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In vitro directed differentiation of mouse embryonic stem cells into insulin-producing cells 总被引:28,自引:0,他引:28
Aims/hypothesis We recently demonstrated that insulin-producing cells derived from embryonic stem cells normalise hyperglycaemia in transplanted diabetic mice. The differentiation and selection procedure, however, was successful in less than 5% of the assays performed. Thus, to improve its effectiveness, new strategies have been developed, which increase the number of islet cells or islet progenitors. Methods Mouse embryonic stem cells transfected with a plasmid containing the Nkx6.1 promoter gene followed by a neomycin-resistance gene, were cultured with factors known to participate in endocrine pancreatic development and factors that modulate signalling pathways involved in these processes. Neomycin was used to select the Nkx6.1-positive cells, which also express insulin. The transfected cells were differentiated using several exogenous agents, followed by selection of Nkx6.1-positive cells. The resulting cells were analysed for pancreatic gene and protein expression by immunocytochemistry, RT-PCR and radioimmunoassay. Also, proliferation assays were performed, as well as transplantation to streptozotocin-induced diabetic mice.Results The protocols yielded cell cultures with approximately 20% of cells co-expressing insulin and Pdx-1. Cell trapping selection yielded an almost pure population of insulin-positive cells, which expressed the beta cell genes/proteins Pdx-1, Nkx6.1, insulin, glucokinase, GLUT-2 and Sur-1. Subsequent transplantation to streptozotocin-induced diabetic mice normalised their glycaemia during the time period of experimentation, proving the efficiency of the protocols.Conclusions/interpretation These methods were both highly efficient and very reproducible, resulting in a new strategy to obtain insulin-containing cells from stem cells with a near 100% success rate, while actively promoting the maturation of the exocytotic machinery.Abbreviations Anti-Shh antibody against sonic hedgehog - D3 undifferentiated D3 stem cell line - EB embryoid bodies - ES embryonic stem - FBS fetal bovine serum - LIF leukaemia inhibitory factor - mES mouse embryonic stem - Ngn3 neurogenin 3 - P gelatine-coated plates - Pdx-1 pancreatic duodenum homeobox 1 相似文献
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Rahul S. Bhansali Malini Rammohan Paul Lee Anouchka P. Laurent Qiang Wen Praveen Suraneni Bon Ham Yip Yi-Chien Tsai Silvia Jenni Beat Bornhauser Aurlie Siret Corinne Fruit Alexandra Pacheco-Benichou Ethan Harris Thierry Besson Benjamin J. Thompson Young Ah Goo Nobuko Hijiya Maria Vilenchik Shai Izraeli Jean-Pierre Bourquin Sbastien Malinge John D. Crispino 《The Journal of clinical investigation》2021,131(1)
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Anouchka Modesto Aurore Siegfried Amelie Lusque Sbastien Vergez Jerome Sarini Laurent Brouchet Emmanuelle Uro-Coste Pierre Graff-Cailleaud Jean Pierre Delord 《Current oncology (Toronto, Ont.)》2021,28(3):1673
Introduction: Recent modifications in the epidemiology of oropharyngeal squamous cell carcinoma (OSCC) have led to the increase of Human papillomavirus (HPV) related metastatic head and neck cancer patients with high life expectancy even at advanced stage, low comorbidity and still restricted systemic therapy opportunities. In the recent era of ablative therapies’ development, oligometastatic HPV OSCC patients are indubitably good candidates for intensified treatment. However, data related to outcomes after optimised management of metastatic sites are dramatically missing. Materials and patients: In our cohort of 186 unselected consecutive OSCC patients treated with curative intent at our institution between 2009 and 2013, we analysed the incidence, treatment and outcomes of distant metastatic (DM) failure according to p16 status. Results: After a median follow-up of 4.2 years (95% CI: 3.8–4.4) from primary diagnosis of OSCC, 21/95 p16− patients (22.1%) vs. 8/91 (8.8%) p16+ patients presented DM failure with a median interval of 11 (range 0–46) and 28 months (range 0–71), respectively (p = 0.10). Overall survival (OS) after DM failure was significantly higher in p16+ patients with a two-year OS rate of 75% and 15% for p16+ and p16−, respectively (p = 0.002). In eight HPV-related metastatic patients, three underwent ablative lung metastasis treatment and are still complete responders four to five years later. Conclusion: This study highlights distinct outcomes of metastatic HPV-related OSCC patients emphasised by three long-term complete responders after lung ablative treatment. In patients with high life expectancy and limited tumour burden, the question of ablative treatment such as metastasectomy or stereotactic ablative radiotherapy (SBRT) should be addressed. 相似文献
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Anouchka Modesto Anne Laprie Laure Vieillevigne Pierre Graff Jérôme Sarini Sébastien Vergez Jean-Pierre Delord Jean-Claude Farenc Emmanuelle Vigarios Thomas Filleron Michel Rives 《Strahlentherapie und Onkologie》2015,191(3):225-233
Purpose
The purpose of this work was to retrospectively determine the value of intensity-modulated radiotherapy (IMRT) in patients with laryngeal and hypopharyngeal squamous cell carcinoma (LHSCC), on outcome and treatment-related toxicity compared to 3-dimensional conformal radiotherapy (3D-CRT).Materials and methods
A total of 175 consecutive patients were treated between 2007 and 2012 at our institution with curative intent RT and were included in this study: 90 were treated with 3D-CRT and 85 with IMRT. Oncologic outcomes were estimated using Kaplan–Meier statistics; acute and late toxicities were scored according to the Common Toxicity Criteria for Adverse Events scale v 3.0.Results
Median follow-up was 35 months (range 32–42 months; 95% confidence interval 95?%). Two-year disease-free survival did not vary, regardless of the technique used (69?% for 3D-CRT vs. 72?%; for IMRT, p?=?0.16). Variables evaluated as severe late toxicities were all statistically lower with IMRT compared with 3D-CRT: xerostomia (0 vs. 12?%; p?<?0.0001), dysphagia (4 vs. 26?%; p?<?0.0001), and feeding-tube dependency (1 vs 13?%; p?=?0.0044). The rates of overall grade ≥?3 late toxicities for the IMRT and 3D-CRT groups were 4.1 vs. 41.4?%, respectively (p?<?0.0001).Conclusion
IMRT for laryngeal and hypopharyngeal cancer minimizes late dysphagia without jeopardizing tumor control and outcome.9.
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