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21. 发明申请

US20090104626A1 METHODS OF ISOLATING BIPOTENT HEPATIC PROGENITOR CELLS 有权
公开(公告)号：US20090104626A1
公开(公告)日：2009-04-23
申请号：US12326514
申请日：2008-12-02
申请人： Hiroshi KUBOTA , Lola M. Reid
发明人： Hiroshi KUBOTA , Lola M. Reid
IPC分类号： G01N33/53 , C12N15/09
CPC分类号： G01N33/5091 , C12N5/0672 , C12N2501/50 , C12N2501/58 , C12N2501/599
摘要： A method of obtaining a mixture of cells enriched in hepatic progenitors is developed which comprises methods yielding suspensions of a mixture of cell types, and selecting those cells that are classical MHC class I antigen(s) negative and ICAM-1 antigen positive. The weak or dull expression of nonclassical MHC class I antigen(s) can be used for further enrichment of hepatic progenitors. Furthermore, the progenitors can be selected to have a level of side scatter, a measure of granularity or cytoplasmic droplets, that is higher than that in non-parenchymal cells, such as hemopoietic cells, and lower than that in mature parenchymal cells, such as hepatocytes. Furthermore, the progeny of the isolated progenitors can express alpha-fetoprotein and/or albumin and/or CK19. The hepatic progenitors, so isolated, can grow clonally, that is an entire population of progeny can be derived from one cell. The clones of progenitors have a growth pattern in culture of piled-up aggregates or clusters. These methods of isolating the hepatic progenitors are applicable to any vertebrates including human. The hepatic progenitor cell population is expected to be useful for cell therapies, for bioartificial livers, for gene therapies, for vaccine development, and for myriad toxicological, pharmacological, and pharmaceutical programs and investigations.

22. 发明申请

US20090053758A1 Processes for clonal growth of hepatic progenitor cells 审中-公开
标题翻译：肝祖细胞克隆生长过程
公开(公告)号：US20090053758A1
公开(公告)日：2009-02-26
申请号：US12289019
申请日：2008-10-17
申请人： Hiroshi Kubota , Lola M. Reid
发明人： Hiroshi Kubota , Lola M. Reid
IPC分类号： C12Q1/18 , C12Q1/02 , C12N5/06
CPC分类号： C12N5/0672 , C12N2500/20 , C12N2500/25 , C12N2500/36 , C12N2500/90 , C12N2501/11 , C12N2501/39 , C12N2502/13
摘要： A method of propagating mammalian endodermally derived progenitors such as hepatic progenitors, their progeny, or mixtures thereof is developed which includes culturing mammalian progenitors, their progeny, or mixtures thereof on a layer of embryonic mammalian feeder cells in a culture medium. The culture medium can be supplemented with one or more hormones and other growth agents. These hormones and other growth agents can include insulin, dexamethasone, transferrin, nicotinamide, serum albumin, β-mercaptoethanol, free fatty acid, glutamine, CuSO4, and H2SeO3. The culture medium can also include antibiotics. Importantly, the culture medium does not include serum.The invention includes means of inducing the differentiation of the progenitors to their adult fates such as the differentiation of hepatic progenitor cells to hepatocytes or biliary cells by adding, or excluding epidermal growth factor, respectively.The method of producing mammalian progenitors is useful in that the progenitors can be used subsequently in one or more of the following processes: identification of growth and differentiation factors, toxicological studies, drug development, antimicrobial studies, or the preparation of an extracorporeal organ such as a bioartificial liver.
摘要翻译：开发了传播哺乳动物内皮祖细胞如肝祖细胞，其后代或其混合物的方法，其包括在培养基中的胚胎哺乳动物饲养细胞层上培养哺乳动物祖细胞，其后代或其混合物。培养基可以补充一种或多种激素和其它生长剂。这些激素和其它生长剂可包括胰岛素，地塞米松，转铁蛋白，烟酰胺，血清白蛋白，β-巯基乙醇，游离脂肪酸，谷氨酰胺，CuSO 4和H 2 SeO 3。培养基也可以包括抗生素。重要的是，培养基不包括血清。本发明包括诱导祖细胞分化成其成体的方法，例如通过分别添加或排除表皮生长因子将肝祖细胞分化为肝细胞或胆汁细胞。生产哺乳动物祖细胞的方法是有用的，因为可以随后在以下一个或多个方法中使用祖细胞：识别生长和分化因子，毒理学研究，药物开发，抗微生物研究或体外器官的制备，例如生物人造肝。

