Observational Study
Copyright ©The Author(s) 2015. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Clin Cases. Jul 16, 2015; 3(7): 640-649
Published online Jul 16, 2015. doi: 10.12998/wjcc.v3.i7.640
In vitro differentiation of human umbilical cord Wharton’s jelly mesenchymal stromal cells to insulin producing clusters
Seideh Masoomeh Nekoei, Negar Azarpira, Ladan Sadeghi, Sulmaz Kamalifar
Seideh Masoomeh Nekoei, Negar Azarpira, Transplant Research Center, Shiraz University of Medical Science, Shiraz 7193711351, Iran
Ladan Sadeghi, Sulmaz Kamalifar, Islamic Azad University, Arsanjan Branch, Shiraz 7193711351, Iran
Author contributions: Nekui SM and Azarpira N were participated in study design; Nekui SM did all experiments with data gathering and statistical analysis; all authors were participated in manuscript preparation and scientific writing; Nekui SM wrote the manuscript with valuable assistance of Azarpira N.
Supported by A grant from Iran National Science Foundation (INSF).
Institutional review board statement: The study design was approved by ethical committee of Shiraz University of Medical Sciences, Shiraz, Iran.
Informed consent statement: Informed consent was received from all patients that participated in this study.
Conflict-of-interest statement: All authors declared that they have no conflict of interest.
Data sharing statement: Technical appendix, statistical code, and dataset available from the corresponding author at Dryad repository, who will provide a permanent, citable and open-access home for the dataset.
Open-Access: This article is an open-access article which was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See: http://creativecommons.org/licenses/by-nc/4.0/
Correspondence to: Negar Azarpira, MD, Transplant Research Center, Shiraz University of Medical Science, Zand Street, Shiraz 7193711351, Iran. negarazarpira@yahoo.com
Telephone: +98-711-6473954 Fax: +98-711-6473954
Received: August 10, 2014
Peer-review started: August 14, 2014
First decision: September 14, 2014
Revised: April 23, 2015
Accepted: May 16, 2015
Article in press: May 18, 2015
Published online: July 16, 2015
Processing time: 351 Days and 13.7 Hours
Abstract

AIM: To investigate the differentiation of human Wharton’s jelly derived mesenchymal stromal cells (WJ-MSCs) to insulin producing clusters (IPC) this study was conducted.

METHODS: The umbilical cords samples were collected from full term caesarian section mothers and the WJ-MSCS were cultured from tissue explants in High glucose-Dulbecco’s Modified Eagle Medium (H-DMEM); H-DMEM supplemented with 10% fetal bovine serum (FBS) and antibiotics. The expression of CD90, CD44, CD105, CD34 and CD133 as well as osteogenic and adipogenic differentiation of cells in appropriate medium were also evaluated. The cells were differentiated toward IPC with changing the culture medium and adding the small molecules such as nicotinic acid, epidermal growth factor, and exendin-4 during 3 wk period. The gene expression of PDX1, NGN3, Glut2, insulin was monitored by reveres transcription polymerase chain reaction method. The differentiated clusters were stained with Dithizone (DTZ) which confirms the presence of insulin granules. The insulin challenge test (low and high glucose concentration in Krebs-Ringer HEPES buffer) was also used to evaluate the functional properties of differentiated clusters.

RESULTS: WJ-MSCS were positive for mesenchymal surface markers (CD90, CD44, CD105), and negative for CD34 and CD133. The accumulation of lipid vacuoles and deposition of calcium mineral in cells were considered as adipogenic and osteogenic potential of WJ-MSCS. The cells also expressed the transcriptional factors such as Nanog and OCT4. During this three step differentiation, the WJ-MSCS morphology was gradually changed from spindle shaped cells in to epithelioid cells and eventually to three dimensional clusters. The clusters expressed PDX1, NGN3, Glut2, and insulin. The cells became bright red color when stained with DTZ and the insulin secretion was also confirmed. In glucose challenge test a significant increase in insulin secretion from 0.91 ± 0.04 μIu/mL (2.8 mmol/L glucose) to to 8.34 ± 0.45 μIu/mL (16.7 mmol/L glucose) was recorded (P < 0.05). The insulin secretion of undifferentiated WJ-MSCS was not changed in this challenge test.

CONCLUSION: WJ-MSCs have the ability to differentiate in to islet-like cells in vitro. However, this process needs further optimization in order to generate efficient and functional IPCs.

Keywords: Mesenchymal stromal cells; Umbilical cord; Beta cell; Islet

Core tip: Diabetes is a major chronic metabolic disorder in the world. Mesenchymal stromal cells (MSCs) has the ability to differentiate in to functional insulin producing cells.In this study, human Wharton’s jelly derived MSCs (The clusters expressed PDX1, NGN3, Glut2, and insulin. The cells became red color when stained with DTZ and the insulin secretion was confirmed Wharton’s jelly derived MSCs were differentiated to insulin producing clusters (IPCs). More efficient differentiation protocoles for generation of functional IPCs will be a potential new source for cell transplantation in diabetic patients.