In vitro differentiation of mouse bone marrow derived mesenchymal stem cells into islet of Langerhans-like cells

Other Title(s)

تمايز الخلايا الجذعية اللحمية المشتقة من نخاع عظم الفار إلى الخلايا الشيبة بجزر لانكرهانس خارج الجسم الحي

Dissertant

al-Qaysi, Bayda Amir Ahmad

Thesis advisor

al-Rikabi, Nahi Yusuf Yasin
al-Alujy, Sabah Nasir

Comitee Members

al-Ubaydi, Salim Rashid
Yanzil, Jabbar H.
Yusuf, Walid Hamid
Rahmah, Abbas Mahdi
Adhiah, Ali Husayn

University

University of Baghdad

Faculty

College of Science

Department

Department of Biology

University Country

Iraq

Degree

Ph.D.

Degree Date

2013

English Abstract

Diabetes mellitus (DM) is an insulin dependent and autoimmune disorder.

The cell therapy for this disorder regarded as a treatment approach option because of the inability to find a medicinal definitive cure for it.

Bone marrow-derived mesenchymal stem cells (BM-MSCs) with ability to transdifferentiate into Langerhans islet-like cells (ILCs) would provide an unlimited source of pancreatic endocrine cells for transplantation.

Although the culture of MSCs has been studied for over 30 years.

Standard protocol for isolation and characterization have yet to be developed because MSCs represent the mainstay of transdifferentiation.

A comparative study was made on three different isolation techniques, which include direct plating, red blood lysis buffer strategies, and density gradient (DG) method to find the optimal isolation and culture condition for adequate amount of MSCs.

The results demonstrate that direct plating of whole bone marrow (BM) suspension provided a suitable alternative for isolation of BM-MSCs, which mainly based on the frequent medium change in primary culture.

The BM cell suspension were isolated by aspiration from albino mice and cultured in fresh complete isolation media (CIM) RPMI-1640/10%FBS.

Adherent cells from BM cell suspension were MSCs which then expanded by complete expansion media (CEM) α-MEM/ 10% FBS.

Immunophenotypic analysis demonstrated that mouse BM-MSCs at passage three uniformly positive for CD44, CD90, CD105, CD106, and low level of nestin positive subpopulation.

However, MSCs were always found to be negative for heamatopoietic specific markers CD34, CD45, and endothelial marker CD31.

To explore the possibility of BM-MSCs in vitro transdifferentiating into functional ILCs, the study developed four-step protocol using two different culture II methods which include; routine two –dimensional (2D) and three-dimensional (3D) pellet culture systems.

The identity of the ILCs in both systems were illustrated by the analysis of morphology using haematoxlyin and eosin (H & E), and immunocytochemistry.

The release of insulin by these cells was confirmed by Dithiozone staining (DTZ) and linked Immunosorbent assay (ELISA).-the closest to reality of islet It can be concluded from the present study that mouse BM-MSCs can successfully isolated by direct plating and transdifferentiated into functional ILCs under certain conditions in vitro developed in this study which may reveal a new procedure for clinical diabetes stem -cell therapy.

Main Subjects

Biology

Topics

• Bone marrow
• Stem cells

No. of Pages

204

Table of Contents

• APA
• MLA
• AMA

Language

English

Data Type

Arab Theses

Record ID

BIM-598983