266 results about "Differentiation induction" patented technology

There is provided a method of inducing differentiation of bone marrow stromal cells to neural cells or skeletal muscle cells by introduction of a Notch gene. Specifically, the invention provides a method of inducing differentiation of bone marrow stromal cells to neural cells or skeletal muscle cells in vitro, which method comprises introducing a Notch gene and / or a Notch signaling related gene into the cells, wherein the finally obtained differentiated cells are the result of cell division of the bone marrow stromal cells into which the Notch gene and / or Notch signaling related gene have been introduced. The invention also provides a method of inducing further differentiation of the differentiation-induced neural cells to dopaminergic neurons or acetylcholinergic neurons. The invention yet further provides a treatment method for neurodegenerative and skeletal muscle degenerative diseases which employs neural precursor cells, neural cells or skeletal muscle cells produced by the method of the invention.

There is provided a method of inducing differentiation of bone marrow stromal cells to neural cells or skeletal muscle cells by introduction of a Notch gene. Specifically, the invention provides a method of inducing differentiation of bone marrow stromal cells to neural cells or skeletal muscle cells in vitro, which method comprises introducing a Notch gene and / or a Notch signaling related gene into the cells, wherein the finally obtained differentiated cells are the result of cell division of the bone marrow stromal cells into which the Notch gene and / or Notch signaling related gene have been introduced. The invention also provides a method of inducing further differentiation of the differentiation-induced neural cells to dopaminergic neurons or acetylcholinergic neurons. The invention yet further provides a treatment method for neurodegenerative and skeletal muscle degenerative diseases which employs neural precursor cells, neural cells or skeletal muscle cells produced by the method of the invention.

The present invention provides a method of examining polycystic kidney disease or a complication of polycystic kidney disease using a gene(s) selected from the group consisting of NTNG1, POSTN, TNC, KAL1, BST1, ACAT2, INSIG1, SCD, HSD3B1, KRT7, USP40, SULT1E1, BMP6, CD274, CTGF, E2F7, EDN1, FAM43A, FRMD3, MMP10, MYEOV, NR2F1, NRCAM, PDCK1, PLXNA2, SLC30A3, SNAI1, SPOCD1, MMP1, TFPI2, HMGA2, KRTAP4-7, KRTAP4-8, KRTAP4-9, MYPN, RPPH1, and SIAE, and a method of screening for a therapeutic agent or a preventive agent therefore, and further vascular endothelial cells or vascular mural cells obtained via differentiation induction from iPS cells formed from a somatic cell of a subject suffered from polycystic kidney disease and having cerebral aneurysm as a complication.

The invention relates to the field of cell engineering and particularly relates to a method for separating human adipose-derived stem cells from human adipose tissues. According to the method, by adding a small amount of low-molecular-weight heparin calcium into normal saline, a mixed solution is obtained and used as a cleaning fluid to clean the human adipose tissues, residual impurities such as red blood cells in the human adipose tissues can be well removed and thus human adipose-derived stem cells with relatively higher purity can be separated, the content of the parenchyma cell is significantly reduced, the differentiation induction of the human adipose-derived stem cells is facilitated, the consumption amounts of collagenase I, trypsase and a serum-free culture medium can be reduced and the cost of the process is also decreased. In addition, by centrifuging and digesting the human adipose tissues for 1 hour at a speed of 150r / min, the human adipose-derived stem cells with significantly better activity can be obtained.

The invention discloses a tissue-engineered bone based on an entochondrostosis system and a construction method thereof. The construction method is characterized in that mesenchymal stem cells are cultivated on a porous bone scaffold material to construct a tissue-engineering complex, subjected to in vitro cartilage differentiation induction cultivation for two weeks and continuously conducted with fat cartilage differentiation induction cultivation for two weeks to obtain the tissue-engineered bone. According to the invention, an in vivo research result shows that the in vivo ectopic bone formation can be realized, bone defects can be successfully repaired, the difficulties that the uniform vascularization of large piece of tissue-engineered bone cannot be realized and enough nutrition can't be supplied are solved, and the construction method has a very good application prospect in the construction of the tissue-engineered bone and bone defect repairing.

