Skip to main page content
Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2012 May 14;3(1-3):51-61.
doi: 10.3727/215517912X639487. eCollection 2012 Jan.

Differentiation of Mouse Pancreatic Stem Cells Into Insulin-Producing Cells by Recombinant Sendai Virus-Mediated Gene Transfer Technology

Affiliations
Free PMC article

Differentiation of Mouse Pancreatic Stem Cells Into Insulin-Producing Cells by Recombinant Sendai Virus-Mediated Gene Transfer Technology

Hiroshi Yukawa et al. Cell Med. .
Free PMC article

Abstract

Islet transplantation, including β-cells, has proven to be effective for diabetes in many recent studies; however, this treatment strategy requires sufficient organ donors. One attractive approach for the generation of β-cells is to utilize the expansion and differentiation of cells from pancreatic stem cells (PSCs), which are closely associated to the β-cells lineage. In this study, we investigated whether important transcription factors (Pdx-1, Ngn3, NeuroD, and MafA) in islet cells could be efficiently transduced into mouse PSCs (mPSCs) using Sendai virus (SeV) vectors and found that the transduced cells were differentiated into insulin-producing pancreatic β-cells. The mPSCs transduced with single transcription factors using SeV vectors could not express the insulin-2 mRNA. When combinations of two transcription factors were transduced using the SeV vectors, including combinations of Pdx-1 + NeuroD, Pdx-1 + MafA, and NeuroD + MafA, the expression of insulin-2 mRNA was low but could be detected. When combinations of three or more transcription factors were transduced using SeV vectors, the expression of insulin-2 mRNA could be detected. In particular, the transduction of the combination of PDX-1, NeuroD, and MafA produced the most effective for the expression of insulin-2 mRNA out of all of the different combinations examined. These data suggest that the transduction of transcription factors using SeV vectors facilitates mPSC differentiation into insulin-producing cells and showed the possibility of regenerating β-cells by using transduced PSCs.

Keywords: Insulin-producing cells; Islet transplantation; Pancreatic stem cells (PSCs); Sendai virus (SeV).

Figures

Figure 1
Figure 1
(A) The method used to generate the β-cells and transcriptional factors expressed in the cells at each steps. (B) The basic gene structure of the F (envelope fusion protein)–defective Sendai virus (SeV) vector. Enhanced green fluorescent protein (EGFP) was inserted between the M and HN locations. (C) The experimental design for infection of mouse pancreatic stem cells (mPSCs) with SeV vectors and for the differentiation of mPSCs transfected with pancreatic and duodenal homeobox factor-1 (Pdx-1), neurogenin 3 (Ngn-3), neurogenic differentiation factor (NeuroD), and/or v-maf musculoaponeurotic fibrosarcoma oncogene homolog A (MafA).
Figure 2
Figure 2
(A) The viability of mPSCs infected with SeV vectors at multiplicity of infections (MOIs) of 0, 50, or 200. No significant differences were observed. (B) The proliferation rate of mPSCs infected with SeV vectors at MOIs of 0, 50, or 200. No significant differences were observed.
Figure 3
Figure 3
(A) The morphologies and GFP expressions levels of mPSCs infected with single transcription factors using SeV vectors on days 2, 4, and 7. These figures were obtained by fluorescence microscopy. Scale bars: 100 μm. (B) The mRNA expression levels of pancreas-related genes in the mPSCs infected with single transcriptional factors using the SeV vectors on day 7. Cells incubated with medium alone were used as a negative control, and the insulinoma MIN6 cells were used as a positive control.
Figure 4
Figure 4
(A) The morphologies and GFP expression levels of mPSCs infected with two transcriptional factors using SeV vectors on days 2, 4, and 7. These figures were obtained by fluorescence microscopy. Scale bars: 100 μm. (B) The mRNA expression levels of pancreas-related genes in the mPSCs infected with two transcriptional factors using the SeV vectors on day 7.
Figure 5
Figure 5
(A) The morphologies and GFP expressions of mPSCs infected with three transcriptional factors using SeV vectors on days 2, 4, and 7. These figures were obtained by fluorescent microscopy. Scale bars: 100 μm. (B) The mRNA expression levels of pancreas-related genes in the mPSCs transduced with three or four transcription factors using SeV vectors on day 7.

Similar articles

See all similar articles

Cited by 1 article

LinkOut - more resources

Feedback