Reference gene expression in human osteosarcoma cell lines treated by EGB and CTX

Our findings provide a suitable approach through which qRT-PCR can be applied to investigate the pharmacological effects and the molecular mechanisms of CTX and EGB on human osteosarcoma.

📌 3 Maddede Bu Makale

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Amaç & Hipotez:The selection of stably expressed reference genes is crucial for quantitative real-time polymerase chain reaction (qRT-PCR) assay when assessing osteosarcoma treated with cyclophosphamide (CTX) and Ginkgo biloba L. extract (EGB).

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Temel Bulgular & Veri:We used qRT-PCR assay to determine the expression levels of 18S, ACTB, ALAS1, GAPDH, TBP, HPRT1, RPL29, HMBS, PPIA, PUM1, GUSB, and B2M in the cell lines MG-63, Saos-2, and U2OS, which were treated with CTX or EGB or both.

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Sonuç & Katkı:Our findings provide a suitable approach through which qRT-PCR can be applied to investigate the pharmacological effects and the molecular mechanisms of CTX and EGB on human osteosarcoma.

🔬Tür:Araştırma Makalesi

📍Kapsam:Genişletilmiş Akademik Örneklem

⚙️Değişkenler:Alan Parametreleri

Abstract

The selection of stably expressed reference genes is crucial for quantitative real-time polymerase chain reaction (qRT-PCR) assay when assessing osteosarcoma treated with cyclophosphamide (CTX) and Ginkgo biloba L. extract (EGB). We aimed to identify the reference genes that are stably expressed in human osteosarcoma cell lines after the above treatments. We used qRT-PCR assay to determine the expression levels of 18S, ACTB, ALAS1, GAPDH, TBP, HPRT1, RPL29, HMBS, PPIA, PUM1, GUSB, and B2M in the cell lines MG-63, Saos-2, and U2OS, which were treated with CTX or EGB or both. We analyzed the expression stability of these genes using geNorm, NormFinder, and BestKeeper. The optimal reference gene combinations were 18S + PPIA + HPRT1 in the total group, HMBS + ALAS1 + 18S in the EGB-treated group, PPIA + RPL29 in the EGB + CTX-treated group, PPIA + HPRT1 in the MG-63 group, 18S + PPIA in the Saos-2 group, and HPRT1 + ALAS1 + GAPDH in the U2OS group. We conclude that no single reference gene was most stably expressed in all three treated cell lines. Our findings provide a suitable approach through which qRT-PCR can be applied to investigate the pharmacological effects and the molecular mechanisms of CTX and EGB on human osteosarcoma.

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Bu Makaleye Atıf Yap

YANG, Q., WANG, Z., REN, M., SONG, Y., WANG, A., WANG, Q., WANG, X., HAO, S., DU, Z., ZHANG, G., WANG, J. (2022). Reference gene expression in human osteosarcoma cell lines treated by EGB and CTX. Turkish Journal of Biology, Cilt: 41 - Sayı: 3, 439–447. https://ajindex.com/articles/600320

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Kaynakça (10)

Ali H, Du Z, Li X, Yang Q, Zhang YC, Wu M, Li Y, Zhang G (2015). Identification of suitable reference genes for gene expression studies using quantitative polymerase chain reaction in lung cancer in vitro. Mol Med Rep 11: 3767-3773.
Andersen CL, Jensen JL, Orntoft TF (2004). Normalization of realtime quantitative reverse transcription-PCR data: a modelbased variance estimation approach to identify genes suited for normalization, applied to bladder and colon cancer data sets. Cancer Res 64: 5245-5250.
Bruge F, Venditti E, Tiano L, Littarru GP, Damiani E (2011). Reference gene validation for qPCR on normoxia- and hypoxia-cultured human dermal fibroblasts exposed to UVA: is beta-actin a reliable normalizer for photoaging studies? J Biotechnol 156: 153-162.
Chang E, Shi S, Liu J, Cheng T, Xue L, Yang X, Yang W, Lan Q, Jiang Z (2012). Selection of reference genes for quantitative gene expression studies in Platycladus orientalis (Cupressaceae) using real-time PCR. PLoS One 7: e33278. Declercq H, Van Den Vreken N, De Maeyer E, Verbeeck R, Schacht E, De Ridder L, Cornelissen M (2004). Isolation, proliferation and differentiation of osteoblastic cells to study cell/biomaterial interactions: comparison of different isolation techniques and source. Biomaterials 25: 757-768.
Derveaux S, Vandesompele J, Hellemans J (2010). How to do successful gene expression analysis using real-time PCR. Methods 50: 227-230.
Dias MC, Furtado KS, Rodrigues MA, Barbisan LF (2013). Effects of Ginkgo biloba on chemically-induced mammary tumors in rats receiving tamoxifen. BMC Complement Altern Med 13: 93. Grigolo B, Lisignoli G, Toneguzzi S, Mazzetti I, Facchini A (1998). Copper/zinc superoxide dismutase expression by different human osteosarcoma cell lines. Anticancer Res 18: 1175-1180.
Hauns B, Haring B, Kohler S, Mross K, Unger C (2001). Phase II study of combined 5-fluorouracil/ Ginkgo biloba extract (GBE 761 ONC) therapy in 5-fluorouracil pretreated patients with advanced colorectal cancer. Phytother Res 15: 34-38.
He YX, Zhang Y, Yang Q, Wang C, Su G (2015). Selection of suitable reference genes for reverse transcription-quantitative polymerase chain reaction analysis of neuronal cells differentiated from bone mesenchymal stem cells. Mol Med Rep 12: 2291-2300.
Huan P, Maosheng T, Zhiqian H, Long C, Xiaojun Y (2012). TLR4 expression in normal gallbladder, chronic cholecystitis and gallbladder carcinoma. Hepatogastroenterology 59: 42-46.
Li X, Yang Q, Bai J, Xuan Y, Wang Y (2015a). I