240 results about "In vitro proliferation" patented technology

PROBLEM

There are provided induced malignant stem cells capable of in vitro proliferation that are useful in cancer research and drug discovery for cancer therapy, as well as processes for production thereof, cancer cells derived from these cells, and applications of these cells.

MEANS FOR SOLVING

An induced malignant stem cell capable of in vitro proliferation are characterized by satisfying the following two requirements:

• (1) having at least one aberration selected from among (a) an aberration of methylation (high or low degree of methylation) in a tumor suppressor gene or a cancer-related genetic region in endogenous genomic DNA, (b) a somatic mutation of a tumor suppressor gene or a somatic mutation of an endogenous cancer-related gene in endogenous genomic DNA, (c) abnormal expression (increased or reduced/lost expression) of an endogenous oncogene or an endogenous tumor suppressor gene, (d) abnormal expression (increased or reduced/lost expression) of a noncoding RNA such as an endogenous cancer-related microRNA, (e) abnormal expression of an endogenous cancer-related protein, (f) an aberration of endogenous cancer-related metabolism (hypermetabolism or hypometabolism), (g) an aberration of endogenous cancer-related sugar chain, (h) an aberration of copy number variations in endogenous genomic DNA, and (i) instability of microsatellites in endogenous genomic DNA in an induced malignant stem cell; and
• (2) expressing genes including POU5F1 gene, NANOG gene, SOX2 gene, and ZFP42 gene.

The invention discloses an AFP[158-166] specific TCR gene, its transgenic T cell, and an in-vitro proliferation method and a use of the transgenic T cell. The AFP[158-166] specificity TCR gene is represented by SEQ ID NO.1 in a sequence table. The transgenic T cell of the AFP[158-166] specific TCR gene has a specific killing effect on AFP positive liver cancer cells. Artificial antigen presentation cells can present AFP[158-166] epitope peptides through the MHCI type molecule of the cell surface when the artificial antigen presentation cells are secreted by IL-15 to induce the AFP specific immune reaction, and the stimulation of the AFP[158-166] specific TCR gene transgenic T cell by the AFP[158-166] specific TCR gene can improve the proportion of the transgenic T cell, enhances the specificity and enhances the T cell activity and the antitumor capability in an auxiliary manner through the secretion of IL-15.

The invention relates to a preparation method of a specific extracellular matrix ECM of periodontium. The preparation method comprises the following steps: 1) separating and culturing a human periodontal ligament cell; and 2) preparing the specific extracellular matrix ECM of periodontium; applying the specific extracellular matrix ECM of periodontium and applying the specific extracellular matrix ECM of periodontium as a human periodontal ligament stem cell in in-vitro amplification; and promoting directional differentiation of the ECM. The preparation method is simple and can remarkably promote in-vitro amplification and directional differentiation of the periodontal ligament stem cell.

The invention relates to the field of protein, in particular to a bioactive peptide SKVLPVPEKAVPYPQ as well as a preparation method and an application thereof. An amino acid sequence of the bioactive peptide SKVLPVPEKAVPYPQ is Ser-Lys-Val-Leu-Pro-Val-Pro-Glu-Lys-Ala-Val-Pro-Tyr-Pro-Gln. An in-vitro anti-oxidation experiment and an in-vitro immunity function promoting experiment prove that the bioactive peptide SKVLPVPEKAVPYPQ has better anti-oxidation biological activity and immunity regulation function, on one hand, the bioactive peptide SKVLPVPEKAVPYPQ has better anti-oxidation activity, can clear free radicals in an organism and improves life quality; on the other hand, by the aid of the bioactive peptide SKVLPVPEKAVPYPQ, in-vitro proliferation capacity of lymphocytes and macrophages can be enhanced, the phagocytosis capacity of the macrophages for neutral red is improved, the organism's capacity of resisting outside pathogen infection is improved, morbidity of the organism is reduced, and therefore, the bioactive peptide SKVLPVPEKAVPYPQ has quite great significance in development of food, healthcare products and drugs with an immune regulation function and an anti-oxidation function.

