Immunologically privileged cells and uses thereof

METHOD FOR PRODUCING ADENOHYPOPHYSIS OR PRECURSOR TISSUE THEREOF

The present invention provides a method for deriving an adenohypophysis or precursor tissue thereof in vitro from human pluripotent stem cells. In this method: a human cell aggregate including hypothalamus neuroepithelial tissue and epidermal ectoderm is obtained by subjecting a human pluripotent stem cell aggregate to suspension culture in a culture medium including a bone morphogenetic factor signal transduction pathway activator and an Shh signal pathway active substance; and by subjecting the obtained human cell aggregate including hypothalamus neuroepithelial tissue and epidermal ectoderm to further suspension culture in a culture medium including a bone morphogenetic factor signal transduction pathway activator and an Shh signal pathway active substance, the formation of a hypophysis placode and/or Rathke's pouch in the epidermal ectoderm is induced to obtain a human cell aggregate including 1) hypothalamus neuroepithelial tissue and 2) a hypophysis placode and/or Rathke's pouch.

METHOD FOR PRODUCING CELL MASS INCLUDING PITUITARY TISSUE, AND CELL MASS THEREOF

The present invention aims to provide a method for efficiently producing a cell mass containing pituitary tissue from pluripotent stem cells. A method for producing a cell mass containing pituitary tissue, including the following steps (1) and (2): (1) a first step of suspension-culturing pluripotent stem cells to form a cell aggregate in the presence of a Wnt signal transduction pathway inhibiting substance, (2) a second step of suspension-culturing the aggregate obtained in the first step in the presence of a BMP signal transduction pathway activating substance and a Sonic hedgehog signal transduction pathway activating substance, thereby obtaining a cell mass comprising pituitary tissue.

DERIVATION OF SOMATOTROPHS FROM STEM CELLS AND USES THEREOF

The presently disclosed subject matter provides for in vitro methods of inducing differentiation of stem cells (e.g., human stem cells) into somatotrophs, and differentiated cells generated by such methods. The presently disclosed subject matter also provides for uses of such cells for treating growth hormone deficiency.

ISOLATION OF MULTIPOTENT HYPOPHYSARY CELLS AND IN VITRO DIFFERENTIATION THEREOF

The invention relates to the isolation of multipotent hypophysary cells and to the in vitro differentiation thereof. The method is based on the presence in multipotent hypophysary cells of a receptor of the external face of the membrane, GFRa2+, which is not present in other cells of the hypophysis. The isolation method uses antibodies directed against said receptor and techniques permitting the separation of the cells to which said antibodies have bound. Once isolated, the cells may be maintained in culture without being differentiated or they may be differentiated in order to produce cells that produce hypophysary hormones or cells having a neurone phenotype. Consequently, the cells isolated using said method can be used for the production of hypophysary hormones. In addition, antibodies directed against GFRa2+, coupled to signalling molecules, can be used to detect the presence of multipotent hypophysary cells in any sample.

IMMORTALIZED CELL LINES AND METHODS OF MAKING THE SAME

La présente invention concerne des procédés permettant d'immortaliser diverses cultures de cellules primaires, y compris des cellules hypophysaires, des neurones, des cellules gliales, des cellules d'épithélium cornéen et des cellules stellaires folliculaires. Les cellules primaires sont transfectées avec un vecteur contenant un oncogène constitutif ceci formant des cellules immortalisées non transformées. Les cellules primaires et/ou les cellules immortalisées subséquentes sont mises en culture dans un milieu défini contenant au moins un facteur environnemental qui commande la prolifération et/ou la différenciation des cellules.

METHODS FOR ISOLATING AND USING PITUITARY ADENOMA STEM CELLS AND PITUITARY ADENOMA CELLS

The present invention describes pituitary adenoma stem cells, pituitary carcinoma stem cells, a method of obtaining the stem cells, and a method of using the stem cells. Uses of the pituitary stem cells include but are not limited to producing pituitary hormones and identifying drugs to treat pituitary disease conditions or pituitary-related disease conditions.

METHOD FOR PRODUCING ADENOHYPOPHYSIS OR PRECURSOR TISSUE THEREOF

The invention provides a method for inducing adenohypophysis or precursor tissue thereof in vitro from human pluripotent stem cells. The method includes culturing an aggregate of human pluripotent stem cells in suspension in a medium containing a bone morphogenetic protein signal transduction pathway activating substance and a substance acting on the Shh signal pathway to obtain a human cell aggregate containing a hypothalamus neuroepithelial tissue and a surface ectoderm, and further culturing the obtained human cell aggregate containing the hypothalamus neuroepithelial tissue and the surface ectoderm in suspension in a medium containing a bone morphogenetic protein signal transduction pathway activating substance and a substance acting on the Shh signal pathway to induce formation of hypophysial placode and/or Rathke's pouch in the surface ectoderm, thus obtaining a human cell aggregate containing 1) hypothalamus neuroepithelial tissue, and 2) hypophysial placode and/or Rathke's pouch ...

METHOD FOR PRODUCING ADENOHYPOPHYSIS OR PRECURSOR TISSUE THEREOF

IMMUNOLOGICALLY PRIVILEGED CELLS AND USES THEREOF

The invention is directed to immunologically privileged cells, e.g., autologous, allogeneic, and xenogeneic intermediate lobe pituitary cells, for delivering polypeptides, e.g., insulin, to a subject, and to methods of using the same.

Generation of multipotent central nervous system stem cells

Methods for generating various cellular phenotypes from central nervous system stem cells are disclosed. Cellular differentiation into phenotypes of organs and tissues within and outside of the central nervous system is induced by co-culture with target cell types or by soluble trophic factors and elements of the extracellular matrix. Established pluripotent CNS stem cell lines are also disclosed.

