405results about "Respiratory/lung cells" patented technology

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 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 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 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 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 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 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.

The invention discloses a construction method and application of a human-derived far-end airway organ-like COPD model, the constructed human-derived far-end airway organ-like COPD model is highly similar to COPD far-end airway tissue on the transcriptional level, and the pathological characteristics of COPD are well restored on the protein expression level and cilia function level. Therefore, the human-derived far-end airway organ-like COPD model constructed in vitro can simulate the COPD lesion of the far-end airway, and an available model is provided for researching the COPD pathological mechanism of the far-end airway. In addition, the requirements on time, temperature and quality inspection indexes of CSE extraction in operation are clear, and the batch-to-batch extraction quality and experiment repeatability of CSE can be well ensured. The cilia function is used as a detection index, the improvement effect of the medicine on the lung function can be better reflected, and data support of the function level can be provided for COPD medicine research.

The invention belongs to the technical field of biological medicine manufacturing, and particularly provides a preparation method of an engineered exosome capable of loading mitochondria for treating pulmonary fibrosis and the engineered exosome. The method comprises the following steps: constructing fusion plasmids for expressing mitochondrial targeting peptide fragment-therapeutic protein genes, dissolving the fusion plasmids in non-enzyme ultrapure water, adding the fusion plasmids into a lipid nano-preparation, and fully mixing to obtain a nano-preparation containing the therapeutic protein genes; the nano preparation is delivered into alveolar epithelial cells and is efficiently transfected, so that the homeostatic balance of the injured alveolar epithelial cells in the development of pulmonary fibrosis is effectively regulated and controlled, the secretion of an exosome with a therapeutic effect is promoted, and the engineered exosome is further obtained through mannose modification. The exosome can effectively load mitochondria and sequentially and accurately target macrophage mitochondria on the basis of a mannose target head and a mitochondrial targeting peptide fragment, so that macrophage mitochondrial homeostatic balance is effectively regulated and controlled, the purpose of reverse treatment of pulmonary fibrosis is achieved, and the exosome has high clinical application value.

The invention provides a lung stem cell peptide for treating pulmonary nodule as well as a preparation method and application thereof, and belongs to the technical field of biological pharmacy. The method comprises the following steps: performing induced differentiation on iPSC to obtain lung stem cells; lung stem cells are cultured and centrifuged, supernatant is collected, and a lung stem cell crude solution is obtained; mixing the lung stem cell crude liquid with protease, performing enzymolysis, centrifuging, concentrating, and performing spray drying to obtain the lung stem cell peptide for treating pulmonary nodule. By optimizing the composition of each culture medium, the iPSC is induced and differentiated into the lung stem cells, then the lung stem cell peptide is extracted and separated from the lung stem cells, and the extracted lung stem cell peptide migrates to the lung in a vein reinfusion mode, so that the generation of lung tumor nodules can be effectively inhibited, the number of the lung tumor nodules is greatly reduced, and the lung tumor nodules can be effectively inhibited. Toxic and side effects and adverse reactions are avoided.

The present disclosure provides methods of making tissue-derived epithelial organoids. In particular, the present disclosure provides a tissue-derived epithelial organoid embedded in a hydrogel suspended in a culture medium. The present disclosure further provides methods of using such organoids.

A method of modifying a surface of a three-dimensional (3D) article includes immersing at least one part of the 3D article in a buffered solution of functionalized peptides, allowing reaction between the functionalized peptides and reactive groups on the surface of the 3D article; and washing the at least one part of the 3D article to remove unreacted functionalized peptides. The surface-modified 3D article includes a plurality of peptides covalently bonded to the surface of the 3D article via a cysteine bridge. The surface- modified 3D article can be used as a scaffold for the formation of biological tissue or bodily implants.

This invention provides a lung stem cell peptide for treating pulmonary nodules, its preparation method, and its application, belonging to the field of biopharmaceutical technology. The invention involves inducing iPSCs to differentiate into lung stem cells; culturing the lung stem cells, centrifuging, collecting the supernatant to obtain crude lung stem cell solution; mixing the crude lung stem cell solution with a protease for enzymatic hydrolysis, centrifuging, concentrating, and spray-drying to obtain the lung stem cell peptide for treating pulmonary nodules. This invention optimizes the composition of each culture medium to induce iPSCs to differentiate into lung stem cells, then extracts and separates lung stem cell peptides from these cells. The extracted lung stem cell peptides are intravenously reinfused into the lungs, effectively inhibiting the formation of pulmonary tumor nodules, significantly reducing the number of pulmonary tumor nodules, and without toxic side effects or adverse reactions.

