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A suspension technique for efficient large-scale cancer organoid culturing and perturbation screens.Scientific Reports Apr 2022Organoid cell culture methodologies are enabling the generation of cell models from healthy and diseased tissue. Patient-derived cancer organoids that recapitulate the...
Organoid cell culture methodologies are enabling the generation of cell models from healthy and diseased tissue. Patient-derived cancer organoids that recapitulate the genetic and histopathological diversity of patient tumours are being systematically generated, providing an opportunity to investigate new cancer biology and therapeutic approaches. The use of organoid cultures for many applications, including genetic and chemical perturbation screens, is limited due to the technical demands and cost associated with their handling and propagation. Here we report and benchmark a suspension culture technique for cancer organoids which allows for the expansion of models to tens of millions of cells with increased efficiency in comparison to standard organoid culturing protocols. Using whole-genome DNA and RNA sequencing analyses, as well as medium-throughput drug sensitivity testing and genome-wide CRISPR-Cas9 screening, we demonstrate that cancer organoids grown as a suspension culture are genetically and phenotypically similar to their counterparts grown in standard conditions. This culture technique simplifies organoid cell culture and extends the range of organoid applications, including for routine use in large-scale perturbation screens.
Topics: Cell Culture Techniques; DNA; Humans; Neoplasms; Organoids
PubMed: 35368031
DOI: 10.1038/s41598-022-09508-y -
Scientific Reports Jun 2021Microphysiological organ-on-chip models offer the potential to improve the prediction of drug safety and efficacy through recapitulation of human physiological...
Microphysiological organ-on-chip models offer the potential to improve the prediction of drug safety and efficacy through recapitulation of human physiological responses. The importance of including multiple cell types within tissue models has been well documented. However, the study of cell interactions in vitro can be limited by complexity of the tissue model and throughput of current culture systems. Here, we describe the development of a co-culture microvascular model and relevant assays in a high-throughput thermoplastic organ-on-chip platform, PREDICT96. The system consists of 96 arrayed bilayer microfluidic devices containing retinal microvascular endothelial cells and pericytes cultured on opposing sides of a microporous membrane. Compatibility of the PREDICT96 platform with a variety of quantifiable and scalable assays, including macromolecular permeability, image-based screening, Luminex, and qPCR, is demonstrated. In addition, the bilayer design of the devices allows for channel- or cell type-specific readouts, such as cytokine profiles and gene expression. The microvascular model was responsive to perturbations including barrier disruption, inflammatory stimulation, and fluid shear stress, and our results corroborated the improved robustness of co-culture over endothelial mono-cultures. We anticipate the PREDICT96 platform and adapted assays will be suitable for other complex tissues, including applications to disease models and drug discovery.
Topics: Cell Communication; Cell Membrane Permeability; Cells, Cultured; Coculture Techniques; Dermis; Endothelium, Vascular; Humans; Microfluidic Analytical Techniques; Pericytes; Retina
PubMed: 34108507
DOI: 10.1038/s41598-021-90833-z -
Stem Cell Research Oct 2019Adipose-derived Stem Cells (ASCs) present great potential for reconstructive procedures. Currently, isolation by enzyme digestion and culturing using xenogenic...
BACKGROUND
Adipose-derived Stem Cells (ASCs) present great potential for reconstructive procedures. Currently, isolation by enzyme digestion and culturing using xenogenic substances remain the gold standard, impairing clinical use.
METHODS
Abdominal lipo-aspirate and blood samples were obtained from healthy patients. A novel mechanical isolation method for ASCs was compared to (the standard) collagenase digestion. ASCs are examined by flowcytometry and multilineage differentiation assays. Cell cultures were performed without xenogenic or toxic substances, using autologous plasma extracted from peripheral blood. After eGFP-transfection, an in vivo differentiation assay was performed.
RESULTS
Mechanical isolation is more successful in isolating CD34/CD31/CD45/CD13/CD73/CD146 ASCs from lipo-aspirate than isolation via collagenase digestion (p <0 .05). ASCs display multilineage differentiation potential in vitro. Autologous plasma is a valid additive for ASCs culturing. eGFP-ASCs, retrieved after 3 months in vivo, differentiated in adipocytes and endothelial cells.
CONCLUSION
A practical method for human ASC isolation and culturing from abdominal lipo-aspirate, without the addition of xenogenic substances, is described. The mechanical protocol is more successful than the current gold standard protocol of enzyme digestion. These results are important in the translation of laboratory-based cell cultures to clinical reconstructive and aesthetic applications.
Topics: Adipose Tissue; Animals; Antigens, CD34; Cell Differentiation; Cell Separation; Cells, Cultured; Culture Media; Female; Humans; Mesenchymal Stem Cells; Mice; Mice, Nude
PubMed: 31421383
DOI: 10.1016/j.scr.2019.101532 -
Tissue Engineering. Part C, Methods May 2017Rapid prototyping and fabrication of elastomeric molds for sterile culture of engineered tissues allow for the development of tissue geometries that can be tailored to...
