Hypericum perforatum ekstraktının insan tiroid kanserine karşı antikanserojenik etkilerinin araştırılması
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Dosyalar
Tarih
2024
Yazarlar
Dergi Başlığı
Dergi ISSN
Cilt Başlığı
Yayıncı
Selçuk Üniversitesi, Fen Bilimleri Enstitüsü
Erişim Hakkı
info:eu-repo/semantics/openAccess
Özet
Kanser, ülkemizde ve dünyada başlıca hastalık nedenlerinden biridir. Genellikle somatik hücrelerdeki mutasyonlar sonucunda vücudun diğer bölgelerine yayılan, mortalite ve morbidite sebebiyle sağlığı olumsuz etkileyen bir hastalık olarak bilinmektedir. Tiroid kanseri endokrin sistemde görülen nadir bir kanser türüdür. Geleneksel tedavilerin beraberinde getirdiği ciddi yan etkiler ve yetersiz başarı oranları, yeni ve alternatif tedavi yöntemlerine ihtiyaç duyulmaktadır. Kanserin önlenmesi, tedavisi ve yan etkilerinin azaltılması için alternatif çözümler ve destekleyici tedaviler sıkça araştırılmakta ve talep görmektedir. Bitkiler kanser ve birçok hastalığın tedavisinde potansiyel olarak yer almaktadır. Hypericum perforatum bitkisinin anti-inflamatuar, antiviral, antioksidan, antialerjik ve antitümör özelliklere sahip olduğu bilinmektedir. Hypericum perforatum'un insan tiroid kanseri TT hücre hattı üzerindeki antioksidan ve antikanserojen etkilerini belirlemek için öncelikle Hypericum perforatum matanol ekstraktı (HPME) elde edildi. HPME'nin antioksidan aktivitesi toplam fenolik madde miktar tayini, DPPH serbest radikal temizleme aktivitesi, ABTS katyon giderim aktivitesi ve CU (II) indirgeyici aktivite (CUPRAC) yöntemleri kullanılarak spektrofotometrik olarak belirlendi. Hypericum perforatum'un fenolik ve flavonoid bileşen profili LC-MS/MS ile belirlendi. Hypericum perforatum'un antikarsinojenik etkileri in vitro koşullar altında tiroid kanseri hücresi ve sağlıklı epitel hücre üzerindeki sitotoksik etkileri Alamar Mavisi reaktifi kullanılarak bulundu. İnsan tiroid kanseri TT hücreleri üzerindeki antimetastatik özelliğini belirlemek için in vitro yara iyileştirme deneyi yapıldı ve TT hücrelerinin farklı dokulardaki hayatta kalma potansiyelinin HPME aracılığıyla inhibisyonunu belirlemek için in vitro koloni oluşturma deneyi yapıldı. Aynı zamanda HPME'nin hücre içerisindeki lokalizasyonunu tespit etmek için hücre içi floresans biyogörüntüleme deneyi yapıldı. HPME'nin insan tiroid kanseri hücreleri üzerindeki genlerin ve proteinlerin ekspresyonunda meydana getirdiği değişimler, qRT-PCR ve Western Blot yöntemiyle bulundu. HPME'nin tiroid kanserindeki apoptoz etkileri ise flow sitometri yöntemiyle Annexin V-APC ve 7-AAD boyaları kullanılarak belirlendi. Yapılan çalışmalar sonucunda yüksek antioksidan etkinlik gösteren HPME'nin sitoplazmada lokalize olduğu bulundu. HPME'nin tiroid kanseri ve sağlıklı epitel hücre HACAT üzerindeki hücre inhibisyonunun sigmoidal grafiğinden yararlanılarak bulunan IC50 değerleri sırasıyla 13.61 µg/mL ve 171.9 µg/mL olarak hesaplandı. Sonuçlara göre HPME TT hücrelerini inhibe ederken HACAT hücreleri üzerinde de minimum sitotoksik etki gösterdiği bulundu. HPME, TT Hücrelerinin koloni oluşumunu ve göçünü önemli ölçüde ***p<0.0001 inhibe ettiği bulundu. Apoptoz yolağında rol alan Bax, BcL-2, Kaspaz-3, Kaspaz-9, Kaspaz-12, Apoe ve Clu'nun gen ve protein ekspresyonlarını önemli ölçüde değiştirdiği bulundu. Aynı zamanda HPME TT hücrelerini control hücresine göre geç apoptozu %18.92 artırdığı görüldü. Bu tez çalışmasında sonuç olarak; HPME'nin hücre sitoplazmasına lokalize olarak etkinlik gösterdiği, gen, protein ve apoptoz çalışmaları sonucunda HPME'nin TT hücreleri üzerinde sitotoksik etkilerinin moleküler düzeyde mekanizması belirlendi. Elde edilen sonuçlar doğrultusunda HPME'nin yüksek antioksidan kapasiteye sahip olduğu ve HPME'nin tiroid kanseri üzerinde metastaz ve koloni oluşumunun inhibisyonuna neden olarak tiroid kanseri tedavisinde alternatif bir tedavi yöntemi olabileceği öngörülmektedir.
