19 results about "Mesoderm Cell" patented technology

The invention provides a cryopreservation and resuscitation method for induced differentiation of pluripotent stem cells to metaphase cells and application, and the cryopreservation method comprises the following steps: (1) the initial cells are pluripotent stem cells, and are induced and differentiated to the metaphase; (2) recovering the cells by using a mild digestive enzyme, counting, and resuspending the cells in a cryopreservation solution; and (3) carrying out programmed cooling to-80 DEG C, and then transferring into liquid nitrogen for preservation for later use. According to the cryopreservation method and the cryopreservation liquid provided by the invention, the cryopreservation survival rate of the differentiated metaphase cells is obviously improved, apoptosis or irreversible stress injury induced by a traditional cryopreservation liquid is avoided, and the problems of large cell injury and low survival rate caused by using a general cryopreservation liquid in the prior art are obviously improved. The resuscitated cells keep good differentiation potential, can smoothly form a mature renal unit structure, and solves the problem of subsequent differentiation failure in the prior art.

The application provides a freezing and recovery method and application of pluripotent stem cell induced differentiation middle stage cells, and the freezing method comprises the following steps: (1) the starting cells are pluripotent stem cells, which are induced to the middle stage; (2) the cells are recovered and counted by using a mild digestion enzyme, and the cells are resuspended in a freezing solution; (3) after programmed cooling to-80 DEG C, the cells are transferred into liquid nitrogen for storage, and are ready for use. The freezing method and the freezing solution provided by the application significantly improve the freezing survival rate of the differentiation middle stage cells, avoid the apoptosis or irreversible stress damage induced by the traditional freezing solution, and obviously improve the problems of cell damage and low survival rate caused by the general freezing solution in the prior art. The differentiation potential of the recovered cells is good, and the cells can successfully form mature kidney unit structures, so that the problem of subsequent differentiation failure in the prior art is solved.

Disclosed herein are improved in vitro methods of making splanchnic mesoderm cell types and subtypes thereof from pluripotent cells. These methods can be used to produced improved foregut- and hindgut-derived organoids containing enriched mesenchyme, which enhances organoid viability, growth, and maturation, both in in vitro culture and in vivo transplantation.

This invention discloses a method for co-culturing and inducing stem cell differentiation into hematopoietic progenitor cells. The method includes the following steps: culturing pluripotent stem cells to obtain embryoid bodies; culturing the embryoid bodies for mesodermal differentiation to obtain mesodermal cells; culturing the mesodermal cells for hematopoietic endothelial differentiation to obtain hematopoietic endothelial cells; and co-culturing the hematopoietic endothelial cells with mesenchymal stem cells to obtain hematopoietic progenitor cells. Through optimization of the culture system, the differentiation efficiency and the number of hematopoietic progenitor cells obtained are significantly improved; by co-culturing mesenchymal stem cells with hematopoietic endothelial cells, the number of hematopoietic progenitor cells is further increased while maintaining high differentiation efficiency and uniform and stable expression of various markers.

The invention discloses a vascular endothelial cell as well as a preparation method and application thereof. The preparation method comprises the following steps: inoculating induced pluripotent stem cells into a culture medium containing an ROCK signal channel inhibitor for culturing, and inducing the cultured induced pluripotent stem cells by using a culture medium containing a Wnt signal channel activator and a BMP signal channel activator, so that the induced pluripotent stem cells are differentiated into mesoderm cells; inducing the mesoderm cells by using a culture medium containing a vascular endothelial growth factor and a cAMP signaling pathway agonist, so that the mesoderm cells are differentiated into endothelial coarse cells; and performing positive sorting of the endothelial cell marker CD144 on the endothelial coarse cells to obtain purified vascular endothelial cells. The induction path of the method is clear and efficient, the period is short, the vascular endothelial cell population with high purity, complete vascular barrier function and mature phenotype can be obtained, and the cell population can be applied to construction of a vascular network in a gel matrix.

The present disclosure relates to a method for induced differentiation of stem cells into hematopoietic progenitor cells. The method comprises the following steps: culturing pluripotent stem cells to obtain an embryoid body; performing mesoderm differentiation culture on the embryoid body to obtain mesoderm cells; performing hemogenic endothelium differentiation culture on the mesoderm cells to obtain hemogenic endothelial cells; and performing hematopoietic progenitor cell differentiation culture on the hemogenic endothelial cells to obtain hematopoietic progenitor cells. According to the present disclosure, the hematopoietic progenitor cells can be rapidly and efficiently prepared, and the prepared hematopoietic progenitor cells have an ability to stably differentiate into a plurality of different blood cells (including simultaneously having erythroid, myeloid, and lymphoid cells); by optimizing a culture system, the differentiation efficiency is significantly improved and the number of hematopoietic progenitor cells obtained is significantly increased.