Rapid prototyping and fabrication of elastomeric molds for sterile culture of engineered tissues allow for the development of tissue geometries that can be tailored to different in vitro applications and customized as implantable scaffolds for regenerative medicine. Commercially available molds offer minimal capabilities for adaptation to unique conditions or applications versus those for which they are specifically designed. Here we describe a replica molding method for the design and fabrication of poly(dimethylsiloxane) (PDMS) molds from laser-etched acrylic negative masters with ∼0.2 mm resolution. Examples of the variety of mold shapes, sizes, and patterns obtained from laser-etched designs are provided. We use the patterned PDMS molds for producing and culturing engineered cardiac tissues with cardiomyocytes derived from human-induced pluripotent stem cells. We demonstrate that tight control over tissue morphology and anisotropy results in modulation of cell alignment and tissue-level conduction properties, including the appearance and elimination of reentrant arrhythmias, or circular electrical activation patterns. Techniques for handling engineered cardiac tissues during implantation in vivo in a rat model of myocardial infarction have been developed and are presented herein to facilitate development and adoption of surgical techniques for use with hydrogel-based engineered tissues. In summary, the method presented herein for engineered tissue mold generation is straightforward and low cost, enabling rapid design iteration and adaptation to a variety of applications in tissue engineering. Furthermore, the burden of equipment and expertise is low, allowing the technique to be accessible to all.
Topics: Animals; Animals, Newborn; Cells, Cultured; Elasticity; Humans; Hydrogels; Lasers; Male; Myocardial Infarction; Myocytes, Cardiac; Rats; Rats, Sprague-Dawley; Tissue Engineering
PubMed: 28457187
DOI: 10.1089/ten.TEC.2017.0068 -
Journal of Cellular and Molecular... 2004The reconstruction of skeletal muscle tissue either lost by traumatic injury or tumor ablation or functional damage due to myopathies is hampered by the lack of... (Review)
Review
The reconstruction of skeletal muscle tissue either lost by traumatic injury or tumor ablation or functional damage due to myopathies is hampered by the lack of availability of functional substitution of this native tissue. Until now, only few alternatives exist to provide functional restoration of damaged muscle tissues. Loss of muscle mass and their function can surgically managed in part using a variety of muscle transplantation or transposition techniques. These techniques represent a limited degree of success in attempts to restore the normal functioning, however they are not perfect solutions. A new alternative approach to addressing difficult tissue reconstruction is to engineer new tissues. Although those tissue engineering techniques attempting regeneration of human tissues and organs have recently entered into clinical practice, the engineering of skeletal muscle tissue ist still a scientific challenge. This article reviews some of the recent findings resulting from tissue engineering science related to the attempt of creation and regeneration of functional skeletal muscle tissue.
Topics: Animals; Biomedical Engineering; Cell Differentiation; Cell Transplantation; Cells, Cultured; Culture Techniques; Female; Humans; Male; Models, Biological; Muscle Fibers, Skeletal; Muscle, Skeletal; Muscles; Rats; Tissue Engineering
PubMed: 15601570
DOI: 10.1111/j.1582-4934.2004.tb00466.x -
Experimental Physiology May 2019What is the central question of this study? Do circulating factors mediate exercise-induced effects on adipose tissue GLUT4 expression? What is the main finding and its...
NEW FINDINGS
What is the central question of this study? Do circulating factors mediate exercise-induced effects on adipose tissue GLUT4 expression? What is the main finding and its importance? Serum (10%) obtained from human volunteers immediately after a single exercise bout increased GLUT4 protein levels in human adipocytes in culture. This result suggests that circulating factors might mediate the effects of exercise on adipose tissue GLUT4 and prompts further effort to identify the specific factor(s) and tissue(s) of origin.
ABSTRACT
In this study, we tested the hypothesis that circulating factors generated during exercise increase adipose tissue GLUT4 expression. Serum was obtained from eight healthy subjects before and after 60 min of cycling exercise, and primary adipocytes were cultured from stromal vascular fractions that were isolated from subcutaneous abdominal adipose tissue samples from one healthy, male volunteer. A 48 h exposure of human primary adipocytes to 10% serum obtained after exercise increased GLUT4 protein expression, on average, by 12% compared with exposure to 10% serum obtained at rest, before exercise. GLUT4 mRNA levels were increased after 12 h of exposure to exercise serum but were unchanged after 6 and 24 h of exposure. Our results suggest that circulating factors might mediate the effects of exercise on adipose tissue GLUT4 expression and encourage further efforts to identify the potential factor(s), tissue(s) of origin and physiological relevance.
Topics: Adipocytes; Bicycling; Exercise; Gene Expression Regulation; Glucose Transporter Type 4; Humans; Male; Primary Cell Culture; RNA, Messenger; Subcutaneous Fat; Young Adult
PubMed: 30758088
DOI: 10.1113/EP087495 -
Stem Cell Research & Therapy Aug 2022Body-on-a-chip (BoC) platforms are established from multiple organs-on-chips (OoCs) to recapitulate the interactions between different tissues. Recently,...
