Indian Institute of Science Education and Research, Kolkata, India.
Indian Association of Cultivation of Science, Kolkata, India.
Cytotherapy. 2014 May;16(5):640-52. doi: 10.1016/j.jcyt.2013.09.003. Epub 2013 Nov 7.
Mesenchymal stromal cells (MSCs) have remarkable clinical potential for cell-based therapy. Wharton's jelly-derived mesenchymal stromal cells (WJ-MSCs) from umbilical cord share unique properties with both embryonic and adult stem cells. MSCs are found at low frequency in vivo, and their successful therapeutic application depends on rapid and efficient large-scale expansion in vitro. Non-muscle myosin II (NMII) has pivotal roles in different cellular activities, such as cell division, migration and differentiation. We performed this study to understand the role of NMII in proliferation and cell cycle progression in WJ-MSCs.
WJ-MSCs were cultured in the presence of blebbistatin, and cell cycle analysis was performed using flow cytometry, proliferation kinetics, senescence assay and gene expression profile using polymerase chain reaction array.
When cultured in the presence of blebbistatin, an inhibitor of NMII adenosine triphosphatase activity, WJ-MSCs exhibited dose-dependent reduction in proliferative potential along with increase in cell size and induction of early senescence. Inhibition of NMII activity also affected cell cycle progression in WJ-MSCs and led to an increase in the percentage of cells in G0/G1 phase with a corresponding reduction in the percentage of cells in G2/M phase. Blebbistatin-induced G0/G1 arrest of WJ-MSCs was further associated with up-regulation of cell cycle inhibitory genes CDKN1A, CDKN2A and CDKN2B and down-regulation of numerous genes related to progression through S and M phases of the cell cycle.
Our study demonstrates that inhibition of NMII activity in WJ-MSCs leads to G0/G1 arrest and alteration in the expression levels of certain key cell cycle-related genes.
间充质基质细胞（MSCs）在基于细胞的治疗中具有显著的临床潜力。源自脐带 Wharton 氏胶的间充质基质细胞（WJ-MSCs）与胚胎和成人干细胞具有独特的性质。MSCs 在体内的频率较低，其成功的治疗应用取决于在体外快速有效地大规模扩增。非肌肉肌球蛋白 II（NMII）在细胞分裂、迁移和分化等不同细胞活动中起着关键作用。我们进行这项研究是为了了解 NMII 在 WJ-MSCs 增殖和细胞周期进展中的作用。
WJ-MSCs 在 blebbistatin 的存在下培养，并使用流式细胞术进行细胞周期分析，使用聚合酶链反应阵列进行增殖动力学、衰老测定和基因表达谱分析。
当在 blebbistatin 的存在下培养时，一种 NMII 三磷酸腺苷酶活性的抑制剂，WJ-MSCs 表现出与增殖潜力呈剂量依赖性降低，同时细胞体积增大并诱导早期衰老。NMII 活性的抑制也影响了 WJ-MSCs 的细胞周期进展，并导致 G0/G1 期细胞百分比增加，而 G2/M 期细胞百分比相应减少。Blebbistatin 诱导的 WJ-MSCs 的 G0/G1 期阻滞与细胞周期抑制基因 CDKN1A、CDKN2A 和 CDKN2B 的上调以及与细胞周期 S 和 M 期通过相关的许多基因的下调进一步相关。
我们的研究表明，抑制 WJ-MSCs 中的 NMII 活性会导致 G0/G1 期阻滞和某些关键细胞周期相关基因表达水平的改变。
