544results about "Respiratory/lung cells" patented technology

The invention discloses a construction method and application of a mouse airway organoid inflammation model, and relates to the technical field of biology. The construction method comprises the following steps: (1) taking a mouse airway tissue, digesting with digestive juice, and centrifuging to obtain a cell suspension; (2) centrifuging the cell suspension, discarding the supernatant, adding matrigel, uniformly mixing, incubating, and culturing by using an airway organoid complete medium; (3) adding a cell recovery solution into the cells cultured in the step (2), incubating, degumming, centrifuging, discarding supernatant, adding matrigel, uniformly mixing, incubating, and adding an airway organ complete culture medium for continuous culture; and (4) adding 1-20 [mu] g / ml of LPS into the airway organoid complete culture medium, and incubating. The AO constructed by the invention covers four main cell types of cilia cells, goblet cells, basal cells and rod-like cells, causes epithelial injury in the AO and expression of AO inflammatory factors, and can provide reference for development of CRD new drugs.

A robust nasal organoid culture system and fully characterized 2D and 3D in vitro produced organoids are provided, as well as methods of using the organoids. The nasal organoids comprise a population of differentiated nasal epithelial cells having cellular heterogeneity and molecular markers similar to mature nasal epithelial tissue. Improved methods for producing two-dimensional and three-dimensional nasal organoids are also provided. These methods generally require a two-phase culture system, i.e., amplification and differentiation culture. The provided nasal organoids are suitable for reproducing nasal tissue complexity, for mimicking respiratory diseases, studying barrier function and drug delivery, high throughput drug screening, and assessing the effects of pathogens and environmental toxins on nasal respiratory epithelium.

The invention provides application of a PDLIM4 gene in preparation of a medicine for inhibiting non-small cell lung cancer proliferation, belongs to the technical field of cancer suppressor gene research, and finds that the PDLIM4 gene is highly expressed in non-small cell lung cancer tumors, and the prognosis of a patient with the highly expressed PDLIM4 gene is good; the proliferation of the non-small cell lung cancer cells overexpressed with the PDLIM4 gene is inhibited, and the proliferation of the non-small cell lung cancer cells knocked down with the PDLIM4 gene is accelerated, so that the reagent capable of realizing the overexpression of the PDLIM4 gene is confirmed to be of great significance for inhibiting the proliferation of the non-small cell lung cancer cells. The overexpression of the PDLIM4 gene in the non-small cell lung cancer cells can enhance the chemotherapy effect of the chemotherapy drug adriamycin, and can be used as a chemotherapy sensitizer. The BRD4 is an upstream regulatory factor for increasing the expression of the PDLIM4 gene in the non-small cell lung cancer, and can be used for promoting the expression of the PDLIM4 gene.

The field of the invention belongs to in vitro assays for producing an in vitro organ culture system, and the use of such an in vitro organ culture system for screening assays in the context of drug screening, patient selection, and personalized medicine. More specifically, the invention relates to an in vitro method of establishing tumor organoids from a cancer tissue sample obtained from a subject, to an in vitro method of providing autologous tumor organoid-activated immune cells, to a method of preparing an in vitro organ culture system for mimicking tumor interaction with an immune system of a subject, as well as to the in vitro organ culture system produced thereby, to a method for determining the responsiveness of a tumor to a treatment with at least one immune- oncologic drug applying said in vitro organ culture system, and a kit-in-parts for preparing an in vitro organ culture system mimicking tumor interaction with an immune system of a subject, as disclosed herein.

The invention belongs to the technical field of biomedical engineering and microfluidics, and particularly discloses a vascularized organ chip and method for large-scale culture, the vascularized organ chip comprises an upper runner layer and a lower culture layer, and the runner layer and the culture layer are fixedly sealed; a liquid storage pipe matched with the flow channel layer is adhered to the upper surface of the flow channel layer; according to the vascularized organ chip, the flow channel layer is used for pouring a culture solution to provide fluid stimulation and nutrients for cells; the three parallel-connected large-scale culture chambers of the culture layer are used for culturing cells, so that blood circulation of a microvascular system in a human body can be simulated, and a microenvironment in tissues can be reproduced more truly; meanwhile, the requirement for culturing a large number of tissue cells through the chip can be met, and the cell yield and the experiment scale can be effectively increased.

