İnsan T hücreli akut lenfoblastik lösemi hücrelerinin kapsaisin ile indüklenmiș apoptozunda rolü olabilecek jak/stat sinyal iletim yolağı elemanlarının gen ekspresyon profillerinin belirlemesi

AMAÇ: Dünya genelinde baharat olarak tüketilen kırmızı acı biberin içerdiği kapsaisin (trans-8-metil-N-vanilil-6-nonenamid), olası tümöro- genez ve genotoksisite özelliğinin yanı sıra; çeșitli karsinojen ile muta- jenlere antagonist olması ve tümöral hücre apoptozunun indüklemesi sebebiyle “iki yüzü keskin bıçak” bir ajan olarak tanımlanmaktadır. Ancak kapsaisin ile indüklenmiș lösemi hücre apoptozunun nedenle- ri halen araștırma konusudur. Bu çalıșmada, akut lenfoblastik lösemi hücre dizisi olan CCRF-CEM’ e kapsaisin uygulandıktan sonra apop- toz analizinin gerçekleștirilmesinin yanı sıra; JAK/STAT (Janus Kinaz/ Sinyal İleticisi Transkripsiyon Aktivatörü) sinyal iletim yolağı elemanla- rından STAT3, STAT5A, -5B, JAK2 ve IL–6’ nın mRNA seviyesindeki gen ekspresyon profillerinin çıkartılarak, kapsaisinin olası moleküler etki mekanizmasının aydınlatılması amaçlanmıștır. YÖNTEM: Kapsaisinin CCRF-CEM hücreleri üzerinde sitotoksik etki olușturduğu IC50 dozu 80 μM uygulandıktan 72 saat sonra hücre- lerin apoptotik durumu değerlendirilmiș; diğer bir hücre grubundan total RNA izole edilip cDNA sentezi gerçekleștirilmiș ve hedef genle- rin ekspresyon seviyeleri gerçek zamanlı qRT-PCR ile belirlenmiștir. Ekspresyon değișimi, madde verilmemiș kontrol grubu hücrelerle kıyaslanarak, kat değișimi ve % değișim olarak istatistiksel analiz ya- pılarak değerlendirilmiștir. BULGULAR: Kapsaisin ile muamele edilmiș lösemi hücreleriyle kontrol grubu hücrelerin apoptotik durumu kıyaslandığında, mua- mele grubu hücrelerde %37.1 oranında anlamlı bir apoptoz indük- siyonu saptanmıștır. Hedef genlere ait mRNA ekspresyon seviyeleri kıyaslandığında, STAT3, STAT5A, -5B, JAK2 ve IL-6 ekspresyon- larında sırasıyla -6.02, -49.6, -2.23, -5.53, -2.51 katlık anlamlı azal- malar saptanmıștır. SONUÇ: Kapsaisin ile indüklenmiș lösemi hücre apoptozunun ola- sı nedenlerinden biri de, JAK/STAT sinyal yolağı elemanlarından; transkripsiyon faktörü olan ve lösemide artmıș ekspresyon ser- gileyen STAT3, STAT5A ve -5B ekspresyon seviyelerinin anlamlı derecede azalmıș olması olabilir. Buna ilave olarak sinyalleșme te- tikleyicileri JAK2 ve IL-6 ekspresyonlarındaki düșüș ise; sinyalleș- menin baskılanarak lösemik hücre proliferasyonunun yavașlaması ve apoptozun indüklenmesiyle sonuçlanmıș olabilir. Bu nedenle, akut lenfoblastik lösemi tedavisinde kapsaisinin de yer alabilecek olması, teröpotik uygulama alanında yeni bir açılım yaratabilecektir.

Determination of the gene expression profiles of jak/stat cascade components for the potential role of capsaicin ınduced apoptosis of acute T-cell lymphoblastic leukemia cells

AIM: Capsaicin, an ingredient of red chili pepper consumed as spice throughout the world is defined as “double-edged sword” agent for having possible tumorigenicity/genotoxicity properties, for being an- tagonist to carcinogens/mutagens for inducing tumor cell apoptosis. However, reasons of capsaicin induced leukemic cell apoptosis are investigated. In this study, following capsaicin treatment to acute lym- phoblastic leukemia cell line CCRF-CEM, detecting apoptosis; de- termining gene expression profiles of JAK/STAT signaling pathway members STAT3, STAT5A,-5B, JAK2,IL-6 mRNA levels and clarifying possible molecular effect mechanism of capsaicin were aimed. METHODS: After 72h capsaicin treatment with IC50 dose that caused cytotoxic effect upon CCRF-CEM; an apoptosis assay was carried out; from another group of cells’, following total RNA isola- tion, cDNA synthesis was revealed, target genes’ expression lev- els were determined by real time qRT-PCR. Expressional changes were evaluated by comparing with untreated control group cells’ as fold or % change by performing statistical analyses. RESULTS: When apoptotic case of capsaicin treated and un- treated control group cells was compared, significant apoptosis induction was detected in treated group by 37.1% increase. When mRNA expression levels of capsaicin treated leukemic cells/ con- trol group cells for target genes were compared; significant down- regulations were detected in STAT3, STAT5A, -5B, JAK2 and IL-6 expressions with -6.02, -49.6, -2.23, -5.53, -2.51 fold respectively. CONCLUSION: One possible reason of capsaicin induced leu- kemic cell apoptosis might be due to the significant decrease in STAT3, STAT5A and -5B expression levels that function as tran- scription factors and exhibit an upregulated expression in leuke- mia. In addition, the down-regulation in signaling accelerators JAK2 and IL-6 expressions could be resulted in deceleration in leukemic cell proliferation and induced apoptosis due to signaling suppression. Therefore, the possibility of capsaicin to take place in the treatment of acute lymphoblastic leukemia as well might create new initiative in therapeutic application area.

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