194 results about "Proliferative capacity" patented technology

The invention provides a humanized PD-L1 tumor cell line MC-38-hPD-L1, a builtanimal tumor model with the same and a method for constructing the humanized PD-L1 tumor cell line. The method particularly includes knocking out animal-origin PD-L1 by the aid of CRISPR-CAS9; carrying out amplification and cultivation to obtain knocked-out cell banks; extracting DNA (deoxyribonucleic acid) and carrying out PCR (polymerase chain reaction) amplification; recycling and cloning amplification products; carrying out over-expression on human-origin PD-L1 in MC-38 cell lines of mPD-L1 KO by the aid of lentivirus systems; packaging lentivirus and screening Puromycin to obtain the humanized MC-38 cell line of PD-L1. The humanized PD-L1 tumor cell line, the animal tumor model and the method have the advantages that as shown by results, high killing efficiency and multiplication capacity are obviously presented by tumor infiltration CD8 T lymphocytes after antibody treatment is carried out, tumor infiltration Treg cells can be obviously inhibited after antibody treatment is carried out, and accordingly the method is proved to be effective and feasible from the aspect of molecular mechanisms.

The invention provides compositions and methods related to human telomerase reverse transcriptase (hTRT), the catalytic protein subunit of human telomerase. The polynucleotides and polypeptides of the invention are useful for diagnosis, prognosis and treatment of human diseases, for changing the proliferative capacity of cells and organisms, and for identification and screening of compounds and treatments useful for treatment of diseases such as cancers.

The present invention is directed to cells comprising a recombinant polynucleotide sequence that encodes a telomerase reverse transcriptase protein, variant, or fragment having telomerase catalytic activity when complexed with a telomerase RNA.

The invention discloses an HEK (human embryonic kidney) 293 cell line applicable to serum-free culture and an application thereof. According to the invention, the HEK 293 cell (293SF) applicable to serum-free culture is obtained by adopting a culture solution progressive substitution method, wherein the preservation number of the HEK 293 cell is CGMCC (China General Microbiological Culture Collection Center) No.5824. Detection shows that the 293SF cell has stable adenovirus proliferation capacity and foreign protein expressing ability; average lesion time is 97 hours and is reduced by 12.9% compared with the HEK 293 cell; virus virulence reaches up to 107.48TCID50/mL and is improved by 43.3% compared with the HEK293 cell; and stability is good while generation number is improved, and the state and susceptibility of the 293SF cell are not changed after the 293SF cell is continuous passage is carried out for sixteen times. The cell line disclosed by the invention can be used for serum-free production of an adenovirus vector vaccine.

The invention relates to an improved mesenchyme stem cell protection solution as well as application and a preparation method thereof. The protection solution can effectively prolong the activity remaining time of mesenchyme stem cells, reduces preparation cost, and has the advantages of wide raw material source, simple preparation, safe and reliable direct clinical application; and after the mesenchyme stem cells are preserved for 48 hours in the protection solution, the cell activity is still above 90 percent, the cell morphology is normal, and the multiplication capacity and mesenchyme stem cell phenotype characteristics are not influenced.

The invention discloses a cultural method for inducing a human embryonic stem cell to directionally differentiate into a corneal limbal stem cell, which comprises the following steps that firstly, a DMEM (Dulbecco's Modified Eagle Medium)/F12 conditioned medium is adopted to culture a human primary corneal limbal stem cell to prepare a corneal limbal stem cell conditioned medium, and then the corneal limbal stem cell conditioned medium is utilized and combined with IV type collagen culture in vitro to induce the human embryonic stem cell to directionally differentiate into the corneal limbal stem cell. According to the corneal limbal stem cell obtained by utilizing the cultural method, through light microscope observation in vitro, electron microscope observation, real-time quantitative polymerase chain reaction, immunofluorescence, flow cytometry, cloning efficiency determination and the like, the induced cell has a similar shape and phenotype with a normal corneal limbal stem cell, has good differentiation and proliferation capacity in vitro, can be transferred in vitro for more than four generations and can be used as a seed cell for preparing a corneal graft.

The invention belongs to the technical field of stem cells and relates to a serum-free medium for placenta-derived mesenchymal stem cells and a preparation method thereof. The serum-free medium comprises a DMEM basic medium and further comprises vitamin H, glutathione, recombinant human insulin, human serum albumin, transferrin, fibroblast growth factors, epidermal growth factors, stem cell growth factors, Human stem cell factors and magnolol. The serum-free medium provided by the invention has the advantages of high safety and capability for significant improvement of proliferative capability of placenta-derived mesenchymal stem cells, the serum-free medium is capable of well keeping cell forms and stem cell characteristics of the placenta-derived mesenchymal stem cells and is convenient to popularize and apply.