METHOD FOR PRODUCING CELL MASS INCLUDING PITUITARY TISSUE, AND CELL MASS THEREOF

The present invention aims to provide a method for efficiently producing a cell mass containing pituitary tissue from pluripotent stem cells. A method for producing a cell mass containing pituitary tissue, including the following steps (1) and (2): (1) a first step of suspension-culturing pluripotent stem cells to form a cell aggregate in the presence of a Wnt signal transduction pathway inhibiting substance, (2) a second step of suspension-culturing the aggregate obtained in the first step in the presence of a BMP signal transduction pathway activating substance and a Sonic hedgehog signal transduction pathway activating substance, thereby obtaining a cell mass comprising pituitary tissue.

DIFFERENTIATION INDUCTION FROM HUMAN PLURIPOTENT STEM CELLS INTO HYPOTHALAMIC NEURONS

Provided is a method for efficiently inducing differentiation from human pluripotent stem cells into hypothalamic neurons. Also provided is a method for constructing a cellular structure that integrates pituitary tissue and hypothalamic tissue from human pluripotent stem cells. A cellular structure including hypothalamic tissue is obtained by a method including a step for suspension culturing human pluripotent stem cell spheroids in a medium including a low concentration of an osteogenic factor signaling pathway activator and a low concentration of a substance capable of acting on the Shh signaling pathway, and a step for also suspension culturing the cell spheroids obtained in that step in a medium including a low concentration of a substance capable of acting on the Shh signaling pathway. A cellular structure including hypothalamic tissue and pituitary tissue is also obtained by a method including a step for suspension culturing human pluripotent stem cell spheroids in a medium including ...

IMMORTALIZED CELL LINES AND METHODS OF MAKING THE SAME

The present invention relates to methods of immortalizing various primary cell cultures, including pituitary cells, neurons, beta islet cells, glial cells, corneal epithelial cells and follicular stellate cells. The primary cells are transfected with a vector containing an establishment oncogene, resulting in non-transformed immortalized cells. The primary cells and/or the subsequent immortalized cells are cultured in a defined media containing one or more environmental factor(s) that control the proliferation and/or differentiation of the cells.

GENERATION OF MULTIPOTENT CENTRAL NERVOUS SYSTEM STEM CELLS

Methods for generating various cellular phenotypes from central nervous system stem cells are disclosed. Cellular differentiation into phenotypes of organs and tissues within and outside of the central nervous system is induced by co- culture with target cell types or by soluble trophic factors and elements of the extracellular matrix. Established pluripotent CNS stem cell lines are also disclosed.

IMMORTALIZED CELL LINES AND METHODS OF MAKING THE SAME

La présente invention concerne des procédés permettant d'immortaliser diverses cultures de cellules primaires, y compris des cellules hypophysaires, des neurones, des cellules gliales, des cellules d'épithélium cornéen et des cellules stellaires folliculaires. Les cellules primaires sont transfectées avec un vecteur contenant un oncogène constitutif ceci formant des cellules immortalisées non transformées. Les cellules primaires et/ou les cellules immortalisées subséquentes sont mises en culture dans un milieu défini contenant au moins un facteur environnemental qui commande la prolifération et/ou la différenciation des cellules.

DERIVATION OF SOMATOTROPHS FROM STEM CELLS AND USES THEREOF

The presently disclosed subject matter provides for in vitro methods of inducing differentiation of stem cells (e.g., human stem cells) into somatotrophs, and differentiated cells generated by such methods. The presently disclosed subject matter also provides for uses of such cells for treating growth hormone deficiency. 1. An in vitro method for inducing differentiation of pituitary precursors , comprising contacting cells expressing at least one pituitary precursor marker with at least one dorsalizing agent and at least one ventralizing agent; and contacting the cells with at least one activator of Wingless (Wnt) signaling (Wnt activator) and at least one molecule that is capable of inducing growth hormone expression (GH inducer) to obtain a cell population of differentiated cells , wherein at least about 50% of differentiated cells are somatotrophs that are capable of producing growth hormone (GH-producing somatotrophs).2. The method of claim 1 , wherein the GH-producing somatotrophs comprise cells expressing a low level of GHRHR immunoreactivity (GHRHRcells) claim 1 , cells expressing a high level of GHRHR immunoreactivity (GHRHRcells) claim 1 , and a combination thereof.3. The method of claim 2 , comprising obtaining a cell population comprising at least about 50% GHRHRcells at least about 10 days claim 2 , or two weeks or three weeks claim 2 , or about 15 days from the initial contact of the cells expressing at least one pituitary precursor marker with the at least one dorsalizing agent.4. The method of claim 2 , comprising obtaining a cell population comprising at least about 50% GHRHRcells at least about 10 days claim 2 , or about three weeks claim 2 , or about 25 days claim 2 , or about 35 days from the initial contact of the cells expressing at least one pituitary precursor marker with the at least one dorsalizing agent.5. An in vitro method for inducing differentiation of stem cells claim 2 , comprising contacting stem cells with at least one BMP molecule ...

Generation of multipotent central nervous system stem cells

Method for producing cell mass including pituitary tissue, and cell mass thereof

The present invention addresses the problem of providing a technique for efficiently producing a cell mass including pituitary tissue from pluripotent stem cells. A method for producing a cell mass including pituitary tissue, the method including the following steps (1) and (2): (1) a first step for suspension-culturing pluripotent stem cells in the presence of a Wnt signaling pathway inhibitor, and forming an aggregate of cells; and (2) a second step for suspension-culturing the aggregate obtained in the first step in the presence of a BMP signaling pathway agonist and a sonic hedgehog signaling pathway agonist, and obtaining a cell mass including pituitary tissue.