The present invention relates to the field of protein, and in particular relates to biologically active polypeptide WNIPMGLIVNQ and a preparation method and the application thereof, and the amino acid sequence of the biologically active polypeptide WNIPMGLIVNQ is Trp-Asn-Ile-Pro-Met-Gly-Leu-Ile-Val-Asn-Gln. In vitro antioxidant experiments and in vitro immune function promotion experiments verify the biologically active polypeptide WNIPMGLIVNQ has good antioxidant biological activity and immunity regulating function, on the one hand, the biologically active polypeptide WNIPMGLIVNQ has good antioxidant activity to remove free radicals in the body and improve the quality of life; on the other hand, the biologically active polypeptide WNIPMGLIVNQ can enhance the in-vitro proliferation ability of lymphocytes and macrophages to promote the increase of macrophage induced nitric oxide, the biologically active polypeptide WNIPMGLIVNQ can improve the external pathogen infection resistance of the body and reduce the disease incidence of the body, and has very important significance for development of food, health care products and drugs with immune function regulating and antioxidant function.

The invention relates to the field of protein, and in particular to a bioactive polypeptide TIASGEPT, and a preparation method and application thereof. An amino acid sequence of the bioactive polypeptide TIASGEPT is Thr-Ile-Ala-Ser-Gly-Glu-Pro-Thr. An in-vitro antioxidant experiment and an in-vitro immunological function promoting experiment verify that the bioactive polypeptide TIASGEPT has better antioxidant biological activity and a better immune modulating function; on one hand, the bioactive polypeptide TIASGEPT has better antioxidant activity and is capable of removing free radicals in a body and increasing the quality of life; on the other hand, the bioactive polypeptide TIASGEPT is capable of enhancing in-vitro proliferation ability of lymphocytes and macrophages, promoting the increment of nitric oxide induction amount of the macrophages, increasing the ability of defending external pathogen infection by the body and reducing the morbidity of the body, and has a very important significance in developing food, healthcare products and drugs, which have an immune modulating function and an antioxidant function.

The invention relates to the field of protein, in particular to a bioactive peptide DRAAHVKQVL as well as a preparation method and an application thereof. An amino acid sequence of the bioactive peptide DRAAHVKQVL is Asp-Arg-Ala-Ala-His-Vla-Lys-Gln-Val-Leu. An in-vitro anti-oxidation experiment and an in-vitro immunity function promoting experiment prove that the bioactive peptide DRAAHVKQVL has better anti-oxidation biological activity and immunity regulation function, on one hand, the bioactive peptide DRAAHVKQVL has better anti-oxidation activity, can clear free radicals in an organism and improves life quality; on the other hand, by the aid of the bioactive peptide DRAAHVKQVL, in-vitro proliferation capacity of lymphocytes and macrophages can be enhanced, the macrophages are promoted to secrete cytokines, the organism's capacity of resisting outside pathogen infection is improved, morbidity of the organism is reduced, and therefore, the bioactive peptide DRAAHVKQVL has quite great significance in development of food, healthcare products and drugs with an immune regulation function and an anti-oxidation function.

The invention discloses a synthesis and medical application of pyrrolo-[2,1-f] [1,2,4] triazine mother nucleus compound. The compound belongs to a novel-structure compound. The synthesis method is high in operating safety, mild in reaction condition and suitable for industrial production. The activity and the selectivity of BTK kinase and the in-vitro proliferation activity of a leukemia cell line are test, it is verified that the compound has the selective and irreversible inhibition effect on the BTK kinase, and has the different-degree inhibition effect on leukemia cells. According to measurement, the compound also has the good anti-arthritic activity on a collagen-induced arthritis (rCIA) model. The pyrrolo-[2,1-f] [1,2,4] triazine mother nucleus compound can be used for preparing medicine for treating arthritis and leukemia.