The invention discloses a method for producing a red blood cell drug containing L-ASPase II (L-asparaginase) by in vitro induction. The method comprises the steps of: 1) using a lentiviral vector system to transform induced pluripotent stem (IPS) cells: I. constructing a pLenti6.3 / V5-GW / Em-GFP VerA-L-ASPase II plasmid; II. using 293T cells to perform packaging so as to generate high titer virus particles containing L-ASPase II; III. using the virus particles to transfect induced pluripotent stem cells; and 2) inducing the induced pluripotent stem cells in vitro to generate L-ASPase II-containing red blood cells: I. subjecting IPS cells to in vitro differentiation induction to form an erythroid body EB; and II. performing induced differentiation on EB to obtain mature red cells. The method provided in the invention performs genetic modification on IPS cells by the lentiviral vector system, and induces them in vitro to generate red blood cells containing L-ASPase II. The generated red blood cells can be used as a sustained release carrier of L-ASPase II, and also can maintain the original shapes and functions of red blood cells. The toxic and side effects of L-ASPase II are further reduced, so that the method lays a solid foundation for future clinical application.

In light of such conventional technical problems, it is an object of the present invention to provide a cell adsorption column which has an adsorbent with a surface area where there is less concern about denaturation and differentiation induction of cells caused by a contact of the adsorbent with the cells, and whose adsorbent-filled volume is small, and which has a compact property for improving operability and reducing a burden on a patient. The present invention provides a cell adsorption column filled with an adsorbent containing a fiber whose fiber diameter is 0.5 μm or more and 10 μm or less and having a surface area of 0.5 m2 or more and less than 10 m2, in which the adsorbent-filled volume is 100 mL or less.

The invention discloses a method for inducing human umbilical cord mesenchymal stem cells in vitro into liver cells and application thereof. The method comprises the following steps of: at the culture logarithmic growth phase of human umbilical cord mesenchymal stem cells, adding a liver cell induced differentiation culture medium and performing differentiated induction to obtain the liver cells formed by the differentiation of the human umbilical cord mesenchymal stem cells. The liver cell induced differentiation culture medium consists of the basic culture medium is an IMDM culture medium containing 10 to 70 ng / ml of liver cell growth factor, 3 to 30 ng / ml of fibroblast growth factor, 4.5 to 50 ng / ml of oncostatin, 2 to 40 ng / ml of alkali fibroblast growth factor, 20 ng / ml of epidermal growth factor, 1 mu M of dexamethasone and 50 mg / ml of ITS+premix. The method has the characteristics of one step method, simplicity, high repeatability, short duration time of differentiation process and good induced differentiation effect. The method can be applied to the research on turning mesenchymal stem cells into liver cells by induced differentiation.

The invention provides a method of efficiently producing corneal endothelial cells, particularly from corneal stroma or iPS cell-derived neural crest stem cells, a method of producing corneal endothelial cells stably in a large amount by inducing more efficient differentiation of stem cells into corneal endothelial cells, and a medicament containing corneal endothelial cells. The method of inducing differentiation of stem cells into corneal endothelial cells includes a step of culturing the stem cells in a differentiation induction medium containing a GSK3 inhibitor (preferably a GSK3β inhibitor) and retinoic acid, with the differentiation induction medium preferably further containing one or more of TGFb2, insulin, a ROCK inhibitor, and the like.

The invention discloses a human mesenchymal stem cell adipogenesis inducing and differentiating culture medium and a preparation method thereof. The human mesenchymal stem cell adipogenesis inducing and differentiating culture medium is produced by the following components of an alpha-MEM / HG-DMEM culture medium, 5-50% of percent by volume of fetal calf serum, 0.5-10% of percent by volume of glutamine, 100-400 [mu]M volume of indomethacin, insulin with the concentration of 0.1-20 [mu]g / ml, 10-200 [mu]M volume of 1-methyl-3-isobutyl xanthine, 10-200 nM volume of dexamethasone and 0.1-20 [mu]M volume of spermine. The preparation method includes the following steps that a culture dish is cleaned; the culture medium is provided, and the alpha-MEM / HG-DMEM culture medium is prepared in the culture dish according to a formula; materials are mixed; and mixed liquor is filtered. Multiple histologic origin human mesenchymal stem cells including human mesenchymal stem cells, umbilical cord mesenchymal stem cells and adipose tissue-derived stromal cells are induced to the directional differentiation of adipogenesis cells; differentiating and inducing time of the human mesenchymal stem cells isshortened, the preparation method is convenient, and the differentiating and the inducing of human mesenchymal stem cell adipogenesis can be achieved stably and efficiently.

