Betülinik asit EAG1 Baskılanması Yoluyla SH-SY5Y Nöroblastom Hücrelerinde Apoptoz ve G2/M Hücre Döngüsü Durmasını İndükler

Öz

Amaç: Nöroblastoma, çocuklarda en sık görülen ekstrakraniyal solid tümör olup özellikle ileri evrelerde yüksek mortaliteye sahiptir. Tekrarlama ve ilaç direnci gibi mevcut tedavi yöntemlerinin sınırlılıkları nedeniyle, tümöre özgü etkisi yüksek ve normal hücrelere minimum toksisite gösteren yeni ajanlara ihtiyaç vardır. Doğal bir pentasiklik triterpen olan betulinik asit (BA), seçici antitümör özellikleri nedeniyle ilgi çekmektedir. Gereç ve Yöntemler: Bu çalışmada, BA’nın SH-SY5Y nöroblastoma ve epitelyal hücre hattı, Vero, üzerindeki sitotoksik ve moleküler etkileri incelendi. Hücre canlılığı ise MTT testi ile değerlendirildi. Morfolojik değişiklikler mikroskop ile gözlemlendi. Apoptoz (Bax, Bcl-2, Caspase-3,p53, p21) ve hücre döngüsü (CDK1, Cyclin B1, Ki-67, EAG1) ile ilişkili genlerin ekspresyon seviyeleri qPCR ile analiz edildi. Bulgular: BA, her iki hücre hattında da zaman ve doz bağımlı olarak hücre canlılığını anlamlı şekilde azalttı; SH-SY5Y hücreleri daha duyarlı bulundu (48 saat için IC₅₀=75 μM). SH-SY5Y hücrelerinde hücre büzülmesi ve hücre yoğunluğunda azalma gibi morfolojik değişiklikler belirgindi. Bu hücre hattında BA tedavisi pro-apoptotik genlerin (Bax, Caspase-3, p53, p21) ifadesini artırırken, anti-apoptotik ve proliferasyonla ilişkili genlerin (Bcl-2, CDK1, Cyclin B1, Ki-67, EAG1) ifadesini azalttı (p<0,05). Vero hücrelerinde ise değişiklikler daha ılımlı olup sınırlı sitotoksisite gözlendi. Sonuç: Betulinik asit, nöroblastoma hücrelerinde seçici olarak apoptozu indükleyip hücre döngüsünü inhibe ederken, normal hücreleri korumaktadır. EAG1 genindeki azalma, BA’nın sitotoksik etkisinde voltaj bağımlı potasyum kanallarının rol oynayabileceğini göstermektedir. Bu bulgular, BA’nın hedefe yönelik nöroblastoma tedavisinde seçici bir ajan olarak terapötik potansiyelini ortaya koymaktadır

Anahtar Kelimeler

• Betulinik asit
• nöroblastoma
• SH-SY5Y
• apoptoz
• hücre döngüsü
• EAG1

Betulinic Acid Induces Apoptosis and G2/M Cell Cycle Arrest in SH-SY5Y Neuroblastoma Cells via EAG1 Suppression

Öz

Aim: Neuroblastoma is the most common extracranial solid tumour in children and is associated with high mortality, particularly in advanced stages. Given the limitations of current treatment approaches, such as recurrence and drug resistance, novel agents with tumour-specific effects and minimal toxicity to normal cells are needed. Betulinic Acid (BA), a naturally occurring pentacyclic triterpene, has gained attention due to its selective antitumor properties. Material and Methods: This study investigated the cytotoxic and molecular effects of BA on SH-SY5Y neuroblastoma and epithelial cell lines, Vero. Cell viability was assessed using the MTT assay. Microscopic observations were conducted to evaluate morphological changes. Quantitative real-time PCR (qPCR) was performed to analyse the gene expression changes associated with apoptosis (Bax, Bcl-2, Caspase-3, p53, p21) and the cell cycle (CDK1, Cyclin B1, Ki-67, EAG1). Results: BA significantly reduced cell viability in both cell lines in a time- and dose-dependent manner, with SH-SY5Y cells showing greater sensitivity (IC₅₀=75 μM at 48 hours). Morphological changes such as cell shrinkage and reduced cell density were more pronounced in SH-SY5Y cells. In this cell line, BA treatment significantly upregulated pro-apoptotic genes (Bax, Caspase-3, p53, p21) and downregulated anti-apoptotic and proliferation-related genes (Bcl-2, CDK1, Cyclin B1, Ki-67, EAG1) (p<0.05). In contrast, Vero cells exhibited moderate changes, indicating limited cytotoxicity. Conclusion: Betulinic acid selectively induces apoptosis and inhibits cell cycle progression in neuroblastoma cells while sparing normal cells. The downregulation of EAG1 suggests that voltage-gated potassium channels may be involved in the mechanism of BA-mediated cytotoxicity. These findings highlight the therapeutic potential of BA as a selective agent for targeted neuroblastoma treatment

Anahtar Kelimeler

• Betulinic acid
• neuroblastoma
• SH-SY5Y
• apoptosis
• cell cycle
• EAG1

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