利用毫流体生物反应器可以长期培养原始淋巴细胞或 CD34+ 造血细胞,同时还能检测肿瘤扩增。

IF 4.3 3区 工程技术 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY Frontiers in Bioengineering and Biotechnology Pub Date : 2024-10-02 eCollection Date: 2024-01-01 DOI:10.3389/fbioe.2024.1388312
Paolo Ritter, Stefania Oliveto, Chiara Cordiglieri, Alessandra Fasciani, Christian Andrea Di Buduo, Lucrezia Della Volpe, Alberto Bocconi, Claudio Conci, Carolina Paula Miguel, Raffaella Di Micco, Alessandra Balduini, Manuela Teresa Raimondi, Stefano Biffo
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引用次数: 0

摘要

原代淋巴细胞和造血干细胞及祖细胞(HSPCs)的长期培养对其扩增和研究至关重要。此外,对上述人类原代细胞进行基因工程有几种安全需求,包括需要高效的体外检测方法来检测不希望发生的致瘤事件。在这项工作中,我们测试并优化了微型可视生物反应器(MOAB)平台。MOAB 由带有三个光学可及培养室的毫流体细胞培养装置组成。我们在 MOAB 中插入了一个与骨髓环境某些特性相似的丝基框架,并在该装置中培养从健康供体水包衣中分离出来的 CD4+ T 淋巴细胞或脐带血造血 CD34+ 细胞。这些细胞中的一部分可存活长达 3 个月。接下来,我们测试了 MOAB 检测致瘤事件的能力。将连续稀释的工程荧光肿瘤细胞与 CD4+ 或 CD34+ 原始细胞混合,并跟踪其生长情况。通过这种方法,我们成功地检测到了100,000个原代细胞中混有100个致瘤细胞。我们发现,非致瘤性原代细胞在丝环境中定植,而肿瘤细胞在经过适应阶段后,扩大并进入血液循环。我们的结论是,毫流体平台可以在人类细胞的长期培养过程中检测到罕见的致瘤事件。
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A millifluidic bioreactor allows the long term culture of primary lymphocytes or CD34+ hematopoietic cells while allowing the detection of tumorigenic expansion.

Long-term culture of primary lymphocytes and hematopoietic stem and progenitor cells (HSPCs) is pivotal to their expansion and study. Furthermore, genetic engineering of the above-mentioned primary human cells has several safety needs, including the requirement of efficient in vitro assays for unwanted tumorigenic events. In this work, we tested and optimized the Miniaturized Optically Accessible Bioreactor (MOAB) platform. The MOAB consists of a millifluidic cell culture device with three optically-accessible culture chambers. Inside the MOAB, we inserted a silk-based framework that resembles some properties of the bone marrow environment and cultivated in this device either CD4+ T lymphocytes isolated from healthy donor buffy coat or cord blood-derived hematopoietic CD34+ cells. A fraction of these cells is viable for up to 3 months. Next, we tested the capability of the MOAB to detect tumorigenic events. Serial dilutions of engineered fluorescent tumor cells were mixed with either CD4+ or CD34+ primary cells, and their growth was followed. By this approach, we successfully detected as little as 100 tumorigenic cells mixed with 100,000 primary cells. We found that non-tumorigenic primary cells colonized the silk environment, whereas tumor cells, after an adaptation phase, expanded and entered the circulation. We conclude that the millifluidic platform allows the detection of rare tumorigenic events in the long-term culture of human cells.

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来源期刊
Frontiers in Bioengineering and Biotechnology
Frontiers in Bioengineering and Biotechnology Chemical Engineering-Bioengineering
CiteScore
8.30
自引率
5.30%
发文量
2270
审稿时长
12 weeks
期刊介绍: The translation of new discoveries in medicine to clinical routine has never been easy. During the second half of the last century, thanks to the progress in chemistry, biochemistry and pharmacology, we have seen the development and the application of a large number of drugs and devices aimed at the treatment of symptoms, blocking unwanted pathways and, in the case of infectious diseases, fighting the micro-organisms responsible. However, we are facing, today, a dramatic change in the therapeutic approach to pathologies and diseases. Indeed, the challenge of the present and the next decade is to fully restore the physiological status of the diseased organism and to completely regenerate tissue and organs when they are so seriously affected that treatments cannot be limited to the repression of symptoms or to the repair of damage. This is being made possible thanks to the major developments made in basic cell and molecular biology, including stem cell science, growth factor delivery, gene isolation and transfection, the advances in bioengineering and nanotechnology, including development of new biomaterials, biofabrication technologies and use of bioreactors, and the big improvements in diagnostic tools and imaging of cells, tissues and organs. In today`s world, an enhancement of communication between multidisciplinary experts, together with the promotion of joint projects and close collaborations among scientists, engineers, industry people, regulatory agencies and physicians are absolute requirements for the success of any attempt to develop and clinically apply a new biological therapy or an innovative device involving the collective use of biomaterials, cells and/or bioactive molecules. “Frontiers in Bioengineering and Biotechnology” aspires to be a forum for all people involved in the process by bridging the gap too often existing between a discovery in the basic sciences and its clinical application.
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