The invention discloses a kit and a method for constructing a human bone organoid from pluripotent stem cells, and relates to the technical field of organoid culture. The kit for constructing the human bone organoid by the pluripotent stem cells comprises a culture medium A, a culture medium B, a culture medium C, a culture medium D and a culture medium E. The culture medium A induces the pluripotent stem cells to form original stripes, the culture medium B induces the original stripes to form paraaxial mesoderm, the culture medium C induces the paraaxial mesoderm to form body wall mesoderm, the culture medium D induces body wall mesoderm cells to differentiate to bone cells, and the culture medium E induces bone cells to form cartilage organs. The kit provided by the invention can induce the pluripotent stem cells into bone organs, and has important significance in research and treatment of pathogenesis of bone related diseases and related research of bone repair and regeneration.

Disclosed herein are improved in vitro methods of making splanchnic mesoderm cell types and subtypes thereof from pluripotent cells. These methods can be used to produced improved foregut- and hindgut-derived organoids containing enriched mesenchyme, which enhances organoid viability, growth, and maturation, both in in vitro culture and in vivo transplantation.

The invention discloses a method for obtaining, activating and amplifying natural killer cells through iPSC differentiation induction, and relates to the technical field of stem cell biology, the method for obtaining, activating and amplifying natural killer cells through iPSC differentiation induction comprises the following steps: S1, preparing and inoculating iPSC cells; s2, differentiating the iPSC to a mesoderm; s3, the mesoderm cells are differentiated to CD34 + hematopoietic endothelial cells; s4, differentiating the CD43 < + > / CD45 < + > hematopoietic stem cells into the iNK cells; s5, maturation of the iNK cells; s6, activating and amplifying the iNK cells; in the step S1, cells are resuspended by using a fresh E8 complete culture medium and are inoculated into a porous culture plate; a plurality of induction culture media are used in the culture process; by adopting a porous culture plate and using an E8 complete culture medium to resuspend cells, efficient inoculation and primary culture of iPSC are realized, and the survival rate and amplification efficiency of cells are improved; the differentiation process of the iPSC towards the mesoderm, the hematopoietic endothelial cells and the CD34 + hematopoietic stem cells is optimized by using various induction culture media, and the differentiation efficiency is improved.

The invention relates to a method for producing a mesenchymal stem cell (MSC), the method comprising culturing a primitive mesoderm cell in a mesenchymal colony forming medium (M-CFM) comprising LiCl and FGF2, but excluding PDGF, under normoxic conditions for sufficient time for a mesenchymal colony to form, and culturing the mesenchymal colony adherently to produce the MSC, wherein the MSC has superior T-cell immunosuppressive properties relative to an MSC not produced in said M-CFM. The invention also relates to an MSC produced by the method, a population of MSCs produced by the method, a therapeutic composition comprising the MSC produced by the method, an M-CFM and an M-CFM in concentrated form, and method and uses of the MSC or population in treating a disease.

The invention relates to a method for producing mesenchymal stem cells with bone differentiation tendency by using extraembryonic mesoderm cells. Specifically, the invention develops a method for inducing and differentiating human pluripotent stem cells into embryonic ectomesoderm cells in vitro, and optimizes induction culture conditions and a purification method. The obtained extraembryonic mesoderm cells are further subjected to differentiation culture, so that the mesenchymal stem cells can be stably obtained, and the obtained mesenchymal stem cells have a bone differentiation tendency and lack the capability of differentiating to adipocytes. The mesenchymal cells obtained through in-vitro culture have application prospects in the field of cell therapy of bone related diseases.

The present invention provides a method for producing mesodermal lineage cells from an embryoid body, the method comprising: (1) a step for inducing an embryoid body into a differentiated embryoid body in which differentiation into a mesodermal cell lineage is promoted; and (2) a step for determining the solidity of the induced differentiated embryoid body.

This invention relates to a method for preparing cardiac organoids derived from human pluripotent stem cells, comprising: Step A: culturing human pluripotent stem cells (hPSCs) to prepare embryonic bodies with a diameter of 100 μm to 130 μm; Step B: culturing the embryonic bodies prepared in Step A to a diameter of 200 μm to 250 μm; Step C: differentiating the embryonic bodies derived from human pluripotent stem cells with a diameter of 200 μm to 250 μm prepared in Step B into constructs containing mesodermal cells and endoderm cells; Step D: differentiating the constructs containing mesodermal cells and endoderm cells differentiated in Step C into constructs containing cardiomyocytes, fibroblasts, and vascular endothelial cells. Self-organized cardiac organoids (cells); and, step E: culturing and maturing the self-organized cardiac organoids differentiated through step D, wherein the cardiac organoids that have completed the maturation process through step E have cardiomyocytes, fibroblasts, and endothelial cells forming a self-organized structure with a cell ratio of 55 to 65:15 to 30:15.

