Kadir TURAN

Prof. Dr. Muhsin KONUK

Prof. Dr. Keriman GÜNAYDIN