Method for improving gonadotropin expression level after vitamin D is added to E cynoglossus semilaevis pituitary gland cells

本发明涉及一种向半滑舌鳎垂体细胞添加维生素E后提高促性腺激素表达水平的方法，步骤如下：(1)半滑舌鳎垂体细胞的分离与培养：首先配制L‑15细胞培养基，然后用所配培养基进行原代细胞的培养：(2)向半滑舌鳎垂体细胞中添加维生素E：配制维生素E溶液，向步骤(1)中所培养的细胞中添加维生素E，在添加后第四天进行样品收集；(3)向半滑舌鳎垂体细胞中添加维生素E后FSH和LH的定量PCR分析；取步骤(2)中的细胞沉淀进行RNA的提取及反转录，然后进行实时荧光定量PCR反应；(4)向半滑舌鳎垂体细胞中添加维生素E后FSH和LH的ELISA分析；以步骤(2)中收集的上清液为材料依次进行标准品的稀释与加样、加样、温育、配液、洗涤、加酶、温育、洗涤、显色、终止、测定。

고강도 전자기장과 음파 또는 초음파를 이용한 신경재생시스템

The present invention handles 100 to 2000mT high intensity electromagnetic field and from 0.5 to 2kHz, high-intensity electromagnetic field of from 80 to sound waves, or 100 to 2000mT of 200dB and 10 to 2000kHz, 0.1 to about 100 mW / cm 2 ultrasound in mesenchymal stem cells or adult stem cells To a method for differentiating mesenchymal stem cells or adult stem cells into neural cells. The present invention also relates to a composition comprising neurons differentiated by said method. The method and composition for neuron differentiation using high-intensity magnetic field and sonic wave or ultrasonic wave according to the present invention induces differentiation of adult stem cells into neuronal cells by using high-frequency electromagnetic field of high frequency and sonic wave or ultrasonic wave, Treatment alone can easily differentiate neurons or neural stem cells or improve recovery of damaged neurons.

Immortalized cell lines and methods of making the same

Method for producing adenohypophysis or precursor tissue thereof

The present invention provides a method for deriving an adenohypophysis or precursor tissue thereof in vitro from human pluripotent stem cells. In this method: a human cell aggregate including hypothalamus neuroepithelial tissue and epidermal ectoderm is obtained by subjecting a human pluripotent stem cell aggregate to suspension culture in a culture medium including a bone morphogenetic factor signal transduction pathway activator and an Shh signal pathway active substance; and by subjecting the obtained human cell aggregate including hypothalamus neuroepithelial tissue and epidermal ectoderm to further suspension culture in a culture medium including a bone morphogenetic factor signal transduction pathway activator and an Shh signal pathway active substance, the formation of a hypophysis placode and/or Rathke's pouch in the epidermal ectoderm is induced to obtain a human cell aggregate including 1) hypothalamus neuroepithelial tissue and 2) a hypophysis placode and/or Rathke's pouch.

SYSTEM FOR REGENERATING NERVE USING HIGH-INTENSITY ELECTROMAGNETIC FIELD, AND SOUND OR ULTRASOUND

The present invention relates to a method for differentiating mesenchymal stem cells or adult stem cells into nerve cells by treating the mesenchymal stem cells or adult stem cells with a high-intensity electromagnetic field (100-2000 mT) and sonic waves (0.5-2 kHz, 80-200 dB) or a high-intensity electromagnetic field (100-2000 mT) and ultrasonic waves (10-2000 kHz, 0.1-100 mW/cm^2). In addition, the present invention relates to a composition containing nerve cells differentiated by the method. According to the method and composition for differentiation into the nerve cells using the high-intensity magnetic field together with sonic or ultrasonic waves of the present invention, the differentiation of the adult stem cells into the nerve cells is induced by using a low-frequency high-intensity electromagnetic field and sonic or ultrasonic waves, thereby facilitating the differentiation into nerve cells or nerve stem cells or improving the restoration of damaged nerves through only the treatment ...

선하수체 또는 이의 전구 조직의 제조 방법

... 본 발명은 인간 다능성 줄기 세포에서의 선하수체 또는 이의 전구 조직의 시험관 내 유도 방법을 제공한다. 본 발명은 인간 다능성 줄기 세포 응집체를 골 형성 단백질 신호 전달 경로 활성화 물질 및 Shh 신호 경로 작용 물질을 함유하는 배지에서 부유 배양하여 시상하부 신경상피 조직 및 표면 외배엽을 함유하는 인간 세포 응집체를 수득하고, 수득된 시상하부 신경상피 조직 및 표면 외배엽을 함유하는 인간 세포 응집체를 골 형성 단백질 신호 전달 경로 활성화 물질 및 Shh 신호 경로 작용 물질을 함유하는 배지에서 추가로 부유 배양하여 표면 외배엽에서 뇌하수체 플라코드 및/또는 라트케낭의 형성을 유도함으로써, 1) 시상하부 신경상피 조직, 및 2) 뇌하수체 플라코드 및/또는 라트케낭을 함유하는 인간 세포 응집체를 수득하는 것을 포함한다.

METHODS FOR ISOLATING AND USING PITUITARY ADENOMA STEM CELLS AND PITUITARY ADENOMA CELLS

The present invention describes pituitary adenoma stem cells, pituitary carcinoma stem cells, a method of obtaining the stem cells, and a method of using the stem cells. Uses of the pituitary stem cells include but are not limited to producing pituitary hormones and identifying drugs to treat pituitary disease conditions or pituitary-related disease conditions.

Differentiation induction from human pluripotent stem cells into hypothalamic neurons

Provided is a method for efficiently inducing differentiation from human pluripotent stem cells into hypothalamic neurons. Also provided is a method for constructing a cellular structure that integrates pituitary tissue and hypothalamic tissue from human pluripotent stem cells. A cellular structure including hypothalamic tissue is obtained by a method including a step for suspension culturing human pluripotent stem cell spheroids in a medium including a low concentration of an osteogenic factor signaling pathway activator and a low concentration of a substance capable of acting on the Shh signaling pathway, and a step for also suspension culturing the cell spheroids obtained in that step in a medium including a low concentration of a substance capable of acting on the Shh signaling pathway. A cellular structure including hypothalamic tissue and pituitary tissue is also obtained by a method including a step for suspension culturing human pluripotent stem cell spheroids in a medium including ...

METHODS FOR ISOLATING AND USING PITUITARY ADENOMA STEM CELLS AND PITUITARY ADENOMA CELLS

The present invention describes pituitary adenoma stem cells, pituitary carcinoma stem cells, a method of obtaining the stem cells, and a method of using the stem cells. Uses of the pituitary stem cells include but are not limited to producing pituitary hormones and identifying drugs to treat pituitary disease conditions or pituitary-related disease conditions.

