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用于Transwell培养小室的开源、一次性3D打印环境暴露系统的开发与验证。

Development and validation of an open-source, disposable, 3D-printed environmental exposure system for Transwell culture inserts.

作者信息

Chandiramohan Abiram, Dabaghi Mohammedhossein, Aguiar Jennifer A, Tiessen Nicholas, Stewart Mary, Cao Quynh T, Nguyen Jenny P, Makhdami Nima, Cox Gerard, Doxey Andrew C, Hirota Jeremy A

机构信息

Firestone Institute for Respiratory Health - Division of Respirology, Dept of Medicine, McMaster University, Hamilton, ON, Canada.

These authors contributed equally.

出版信息

ERJ Open Res. 2021 Feb 15;7(1). doi: 10.1183/23120541.00705-2020. eCollection 2021 Jan.

Abstract

Accessible models recapitulating the human airway that are amenable to study whole cannabis smoke exposure are needed for immunological and toxicological studies that inform public health policy and recreational cannabis use. In the present study, we developed and validated a novel three-dimensional (3D)-printed exposure system (IVES) that can be directly applied to study the effect of cannabis smoke exposure on primary human bronchial epithelial cells. Using commercially available design software and a 3D printer, we designed a four-chamber Transwell insert holder for exposures to whole smoke. COMSOL Multiphysics software was used to model gas distribution, concentration gradients, velocity profile and shear stress within IVES. Following simulations, primary human bronchial epithelial cells cultured at the air-liquid interface on Transwell inserts were exposed to whole cannabis smoke using a modified version of the Foltin puff procedure. Following 24 h, outcome measurements included cell morphology, epithelial barrier function, lactate dehydrogenase (LDH) levels, cytokine expression and gene expression. Whole smoke delivered through IVES possesses velocity profiles consistent with uniform gas distribution across the four chambers and complete mixing. Airflow velocity ranged between 1.0 and 1.5 µm·s and generated low shear stresses (<<1 Pa). Human airway epithelial cells exposed to cannabis smoke using IVES showed changes in cell morphology and disruption of barrier function without significant cytotoxicity. Cannabis smoke elevated interleukin-1 family cytokines and elevated CYP1A1 and CYP1B1 expression relative to control, validating IVES smoke exposure impacts in human airway epithelial cells at a molecular level. The growing legalisation of cannabis on a global scale must be paired with research related to potential health impacts of lung exposures. IVES represents an accessible, open-source, exposure system that can be used to model varying types of cannabis smoke exposures with human airway epithelial cells grown under air-liquid interface culture conditions.

摘要

免疫和毒理学研究需要可用于研究全大麻烟雾暴露的、能概括人类气道的可及模型,这些研究为公共卫生政策和休闲大麻使用提供信息。在本研究中,我们开发并验证了一种新型的三维(3D)打印暴露系统(IVES),该系统可直接用于研究大麻烟雾暴露对原代人支气管上皮细胞的影响。使用商用设计软件和3D打印机,我们设计了一个四腔Transwell插入物支架,用于全烟雾暴露。使用COMSOL Multiphysics软件对IVES内的气体分布、浓度梯度、速度剖面和剪切应力进行建模。模拟之后,使用改良版的福尔廷吹气程序,将在Transwell插入物上的气液界面培养的原代人支气管上皮细胞暴露于全大麻烟雾中。24小时后,结果测量包括细胞形态、上皮屏障功能、乳酸脱氢酶(LDH)水平、细胞因子表达和基因表达。通过IVES输送的全烟雾具有与四个腔室中均匀气体分布和完全混合一致的速度剖面。气流速度在1.0至1.5 µm·s之间,产生的剪切应力较低(<<1 Pa)。使用IVES暴露于大麻烟雾的人气道上皮细胞显示出细胞形态变化和屏障功能破坏,但无明显细胞毒性。相对于对照,大麻烟雾使白细胞介素-1家族细胞因子升高,并使CYP1A1和CYP1B1表达升高,这在分子水平上验证了IVES烟雾暴露对人气道上皮细胞的影响。全球范围内大麻合法化的不断增加必须与肺部暴露潜在健康影响的相关研究相结合。IVES代表了一种可及的、开源的暴露系统,可用于模拟在气液界面培养条件下生长的人气道上皮细胞对不同类型大麻烟雾的暴露。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/73df/7882787/d0e6931564e4/00705-2020.01.jpg

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