The invention relates to a method for extracting and purifying neutral pseudo-ginseng polysaccharide, a research and an application for pharmacological activity for promoting cell proliferation. The pseudo-ginseng waste residue of the notoginsenoside extracted in the production process is taken as a raw material, a water extract and alcohol precipitation method is adopted for acquiring crude pseudo-ginseng polysaccharide and the DEAE Sepharose Fast Flow anion exchange chromatography is adopted for purifying the crude pseudo-ginseng polysaccharide so as to acquire five components: neutral pseudo-ginseng polysaccharide PNPS I and acidic pseudo-ginseng polysaccharide PNPS II, PNPS III, PNPS IV and PNPS V five samples. The research on the influence of the pseudo-ginseng polysaccharide on human periodontal ligament stem cell, mice osteoblast and human skin epidermis cell in vitro proliferation proves that the neutral pseudo-ginseng polysaccharide PNPS I is effective in boosting the periodontal ligament stem cell, mice osteoblast and human skin epidermis cell in vitro proliferation and the neutral pseudo-ginseng polysaccharide PNPS I can be used as an important drug intermediate for developing a new drug and a natural raw material for a functional healthcare food and also can be used as an active raw material of the household chemicals.

The invention relates to the field of proteins and particularly relates to bioactive polypeptide ATLEDSPEVI as well as a preparation method and application thereof. The amino acid sequence of the bioactive polypeptide ATLEDSPEVI is Ala-Thr-Leu-Glu-Asp-Ser-Pro-Glu-Val-Ile. In-vitro anti-oxidation experiments and in-vitro immune function promotion experiments verify that the polypeptide ATLEDSPEVI has better antioxidant biological activity and immune modulating function, so that on one hand, the polypeptide ATLEDSPEVI has better antioxidant activity, so that free radicals in a body can be removed, and the quality of life is improved; on the other hand, the polypeptide ATLEDSPEVI can enhance the in-vitro multiplication capacity of lymphocytes and macrophages, the macrophages are promoted to secrete cytokines, the capability of the body in defense for external pathogen infection is enhanced, and the incidence rate of the body is reduced. Therefore, the polypeptide ATLEDSPEVI has great significance in development of food, health-care products and drugs with immune modulating function and antioxidant function.

The invention relates to the field of anti-tumor drugs, and specifically discloses an indole alkaloid adduct, and a preparation method and an application thereof in preparing an anti-tumor drug. The indole alkaloid adduct has a structure shown as a formula I, wherein R is formed in the way that an indole alkaloid is subjected to hydrazinolysis and the hydrazinolysis product is bonded with hydrazine group. The indole alkaloid adduct can significantly reduce toxicity to normal cells and in-vivo toxicity and inhibit in-vitro proliferation of various tumor cell lines and the growth of tumor in the nude mouse bearing the tumor.

The invention discloses a molecular marker for diagnosing and treating the bladder cancer, namely LncRNA DANCR, and also discloses application of the molecular marker to screening or preparation of reagents used for diagnosing the bladder cancer diagnosing, bladder cancer lymphatic metastasis, bladder cancer prognosis relapse or bladder cancer clinical stages and application of the molecular marker to screening or preparation of drugs used for treating the bladder cancer. It is found that the LncRNA DANCR has high expression in bladder cancer cases with lymphatic metastasis, has negative correlation with overall survival prognosis of a patient and is an independent indicator for diagnosing the bladder cancer and judging bladder cancer development and survival prognosis; silencing of the LncRNA DANCR can restrain in-vitro proliferation, cell migration and invasion and in-vivo tumor formation and lymphatic metastasis of bladder cancer cells. It is verified that the LncRNA DANCR is an important cancerigenic factor of the bladder cancer for the first time, and can be used as a molecular marker for bladder cancer prognosis diagnosis and a new target spot for treatment.

The present invention provides an induced cancer cell capable of self-replication in vitro which is useful in cancer therapy research and the research for cancer-related drug discovery, processes for production thereof, cancer cells induced by the malignant cells, and applications of these cells.

The present invention provides an induced cancer stem cell capable of proliferation (self-replication) in vitro, wherein the induced cancer stem cell has the following two characteristics:

• (1) expressing the six genes (self-renewal related genes) POU5F1, NANOG, SOX2, ZFP42, LIN28, and TERT selected from a certain group of genes; and
• (2) having an aberration which is either (a) a mutation in an endogenous tumor suppressor gene or (b) increased expression of an endogenous cancer-related gene.