23. 发明授权

US6069005A Hapatoblasts and method of isolating same 失效
公开(公告)号：US6069005A
公开(公告)日：2000-05-30
申请号：US757336
申请日：1996-11-27
申请人： Lola M. Reid , Samuel H. Sigal , Shlomo Brill , Patricia A. Holst
发明人： Lola M. Reid , Samuel H. Sigal , Shlomo Brill , Patricia A. Holst
IPC分类号： A61K35/12 , C12N5/00 , C12N5/074 , G01N33/50 , G01N33/574 , C12N5/02
CPC分类号： C12N5/0068 , C12N5/0672 , G01N33/5017 , G01N33/57438 , A61K35/12 , C12N2500/20 , C12N2500/22 , C12N2500/25 , C12N2500/36 , C12N2500/90 , C12N2501/11 , C12N2501/305 , C12N2501/315 , C12N2501/39 , C12N2501/395 , C12N2509/00 , C12N2510/00
摘要： This invention relates to methods of isolating hepatoblasts utilizing panning techniques and fluorescence activated cell sorting. This invention further relates to isolated hepatoblasts and to a method of treating liver dysfunction as well as to methods of forming artificial livers.

24. 发明授权

US5576207A Method of expanding hepatic precursor cells 失效
标题翻译：扩大肝前体细胞的方法
公开(公告)号：US5576207A
公开(公告)日：1996-11-19
申请号：US265696
申请日：1994-06-24
申请人： Lola M. Reid , Maria Agelli , Andreas Ochs
发明人： Lola M. Reid , Maria Agelli , Andreas Ochs
IPC分类号： A61K35/12 , C12N5/00 , C12N5/074 , C12N15/09 , C12N5/06 , C12N5/08
CPC分类号： C12N5/0672 , C12N5/0068 , A61K35/12 , C12N2500/20 , C12N2500/22 , C12N2500/25 , C12N2500/36 , C12N2500/90 , C12N2501/11 , C12N2501/305 , C12N2501/315 , C12N2501/39 , C12N2501/395 , C12N2502/02 , C12N2510/00 , C12N2533/52 , C12N2533/54 , C12N2533/70 , C12N2533/90
摘要： A composition which comprises an animal cell population, and which contains immature animal cells. The immature animal cells are characterized by expression of alpha-fetoprotein or lack of essential expression of alpha-fetoprotein and albumin, and at least a portion of said immature animal cells or at least a portion of the progeny of said immature cells is capable of differentiating into cells which express albumin. The cell population is cultured under conditions which result in expansion of the cells. Expansion of the cells may be achieved by culturing the cells in the presence of an extracellular matrix and liver stromal cells; and preferably in the presence of growth factors. Such cells may be used for liver transplantation, artificial livers, and for toxicology and pharmacology studies. Such cells may also be genetically engineered to express proteins or polypeptides of interest.
摘要翻译：包含动物细胞群并且含有未成熟动物细胞的组合物。未成熟动物细胞的特征在于甲胎蛋白的表达或甲胎蛋白和白蛋白的基本表达的缺失，并且所述未成熟动物细胞的至少一部分或所述未成熟细胞的后代的至少一部分能够分化进入表达白蛋白的细胞。细胞群在导致细胞扩增的条件下培养。可以通过在细胞外基质和肝脏基质细胞的存在下培养细胞来实现细胞的扩增; 并且优选在生长因子存在下。这些细胞可用于肝移植，人肝，以及毒理学和药理学研究。这样的细胞也可以被遗传工程化以表达感兴趣的蛋白质或多肽。