Method for producing adenohypophysis or precursor tissue thereof

The present invention provides a method for deriving an adenohypophysis or precursor tissue thereof in vitro from human pluripotent stem cells. In this method: a human cell aggregate including hypothalamus neuroepithelial tissue and epidermal ectoderm is obtained by subjecting a human pluripotent stem cell aggregate to suspension culture in a culture medium including a bone morphogenetic factor signal transduction pathway activator and an Shh signal pathway active substance; and by subjecting the obtained human cell aggregate including hypothalamus neuroepithelial tissue and epidermal ectoderm to further suspension culture in a culture medium including a bone morphogenetic factor signal transduction pathway activator and an Shh signal pathway active substance, the formation of a hypophysis placode and/or Rathke's pouch in the epidermal ectoderm is induced to obtain a human cell aggregate including 1) hypothalamus neuroepithelial tissue and 2) a hypophysis placode and/or Rathke's pouch.

Method for producing hypophysis precursor tissue

The present invention provides a method of obtaining aggregates containing a rostral hypothalamus tissue and a rostral head ectodermal tissue, a hypophysis precursor tissue and a hypophysis hormone producing cell, by using a serum-free medium (preferably substantially free of growth factor and insulins), forming homogeneous aggregates of stem cells from pluripotent stem cells such as ES cell and the like, which are plated at a high cell concentration, and subjecting the formed aggregates to floating-culture.

METHOD FOR PRODUCING CELL MASS INCLUDING PITUITARY TISSUE, AND CELL MASS THEREOF

METHOD FOR CULTURING STEM CELL

Immortalized cell lines and methods of making the same

The present invention relates to methods of immortalizing various primary cell cultures, including pituitary cells, neurons, beta islet cells, glial cells, corneal epithelial cells and follicular stellate cells. The primary cells are transfected with a vector containing an establishment oncogene, resulting in non-transformed immortalized cells. The primary cells and/or the subsequent immortalized cells are cultured in a defined media containing one or more environmental factor(s) that control the proliferation and/or differentiation of the cells.

인간 다능성줄기세포로부터 시상하부 뉴런으로의 분화 유도

... 인간 다능성줄기세포(pluripotent stem cell)로부터 시상하부 뉴런으로 효율적으로 분화 유도하는 방법을 제공한다. 또한, 인간 다능성줄기세포로부터 시상하부 조직과 하수체 조직이 일체화한 세포 구조체를 구축하는 방법을 제공한다. 인간 다능성줄기세포의 응집괴를, 저농도의 골 형성 인자 시그널 전달 경로 활성화 물질 및 저농도의 Shh 시그널 경로 작용 물질을 포함하는 배지 중에서 부유 배양하는 스텝과, 상기 스텝에서 얻어진 세포 응집괴를, 저농도의 Shh 시그널 경로 작용 물질을 포함하는 배지 중에서 더욱 부유 배양하는 스텝을 포함하는 방법에 의해, 시상하부 조직을 포함하는 세포 구조체를 얻는다. 또한, 인간 다능성줄기세포의 응집괴를, 골 형성 인자 시그널 전달 경로 활성화 물질 및 Shh 시그널 경로 작용 물질을 포함하는 배지 중에서 부유 배양하는 스텝과, 상기 스텝에서 형성된 세포 응집괴를, 골 형성 인자 시그널 전달 경로 활성화 물질 및 Shh 시그널 경로 작용 물질을 포함하는 배지 중에서 더욱 부유 배양하는 스텝과, 상기 스텝에서 얻어진 세포 응집괴를, 하수체 및 시상하부의 동시 유도에 적합한 배지 중에서 부유 배양하는 스텝을 포함하는 방법에 의해, 시상하부 조직과 하수체 조직을 포함하는 세포 구조체를 얻는다.

Process for the large scale production of human growth hormone by serial secondary suspension culture

A new system of serially culturing human anterior pituitary gland cells in a new nutrient medium to produce large amounts of human growth hormone. Since only human growth hormone can be used to treat growth deficiencies in man, there is a great demand for the hormone, which is in relative short supply since only 2 to 3 mgs. of the human growth hormone can be extracted from one human pituitary gland obtained at autopsy. Using the new system of culture and new nutrient media, the human pituitary cells can be grown in vitro to produce in approximately three weeks, more than 20 times the amount of extractable growth hormone than that which was originally present in the original tissue now used for extracting the 2 to 3 mgs. of the hormone.

METHOD FOR CULTURING STEM CELL

The present invention provides methods for producing an aggregate containing a rostal hypothalamic tissue and a rostal cranial ectodermal tissue, a pituitary progenitor tissue and a pituitary hormone-producing cell by forming a homogeneous aggregate of a stem cell from a pluripotent stem cell, e.g., an ES cell, seeded at a high cell concentration using a serum-free culture medium (preferably containing substantially no proliferation factor or no insulin component)and subjecting the formed aggregate to floating culture.

GENERATION OF MULTIPOTENT CENTRAL NERVOUS SYSTEM STEM CELLS

Methods for generating various cellular phenotypes from central nervous system stem cells are disclosed. Cellular differentiation into phenotypes of organs and tissues within and outside of the central nervous system is induced by co-culture with target cell types or by soluble trophic factors and elements of the extracellular matrix. Established pluripotent CNS stem cell lines are also disclosed.

METHODS OF INCREASING PRODUCTION OF PITUITARY HORMONES

METHOD FOR PRODUCING ADENOHYPOPHYSIS OR PRECURSOR TISSUE THEREOF

Process for the large scale production of human growth hormone by serial secondary suspension culture

선하수체 또는 이의 전구 조직의 제조 방법

... 본 발명은 인간 다능성 줄기 세포에서의 선하수체 또는 이의 전구 조직의 시험관 내 유도 방법을 제공한다. 본 발명은 인간 다능성 줄기 세포 응집체를 골 형성 단백질 신호 전달 경로 활성화 물질 및 Shh 신호 경로 작용 물질을 함유하는 배지에서 부유 배양하여 시상하부 신경상피 조직 및 표면 외배엽을 함유하는 인간 세포 응집체를 수득하고, 수득된 시상하부 신경상피 조직 및 표면 외배엽을 함유하는 인간 세포 응집체를 골 형성 단백질 신호 전달 경로 활성화 물질 및 Shh 신호 경로 작용 물질을 함유하는 배지에서 추가로 부유 배양하여 표면 외배엽에서 뇌하수체 플라코드 및/또는 라트케낭의 형성을 유도함으로써, 1) 시상하부 신경상피 조직, 및 2) 뇌하수체 플라코드 및/또는 라트케낭을 함유하는 인간 세포 응집체를 수득하는 것을 포함한다.