The invention discloses a method for transfecting bovine somatic cells into inducted pluripotent stem cells (iPSCs) by adopting transcription factors, concretely comprising the following steps: 1) isolation and culture of bovine dermal fibroblasts; 2) cloning of bovine transcription factor genes maintaining pluripotency and construction of retroviral vectors of the transcription factor genes; 3) retrovirus packaging and preparation; and 4) infection of bovine dermal fibroblasts by retrovirus and acquisition of bovine inducted pluripotent stem cells. The invention first adopts the acellular human amniotic membranes as support to replace the mouse embryonic fibroblasts (MEFs) to maintain in vitro proliferation of the bovine iPSCs and keep the bovine iPSCs in undifferentiated pluripotent states for long time. The method which adopts gene inducing to generate the PSCs not only solves the technical problem that the PSCs of the animals such as cattle, sheep, pigs and other livestock are difficult to obtain but also avoids the problem of heterogeneous animal cell pollution caused by the feeder layer which applies the MEFs as the stem cells.

The present invention relates to an application of bone morphogenetic protein 4 in cancer inhibition. Specifically bone morphogenetic protein (BMP) has a liver cancer stem cell differentiation induction effect, wherein CD133 protein expression of liver cancer cell lines can be down-regulated and liver cancer cell line CD133+ liver cancer stem cell differentiation can be promoted with BMP4, and BMP4 can further be provided for inhibiting in vitro proliferation and self-renewal of liver cancer cell lines and inhibiting tumorigenicity of immune-deficient mice. The present invention further provides BMP4-induced liver cancer stem cell differentiation action mechanism.

Methods are provided for the establishment and maintenance in long term culture of hormone secreting cells. Cells are derived from tumorous or non-tumorous animal or human tissues, including ovary, endometrium, trophoblast, pituitary, thyroid, and pancreas. The cells secrete into the culture medium hormones such as estrogens, progestins, follicle-stimulating hormone, luteinizing hormone, human chorionic gonadotrophin, thyroxin, glucagon, and insulin, depending on the tissue of origin of individual cell cultures. Contact with an appropriate secretogogue causes the cells to respond with increased hormone secretion. For instance, ovarian follicular cells respond to follicle-stimulating hormone with increased estrogen and progesterone secretion. Pancreatic cells respond to elevated glucose with increased insulin secretion. The cells proliferate in in vitro for up to one year or longer, during which time they retain their hormone-secretion profile. The cells may be frozen for storage, and retain their hormone-secretion profile after thawing. The cell cultures are useful for the production of human hormones, for the bio-assay of drugs such as therapeutic gonadotrophin, for the testing of drug efficacy and design, and for toxicity testing of drugs and chemicals. The cells may also be implanted in an individual to replace deficient hormone secretion. For instance, insulin secreting pancreatic cells may be implanted in a diabetic individual as an adjunct or replacement therapy for exogenously administered insulin.

The invention discloses a culture medium and a culture method for constructing a liver tumor scaffoldless organoid. The culture medium comprises the following components: Advanced DMEM/F12, FBS, diabody, N-2, Noggin, B-27, EGF, FGF-10, Y-27632, A83-01, SB202190, R-Spondin, N-acetylcysteine, Nicotinamide, Wnt3A, HGF and PrimocinTM. The culture method comprises the steps of adding primary tumor cells separated and obtained into the above-mentioned special culture medium for resuspension, inoculating into an ultra-low adsorption U-shaped well plate after counting, and centrifugally culturing until organoids are formed. The culture medium and the culture method can realize in-vitro proliferation of primary liver cancer cells, can quickly construct the liver tumor scaffoldless organoids, and can maintain long-term stable growth of the liver tumor organoids. The tumor organoids formed by the culture medium and the culture method retain heterogeneity of tumor tissues of patients, and are suitable for drug sensitivity detection, tumor generation and development related research of in-vitro liver cancer chemotherapeutic drugs.