Body-on-a-chip (BoC) platforms are established from multiple organs-on-chips (OoCs) to recapitulate the interactions between different tissues. Recently, Vunjak-Novakovic and colleagues reported the creation of a BoC system comprising four fluidically linked OoCs. Herein, the major innovations in their BoC system are discussed, followed by our future perspectives on enhancing the physiological relevance and scalability of BoCs for applications in studying disease mechanisms, testing potential therapeutics, and developing personalized medicine.
Topics: Human Body; Humans; Lab-On-A-Chip Devices; Precision Medicine
PubMed: 35987699
DOI: 10.1186/s13287-022-03130-5 -
Journal of Ethnopharmacology Jan 2021Cereus hildmannianus (K.) Schum. (syn. C. peruvianus) is a native medicinal plant in the Neotropical region. C. hildmannianus cladodes extracts are used in folk medicine... (Review)
Review
ETHNOPHARMACOLOGICAL RELEVANCE
Cereus hildmannianus (K.) Schum. (syn. C. peruvianus) is a native medicinal plant in the Neotropical region. C. hildmannianus cladodes extracts are used in folk medicine for weight loss, reducing cholesterol, low-density lipoprotein (LDL) levels, as diuretic and cardiotonic, and to treat various diseases, including pulmonary disorders, rheumatism, and in topical treatment for wounds and lithiasis. Fruits and flowers of C. hildmmanianus have high nutritional value.
AIM OF THE REVIEW
In this review, previous reports on C. hildmannianus (syn. C. peruvianus) concerning its botanical description, geographical distribution, ethnomedicinal use, phytochemistry, in vitro and in vivo pharmacological properties, food benefits and plant biotechnology were summarized.
MATERIALS AND METHODS
Scientific search engines, including ScienceDirect, Capes Journals Portal, Google Scholar, PubMed, Scielo, and Scifinder, were consulted to gather data on C. hildmannianus. The present review is an up-to-date and comprehensive analysis of phytochemical compounds, ethnomedicinal uses, phytopharmacological activities, and food value of cladodes, flowers, fruits, seeds, and tissue culture from C. hildmannianus.
RESULTS
In traditional medicine, C.hildmannianus is used to treat various diseases, including pulmonary disorders, rheumatism, and in topical treatment for wounds and lithiasis. The extracts from the cladodes of C. hildmannianus exhibited numerous in vitro and in vivo pharmacological activities such as gastroprotective, antioxidant, antifungal, ovicidal, hemagglutinating and slimming, and anticancer activity. These extracts' substances belong to different classes of chemical compounds such as fatty acids, polysaccharides, terpenes, alkaloids, phenolic acids, and flavonoids CONCLUSIONS: Ethnomedicinal surveys have indicated the use of C. hildmannianus, an important medicinal plant in South America, for the treatment of various diseases. The ethnobotanical, phytochemical, pharmacological, and ethnomedicinal properties from various morphological parts of the plant of C. hildmannianus were highlighted in this review, which provides information for future studies, commercial exploration and reveals that this plant has a huge potential for pharmaceutical and nutraceutical applications.
Topics: Animals; Antifungal Agents; Antioxidants; Cactaceae; Ethnopharmacology; Humans; Medicine, Traditional; Phytochemicals; Plant Extracts; Plants, Medicinal; South America
PubMed: 32898627
DOI: 10.1016/j.jep.2020.113339 -
Alternative Therapies in Health and... Sep 2023Mammary gland hyperplasia is a common gynecological disease, which seriously affects the patient's physical and mental health. Therapeutic strategies to treat the... (Review)
Review
OBJECTIVE
Mammary gland hyperplasia is a common gynecological disease, which seriously affects the patient's physical and mental health. Therapeutic strategies to treat the disease include endocrine therapy and surgery. Compared to Western treatment, traditional Chinese medicine prescription shows its superiority in treatment. The purpose of this review was to provide a reference for the determination of the pathogenesis, treatment principles, and treatment methods of mammary gland hyperplasia.
METHOD
This article comprehensively reviewed the records on mammary gland hyperplasia in ancient Chinese medical literature.
RESULTS
The present review discussed the disease and summarizes the information on mammary gland hyperplasia, including the disease name, the traditional Chinese medicine analysis, etiology, pathogenesis, treatment methods, prognosis, and nursing care.
CONCLUSION
We clearly described the research history of mammary gland hyperplasia, and the analysis and treatment of this disease by physicians in past dynasties. This information will help modern physicians to fully understand the disease development and treatment process.
Topics: Humans; Hyperplasia; Mammary Glands, Human; Medicine, Chinese Traditional
PubMed: 37235489
DOI: No ID Found -
Frontiers in Endocrinology 2022
Topics: Bone and Bones; Drugs, Chinese Herbal; Homeostasis; Medicine, Chinese Traditional
PubMed: 35873007
DOI: 10.3389/fendo.2022.969729