A straightforward cell culture insert model to incorporate biochemical and biophysical stromal properties into transplacental transport studies

IF 3 2区 医学 Q2 DEVELOPMENTAL BIOLOGY
Katherine M. Nelson, Bryan J. Ferrick, Hassan Karimi, Christine L. Hatem, Jason P. Gleghorn
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Abstract

The placental extracellular matrix (ECM) dynamically remodels over pregnancy and in disease. How these changes impact placental barrier function is poorly understood as there are limited models of the placenta with a modifiable stromal compartment to mechanistically investigate these extracellular factors. We developed a straightforward method to incorporate uniform hydrogels into standard cell culture inserts for transplacental transport studies. Uniform polyacrylamide (PAA) gels were polymerized within cell culture inserts by (re)using the insert packaging to create a closed, controllable environmental chamber. PAA pre-polymer solution was added dropwise via a syringe to the cell culture insert and the atmosphere was purged with an inert gas. Transport and cell culture studies were conducted to validate the model. We successfully incorporated ECM-functionalized uniform PAA gels into cell culture inserts, enabling cell adhesion and monolayer formation. Imaging and analyte transport studies validated gel formation and expected mass transport results, and successful cell studies confirmed cell viability, stiffness-mediated YAP translocation, and that the model could be used in transplacental transport studies. Detailed methods and validation protocols are included. The incorporation of a PAA gel within a cell culture insert enables independent study of placental ECM biophysical and biochemical properties in the context of transplacental transport. These straightforward and low-cost methods to build three-dimensional cellular models are readily adoptable by the wider scientific community.
将生物化学和生物物理基质特性纳入经胎盘运输研究的直接细胞培养插入模型
胎盘细胞外基质(ECM)在妊娠期和疾病期间会发生动态重塑。人们对这些变化如何影响胎盘屏障功能知之甚少,因为目前只有有限的胎盘模型具有可改变的基质区,可以从机理上研究这些细胞外基质。我们开发了一种简单易行的方法,将均匀水凝胶加入标准细胞培养插入物中,用于经胎盘运输研究。均匀聚丙烯酰胺(PAA)凝胶通过(重新)使用插入式包装在细胞培养插入物中聚合,以创建一个封闭、可控的环境室。通过注射器将 PAA 预聚物溶液滴加到细胞培养插片中,并用惰性气体吹扫周围环境。为验证模型,我们进行了运输和细胞培养研究。我们成功地将 ECM 功能化的均匀 PAA 凝胶加入到细胞培养插片中,实现了细胞粘附和单层形成。成像和分析物运输研究验证了凝胶的形成和预期的质量运输结果,成功的细胞研究证实了细胞的活力、硬度介导的 YAP 转位以及该模型可用于经胎盘运输研究。详细的方法和验证协议也包括在内。在细胞培养插入物中加入 PAA 凝胶可独立研究胎盘 ECM 在经胎盘运输过程中的生物物理和生物化学特性。这些建立三维细胞模型的方法简单易行、成本低廉,很容易被更广泛的科学界采用。
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来源期刊
Placenta
Placenta 医学-发育生物学
CiteScore
6.30
自引率
10.50%
发文量
391
审稿时长
78 days
期刊介绍: Placenta publishes high-quality original articles and invited topical reviews on all aspects of human and animal placentation, and the interactions between the mother, the placenta and fetal development. Topics covered include evolution, development, genetics and epigenetics, stem cells, metabolism, transport, immunology, pathology, pharmacology, cell and molecular biology, and developmental programming. The Editors welcome studies on implantation and the endometrium, comparative placentation, the uterine and umbilical circulations, the relationship between fetal and placental development, clinical aspects of altered placental development or function, the placental membranes, the influence of paternal factors on placental development or function, and the assessment of biomarkers of placental disorders.
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