GENERATION OF MULTIPOTENT CENTRAL NERVOUS SYSTEM STEM CELLS

Methods for generating various cellular phenotypes from central nervous system stem cells are disclosed. Cellular differentiation into phenotypes of organs and tissues within and outside of the central nervous system is induced by co-culture with target cell types or by soluble trophic factors and elements of the extracellular matrix. Established pluripotent CNS stem cell lines are also disclosed.

Immunologically privileged cells and uses thereof

The invention is directed to immunologically privileged cells, e.g., autologous, allogeneic, and xenogeneic intermediate lobe pituitary cells, for delivering polypeptides, e.g., insulin, to a subject, and to methods of using the same.

Immortalized cell lines and methods of making the same

The present invention relates to methods of immortalizing various primary cell cultures, including pituitary cells, neurons, beta islet cells, glial cells, corneal epithelial cells and follicular stellate cells. The primary cells are transfected with a vector containing an establishment oncogene, resulting in non-transformed immortalized cells. The primary cells and/or the subsequent immortalized cells are cultured in a defined media containing one or more environmental factor(s) that control the proliferation and/or differentiation of the cells.

Immunologically privileged cells and uses thereof

The invention is directed to immunologically privileged cells, e.g., autologous, allogeneic, and xenogeneic intermediate lobe pituitary cells, for delivering polypeptides, e.g., insulin, to a subject, and to methods of using the same.

DIFFERENTIATION INDUCTION FROM HUMAN PLURIPOTENT STEM CELLS INTO HYPOTHALAMIC NEURONS

There is provided a method for efficient differentiation induction from human pluripotent stem cells into hypothalamic neurons. Also, provided is a method for constructing, from human pluripotent stem cells, a cellular structure in which hypothalamic tissue and pituitary tissue are integrated. A cellular structure including hypothalamic tissue is obtained by a method including the steps of: culturing an aggregate of human pluripotent stem cells in suspension in a medium containing a low concentration of a bone morphogenetic protein signal transduction pathway activating substance and a low concentration of a substance acting on the Shh signaling pathway; and further culturing the cell aggregate obtained in the step in suspension in a medium containing a low concentration of a substance acting on the Shh signaling pathway.

METHOD FOR CULTURING STEM CELL

The present invention provides a method of obtaining aggregates containing a rostral hypothalamus tissue and a rostral head ectodermal tissue, a hypophysis precursor tissue and a hypophysis hormone producing cell, by using a serum-free medium (preferably substantially free of growth factor and insulins), forming homogeneous aggregates of stem cells from pluripotent stem cells such as ES cell and the like, which are plated at a high cell concentration, and subjecting the formed aggregates to floating-culture.

DIFFERENTIATION INDUCTION FROM HUMAN PLURIPOTENT STEM CELLS INTO HYPOTHALAMIC NEURONS

DIFFERENTIATION INDUCTION FROM HUMAN PLURIPOTENT STEM CELLS INTO HYPOTHALAMIC NEURONS 5 There is provided a method for efficient differentiation induction from human pluripotent stem cells into hypothalamic neurons. Also, provided is a method for constructing, from human pluripotent stem cells, a cellular structure in which hypothalamic tissue and pituitary tissue are integrated. A cellular structure including hypothalamic tissue is obtained by a method including the steps of: culturing an aggregate of human pluripotent stem 10 cells in suspension in a medium containing a low concentration of a bone morphogenetic protein signal transduction pathway activating substance and a low concentration of a substance acting on the Shh signaling pathway; and further culturing the cell aggregate obtained in the step in suspension in a medium containing a low concentration of a substance acting on the Shh signaling pathway. Also, a cellular structure including hypothalamic tissue 15 and pituitary tissue ...

Methods of increasing production of pituitary hormones

A method for increasing the production of pituitary hormones, including particularly growth hormone, is described which comprises contacting a culture of pituitary cells with an effective amount of cholera enterotoxin.

Immortalized cell lines and methods of making the same

The present invention relates to methods of immortalizing various primary cell cultures, including pituitary cells, neurons, beta islet cells, glial cells, corneal epithelial cells and follicular stellate cells. The primary cells are transfected with a vector containing an establishment oncogene, resulting in non-transformed immortalized cells. The primary cells and/or the subsequent immortalized cells are cultured in a defined media containing one or more environmental factor(s) that control the proliferation and/or differentiation of the cells.

METHOD FOR CULTURING STEM CELL

The present invention provides a method of obtaining aggregates containing a rostral hypothalamus tissue and a rostral head ectodermal tissue, a hypophysis precursor tissue and a hypophysis hormone producing cell, by using a serum-free medium (preferably substantially free of growth factor and insulins), forming homogeneous aggregates of stem cells from pluripotent stem cells such as ES cell and the like, which are plated at a high cell concentration, and subjecting the formed aggregates to floating-culture.

Immunologically privileged cells and uses thereof

The invention is directed to immunologically privileged cells, e.g., autologous, allogeneic, and xenogeneic intermediate lobe pituitary cells, for delivering polypeptides, e.g., insulin, to a subject, and to methods of using the same.

Generation of multipotent central nervous system stem cells

Methods for generating various cellular phenotypes from central nervous system stem cells are disclosed. Cellular differentiation into phenotypes of organs and tissues within and outside of the central nervous system is induced by co-culture with target cell types or by soluble trophic factors and elements of the extracellular matrix. Established pluripotent CNS stem cell lines are also disclosed.

