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Developmental Biology Dec 2016The liver and pancreas are critical organs maintaining whole body metabolism. Historically, the expansion of adult-derived cells from these organs in vitro has proven... (Review)
Review
The liver and pancreas are critical organs maintaining whole body metabolism. Historically, the expansion of adult-derived cells from these organs in vitro has proven challenging and this in turn has hampered studies of liver and pancreas stem cell biology, as well as being a roadblock to disease modelling and cell replacement therapies for pathologies in these organs. Recently, defined culture conditions have been described which allow the in vitro culture and manipulation of adult-derived liver and pancreatic material. Here we review these systems and assess their physiological relevance, as well as their potential utility in biomedicine.
Topics: Animals; Cell- and Tissue-Based Therapy; Genetic Therapy; Humans; Liver; Liver Regeneration; Models, Biological; Organ Culture Techniques; Organogenesis; Organoids; Pancreas; Signal Transduction; Stem Cells
PubMed: 27364469
DOI: 10.1016/j.ydbio.2016.06.039 -
Bosnian Journal of Basic Medical... Jul 2022Although DNA methylation epigenetically regulates development, data on global DNA methylation during development of limb buds (LBs) are scarce. We aimed to investigate...
Although DNA methylation epigenetically regulates development, data on global DNA methylation during development of limb buds (LBs) are scarce. We aimed to investigate the global DNA methylation developmental dynamics in rat LBs cultivated in a serum-supplemented (SS) and in chemically defined serum- and protein-free (SF) three-dimensional organ culture. Fischer rat front- and hind-LBs at 13th and 14th gestation days (GD) were cultivated at the air-liquid interface in Eagle's Minimal Essential Medium (MEM) or MEM with 50% rat serum for 14 days, as SF and SS conditions, respectively. The methylation of repetitive DNA sequences (SINE rat ID elements) was assessed by pyrosequencing. Development was evaluated by light microscopy and extracellular matrix glycosaminoglycans staining by Safranin O. Upon isolation, weak Safranin O staining was present only in more developed GD14 front-LBs. Chondrogenesis proceeded well in all cultures towards day 14, except in the SF-cultivated GD13 hind-LBs, where Safranin O staining was almost absent on day 3. That was associated with a higher percentage of DNA methylation than in SF-cultivated GD13 front-LBs on day three. In SF-cultivated front-LBs, a significant methylation increase between the 3rd and 14th day was detected. In SS-cultivated GD13 front-LBs, methylation increased significantly on day three and then decreased. In older GD14 SS-cultivated LBs, there was no increase of DNA methylation, but they were significantly hypomethylated relative to the SS-cultivated GD13 at days 3 and 14. We confirmed that the global DNA methylation increase is associated with less developed limb organ primordia that strive towards differentiation in vitro, which is of importance for regenerative medicine strategies.
Topics: Animals; Cell Differentiation; Chondrogenesis; DNA Methylation; Extracellular Matrix; Limb Buds; Organ Culture Techniques; Rats
PubMed: 35188093
DOI: 10.17305/bjbms.2021.6584 -
Cell Sep 2014
Topics: Animals; Brain; Embryonic Stem Cells; History, 20th Century; History, 21st Century; Humans; Japan; Neurons; Organ Culture Techniques; Stem Cell Research
PubMed: 25349875
DOI: 10.1016/j.cell.2014.08.024 -
Cell and Tissue Banking Dec 2022To determine the effectiveness of two methods to improve the microbial safety of human corneas preserved in organ culture. We compared the number of positive...