Cancer is one of the main causes of disease in our country and in the world. It is generally known as a disease that spreads to other parts of the body as a result of mutations in somatic cells and negatively affects health due to mortality and morbidity. Thyroid cancer is a rare type of cancer seen in the endocrine system. Serious side effects and insufficient success rates caused by traditional treatments necessitate the need for new and alternative treatment methods. Alternative solutions and supportive treatments for the prevention, treatment and reduction of side effects of cancer are frequently researched and in demand. Plants are potentially involved in the treatment of cancer and many diseases. Hypericum perforatum plant is known to have anti-inflammatory, antiviral, antioxidant, antiallergic and antitumor properties. To determine the antioxidant and anticarcinogenic effects of Hypericum perforatum on the human thyroid cancer TT cell line, firstly, Hypericum perforatum methanol extract (HPME) was obtained. The antioxidant activity of HPME was determined spectrophotometrically using total phenolic substance quantification, DPPH free radical scavenging activity, ABTS cation removal activity and CU(II) reducing activity (CUPRAC) methods. The phenolic and flavonoid component profile of Hypericum perforatum was determined by LC-MS/MS. The anticarcinogenic effects of Hypericum perforatum were found using the Alamar Blue reagent under in vitro conditions and its cytotoxic effects on thyroid cancer cells and healthy epithelial cells. In vitro wound healing assay was performed to determine the antimetastatic property of human thyroid cancer on TT cells, and in vitro colony formation assay was performed to determine the inhibition of survival potential of TT cells in different tissues via HPME. At the same time, an intracellular fluorescence bioimaging experiment was performed to detect the localization of HPME within the cell. The changes caused by HPME in the expression of genes and proteins on human thyroid cancer cells were found by qRT-PCR and Western Blot method. The apoptosis effects of HPME in thyroid cancer were determined by flow cytometry using Annexin V-APC and 7-AAD dyes. As a result of the studies, it was found that HPME, which shows high antioxidant activity, is localized in the cytoplasm. Using the sigmoidal graph of HPME's cell inhibition on thyroid cancer and healthy epithelial cell HACAT, the IC50 values were calculated as 13.61 µg/mL and 171.9 µg/mL, respectively. According to the results, it was found that while HPME inhibited TT cells, it also had a minimal cytotoxic effect on HACAT cells. HPME was found to significantly inhibit colony formation and migration of TT Cells ***p<0.0001. It was found that Bax, BcL-2, Caspase-3, Caspase-9, Caspase-12, Apoe and Clu, which are involved in the apoptosis pathway, significantly changed gene and protein expressions. At the same time, it was observed that HPME TT cells increased late apoptosis by 18.92% compared to control cells. As a result of this thesis study; As a result of gene, protein, and apoptosis studies, the mechanism of HPME's cytotoxic effects on TT cells was determined at the molecular level, where HPME showed activity localized to the cell cytoplasm. In line with the results obtained, it is predicted that HPME has a high antioxidant capacity and HPME may be an alternative treatment method in the treatment of thyroid cancer by causing the inhibition of metastasis and colony formation in thyroid cancer.