The object of the present invention is to provide a method of preparing excellent cultured muscle cells having high metabolic capacity and insulin responsiveness, and further provide a method for the measurement of sensitive metabolic capacity using the cells. Moreover, its purpose is to provide a culture system / culture apparatus that can smoothly translocate such highly advanced cultured muscle cells intact to activity evaluation systems of a number of drugs. Moreover, the object of the present invention is to provide cultured muscle cells that are very suitable for measurement of the membrane-translocation activity of GLUT4 in an extraneous stimulus-dependent manner such as insulin, etc., and to provide a method for the measurement of the membrane-translocation activity of GLUT4 using the cells. The present invention is a method of preparing myotube cells, comprising a step (1) of culturing myoblast cells, a step (2) of differentiation-inducing the myotube cells into the myoblast cells in a culture medium with a high content of amino acids, and a step (3) of applying an electric pulse to the differentiation-induced myotube cells, and a method for the measurement of insulin-dependent sugar uptake using the myotube cells prepared by said method, and relates to the method for the measurement, comprising applying insulin stimulation by culturing the cells in a culture medium containing insulin, culturing the cells in the culture medium further supplemented with sugar, and measuring the sugar uptake. Furthermore, the present invention relates to a differentiation-type culture myotube cell constitutively expressing a recombinant GLUT4 having a labeled substance at its extra-cellular site, which is prepared by co-culturing wild-type myoblast cells and recombinant myoblast cells constitutively expressing said recombinant GLUT4, and a method for the measurement of membrane-translocation activity of the recombinant GLUT4 using the cells, and particularly a method for the measurement of insulin-dependent sugar uptake activity.

Provided is a differentiation-inducing culture medium additive for inducing bone differentiation of at least one type of cell selected from the group consisting of a stem cell, a dental pulp cell, a periodontal ligament cell, a placenta, an amnion, and a fibroblast under a serum-free condition, and a use of the differentiation-inducing culture medium additive. The differentiation-inducing culture medium additive of the present invention for inducing differentiation of a stem cell under a serum-free condition at least contains at least one growth factor selected from the group consisting of EGF, FGF, and PDGF; dexamethasone; and β-glycerophosphate. The differentiation-inducing culture medium additive of the present invention does not require ascorbic acid 2-phosphate and ITS, which are normally essential for bone differentiation. Further, bone differentiation can be promoted by adding phospholipid.

The present invention relates to a method for preparing corneal endothelial progenitor cells by adherent culturing a cell population isolated from corneal endothelial cell tissue at a low density by using a serum-free medium. The present invention also relates to a method for preparing corneal endothelial cells by differentiation-inducing the corneal endothelial progenitor cells obtained by the aforementioned method. According to the present invention, corneal endothelial progenitor cells can be selectively grown from a corneal tissue-derived cell population, and corneal endothelial cells obtained by inducing the corneal endothelial progenitor cells can be applied to treatment of corneal endothelial diseases. As a result, problems of corneal transplantation such as shortage of donors and occurrence of rejection can be solved.

The invention provides a method for tissue culture propagation of Acer fabri Hance by axillary buds, which comprises the following steps: disinfecting explants, inoculating the disinfected explants onto an induction medium for differentiation induction of buds, and normally culturing for 25 to 35 days under the conditions of a culture temperature between 20 and 26 DEG C, illumination time between 10 to 16 hours per day and an illumination intensity between 1,600 and 20,001x; cutting obtained tender shoots of Acer fabri Hance, and inoculating the cut tender shoots onto a subculture medium to perform subculture for 25 to 35 days; inoculating the sterile seedlings of the Acer fabri Hance into a rooting culture medium to perform rooting culture for 10 to 15 days under the conditions of a culture temperature between 20 and 26 DEG C, illumination time between 10 and 16 hours per day and illumination intensity between 1,600 and 20,001x; and transferring the obtained rotted sterile seedlings to a seedling bed to perform domestication culture for 30 to 60 days. The method can effectively solve the quick promotion problem of Acer fabri Hance.