The invention provides a small cell lung cancer organoid culture medium and an application method thereof. The small cell lung cancer organoid culture medium is prepared from the following components: 10 to 500 [mu] g / mL of R-spondin-1, 10 to 500 [mu] g / mL of Noggin, 0.01 to 0.2 mg / mL of EGF (Epidermal Growth Factor), 0.05 to 0.2 [mu] g / mL of FGFb (Fibroblast Growth Factor b), 0.1 to 10 [mu] g / mL of FGF10, 0.02 to 0.5 mM of A83-01, 0.5 to 20 mM of Y-27632, 1 to 500 mM of SB202190, 0.1 to 200 mM of Nicotinamide, 0.1 to 500 mM of Insulin, 0.1 to 50 nM of protodioscin, 1 to 100 [mu] All the components are dissolved in a DMEM / F12 culture solution. The small cell lung cancer organoid culture medium provided by the invention is suitable for small cell lung cancer tissue organoid culture after component concentration optimization design, good in stability, high in growth speed and remarkably improved in passage frequency, and can still maintain the same tissue structure and genome characteristics of primary tissues after multiple passages; and an ideal model can be provided for research on the occurrence and development mechanism of small cell lung cancer, detection and research on drug sensitivity and the like.

The invention relates to a pathological pulmonary fibrosis model and a construction method and application thereof. The pathological pulmonary fibrosis model provided by the invention is good in stability and controllability and high in maturity, not only can better simulate the function and structural characteristics of pulmonary alveoli-capillary barrier, but also has the pathological characteristics and immune microenvironment of pulmonary fibrosis lesions, has the advantage of high simulation degree, and has good application prospects in simulation and research of pulmonary fibrosis diseases. Good application prospects are realized in the aspects of drug detection, screening, evaluation, research and development and the like.

The present invention uses ex vivo human airway cultures to assess the human transmissibility and replication competence of influenza and coronavirus strains. By comparing pandemic influenza A subtype H1N1 and highly pathogenic avian influenza H5N1 as reference strains, the transmissibility risk of various viruses was evaluated and categorized. Additionally, an in vitro model evaluated virus-induced impairment of alveolar fluid clearance (AFC) as an indicator of disease severity. The study revealed correlations between bronchus viral replication, human transmission, AFC impairment, and clinical disease severity across different influenza and coronavirus strains.

The invention belongs to the technical field of biology, and particularly relates to a human metapneumovirus mouse adapted strain and application thereof. Compared with a wild strain, the strain has higher susceptibility to animals; the strain can be used for treating BALB / c mice and can cause typical clinical symptoms and even death, for example, after the strain infects the BALB / c mice, the disease symptoms are obvious, the weight of 83% infected mice is severely reduced to 75% or below and reaches the humanity killing standard, and the death rate reaches 50% or above 4 days after the strain infects the BALB / c mice; the strain can be used for constructing a cell model or an animal model of human metapneumovirus infection or diseases caused by the human metapneumovirus infection, and is further used for screening drugs for preventing and / or treating the human metapneumovirus infection or the diseases caused by the human metapneumovirus infection. In addition, the strain can also be used for preparing medicines for preventing and / or treating human metapneumovirus infection or diseases caused by the human metapneumovirus infection, preparing antibodies or antiserum for resisting the human metapneumovirus and the like.

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 invention discloses a mouse lung organ and fibroblast co-culture method, which comprises the following steps of: performing in-vitro mouse lung organ culture on mouse lung tissue primary epithelial stem cell spheres (pneumospheroids) in a Transwell upper chamber, and after the lung organs in the Transwell upper chamber are differentiated, replacing with a lower chamber paved with mouse lung fibroblasts, therefore, a gas-liquid interface co-culture system of the mouse lung organ (upper chamber) and the fibroblast (lower chamber) is formed, and the hierarchical structure, morphological characteristics and regeneration function of the upper chamber organ and the phenotype, function and related signal path activation level of the lower chamber fibroblast can be dynamically researched.

The invention discloses a lung cancer PDL1 immunotherapy drug-resistant cell strain LLC-R and application thereof, and relates to the technical field of cell biology. The cell strain LLC-R is preserved in the China Center for Type Culture Collection on November 6, 2024, the preservation address is Wuhan University, Wuhan, China, and the preservation number is CCTCC NO: C2024374. The invention provides a reliable experimental model for researching the drug resistance mechanism of lung cancer on PDL1 immunotherapy and related signal channels, so that technicians in the field can more deeply understand the reaction of lung cancer cells on PDL1 immunotherapy, which is helpful for exploring new biomarkers and researching and developing new combined treatment strategies, and has a wide application prospect. The treatment effect is improved; the occurrence of drug resistance is reduced. The invention has important scientific and clinical significance for deeply understanding the lung cancer PDL1 immunotherapy drug resistance and improving the clinical treatment effect of patients.

