722results about "Gastrointestinal cells" patented technology

The invention belongs to the technical field of biology, and discloses a reagent combination or kit for constructing intestinal organs and application of the reagent combination or kit. According to the reagent combination or the kit, intestinal organs can be obtained from cell-derived epithelial cells obtained from a donor in a non-invasive manner, and the obtained intestinal organs can be cryopreserved and recovered and can be amplified in vitro for a long time; transcriptome characteristics are similar to those of real human small intestine tissues, and typical marker genes of various small intestine pedigree cell types are highly expressed; compared with intestinal organs obtained through induction of pluripotent stem cells, the intestinal organs have more intestinal pedigree characteristics, and the intestinal function development is more mature; after being promoted to be mature, the intestinal organ also highly expresses genes related to drug absorption and metabolism, has drug absorption capability similar to that of an immortalized intestinal cell line, but more prominently shows intestinal cell lineage characteristics, is closer to an intestinal environment in a real human body, and can be used for screening intestinal disease drugs; the intestinal barrier function is realized.

The invention discloses a personalized in-vitro total digestion glycolysis simulation system, and belongs to the technical field of in-vitro digestion simulation. According to the invention, by dynamically regulating and controlling the emptying amount and the flow velocity of digestive juice, the process of'digesting and absorbing at the same time 'in the small intestine is simulated, and the interaction of colon flora and colon organs is enhanced in combination with a micro-fluidic chip technology, so that the accurate detection of the digestion process and digestion products is realized. The digestion product simulated by the method disclosed by the invention and the flora structure are very close to the real in-vivo structure, so that the method has relatively high authenticity and credibility.

The invention discloses a 3D intestinal organ differentiation method based on human pluripotent stem cells and an induction culture medium and application thereof, and relates to the technical field of stem cells. According to a culture medium formula combination, intestinal organs can be differentiated into various cell types such as epithelial cells, neuroendocrine cells and endothelial cells; the method is a key mark for successful differentiation and functional maturation of intestinal organs. According to the 3D intestinal organ differentiation method disclosed by the invention, histological structures such as intestinal crypts are differentiated from intestinal organs generated by differentiation, and the intestinal organs can creep in a maintenance stage, so that the intestinal organs are changed from structural bionics to functional simulation, and the significance of the 3D intestinal organ differentiation method is far better than that of pure morphological simulation. Through a systematic culture medium formula, a clear operation process and a multi-stage induction strategy, the 3D intestinal organ with structural integrity, cell diversity and functional activity is successfully constructed, and the system provides an efficient, reliable and extensible in-vitro model platform for intestinal biological research and related application.

The invention discloses a metabolite screening method for promoting expression of porcine intestinal defensin pBD1 and application of the metabolite screening method. The screening method comprises the following steps: 1) constructing a porcine defensin pBD1 gene fluorescence report intestinal epithelial cell line; 2) evaluating the promotion effects of different metabolites based on the pBD1 gene fluorescence report intestinal epithelial cell line; and 3) further evaluating the promotion effects of the metabolites screened in the step 2) based on a porcine intestinal epithelial cell IPEC-J2 model. And 4) further evaluating the promotion effect of the metabolite screened in the step 3) based on the pig intestinal organ. The invention can be used for preparing or screening drugs influencing the expression of the porcine intestinal defensin pBD1. According to the application disclosed by the invention, the effect of the D-malic acid for improving the immune function of the pig intestinal tract by promoting the expression of the defensin pBD1 is found for the first time, verification is carried out on a pig intestinal tract organ-like model, and meanwhile, the optimal treatment concentration of the D-malic acid for promoting the expression of the defensin pBD1 in the pig intestinal tract is optimized.

The invention discloses an application of an MDH2 knockout system or inhibitor in reducing toxicity of T-2 toxin, and belongs to the technical field of biological agents. An MDH2 knockout IPEC-J2 cell line is constructed through a CRISPR / Cas9 gene editing system, and it is found that the cell activity of the cell line is remarkably improved under the condition of 20 ng / mLT-2 toxin concentration, the cell death rate is remarkably reduced, and the cell line has obvious resistance to T-2 toxin. The MDH2 inhibitor LW6 can also enhance the resistance of the cells to T-2 toxin when being used for treating the cells. Therefore, the toxicity of the T-2 toxin in the cell can be reduced by knocking out the MDH2 gene in the IPEC-J2 cell and inhibiting the activity of the MDH2, the tolerance of the cell to the T-2 toxin is improved, and a theoretical basis is provided for prevention and control of harm of the T-2 toxin to human and animals.

