HCT-116 ve HT-29 kolon kanseri hücre hattında kuersetinin hücre ölüm yolakları üzerine etkisinin incelenmesi
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Dosyalar
Tarih
2023
Yazarlar
Dergi Başlığı
Dergi ISSN
Cilt Başlığı
Yayıncı
Düzce Üniversitesi
Erişim Hakkı
info:eu-repo/semantics/openAccess
Özet
Kanser çok basamaklı, uzun süreli bir süreçtir ve sağlıksız bir hücrenin kontrolsüz bölünüp büyümesinin ardından tümör üreten hücre ölümünün yokluğu ile karakterizedir. Kolorektal kanserler dünyada en sık görülen kanserlerdendir Literatüre kalın bağırsağın iç yüzeyindeki hücrelerin kontrolsüz bölünmesiyle karakterize, yaygın gastrointestinal malign tümör olarak geçmiştir. Ülkemizde oluşan kanser türleri arasında ilk beşte yer alır ve görülme sıklığı giderek artmaktadır. Kanser araştırmalarındaki önemli zorluklardan biri sağlıklı hücreler bozulmadan, kanser hücrelerinin etkili bir şekilde nasıl öldürüleceğidir ve bu noktada kanserli hücreleri tedavi etmek için ölüm yolaklarının bilinmesi kritik bir önem taşır. Son yıllarda keşfedilmiş olan ferroptoz, demir bağımlılığı ile ortaya çıkan yeni bir programlanmış hücre ölümü türü olarak bilinir ve kanser hücreleri büyümeyi sağlamak için normal hücrelere kıyasla daha fazla demir talebi sergilediğinden kanser hücrelerinde ferroptozun indüklenmesinin çoklu kemoterapötiklerin direncini tersine çevirdiği ve kanser direncini azalttığı doğrulanmıştır. Bu çalışmalar ferroptozun tömör oluşumunda, gelişmesinde önemli bir düzenleyici olarak rol oynadığını göstermiştir. Flavonoidlerden kuersetin kanser önleme, ilerlemesini durdurma ve geleneksel kanser tedavilerinin yan etkilerini azaltılmasında başarılı olduğu bilinen metabolittir. Kuersetinin kolon kanserini engelleyici etkilerinin hücre ölüm yolakları üzerine etkilerinin ortaya çıkarılabilmesi için yapılan bu çalışmada kuersetin molekülünün kolon kanseri hücre hatlarında kemoterapik ajan ile kombine kullanımlarına ve tek kullanımlarının hücre canlılığı üzerindeki etkilerinin belirlenerek hücre ölüm mekanizmalarından apoptoz ve ferroptoz üzerindeki potansiyel etkilerinin değerlendirilmesi amaçlandı. Çalışmamızda öncelikle kuersetin ve kemoterapik ajan sitotoksik konsantrasyonları 3-(4,5-dimetiltiazol-2-il)- 2,5-difenil tetrazolyum bromür (MTT) testi ile HT-29 ve HCT-116 hücrelerinde 14 belirlendi. Daha sonra sitotoksik kuersetin ve kemoterapik ajan ile 24 saat inkübe edilen HCT-116 ve HT-29 hücreleri Apoptotik Proteaz Aktive Edici Faktör-1 (APAF-1), İnsan Kaspaz-3 Proteini (CASP-3), Uzun Zincirli Yağ Asidi-KoA Ligaz (ACSL4) ve Glutatyon Peroksidaz 4 (GPx4) seviyeleri enzime bağlı immünosorbent testi yöntemiyle (ELİSA) belirlendi. Çalışmada kuersetin molekülünün her iki kolon kanseri hücresinde tek başına APAF-1 ve CASP-3 biyobelirteçlerini kontrol grubuna göre arttırarak apoptotik yolağı aktive ettiği belirlenmiştir. GPx4 ve ACSL4 biyobelirteçlerinde ise kombine uygulamanın ACSL4'ü arttırıp GPx4'ü azaltarak ferroptotik yolağı indükleyip kanserli hücrede sinerjik etki yaptığı belirlenmiştir. Yapılan bu çalışma sonucundaki veriler kolon kanserine karşı kombine veya diğer alternatif yaklaşımlarına temel veri sağlayıp kanser tedavisi için ferroptoz sinyal yolağının daha detaylı araştırılmasında etken maddenin kullanımına yönelik çalışmalara katkı sağlayabilir.
