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The combined effect ofPdx1overexpression and Shh manipulation on the function of insulin-producing cells derived from adipose-tissue stem cells
- Source :
- FEBS Open Bio
- Publication Year :
- 2018
- Publisher :
- Wiley, 2018.
-
Abstract
- Pancreatic and duodenal homeobox 1 (Pdx1) and Sonic hedgehog (Shh) are the key regulators of beta‐cell function. In vitro experiments have shown that there is significant cooperation between Pdx1 and Shh with regard to the production and maintenance of insulin‐producing cells (IPCs). In this study, the combined effect of Pdx1 overexpression and Shh manipulation on the function of adipose tissue‐derived IPCs was determined. A eukaryotic expression vector (Pdx1‐ pCDNA3.1(+)) was constructed and transfected into a Chinese hamster ovary (CHO) cell line. Adipose tissue‐derived mesenchymal stem cells (ADMSCs) obtained from rats were assigned to two groups [control (C) and manipulated (M)] and differentiated into IPCs. Manipulated cells were treated with a mixture of FGF‐β and cyclopamine and recombinant Shh protein at days 3 and 11, respectively, and transfected with Pdx1‐ pCDNA3.1(+) at day 10. The expression of multiple genes related to function of beta cells was analyzed using real‐time PCR. The functionality of IPCs in vitro was analyzed through dithizone (DTZ) staining and ELISA. IPCs were injected into the tail vein of diabetic rats, and blood glucose and insulin concentrations were measured. CHO cells transfected with Pdx1‐ pCDNA3.1(+) showed a significantly higher expression of Pdx1 compared with nontransfected cells. Manipulated IPCs exhibited a significantly higher expression of MafA, Nkx2.2, Nkx6.1, Ngn3, insulin, and Isl1 and a higher insulin secretion in response to glucose challenge in relation to control cells. Rats that received manipulated IPCs exhibited a higher ability to normalize blood glucose and insulin secretion when compared to controls. Our protocol might be used for more efficient cell therapy of patients with diabetes in the future.
- Subjects :
- 0301 basic medicine
endocrine system
medicine.medical_specialty
medicine.medical_treatment
Adipose tissue
030209 endocrinology & metabolism
General Biochemistry, Genetics and Molecular Biology
Cell therapy
sonic hedgehog pathway
03 medical and health sciences
0302 clinical medicine
Internal medicine
medicine
Research Articles
adipose tissue‐derived mesenchymal stem cells
Pdx1
Chemistry
Chinese hamster ovary cell
Insulin
Transfection
030104 developmental biology
Endocrinology
Cell culture
embryonic structures
PDX1
insulin‐producing cells
Stem cell
Research Article
Subjects
Details
- ISSN :
- 22115463
- Volume :
- 8
- Database :
- OpenAIRE
- Journal :
- FEBS Open Bio
- Accession number :
- edsair.doi.dedup.....11187eaf0ea0b4ed2a915c4dcd6b2e56