25. 发明授权

US08709800B2 Methods of isolating bipotent hepatic progenitor cells 有权
公开(公告)号：US08709800B2
公开(公告)日：2014-04-29
申请号：US12265381
申请日：2008-11-05
申请人： Hiroshi Kubota , Lola M. Reid
发明人： Hiroshi Kubota , Lola M. Reid
IPC分类号： C12N5/00 , C12N15/09
CPC分类号： G01N33/5091 , C12N5/0672 , C12N2501/50 , C12N2501/58 , C12N2501/599
摘要： A method of obtaining a mixture of cells enriched in hepatic progenitors is developed which comprises methods yielding suspensions of a mixture of cell types, and selecting those cells that are classical MHC class I antigen(s) negative and ICAM-1 antigen positive. The weak or dull expression of nonclassical MHC class I antigen(s) can be used for further enrichment of hepatic progenitors. Furthermore, the progenitors can be selected to have a level of side scatter, a measure of granularity or cytoplasmic droplets, that is higher than that in non-parenchymal cells, such as hemopoietic cells, and lower than that in mature parenchymal cells, such as hepatocytes. Furthermore, the progeny of the isolated progenitors can express alpha-fetoprotein and/or albumin and/or CK19. The hepatic progenitors, so isolated, can grow clonally, that is an entire population of progeny can be derived from one cell. The clones of progenitors have a growth pattern in culture of piled-up aggregates or clusters. These methods of isolating the hepatic progenitors are applicable to any vertebrates including human. The hepatic progenitor cell population is expected to be useful for cell therapies, for bioartificial livers, for gene therapies, for vaccine development, and for myriad toxicological, pharmacological, and pharmaceutical programs and investigations.

26. 发明授权

US07759118B2 Proliferation of hepatocyte precursors 失效
标题翻译：肝细胞前体的增殖
公开(公告)号：US07759118B2
公开(公告)日：2010-07-20
申请号：US11622902
申请日：2007-01-12
申请人： Lola M. Reid , Maria Agelli , Andreas Ochs
发明人： Lola M. Reid , Maria Agelli , Andreas Ochs
IPC分类号： C12N5/00
CPC分类号： C12N5/0672 , A61K35/12 , C12N5/0068 , C12N2500/20 , C12N2500/22 , C12N2500/25 , C12N2500/36 , C12N2500/90 , C12N2501/11 , C12N2501/305 , C12N2501/315 , C12N2501/39 , C12N2501/395 , C12N2502/02 , C12N2510/00 , C12N2533/52 , C12N2533/54 , C12N2533/70 , C12N2533/90
摘要： A composition which comprises an animal cell population which contains immature animal cells. The immature animal cells are characterized by expression of alpha-fetoprotein or lack of essential expression of alpha-fetoprotein and albumin, and at least a portion of said immature animal cells or at least a portion of the progeny of said immature animal cells is capable of differentiating into cells which express albumin. The cell population is cultured under conditions which result in expansion of the cells. Expansion of the cells may be achieved by culturing the cells in the presence of an extracellular matrix and liver stromal cells; and preferably in the presence of growth factors. Such cells may be used for liver transplantation, artificial livers, and for toxicology and pharmacology studies. Such cells may also be genetically engineered to express proteins or polypepetides of interest.
摘要翻译：包含含有未成熟动物细胞的动物细胞群的组合物。未成熟动物细胞的特征在于甲胎蛋白的表达或甲胎蛋白和白蛋白的必需表达的缺失，并且所述未成熟动物细胞的至少一部分或所述未成熟动物细胞的后代的至少一部分能够分化成表达白蛋白的细胞。细胞群在导致细胞扩增的条件下培养。可以通过在细胞外基质和肝脏基质细胞的存在下培养细胞来实现细胞的扩增; 并且优选在生长因子存在下。这些细胞可用于肝移植，人肝，以及毒理学和药理学研究。这样的细胞也可以被遗传工程化以表达感兴趣的蛋白质或多肽。