The embodiment of the invention provides a culture solution for delaying aging of mesenchymal stem cells and a preparation method thereof, and relates to the technical field of biomedical materials. The culture solution reduces the aging rate of the mesenchymal stem cells as much as possible, maintains cell phenotypes and functions of the stem cells and improves the utilization efficiency of the stem cells while in-vitro proliferation of the mesenchymal stem cells is increased, so that requirements of scientific research and clinical application are met. According to the culture solution for delaying aging of the mesenchymal stem cells and the preparation method thereof, the culture solution comprises a basal culture solution and exogenous additives, wherein the exogenous additives include5-15g/L of glucan, 0.1-0.5mL/L of a lipid-based concentrating agent, 0.05-5[mu]g/L of hydrocortisone, 0.1-10[mu]g/L of dexamethasone, 1-5[mu]g/L of sodium selenite, 10-50[mu]M of beta-mercaptoethanol, 10-100mg/L of vitamin C (VC), 10-50[mu]g/L of vitamin E (VE), 1-20mg/L of transferrin, 5-50[mu]g/L of insulin, 10-50[mu]g/L of growth hormone (GH), 10-50[mu]g/mL of glutathione and 1% of non-essential amino acids.

The invention discloses new application of a substance for inhibiting genetic expression and/or protein activity of acyl coenzyme A/cholesterol acyltransferase-1. According to the application, the inhibitor can be used for preparing drugs for preventing and/or treating cancer; the inhibitor can be used for preparing drugs for preventing and/treating cancer spreading and metastasis; the inhibitor can be used for preparing drugs for promoting apoptosis of cancer cells; the inhibitor can be used for preparing drugs for inhibiting cancer cells from being transformed into cancer; the inhibitor canbe used for preparing drugs for inhibiting in vitro proliferation growth of the cancer cells. Through experiments, the inventor proves that the acyl coenzyme A/cholesterol acyltransferase-1 (ACAT1/SOAT1) is obviously highly expressed in liver cancer tissue, and the high enrichment value of the acyl coenzyme A/cholesterol acyltransferase-1 shows that prognosis of a liver cancer patient is poor. TheACAT1/SOAT1 inhibitor can effectively inhibit growth of human liver cancer cells and other cancer cells in the cellular level, and the inhibitor can used as a candidate drug target of cancers, especially the liver cancer.

The invention discloses a method for transfecting bovine somatic cells into inducted pluripotent stem cells (iPSCs) by adopting transcription factors, concretely comprising the following steps: 1) isolation and culture of bovine dermal fibroblasts; 2) cloning of bovine transcription factors maintaining pluripotent genes and construction of retroviral vectors of the transcription factors; 3) retrovirus packaging and preparation; and 4) infection of bovine dermal fibroblasts by retrovirus and acquisition of bovine inducted pluripotent stem cells. The invention first adopts the acellular human amniotic membranes as support to replace the mouse embryonic fibroblasts (MEFs) to maintain in vitro proliferation of the bovine iPSCs and keep the bovine iPSCs in undifferentiated pluripotent states for long time. The method which adopts gene inducing to generate the PSCs not only solves the technical problem that the PSCs of the animals such as cattle, sheep, pigs and other livestock are difficult to obtain but also avoids the problem of heterogeneous animal cell pollution caused by the feeder layer which applies the MEFs as the stem cells.

The invention discloses a mouse primary hepatocyte perfusion type separating and in-vitro culturing method. The perfusion type separating of mouse primary hepatocytes is conduced through an in-situ living body perfusion method, a high-yield single-cell suspension can be obtained, and therefore a basis is provided for in-vitro long-time culturing of the primary hepatocytes. An applied stimulating culture solution contains mouse colony stimulating factors (CSF), mouse IL-2, mouse IL-6 and mouse recombinant hepatocyte growth factors r-mHGF and other cell factors; the primary hepatocytes can be separated through stimulation according to the reasonable ratio for in-vitro proliferation, and the number can reach 2.25 times the number of initially-added primary hepatocytes 72 hours later. The cellproliferation activity can be kept for at least 1-2 weeks under the in-vitro conditions, the problem that the primary hepatocytes cannot be cultured for a long time in vitro is solved, and thereforethe method can be used for infection of retroviruses, provides a basis for over-expression target genes or knockout target genes in in-vitro primary cells, and greatly expands the application field ofthe primary hepatocytes.