Immunologically privileged cells and uses thereof

The invention is directed to immunologically privileged cells, e.g., autologous, allogeneic, and xenogeneic intermediate lobe pituitary cells, for delivering polypeptides, e.g., insulin, to a subject, and to methods of using the same.

DIFFERENTIATION INDUCTION FROM HUMAN PLURIPOTENT STEM CELLS INTO HYPOTHALAMIC NEURONS

METHOD FOR CULTURING STEM CELL

The present invention provides a method of obtaining aggregates containing a rostral hypothalamus tissue and a rostral head ectodermal tissue, a hypophysis precursor tissue and a hypophysis hormone producing cell, by using a serum-free medium (preferably substantially free of growth factor and insulins), forming homogeneous aggregates of stem cells from pluripotent stem cells such as ES cell and the like, which are plated at a high cell concentration, and subjecting the formed aggregates to floating-culture.

Method for culturing stem cell

The present invention provides a method of obtaining aggregates containing a rostral hypothalamus tissue and a rostral head ectodermal tissue, a hypophysis precursor tissue and a hypophysis hormone producing cell, by using a serum-free medium (preferably substantially free of growth factor and insulins), forming homogeneous aggregates of stem cells from pluripotent stem cells such as ES cell and the like, which are plated at a high cell concentration, and subjecting the formed aggregates to floating-culture.

Method for producing cell mass including pituitary tissue, and cell mass thereof

The present invention addresses the problem of providing a technique for efficiently producing a cell mass including pituitary tissue from pluripotent stem cells. A method for producing a cell mass including pituitary tissue, the method including the following steps (1) and (2): (1) a first step for suspension-culturing pluripotent stem cells in the presence of a Wnt signaling pathway inhibitor, and forming an aggregate of cells; and (2) a second step for suspension-culturing the aggregate obtained in the first step in the presence of a BMP signaling pathway agonist and a sonic hedgehog signaling pathway agonist, and obtaining a cell mass including pituitary tissue.

Method for preparing adenohypophysis or precursor tissue thereof

本发明提供用于在体外从人多能干细胞衍生腺垂体或其前体组织的方法。在该方法中：通过在含有骨形态发生因子信号转导途径激活剂和Shh信号途径活性物质的培养基中对人多能干细胞聚集体进行悬浮培养而得到包含下丘脑神经上皮组织和表皮外胚层的人细胞聚集体；并且通过在含有骨形态发生因子信号转导途径激活剂和Shh信号途径活性物质的培养基中对所得到的包含下丘脑神经上皮组织和表皮外胚层的人细胞聚集体进行进一步悬浮培养，在表皮外胚层中诱导垂体基板和/或拉特克囊的形成，从而得到包含1)下丘脑神经上皮组织和2)垂体基板和/或拉特克囊的人细胞聚集体。

Method for producing glandular pituitary gland or precursor tissue thereof

本発明は、ヒト多能性幹細胞から、腺性下垂体又はその前駆組織をインビトロで誘導する方法を提供する。該方法においては、ヒト多能性幹細胞の凝集塊を、骨形成因子シグナル伝達経路活性化物質及びShhシグナル経路作用物質を含む培地中で浮遊培養することにより、視床下部神経上皮組織及び表皮外胚葉を含むヒト細胞凝集塊を得て、更に得られた視床下部神経上皮組織及び表皮外胚葉を含むヒト細胞凝集塊を、骨形成因子シグナル伝達経路活性化物質及びShhシグナル経路作用物質を含む培地中で更に浮遊培養することにより、表皮外胚葉中に下垂体プラコード及び／又はラトケ嚢の形成を誘導し、1）視床下部神経上皮組織、及び2）下垂体プラコード及び／又はラトケ嚢を含むヒト細胞凝集塊を得る。

Method for producing glandular pituitary gland or its precursor tissue

Construction method and application of human adenohormone type pituitary adenoma stem cell line

本发明公开了一种人源性腺激素型垂体腺瘤干细胞细胞系的构建方法及其应用，通过前期探索及优化培养条件，成功建立了来源于人的性腺激素型垂体腺瘤干细胞细胞系，该细胞系具有稳定的生物学特性、高体外成球能力、干细胞特性，可稳定多次传代，对其进行分化可产生分化后的肿瘤细胞，并且可持续自我更新，为针对性腺激素型垂体腺瘤的基础科研、药物的研发与筛选、耐药机理的研究、以及诊断试剂的研发提供了实验材料，为临床上进一步提高性腺激素型垂体腺瘤的治疗效果奠定了基础。

Method for producing adenohypophysis or precursor tissue thereof

Differentiation induction from human pluripotent stem cells into hypothalamic neurons

Provided is a method for efficiently inducing differentiation from human pluripotent stem cells into hypothalamic neurons. Also provided is a method for constructing a cellular structure that integrates pituitary tissue and hypothalamic tissue from human pluripotent stem cells. A cellular structure including hypothalamic tissue is obtained by a method including a step for suspension culturing human pluripotent stem cell spheroids in a medium including a low concentration of an osteogenic factor signaling pathway activator and a low concentration of a substance capable of acting on the Shh signaling pathway, and a step for also suspension culturing the cell spheroids obtained in that step in a medium including a low concentration of a substance capable of acting on the Shh signaling pathway. A cellular structure including hypothalamic tissue and pituitary tissue is also obtained by a method including a step for suspension culturing human pluripotent stem cell spheroids in a medium including an osteogenic factor signaling pathway activator and a substance capable of acting on the Shh signaling pathway, a step for also suspension culturing the cell spheroids formed in that step in a medium including an osteogenic factor signaling pathway activator and a substance capable of acting on the Shh signaling pathway, and a step for suspension culturing the cell spheroids obtained in that step in a medium suitable for simultaneous pituitary and hypothalamic induction .

