Department of Neurosurgery, First Affiliated Hospital of Xi'an Jiaotong University, Xi'an, Shaanxi 710061 P.R. China.
Department of Ophthalmology, First Affiliated Hospital of Xi'an Jiaotong University, Xi'an, Shaanxi 710061 P.R. China.
Mol Med Rep. 2023 Nov;28(5). doi: 10.3892/mmr.2023.13102. Epub 2023 Sep 29.
Malignant glioma is a highly vascularized tumor. Therefore, inhibition of angiogenesis is an effective treatment strategy for it. Alphastatin is a 24‑amino acid peptide that has been demonstrated to inhibit glioma angiogenesis and tumor growth. Adipose‑derived stem cells (ADSCs) are considered an ideal targeted drug delivery system for glioma therapy due to their targeted tropism for cancer and the intrinsic attribute of autologous transplantation. The aim of the present study was to construct an ADSC‑mediated alphastatin targeted delivery system and investigate its effects on angiogenesis in glioma. The sequence encoding the human neurotrophin‑4 signal peptide and alphastatin fusion gene fragment was transferred into ADSCs using a lentiviral vector to construct the ADSC‑mediated alphastatin targeted delivery system (Al‑ADSCs). Flow cytometry was used to detect the stem cell surface markers of Al‑ADSCs. Western blot analysis and ELISA were used to detect the expression and secretion of alphastatin peptide in Al‑ADSCs. Cell migration assay was used to detect the tendency of Al‑ADSCs to target CD133 glioma stem cells (GSCs). The effects of Al‑ADSCs on angiogenesis were detected by tube formation assay. A Cell Counting Kit‑8 assay was used to detect the effects of Al‑ADSCs on endothelial cell (EC) proliferation. Wound healing assay was used to examine the effects of Al‑ADSCs on EC migration. Intracranial xenograft models were constructed and fluorescence imaging was used to examine the effects of Al‑ADSCs on glioma growth. Fluorescence microscopy was used to detect the distribution of Al‑ADSCs in glioma tissue and CD133 immunofluorescence staining was used to detect the effects of Al‑ADSCs on GSCs in glioma tissue. The results revealed that ADSCs exhibited more marked tropism to GSCs than to other types of cells (P<0.01). Al‑ADSCs maintained the surface markers of ADSCs and there was no significant difference between the ADSCs and Al‑ADSCs regarding tropism to GSCs (P=0.639 for GSCs‑SHG44 cells; and P=0.386 for GSCs‑U87 cells). Al‑ADSCs were able to successfully secrete and express alphastatin peptide and inhibited EC‑mediated angiogenesis (P<0.01) and EC migration (P<0.01) and proliferation (P<0.01) . , Al‑ADSCs were detected in glioma tissue and were able to inhibit tumor growth. In addition, the Al‑ADSCs reduced the number of GSCs and microvascular density (P<0.01) in the tumors. Overall, the results of the present study indicated that the Al‑ADSCs were able to target glioma tissue and inhibit glioma angiogenesis and tumor growth. This anti‑angiogenic targeted therapy system may provide a new strategy for the treatment of glioma.
恶性脑胶质瘤是一种高度血管化的肿瘤。因此，抑制血管生成是治疗它的有效策略。阿尔法他汀是一种 24 个氨基酸的肽，已被证明可抑制神经胶质瘤的血管生成和肿瘤生长。脂肪干细胞（ADSCs）因其对癌症的靶向亲嗜性和自体移植的内在属性，被认为是神经胶质瘤治疗的理想靶向药物输送系统。本研究的目的是构建 ADSC 介导的阿尔法他汀靶向递药系统，并研究其对神经胶质瘤血管生成的影响。采用慢病毒载体将编码人神经营养因子-4 信号肽和阿尔法他汀融合基因片段的序列转染至 ADSCs 中，构建 ADSC 介导的阿尔法他汀靶向递药系统（Al-ADSCs）。采用流式细胞术检测 Al-ADSCs 的干细胞表面标志物。采用 Western blot 分析和 ELISA 检测 Al-ADSCs 中阿尔法他汀肽的表达和分泌。采用细胞迁移实验检测 Al-ADSCs 向 CD133 神经胶质瘤干细胞（GSCs）的靶向迁移趋势。采用管形成实验检测 Al-ADSCs 对血管生成的影响。采用细胞计数试剂盒-8 实验检测 Al-ADSCs 对内皮细胞（EC）增殖的影响。采用划痕愈合实验检测 Al-ADSCs 对 EC 迁移的影响。构建颅内异种移植模型，并采用荧光成像检测 Al-ADSCs 对神经胶质瘤生长的影响。采用荧光显微镜检测 Al-ADSCs 在神经胶质瘤组织中的分布，并采用 CD133 免疫荧光染色检测 Al-ADSCs 对神经胶质瘤组织中 GSCs 的影响。结果显示，ADSCs 对 GSCs 的亲嗜性明显高于其他类型的细胞（P<0.01）。Al-ADSCs 保持了 ADSCs 的表面标志物，并且在 GSCs 亲嗜性方面，ADSCs 与 Al-ADSCs 之间无显著差异（SHG44 细胞：P=0.639；U87 细胞：P=0.386）。Al-ADSCs 能够成功分泌和表达阿尔法他汀肽，并抑制 EC 介导的血管生成（P<0.01）和 EC 迁移（P<0.01）以及增殖（P<0.01）。此外，在神经胶质瘤组织中检测到 Al-ADSCs，并能够抑制肿瘤生长。此外，Al-ADSCs 还降低了肿瘤中的 GSCs 数量和微血管密度（P<0.01）。综上所述，本研究结果表明，Al-ADSCs 能够靶向神经胶质瘤组织，并抑制神经胶质瘤的血管生成和肿瘤生长。这种抗血管生成靶向治疗系统可能为神经胶质瘤的治疗提供一种新策略。
