Developing a soft micropatterned substrate to enhance maturation of human induced pluripotent stem cell-derived cardiomyocytes

IF 9.4 1区 医学 Q1 ENGINEERING, BIOMEDICAL Acta Biomaterialia Pub Date : 2024-12-01 DOI:10.1016/j.actbio.2024.10.029
Yasaman Maaref , Shayan Jannati , Farah Jayousi , Philipp Lange , Mohsen Akbari , Mu Chiao , Glen F Tibbits
{"title":"Developing a soft micropatterned substrate to enhance maturation of human induced pluripotent stem cell-derived cardiomyocytes","authors":"Yasaman Maaref ,&nbsp;Shayan Jannati ,&nbsp;Farah Jayousi ,&nbsp;Philipp Lange ,&nbsp;Mohsen Akbari ,&nbsp;Mu Chiao ,&nbsp;Glen F Tibbits","doi":"10.1016/j.actbio.2024.10.029","DOIUrl":null,"url":null,"abstract":"<div><div>Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC<img>CMs) offer numerous advantages as a biological model, yet their inherent immaturity compared to adult cardiomyocytes poses significant limitations. This study addresses hiPSC<img>CM immaturity by introducing a physiologically relevant micropatterned substrate for long-term culture and maturation. An innovative microfabrication methodology combining laser etching and casting creates a micropatterned polydimethylsiloxane (PDMS) substrate with varying stiffness, from 2 to 50 kPa, mimicking healthy and fibrotic cardiac tissue. Platinum electrodes were integrated into the cell culture chamber enable pacing of cells at various frequencies. Subsequently, cells were transferred to the incubator for time-course analysis, ensuring contamination-free conditions. Cell contractility, cytosolic Ca<sup>2+</sup> transient, sarcomere orientation, and nucleus aspect ratio were analyzed in a 2D hiPSC<img>CM monolayer up to 90 days post-replating in relation to substrate micropattern dimensions. Culturing hiPSC<img>CMs for three weeks on a micropatterned PDMS substrate (2.5–5 µm deep, 20 µm center-to-center spacing of grooves, 2–5 kPa stiffness) emerges as optimal for cardiomyocyte alignment, contractility, and cytosolic Ca<sup>2+</sup> transient. The study provides insights into substrate stiffness effects on hiPSC<img>CM contractility and Ca<sup>2+</sup> transient at immature and mature states. Maximum contractility and fastest Ca<sup>2+</sup>transient kinetics occur in mature hiPSC<img>CMs cultured for two to four weeks, with the optimum at three weeks, on a soft micropatterned PDMS substrate. MS proteomic analysis further revealed that hiPSC<img>CMs cultured on soft micropatterned substrates exhibit advanced maturation, marked by significant upregulation of key structural, electrophysiological, and metabolic proteins. This new substrate offers a promising platform for disease modeling and therapeutic interventions.</div></div><div><h3>Statement of Significance</h3><div>Human induced pluripotent stem cell derived cardiomyocytes (hiPSC<img>CMs) have been transformative to disease-in-a-dish modeling, drug discovery and testing, and autologous regeneration for human hearts and their role will continue to expand dramatically. However, one of the major limitations of hiPSC<img>CMs is that without intervention, the cells are immature and represent those in the fetal heart. We developed protocols for the fabrication of the PDMS matrices that includes variations in its stiffness and micropatterning. Growing our hiPSC<img>CMs on matrices of comparable stiffness to a healthy heart (5 kPa) and grooves of 20 μm, generate heart cells typical of the healthy adult human heart.</div></div>","PeriodicalId":237,"journal":{"name":"Acta Biomaterialia","volume":"190 ","pages":"Pages 133-151"},"PeriodicalIF":9.4000,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Acta Biomaterialia","FirstCategoryId":"5","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S1742706124006214","RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"ENGINEERING, BIOMEDICAL","Score":null,"Total":0}
引用次数: 0

Abstract

Human induced pluripotent stem cell-derived cardiomyocytes (hiPSCCMs) offer numerous advantages as a biological model, yet their inherent immaturity compared to adult cardiomyocytes poses significant limitations. This study addresses hiPSCCM immaturity by introducing a physiologically relevant micropatterned substrate for long-term culture and maturation. An innovative microfabrication methodology combining laser etching and casting creates a micropatterned polydimethylsiloxane (PDMS) substrate with varying stiffness, from 2 to 50 kPa, mimicking healthy and fibrotic cardiac tissue. Platinum electrodes were integrated into the cell culture chamber enable pacing of cells at various frequencies. Subsequently, cells were transferred to the incubator for time-course analysis, ensuring contamination-free conditions. Cell contractility, cytosolic Ca2+ transient, sarcomere orientation, and nucleus aspect ratio were analyzed in a 2D hiPSCCM monolayer up to 90 days post-replating in relation to substrate micropattern dimensions. Culturing hiPSCCMs for three weeks on a micropatterned PDMS substrate (2.5–5 µm deep, 20 µm center-to-center spacing of grooves, 2–5 kPa stiffness) emerges as optimal for cardiomyocyte alignment, contractility, and cytosolic Ca2+ transient. The study provides insights into substrate stiffness effects on hiPSCCM contractility and Ca2+ transient at immature and mature states. Maximum contractility and fastest Ca2+transient kinetics occur in mature hiPSCCMs cultured for two to four weeks, with the optimum at three weeks, on a soft micropatterned PDMS substrate. MS proteomic analysis further revealed that hiPSCCMs cultured on soft micropatterned substrates exhibit advanced maturation, marked by significant upregulation of key structural, electrophysiological, and metabolic proteins. This new substrate offers a promising platform for disease modeling and therapeutic interventions.

