1,721,026 research outputs found
Adult liver stem cells in hepatic regeneration and cancer
No full textAn alternative source of livers for transplantation in patients with (genetic) liver diseases and liver failure is needed because liver donors are scarce. HPC-derived hepatocyte-like cells could be one of the options. Because dogs and humans share liver-pathologies and disease-pathways, the dog is considered the best model for human liver disease. We can use dogs to test safety and efficacy of a new treatment before considering to apply it in human clinics. We developed the long-term culture system of adult canine hepatic progenitor cells in three-dimensions, called organoids. Canine liver organoids showed characteristics of stem cells, cholangiocytes and hepatocytes, mimicking in vivo liver progenitor cell phenotype, and had potential to differentiate towards functional hepatocytes. Organoids derived from COMMD1-deficient dogs presented a copper excretion defect similar to the in vivo situation in dogs with copper storage disease. Upon transduction with a COMMD1 construct, successful restoration of copper excretion was achieved. Patient-derived liver organoids are a powerful tool to perform disease modeling and demonstrate the feasibility to perform gene correction in liver organoids, a key prerequisite for autologous transplantation for gene-therapy. Upon liver injury either hepatocytes themselves or progenitor cells proliferate to regenerate the liver. However, in many virus-induced liver diseases and cancers the hepatocytes harbor mutations in the tumor suppressor genes, Rb and p53. Therefore the regenerative response may differ from regeneration under normal situations. We analyzed the cooperative role of Rb and p53 in accordance with the 3, 5-diethoxycarbonyl-1, 4-dihydrocollidine (DDC) diet, a xenobiotic inducing hepatobiliary injury and oval cell proliferation. DDC is metabolized by Cyp3a. Deletion of both Rb and p53 deregulates Cyp3a activity causing less porphyrin accumulation, therefore mild oval cell proliferation is induced. In addition, Rb/p53 deficient livers experience less biliary injury, but instead more hepatocellular injury under DDC exposure. Finally, inactivation of Rb/p53 shortens life span and accelerates HCC development after DDC exposure. Rb and p53 are not only important cell cycle regulators, but also influence xenobiotic metabolism of cells. Liver resection and radiofrequency ablation (RFA) are the treatment of choice for liver cancers. Yet, cancer recurrence rate after resection is high. We focused on identifying the mechanism of liver tumor formation after surgical interventions. Partial hepatectomy and RFA were performed in mice with p53 and Rb-specific deletion in the livers. Cancer developed at the necrotic injury site, and cancer initiation was associated with migration, expansion, and transformation of bile duct cells within the injury site. This resulted in the formation of undifferentiated carcinomas. A necrotic environment induced by surgery combined with loss of tumor suppressor genes causes liver cancer and epithelial-mesenchymal transition at the surgery site. Bile duct cells or HPCs are the cell-of-origin of liver tumors in this model. Polyploidization is a normal physiological event for livers. Atypical E2Fs, especially E2F8, are essential for hepatocyte binucleation and polyploidization. E2F8 inactivation resulted in up-regulation of E2F-target genes promoting DNA replication, DNA repair, mitosis and cytokinesis. Prevention of polyploidization through inactivation of atypical E2Fs did not have any impact on liver differentiation, zonation, metabolism and regeneration
Radiographic and MRI characteristics of lumbar disseminated idiopathic spinal hyperostosis and spondylosis deformans in dogs
No full textOBJECTIVES To evaluate clinical signs, describe lesions and differences in the magnetic resonance imaging appearance of spinal new bone formations classified as disseminated idiopathic spinal hyperostosis and/or spondylosis deformans on radiographs and compare degeneration status of the intervertebral discs using the Pfirrmann scale. METHODS Retrospective analysis of 18 dogs presented with spinal disorders using information from radiographic and magnetic resonance imaging examinations. RESULTS All dogs were found to be affected with both disseminated idiopathic spinal hyperostosis and spondylosis deformans. Neurological signs due to foraminal stenosis associated with disseminated idiopathic spinal hyperostosis were found in two dogs. Spondylosis deformans was associated with foraminal stenosis and/or disc protrusion in 15 cases. The Pfirrmann score on magnetic resonance imaging was significantly higher in spondylosis deformans compared with disseminated idiopathic spinal hyperostosis and signal intensity of new bone due to disseminated idiopathic spinal hyperostosis was significantly higher compared to spondylosis deformans. CLINICAL SIGNIFICANCE Differences between disseminated idiopathic spinal hyperostosis and spondylosis deformans found on magnetic resonance imaging contribute to an increased differentiation between the two entities. Clinically relevant lesions in association with disseminated idiopathic spinal hyperostosis were rare compared to those seen with spondylosis deformans
Keratin 19 marks poor differentiation and a more aggressive behaviour in canine and human hepatocellular tumours
Get PDFKeratin 19 marks poor differentiation and a more aggressive behaviour in canine and human hepatocellular tumours Renee GHM van Sprundel1, Ted SGAM van den Ingh2, Valeer J Desmet3, Azeam Katoonizadeh3, Louis C Penning1, Jan Rothuizen1, Tania Roskams3 and Bart Spee13* * Corresponding author: Bart Spee [email protected] Author Affiliations 1 Department of Clinical Sciences of Companion Animals, Faculty of Veterinary medicine, Utrecht University, Utrecht, The Netherlands 2 TCCI Consultancy BV, Utrecht, The Netherlands 3 Department of Morphology and Molecular Pathology, University Hospitals Leuven, Leuven, Belgium For all author emails, please log on. Comparative Hepatology 2010, 9:4 doi:10.1186/1476-5926-9-4 The electronic version of this article is the complete one and can be found online at: http://www.comparative-hepatology.com/content/9/1/4 Received: 23 November 2009 Accepted: 18 February 2010 Published: 18 February 2010 © 2010 van Sprundel et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited
Clinical outcome after traumatic spinal fractures in patients with ankylosing spinal disorders compared with control patients.
