双氢青蒿素对食管癌细胞光动力治疗敏感性影响的研究

Photodynamic therapy (PDT) can relieve esophageal obstruction, and improve life quality of patients. However, PDT can cause the formation of new vasculature, which reduces the effect of PDT. We have confirmed that PDT could induce the expression of NF-κB and hypoxia-inducible factor-1α(HIF-1α)，and dihydroartemisinin (DHA) as a potential anticancer drug, could significantly improve the sensitivity of esophageal cancer cells to PDT. Therefore, we proposed a hypothesis: PDT could cause tumor vascularization by NF-κB/HIF-1α/VEGF pathway, while DHA could enhance the sensitivity of cancer cells to PDT by inhibition of this pathway. To test this hypothesis, we will investigate the mechanism of vascularization from the molecular, cellular and animal levels in human esophageal cancer cell lines Eca109, EC9706 and animal model. We will take measures such as real time PCR, western blot, RNA interference and so on, to explore the role of NF-κB/HIF-1α/VEGF pathway in vascularization, to verify the mechanism of sensitizing PDT. This study elaborates the vascular mechanism caused by PDT through NF-κB/HIF-1α/VEGF pathway. And this study enhances the effect of PDT on the concept of conjoined treatment, and provides new ideas for the PDT of esophageal cancer.

光动力疗法（PDT）可以解除食管梗阻，提高生存质量。但PDT可引起血管化，从而降低杀伤率。我们已证实：PDT可激活食管癌细胞中NF-κB及HIF-1α的表达，且双氢青蒿素（DHA）有抗癌活性，并可明显提高食管癌细胞PDT敏感性。为此，我们提出假说：PDT可通过NF-κB/HIF-1α/VEGF路径来引起血管化，而DHA通过抑制该路径来增强细胞对PDT的敏感性。为了验证这一假说，我们通过人食管癌细胞系Eca109、Ec9706和动物模型，采用real time PCR、western blot、RNA干扰等手段，从分子、细胞及动物水平等方面探讨NF-κB/HIF-1α/VEGF路径在PDT引起血管化中的作用，明确DHA增敏PDT的机制。本研究通过NF-κB/HIF-1α/VEGF这个新视点来阐述PDT引起血管化的机制，并通过DHA抑制该通路来增强PDT作用，为食管癌的PDT应用提供新的思路。

背景：光动力疗法（Photodynamic therapy，PDT）能够解除食管癌梗阻，提高患者的生存质量，延长生存期。然而，在各种光敏剂介导的光动力过程中，都能激活核转录因子NF-κB。而文献中已经证实，NF-κB是光动力作用的负性调节因子，抑制NF-κB活性可能是增强光动力治疗效果的一种方式。双氢青蒿素（dihydroartemisinin，DHA）是活性最强的青蒿素衍生物，在体内外可抑制多种肿瘤细胞的生长，在之前的研究中，我们已经证实DHA对体内外食管癌细胞都有抑制作用。文献报道DHA可以使NF-κB的活性降低。目的：本实验将通过检测细胞增殖、凋亡、NF-κB的活性及其调节的下游基因表达产物来探究DHA抑制NF-κB的活性能否提高光动力对食管癌细胞的治疗作用；探讨PDT是否通过NF-κB/HIF-1α/VEGF路径介导，最终引起血管化；探索DHA是否通过抑制该路径来增加食管癌细胞对PDT的敏感性。结果：体内外实验均证实，PDT组和DHA组与对照组相比较，均有抑制Eca109和Ec9706细胞增殖的作用，联合组这种抑制作用更加明显；联合组凋亡率明显高于单纯作用组。DHA能够降低NF-κB的表达。PDT组的NF-κB活性、P65、HIF-1α及VEGF的活性及蛋白表达均较对照组明显增加，DHA可以减少PDT的这些作用，沉默NF-κB后PDT对NF-κB的诱导作用也被逆转，HIF-1α蛋白和VEGF蛋白表达减少；沉默HIF-1α后VEGF蛋白表达减少。体内实验证实沉默NF-κB、HIF-1α组及DHA联合PDT组中肿瘤细胞增殖相关蛋白Ki-67表达明显减少。结论：以上实验结果提示我们，PDT可通过NF-κB/HIF-1α/VEGF路径来引起血管化，从而促进食管癌的生长；DHA可以通过抑制该路径来增加食管癌细胞对PDT的敏感性。