Statement of Significance

Human induced pluripotent stem cell derived cardiomyocytes (hiPSCCMs) have been transformative to disease-in-a-dish modeling, drug discovery and testing, and autologous regeneration for human hearts and their role will continue to expand dramatically. However, one of the major limitations of hiPSCCMs is that without intervention, the cells are immature and represent those in the fetal heart. We developed protocols for the fabrication of the PDMS matrices that includes variations in its stiffness and micropatterning. Growing our hiPSCCMs on matrices of comparable stiffness to a healthy heart (5 kPa) and grooves of 20 μm, generate heart cells typical of the healthy adult human heart.

Abstract Image

查看原文
分享 分享
微信好友 朋友圈 QQ好友 复制链接
本刊更多论文
开发软性微图案基底,促进人类诱导多能干细胞衍生心肌细胞的成熟。
人诱导多能干细胞衍生的心肌细胞(hiPSC-CMs)作为一种生物模型具有诸多优势,但与成体心肌细胞相比,其固有的不成熟性带来了很大的局限性。本研究通过引入与生理相关的微图案基底来解决 hiPSC-CM 的不成熟问题,以便进行长期培养和成熟。创新的微加工方法结合了激光蚀刻和铸造技术,创造出了微图案化的聚二甲基硅氧烷(PDMS)基底,其硬度从 2 千帕到 50 千帕不等,可模拟健康和纤维化的心脏组织。细胞培养室中集成了铂电极,可对细胞进行不同频率的起搏。随后,将细胞转移到培养箱中进行时程分析,确保无污染条件。在二维 hiPSC-CM 单层中分析了细胞收缩力、细胞膜 Ca2+ 瞬态、肌节定向和细胞核长宽比,分析结果与基底微图案尺寸有关,分析时间长达复制后 90 天。在微图案 PDMS 基底(2.5-5 微米深,沟槽中心到中心间距为 20 微米,硬度为 2-5 千帕)上培养 hiPSC-CMs 三周后,发现其对心肌细胞排列、收缩力和细胞膜 Ca2+ 瞬态都是最佳的。该研究深入探讨了基质硬度对未成熟和成熟状态的 hiPSC-CM 收缩能力和 Ca2+ 瞬态的影响。在柔软的微图案 PDMS 基质上培养 2 至 4 周后,成熟的 hiPSC-CMs 的收缩力最大,Ca2+瞬态动力学速度最快,3 周时最佳。质谱蛋白质组分析进一步显示,在柔软的微图案基底上培养的 hiPSC-CMs 表现出高级成熟,其标志是关键结构蛋白、电生理蛋白和代谢蛋白的显著上调。这种新型基质为疾病建模和治疗干预提供了一个前景广阔的平台。意义声明:人类诱导多能干细胞衍生的心肌细胞(hiPSC-CMs)在 "皿中疾病 "建模、药物发现和测试以及人类心脏自体再生方面具有变革性作用,其作用将继续显著扩大。然而,hiPSC-CMs 的一个主要局限性是,如果不进行干预,这些细胞是不成熟的,只能代表胎儿心脏中的细胞。我们制定了制作 PDMS 基质的方案,包括改变其硬度和微图案。在硬度与健康心脏相当(5 kPa)、沟槽长度为 20 μm 的基质上培育 hiPSC-CM,可产生典型的健康成人心脏细胞。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
求助全文
约1分钟内获得全文 去求助
来源期刊
Acta Biomaterialia
Acta Biomaterialia 工程技术-材料科学:生物材料
CiteScore
16.80
自引率
3.10%
发文量
776
审稿时长
30 days
期刊介绍: Acta Biomaterialia is a monthly peer-reviewed scientific journal published by Elsevier. The journal was established in January 2005. The editor-in-chief is W.R. Wagner (University of Pittsburgh). The journal covers research in biomaterials science, including the interrelationship of biomaterial structure and function from macroscale to nanoscale. Topical coverage includes biomedical and biocompatible materials.
期刊最新文献
Editorial Board A data-driven microstructure-based model for predicting circumferential behavior and failure in degenerated human annulus fibrosus A multifunctional nanosystem catalyzed by cascading natural glucose oxidase and Fe3O4 nanozymes for synergistic chemodynamic and photodynamic cancer therapy Bioengineering strategy to promote CNS nerve growth and regeneration via chronic glutamate signaling Cellular fibronectin-targeted fluorescent aptamer probes for early detection and staging of liver fibrosis
×
引用
GB/T 7714-2015
复制
MLA
复制
APA
复制
导出至
BibTeX EndNote RefMan NoteFirst NoteExpress
×
×
提示
您的信息不完整,为了账户安全,请先补充。
现在去补充
×
提示
您因"违规操作"
具体请查看互助需知
我知道了
×
提示
现在去查看 取消
×
提示
确定
0
微信
客服QQ
Book学术公众号 扫码关注我们
反馈
×
意见反馈
请填写您的意见或建议
请填写您的手机或邮箱
已复制链接
已复制链接
快去分享给好友吧!
我知道了
×
扫码分享
扫码分享
Book学术官方微信
Book学术文献互助
Book学术文献互助群
群 号:481959085
Book学术
文献互助 智能选刊 最新文献 互助须知 联系我们:info@booksci.cn
Book学术提供免费学术资源搜索服务,方便国内外学者检索中英文文献。致力于提供最便捷和优质的服务体验。
Copyright © 2023 Book学术 All rights reserved.
ghs 京公网安备 11010802042870号 京ICP备2023020795号-1