Ekspresi Caspase-3 pada Sel Epitel Rongga Mulut (Kb Cell Line) setelah Paparan Ekstrak Kopi

https://doi.org/10.22146/majkedgiind.8739

Suryani Hutomo(1*), Yanti Ivana Suryanto(2), Heni Susilowati(3), Agustinus Rudolf Phym(4), Devi Chretella Maheswara(5)

(1) Universitas Kristen Duta Wacana
(2) 
(3) 
(4) 
(5) Fakultas Kedokteran, Universitas Kristen Duta Wacana, Yogyakarta, Indonesia
(*) Corresponding Author

Abstract


Kopi adalah minuman yang biasa dikonsumsi oleh masyarakat sehari-hari. Telah diketahui bahwa kopi mengandung kafein seperti yang terdapat juga pada teh dan coklat. Kandungan terbanyak kafein terdapat pada kopi. Kafein mempunyai struktur kimia 1, 3, 7- trimethylxanthine dan merupakan derivat xanthine. Senyawa ini dapat menginduksi kematian sel yang mengarah pada apoptosis, namun mekanisme yang terlibat belum diketahui dengan jelas. Tingginya
konsumsi kopi di dunia yang selalu meningkat mengindikasikan perlunya dilakukan penelitian untuk mengetahui efek kafein pada epitel rongga mulut yang berkontak langsung dengan kafein. Penelitian terdahulu melaporkan bahwa
ekstrak kopi menyebabkan kerusakan sel yang sebagian besar mengarah pada apoptosis, tetapi mekanismenya belum jelas. Tujuan penelitian ini adalah untuk menganalisis mekanisme kematian sel KB yang diinduksi oleh kafein melalui
aktivasi caspase-3. Sel KB sebagai model epitel oral (5x10⁴ sel) dikultur dalam DMEM menggunakan 24 wells microplate selama 24 jam sebelum perlakuan. Sel selanjutnya dipapar dengan kafein dengan konsentrasi 100 μg/ml, 200 μg/ml, 400 μg/ml dan diinkubasi selama 24 dan 48 jam dalam DMEM. Doxorubicin (0,5625 μg/ml) digunakan sebagai kontrol positif induksi apoptosis. Teknik imunositokimia terhadap caspase-3 dilakukan pada sel setelah dipapar kafein
untuk mengamati adanya ekspresi caspase-3 sebagai ciri apoptosis. Identifikasi caspase-3 dilakukan menggunakan mikroskop fase kontras. Ekspresi protein caspase-3 terdeteksi pada sitoplasma sel KB. Hasil penelitian ini menunjukkan
adanya ekspresi caspase-3 aktif yang ditandai dengan warna cokelat dengan intensitas kuat pada sitoplasma sebagian besar sel setelah dipapar kafein dengan konsentrasi 100 μg/ml dan 200 μg/ml selama 24 jam. Disimpulkan bahwa ekstrak kopi menyebabkan apoptosis sel KB melalui jalur aktivasi caspase-3.

 

ABSTRACT: The Expression of Caspase-3 in Oral Cavity (Kb Cell Line) after Exposure to Coffee Extract. People widely consume coffee in daily meals. It is known there is caffeine found in coffee like it is found in tea and chocolate.
Caffeine is found in the greatest amount of coffee. This 1, 3, 7- trimethyl xanthine substance is a derivate of xanthine that is consumed by almost all people in the world. This substance could induce cell death that mainly is apoptosis, but how the mechanism has not been clearly understood. Considering that coffee is widely consumed in the whole world, it is necessary to conduct an experiment to find any possible effect of caffeine to oral epitel that make direct exposure to caffeine. This experiment is targeted to analyze the mechanism of cell death which caused by caffeine through activation of caspase-3. KB cells as oral epithelial model (5x10⁴ sel) were cultured in DMEM using 24 well microplate for 24 hours before treatment. Then caffeine was given with concentration of 100 μg/ml, 200 μg/ml and 400 μg/ml. Cells were then incubated for 24 and 48 hours period in DMEM. Doxorubicin (0,5625 μg/ml) was used as a positive control of apoptosis induction. Immunocytochemistry technique was then done to observe any caspase three expression as a
marker for apoptosis. Identification of active caspase-3 was then done using contrast phase microscope. The results showed expression of caspase-3 in KB cells cytoplasm which observed as high intensity of brown colored molecules in
cell cytoplasm after 100 μg/ml and 200 μg/ml caffeine exposure in 24 hours. It was concluded that coffee extract induce KB cells apoptosis through caspase-3 activation mechanism.


Keywords


kopi, kafein, sel KB, apoptosis, caspase-3; coffee, caffeine, KB cells, apoptosis, caspase-3

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DOI: https://doi.org/10.22146/majkedgiind.8739

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