6-hidroksi dopamin (6-OHDA) ile oluşturulan ın vitro parkinson hastalığı modelinde humanin'in nöroprotektif etkisinin araştırılması
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Dosyalar
Tarih
2019
Yazarlar
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Yayıncı
Ege Üniversitesi, Sağlık Bilimleri Enstitüsü
Erişim Hakkı
info:eu-repo/semantics/openAccess
Özet
24 amino asitli bir peptid olan Humanin (HN), keşfedildikten sonra Alzheimer hastalığı da dahil olmak üzere birçok nörodejeneratif hastalıkta incelenmiştir. HN çok biyoaktif bir peptittir. Böylece, birçok hücre içi ve hücre dışı sinyalleşme mekanizmasını etkiler. HN, PI3k / Akt, JAK / STAT, MAPK / ERK yolları ve IGFBP, Bax proteinleri üzerinden hücre büyümesini, çoğalmasını ve hayatta kalmasını düzenler. Bu çalışmanın amacı, HN'nin 6 hidroksidopamin (6-OHDA) ile indüklenen in vitro Parkinson hastalığı modelinde nöroprotektif bir etkisinin olup olmadığını araştırmaktır. Bu çalışmada Parkinson hastalığının deneysel olarak taklit edilmesi için 6-OHDA ile indüklenen insan nöroblastom hücre hattı (SH-SY5Y) kullanıldı. HN'nin 6-OHDA nörotoksisitesine karşı nöroprotektif etkisi ise mitokondriyal disfonksiyon, apoptoz ve sitotoksisite parametreleri ile değerlendi. Mitokondriyal disfonksiyonun belirlenmesinde MTT, apoptoz'un belirlenmesi için kaspaz-3 ve sitotoksisitenin belirlenmesi için LDH testleri kullanıldı. İstatistiksel analiz için GraphPad Prism 8.0 programı kullanıldı ve gruplar arasındaki farklar tek yönlü ANOVA istatistikleri ile değerlendirildi. Yapılan analiz sonucunda IC50 dozu 233.7 µM olarak belirlendi. MTT testinde elde edilen bulgulara göre, HN tek başına uygulandığında proliferatif bir etkiye sahip değildi (p>0,05). Bununla birlikte, 10 µM ve 20 µM HN içeren 24 saatlik ön işleme hücrelerinin 6-OHDA nörotoksisitesine karşı koruduğu görüldü (p<0,01). LDH testinde, herhangi bir HN doz grubunda laktat dehidrojenaz salınımı saptanmadı. Kaspaz-3 testinde, 20 μM HN ile ön muamelenin, 6-OHDA nörotoksisitesine karşı kaspaz-3 seviyesini düşürdüğü, ancak istatistiksel bir fark yaratmadığı görüldü (p> 0, 05).
Humanin (HN), a 24 amino acid peptide, has been studied in several neurodegenerative diseases including Alzheimer's disease following its discovery. HN is a very bioactive peptide. Thus, it affects many intracellular and extracellular signaling mechanisms. HN, PI3k / Akt, JAK / STAT, MAPK / ERK pathways and IGFBP regulate cell growth, proliferation and survival through Bax proteins. The aim of this study was to investigate whether HN has a neuroprotective effect in an in vitro Parkinson's disease model induced by 6-hydroxydopamine (6-OHDA). In this study 6-OHDA induced human neuroblastoma cell line (SH-SY5Y) was used to experimentally mimic Parkinson's disease. The neuroprotective effect of HN against 6-OHDA neurotoxicity was evaluated by mitochondrial dysfunction, apoptosis and cytotoxicity parameters. MTT test for determination of mitochondrial dysfunction, caspase-3 test for apoptosis and LDH test for cytotoxicity were used. GraphPad Prism 8 program was used for statistical analysis and the differences between the groups were evaluated with one-way ANOVA. IC50 dose was 233.7 μM. In MTT test, HN did not have a proliferative effect when administered alone (p>0.05). However, 24 h pre-treatment cells containing 10 μM and 20 μM HN were shown to protect against 6-OHDA neurotoxicity (p<0.01). In the LDH assay, no lactate dehydrogenase release was detected in any HN dose group. In caspase-3 test, pretreatment with 20 μM HN decreased caspase-3 against 6-OHDA neurotoxicity, but did not make any statistical difference (p>0,05).
Humanin (HN), a 24 amino acid peptide, has been studied in several neurodegenerative diseases including Alzheimer's disease following its discovery. HN is a very bioactive peptide. Thus, it affects many intracellular and extracellular signaling mechanisms. HN, PI3k / Akt, JAK / STAT, MAPK / ERK pathways and IGFBP regulate cell growth, proliferation and survival through Bax proteins. The aim of this study was to investigate whether HN has a neuroprotective effect in an in vitro Parkinson's disease model induced by 6-hydroxydopamine (6-OHDA). In this study 6-OHDA induced human neuroblastoma cell line (SH-SY5Y) was used to experimentally mimic Parkinson's disease. The neuroprotective effect of HN against 6-OHDA neurotoxicity was evaluated by mitochondrial dysfunction, apoptosis and cytotoxicity parameters. MTT test for determination of mitochondrial dysfunction, caspase-3 test for apoptosis and LDH test for cytotoxicity were used. GraphPad Prism 8 program was used for statistical analysis and the differences between the groups were evaluated with one-way ANOVA. IC50 dose was 233.7 μM. In MTT test, HN did not have a proliferative effect when administered alone (p>0.05). However, 24 h pre-treatment cells containing 10 μM and 20 μM HN were shown to protect against 6-OHDA neurotoxicity (p<0.01). In the LDH assay, no lactate dehydrogenase release was detected in any HN dose group. In caspase-3 test, pretreatment with 20 μM HN decreased caspase-3 against 6-OHDA neurotoxicity, but did not make any statistical difference (p>0,05).
Açıklama
Anahtar Kelimeler
Apoptoz, Humanin, Mitokondriyal Disfonksiyon, Nöroprotektif Etki, Parkinson Hastalığı, Sitotoksisite, Apoptosis, Cytotoxicity, Mitochondrial Dysfunction, Neuroprotective Effect, Parkinson’s Disease
