Laboratory of Epithelial Biology, Department of Periodontics and Oral Medicine, University of Michigan School of Dentistry, Ann Arbor, MI 48109-1078, USA.
Genetics and Molecular Biology Research Unit (UPGEM), Department of Molecular Biology, School of Medicine of São José do Rio Preto, São José do Rio Preto 15090-000, SP, Brazil.
Int J Mol Sci. 2022 May 18;23(10):5633. doi: 10.3390/ijms23105633.
The human body is endowed with an extraordinary ability to maintain different oxygen levels in various tissues and organs. The maintenance of physiological levels of oxygen is known as physoxia. The development of hypoxic conditions plays an important role in the biology of several pathologies, including cancer. In vitro studies using normal and neoplastic cells require that culture conditions be carried out under appropriate oxygen levels, either physoxic or hypoxic conditions. Such requirements are difficult to widely implement in laboratory practice, mainly due to the high costs of specialized equipment. In this work, we present and characterize a cost-effective method to culture cells under a range of oxygen levels using deoxidizing pouches. Our results show that physoxic and hypoxic levels using deoxidizing absorbers can be achieved either by implementing a gradual change in oxygen levels or by a regimen of acute depletion of oxygen. This approach triggers the activation of an epithelial-mesenchymal transition in cancer cells while stimulating the expression of HIF-1α. Culturing cancer cells with deoxidizing agent pouches revealed PI3K oncogenic pathway exacerbations compared to tumor cells growing under atmospheric levels of oxygen. Similar to the PI3K signaling disturbance, we also observed augmented oxidative stress and superoxide levels and increased cell cycle arrest. Most interestingly, the culture of cancer cells under hypoxia resulted in the accumulation of cancer stem cells in a time-dependent manner. Overall, we present an attractive, cost-effective method of culturing cells under appropriate physoxic or hypoxic conditions that is easily implementable in any wet laboratory equipped with cell culture tools.
人体具有在不同组织和器官中维持不同氧水平的非凡能力。维持氧的生理水平被称为氧合作用。缺氧条件的发展在包括癌症在内的几种病理学的生物学中起着重要作用。在使用正常和肿瘤细胞的体外研究中，需要在适当的氧水平下进行培养条件，无论是氧合作用还是缺氧条件。在实验室实践中，这些要求很难广泛实施，主要是因为专门设备的成本高。在这项工作中，我们提出并描述了一种使用脱氧袋在一系列氧水平下培养细胞的经济有效的方法。我们的结果表明，使用脱氧吸收剂可以通过实施氧水平的逐渐变化或通过急性耗氧方案来实现氧合和缺氧水平。这种方法会触发癌细胞中上皮-间充质转化的激活，同时刺激 HIF-1α 的表达。与在大气氧水平下生长的肿瘤细胞相比，用脱氧剂袋培养的癌细胞显示出 PI3K 致癌途径的加剧。与 PI3K 信号干扰相似，我们还观察到氧化应激和超氧化物水平增加以及细胞周期停滞增加。最有趣的是，癌细胞在缺氧条件下培养会以时间依赖性方式积累癌症干细胞。总的来说，我们提出了一种有吸引力的、经济有效的方法，可以在任何配备细胞培养工具的湿实验室中轻松实施，以在适当的氧合或缺氧条件下培养细胞。