The present invention relates to an intestinal epithelium model accurately reproducing the pathophysiological mechanisms observed in vivo in the context an inflammatory condition of the intestinal barrier. The model according to the present invention comprises two compartments separated by a semi-permeable membrane. The first compartment, corresponding to the apical pole of the intestinal epithelium, comprises a coculture of Caco-2 cells differentiated into enterocytes and HT29-MTX cells differentiated into goblet cells. The second compartment, corresponding to the basolateral pole of the intestinal epithelium, comprises a culture of THP-1 monocytic cells differentiated into macrophages. The model according to the present invention is characterized in that the cells contained in the first compartment produce interleukin 6 (IL-6) at a concentration of more than 100 pg / ml and interleukin 8 (IL-8) at a concentration of more than 150 pg / ml, and the cells contained in the second compartment produce tumour necrosis factor α (TNF-α) at a concentration of more than 40 pg / ml and interleukin 1β (IL-1β) at a concentration of more than 90 pg / ml. The present invention also relates to the method for obtaining this model and also to a method for selecting candidate compounds for the treatment of inflammatory bowel diseases by verifying their ability to stop the inflammation as soon as the permeability is increased, or verifying the restoration of the permeability and of the mucus layer.

The invention relates to the technical field of tumor biomedical engineering and organ chips, in particular to a vascularized gastric cancer organ chip and a preparation method thereof.The chip is composed of an integrated micro-fluidic main body, an annular micro-column array and an optical sealing film, and a central culture cavity is divided into a tumor area and a blood vessel area by micro-columns; the preparation method comprises the following steps: injecting a fibrous protein solution containing human umbilical vein endothelial cells and cancer-related fibroblasts into a vascular region, and adding thrombin for in-situ gelation to form a pre-vascularized network; the method comprises the following steps: mixing a patient-derived gastric cancer organ with a methacrylic acid esterified gelatin pre-polymerized solution, injecting the mixture into a tumor area, and carrying out photo-crosslinking immobilization; culturing for 7-14 days under the dynamic perfusion condition of a mixed culture medium to obtain a three-dimensional gastric cancer organ model containing a capillary network; the method is expected to be used for rapid, low-cost and high-throughput screening of chemotherapy or anti-angiogenesis drugs and accurate prediction of individual curative effects of patients.

The invention discloses an inflammatory bowel disease model, a construction method and application thereof and a drug evaluation method based on the inflammatory bowel disease model, and belongs to the technical field of inflammatory bowel disease model construction. A highly bionic intestinal crypt-villus three-dimensional structure is formed based on self-organization of an intestinal cell line C2BBe1, the intestinal crypt-villus three-dimensional structure and a DMEM complete medium containing dextran sulfate sodium are added into an upper chamber of a double-layer culture chamber Transwell, the DMEM complete medium is added into a lower chamber of the double-layer culture chamber Transwell, culture is performed for 2 days, and the inflammatory bowel disease model is obtained. The method takes effect quickly and is highly simulated, the obtained model can simulate the space-time dynamic development process of intestinal inflammation molecular characteristics, and the natural recovery model and the drug treatment model constructed based on the model can provide a brand new platform for researching an inflammation regression mechanism and a drug intervention effect.

The present invention relates to inflammatory bowel disease model production using patient-derived intestinal organoids, and use thereof, wherein the inflammatory bowel disease model produced by an optimized production method for inflammatory bowel disease model production can be used as a patient-customized bowel disease model by constructing an evaluation method for inflammatory bowel disease drugs.