To determine the effectiveness of two methods to improve the microbial safety of human corneas preserved in organ culture. We compared the number of positive preservation solutions of corneas in organ culture in which the initial short-term hypothermic corneal maintenance solution was supplemented with amphotericin B 2.5 µg/mL and the historical data of microbial test results (2015-2019). In addition, we appraised the efficacy of Gram stain to detect bacterial or fungal contamination in the organ culture solutions of corneas from at-risk donors compared to the culture tests of corneas from not-at-risk donors. Statistical analysis was performed using STATA and statistical significance set at p < 0.05. The number of positive culture tests after preservation was 15 (0.5%) in 2020 compared to a mean of 37 (1.2%) in the period 2015-2019 (p < 0.01), with 10 (1.0%) positive samples in the cohort of 998 corneas from at-risk donors and 5 (0.2%) in the 2046 corneas from not-at-risk donors (p < 0.01). All corneas from at-risk donors tested positive at Gram stain and the results were available 1-3 days before those of the conventional culture tests. Amphotericin B supplementation in the short-term maintenance solution markedly reduced the number of positive microbial tests after organ culture and the early detection of contaminants, including slow-growing microorganisms, by Gram stain before the standard culture results. This meant fewer corneas being discarded and a greater likelihood of preventing post-graft infections.
Topics: Humans; Amphotericin B; Cornea; Tissue Donors; Organ Culture Techniques; Bacteria; Organ Preservation; Corneal Transplantation; Eye Banks
PubMed: 34791554
DOI: 10.1007/s10561-021-09981-1 -
Journal of Clinical Laboratory Analysis Jan 2018Anti-tissue transglutaminase (anti-tTG) and endomysium antibodies (EMA) are detectable in duodenal culture media of celiac disease (CD) patients. To improve the...
One-step immunochromatographic visual assay for anti-transglutaminase detection in organ culture system: An easy and prompt method to simplify the in vitro diagnosis of celiac disease.
BACKGROUND
Anti-tissue transglutaminase (anti-tTG) and endomysium antibodies (EMA) are detectable in duodenal culture media of celiac disease (CD) patients. To improve the management of this organ culture system, we evaluated the anti-tTG occurrence by immunochromatographic assay (ICA).
METHODS
A total of 103 CD patients and 41 disease controls underwent duodenal biopsy for the organ culture. In culture supernatants, IgA anti-tTG were tested by both enzyme-linked immunosorbent assay (ELISA) and ICA, IgA EMA were searched by indirect immunofluorescence analysis (iIFA).
RESULTS
Endomysium antibodies and anti-tTG measured by ELISA were positive in culture media of all CD patients, while anti-tTG detected by ICA were positive in culture media of 87/103 CD patients. Anti-tTG ICA scores significantly correlated with anti-tTG ELISA values (r=.71, P<.0001). Sensitivity, specificity and diagnostic accuracy of anti-tTG detected by ICA were 84.5%, 100% and 88.9%, respectively.
CONCLUSIONS
Using ICA, anti-tTG are detectable in duodenal culture media of most CD patients and the intensity of indicative lines depends on the anti-tTG concentration. Sensitivity and diagnostic accuracy achieved with ICA are lower than those obtained with ELISA but, given that the first is a more easy and prompt method, data suggest the possibility of utilizing it in the in vitro diagnosis of CD.
Topics: Adolescent; Adult; Aged; Autoantibodies; Biopsy; Case-Control Studies; Celiac Disease; Chromatography, Affinity; Endoscopy, Gastrointestinal; Enzyme-Linked Immunosorbent Assay; Female; Humans; Male; Middle Aged; Organ Culture Techniques; Sensitivity and Specificity; Transglutaminases; Young Adult
PubMed: 28276604
DOI: 10.1002/jcla.22195 -
The American Journal of Pathology Sep 2018Colorectal cancer (CRC) diagnosis and prognostic stratification are based on histopathologic assessment of cell or nuclear pleomorphism, aberrant mitotic figures,... (Review)
Review
Colorectal cancer (CRC) diagnosis and prognostic stratification are based on histopathologic assessment of cell or nuclear pleomorphism, aberrant mitotic figures, altered glandular architecture, and other phenomic abnormalities. This complexity is driven by oncogenic perturbation of tightly coordinated spatiotemporal signaling to disrupt multiple scales of tissue organization. This review clarifies molecular and cellular mechanisms underlying common CRC histologic features and helps understand how the CRC genome controls core aspects of tumor aggressiveness. It further explores a spatiotemporal framework for CRC phenomics based on regulation of living cells in fundamental and organotypic model systems. The review also discusses tissue homeostasis, considers distinct classes of oncogenic perturbations, and evolution of cellular or multicellular cancer phenotypes. It further explores the molecular controls of cribriform, micropapillary, and high-grade CRC morphology in organotypic culture models and assesses relevant translational studies. In addition, the review delves into complexities of morphologic plasticity whereby a single molecular signature generates heterogeneous cancer phenotypes, and, conversely, morphologically homogeneous tumors show substantive molecular diversity. Principles outlined may aid mechanistic interpretation of omics data in a setting of cancer pathology, provide insight into CRC consensus molecular subtypes, and better define principles for CRC prognostic stratification.