Method for producing adenohypophysis or precursor tissue thereof

ヒト多能性幹細胞から視床下部ニューロンへの分化誘導

Method for producing adenohypophysis or precursor tissue thereof

The present invention provides a method for deriving an adenohypophysis or precursor tissue thereof in vitro from human pluripotent stem cells. In this method: a human cell aggregate including hypothalamus neuroepithelial tissue and epidermal ectoderm is obtained by subjecting a human pluripotent stem cell aggregate to suspension culture in a culture medium including a bone morphogenetic factor signal transduction pathway activator and an Shh signal pathway active substance; and by subjecting the obtained human cell aggregate including hypothalamus neuroepithelial tissue and epidermal ectoderm to further suspension culture in a culture medium including a bone morphogenetic factor signal transduction pathway activator and an Shh signal pathway active substance, the formation of a hypophysis placode and/or Rathke's pouch in the epidermal ectoderm is induced to obtain a human cell aggregate including 1) hypothalamus neuroepithelial tissue and 2) a hypophysis placode and/or Rathke's pouch.

一种长双歧杆菌及其培养方法和其在高产γ-氨基丁酸和5-羟色胺中的应用

本发明公开了一种长双歧杆菌及其培养方法和其在高产γ‑氨基丁酸和5‑羟色胺中的应用，涉及微生物领域，本发明提供了一种长双歧杆菌菌株MP‑569，可高产γ‑氨基丁酸和5‑羟色胺，它们是神经系统中的神经抑制剂，对神经元的兴奋程度有着潜在的影响，具有改善人的抑郁症状，增强记忆力，防止人脑损害的潜在能力；并且还能促进生长激素的分泌，在人体生长发育中起着关键性作用。

一种长双歧杆菌及其培养方法和其在高产γ-氨基丁酸和5-羟色胺中的应用

本发明公开了一种长双歧杆菌及其培养方法和其在高产γ‑氨基丁酸和5‑羟色胺中的应用，涉及微生物领域，本发明提供了一种长双歧杆菌菌株MP‑569，可高产γ‑氨基丁酸和5‑羟色胺，它们是神经系统中的神经抑制剂，对神经元的兴奋程度有着潜在的影响，具有改善人的抑郁症状，增强记忆力，防止人脑损害的潜在能力；并且还能促进生长激素的分泌，在人体生长发育中起着关键性作用。

幹細胞の培養方法

本発明は、無血清培地（好ましくは増殖因子及びインシュリン類を実質的に含まない）を用い、高細胞濃度で播種したＥＳ細胞等の多能性幹細胞から、均一な幹細胞の凝集体を形成し、形成された凝集体を浮遊培養することにより、吻側視床下部組織及び吻側頭部外胚葉組織を含む凝集体、下垂体前駆組織及び下垂体ホルモン産生細胞を得る方法を提供する。

뇌하수체 호르몬 생산 세포를 포함하는 세포 응집체 및 그 생산 방법

본 발명은 다능성 줄기 세포에서 뇌하수체 호르몬 생산 세포를 함유하는 세포 응집체를 효율적으로 생산하는 방법을 제공한다.

一种向半滑舌鳎垂体细胞添加维生素 e 后提高促性腺激素表达水平的方法

本发明涉及一种向半滑舌鳎垂体细胞添加维生素E后提高促性腺激素表达水平的方法，步骤如下：(1)半滑舌鳎垂体细胞的分离与培养：首先配制L‑15细胞培养基，然后用所配培养基进行原代细胞的培养：(2)向半滑舌鳎垂体细胞中添加维生素E：配制维生素E溶液，向步骤(1)中所培养的细胞中添加维生素E，在添加后第四天进行样品收集；(3)向半滑舌鳎垂体细胞中添加维生素E后FSH和LH的定量PCR分析；取步骤(2)中的细胞沉淀进行RNA的提取及反转录，然后进行实时荧光定量PCR反应；(4)向半滑舌鳎垂体细胞中添加维生素E后FSH和LH的ELISA分析；以步骤(2)中收集的上清液为材料依次进行标准品的稀释与加样、加样、温育、配液、洗涤、加酶、温育、洗涤、显色、终止、测定。

Cell aggregate including pituitary hormone-producing cells and method for producing same

The present invention provides a method for efficiently producing a cell aggregate including pituitary hormone-producing cells from pluripotent stem cells.

Differentiation induction from human pluripotent stem cells into hypothalamic neurons

Cell aggregate including pituitary hormone-producing cells and method for producing same

Differentiation induction from human pluripotent stem cells into hypothalamic neurons

Cell aggregate including pituitary hormone-producing cells and method for producing same

The present invention provides a method for efficiently producing a cell aggregate containing pituitary hormone-producing cells from pluripotent stem cells.

Method for separating pituitary hormone-producing cells and progenitor cells thereof

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一种将多能干细胞诱导为三维垂体类器官的培养方法

本发明提供了一种将多能干细胞诱导为三维垂体类器官的培养方法，包括：在一系列信号通路激活剂和抑制剂的作用下将多能干细胞在体外诱导为三维垂体基板类器官；以及将三维垂体基板类器官在Sonic hedgehog信号通路激活剂和AKT信号通路抑制剂的持续作用下逐步发育成熟，形成三维垂体类器官。本发明方法可以稳定且高效地在体外构建三维垂体类器官，为垂体类器官在垂体发病机制研究、再生医学治疗、毒药物筛选等领域中的应用提供坚实的模型基础。

一种将多能干细胞诱导为三维垂体基板类器官的培养方法

本发明提供了一种将多能干细胞诱导为三维垂体基板类器官的培养方法，包括：将多能干细胞在一系列信号通路激活剂和抑制剂的作用下逐步发育成三维垂体基板类器官；一系列信号通路激活剂和抑制剂包括BMP信号通路抑制剂、TGF‑β信号通路抑制剂、细胞凋亡抑制剂、Sonic hedgehog信号通路激活剂、BMP信号通路激活剂、FGF信号通路激活剂和AKT信号通路抑制剂。本发明方法可以稳定且高效地在体外构建三维垂体基板类器官，为垂体发育研究、垂体基板类器官广泛应用潜力的开发和后续垂体类器官分化方法的优化提供坚实的模型基础。

使用细胞表面标记物的垂体细胞和下丘脑细胞的分离方法

本发明提供分离垂体激素产生细胞和/或其前体细胞的方法，所述方法包括由包含腺垂体和/或其前体组织以及下丘脑神经上皮组织的细胞聚集块分离表达CD49c的细胞的步骤。

Method for separating pituitary cells and hypothalamic cells using cell surface marker

The present invention provides a method for separating pituitary hormone-producing cells and/or progenitor cells thereof, said method comprising a step for separating CD49c-expressing cells from a cell aggregate including adenohypophysis and/or a progenitor tissue thereof, and a hypothalamic neuroepithelial tissue.