The invention relates to Chinese toon tissue culture and rapid propagation technology. The technology comprises the following steps: clipping an annual semi-lignified branch with plump axillary buds from a growing plant, clipping the branch into a stem section containing 1 to 2 axillary buds after sterilization, and rapidly inoculating the stem section to a differentiation induction culture medium for differentiation culture; placing the plant obtained through the differentiation culture into a propagation culture medium for propagation and rapid propagation; and when a certain quantity is reached through the propagation, putting the plant into a rootage culture medium for sound seedling rootage culture, and hardening the seedling after the sound seedling takes root. The method carries out the asexual tissue rapid propagation and disinfection through the stem apex and stem section of Chinese toon. The propagated filial generation maintains the good seed nature of the original variety, ensures the seed purity and has no diseases and insects. The technology overcomes the defect of low propagation speed lying in the conventional seed and cut root propagation, enables the artificial control on the culture and growth conditions, is not limited by natural conditions, and has a small amount of drawn material. The technology can realize the industrialized seedling culture and high-efficiency production.

A directional differentiation induction method of human embryonic stem cells to obtain corneal endothelial cells is provided. The method includes inducing the human embryonic stem cells to differentiate into human neural crest stem cells, inducing the human neural crest stem cells to differentiate into the human corneal endothelial cells, and other steps. The method adopts a primary human corneal stroma cell culture supernatant fluid, a human corneal endothelial cell culture supernatant fluid and a human lens cell culture supernatant fluid as a conditioned medium. Through adding retinoic acid and a plurality of recombinant proteins into the medium, and combining three-dimensional and two-dimensional culture methods, a corneal endothelial cell development process is simulated to induce the human embryonic stem cells to directionally differentiate into the human corneal endothelial cells, the corneal endothelial cells morphological structures of which are similar to those of normal human corneal endothelial cells can be obtained, and in-vitro subculture results show that proliferation of the obtained human corneal endothelial cells is consistent with proliferation of the normal human corneal endothelial cells.

The invention relates to a method for in-vitro directional differentiation inducing of a mesenchymal stem cell towards melanocyte, particularly relates to a method for in-vitro directional differentiation inducing of the mesenchymal stem cell with an umbilical cord source towards the melanocyte, and belongs to the technical field of biology. The mesenchymal stem cell with the umbilical cord source is cultivated by adopting an explants method, and then the mesenchymal stem cell with the umbilical cord source is identified, so as to determine the characteristics of the mesenchymal stem cell; a culture medium is induced to carry out in-vitro inducing differentiation to form the melanocyte by using improved melanine; and finally the cultivated melanocyte is identified. An ES cell source and a marrow or adipose tissue-derived stromal cell source are limited or not easy to obtain while the stem cell can be easily obtained from the umbilical cord, the immunogenicity is very low, and an individual difference is not easily generated, so that the method is suitable for large-scale clinical application, and the induced melanocyte can be applied to epichrosis leucasmus therapy.

Provided are: a method for producing an immortalized human erythroid progenitor cell line, enabling efficient and stable production of enucleated red blood cells; and a method for producing human enucleated red blood cells from a human erythroid progenitor cell line obtained by the aforementioned production method. An expression cassette capable of inducing expression of HPV-E6 / E7 genes in the presence of DOX was introduced into the genomic DNA of blood stem cells. Then, the blood stem cells were cultured in the presence of DOX and a blood growth factor. Thereby, immortalized cell lines of human erythroid progenitor cells were established. Further, it was revealed that culturing the cell lines under a condition where the expression of the HPV-E6 / E7 genes was not induced enabled differentiation induction into enucleated red blood cells at a high ratio.