Topics: Animals; Colorectal Neoplasms; Disease Models, Animal; Humans; Organ Culture Techniques
PubMed: 30028958
DOI: 10.1016/j.ajpath.2018.05.021 -
Scientific Reports Apr 2021Open-globe injuries have poor visual outcomes and have increased in frequency. The current standard of care is inadequate, and a therapeutic is needed to stabilize the...
Open-globe injuries have poor visual outcomes and have increased in frequency. The current standard of care is inadequate, and a therapeutic is needed to stabilize the injury until an ophthalmic specialist is reached. Unfortunately, current models or test platforms for open-globe injuries are insufficient. Here, we develop and characterize an open-globe injury model using an anterior segment organ-culture platform that allows therapeutic assessment for up to 72 h post-injury. Anterior segments maintained in organ culture were kept at physiological intraocular pressure throughout, and puncture injuries were created using a novel pneumatic-powered system. This system can create high-speed, military-relevant injuries up to 4.5 mm in diameter through the cornea. From intraocular pressure readings, we confirmed a loss of pressure across the 72 h after open-globe injury. Proof-of-concept studies with a Dermabond tissue adhesive were performed to show how this model system could track therapeutic performance for 72 h. Overall, the organ-culture platform was found to be a suitable next step towards modeling open-globe injuries and assessing wound closure over the critical 72 h post-injury. With improved models such as this, novel biomaterial therapeutics development can be accelerated, improving care, and, thus, improving the prognosis for the patients.
Topics: Animals; Cyanoacrylates; Eye; Eye Injuries, Penetrating; Intraocular Pressure; Models, Theoretical; Organ Culture Techniques; Swine; Tissue Adhesives; Visual Acuity
PubMed: 33879808
DOI: 10.1038/s41598-021-87910-8 -
International Journal of Molecular... Jul 2021Organoids represent one of the most important advancements in the field of stem cells during the past decade. They are three-dimensional in vitro culturing models that... (Review)
Review
Organoids represent one of the most important advancements in the field of stem cells during the past decade. They are three-dimensional in vitro culturing models that originate from self-organizing stem cells and can mimic the in vivo structural and functional specificities of body organs. Organoids have been established from multiple adult tissues as well as pluripotent stem cells and have recently become a powerful tool for studying development and diseases in vitro, drug screening, and host-microbe interaction. The use of stem cells-that have self-renewal capacity to proliferate and differentiate into specialized cell types-for organoids culturing represents a major advancement in biomedical research. Indeed, this new technology has a great potential to be used in a multitude of fields, including cancer research, hereditary and infectious diseases. Nevertheless, organoid culturing is still rife with many challenges, not limited to being costly and time consuming, having variable rates of efficiency in generation and maintenance, genetic stability, and clinical applications. In this review, we aim to provide a synopsis of pluripotent stem cell-derived organoids and their use for disease modeling and other clinical applications.