The invention discloses a co-culture method of gastric cancer organoid and gastric cancer-related myofibroblast and application of the co-culture method, gastric cancer organoid single-cell suspension and gastric cancer-related myofibroblast single-cell suspension are mixed according to the number ratio of 1: 1, then matrigel is added, after uniform mixing, the mixture is put into a CO2 cell incubator to solidify the matrigel, and the gastric cancer-related myofibroblast co-culture medium is obtained. The co-cultured gastric cancer organoid is faster in growth speed and stronger in resistance to chemotherapeutic drugs, the drug resistance of tumor cells in the body of a gastric cancer patient to the chemotherapeutic drugs is better simulated, and the co-cultured gastric cancer organoid can become a screening platform of individualized therapeutic drugs for tumor patients.

The invention relates to the technical field of tissue isolation, and particularly discloses a high-activity intestinal crypt and an isolation method and application thereof. The separation method comprises the following steps: S1, taking fresh intestinal tracts, cleaning contents, and scraping mucous membrane layer tissues to obtain intestinal mucous membrane tissues; s2, shearing intestinal mucosa layer tissues, mixing the intestinal mucosa layer tissues with the separation liquid after cleaning, and digesting and dissociating to obtain intestinal mucosa tissue digestive juice; s3, centrifuging the intestinal mucosa tissue digestive juice, removing supernate, mixing the supernate with the protective liquid to obtain intestinal crypt mixed liquid, filtering, centrifuging, and taking precipitate to obtain high-activity intestinal crypts; wherein the separation liquid comprises collagenase I, collagenase II, collagenase IV, elastase, DNA enzyme I and a ROCK inhibitor; the protection liquid comprises an RPMI1640 culture medium, antibiotics, fetal calf serum, bovine serum albumin and a ROCK inhibitor. The technical scheme provided by the invention aims to shorten the intestinal tract crypt separation time and improve the cell viability.

The invention discloses a mouse-derived gastric adenocarcinoma cell line MKC1 and application thereof. The mouse gastric adenocarcinoma cell line MKC1 (Mus musculus) is preserved in the China Center for Type Culture Collection, the preservation date is June 15, 2024, and the preservation number is CCTCC (China Center for Type Culture Collection) NO: C2024183. The cell line belongs to a murine gastric adenocarcinoma cell line, can be stably passaged, is high in growth speed, and can form primary tumors in a C57BL / 6 mouse body with a sound immune system. In addition, the cell line has the advantages of being high in repeatability, stable and single in property and the like, and is suitable for related research of gastric cancer immune microenvironment, tumor drug resistance, invasion and metastasis and the like.

The invention relates to the technical field of cell biology, in particular to a construction method and application of intestinal organs for children with Crohn's disease, the intestinal organs for children with Crohn's disease are successfully constructed by utilizing intestinal tissues from children with Crohn's disease, and the intestinal organs have various characteristic cells of the intestinal tissues; besides, immune cells are extracted from peripheral blood of a child patient homologous with intestinal tissues, a co-culture system is established with the organ-like model, the intestinal immune microenvironment of the child patient suffering from the Crohn's disease can be reproduced, and the system provides a new model for related mechanism exploration and drug screening of the child Crohn's disease.

The invention discloses a construction method and application of a gastric cancer treatment effect evaluation model of CAR-T cells, and the constructed gastric cancer treatment effect evaluation model of the CAR-T cells comprises a CAR-T cell and gastric cancer organ co-culture killer dynamic detection system and a CAR-T cell, intestine organ and lung organ co-culture safety evaluation system. Based on a CAR-T cell and gastric cancer organoid co-culture lethality dynamic detection system, the process that CAR-T cells act on gastric cancer organoid can be dynamically observed through a living cell imaging method, then the action mechanism of the CAR-T cells for treating solid tumors is studied, and meanwhile, based on a CAR-T cell, intestine organoid and lung organoid co-culture safety evaluation system, the CAR-T cells and the gastric cancer organoid co-culture lethality dynamic detection system can be used for evaluating the CAR-T cells and the gastric cancer organoid co-culture lethality. Compared with the prior art, the preclinical model has the advantages that the effect of the CAR-T cells on gastric cancer para-tissues and other tissue-derived organs can be detected, and safety evaluation is carried out, so that the preclinical model for more comprehensively detecting the gastric cancer treatment effect of the CAR-T cells is obtained, and the research on CAR-T cell targets is facilitated.

