Human Cells and Tissues Core: 3D Models of Engineered Human iPS Cells to Investigate Neurotropic Virus Infections
人类细胞和组织核心:用于研究嗜神经病毒感染的工程化人类 iPS 细胞 3D 模型
基本信息
- 批准号:9312528
- 负责人:
- 金额:$ 25.87万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2017
- 资助国家:美国
- 起止时间:2017-04-01 至 2022-03-31
- 项目状态:已结题
- 来源:
- 关键词:AddressAdherent CultureAstrocytesCD44 geneCell Differentiation processCell SeparationCellsCerebrumCiliary Neurotrophic FactorCommunicable DiseasesCore FacilityDimensionsEGF geneElectrophysiology (science)EmbryoExcisionExposure toFGF2 geneFibroblastsFlavivirus InfectionsGenesGlial Fibrillary Acidic ProteinGoalsGrowth FactorHumanHuman EngineeringImmunofluorescence ImmunologicInstitutesMagnetismMeasuresMicrocephalyMicrospheresNCAM1 geneNeuronsOligodendrogliaOrganoidsOutputPathogenesisPopulationProductionProtocols documentationResearchResearch PersonnelResearch Project GrantsTestingTissue ModelTissuesTriiodothyronineVirus DiseasesWorkZika Virusbasedrug candidatehuman embryonic stem cellhuman tissueinduced pluripotent stem cellinfectious disease modelmulti-electrode arraysmutantnerve stem cellnestin proteinneurotropic virusoligodendrocyte precursorprogenitorrelating to nervous systemscreeningstemsuccessthree dimensional cell culturethree-dimensional modelingvirus tropism
项目摘要
The MIT Center for Human Tissue Models of Infectious Diseases (MIT.HTMID) will study viral infections of five
types of neural cells that are differentiated from human embryonic stem (ES) cells or human induced
pluripotent stem (iPS) cells. In addition, MIT.HTMID research will address viral infections of human three
dimensional (3D) cerebral organoids toward understanding Zika virus microcephaly. All cell work and tissue
work for the Center’s research will be supported by the Human Cells and Tissues Core facility. The Core will
be located at the Whitehead Institute, and will provide a continuous supply of cells for both HT.MID research
projects. To prepare cells and organoids from iPS cells, human fibroblasts are reprogrammed into pluripotent
iPS cells. Next, the pluripotent iPS cells are differentiated into multipotent neural progenitors (NPs). Neurons,
astrocytes and oligodendrocyte are differentiated from NPs. To generate a homogenous population of NPs, a
SMADinhibitionbased neural differentiation protocol is routinely used. Adherent cultures of control and mutant
ESCs/iPSCs will be differentiated into neural rosettes expressing Pax6 and Nestin as detected by
immunofluorescence, and further expanded as neuronal progenitors in the presence of bFGF. Multipotent NPs
are differentiated into neurons. To generate neurons, NPs are cultured in a growth factor depleted medium that
promotes terminal differentiation and maturation of a mixed populations of neurons. These neurons are
electrophysiologically active, as measured on multielectrode arrays. Multipotent neural progenitors are
separately differentiated into astrocytes or oligodendrocytes. To derive astrocytes from human NPs, we have
developed protocols that allow the production of highly homogeneous populations. Specifically, FACSsorted
PSANCAM/A2B5 glial progenitors, derived from NPs, are expanded in the presence of EGF and bFGF to
+
derive S100b+ immature astrocytes. Further maturation of this FACSsorted population, in the presence of
CNTF, yields GFAP+ astrocytes. The cells can be sorted to high purity by magnetic cell sorting (MACS) against