No full textBackground context The clinical outcome of patients with ankylosing spinal disorders (ASDs) sustaining a spinal fracture has been described to be worse compared with the general trauma population. Purpose To investigate clinical outcome (neurologic deficits, complications, and mortality) after spinal injury in patients with ankylosing spondylitis (AS) and diffuse idiopathic skeletal hyperostosis (DISH) compared with control patients. Study design Retrospective cohort study. Patient sample All patients older than 50 years and admitted with a traumatic spinal fracture to the Emergency Department of the University Medical Center Utrecht, the Netherlands, a regional level-1 trauma center and tertiary referral spine center. Outcome measures Data on comorbidity (Charlson comorbidity score), mechanism of trauma, fracture characteristics, neurologic deficit, complications, and in-hospital mortality were collected from medical records. Methods With logistic regression analysis, the association between the presence of an ASD and mortality was investigated in relation to other known risk factors for mortality. Results A total of 165 patients met the inclusion criteria; 14 patients were diagnosed with AS (8.5%), 40 patients had DISH (24.2%), and 111 patients were control patients (67.3%). Ankylosing spinal disorder patients were approximately five years older than control patients and predominantly of male gender. The Charlson comorbidity score did not significantly differ among the groups, but Type 2 diabetes mellitus and obesity were more prevalent among DISH patients. In many AS and DISH cases, fractures resulted from low-energy trauma and showed a hyperextension configuration. Patients with AS and DISH were frequently admitted with a neurologic deficit (57.1% and 30.0%, respectively) compared with controls (12.6%; p=.002), which did not improve in the majority of cases. In AS and DISH patients, complication and mortality rates were significantly higher than in controls. Logistic regression analysis showed the parameters age and presence of DISH to be independently, statistically significantly related to mortality. Conclusions Many patients with AS and DISH showed unstable (hyperextension) fracture configurations and neurologic deficits. Complication and mortality rates were higher in patients with ASD compared with control patients. Increasing age and presence of DISH are predictors of mortality after a spinal fracture
Chondrocyte redifferentiation and construct mechanical property development in single-component photocrosslinkable hydrogels.
No full textHydrogels are promising materials for cartilage repair, but the properties required for optimal functional outcomes are not yet known. In this study, we functionalized four materials that are commonly used in cartilage tissue engineering and evaluated them using in vitro cultures. Gelatin, hyaluronic acid, polyethylene glycol, and alginate were functionalized with methacrylic anhydride to make them photocrosslinkable. We found that the responses of encapsulated human chondrocytes were highly dependent on hydrogel type. Gelatin hydrogels supported cell proliferation and the deposition of a glycosaminoglycan rich matrix with significant mechanical functionality. However, cells had a dedifferentiated phenotype, with high expression of collagen type I. Chondrocytes showed the best redifferentiation in hyaluronic acid hydrogels, but the newly formed matrix was highly localized to the pericellular regions, and these gels degraded rapidly. Polyethylene glycol hydrogels, as a bioinert control, did not promote any strong responses. Alginate hydrogels did not support the deposition of new matrix, and the stiffness decreased during culture. The markedly different response of chondrocytes to these four photocrosslinkable hydrogels demonstrates the importance of material properties for chondrogenesis and extracellular matrix production, which are critical for effective cartilage repair
A biomimetic extracellular matrix for cartilage tissue engineering centered on photocurable gelatin, hyaluronic acid and chondroitin sulfate.
