Combination of GLP-1 and sodium butyrate promote differentiation of pancreatic progenitor cells into insulin-producing cells

Citations:

Tissue and Cell, Volume 40, Issue 6, December 2008, Pages 437-445Li Li, Ren Lili, Qi Hui, Wang Min, Wen Xue, Shen Xin, Liu Jing, Li Yan, Liu Yeqiang, He Fenrong, Li Furong, Shen Guanxin


Summary:

In this study, the researchers isolated pancreatic progenitor cells from pancreateic precursor cells. To do so, they relied on the hypothesis of a previous experiment showing that cells expressing nestin (a protein of the adult pancreas) were pancreatic progenitor cells. To do this, the reserachers created transgenic mice which expressed a recombinant form of nestin enhanced with a green fluorescent protein tag (nestin-EGFP). Nestin expression could then be quantified by fluorescence activated cell sorting (FACS). This method analyzes fluorescence on a per-cell basis, allowing researchers to quantify nestin expression in single cells. Neonatal pancreatic cells (NPPCs), which are multipotent, have the ability to differentiate into pancreatic or neural cells. Such cells were selected from the islets of six week old transgenic mice and then plated. After 5 days in differentiation medium, the cells had changed morphology and most were expressing nestin. Nestin expressing cells were grown in differentiation medium. Using antibody staining and real time PCR, the researchers found that after 25 days of differentiation, cells were marked to become either pancreatic or neuronal cells, suggesting that nestin-positive cells could differentiate into pancreatic islet or neural cells.
The researchers focused on insulin-producing cells. The researchers hypothesized that sodium butyrate, a fatty acid that inhibits histone deacetylase, allowing for increased DNA transcription would cause differentiation towards insulin-producing pancreatic cells. They also hypothesized that glucagon-like peptide-1 (GLP-1), which plays a role in the regeneration of pancreatic beta cells (the insulin-releasing cells) would cause differentaition towards insulin-producing pancreatic cells. By treating cells with sodium butyrate alone, the percent of insulin-producing cells formed did not change from the control (8%). GLP-1 alone increased the percent of insulin-producing cells to 20%. However, combining sodium butyrate with GLP-1 caused the percent to increase to 45%. These percentages were confirmed by immunofluorescence and real-time PCR to determine gene expression of islet and neural developmental control genes. In addition, the cells were cultured at varying levels of glucose concentration. Cells cultured with high glucose concentration released more insulin than cells cultured at lower levels. Though not confirmed, the results suggested that insulin secretion was glucose dose-dependent. Thus, through application of sodium butyrate and GLP-1, researchers were able to induce differentiation of cells that had similar gene expression and insulin responses to pancreatic insulin-producing cells.


Importance:

The ability to engineer insulin-producing cells for transplantation into humans is desirable for the treatment of Type 1 diabetes in which the immune system destroys the insulin-producing cells of the pancreas. The use of pancreatic precursor cells to produce the desired insulin-producing cells is a more controllable option than embryonic stem cells. This study developed a method to find pancreatic precursor cells using nestin expression as a marker. The ability to select these cells allows for further control to create insulin-producing cells. This study did so, growing cells with sodium butyrate and GLP-1 to select for insulin-producing cells. This is significant because the natural rate of differentation into insulin-producing cells by neonatal pancreatic cells is low and therefore not useful for large-scale production. The results of this study could be used in the generation of implantable insulin-producing cells to treat Type 1 diabetes. Such an implant would allow for much tighter regulation of blood-glucose levels than current diabetes treatments allow.