Department of Orthopedics, Graduate School of Medical Science, Kyoto Prefectural University of Medicine, Kyoto 602‑8566, Japan.
Department of Sports and Para‑Sports Medicine, Graduate School of Medical Science, Kyoto Prefectural University of Medicine, Kyoto 602‑8566, Japan.
Mol Med Rep. 2022 Jul;26(1). doi: 10.3892/mmr.2022.12745. Epub 2022 May 20.
While cartilage can be produced from induced pluripotent stem cells (iPSCs), challenges such as long culture periods and compromised tissue purity continue to prevail. The present study aimed to determine whether cartilaginous tissue could be produced from iPSCs under hypoxia and, if so, to evaluate its effects on cellular metabolism and purity of the produced tissue. Human iPSCs (hiPSCs) were cultured for cartilage differentiation in monolayers under normoxia or hypoxia (5% O), and chondrocyte differentiation was evaluated using reverse transcription‑quantitative PCR and fluorescence‑activated cell sorting. Subsequently, cartilage differentiation of hiPSCs was conducted in 3D culture under normoxia or hypoxia (5% O), and the formed cartilage‑like tissues were evaluated on days 28 and 56 using histological analyses. Hypoxia suppressed the expression levels of the immature mesodermal markers brachyury () and forkhead box protein F1; however, it promoted the expression of the chondrogenic markers and . The number of sex‑determining region Y‑box 9‑positive cells and the percentages of safranin O‑positive and type 2 collagen‑positive tissues increased under hypoxic conditions. Moreover, upon hypoxia‑inducible factor (HIF)‑1α staining, nuclei of tissues cultured under hypoxia stained more deeply compared with those of tissues cultured under normoxia. Overall, these findings indicated that hypoxia not only enhanced cartilage matrix production, but also improved tissue purity by promoting the expression of HIF‑1α gene. Potentially, pure cartilage‑like tissues could be produced rapidly and conveniently using this method.
虽然软骨可以由诱导多能干细胞（iPSCs）产生，但仍然存在培养周期长和组织纯度受损等挑战。本研究旨在确定 iPSCs 是否可以在低氧条件下产生软骨组织，如果可以，评估其对细胞代谢和产生组织纯度的影响。将人 iPSCs（hiPSCs）在单层培养物中于常氧或低氧（5% O）下培养用于软骨分化，并通过逆转录-定量 PCR 和荧光激活细胞分选评估软骨细胞分化。随后，在常氧或低氧（5% O）下进行 hiPSCs 的 3D 培养中的软骨分化，并在第 28 天和第 56 天通过组织学分析评估形成的软骨样组织。低氧抑制了未成熟中胚层标志物 brachyury（）和叉头框蛋白 F1 的表达水平；然而，它促进了软骨形成标志物和的表达。在低氧条件下，性别决定区 Y 盒 9 阳性细胞的数量以及番红 O 阳性和 II 型胶原蛋白阳性组织的百分比增加。此外，在缺氧诱导因子（HIF）-1α染色中，与在常氧下培养的组织相比，在低氧下培养的组织的细胞核染色更深。总体而言，这些发现表明，低氧不仅增强了软骨基质的产生，而且通过促进 HIF-1α 基因的表达提高了组织纯度。通过这种方法，有可能快速方便地产生纯软骨样组织。
