Nestin positive cells vs human islets
Ontology highlight
ABSTRACT:
INSTRUMENT(S): G2500A Hewlett Packard
ORGANISM(S): Homo sapiens
SUBMITTER: Joel Habener
PROVIDER: E-MTAB-3049 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:We have developed an in vitro culture system that allows us to expand progenitor cells from human islet preparations and differentiate them into insulin-producing cells. We noticed however, that cultures from individual islet preparations had very heterogeneous outcomes, from good differentiation to almost none. We therefore speculated that our progenitor cell cultures contained different kinds of cells and that the true endocrine progenitor cells are present in the successful cultures, but not in the unsuccessful ones. To address this issue and to begin to identify markers for the true endocrine progenitor cells we compared global gene expression between a very successful culture (final insulin-expression 10% of islets) and an unsuccessful one (final insulin-expression 0%). We also included RNA from freshly isolated islets for control purposes. The cultures from donor A yielded substantial differentiation, while the cultures from donor B showed no successful differentiation.
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Project description:Background: Survival and function of insulin-secreting pancreatic β-cells are markedly altered by changes in nutrient availability. In vitro, culture in 10 rather than 2mM glucose improves rodent β-cell survival and function whereas glucose concentrations above 10mM are deleterious. Aim-Method: To identify the mechanisms of such β-cell plasticity, we tested the effects of a 18h culture at 2, 5, 10 and 30mM glucose on the transcriptome of rat islets precultured for 1 week at 10mM glucose (Affymetrix Rat 230.2 arrays). Results: Culture in either 2-5mM or 30mM instead of 10mM glucose markedly impaired β-cell function without affecting islet cell survival. Of ~16000 probe sets reliably detected in islets, ~5000 were significantly regulated at least 1.4-fold by glucose. Analysis of these probe sets with GeneCluster software identified 10 mRNA profiles with unidirectional up- or down-regulation between 2 and 10, 2 and 30, 5 and 10, 5 and 30 or 10 and 30 mM glucose, and 8 complex V-shaped or inverse V-shaped profiles with a nadir or peak level of expression in 5 or 10mM glucose. Analysis of genes belonging to these various clusters with Onto-express and GenMapp software revealed several signaling and metabolic pathways that may contribute to the induction of β-cell dysfunction and apoptosis after culture in low or high vs. intermediate glucose concentration. Conclusion: We have identified 18 distinct mRNA profiles of glucose-induced changes in islet gene mRNA levels that should help understanding the mechanisms by which glucose affects β-cell survival and function under states of chronic hypo- or hyperglycemia. Experiment Overall Design: Effect of 18h culture in 2, 5, 10 and 30 mmol/l glucose on the transcriptome of rat pancreatic islets precultured for 1 week in 10 mmol/l glucose. Four experiments were done on different islet preparations over a two-months period.
2009-01-31 | E-GEOD-12817 | biostudies-arrayexpress
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Project description:Intensive efforts are focused on identifying regulators of human pancreatic islet cell growth and maturation to accelerate development of therapies for diabetes. In the present study we evaluate the contribution of the physical properties of the extracellular matrix in supporting long term culture of human islets in vitro. We used metal oxide layers with tailored nanoscale roughness to fabricate scaffolds. Their suitability to support long term culture of human islets was investigated through assessment of β-cell survival, differentiation and function by using a proteomic approach to evaluate the molecular mechanisms involved. Proteomic analysis confirmed that ns-ZrOx promoted the activation of a transcriptional and translational program leading to activation of pro-survival and pro-differentiation signalling pathways. The process was driven by a mechanotransductive pathway driven by nanostructured topology, via remodelling of the actin cytoskeleton nuclear architecture and increase of oxygen sensor PDH2 Our data suggest that β-cells relay on nanotopography features to regulate their differentiation and survival. Tailored nanostructured substrates may provide a unique strategy to identify novel hints for tissue engineering and molecular /pharmacological targets of intervention to treat diabetes mellitus.
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