The invention discloses a method for inducing and differentiating a human embryonic stem cell into nerve cells. The method comprises the following steps of: separating and purifying a human embryonic stem cell to obtain human embryonic stem cell monomers; and inoculating the human embryonic stem cell monomers according to the cell density of (1-5)*104 / cm<2>, and performing single-layer adherence induced culturing, wherein in the induced culturing, nerve cells are obtained by pre-inducing the human embryonic stem cell with a sodium butyrate and an inducting culture medium I and performing differentiated induced culturing with an inducing culture medium. According to the method, the defects of high nerve cell heterogeneous degree, low differentiating efficiency, inconvenience for observing and operating in real time, need of a subsequent sorting process and the like existing in the conventional differentiating method can be overcome; the method has the advantages of easiness and convenience for operating, short inducting period (10-12) days, high nerve cell uniformity (the ratio is over 85 percent), and the like; and according to electrophysiologic monitoring, the nerve cells have convulsion discharging functions.

The invention discloses a propagation method for keeping springness of an inbred line of a spring common head cabbage breed. The method comprises the following steps: selecting external-layer green leaves of mature leaf heads of the spring common head cabbage inbred line in spring, inoculating the leaves on an adventitious bud differentiation induction culture medium according to a certain size for culture, and inoculating adventitious buds to an adventitious root differentiation induction culture medium for culture when the adventitious buds grow to 1.5-2.5cm high to obtain complete regenerated seedlings; then transplanting the regenerated seedlings to culture soil for culture, and permanently planting the seedlings in a field (open field) when the seedlings grow to 5-7 true leaves and are adaptively hardened for 7 days, and by which the survival rate of the seedlings reaches over 95%. The method helps improve the breeding effect of the spring common head cabbage, ensure the spring common head cabbage inbred line to complete a heading stage in spring and agronomic character selection of plants of the spring common head cabbage, maintain the characters (springness) of the selected plants unchanged, ensure the inbred line to complete a life cycle, greatly improve the number of the plants with the same genotype of the inbred line, and proliferate the number of the plants over 50 times.

The invention belongs to the field of cell extraction and differentiation authentication, and discloses a method for extracting myelomonocyte and differentiating to osteoclast. The method comprises the following steps: (A) performing sterile myelomonocyte separation, culturing in a complete medium of 20-100ng / mLM-CSF, replacing the complete medium every other day, and observing the morphological characteristics of monocyte; (B) when the density of monocyte is 80-90%, taking out a part of the cells for surface antigen authentication; (C) drawing a growth curve of other cells and performing differentiation induction for osteoclast on other cells. The method for extracting myelomonocyte and differentiating to osteoclast is simple, convenient and reliable, low in requested condition requirements, low in cost and short in time, the extracted monocyte is stable in property, and C57BL / 6 mice myelomonocyte which is reliable and stable in resource can be provided for medical basis, clinical research, tissue engineering study, and the like.

The invention provides a method of efficiently producing corneal endothelial cells, particularly from corneal stroma or iPS cell-derived neural crest stem cells, a method of producing corneal endothelial cells stably in a large amount by inducing more efficient differentiation of stem cells into corneal endothelial cells, and a medicament containing corneal endothelial cells. The method of inducing differentiation of stem cells into corneal endothelial cells includes a step of culturing the stem cells in a differentiation induction medium containing a GSK3 inhibitor (preferably a GSK3β inhibitor) and retinoic acid, with the differentiation induction medium preferably further containing one or more of TGFb2, insulin, a ROCK inhibitor, and the like.

The invention provides a culture and reproduction method of Lagerstroemia fauriei Keohne tissues, which comprises the following steps: firstly sterilizing exophyte of stalk sections of Lagerstroemia fauriei Keohne, culturing in an inducing culture medium to carry out bud differentiation induction, culturing in the dark at controlled culture temperature, normally culturing under illumination, cutting off young tips of Lagerstroemia fauriei Keohne, subculturing in a successive transfer culture medium, and finally carrying out the rooting culture on the Lagerstroemia fauriei Keohne sterile seedlings in a rooting culture medium, thereby obtaining the Lagerstroemia fauriei Keohne sterile seedlings. The method can effectively solve the problem of resource shortage of Lagerstroemia fauriei Keohne.