Cancer is a multi-step and long-term process and is characterized by the uncontrolled division and growth of an unhealthy cell followed by the absence of tumor-producing cell death. Colorectal cancers are among the most common cancers in the world. It is one of the first five types of cancer in our country and its incidence is increasing. One of the major challenges in cancer research is how to kill cancer cells effectively before healthy cells are destroyed, and at this point, knowing the death pathways is critical to treating cancerous cells. Discovered in recent years, ferroptosis is known as a new type of programmed cell death induced by iron dependence, and it has been confirmed that induction of ferroptosis in cancer cells reverses the resistance of multiple chemotherapeutics and reduces cancer resistance, as cancer cells exhibit greater demand for iron than normal cells to promote growth. These studies have shown that ferroptosis plays an important role as an important regulator in tumor formation and development. Quercetin, one of the flavonoids, is a metabolite known to be successful in preventing cancer, stopping its progression, and reducing the side effects of conventional cancer treatments. In this study, which was conducted to reveal the effects of quercetin inhibitory effects on colon cancer, on cell death pathways, it was aimed to evaluate the potential effects of quercetin molecule on apoptosis and ferroptosis by determining the effects of quercetin molecule in combination with a chemotherapy agent in colon cancer cell lines and the effects of single use on cell viability. In our study, first of all, quercetin and chemotherapeutic agent cytotoxic concentrations were determined in HT- 29 and HCT-116 cells by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) test. HCT-116 and HT-29 cells, which were then incubated for 24 hours with cytotoxic quercetin and chemotherapeutic agent, Apoptotic Protease Activating 16 Factor-1(APAF-1), Human Caspase-3 Protein (CASP-3), Long Chain Fatty Acid-CoA Ligase (ACSL4) and Glutathione Peroxidase 4 (GPx4) levels were determined by enzyme-linked immunosorbent assay method (ELISA). In the study, it was determined that the quercetin molecule alone activates the apoptotic pathway by increasing APAF-1 and CASP-3 biomarkers in both colon cancer cells compared to the control group. In GPx4 and ACSL4 biomarkers, it was determined that the combined application had a synergistic effect on cancer cells by increasing ACSL4 and decreasing GPx4, inducing the ferroptotic pathway. The data from this study can provide basic data for combined or other alternative approaches against colon cancer. Further investigation of the ferroptosis signaling pathway for cancer treatment may contribute to studies on the use of the active substance.
Cancer is a multi-step and long-term process and is characterized by the uncontrolled division and growth of an unhealthy cell followed by the absence of tumor-producing cell death. Colorectal cancers are among the most common cancers in the world. It is one of the first five types of cancer in our country and its incidence is increasing. One of the major challenges in cancer research is how to kill cancer cells effectively before healthy cells are destroyed, and at this point, knowing the death pathways is critical to treating cancerous cells. Discovered in recent years, ferroptosis is known as a new type of programmed cell death induced by iron dependence, and it has been confirmed that induction of ferroptosis in cancer cells reverses the resistance of multiple chemotherapeutics and reduces cancer resistance, as cancer cells exhibit greater demand for iron than normal cells to promote growth. These studies have shown that ferroptosis plays an important role as an important regulator in tumor formation and development. Quercetin, one of the flavonoids, is a metabolite known to be successful in preventing cancer, stopping its progression, and reducing the side effects of conventional cancer treatments. In this study, which was conducted to reveal the effects of quercetin inhibitory effects on colon cancer, on cell death pathways, it was aimed to evaluate the potential effects of quercetin molecule on apoptosis and ferroptosis by determining the effects of quercetin molecule in combination with a chemotherapy agent in colon cancer cell lines and the effects of single use on cell viability. In our study, first of all, quercetin and chemotherapeutic agent cytotoxic concentrations were determined in HT- 29 and HCT-116 cells by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) test. HCT-116 and HT-29 cells, which were then incubated for 24 hours with cytotoxic quercetin and chemotherapeutic agent, Apoptotic Protease Activating 16 Factor-1(APAF-1), Human Caspase-3 Protein (CASP-3), Long Chain Fatty Acid-CoA Ligase (ACSL4) and Glutathione Peroxidase 4 (GPx4) levels were determined by enzyme-linked immunosorbent assay method (ELISA). In the study, it was determined that the quercetin molecule alone activates the apoptotic pathway by increasing APAF-1 and CASP-3 biomarkers in both colon cancer cells compared to the control group. In GPx4 and ACSL4 biomarkers, it was determined that the combined application had a synergistic effect on cancer cells by increasing ACSL4 and decreasing GPx4, inducing the ferroptotic pathway. The data from this study can provide basic data for combined or other alternative approaches against colon cancer. Further investigation of the ferroptosis signaling pathway for cancer treatment may contribute to studies on the use of the active substance.
Açıklama
Anahtar Kelimeler
Biyokimya, Biochemistry