No full textThe development of hydrogels tailored for cartilage tissue engineering has been a research and clinical goal for over a decade. Directing cells towards a chondrogenic phenotype and promoting new matrix formation are significant challenges that must be overcome for the successful application of hydrogels in cartilage tissue therapies. Gelatin-methacrylamide (Gel-MA) hydrogels have shown promise for the repair of some tissues, but they have not been extensively investigated for cartilage tissue engineering. We encapsulated human chondrocytes in gel-MA based hydrogels, and show that with the incorporation of small quantities of photo-crosslinkable hyaluronic acid methacrylate (HA-MA), and to a lesser extent chondroitin sulfate methacrylate (CS-MA), chondrogenesis and mechanical properties can be enhanced. The addition of HA-MA to Gel-MA constructs resulted in more rounded cell morphologies, enhanced chondrogenesis as assessed by gene expression and immunofluorescence, and increased quantity and distribution of the newly synthesised ECM throughout the construct. Consequently, while the compressive moduli of control Gel-MA constructs increased by 26 kPa after 8 weeks culture, constructs with HA-MA and CS-MA increased by 96 kPa. The enhanced chondrogenic differentiation, distribution of ECM, and improved mechanical properties make these materials potential candidates for cartilage tissue engineering applications
INCREASED OSMOLARITY AND CELL CLUSTERING PRESERVES CANINE NOTOCHORDAL CELL PHENOTYPE IN CULTURE
No full textAbstract Degeneration of the intervertebral disc (IVD) is associated with a loss of notochordal cells (NCs) from the nucleus pulposus (NP) and their replacement by chondrocyte-like cells. NCs are known to maintain extracellular matrix quality and stimulate the chondrocyte-like NP cells, making NCs attractive for designing new tissue engineering approaches for IVD regeneration. However, optimal conditions, such as osmolarity and other characteristics of the culture media, for long-term culture of NCs are not known. The purpose of this study was to investigate the effects of different culture media and osmolarity on the physiology of NCs in vitro. NC clusters isolated from canine IVDs were suspended in alginate beads and cultured at 37°C under normoxic conditions for 28 days. Three different culture conditions were investigated; (1) DMEM/F12 (300 mOsm/L), (2) α-MEM (300 mOsm/L) and (3) α-MEM adjusted to 400 mOsm/L to mimic a hyperosmolar environment. NC morphology, expression of genes related to NC markers, matrix production and remodelling, and DNA- and GAG analyses were performed on 1, 7, 14, and 28 days in culture. Large, vesicle-containing cells organised in clusters, characterized as NCs, remained present during 28 days for all culture conditions. However, the proportion of the NC clusters decreased over time, whereas the proportion of spindle-shaped cells increased. Gene expression profiling at 7, 14, and 28 days in culture compared to day 1 indicated a initial loss of NC phenotype followed by some recovery of brachyury and aggrecan expression after 28 days of culture supporting a potential recovery of NC phenotype. NCs cultured in α-MEM adjusted to 400 mOsm/L showed the highest expression of brachyury, cytokeratin 18, and aggrecan, the highest GAG production, and the lowest collagen 1α1 gene expression. In conclusion, NCs cultured in alginate in native cell clusters, partially retained their characteristic morphology and recovered their phenotype in long-term culture. The type of culture medium and medium osmolarity appear to be important factors for culturing NC clusters. These findings provide additional information concerning the maintenance of NCs in vitro that may aid further mechanistic inquiry into the biology of NCs. PMID:24304309[PubMed - as supplied by publisher
The Stimulatory Effect of Notochordal-Cell Conditioned Medium in a Nucleus Pulposus Explant Culture
Get PDFOBJECTIVES: Notochordal cell-conditioned medium (NCCM) has previously shown to have a stimulatory effect on nucleus pulposus cells (NPCs) and bone marrow stromal cells (BMSCs) in alginate and pellet cultures. These culture methods provide a different environment than the nucleus pulposus (NP) tissue, in which the NCCM ultimately should exert its effect. The objective of this study is to test whether NCCM stimulates NPCs within their natural environment, and whether combined stimulation with NCCM and addition of BMSCs has a synergistic effect. METHODS: Bovine NP tissue was cultured in an artificial annulus in base medium (BM), porcine NCCM, or BM supplemented with 1 µg/ml Link N. Furthermore, BM and NCCM samples were injected with 1 million BMSCs per NP sample. Samples were cultured for 4 weeks, and analyzed for biochemical contents (water, GAG, hydroxyproline, and DNA), gene expression (COL1A1, COL2A1, ACAN, and SOX9), and histology by Safranin O/Fast Green staining. RESULTS: Culture in NCCM resulted in increased proteoglycan content compared to day 0 and BM, similar to Link N. However, only minor differences in gene expression compared to day 0 were observed. Addition of BMSCs did not result in increased GAG content, and surprisingly, DNA content in BMSC injected groups was not higher than in the other groups after 4 weeks of culture. DISCUSSION: This study shows that indeed, NCCM is capable of stimulating NPC matrix production within the NP environment. The lack of increased DNA content in the BMSC-injected groups indicates that BMSCs have died over time. Identification of the bioactive factors in NCCM is crucial for further development of a NCCM-based treatment for intervertebral disc regeneration
Musculoskeletal health from the “One Medicine” perspective – what can we learn rom large and small animal models (with emphasis on articular cartilage)?
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