Differentiation of a large number of embryonic stem cells or artificial pluripotent stem cells such as induced pluripotent stem cells can be readily induced by the following method. A substrate surface having a cell non-adhesive region on the periphery of a cell adhesive region is used, and(1) the cells are allowed to adhere to the cell adhesive region of the substrate surface in a differentiation-inducing medium;(2) the cells are subsequently allowed to grow on the cell adhesive surface to reach confluence;(3) the culture is further continued to three-dimensionally stratify the cells from the boundary with the cell non-adhesive region surrounding the cell adhesive region; and(4) a plurality of the stratified portions formed in the cell adhesive region is finally bound to one another to produce a cell agglomerate spreading over the entire cell adhesive region with a certain thickness.

Provided is a method of inducing the differentiation of a stem cell into nerve progenitor cells, comprising the step (1) of forming a homogenous aggregate of stem cells in a serum-free medium (1) and the step (2) of suspension-culturing the homogenous aggregate of stem cells in the presence of a basement membrane reference standard.

The invention discloses a propagation method for keeping a cabbage RGMS male sterile line, which comprises the following steps: selecting a scape leaf blade of the cabbage male sterile line, inoculating the scape leaf blade on an adventitious bud differentiation induction culture medium for cultivation, and when adventitious buds grow to a height of between 2.0 and 4.0 centimeters, transplanting the adventitious buds to an adventitious root differentiation induction culture medium for cultivation to obtain a complete regrowth seedling; transplanting the regrowth seedling to cultivating soil for cultivation; when the seedling grows to have 6 to 8 true leaves, hardening off the seedling adaptively for 3 to 5 days and then planting the seedling permanently in a field (an open ground), wherein the survival rate of the seedling is over 95 percent. By adopting the method, the breeding effect of the cabbage can be improved, and the normal breeding of the male sterility of the cabbage is ensured and enlarged. The method overcomes the complexity of the normal breeding of the male sterility of the cabbage, not only keeps the sterile line but also greatly improves the propagation coefficient, and provides guarantees for mass seed production.

The present invention provides a method for subculturing primate embryonic stem cells, and a method for inducing differentiation of the same cell into a vascular endothelial cell and a blood cell. The present invention provides a method comprising culturing primate embryonic stem cells in a medium containing a protein component without using feeder cells and cytokines in a container coated with an extracellular matrix, detaching colonies of the resulting embryonic stem cells in the presence of a cytodetachment agent, and plating the colonies in the similar medium, and a method comprising culturing primate embryonic stem cells in a serum-containing or not containing medium in the presence of cytokine, adhesion-culturing the resulting embryoid body or embroyid body-analogous cellular aggregate in the presence of a cytokine to obtain specific precursor cells, and separating non-adherent cells and adherent cells from the specific precursor cells to obtain blood cells and vascular endothelial precursor cells.

The present invention provides a method for inducing differentiation of cardiomyocytes efficiently and selectively from stem cells.A method for inducing differentiation of cardiomyocytes from pluripotent stem cells, which comprises: (i) culturing the pluripotent stem cells in a culture medium containing no substance that promotes activation of the canonical Wnt signaling pathway during the time period between initiation of differentiation induction and 24 hours before the period of elevated canonical Wnt gene expression; and then (ii) culturing the pluripotent stem cells in a culture medium containing a substance that promotes activation of the canonical Wnt signaling pathway during a time period of 24 to 96 hours, starting from 24 to 0 hours before the period of elevated canonical Wnt gene expression.

Disclosed are: a gene transduction method for use in the induction of the differentiation of stem cells such as ES cells or iPS cells into hepatocytes effectively; stem cells into each of which a gene useful for the induction of the differentiation into hepatocytes is introduced; and hepatocytes produced from stem cells each having the gene introduced therein. A specific gene can be introduced into stem cells such as ES cells or iPS cells using an adenovirus vector. The effective induction of the differentiation into hepatocytes can be achieved by introducing the gene. Specifically, the effective induction of the differentiation of stem cells such as ES cells or iPS cells into hepatocytes can be achieved by introducing at least one gene selected from HEX gene, HNF4A gene and SOX17 gene into the stem cells.