Patient-specific iPS cells to study myeloproliferative disease.

描述（由申请人提供）：最近通过确定的因子将体细胞重编程为诱导多能干细胞（iPS）的成功不仅为在培养皿中研究复杂的人类疾病，而且为移植治疗的最终应用开辟了令人兴奋的可能性。直接重编程首次提供了使用患者特异性细胞产生复杂疾病（如家族性和散发性帕金森病）的体外和体内模型的机会。本申请中提出的实验旨在使用iPS技术来建立实验系统以研究造血系统的人类遗传疾病。该提案有三个目标： 1.从外周血中衍生iPS细胞：我们将建立使用可通过瞬时Cre表达缺失的载体从人外周血的髓样细胞中分离iPS细胞的方案。 2. iPS细胞衍生的造血细胞（HSC）的表征：我们将建立允许iPS细胞衍生的HSC体外分化的稳健方案，并评估它们在造血干细胞中移植的潜力。 NOD/SCID小鼠。为了提高植入效率，我们将通过瞬时表达重编程因子或癌基因来预处理细胞。 3.骨髓增生性肿瘤（MPN）的生物学和遗传学：我们将建立一个实验范式来研究骨髓增生性肿瘤疾病的生物学和遗传易感性 (MPN)这种疾病的特征是特异性JAK 2突变。目的1和2中开发的方法将用于研究MPN的病理学，例如评估JAK 2突变是否为造血细胞的生长提供选择性优势，以及当在体外分化时或当移植到NOD/SCID小鼠中时，源自同一患者的皮肤活检的造血干细胞是否获得突变。

DESCRIPTION (provided by applicant): The recent success in reprogramming somatic cells into induced Pluripotent Stem (iPS) cells by defined factors has opened exciting possibilities not only for the investigation of complex human diseases in the Petri dish but also for the ultimate application in transplantation therapy. Direct reprogramming provides for the first time the opportunity to generate in vitro and in vivo models of complex disorders such as familial and sporadic Parkinson's disease using patient-specific cells. The experiments proposed in this application are aimed at using the iPS technology to set up an experimental system to study a human genetic disease of the hematopoietic system. The proposal has three aims: 1. Derivation of iPS cells from peripheral blood: We will establish protocols to isolate iPS cells from myeloid cells of human peripheral blood using vectors that can be deleted by transient Cre expression. 2. Characterization of iPS cell-derived hematopoietic cells (HSCs): We will establish robust protocols allowing in vitro differentiation of iPS cell-derived HSCs and assess their potential to engraft in NOD/SCID mice. To improve engraftment efficiency we will precondition the cells by transient expression of reprogramming factors or oncogenes. 3. Biology and genetics if myeloproliferative neoplasm (MPN): We will establish an experimental paradigm to study the biology and genetic predisposition of myeloproliferative neoplastic disorder (MPN), a disorder characterized by a specific JAK2 mutation. The methods developed in aims 1 and 2 will be used to investigate the pathology of MPN such as assessing whether the JAK2 mutation provides a selective advantage to the growth of hematopoietic cells and whether hematopoietic stem cells derived from skin biopsies of the same patient acquire the mutation when differentiated in vitro or when transplanted into NOD/SCID mice.