27. 发明申请

US20080293133A1 Textiles for use in bioreactors for expansion and maintenance of cells 审中-公开
标题翻译：生物反应器用于细胞扩增和维持的纺织品
公开(公告)号：US20080293133A1
公开(公告)日：2008-11-27
申请号：US12153170
申请日：2008-05-14
申请人： Lola M. Reid , Bhupender S. Gupta , Jeffrey M. MacDonald
发明人： Lola M. Reid , Bhupender S. Gupta , Jeffrey M. MacDonald
IPC分类号： C12M3/00 , C12M1/00
CPC分类号： C12M23/06 , A61F2/06 , C12M21/08 , C12M23/20 , C12M23/34 , C12M23/58 , C12M25/06 , C12M29/10 , C12N5/0671 , C12N2501/11 , C12N2501/12 , C12N2501/148 , C12N2533/30 , C12N2535/10
摘要： A bioreactor for three-dimensional culture of liver cells is disclosed. The device is characterized by the use of textile vasculatures. A model and method for optimizing vasculature parameters is also disclosed. Liver acinar structure and physiological parameters are mimicked by sandwiching cells in the space between the two innermost woven textile hollow fibers, and creating radial flow of media from an outer compartment, through the cell mass compartment, and to an inner compartment. The theoretical optimum hydraulic permeability for the two innermost semi-permeable membranes is determined based on physiological hepatic sinusoidal blood flow and pressures. Experimental studies using a flow rate and pressure monitoring systems in conjunction with phase-contrast velocity-encoded MRI confirm theoretical results. Novel woven vascular tubes with optimum hydraulic permeability are disclosed for culturing hepatocytes in the multi-coaxial bioreactor.
摘要翻译：公开了用于肝细胞三维培养的生物反应器。该装置的特征在于使用纺织品血管系统。还公开了优化血管系统参数的模型和方法。通过将细胞夹在两个最内层的织造中空纤维之间的空间中，并且从外隔室，通过细胞块隔室和内隔室产生介质的径向流动来模拟肝腺泡结构和生理参数。基于生理肝脏正弦血流和压力确定两个最内半透膜的理论最佳水力渗透性。使用流速和压力监测系统结合相位对比度速度编码MRI的实验研究证实了理论结果。公开了具有最佳水力渗透性的新型编织血管，用于在多同轴生物反应器中培养肝细胞。

28. 发明授权

US07332589B2 Variants of alpha-fetoprotein coding and expression sequences 有权
标题翻译：甲胎蛋白编码和表达序列的变体
公开(公告)号：US07332589B2
公开(公告)日：2008-02-19
申请号：US10254676
申请日：2002-09-26
申请人： Hiroshi Kubota , Robert Storms , Lola M. Reid
发明人： Hiroshi Kubota , Robert Storms , Lola M. Reid
IPC分类号： C07H21/02 , C07H21/04 , C12Q1/68
CPC分类号： C07K14/4715
摘要： The invention discloses the sequences of variant forms of alpha-fetoprotein transcripts that have been identified in human hemopoietic progenitors but not in differentiated mature cells. The variant forms of AFP (vAFP) cDNA sequences isolated from a multipotent hemopoietic cell line, K562, differ from the authentic AFP transcript, consisting of 15 exons, by lacking only exon 1. Instead of exon 1, vAFP transcripts use an additional one or two exons located in the 5′-untranslated region of the AFP gene. K562 expressed selectively vAFP, whereas a hepatocellular carcinoma cell line, HepG2, showed no detectable expression of vAFP. In normal adult tissues, vAFP transcripts is detected in the bone marrow, thymus and brain, but not the spleen, suggesting the expression occurs in normal hemopoietic progenitors. Moreover, CD34+Lin− hemopoietic stem/progenitor cells purified by flow cytometric sorting also express the variant transcripts.
摘要翻译：本发明公开了已经在人造血祖细胞中鉴定但不在分化的成熟细胞中的甲胎蛋白转录物的变体形式的序列。从多能造血细胞系K562分离的AFP（vAFP）cDNA序列的变体形式与仅由外显子1缺失的15个外显子组成的真正的AFP转录物不同。代替外显子1，vAFP转录物使用另外一个或位于AFP基因的5'非翻译区的两个外显子。 K562选择性表达vAFP，而肝细胞癌细胞系HepG2则显示无可检测的vAFP表达。在正常的成年组织中，在骨髓，胸腺和脑中检测到vAFP转录物，而不是脾脏，表明在正常造血祖细胞中发生表达。此外，通过流式细胞分选纯化的CD34 +造血干细胞/祖细胞也表达变体转录物。