Provided is a method for producing an osteoblast construct from an iPS cell, the method comprising the steps of: (1) inducing formation of embryoid bodies by subjecting undifferentiated iPS cells to non-adherent culture; (2) inducing differentiation of the iPS cells into mesoderm cells by subjecting the embryoid bodies of the iPS cells obtained in step (1) to non-adherent culture; and (3) inducing differentiation into osteoblasts by subjecting the mesoderm cells of the iPS cells obtained in step (2) to non-adherent culture, wherein both steps (1) and (2) are performed using a culture vessel comprising a bottom surface and a ring-shaped side wall arranged vertically on the bottom surface, the bottom surface having a plurality of recesses arranged independently of one another.

The invention relates to a vein vessel organoid based on human pluripotent stem cell induction and a construction method thereof. The construction method comprises the following steps: S1, culturing human pluripotent stem cells in a pore plate by using an mTESR1 culture medium; s2, preparing the human pluripotent stem cells into a single-cell suspension, and forming uniform cell aggregates in an ultra-low adsorption culture plate; step S3, inducing mesoderm cell spheres, collecting cell aggregates, discarding supernatant, adding into a first culture medium which contains a Glutamax additive, L-ascorbic acid, CHIR99021 and BMP4, culturing for 2-3 days, and inducing to form the mesoderm cell spheres; s4, vein vessel organoid differentiation: adding a second culture medium for culture, promoting mesoderm cells to continue to differentiate to the vessel pedigree, and inducing to form a vein vessel network. According to the invention, the directional differentiation of the human pluripotent stem cells to the vein vessel organoid is realized, and the capillary vessel organoid with vein characteristics is successfully constructed.

The invention discloses a method for inducing directional differentiation of human induced pluripotent stem cells into mesenchymal stem cells in stages and application of the method, and develops a mesenchymal induction system which has definite components, is reasonably added with induced differentiation factors and has a reasonable concentration and is subjected to induced differentiation in stages. The induced pluripotent stem cells are regulated to efficiently and directionally induce and differentiate into the mesoderm cells, and then the mesoderm cells are directionally induced to induce and convert into the MSCs, so that the differentiation period of inducing and differentiating the human induced pluripotent stem cells into the mesenchymal stem cells can be shortened to 2-3 weeks, the differentiation rate is high, and the batch consistency can be ensured.

This invention provides a method for differentiating SAN-like cells from pluripotent stem cells in vitro. The method for differentiating SAN-like cells from pluripotent stem cells in vitro includes S1, providing a population of pluripotent stem cells; S2, performing a first contact culture on the pluripotent stem cell population to obtain mesodermal cells; S3, performing a second contact culture on the mesodermal cells to obtain pSHF cells; and S4, performing a third contact culture on the pSHF cells to obtain SAN-like cells. This invention solves at least one of the following problems in the prior art: the induction efficiency of sinoatrial node (SAN)-like cells from pluripotent stem cells is generally low, usually only 30%–50%; and the differentiation cycle is long, the operation steps are cumbersome, and a variety of small molecule additives are required. In addition, the differentiation products contain a large number of NKX2.5 positive atrial or ventricular myocytes, resulting in insufficient purity, which still requires further cell separation and purification.

To provide a method for efficiently inducing differentiation in vitro of spherical human cell aggregates containing axial mesodermal cells from human pluripotent stem cells, in order to realize the transplant application to patients with degenerative disc disease; and to provide the spherical human cell aggregates containing axial mesodermal cells induced to differentiate in vitro.SOLUTION: A spherical human cell aggregate has the following characteristics: (1) it has two distinct regions, a primitive node area and a non-node area; (2) the primitive node area and the non-node area have a structure consisting of at least two layers; (3) axial mesoderm cells are present in the first layer on the surface of the primitive node area, and endoderm cells are substantially absent in the primitive node area; and (4) endoderm cells are present in the first layer on the surface of the non-node area.SELECTED DRAWING: None

To enable the construction of specific disease models involving multiple cell types and the development of screening systems for specific diseases, we provide a method for producing organoids containing ectoderm and mesoderm cells. [Solution] A method for producing organoids containing ectoderm cells and mesoderm cells, comprising the step (1) of culturing a cell aggregate containing pluripotent stem cells in suspension for at least two days in a culture medium containing a ROCK inhibitor and a TGFβ inhibitor.