一种将多能干细胞诱导为三维垂体类器官的培养方法

本发明提供了一种将多能干细胞诱导为三维垂体类器官的培养方法，包括：在一系列信号通路激活剂和抑制剂的作用下将多能干细胞在体外诱导为三维垂体基板类器官；以及将三维垂体基板类器官在Sonic hedgehog信号通路激活剂和AKT信号通路抑制剂的持续作用下逐步发育成熟，形成三维垂体类器官。本发明方法可以稳定且高效地在体外构建三维垂体类器官，为垂体类器官在垂体发病机制研究、再生医学治疗、毒药物筛选等领域中的应用提供坚实的模型基础。

一种将多能干细胞诱导为三维垂体基板类器官的培养方法

本发明提供了一种将多能干细胞诱导为三维垂体基板类器官的培养方法，包括：将多能干细胞在一系列信号通路激活剂和抑制剂的作用下逐步发育成三维垂体基板类器官；一系列信号通路激活剂和抑制剂包括BMP信号通路抑制剂、TGF‑β信号通路抑制剂、细胞凋亡抑制剂、Sonic hedgehog信号通路激活剂、BMP信号通路激活剂、FGF信号通路激活剂和AKT信号通路抑制剂。本发明方法可以稳定且高效地在体外构建三维垂体基板类器官，为垂体发育研究、垂体基板类器官广泛应用潜力的开发和后续垂体类器官分化方法的优化提供坚实的模型基础。

分离垂体激素产生细胞及其祖细胞的方法

本发明提供一种从来自多能干细胞的分化组织中有效地分离、纯化功能性垂体激素产生细胞和/或其祖细胞的方法。

制备腺垂体或其前体组织的方法

本发明提供用于在体外从人多能干细胞衍生腺垂体或其前体组织的方法。在该方法中：通过在含有骨形态发生因子信号转导途径激活剂和Shh信号途径活性物质的培养基中对人多能干细胞聚集体进行悬浮培养而得到包含下丘脑神经上皮组织和表皮外胚层的人细胞聚集体；并且通过在含有骨形态发生因子信号转导途径激活剂和Shh信号途径活性物质的培养基中对所得到的包含下丘脑神经上皮组织和表皮外胚层的人细胞聚集体进行进一步悬浮培养，在表皮外胚层中诱导垂体基板和/或拉特克囊的形成，从而得到包含1)下丘脑神经上皮组织和2)垂体基板和/或拉特克囊的人细胞聚集体。

一种长双歧杆菌及其培养方法和其在高产γ-氨基丁酸和5-羟色胺中的应用

本发明公开了一种长双歧杆菌及其培养方法和其在高产γ‑氨基丁酸和5‑羟色胺中的应用，涉及微生物领域，本发明提供了一种长双歧杆菌菌株MP‑569，可高产γ‑氨基丁酸和5‑羟色胺，它们是神经系统中的神经抑制剂，对神经元的兴奋程度有着潜在的影响，具有改善人的抑郁症状，增强记忆力，防止人脑损害的潜在能力；并且还能促进生长激素的分泌，在人体生长发育中起着关键性作用。

一种人源性腺激素型垂体腺瘤干细胞细胞系的构建方法及其应用

本发明公开了一种人源性腺激素型垂体腺瘤干细胞细胞系的构建方法及其应用，通过前期探索及优化培养条件，成功建立了来源于人的性腺激素型垂体腺瘤干细胞细胞系，该细胞系具有稳定的生物学特性、高体外成球能力、干细胞特性，可稳定多次传代，对其进行分化可产生分化后的肿瘤细胞，并且可持续自我更新，为针对性腺激素型垂体腺瘤的基础科研、药物的研发与筛选、耐药机理的研究、以及诊断试剂的研发提供了实验材料，为临床上进一步提高性腺激素型垂体腺瘤的治疗效果奠定了基础。

从干细胞中衍生促生长激素细胞及其用途

本发明公开的主题提供诱导干细胞(例如，人干细胞)分化为促生长激素细胞的体外方法，以及通过这些方法产生的分化细胞。本发明公开的主题还提供这些细胞用于治疗生长激素缺乏症的用途。

Method for producing adenohypophysis or precursor tissue thereof

The invention provides a method for inducing adenohypophysis or precursor tissue thereof in vitro from human pluripotent stem cells. The method includes culturing an aggregate of human pluripotent stem cells in suspension in a medium containing a bone morphogenetic protein signal transduction pathway activating substance and a substance acting on the Shh signal pathway to obtain a human cell aggregate containing a hypothalamus neuroepithelial tissue and a surface ectoderm, and further culturing the obtained human cell aggregate containing the hypothalamus neuroepithelial tissue and the surface ectoderm in suspension in a medium containing a bone morphogenetic protein signal transduction pathway activating substance and a substance acting on the Shh signal pathway to induce formation of hypophysial placode and/or Rathke's pouch in the surface ectoderm, thus obtaining a human cell aggregate containing 1) hypothalamus neuroepithelial tissue, and 2) hypophysial placode and/or Rathke's pouch.

包含垂体组织的细胞团块的制备方法及该细胞团块

本发明的课题在于提供由多潜能干细胞有效地制备包含垂体组织的细胞团块的手段。包含垂体组织的细胞团块的制备方法，包括下述步骤(1)及(2)：(1)第一步骤，将多潜能干细胞在Wnt信号转导途径抑制物质的存在下悬浮培养，使细胞的聚集体形成；(2)第二步骤，将第一步骤得到的聚集体在BMP信号转导途径作用物质及Sonic hedgehog信号转导途径作用物质的存在下悬浮培养，得到包含垂体组织的细胞团块。