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 present disclosure relates to a method for preparing a respiratory organoid by proliferating and differentiating respiratory epithelial cells. Unlike the conventional respiratory organoids in which epithelial cells existing in the apical region of the respiratory organ are generated on the interior of the organoid, the respiratory organoid of the present disclosure is shown in a form in which ciliated structures, which are epithelial cells, are located in the outer layer of the organoid, secretory cells are present on the interior, and basal stem cells existing at the base of respiratory tissue are also present on the interior of the organoid. It has a form similar to an actual respiratory tissue, and it was observed that the epithelial cells exhibited normal function, such as motility of ciliated cells. Therefore, when an organoid is prepared using the preparation method of the present disclosure, a respiratory organoid that can well mimic actual tissue can be prepared.

The invention provides a co-culture system of human alveolar epithelial cells and endothelial cells, which is used for researching interaction in an epithelial-endothelial barrier in lungs. It is indicated that infection with the highly pathogenic influenza A virus A / HK / 483 / 97 (H5N1) can significantly reduce alveolar fluid clearance (AFC) and increase protein permeability (APP). Administration of therapeutic extracellular vesicles (EV) after infection effectively restores AFC and APP, reduces viral replication, and enhances expression of antiviral genes. According to the invention, the treatment potential of EV is highlighted, and a valuable tool is provided for researching acute lung injury and improving a lung drug delivery technology.

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 invention relates to a lung in-vitro model and a construction method and application thereof. The lung in-vitro model provided by the invention is good in stability and controllability and high in maturity, can better simulate functions and structural characteristics of pulmonary alveoli-capillary barriers, has the advantage of high simulation, and has a good application prospect in the aspects of disease simulation, drug detection, screening, evaluation, research and development and the like.

The invention discloses a lung cancer organoid culture medium, a culture medium system and a culture method thereof, the culture medium system comprises an initial culture medium, an amplification culture medium and a differential culture medium, and the initial culture medium comprises an Advanced DMEM / F12 basal culture medium, Glutamax, nicotinamide, R-spondin-1, rhNoggin, FGF-7, FGF-10, B27, Y-27632, A83-01, SB202190 and prostaglandin E2. Hydrocortisone and Forskolin are added into the amplification culture medium; hydrocortisone and dexamethasone are added into the differential culture medium, and the culture medium system is adopted for lung cancer organoid culture. Cells obtained from lung cancer tissues are subjected to primary lung cancer organoid culture in the initial culture medium; then, transferring the obtained lung cancer organoid cells into an amplification culture medium for amplification culture; and then, transferring the lung cancer cells into a differential culture medium for differential culture to obtain the lung cancer organoid.

The invention relates to a pulmonary fibrosis model and a construction method and application thereof. The pulmonary fibrosis model provided by the invention is good in stability and controllability and high in maturity, can better simulate functions and structural characteristics of pulmonary alveoli-capillary barriers, has the advantage of high simulation, and has a good application prospect in the aspects of disease simulation, drug detection, screening, evaluation, research and development and the like.

The invention provides an organoid based on a micro-fluidic chip as well as a preparation method and application of the organoid. The organoid based on the micro-fluidic chip comprises the micro-fluidic chip and organoid microspheres, the micro-fluidic chip comprises an inner cavity, the inner cavity comprises capture card embedding structures which are alternately arranged in a quincunx shape, and each capture card embedding structure comprises a capture card embedding micro-column and a liquid guide channel; the organ-like microspheres are hydrogel microspheres loaded with cells. Through a 3D biological printing technology, batch organoid microspheres uniform in structure and stable in function can be rapidly prepared, and the situation that the experiment progress is slow due to the difference between models in a drug screening experiment is improved; the organ-like model with the characteristics simulates the flowing of human blood and better reproduces a human microenvironment; the capture rate of organoid microspheres is increased, the space utilization rate in the chip is improved, the usage amount of reagents and drugs in a drug screening experiment is reduced, and the scientific research cost is reduced; the problem that the micro-fluidic chip type organs are difficult to visualize and traceable can be solved.