Topics: Animals; Drug Evaluation, Preclinical; Humans; Models, Biological; Organ Culture Techniques; Organoids; Pluripotent Stem Cells
PubMed: 34299287
DOI: 10.3390/ijms22147667 -
Annual Review of Biomedical Engineering Aug 2011The definitive treatment for end-stage organ failure is orthotopic transplantation. However, the demand for transplantation far exceeds the number of available donor... (Review)
Review
The definitive treatment for end-stage organ failure is orthotopic transplantation. However, the demand for transplantation far exceeds the number of available donor organs. A promising tissue-engineering/regenerative-medicine approach for functional organ replacement has emerged in recent years. Decellularization of donor organs such as heart, liver, and lung can provide an acellular, naturally occurring three-dimensional biologic scaffold material that can then be seeded with selected cell populations. Preliminary studies in animal models have provided encouraging results for the proof of concept. However, significant challenges for three-dimensional organ engineering approach remain. This manuscript describes the fundamental concepts of whole-organ engineering, including characterization of the extracellular matrix as a scaffold, methods for decellularization of vascular organs, potential cells to reseed such a scaffold, techniques for the recellularization process and important aspects regarding bioreactor design to support this approach. Critical challenges and future directions are also discussed.
Topics: Amniotic Fluid; Animals; Bioreactors; Cells, Cultured; Embryonic Stem Cells; Extracellular Matrix; Fetal Blood; Humans; Induced Pluripotent Stem Cells; Organ Culture Techniques; Tissue Engineering; Tissue Scaffolds
PubMed: 21417722
DOI: 10.1146/annurev-bioeng-071910-124743 -
Acta Ophthalmologica Mar 2017Endothelial assessment is crucial in the release of corneas for grafting. We retrospectively analysed the role of endothelial morphology parameters in predicting...
PURPOSE
Endothelial assessment is crucial in the release of corneas for grafting. We retrospectively analysed the role of endothelial morphology parameters in predicting endothelial cell loss during organ culture.
METHODS
Human donor corneas were cultured in minimal essential medium with 2% fetal calf serum and antibiotics. Initial endothelial morphology was assessed microscopically using score parameters polymegethism (POL), pleomorphism (PLE), granulation (GRA), vacuolization (VAC), segmentation of cell membranes (SEG), Descemet's folds (DF), trypan blue-positive cells (TBPC) and endothelial cell-free areas (ECFA). Some corneas were primarily rejected based on endothelial assessment. Endothelial cell density (ECD) was assessed at the beginning (I-ECD) and end of culture. Corneas were then placed in dehydration medium (as above + 5% dextran 500). In a subgroup, ECD was reassessed after dehydration. Endothelial cell loss during culture (ECL@Culture) and culture+dehydration (ECL-Culture&Dehydration) were calculated. Data were given as mean ± SD and analysed using multiple linear and logistic regression. Odds ratios (OR) and 95% confidence intervals (CI) were calculated.
RESULT
I-ECD was 2812 ± 360/mm (n = 2356). The decision to reject a cornea due to endothelial assessment was associated negatively with I-ECD (OR = 0.77/100 cells, CI 0.7-0.82) and positively with ECFA (OR = 2.7, CI 1.69-4.35), SEG (OR =1.3, CI 1.01-1.68) and donor age (OR = 1.26/decade, CI 1.33-1.41). ECL@Culture was 153 ± 201/mm (n = 1277), ECL@Culture&Dehydration was 169 ± 183/mm (n = 918). ECL@Culture was associated positively with donor age, I-ECD, GRA and TBPC, and negatively with PLE, and DF. ECL@Culture&Dehydration was associated positively with age, sex, initial ECD, POL, PLE, VAC and TBPC.
CONCLUSION
Morphological parameters displayed associations with the exclusion of corneas from culture and with endothelial cell loss. Appropriate parameter selection for screening purposes may help improve graft quality.
Topics: Cell Count; Cell Survival; Corneal Endothelial Cell Loss; Endothelium, Corneal; Eye Banks; Female; Follow-Up Studies; Humans; Male; Middle Aged; Organ Culture Techniques; Organ Preservation; Retrospective Studies; Time Factors
PubMed: 27233584
DOI: 10.1111/aos.13108