The invention relates to a preparation method of CIK (Cytokine Induced Killer) cells with high proliferation capacity, high cytotoxic activity and high survival rate. The preparation method comprises the steps of: (1) sorting and removing CD4<+>CD25<+>Treg cells of peripheral blood mononuclear cell to obtain CIK pre-cells; (2) cultivating the CIK pre-cells in a cell culture fluid containing 100ng/ml of PHA, 100ng/ml of IL-6 and 10ng/ml of PGE2 for 24h; (3) transferring the CIK pre-cells to a cell culture bottle coated with 1microgramme/ml of CD3 monoclonal antibody, and adding 1000U/ml of IFN-gamma for cultivating for 48h; (4) adding 1000U/ml of IL-2 and 100ng/ml of IL-1alpha for cultivating for 4 days; and (5) adding 1microgramme/ml of insulin to continuously cultivate for 7-14 days. The invention further provides associated CIK cells, a method for inhibiting the peripheral blood mononuclear cell to be differentiated to the CD4<+>CD25<+>Treg cells and a method for promoting the proliferation of the CIK cells. The preparation method of the CIK cells provided by the invention is skillful in design, and the tumor killing cells-CIK cells prepared have stronger proliferation capacity, higher cytotoxic activity and better tumor killing efficiency, so that the clinical efficacy is improved, and the preparation method is appropriate for wide application in a large scale.

A product and process for extending the replicative capacity of metazoan somatic cells using targeted genetic amendments to abrogate inhibition of cell-cycle progression during replicative senescence and derive clonal cell lines for scalable applications and industrial production of metazoan cell biomass. An insertion or deletion mutation using guide RNAs targeting the first exon of the transcript encoding each protein is created using CRISPR/Cas9. Targeted amendments result in inactivation of p15 and p16 proteins which increases the proliferative capacity of the modified cell populations relative to their unaltered parental populations. Combining these amendments with ancillary telomerase activity from a genetic construct directing expression of a telomerase protein homolog from a TERT gene, increases the replicative capacity of the modified cell populations indefinitely. One application is to manufacture skeletal muscle for dietary consumption using cells from the poultry species Gallus gallus; another is from the livestock species Bos taurus.

The invention provides compositions and methods related to human telomerase reverse transcriptase (hTRT), the catalytic protein subunit of human telomerase. The polynucleotides and polypeptides of the invention are useful for diagnosis, prognosis and treatment of human diseases, for changing the proliferative capacity of cells and organisms, and for identification and screening of compounds and treatments useful for treatment of diseases such as cancers.

The invention provides compositions and methods related to human telomerase reverse transcriptase (hTRT), the catalytic protein subunit of human telomerase. The polynucleotides and polypeptides of the invention are useful for diagnosis, prognosis and treatment of human diseases, for changing the proliferative capacity of cells and organisms, and for identification and screening of compounds and treatments useful for treatment of diseases such as cancers.

A method of producing a cell mass by three-dimensional culture of primary cancer cells having proliferative ability and properties of handleability, versatility, and high-throughput performance, in which a tumor tissue is used as a starting material, proliferation of cells such as fibroblasts other than cancer cells is inhibited, and the cell mass includes primary cancer cells as a main component. The object is achieved by providing a method of producing a cell mass by three-dimensional culture of primary cancer cells using a tumor tissue, including: a three-dimensional culture step of culturing cells obtained from the tumor tissue in a medium containing a 5% v/v or less extracellular matrix on a substantially low-adhesive cell culture substrate.

The invention discloses a cell freezing medium. The cell freezing medium comprises a basal culture medium, glycerin, fetal calf serum and a non-permeable protective agent, and the contents of the components are 80-94 v/v% of the basal culture medium, 5-10 v/v% of glycerol, 1-10 v/v% of the fetal calf serum and 0.01-0.10 w/v% of the non-permeable protective agent, and the basal culture medium comprises RPMI-1640, MEM, high-sugar DMEM, low-sugar DMEM or DMEM-F12; and the non-permeable protective agent comprises sodium alginate, sodium hyaluronate or gamma-polyglutamic acid. Dimethyl sulfoxide isnot contained, a good freezing effect is achieved through different matching of the non-permeable protective agent with glycerin and fetal calf serum, a strong water molecule complexing ability is achieved, the freezing function is similar to the freezing function of dimethyl sulfoxide, but formation of intracellular ice crystals is reduced during freezing; the cell freezing medium has clear added ingredients, high safety, small damage to cells, consistent morphology before and after freezing, a high cell survival rate after cell resuscitation and good proliferative capacity.