The invention discloses an application of a BAG6 gene in regulation of influenza virus replication. The invention provides any one of the following applications of the substance for inhibiting expression of the BAG6 gene or the encoded protein thereof: 1) preparing a product for expanding propagation of influenza virus; 2) preparing a product for promoting influenza virus replication; and 3) preparing a product for improving the expression quantity of the influenza virus related protein. Experiments prove that the function of the BAG6 gene encoding protein is lost, and the BAG6 gene encoding protein can be used for researching the cell biological function of the BAG6 protein and the action and mechanism in other viruses; in addition, the strain has the potential of being applied to influenza vaccine production. Besides, overexpression is carried out on the BAG6 protein, it is found that the BAG6 protein can inhibit the replication titer of the influenza virus and the expression of the protein, and the effect of the BAG6 protein in influenza virus replication is explored from the two aspects of knockout and overexpression.

The invention relates to the technical field of biological medicine, in particular to a drug screening model construction method based on airway organs, a model and application. The construction method of the drug screening model of the airway organoid comprises the steps of obtaining 2D airway epithelial cells, culturing the airway organoid, establishing an airway organoid inflammation model and / or establishing an airway organoid mucus secretion / goblet cell metaplasia model. Wherein the model is established according to the culture mode of the airway organ in cooperation with a specific stimulant for proliferation and differentiation. Compared with a traditional animal model, the organ-like model is low in cost and short in construction period, can better simulate the pathological process, can realize airway organ-like culture and establishment of an inflammation model and a mucus secretion / goblet cell metaplasia model at the same time, and has a good application prospect in drug detection of asthma and COPD diseases.

The invention relates to the field of biology, in particular to a fluorescence and bar code pedigree tracking alveolar organ and a construction method and application thereof. The invention particularly aims at the problems of specific sorting of AT2 cells, culture of organoids and pedigree retracking in the disease process, and is used for simulating and researching the in-vitro pathological process of chronic obstructive pulmonary disease (COPD). The alveolar organ with fluorescence and bar code pedigree tracking is constructed and can track the fate transformation of alveolar type II cells (AT2) in a disease model, and the cell pedigree is further decoded, so that a new research basis is provided for early diagnosis and treatment of COPD (chronic obstructive pulmonary disease).

The invention discloses an application of a ZDHHC11 gene in preparation of a medicine for treating non-small cell lung cancer, and relates to the technical field of biological medicines, and the research finds that the ZDHHC11 protein level is increased by over-expressing the ZDHHC11 gene in NSCLC cells, the cell viability, migration ability and multiplication ability can be inhibited, and cell apoptosis can be promoted. The activity of NSCLC cells can be increased by knocking out the ZDHHC11 gene, and the result shows that the ZDHHC11 gene plays an important role in controlling the development process of the non-small cell lung cancer, provides a new action target for treating the NSCLC, and has potential application value in preparing the medicine for treating the non-small cell lung cancer.

A method for rapidly and efficiently inducing multiple human airway epithelium in vitro models for a long time relates to the technical field of organoid culture.The method comprises the steps that firstly, a clinical lung tissue sample is induced and differentiated to form a cilia inward lung airway organoid, the cilia inward lung airway organoid is digested into single cells, and multiplication culture is conducted through a basal cell culture medium; the obtained basal cells are subjected to induced differentiation to form cilia outward lung airway organs and a gas-liquid interface culture model. The two types of models can successfully simulate the functional cellular morphology of the human airway epithelium in vitro, can be used as experimental models for disease modeling, drug screening, toxicity testing and the like, and can simulate biological models of the human airway structure and function. The method can solve the problems that clinical samples are difficult to obtain and primary lung epithelial cells have limited passage times in the construction process of lung airway organs, and realizes long-term rapid and efficient induction of various human airway epithelium in-vitro models.

The invention discloses a method for directly inducing generation of organoids from IPS and application, and belongs to the field of cell engineering. The invention provides a method for directly and rapidly generating a target organ based on human IPS, and the lung organ, the placenta organ and the blood vessel organ are obtained. According to the method, the organoid represented by three germ layers is induced, so that the method is suitable for constructing various organoid. Compared with the prior art, the method has the advantages that step-by-step induction is not needed, one-step construction is successful, the operation is simple, the construction time is greatly shortened, the process is repeatable, long-term stable culture and passage are realized under in-vitro conditions, and the freezing survival rate is high; the method has the advantages that the method is simple in operation, the cultured organoid is good in homogeneity, RNA-seq results show that high expression of the blood vessel related genes appears in the organoid, and the maximum length of the organoid can reach 400 micrometers, so that the culture method can spontaneously induce expression of part of the blood vessel related genes to support growth of the organoid. The invention provides an important tool for researching pathogenesis of diseases and subsequent drug development.

