Publication: Hepatosit hücrelerinde kolesterol ile indüklenen endoplazmik retikulum stres aracılı apoptozun mitojenle aktive edilmiş protein kinazlar ile ilişkisi
Abstract
Amaç: AML12 fare hepatosit hücre hattında kolesterol içeren lipozomun indüklediği in vitro kolesterol birikimi sonucu meydana gelen ER stres aracılı apoptoz mekanizmasında CHOP aktivasyonunun mitojenle-aktive edilmiş protein kinazlar ile ilişkisinin incelenmesi. Gereç ve Yöntem: Çalışmamızda lipozom ve kolesterol içeren lipozom kompleksleri hazırlanarak farklı inkübasyon süreleri ile AML12 hücre hattına uygulandı. In vitro kolesterol birikimi Filipin boyama ile belirlenirken, fosfo-IRE1, CHOP, p38 ve JNK protein ekpresyonları immunfloresans yöntemi ile saptandı. Kolesterol aracılı CHOP aktivasyonunun mitojenle-aktive edilmiş protein kinazlar ile ilişkisinin belirlenmesi için JNK ve p38 inhibitör uygulamalarını takiben CHOP protein ekspresyonları immünfloresans yöntemi ile saptandı. Bulgular: Kolesterol içeren lipozom uygulanan gruplarda in vitro kolesterol birikiminin sağlandığı gözlendi. Sekiz saat boyunca kolesterol içeren lipozom uygulanan grup kontrol grubu ile kıyaslandığında fosfo-IRE1, CHOP, p38, JNK protein ekpresyonlarında anlamlı artış saptandı. Kolesterol içeren lipozom uygulamasına ilaveten p38 inhibitörü uygulanan grupta ise, kolesterol içeren lipozom uygulanan gruba kıyasla, CHOP protein ekspresyonunda anlamlı düzeyde düşüş belirlendi. Sonuçlar: Çalışmamızda kolesterol içeren lipozom verilen AML12 hücre hattında JNK ve p38 protein ekspresyon düzeylerinin fosfo-IRE1 protein düzeyleri ile birlikte arttığı saptandı. CHOP aktivasyon düzeylerinde kolesterol içeren lipozoma ek olarak p38 inhibitörü verilen hücrelerde düşüş gözlenirken JNK inhibitörü verilen hücrelerde düşüş gözlenmedi.
Aim: Investigation of the relationship between CHOP activation and mitogen-activated protein kinases in the ER stress-mediated apoptosis mechanism induced by cholesterol-containing liposome via cholesterol deposition in AML12 mouse hepatocyte cell line. Methods: Liposome and cholesterol-containing liposome complexes were prepared and applied to the AML12 cell with different incubation times. In vitro cholesterol accumulation was determined by Philippine staining, while phospho-IRE1, CHOP, p38 and JNK protein expressions were detected by immunofluorescence method. In order to determine the relationship of cholesterol-mediated CHOP activation with mitogen-activated protein kinases, CHOP protein expressions were determined by immunofluorescence method following JNK and p38 inhibitor applications. Results: Cholesterol accumulation was observed in the groups treated with cholesterol-containing liposomes. A significant increase in phospho-IRE1, CHOP, p38, and JNK protein expressions was detected in the group that was treated with cholesterol-containing liposomes for eight hours compared to the control group. In the group in which p38 inhibitor was applied in addition to the cholesterol-containing liposome application, a significant decrease was determined in CHOP protein expression compared to the cholesterol-containing liposome administered group. Conclusion: It was determined that JNK and p38 protein expression levels increased together with phospho-IRE1 levels in the AML12 cells given cholesterol-containing liposomes. There was a decrease in CHOP activation levels in cells given p38 inhibitor in addition to cholesterol-containing liposome while any differences was not observed in cells given JNK inhibitor.
Aim: Investigation of the relationship between CHOP activation and mitogen-activated protein kinases in the ER stress-mediated apoptosis mechanism induced by cholesterol-containing liposome via cholesterol deposition in AML12 mouse hepatocyte cell line. Methods: Liposome and cholesterol-containing liposome complexes were prepared and applied to the AML12 cell with different incubation times. In vitro cholesterol accumulation was determined by Philippine staining, while phospho-IRE1, CHOP, p38 and JNK protein expressions were detected by immunofluorescence method. In order to determine the relationship of cholesterol-mediated CHOP activation with mitogen-activated protein kinases, CHOP protein expressions were determined by immunofluorescence method following JNK and p38 inhibitor applications. Results: Cholesterol accumulation was observed in the groups treated with cholesterol-containing liposomes. A significant increase in phospho-IRE1, CHOP, p38, and JNK protein expressions was detected in the group that was treated with cholesterol-containing liposomes for eight hours compared to the control group. In the group in which p38 inhibitor was applied in addition to the cholesterol-containing liposome application, a significant decrease was determined in CHOP protein expression compared to the cholesterol-containing liposome administered group. Conclusion: It was determined that JNK and p38 protein expression levels increased together with phospho-IRE1 levels in the AML12 cells given cholesterol-containing liposomes. There was a decrease in CHOP activation levels in cells given p38 inhibitor in addition to cholesterol-containing liposome while any differences was not observed in cells given JNK inhibitor.