The invention discloses double-layer cross-linked artificial matrigel and an application method of the double-layer cross-linked artificial matrigel in a small intestine micro-physiological system. Low-concentration calcium ions are adopted for primary crosslinking to form a soft inner layer structure, and then high-concentration calcium ions are used for secondary crosslinking to improve the overall rigidity. The matrigel is used for dynamic culture of small intestine organoid and is combined with a micro-fluidic chip to form a small intestine micro-physiological system. In the system, the digestive absorption process of the small intestine is dynamically simulated by controlling the flow ratio and rate of digestive fluid and culture fluid. Compared with the traditional natural matrigel, the matrigel disclosed by the invention has more appropriate mechanical strength and structural stability, supports long-term growth of the organoids, and improves the multiplication capacity, tight connection and structural integrity of the organoids in an in-vitro environment. The method can be used for researching intestinal diseases, food digestion and drug absorption, and is an in-vitro digestion simulation platform with high biocompatibility and stability.

The invention relates to the technical field of organoid culture, and relates to a culture system and a culture method for gastrointestinal tumor organoids for the first time, the culture system is formed by adding antioxidant active traditional Chinese medicine extracts such as naringin or polydatin on the basis of an existing culture system, when the culture system is used for culturing the gastrointestinal tumor organoid, the primary survival rate of the gastrointestinal tumor organoid and the proliferation rate after passage can be improved. Besides, the invention further provides application of the culture system of the gastrointestinal tumor organoid, the culture system and the cultured gastrointestinal tumor organoid are used for screening small-molecule targeted anti-tumor drugs, and a better in-vitro research model is provided for subsequent anti-tumor treatment.

The present invention relates to a method for constructing a gene network through a single-cell transcriptome and a method for discovering key genes in differentiation using same, and a composition for the prevention, alleviation, or treatment of colon cancer using the target discovered through the method. The method for constructing a gene network of key genes according to the present invention employs single-cell transcriptome data and thus can be applied to all single-cell transcriptome data. The combination of MYB / HDAC2 / FOXA2 discovered upon application to colon cells can serve as a cancer treatment target that promotes the differentiation of colon cancer cells to revert same into differentiated normal cells.

The invention discloses an application of an MDH2 gene and an inhibitor thereof in resisting ZEA toxin, and relates to the technical field of biology. An MDH2 gene deletion cell line is obtained through CRISPR gene editing, and compared with a wild type MDH2 gene expression cell, under the influence of ZEA, the cell activity of the MDH2 gene deletion cell line is remarkably improved, and the cell apoptosis rate is remarkably reduced. In addition, the LW6 inhibitor is used for inhibiting the MDH2 activity of wild type IPEC-J2 cells, and the tolerance of the cells to ZEA can also be improved. Therefore, through knockout, mutation or activity inhibition of the MDH2 gene, the ZEA resistance of people, animals and plants can be improved. The invention provides a potential solution way for solving untoward effects caused by ZEA (Zinc Ethylenediamine).

The invention provides a method for separating, culturing and activating NK (Natural Killer) cells and an application method of the NK cells in tumor immunity. The method comprises the following steps: separating mononuclear cells from peripheral blood of healthy adults, removing non-NK cell populations through a negative selection method, and enriching and purifying NK cells. Through stimulation of specific cytokines, NK cells are successfully amplified, and cytotoxicity of the NK cells is activated. By optimizing the preparation and activation method of the NK cells, the NK cells with efficient killing activity are obtained in vitro, and the tumor treatment effect is remarkably improved through combined use of the NK cells and the immune checkpoint inhibitor. In combination with the immune memory effect of the NK cells, the invention provides a new anti-tumor immunotherapy method which can be applied to preparation of new anti-tumor drugs and provides an innovative thought for tumor treatment.