The invention relates to a lung cancer organoid and peripheral blood source immune cell co-culture model and a construction method thereof. Specifically, the invention provides a construction method of a macrophage and tumor organoid co-culture model for evaluating anti-tumor activity, monocytes of autologous or allogeneic peripheral blood of a patient are induced and differentiated into high-purity macrophages in vitro, and the high-purity macrophages and lung cancer tumor organoid are subjected to three-dimensional co-culture in matrigel; the model aims at highly reducing a core interaction network of tumor cells and macrophages in TME; the dynamic change of the polarization state of the macrophage is simulated and observed; the problem that an organ-like model is incomplete due to immune component deficiency or spatial positioning distortion of an existing model is effectively solved, an experimental platform closer to the physiological state is provided for tumor immune microenvironment research, and therefore development of the tumor immune treatment field is promoted.

The invention discloses a lung airway organoid culture system with a trachea branch morphological structure as well as application and a culture method of the lung airway organoid culture system, and relates to the technical field of lung airway organoid culture. The hydrogel culture system is prepared from I-type rat tail collagen, gelatin, hyaluronic acid, laminin, low-melting agarose, chondroitin sulfate, fibronectin, elastin, epidermal growth factors, fibronectin L and transglutaminase; the induction culture medium is prepared from a 1XB27 supplement, a fiber growth factor, glutamine, glutathione, A8301, SB202190, IBMX, 8-Br-cAMP, FGF7, FGF10, Shh and ascorbic acid. The culture system is clear in component, low in price and cost and simple and convenient in preparation method, more importantly, the lung airway organoid which is closer to the physiological state can be efficiently induced to be formed within a short time, and the generated organoid has a branch form similar to an in-vivo trachea and contains various types of functional airway epithelial cells at the same time; and a more preferable scheme is provided for constructing an airway organ with an airway branch form based on the adult lung stem cells.

The invention discloses a culture system for generating a pulmonary alveolar organ with an alveolar vesicle form based on adult lung stem cells and application of the culture system. Belongs to the technical field of lung organ three-dimensional culture. By utilizing the culture system, the adult lung stem cells can be efficiently and stably induced to be differentiated to generate the pulmonary alveolar organ with a near-physiological state pulmonary alveolar sac structure. Compared with a Matrigel system dependent on pulmonary alveolar organ induction culture in the prior art, the culture system established by the invention has the following outstanding advantages: 1) the generated pulmonary alveolar organ has a near-physiological state pulmonary alveolar sac structure and also contains a functional pulmonary alveolar epithelial cell system; 2) the proliferation and differentiation speed of the adult lung stem cells in the culture system is high, the generation period of the alveolar organs is short and only needs 2-3 weeks, and the generation period of the alveolar organs based on a Matrigel system needs 4-12 weeks; and 3) the culture system is clear in composition, low in price and cost and simple in preparation method.

The invention belongs to the technical field of organoid culture, and particularly relates to a pulmonary alveolar organoid with an endogenous vascular network and a construction method of the pulmonary alveolar organoid. Aiming at the problems that mesoderm and endoderm are difficult to induce at the same time, the operation process is long, the difficulty is high and the like when a vascularized alveolar organ is constructed by an existing method, the invention provides the construction method of the alveolar organ with the endogenous vascular network. The method comprises the following steps: simultaneously and directionally inducing and differentiating human induced pluripotent stem cells (hiPSC) into fixed endoderm and mesoderm, anterior intestine endoderm, abdominal side anterior intestine endoderm and lung progenitor cells by adopting a one-step method, and promoting endothelial cell proliferation in an endogenous induced differentiation process through a specific growth factor combination. A functional vascular network is formed; and vascularized alveolar organs are induced to be generated. According to the invention, a qi-blood barrier structure in which vascular endothelial cells and pulmonary epithelial cells are tightly connected is successfully reproduced, the gas exchange function of lung tissues can be simulated, and a new way is provided for construction of vascularized alveolar organs.