姜黄素/明胶纳米纤维膜抑制胃癌的作用研究

Inhibitory effect of curcumin/gelatin nanofibrous membrane on gastric cancer

摘要:

背景与目的 姜黄素是一种安全且廉价的天然化合物，但其水溶性差、生物利用度低，严重限制了其在胃癌治疗中的应用。本研究构建一种可植入的姜黄素/明胶纳米纤维膜，以实现药物的局部缓释，并探究其抑制胃癌细胞增殖的分子机制。方法 采用静电纺丝技术制备姜黄素/明胶纳米纤维膜，通过扫描电镜观察其形貌，高效液相色谱-质谱联用法测定体外药物释放曲线。以人胃黏膜细胞GES-1评估生物安全性；以人胃癌细胞系MKN-45、SGC-7901和AGS为靶细胞，采用克隆形成实验、活性氧（ROS）检测、Western blot等方法检测细胞增殖、ROS水平及内质网应激通路（BiP/p-PERK/p-eIF2α）和STAT3磷酸化水平的变化。建立裸鼠皮下移植瘤模型，将纤维膜局部贴敷于肿瘤表面，观察体内抑瘤效果；通过免疫组化检测肿瘤组织中BrdU、BiP及p-STAT3的表达。结果 制备的纳米纤维膜表面光滑，姜黄素呈无定形均匀分散；体外释放可维持超过120 h。该材料对正常GES-1细胞无明显毒性（P>0.05）；能显著抑制MKN-45、SGC-7901和AGS细胞克隆形成（抑制率约60%）。机制上，姜黄素/明胶纳米纤维膜处理后，胃癌细胞内ROS水平显著升高（P<0.05），BiP、p-PERK、p-eIF2α和裂解型caspase-3表达上调，p-STAT3表达下调；加入ROS清除剂NAC可逆转上述变化（均P<0.05）。体内实验显示，局部应用该纤维膜能明显抑制移植瘤生长，肿瘤体积减小，BrdU阳性细胞减少（均P<0.05），BiP表达升高，p-STAT3水平下降。结论 姜黄素/明胶纳米纤维膜通过局部缓释姜黄素，显著提高其生物利用度；通过诱导胃癌细胞ROS产生、激活内质网应激并抑制STAT3磷酸化，从而有效抑制胃癌细胞增殖并促进其凋亡。该材料生物相容性好、制备简便、成本低廉，有望成为胃癌术后局部辅助治疗的新型安全制剂。

Abstract:

Background and Aims Curcumin is a safe and inexpensive natural compound, but its clinical application in gastric cancer therapy is severely limited by poor water solubility and low bioavailability. This study aims to construct an implantable curcumin/gelatin nanofibrous membrane for local sustained drug release and to investigate its molecular mechanisms in inhibiting GC cell proliferation.Methods Curcumin/gelatin nanofibrous membranes were prepared by electrospinning. Morphology was observed by scanning electron microscopy, and the in vitro drug release profile was determined by high-performance liquid chromatography-tandem mass spectrometry. Biocompatibility was evaluated using human gastric mucosal cells GES-1. Human gastric cancer cell lines MKN-45, SGC-7901, and AGS were treated with conditioned media from the nanofibrous membranes. Colony formation assay, reactive oxygen species (ROS) detection, and Western blot were performed to assess cell proliferation, ROS levels, and the expression of endoplasmic reticulum stress pathway proteins (BiP/p-PERK/p-eIF2α) and phosphorylated STAT3. A subcutaneous xenograft model in nude mice was established, and the membrane was placed locally on the tumor surface to evaluate in vivo anti-tumor efficacy. Immunohistochemistry was used to detect the expression of BrdU, BiP, and p-STAT3 in tumor tissues.Results The prepared nanofibrous membrane had a smooth surface with amorphous and uniform dispersion of curcumin. In vitro release lasted for over 120 h. The material showed no obvious toxicity to normal GES-1 cells (P>0.05). It significantly inhibited colony formation of MKN-45, SGC-7901, and AGS cells (inhibition rate 60%). Mechanistically, treatment with curcumin/gelatin nanofibrous membrane significantly increased intracellular ROS levels, upregulated the expression of BiP, p-PERK, p-eIF2α, and cleaved caspase-3, and downregulated p-STAT3. At the same time, these changes were reversed by the ROS scavenger NAC (all P<0.05). In vivo, local application of the membrane significantly suppressed xenograft tumor growth, reduced tumor volume, decreased BrdU-positive cells (all P<0.05), increased BiP expression, and lowered p-STAT3 levels.Conclusion The curcumin/gelatin nanofibrous membrane significantly enhances the bioavailability of curcumin through localized sustained release. It effectively inhibits the proliferation of gastric cancer cells and promotes their apoptosis by inducing ROS generation, activating endoplasmic reticulum stress, and suppressing STAT3 phosphorylation. With favorable biocompatibility, simple preparation, and low cost, this material shows promise as a novel and safe formulation for postoperative local adjuvant therapy of gastric cancer.