Bagherpoor Alireza Jian, Kučírek Martin, Fedr Radek, Sani Soodabeh Abbasi, Štros Michal
Institute of Biophysics of the Czech Academy of Sciences, Královopolská 135, 612 00 Brno, Czech Republic.
Biomolecules. 2020 Oct 15;10(10):1450. doi: 10.3390/biom10101450.
HMGB1 and HMGB2 proteins are abundantly expressed in human embryonic stem cells (hESCs) and hESC-derived progenitor cells (neuroectodermal cells, hNECs), though their functional roles in pluripotency and the mechanisms underlying their differentiation in response to the anticancer drug etoposide remain to be elucidated. Here, we show that and/or knockdown (KD) by shRNA in hESCs did not affect the cell stemness/pluripotency regardless of etoposide treatments, while in hESC-derived neuroectodermal cells, treatment resulted in differential effects on cell survival and the generation of rosette structures. The objective of this work was to determine whether HMGB1/2 proteins could modulate the sensitivity of hESCs and hESC-derived progenitor cells (hNECs) to etoposide. We observed that KD knockdown (KD) and, to a lesser extent, KD enhanced the sensitivity of hESCs to etoposide. Enhanced accumulation of 53BP1 on telomeres was detected by confocal microscopy in both untreated and etoposide-treated KD hESCs and hNECs, indicating that the loss of could destabilize telomeres. On the other hand, decreased accumulation of 53BP1 on telomeres in etoposide-treated KD hESCs (but not in KD hNECs) suggested that the loss of promoted the stability of telomeres. Etoposide treatment of hESCs resulted in a significant enhancement of telomerase activity, with the highest increase observed in the KD cells. Interestingly, no changes in telomerase activity were found in etoposide-treated control hNECs, but KD (unlike KD) markedly decreased telomerase activity in these cells. Changes in telomerase activity in the etoposide-treated KD hESCs or hNECs coincided with the appearance of DNA damage markers and could already be observed before the onset of apoptosis. Collectively, we have demonstrated that HMGB1 or HMGB2 differentially modulate the impact of etoposide treatment on human embryonic stem cells and their progenitor cells, suggesting possible strategies for the enhancement of the efficacy of this anticancer drug.
HMGB1和HMGB2蛋白在人胚胎干细胞（hESCs）和hESC衍生的祖细胞（神经外胚层细胞，hNECs）中大量表达，尽管它们在多能性中的功能作用以及它们在响应抗癌药物依托泊苷时的分化机制仍有待阐明。在此，我们表明，在hESCs中通过shRNA敲低（KD）HMGB1和/或HMGB2，无论是否进行依托泊苷处理，均不影响细胞干性/多能性，而在hESC衍生的神经外胚层细胞中，处理对细胞存活和玫瑰花结结构的形成产生不同影响。这项工作的目的是确定HMGB1/2蛋白是否可以调节hESCs和hESC衍生的祖细胞（hNECs）对依托泊苷的敏感性。我们观察到，HMGB1敲低（KD）以及在较小程度上HMGB2敲低增强了hESCs对依托泊苷的敏感性。通过共聚焦显微镜在未处理和经依托泊苷处理的HMGB1 KD hESCs和hNECs中均检测到53BP1在端粒上的积累增加，表明HMGB1的缺失可能会破坏端粒的稳定性。另一方面，在经依托泊苷处理的HMGB2 KD hESCs（但不是在HMGB2 KD hNECs中）中端粒上53BP1的积累减少，表明HMGB2的缺失促进了端粒的稳定性。用依托泊苷处理hESCs导致端粒酶活性显著增强，在HMGB1 KD细胞中观察到的增加最为明显。有趣的是，在经依托泊苷处理的对照hNECs中未发现端粒酶活性的变化，但HMGB2 KD（与HMGB1 KD不同）显著降低了这些细胞中的端粒酶活性。在经依托泊苷处理的HMGB1 KD hESCs或hNECs中端粒酶活性的变化与DNA损伤标志物的出现一致，并且在细胞凋亡开始之前就已经可以观察到。总的来说，我们已经证明HMGB1或HMGB2对依托泊苷处理对人胚胎干细胞及其祖细胞的影响具有不同的调节作用，这为提高这种抗癌药物的疗效提供了可能的策略。