Cancer is one of the main causes of disease in our country and in the world. It is generally known as a disease that spreads to other parts of the body as a result of mutations in somatic cells and negatively affects health due to mortality and morbidity. Thyroid cancer is a rare type of cancer seen in the endocrine system. Serious side effects and insufficient success rates caused by traditional treatments necessitate the need for new and alternative treatment methods. Alternative solutions and supportive treatments for the prevention, treatment and reduction of side effects of cancer are frequently researched and in demand. Plants are potentially involved in the treatment of cancer and many diseases. Hypericum perforatum plant is known to have anti-inflammatory, antiviral, antioxidant, antiallergic and antitumor properties. To determine the antioxidant and anticarcinogenic effects of Hypericum perforatum on the human thyroid cancer TT cell line, firstly, Hypericum perforatum methanol extract (HPME) was obtained. The antioxidant activity of HPME was determined spectrophotometrically using total phenolic substance quantification, DPPH free radical scavenging activity, ABTS cation removal activity and CU(II) reducing activity (CUPRAC) methods. The phenolic and flavonoid component profile of Hypericum perforatum was determined by LC-MS/MS. The anticarcinogenic effects of Hypericum perforatum were found using the Alamar Blue reagent under in vitro conditions and its cytotoxic effects on thyroid cancer cells and healthy epithelial cells. In vitro wound healing assay was performed to determine the antimetastatic property of human thyroid cancer on TT cells, and in vitro colony formation assay was performed to determine the inhibition of survival potential of TT cells in different tissues via HPME. At the same time, an intracellular fluorescence bioimaging experiment was performed to detect the localization of HPME within the cell. The changes caused by HPME in the expression of genes and proteins on human thyroid cancer cells were found by qRT-PCR and Western Blot method. The apoptosis effects of HPME in thyroid cancer were determined by flow cytometry using Annexin V-APC and 7-AAD dyes. As a result of the studies, it was found that HPME, which shows high antioxidant activity, is localized in the cytoplasm. Using the sigmoidal graph of HPME's cell inhibition on thyroid cancer and healthy epithelial cell HACAT, the IC50 values were calculated as 13.61 µg/mL and 171.9 µg/mL, respectively. According to the results, it was found that while HPME inhibited TT cells, it also had a minimal cytotoxic effect on HACAT cells. HPME was found to significantly inhibit colony formation and migration of TT Cells ***p<0.0001. It was found that Bax, BcL-2, Caspase-3, Caspase-9, Caspase-12, Apoe and Clu, which are involved in the apoptosis pathway, significantly changed gene and protein expressions. At the same time, it was observed that HPME TT cells increased late apoptosis by 18.92% compared to control cells. As a result of this thesis study; As a result of gene, protein, and apoptosis studies, the mechanism of HPME's cytotoxic effects on TT cells was determined at the molecular level, where HPME showed activity localized to the cell cytoplasm. In line with the results obtained, it is predicted that HPME has a high antioxidant capacity and HPME may be an alternative treatment method in the treatment of thyroid cancer by causing the inhibition of metastasis and colony formation in thyroid cancer.
Açıklama
Anahtar Kelimeler
Antioksidan Aktivite, Apoptoz, Gen/Protein Ekspresyonu, Hypericum perforatum, Sitotoksisite, Tiroid Kanseri, Antioxidant Activity, Apoptosis, Gene/Protein Expression, Cytotoxicity, Thyroid Cancer
Kaynak
WoS Q Değeri
Scopus Q Değeri
Cilt
Sayı
Künye
Altunkaya, G. (2024). Hypericum perforatum ekstraktının insan tiroid kanserine karşı antikanserojenik etkilerinin araştırılması. (Yüksek Lisans Tezi). Selçuk Üniversitesi, Fen Bilimleri Enstitüsü, Konya.
