GREEN SYNTHESIS OF GOLD NANOPARTICLES USING VITIS VINIFERA SEED EXTRACT AND EVALUATION OF ANTI-CANCER PROPERTIES IN COLON CANCER (HT-29) CELLS

Öz

Objective In recent years, metallic nanoparticles have attracted the attention in the field of nanomedicine with their antioxidant, antimicrobial and anticancer activities. Especially, metal nanoparticles produced by green synthesis method from medicinal plants with known pharmacological efficacy in cancer treatment have been spotlighted by researchers.. Gold nanoparticles; become prominentamong metal nanoparticles with their unique chemical, physical and non-toxic properties. In this study, it was aimed to perform green synthesis of gold nanoparticles using Vitis vinifera (grape seed) aqueous extract and to evaluate cytotoxic, anti-proliferative and apoptotic effects of gold nanoparticles (Vv-AuNP) alone and with Gemcitabine co-administration on HT-29 human colon cancer cell line. Materials and Methods Green synthesis of gold nanoparticles was carried out using the aqueous extract of Vitis vinifera. Vv-AuNPs were characterized by various analytical techniques such as UV-visible spectrophotometer, scanning electron microscope (SEM) and Energy Dispersive X-ray Spectroscopy (EDX). Vv-AuNPs were treated with HT-29 cells at a dose range of 0–80 μg / mL for 72 hours, alone and in combination with Gemcitabine (100 μg / mL or 200 μg / mL fixed dose). Cytotoxic effects were evaluated by MTT assay. In addition, the antiproliferative effects of Vv-AuNPs at increasing doses (0-400 μg / mL) alone and combined with Gemcitabine were determined by the BrdU test. After treatment of gold nanoparticles with HT-29 cells, the level of Caspase-3 activation was analyzed by Western-blot method. Results According to MTT results, when the dose of Vv- AuNPs was increased to 80 μg / mL, it was determined that HT-29 cells did not show the expected level of cytotoxic effect. According to the results of the BrdU cell proliferation test, IC50 values of Vv-AuNPs alone and combined with Gemcitabine were determined as 147.9 and 39.43 μg / mL, respectively. It was observed that the activation of caspase-3 due to Vv- AuNP doses increased compared to control, and its combined use with Gemcitabine reinforced this effect. Conclusion As a result, green synthesized AuNPs exhibited anti-cancer properties on colon cancer cells in vitro. The results of this study show that Vv-AuNPs can be considered as a potential option for colon cancer treatment.

Anahtar Kelimeler

• Vitis vinifera
• Gold Nanoparticle
• Gemcitabine
• Colon Cancer
• HT-29

VİTİS VİNİFERA ÇEKİRDEK ÖZÜTÜ KULLANILARAK ALTIN NANOPARTİKÜLLERİN YEŞİL SENTEZİ VE KOLON KANSERİ (HT-29) HÜCRELERİNDE ANTİ- KANSER ÖZELLİKLERİNİN DEĞERLENDİRİLMESİ

Öz

Amaç Son yıllarda metalik nanopartiküller antioksidan, antimikrobiyal ve antikanser etkinlikleri ile nanotıp alanında oldukça dikkat çekmektedir. Özellikle, kanser tedavisinde farmakolojik etkinliği bilinen tıbbi bitkilerden yeşil sentez metodu ile üretilen metal nanopartiküller araştırmacıların ilgi odağı haline gelmiştir. Altın nanopartiküller; kendilerine özgü kimyasal, fiziksel ve nontoksik özellikleri ile ön plana çıkan metal nanopartiküllerden biridir. Çalışmamızda, üzüm (Vitis vinifera) çekirdeği sulu ekstraktı kullanılarak yeşil sentezi gerçekleştirilen altın nanopartiküllerin (Vv-AuNP) tek başına ve kemoterapötik bir ajan olan Gemsitabin ile kombine kullanımının HT-29 kolon kanseri hücre hattı üzerindeki sitotoksik, anti-proliferatif ve apoptotik etkilerinin değerlendirilmesi amaçlanmıştır. Gereç ve Yöntem Altın nanopartiküllerin yeşil sentezi, Vitis vinifera sulu özütü kullanılarak gerçekleştirildi. Vv-AuNP’lerin UV-görünür spektrofotometre, taramalı elektron mikroskobu (SEM) ve Enerji Dağılımlı X-ışını Spektroskopisi (EDX) gibi çeşitli analitik tekniklerle karakterizasyonu yapıldı. Vv-AuNP'ler 0–80 μg/mL doz aralığında tek başına ve Gemsitabin (100 μg/mL veya 200 μg/mL sabit doz) ile kombine olacak şekilde HT- 29 hücreleri ile 72 saat süresince muamele edildi. Sitotoksik etkileri MTT testi ile değerlendirildi. Ayrıca, HT-29 hücreleri üzerine Vv-AuNP’lerin artan dozlarda (0-400μg/mL) tek başına ve Gemsitabin ile kombine antiproliferatif etkileri ise BrdU testi ile belirlendi. Altın nanopartiküllerin HT-29 hücreleri ile muamelesi sonrası Kaspaz-3 aktivasyon düzeyi Western-blot yöntemi ile analiz edildi. Bulgular MTT sonuçları değerlendirildiğinde Vv-AuNP’lerin 80 μg/mL doza kadar çıkıldığında HT-29 hücrelerinde istenilen düzeyde sitotoksik etkiler göstermediği belirlendi. BrdU hücre proliferasyon testi sonuçlarına dayalı olarak, Vv-AuNP’lerin tek başına ve Gemsitabin ile birlikte uygulamalarında IC50 değerleri sırasıyla 147.9 ve 39.43 μg/mL olarak belirlendi. Vv-AuNP dozlarına bağlı kaspaz-3 aktivasyonunun kontrole kıyasla arttığı, Gemsitabin ile kombine kullanımının bu etkiyi güçlendirdiği gözlendi. Sonuç Sonuç olarak, yeşil sentezlenmiş AuNP’ler in vitro koşullarda kolon kanseri hücreleri üzerinde anti-kanser özellikler sergilemiştir. Bu çalışmanın sonuçları, Vv-AuNP'lerin kolon kanseri tedavisi için potansiyel bir seçenek olarak düşünülebileceğini göstermektedir.

Anahtar Kelimeler

• Vitis vinifera
• Altın Nanopartikül
• Gemsitabin
• Kolon Kanseri
• HT-29

Kaynakça

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