血脑屏障的通透性会随着胶质母细胞瘤细胞在体外的分化而增加。

IF 5.9 1区 医学 Q1 NEUROSCIENCES
Sabrina Digiovanni, Martina Lorenzati, Olga Teresa Bianciotto, Martina Godel, Simona Fontana, Muhlis Akman, Costanzo Costamagna, Pierre-Olivier Couraud, Annalisa Buffo, Joanna Kopecka, Chiara Riganti, Iris Chiara Salaroglio
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引用次数: 0

摘要

背景:多形性胶质母细胞瘤(GBM)是一种侵袭性肿瘤,由于血脑屏障(BBB)富含 ATP 结合盒(ABC)转运体和紧密连接(TJ)蛋白,因此很难进行药物治疗。血脑屏障在 GBM 体积内被破坏,但在脑部与肿瘤相邻的区域却能发挥作用,在这些区域,最终的 GBM 病灶可能会引发肿瘤复发。GBM 细胞如何影响 BBB 的通透性,目前还没有深入研究:为了弄清这一点,我们在从每个肿瘤中分离出干细胞/神经球(SC/NS)和分化/粘附细胞(AC)成分后,用 3 个源自患者的 GBM 细胞共同培养了人类 BBB 模型。此外,我们还用富含或去除 IL-6 的 NS 或 AC 条件培养基培养 BBB 细胞。细胞外细胞因子通过蛋白质阵列和酶联免疫吸附进行测定。在 STAT3 药物抑制剂或特异性 PROTAC 的作用下,通过免疫印迹法测定 BBB 细胞的胞内信号传导。通过渗透性实验和 TEER 测量评估了 BBB 的能力:结果:GBM 细胞或其条件培养基的存在增加了多柔比星、米托蒽醌和右旋糖酐-70 的通透性,降低了 TEER,在转录水平下调了 ABC 转运体和 TJ 蛋白。AC 或其培养基对这些效应的影响高于 NS。分泌组分析发现,AC 产生的 IL-6 明显多于 NS。值得注意的是,用IL-6中和抗体处理的AC调节培养基降低了BBB对NS水平的通透性,而富含IL-6的NS调节培养基增加了BBB对AC水平的通透性。从机制上讲,AC GBM 细胞释放的 IL-6 激活了 BBB 细胞中的 STAT3。反过来,STAT3 下调了 ABC 转运体和 TJ 的表达,增加了通透性并降低了 TEER。用 STAT3 的药理抑制剂 STA-21 或靶向 STAT3 的 PROTAC 处理 BBB 细胞也能获得同样的效果:我们的研究首次证明,GBM 的分化程度会影响 BBB 的通透性。释放 IL-6 的 GBM 细胞和通过激活 STAT3 来做出反应的 BBB 细胞之间的串扰控制着 BBB 上 ABC 转运体和 TJ 蛋白的表达。这些结果可能为调整 BBB 通透性和改善 GBM 药物输送的新型治疗工具铺平了道路。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Blood-brain barrier permeability increases with the differentiation of glioblastoma cells in vitro.

Background: Glioblastoma multiforme (GBM) is an aggressive tumor, difficult to treat pharmacologically because of the blood-brain barrier (BBB), which is rich in ATP-binding cassette (ABC) transporters and tight junction (TJ) proteins. The BBB is disrupted within GBM bulk, but it is competent in brain-adjacent-to-tumor areas, where eventual GBM foci can trigger tumor relapse. How GBM cells influence the permeability of BBB is poorly investigated.

Methods: To clarify this point, we co-cultured human BBB models with 3 patient-derived GBM cells, after separating from each tumor the stem cell/neurosphere (SC/NS) and the differentiated/adherent cell (AC) components. Also, we set up cultures of BBB cells with the conditioned medium of NS or AC, enriched or depleted of IL-6. Extracellular cytokines were measured by protein arrays and ELISA. The intracellular signaling in BBB cells was measured by immunoblotting, in the presence of STAT3 pharmacological inhibitor or specific PROTAC. The competence of BBB was evaluated by permeability assays and TEER measurement.

Results: The presence of GBM cells or their conditioned medium increased the permeability to doxorubicin, mitoxantrone and dextran-70, decreased TEER, down-regulated ABC transporters and TJ proteins at the transcriptional level. These effects were higher with AC or their medium than with NS. The secretome analysis identified IL-6 as significantly more produced by AC than by NS. Notably, AC-conditioned medium treated with an IL-6 neutralizing antibody reduced the BBB permeability to NS levels, while NS-conditioned medium enriched with IL-6 increased BBB permeability to AC levels. Mechanistically, IL-6 released by AC GBM cells activated STAT3 in BBB cells. In turn, STAT3 down-regulated ABC transporter and TJ expression, increased permeability and decreased TEER. The same effects were obtained in BBB cells treated with STA-21, a pharmacological inhibitor of STAT3, or with a PROTAC targeting STAT3.

Conclusions: Our work demonstrates for the first time that the degree of GBM differentiation influences BBB permeability. The crosstalk between GBM cells that release IL-6 and BBB cells that respond by activating STAT3, controls the expression of ABC transporters and TJ proteins on BBB. These results may pave the way for novel therapeutic tools to tune BBB permeability and improve drug delivery to GBM.

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来源期刊
Fluids and Barriers of the CNS
Fluids and Barriers of the CNS Neuroscience-Developmental Neuroscience
CiteScore
10.70
自引率
8.20%
发文量
94
审稿时长
14 weeks
期刊介绍: "Fluids and Barriers of the CNS" is a scholarly open access journal that specializes in the intricate world of the central nervous system's fluids and barriers, which are pivotal for the health and well-being of the human body. This journal is a peer-reviewed platform that welcomes research manuscripts exploring the full spectrum of CNS fluids and barriers, with a particular focus on their roles in both health and disease. At the heart of this journal's interest is the cerebrospinal fluid (CSF), a vital fluid that circulates within the brain and spinal cord, playing a multifaceted role in the normal functioning of the brain and in various neurological conditions. The journal delves into the composition, circulation, and absorption of CSF, as well as its relationship with the parenchymal interstitial fluid and the neurovascular unit at the blood-brain barrier (BBB).
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