CD44 microbeads. To generate oligodendrocytes, A2B5+ glial progenitors will be expanded in bFGF and
PDGFα, and further sorted by MACS against O4 and CD140a markers. These fated oligodendrocyte
precursors are matured into MBP+
oligodendrocytes after growth factor removal and exposure to the thyroid
hormone T3. The project investigators and core staff will meet regularly to plan work flow and anticipate cell
needs for Projects 1 and 2. The Human Cells and Tissues core will be an integral and essential part of the
MIT.HTMID Center, and its output will enhance the project’s potential for success
麻省理工学院传染病人类组织模型中心(MIT.HTMID)将研究五种病毒感染
从人胚胎干(ES)细胞分化或人诱导的神经细胞类型
此外,麻省理工学院的HTMID研究将解决人类三种病毒感染的问题,
三维(3D)大脑类器官,以了解寨卡病毒小头畸形。所有细胞工作和组织
该中心的研究工作将得到人类细胞和组织核心设施的支持。该核心设施将
位于Whitehead研究所,并将为HT.MID研究提供持续的细胞供应。
为了从iPS细胞制备细胞和类器官,将人成纤维细胞重编程为多能干细胞。
接下来,将多能iPS细胞分化成多能神经祖细胞(NP)。 神经元,
星形胶质细胞和少突胶质细胞从NP分化。
常规使用基于SMAD抑制剂的神经分化方案。
ESC/iPSC将分化为表达Pax 6和Nestin的神经细胞,如通过免疫组织化学检测的。
用免疫荧光法检测细胞增殖,并在bFGF存在下进一步扩增为神经元祖细胞。
为了产生神经元,将NP在生长因子耗尽的培养基中培养,
促进神经元混合群体的终末分化和成熟。这些神经元是
电生理活性,如在多个电极阵列上测量的。 多能神经祖细胞是
为了从人NP中获得星形胶质细胞,我们
开发了允许产生高度同质群体的方案。具体地说,
在EGF和bFGF存在下扩增来源于NP的PSA/NCAM β/A2 B5神经胶质祖细胞,
+
进一步成熟的这种FACS分选的群体,在S100 b+的存在下,
CNTF,产生GFAP+星形胶质细胞。
为了产生少突胶质细胞,A2 B5+神经胶质祖细胞将在bFGF中扩增,
用O 4和CD 140 a标记的MACS进一步分选少突胶质细胞,
前体成熟为MBP+
去除生长因子并暴露于甲状腺后的少突胶质细胞
激素T3。项目调查人员和核心人员将定期开会,计划工作流程,并预测细胞
人类细胞和组织核心将是项目1和2的一个不可分割的重要组成部分。
HTMID中心,其输出将提高项目的成功潜力
项目成果
期刊论文数量(0)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
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Lee Gehrke其他文献
Lee Gehrke的其他文献
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{{ truncateString('Lee Gehrke', 18)}}的其他基金
Administrative Core: 3D Models of Engineered Human iPS Cells to Investigate Neurotropic Virus Infections
管理核心:用于研究嗜神经病毒感染的工程化人类 iPS 细胞 3D 模型
- 批准号:
9312526 - 财政年份:2017
- 资助金额:
$ 25.87万 - 项目类别:
3D Models of Engineered Human iPS Cells to Investigate Neurotropic Virus Infections
用于研究嗜神经病毒感染的工程化人类 iPS 细胞 3D 模型
- 批准号:
9903201 - 财政年份:2017
- 资助金额:
$ 25.87万 - 项目类别:
Project 2: Use of 2D cultures and 3D organoids to identify candidate antiviral compounds; to use genetic approaches to identify host genes that promote or protect against flavivirus infection
项目 2:使用 2D 培养物和 3D 类器官来鉴定候选抗病毒化合物;
- 批准号:
9312530 - 财政年份:2017
- 资助金额:
$ 25.87万 - 项目类别:
MMDx: A rapid multiplexed matrix code diagnostic for real time epidemiology
MMDx:用于实时流行病学的快速多路复用矩阵代码诊断
- 批准号:
8915035 - 财政年份:2012
- 资助金额:
$ 25.87万 - 项目类别:
Synergistic innate immune activation and cell killing by RIG-I ligands in HCV-HCC
HCV-HCC 中 RIG-I 配体的协同先天免疫激活和细胞杀伤
- 批准号:
8441526 - 财政年份:2012
- 资助金额:
$ 25.87万 - 项目类别:
Synergistic innate immune activation and cell killing by RIG-I ligands in HCV-HCC
HCV-HCC 中 RIG-I 配体的协同先天免疫激活和细胞杀伤
- 批准号:
8238622 - 财政年份:2012
- 资助金额:
$ 25.87万 - 项目类别:
MMDx: A rapid multiplexed matrix code diagnostic for real time epidemiology
MMDx:用于实时流行病学的快速多路复用矩阵代码诊断
- 批准号:
8467676 - 财政年份:2012
- 资助金额:
$ 25.87万 - 项目类别:
MMDx: A rapid multiplexed matrix code diagnostic for real time epidemiology
MMDx:用于实时流行病学的快速多路复用矩阵代码诊断
- 批准号:
8901539 - 财政年份:2012
- 资助金额:
$ 25.87万 - 项目类别:
MMDx: A rapid multiplexed matrix code diagnostic for real time epidemiology
MMDx:用于实时流行病学的快速多路复用矩阵代码诊断
- 批准号:
8301236 - 财政年份:2012
- 资助金额:
$ 25.87万 - 项目类别:
CONTROL OF CELL FUNCTION VIA SELECTIVE MRNA TRANSLATION
通过选择性 mRNA 翻译控制细胞功能
- 批准号:
2857127 - 财政年份:1989
- 资助金额:
$ 25.87万 - 项目类别:
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