29. 发明授权

US6146889A Proliferation of hepatocyte precursors 失效
标题翻译：肝细胞前体的增殖
公开(公告)号：US6146889A
公开(公告)日：2000-11-14
申请号：US115920
申请日：1998-07-15
申请人： Lola M. Reid , Maria Agelli
发明人： Lola M. Reid , Maria Agelli
IPC分类号： A61K35/12 , C12N5/00 , C12N5/074 , C12N15/09 , C12N5/08 , C12N5/10
CPC分类号： C12N5/0672 , C12N5/0068 , A61K35/12 , C12N2500/20 , C12N2500/22 , C12N2500/25 , C12N2500/36 , C12N2500/90 , C12N2501/11 , C12N2501/305 , C12N2501/315 , C12N2501/39 , C12N2501/395 , C12N2502/02 , C12N2510/00 , C12N2533/52 , C12N2533/54 , C12N2533/70 , C12N2533/90
摘要： A composition which comprises an animal cell population which contains immature animal cells. The immature animal cells are characterized by expression of alpha-fetoprotein or lack of essential expression of alpha-fetoprotein and albumin, and at least a portion of said immature animal cells or at least a portion of the progeny of said immature animal cells is capable of differentiating into cells which express albumin. The cell population is cultured under conditions which result in expansion of the cells. Expansion of the cells may be achieved by culturing the cells in the presence of an extracellular matrix and liver stromal cells; and preferably in the presence of growth factors. Such cells may be used for liver transplantation, artificial livers, and for toxicology and pharmacology studies. Such cells may also be genetically engineered to express proteins or polypepetides of interest.
摘要翻译：包含含有未成熟动物细胞的动物细胞群的组合物。未成熟动物细胞的特征在于甲胎蛋白的表达或甲胎蛋白和白蛋白的必需表达的缺失，并且所述未成熟动物细胞的至少一部分或所述未成熟动物细胞的后代的至少一部分能够分化成表达白蛋白的细胞。细胞群在导致细胞扩增的条件下培养。可以通过在细胞外基质和肝脏基质细胞的存在下培养细胞来实现细胞的扩增; 并且优选在生长因子存在下。这些细胞可用于肝移植，人肝，以及毒理学和药理学研究。这样的细胞也可以被遗传工程化以表达感兴趣的蛋白质或多肽。

30. 发明授权

US5789246A Compositions comprising hepatocyte precursors 失效
标题翻译：包含肝细胞前体的组合物
公开(公告)号：US5789246A
公开(公告)日：1998-08-04
申请号：US751546
申请日：1996-11-18
申请人： Lola M. Reid , Maria Agelli , Andreas Ochs
发明人： Lola M. Reid , Maria Agelli , Andreas Ochs
IPC分类号： A61K35/12 , C12N5/00 , C12N5/074 , C12N15/09 , C12N5/06 , C12N5/08 , C12N5/10
CPC分类号： C12N5/0672 , C12N5/0068 , A61K35/12 , C12N2500/20 , C12N2500/22 , C12N2500/25 , C12N2500/36 , C12N2500/90 , C12N2501/11 , C12N2501/305 , C12N2501/315 , C12N2501/39 , C12N2501/395 , C12N2502/02 , C12N2510/00 , C12N2533/52 , C12N2533/54 , C12N2533/70 , C12N2533/90
摘要： A composition which comprises an animal cell population, and which contains immature animal cells. The immature animal cells are characterized by expression of alpha-fetoprotein or lack of essential expression of alpha-fetoprotein and albumin, and at least a portion of said immature animal cells or at least a portion of the progeny of said immature cells is capable of differentiating into cells which express albumin. The cell population is cultured under conditions which result in expansion of the cells. Expansion of the cells may be achieved by culturing the cells in the presence of an extracellular matrix and liver stromal cells; and preferably in the presence of growth factors. Such cells may be used for liver transplantation, artificial livers, and for toxicology and pharmacology studies. Such cells may also be genetically engineered to express proteins or polypeptides of interest.
摘要翻译：包含动物细胞群并且含有未成熟动物细胞的组合物。未成熟动物细胞的特征在于甲胎蛋白的表达或甲胎蛋白和白蛋白的基本表达的缺失，并且所述未成熟动物细胞的至少一部分或所述未成熟细胞的后代的至少一部分能够分化进入表达白蛋白的细胞。细胞群在导致细胞扩增的条件下培养。可以通过在细胞外基质和肝脏基质细胞的存在下培养细胞来实现细胞的扩增; 并且优选在生长因子存在下。这些细胞可用于肝移植，人肝，以及毒理学和药理学研究。这样的细胞也可以被遗传工程化以表达感兴趣的蛋白质或多肽。

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