Phospholipid scramblase 3 (PLS3) is a newly recognized member of a family of proteins responsible for phospholipid translocation between two lipid compartments. A novel isoform of PLS3 is identified and characterized herein. The function of PLS3 in mitochondria was disrupted, yielding an inactive mutant PLS3(F258V). Cells transfected with PLS3(F258V) exhibited reduced proliferative capacity that was unaffected by the presence of Na₃N. PLS3(F258V)-expressing cells exhibit abnormal mitochondrial metabolism and structure and were associated with decreased sensitivity to UV- and tBid-induced apoptosis, and diminished translocation of cardiolipin to the outer mitochondrial membrane. Cells transfected with wild-type PLS3 displayed increased sensitivity to apoptosis and enhanced cardiolipin translocation. These studies identify PLS3 as a regulator of mitochondrial structure and respiration, and cardiolipin transport in apoptosis.

The present invention relates to the identification of the genomic promoter region of the human and mouse telomerase RNA gene. Telomerase activity is necessary for the unrestricted proliferative capacity of many human cancers. It is proposed that mutation or dysregulation of the telomerase repression pathway may cause reactivation or upregulation of telomerase expression in cancer. The invention provides details of elements important for the regulation of telomerase RNA genes, including the Sp family of transcription factors. There is further provided methods for screening elements having the ability for suppressing telomerase RNA gene promoter activity and use of such elements in the treatment of cancers. In addition, evidence is also provided for the development of new transcription based therapies for cancer and for genetic approaches to targeting therapeutic genes to cancer cells. Namely, (1) transcriptional repression and the disruption of signal transduction pathways regulating telomerase activation. (2) Tumour specific gene expression for genetic therapy via telomerase RNA gene promoters.

The invention discloses culturing, screening and amplifying techniques for hair follicle stem cells for clinic treatment level cell therapy. The bottlenecks are always research hotspots and difficulties in the biological stem cell field; currently, by carrying out separation and purification by virtue of remarkable adhering properties of a basement membrane, the hair follicle stem cells are obtained by virtue of three-dimensional culture and amplification measures. By depending on amplification anchorage-dependent cells, a three-dimensional high-simulation in-vivo extracellular matrix system is provided for screening and separating the levels of target cells. The method is a novel culture technique for culture in vitro of biological cells of stem cells of adults (embryo skin) in a high-simulation in-vivo adhesion environment. Skin seed hair follicle cells of targeted tissue engineering cultured through adhesion screening in the environment of a high-simulation substrate have high abdication capacity and multiplication capacity and can form a differentiated cortex structure in vitro. By screening immunogenic cells, immune escape is safely achieved, the transplant time is prolonged, the clinic application of tissue engineering is increased, the healing of wounds is promoted, and a solution with absolute advantages for the clinic treatment level cell therapy is provided for diseases of skin.

The invention provides compositions and methods related to human telomerase reverse transcriptase (hTRT), the catalytic protein subunit of human telomerase. The polynucleotides and polypeptides of the invention are useful for diagnosis, prognosis and treatment of human diseases, for changing the proliferative capacity of cells and organisms, and for identification and screening of compounds and treatments useful for treatment of diseases such as cancers.

The invention belongs to the field of cells and bioengineering, and particularly relates to a stem cell cryopreservation liquid and a cryopreservation method. The stem cell cryopreservation liquid iscomposed of a main component and an additive component, the main component is PBS, normal saline or a basal culture medium, and the additive component is human serum albumin, stem cell growth factors,gamma-polyglutamic acid and glycerin. A stem cell suspension is centrifuged, supernatant is removed, the product is mixed with the stem cell cryopreservation liquid, and subpackaging and cryopreserving are carried. According to the invention, DMSO and FBS are not contained; gamma-polyglutamic acid glycerol is matched differently; the cryopreservation liquid can obtain better freezing effect by combining with human serum albumin, has strong water molecule complexing capacity, can reduce the formation of ice crystals in cells in the freezing process, can be used for cryopreservation of cells, and has the positive effects of consistent forms before and after cell cryopreservation, high survival rate after cell resuscitation and good proliferation capacity of cells after cryopreservation resuscitation.

The invention provides a human placenta mesenchymal stem cell, a preparation method and an application thereof. The preparation method comprises the following steps: collecting, separating, culturing,cryopreserving, detecting, recovering, and the like. A high-purity high-activity mesenchymal stem cell can be acquired by only performing slide adherent culture after a human placenta chorion tissueblock is acquired in the separating process; the technological process is simplified; the cost of enzymic digestion is saved; the adherence and growth rate of primary cells are accelerated; the cell culture period is shortened; the introduction of more external interference factors is avoided, so that the process stability can be easily controlled. The multiplication capacity of the human placentamesenchymal stem cell acquired according to the invention is more stable than that of other mesenchymal stem cells; after the human placenta mesenchymal stem cell passes to P20 generation, the cell still can stably proliferate, and the cellular morphology, molecular surface antigen and adipogenesis osteogenesis differentiative potential thereof all meet the regulations for minimum standard of MSCidentification from international cell therapy association.