A method of testing a test substance in an in vitro model of a human tracheobronchial respiratory tract, includes: providing the in vitro model including a cell culture including airway epithelial cells (AECs), a basolateral compartment below the AECs and an apical compartment above the AECs, wherein the AECs form a barrier between the basolateral AND apical compartments; adding the test substance to the apical and / or basolateral compartment; adding a tracer to the basolateral compartment, which is fluorescent, has a molecular weight within 5 kDa of human albumin and is added before, during or after adding the test substance; incubating the cell culture system in a presence of the tracer; collecting at least one sample from the apical compartment; and measuring a fluorescence thereof to determine an effect of the test substance on the AECs. A kit is also disclosed.

The invention discloses a culture medium system for inducing airway stem cells to differentiate into airway epithelial cells based on ALI and application of the culture medium system. Belongs to the technical field of ALI culture. By means of the culture medium system, the airway stem cells can be efficiently and stably induced to be differentiated into multiple types of airway epithelial cells in the ALI culture process, and particularly, the generated cilia cells are close to the physiological state in vivo. Compared with an imported culture medium, the culture medium has the advantages of definite components, high batch stability, low cost and simple preparation method; the generated airway epithelial cells have a plurality of cell lineages, can well simulate airway epithelium, and show forms and functional characteristics similar to those of an in-vivo airway. According to the system, a respiratory tract epithelium-induced ALI culture system is remarkably improved, the research cost is reduced, a preferable scheme is provided for scholars engaged in studying physiological and pathological characteristics of respiratory tract epithelial cells based on ALI, and researchers are helped to establish a stable repeatable airway model.

Disclosed herein are lung progenitor cells that are in the form of lung organoids and methods of generating the lung progenitor cells. The lung organoids can be an important resource for treating lung disorders and injuries, studies in human lung regeneration, disease modeling, and drug target identification and validation.

Methods and systems are described for a high-level approach to organ generation by self-assembly using bioprinting, overcoming longstanding challenges in replicating the intricate architecture and function of complex organs. In some embodiments, these methods are used to produce lung tissues and microtissues, demonstrating how mini-lung constructs can be produced on a small scale through specialized processes and media. In such embodiments, methods and systems for generating bioprinted lung tissues and microtissues are provided, as are tissues made by such methods. The tissues find use in implantation, drug discovery, personalized medicine, and other applications. Methods for the large-scale manufacturing of lung epithelial stem cells are also provided.

The application discloses a combined drug for treating lung cancer. It is found for the first time that lomethixol can be used alone or in combination with other drugs to have a significant anticancer effect on NSCLC patients and NSCLC patients resistant to osimertinib, and in addition, it is found that lomethixol is a potential autophagy inducer and can activate cell autophagy. The application provides a new clinical treatment method for NSCLC patients and NSCLC patients resistant to osimertinib, and has a wide application prospect.

The invention discloses an inflammatory pulmonary epithelium injury treatment evaluation method based on alveolar organs, and relates to the technical field of medical biology. The invention mainly aims at alveolar epithelium injury and repair in inflammatory lung injury, and establishes an organ-like model for simulating alveolar epithelium injury. By using the model, the efficacy of related drugs on alveolar epithelium injury repair can be detected in a targeted manner, and an efficient drug screening and evaluation platform and method are provided for pulmonary epithelium repair in inflammatory lung injury. According to the technical scheme, the method comprises the steps that mouse alveolar epithelial stem cells are separated through magnetic bead sorting, an alveolar organoid inflammatory lung injury model is established through LPS exposure, alveolar organoid is treated through drugs, the organoid generation number, the organoid formation rate, the epithelial cell proliferation rate and the AT2-AT1 differentiation efficiency are detected, and the drug treatment effect is evaluated.

The purpose of the present invention is to provide a novel method for culturing tissue-derived cells. The method uses a cell culture method including a step for co-culturing cells derived from the skin and at least one type of cell selected from among cells derived from the intestine, cells derived from the heart, cells derived from the brain, cells derived from the stomach, cells derived from the embryonic membrane, cells derived from the kidney, and cells derived from the lung. And culturing the skin-derived cells.

Provided herein are artificial lung organoids. The artificial lung organoids may include an epithelial cell layer comprising mammalian lung epithelial cells, a stromal cell layer comprising mammalian lung fibroblast cells and an endothelial cell layer comprising mammalian endothelial cells. The artificial lung organoids may optionally include a porous membrane between said epithelial cell layer and said stromal cell layer and / or between said stromal cell layer and said endothelial lung cell layer.