Akbarzadeh Maryam, Rahbarghazi Reza, Nabat Elahe, Movassaghpour Ali Akbar, Shanehbandi Daruish, Faramarzian Azimi Maragheh Behnaz, Matluobi Danial, Barazvan Balal, Kazemi Masoume, Samadi Nasser, Nouri Mohammad
Stem Cell Research Center, Tabriz University of Medical Sciences, Tabriz, Iran; Department of Biochemistry and Clinical Laboratories, School of Medicine, Tabriz University of Medical Sciences, Tabriz, Iran.
Stem Cell Research Center, Tabriz University of Medical Sciences, Tabriz, Iran.
Biomed Pharmacother. 2017 Mar;87:288-295. doi: 10.1016/j.biopha.2016.12.119. Epub 2017 Jan 4.
Endogenous melatonin has numerous physiological roles on modulating the function of different organs. Recent studies revealed oncostatic and protective effects of this molecule on tumor development. In this study, we examined the impact of melatonin and key underlying mechanisms on stemness, morphology, invasiveness and viability of SKOV3 ovarian cancer cells in different types of extracellular matrix.
Cell viability was evaluated by MTT Assay. Colony-forming assay was performed by seeding 4×10 cells on different matrices in six well-plate. The percentage of cancer stem like cells was determined by flow cytometric assay after applying antibodies against stemness markers, CD133 and CD44. Different types of extracellular matrix including fibronectin, gelatin, collagen and matrigel were applied to incubate the cells in the presence of melatonin. Downstream gene expressions including VEGF and E-cadherin were determined by Real-time PCR.
Melatonin (0.1mM) decreased the percentage of viable cells up to 61.79±8.2% (p<0.05). Colony formation assay revealed the significant impact of melatonin in inhibition of colony formation in these cells. The maximum effect was shown in the cells incubated with melatonin on gelatin (p<0.05). Identification of stemness markers showed that applying matrigel caused significant increase in the percentage of cancer stem like cells compared to other types of extracellular matrix (p<0.05). However melatonin slightly diminished the number of stem cell like cells in all incubated matrices. Our results from gene expression assays revealed that melatonin induced a marked increase in E-cadherin along with decrease in VEGF expression levels (p<0.05).
Our results suggest that interaction between ovarian cancer cells and neighboring matrices determines the subsequent anti invasive activities of melatonin.
内源性褪黑素在调节不同器官功能方面具有多种生理作用。最近的研究揭示了该分子对肿瘤发展的抑癌和保护作用。在本研究中，我们研究了褪黑素的影响及其关键潜在机制对不同类型细胞外基质中SKOV3卵巢癌细胞的干性、形态、侵袭性和活力的影响。
通过MTT法评估细胞活力。将4×10个细胞接种于六孔板中的不同基质上进行集落形成试验。在应用针对干性标志物CD133和CD44的抗体后，通过流式细胞术测定癌干细胞样细胞的百分比。应用包括纤连蛋白、明胶、胶原蛋白和基质胶在内的不同类型细胞外基质，在褪黑素存在的情况下孵育细胞。通过实时PCR测定包括VEGF和E-钙黏蛋白在内的下游基因表达。
褪黑素（0.1mM）使活细胞百分比降低至61.79±8.2%（p<0.05）。集落形成试验显示褪黑素对这些细胞的集落形成有显著抑制作用。在明胶上用褪黑素孵育的细胞中显示出最大效应（p<0.05）。干性标志物鉴定表明，与其他类型的细胞外基质相比，应用基质胶导致癌干细胞样细胞百分比显著增加（p<0.05）。然而，褪黑素在所有孵育基质中略微减少了干细胞样细胞的数量。我们的基因表达试验结果表明，褪黑素诱导E-钙黏蛋白显著增加，同时VEGF表达水平降低（p<0.05）。
我们的结果表明，卵巢癌细胞与邻近基质之间的相互作用决定了褪黑素随后的抗侵袭活性。