Provided are chemical inducers of pluripotency (CIP) which include glycogen synthase kinase inhibitors, TGFβ receptor inhibitors, cyclic AMP agonists and S-adenosylhomocysteine hydrolase (SAH) inhibitors or histone acetylators. A method of inducing pluripotency in a partially or completely differentiated cell by using such chemical inducers of pluripotency is also provided. The method includes: (i) contacting a cell with the CIPs for a sufficient period of time to result in reprograming the cell into a pluripotent stem cell having ESC-like characteristics (CiPSC). Isolated chemically induced pluripotent stem cells (CiPSCs) and their progeny, produced by inducing differentiation of the CiPSCs, can be used in a number of applications, including but not limited to cell therapy and tissue engineering.

The invention discloses a preparation method and application of a multi-material cell-loaded 3D scaffold, and the preparation method comprises the following steps: S1, preparing at least two kinds of printing ink; s2, various printing inks prepared in the step S1 are added into 3D printing equipment, and a 3D support is printed according to a designed model; the printing ink comprises a main material and a photoinitiator, the main material comprises one or a mixture of a photosensitive material and a non-photosensitive material, and at least one kind of printing ink further comprises a cell-loaded suspension. The processing and manufacturing efficiency and convenience can be improved, the manufacturing cost is reduced, meanwhile, dynamic culture and testing of the carrier cells are achieved, and the scientific research efficiency is improved.

The present invention discloses a buffer for tissue separation and / or preservation and its application, and a method for culturing tumor organoids, and relates to the field of organoid culture. The buffer is used to preserve tumor tissue samples, which are of liquid origin; the buffer includes: a basal culture medium and specific additive factors, and the specific additive factors include: superoxide dismutase, catalase, penicillin, and streptomycin; based on the volume of the basal culture medium, the concentration of superoxide dismutase is 1 to 100 μg / mL, the concentration of catalase is 50 to 500 μg / mL, the concentration of penicillin is 50 to 200 U / mL, and the concentration of streptomycin is 50 to 200 μg / mL. The tissue preservation solution of the present invention is used for the transportation and protection of liquid-derived tumor tissue samples, and has a high tumor cell activity retention rate. It is used to construct tumor organoids with a high success rate and a large number, which can meet the needs of clinical detection and disease research.

The invention belongs to the field of organoid culture, and particularly relates to a mixed type gastric cancer organoid culture medium and a culture method and application thereof. Wherein the culture medium comprises a basic culture medium and a specific additive factor; the culture method comprises the following specific steps: treating mixed gastric cancer tissues with tissue digestive juice to obtain mixed gastric cancer primary cells, and culturing with a mixed gastric cancer organ culture medium to obtain mixed gastric cancer organs. The mixed type organ-like form corresponding to the mixed type gastric cancer tissue can be cultured by using the mixed type gastric cancer organ-like culture medium disclosed by the invention. By means of methods such as organoid passage and cryopreservation technologies, the culture success rate of the mixed type gastric cancer organoid is high, sufficient mixed type organoids can be obtained within limited time, an accurate tumor model is provided for individualized tumor therapy, and valuable samples are preserved for scientific research.

The invention discloses a culture system, culture method and application of gastrointestinal tumor chemotherapy drug-resistant organoid, the culture system comprises: a basic culture medium, which is a DMEM / F12 culture medium, and is additionally added with a special growth promoting factor obtained from a natural extract; the multiple growth factors comprise an epidermal growth factor (EGF), a fibroblast growth factor (FGF), a bone morphogenetic protein (BMP) and a vascular endothelial growth factor receptor inhibitor (VEGFRi), and the concentration of each growth factor is accurately screened and optimized; the extracellular matrix is Matrigel modified by physical and chemical methods; the specific additive comprises an antioxidant N-acetylcysteine (NAC). According to the culture system, the culture method and the application of the gastrointestinal tumor chemotherapy drug-resistant organoid, the culture system which is closer to a real environment in vivo is constructed by optimizing basic culture medium components, setting various growth factors and specific additive concentrations and adopting modified Matrigel as an extracellular matrix.

