基于空间压力梯度设计的微针装置实现快速真皮间质液采样及其机制研究

本刊推荐：为解决真皮间质液（ISF）采样中收集时间长、失败率高、体积变异大等挑战，研究人员开发了基于达西定律空间压力梯度设计的POP微针装置。该装置在5分钟内平均采集15.5 mg ISF，近乎零失败率，并阐明了穿透深度、压力、年龄对ISF采集的影响，提出了<1%血液污染的接受标准，为ISF诊断技术的稳健发展奠定基础。

　　

在医学诊断领域，血液检测一直是金标准，但它的侵入性、疼痛感以及需要专业人员进行操作的特点，限制了其频繁使用和可及性。因此，科学家们将目光投向了另一种潜在的生物样本——真皮间质液（Interstitial Fluid, ISF）。这种存在于皮肤细胞之间的液体，与血液进行着持续的分子交换，几乎包含了所有的血浆分析物，甚至还有一些独特的、可能源于皮肤微环境的生物标志物。这使其成为一种极具前景的微创诊断替代方案，不仅可用于全身性疾病的监测，也是评估皮肤局部状况的有力工具。

然而，通往ISF广泛应用的道路上横亘着巨大的障碍：采样技术。ISF驻留在皮肤细胞外基质（Extracellular Matrix, ECM）的复杂网络中，其采样面临多重严峻挑战。首先，超过40%的ISF是与ECM非共价结合的“束缚水”，无法被采集，理论上可获取的ISF在皮肤最厚区域也仅有约120 μl/cm²。现实中，现有设备通常只能收集1到20 μl。其次，ECM对ISF流动产生巨大阻力，导致采样过程极其漫长，往往需要20分钟或更久。再者，有假说认为ECM在ISF流动过程中会对分析物产生“色谱效应”，导致大小分子分离，造成检测结果的不可靠性。此外，个体的年龄、性别、种族、身体部位和生活方式的差异都会影响ECM的状态，进一步增加了ISF采样的变异性。这些复杂性使得ISF采样远比采血更具挑战性。传统的ISF采样方法，如wick提取、 suction blister（ suction blister（吸疱法）和微透析，都颇具侵入性。微针（Microneedle）方法虽属微创，但历史上一直受困于采集体积过低（通常<1 μl）。近年来，将压力与微针结合的关键见解显著提升了此类设备的采集速度和体积，使其成为当前ISF研究的主流方法。然而，即便是最新的真空辅助微针阵列技术，也需要1200至1600个穿刺点，耗时25分钟才能收集平均20.8 μl的ISF。采样速度慢、失败率高、收集体积不定以及生物分析物水平不一致等诸多问题，其根本原因很多尚不明确，严重阻碍了ISF测试的理解和采纳进程。

为了解决这些难题，并推动ISF诊断学的发展，一项发表于《SCIENCE ADVANCES》的研究报道了一种名为POP（Puncture-Out-and-Press）的微针装置，它基于达西定律（Darcy's law）进行第一性原理设计，通过精巧设计空间压力梯度而非单纯增大施加压力，实现了ISF的快速、稳健采集。该研究不仅展示了设备的优异性能，更借此深入探究了ISF采样的内在机制，量化了血液污染的影响，并对长期以来关于蛋白质尺寸依赖性运输的观点进行了重新审视。

研究人员为开展此项研究，主要应用了以下几项关键技术：利用高精度3D打印技术（Carbon 3D S2及M2打印机）制造不同版本的微针阵列（MAP）和收集板；通过离体人皮肤（源自Zen-Bio和NDRI）和猪皮肤模型进行ISF采样效率与机制研究；结合有限元分析（FEA）模拟不同设计下的组织内压力分布；开展一项纳入10名参与者的人体先导研究，评估设备安全性、疼痛感及采集性能；运用蛋白质组学技术（基于Orbitrap Eclipse Tribrid质谱仪的液相色谱-质谱联用（LC-MS/MS）与MaxQuant软件分析）对ISF、血浆和红细胞（RBC）样本进行深度分析；并采用统计学方法（如Storey程序）处理组学数据并量化血液污染导致的误差。

POP设计维度

研究的核心是POP设备。其基础设计包含一个7x5的实心微针阵列（MAP），针尖为400 μm直径的圆锥形，以及一个带有500 μm穿孔的收集板。当两者配合时，有1100 μm的微针暴露出来。根据不同实验需求，通过3D打印制造了略微不同的版本。

空间压力梯度设计

为了验证空间压力梯度调控ISF流动的设想，研究团队在离体人皮肤上通过改变POP的使用配置和设计细节来改变压力分布，并比较ISF收集量。他们测试了POP设计、一种不取出MAP而将压力同时施加于MAP和收集板的“needle-in”变体、真空变体以及一种施加环状压力分布的变体。有限元分析（FEA）使用简化的真皮模型模拟了不同的压力梯度。结果显示，在50 kPa压力下，POP提取的ISF量是其他方案的1.9到4倍。模拟的压力分布显示，正如设计预期，除了穿刺点外，整个组织都处于压缩状态。 needle-in变体因留在穿刺点内的微针导致组织扩张，削弱了穿刺点与周围组织之间的压力梯度，从而减少了ISF流动。真空变体虽然FEA预测了比POP更大的压力梯度，但实验结果与之不符， likely due to difficulties in maintaining vacuum seals across multiple openings（ likely due to difficulties in maintaining vacuum seals across multiple openings（ likely due to difficulties in maintaining vacuum seals across multiple openings（ likely due to difficulties in maintaining vacuum seals across multiple openings（ likely due to difficulties in maintaining vacuum seals across multiple openings（ likely due to difficulties in maintaining vacuum seals across multiple openings（ likely due to difficulties in maintaining vacuum seals across multiple openings（ likely due to difficulties in maintaining vacuum seals across multiple openings（ likely due to 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