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
{"title":"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。","authors":"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","doi":"10.3389/fbioe.2024.1388312","DOIUrl":null,"url":null,"abstract":"<p><p>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 <i>in vitro</i> 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<sup>+</sup> T lymphocytes isolated from healthy donor buffy coat or cord blood-derived hematopoietic CD34<sup>+</sup> 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<sup>+</sup> or CD34<sup>+</sup> 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.</p>","PeriodicalId":12444,"journal":{"name":"Frontiers in Bioengineering and Biotechnology","volume":null,"pages":null},"PeriodicalIF":4.3000,"publicationDate":"2024-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11479935/pdf/","citationCount":"0","resultStr":"{\"title\":\"A millifluidic bioreactor allows the long term culture of primary lymphocytes or CD34<sup>+</sup> hematopoietic cells while allowing the detection of tumorigenic expansion.\",\"authors\":\"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\",\"doi\":\"10.3389/fbioe.2024.1388312\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><p>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 <i>in vitro</i> 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<sup>+</sup> T lymphocytes isolated from healthy donor buffy coat or cord blood-derived hematopoietic CD34<sup>+</sup> 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<sup>+</sup> or CD34<sup>+</sup> 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.</p>\",\"PeriodicalId\":12444,\"journal\":{\"name\":\"Frontiers in Bioengineering and Biotechnology\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":4.3000,\"publicationDate\":\"2024-10-02\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11479935/pdf/\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Frontiers in Bioengineering and Biotechnology\",\"FirstCategoryId\":\"5\",\"ListUrlMain\":\"https://doi.org/10.3389/fbioe.2024.1388312\",\"RegionNum\":3,\"RegionCategory\":\"工程技术\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"2024/1/1 0:00:00\",\"PubModel\":\"eCollection\",\"JCR\":\"Q1\",\"JCRName\":\"BIOTECHNOLOGY & APPLIED MICROBIOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Frontiers in Bioengineering and Biotechnology","FirstCategoryId":"5","ListUrlMain":"https://doi.org/10.3389/fbioe.2024.1388312","RegionNum":3,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"2024/1/1 0:00:00","PubModel":"eCollection","JCR":"Q1","JCRName":"BIOTECHNOLOGY & APPLIED MICROBIOLOGY","Score":null,"Total":0}
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.
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.