Hypoxic chamber system for cost-effective and efficient screening of the effect of different oxygen levels on cellular growth and function

低氧室系统可经济有效地筛选不同氧气水平对细胞生长和功能的影响

• 批准号: 10324537
• 负责人: Arshak R Alexanian
• 金额: $ 22.5万
• 依托单位: CELL REPROGRAMMING AND THERAPEUTICS, LLC
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-08-15 至 2023-08-14
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10324537
• 关键词: Air; Android; Attention; Biological; Buffers; Carbon Dioxide; Cell Culture Techniques; Cell Lineage; Cell physiology; Cells; Chemicals; Consumption; Crowding; Culture Media; Development; Devices; Differentiation and Growth; Dimensions; Embryo; Environment; Equilibrium; Exhibits; Gases; Genetic; Genetic Transcription; Goals; Growth; Growth Factor; Hematopoietic; Human; Hypoxia; In Vitro; Incubators; Individual; Industrial Microbiology; Maintenance; Measurement; Mesenchymal; Mesenchymal Stem Cells; Metabolic; Methodology; Methods; Nitrogen; Nutrient; Operating System; Oxygen; Performance; Phase; Play; Production; Protocols documentation; Provider; Publications; Pump; Recombinants; Regenerative Medicine; Research; Role; Running; Signal Transduction; Stem Cell Research; Stem cell pluripotency; System; Testing; Therapeutic; Time; Tissues; Universities; Validation; base; cell growth; cost effective; drug production; experimental study; improved; in vivo; nerve stem cell; phase 2 study; pluripotency; pressure; programs; remote control; research and development; research study; response; screening; self-renewal; stem cell biology; stem cell niche; stem cells; tissue regeneration; tissue/cell culture

Regenerative medicine relies on harnessing the capacity of stem cells to grow, divide and differentiate safely and predictably. This may be in the context of expanding stem cells in vitro or encouraging their expansion, mobilization and capacity to regenerate tissues either locally or remotely in vivo. In either case, understanding the stem cell niche is fundamental to recapitulating or manipulating conditions to enable therapy. Recent studies have shown that hypoxia plays a fundamental role in the maintenance of the stem cell niche. Low oxygen (O2) conditions benefit the self-renewal of human embryonic, hematopoietic, mesenchymal, and neural stem cells, as well as improving the efficiency of genetic reprogramming to induce pluripotency. There is emerging evidence that harnessing or manipulating the hypoxic response can result in safer, more efficacious methodologies for regenerative medicine. In order to accurately reproduce different hypoxic conditions in cell reprogramming research an efficient and cost-effective system that will allow testing of multiple hypoxic conditions for a given reprogramming protocol is required. CO2/O2 incubators, currently available on the market, are too expensive and can be setup for only one hypoxic condition at a time. In addition, the consumption of nitrogen (N2) gas, used to suppress oxygen to desired low levels, is very high and in crowded labs requires the N2 gas cylinder to be replaced every two or three days. Thus, the use of CO2/O2 incubators for running experiments at different O2 levels is time consuming and expensive. Commercially available hypoxic chambers do not include gas mixers and thus rely on gas providers. To conduct multiple experiments at different O2 levels within these hypoxic chambers requires gas cylinders with different gas mixtures, the accuracy of which cannot be guaranteed. Thus, the use of these systems are time consuming and inaccurate. To resolve these problems recently we developed a multi-chamber hypoxic apparatus with remotely controlled air pump injectors that can be used to simultaneously run several experiments with different O2 levels in a single CO2/O2 incubator. The principle method of this system allows filling of the device chambers by the desired O2 concentration by coordinated opening/closing of remotely controlled air pumps as the oxygen concentration is gradually decreased (for example 5%, 4%, 3%, 2%, and 1%) in the incubator. At each O2 level, the air pump associated with each chamber will be remotely switched on for 5 min to allow equilibration of gases in the incubator and chamber. By switching off the air pump after 5 min, the air valves (with a cracking pressure of 0.087 psi) connected to inlet and outlet ports will be shut off due to absence of pump generated pressure. Thus, this will be an easy, cost-effective, and efficient approach for conducting multiple experiments simultaneously at different O2 levels in a single incubator. The goal of this Phase I proposal is to assemble a five chamber hypoxic device with individual air pump injectors that will be operated with an Android operating system based remote controller as well as to test their performance for cell culture studies.

再生医学依赖于利用干细胞安全生长、分裂和分化的能力 并且可以预见。这可能是在体外扩增干细胞或鼓励其扩增的背景下， 体内局部或远程组织的动员和再生能力。无论哪种情况，理解 干细胞生态位对于重现或操纵条件以实现治疗至关重要。最近的研究 研究表明，缺氧在干细胞生态位的维持中起着重要作用。低氧 (O2) 条件有利于人类胚胎、造血、间充质和神经干细胞的自我更新， 以及提高基因重编程诱导多能性的效率。有新的证据 利用或操纵缺氧反应可以产生更安全、更有效的方法 再生医学。为了准确再现细胞重编程中的不同缺氧条件 研究一种高效且具有成本效益的系统，该系统将允许测试给定的多种缺氧条件 需要重新编程协议。目前市场上的CO2/O2培养箱价格太贵 并且一次只能针对一种缺氧条件进行设置。另外，氮气（N2）的消耗量， 用于将氧气抑制到所需的低水平，但非常高，并且在拥挤的实验室中需要氮气瓶来 每两三天更换一次。因此，使用 CO2/O2 培养箱在不同的 O2 条件下进行实验 级别既耗时又昂贵。市售缺氧室不包括气体混合器 从而依赖天然气供应商。在这些低氧环境中进行不同 O2 水平的多次实验 室需要装有不同气体混合物的气瓶，其精度无法保证。因此， 这些系统的使用既耗时又不准确。为了解决这些问题，我们最近开发了 带有远程控制气泵注射器的多室缺氧装置，可用于 在单个 CO2/O2 培养箱中同时运行多个具有不同 O2 水平的实验。原理 该系统的方法允许通过协调以所需的 O2 浓度填充设备室 随着氧气浓度逐渐降低（对于 例如 5%、4%、3%、2% 和 1%）在培养箱中。在每个 O2 水平，与每个室相关的气泵 将远程开启 5 分钟，以平衡培养箱和腔室中的气体。通过切换 5 分钟后关闭气泵，空气阀（开启压力为 0.087 psi）连接到入口和出口 由于没有泵产生的压力，端口将被关闭。因此，这将是一种简单、具有成本效益且 在单个培养箱中以不同 O2 水平同时进行多个实验的有效方法。 第一阶段提案的目标是组装一个带有独立气泵注射器的五室低氧装置 将使用基于 Android 操作系统的遥控器进行操作并测试其 细胞培养研究的性能。