Doctor of Philosophy Program in Medical Sciences, Faculty of Medicine, Chulalongkorn University, Bangkok, Thailand.
Department of Medical Technology, Faculty of Allied Health Sciences, Naresuan University, Phitsanulok, Thailand.
Sci Rep. 2021 Mar 4;11(1):5255. doi: 10.1038/s41598-021-83895-6.
Activating mutations affecting the JAK-STAT signal transduction is the genetic driver of myeloproliferative neoplasms (MPNs) which comprise polycythemia vera (PV), essential thrombocythemia (ET) and myelofibrosis. The JAK2p.V617F mutation can produce both erythrocytosis in PV and thrombocytosis in ET, while JAK2 exon 12 mutations cause only erythrocytosis. We hypothesized that these two mutations activated different intracellular signals. In this study, the induced pluripotent stem cells (iPSCs) were used to model JAK2-mutated MPNs. Normal iPSCs underwent lentiviral transduction to overexpress JAK2p.V617F or JAK2p.N542_E543del (JAK2exon12) under a doxycycline-inducible system. The modified iPSCs were differentiated into erythroid cells. Compared with JAK2V617F-iPSCs, JAK2exon12-iPSCs yielded more total CD71GlycophorinA erythroid cells, displayed more mature morphology and expressed more adult hemoglobin after doxycycline induction. Capillary Western immunoassay revealed significantly higher phospho-STAT1 but lower phospho-STAT3 and lower Phospho-AKT in JAK2exon12-iPSCs compared with those of JAK2V617F-iPSCs in response to erythropoietin. Furthermore, interferon alpha and arsenic trioxide were tested on these modified iPSCs to explore their potentials for MPN therapy. Both agents preferentially inhibited proliferation and promoted apoptosis of the iPSCs expressing mutant JAK2 compared with those without doxycycline induction. In conclusion, the modified iPSC model can be used to investigate the mechanisms and search for new therapy of MPNs.
激活影响 JAK-STAT 信号转导的突变是骨髓增殖性肿瘤（MPN）的遗传驱动因素，包括真性红细胞增多症（PV）、特发性血小板增多症（ET）和骨髓纤维化。JAK2p.V617F 突变可导致 PV 红细胞增多和 ET 血小板增多，而 JAK2 外显子 12 突变仅导致红细胞增多。我们假设这两种突变激活了不同的细胞内信号。在这项研究中，诱导多能干细胞（iPSC）被用于模拟 JAK2 突变的 MPN。正常的 iPSC 通过慢病毒转导，在四环素诱导系统下过表达 JAK2p.V617F 或 JAK2p.N542_E543del（JAK2 外显子 12）。修饰后的 iPSC 分化为红细胞。与 JAK2V617F-iPSC 相比，JAK2exon12-iPSC 在四环素诱导后产生更多的总 CD71GlycophorinA 红细胞，显示出更成熟的形态，并表达更多的成人血红蛋白。毛细管 Western 免疫印迹显示，JAK2exon12-iPSC 在对红细胞生成素的反应中，磷酸化 STAT1 显著升高，但磷酸化 STAT3 和磷酸化 AKT 降低。此外，还在这些修饰的 iPSC 上测试了干扰素 alpha 和三氧化二砷，以探索它们在 MPN 治疗中的潜力。与未诱导四环素的 iPSC 相比，这两种药物都优先抑制表达突变 JAK2 的 iPSC 的增殖，并促进其凋亡。总之，修饰后的 iPSC 模型可用于研究 MPN 的机制并寻找新的治疗方法。