The invention relates to the technical field of organoid culture, in particular to tumor organoid culture hydrogel with definite components and a preparation method and application thereof, and the tumor organoid culture hydrogel with the definite components comprises a gel precursor and laminin 521; the gel precursor is prepared by dissolving sodium alginate and gelatin in PBS; the concentration of the laminin 521 is 0.10 mg / ml to 0.15 mg / ml. The hydrogel provided by the invention is clear in component, the performance of the hydrogel for culturing the tumor organoid is equivalent to that of Matrigel, and the technical problems that Matrigel is complex in component, different in batches and high in price, and consequently conversion and application of a tumor patient source organoid model are limited are solved. Besides, the hydrogel provided by the invention is high in component industrialization degree, good in stability and easy to obtain, standardized and economic production can be realized, and support is provided for construction of a tumor organ model.

The present invention relates to a method for preparing organ-specific mesenchymal cells from pluripotent stem cell-derived intestinal organoid stromal cells. By using cells derived from stromal cell layers adjacent to intestinal organoids for differentiation into organ-specific mesenchymal cells, the present invention can greatly increase the efficiency of differentiation into stromal cells through the regulation of retinoic acid (RA) and hedgehog (HH) signaling pathways, and can increase the expression of organ-specific markers without exhibiting undifferentiated state cell characteristics, and thus mesenchymal cells having well-simulated biological characteristics can be prepared.

The invention provides an organoid basic culture medium and a method for maintaining and promoting organoid growth, and the organoid basic culture medium comprises a basic culture medium and an additive, the additives comprise taurine sodium 3-hydroxybutyrate, taurine, sodium hypoxanthate, creatinine, L-ornithine, urea, L-sodium lactate, putrescine dihydrochloride, anhydrous citric acid, ethanolamine, N-acetylglycine, alpha ketoglutaric acid, formic acid, malic acid, malonic acid, succinic acid, uric acid, ammonium metavanadate, manganese dichloride, sodium selenite, creatine, glycerol and uridine. Anhydrous sodium acetate and para aminobenzoic acid. When the organ-like basic culture medium is used for treating the organ-like, the activity of the organ-like can be effectively maintained, and basic nutrition and metabolism requirements of the organ-like can be provided, so that the organ-like has the advantages of high activity, fast proliferation and long-term stable passage, and is suitable for various types of organs and wide in application.

The invention discloses a 3D (three-dimensional) visual bionic intestine-blood vessel-nerve interface organ simulation chip, which is characterized in that a micro-fluidic chip is used as a platform to construct an intestinal tract-blood vessel-brain system which simulates biological transformation of perfluorinated compounds in intestinal tracts under near physiological conditions and biologically transports the perfluorinated compounds through blood vessels; a neuronal cell culture chamber, a vascular endothelial cell culture chamber and an intestinal tract cell culture chamber are respectively arranged on the micro-fluidic chip, the culture chambers are communicated through micro-columns, and the vascular endothelial cell culture chamber serves as a barrier in the middle; perfluorinated compounds enter corresponding cell culture cavities for culture through reagent injection channels of different cell culture channels, generated metabolites are discharged through the channels, and multi-organ interaction and toxicity induced by the perfluorinated compounds are analyzed. The chip can simulate an intestinal barrier structure, a blood vessel and a neural network structure, can highly simulate biological transport and transformation of a perfluorinated compound between intestines and brains, and effectively reduces cost and time.

The invention belongs to the technical field of biological medicine, and particularly relates to a construction method of a gastric mucosa intestinal epithelial metaplasm type organ model with goblet cell phenotype. The model is characterized by MUC2 + MUC6 + goblet cells, and histological characteristics of gastric mucosa intestinal metaplasia are more deeply simulated from the aspect of cell phenotype. The construction method comprises the following steps: (1) obtaining a human stomach gland to prepare an organoid; (2) subculturing the organoid; (3) construction of a pCDH-CMV-MCS-EF1-CDX2-Puro overexpression lentiviral vector, and construction of a pCDH- (4) virus packaging; (5) constructing CDX2 overexpression stably transfected cells and organoid; and (6) carrying out gradient differentiation on the CDX2 overexpression organoid to prepare the intestinal metaplasia organoid. According to the invention, the pathological transformation of the cytology level in the gastrointestinal mucosa intestinal epithelial metaplasia process is realized in the in-vitro model for the first time, and a new research model and theoretical guidance are provided for the diagnosis, prevention and individualized precise treatment of precancerous lesions of gastric cancer.