טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
M.Sc Thesis
M.Sc StudentLior Limor
SubjectEffect of Placenta Derived Mesenchymal Stem Cells Cultured
in Bioreactor, on Engraftment of Human
Hematopoietic Stem Cells Derived from
Umbilical Cord Blood in NOD-
DepartmentDepartment of Biotechnology
Supervisors Ms. Erella Livne (Deceased)
Dr. Anna Weiss
Full Thesis text - in Hebrew Full thesis text - Hebrew Version


Abstract

Bone marrow transplantation (BMT) has been a standard of care for the last forty years. However, only 30% of candidates eligible for hematopoietic stem cells (HSC) transplantation will have such a donor available. Therefore, alternative sources of HSC are being investigated. Among those the umbilical cord blood (UCB) possesses several advantages, such as abundant availability, no risk to the donor and naïve nature that enables the use of HLA-mismatch grafts without a high risk and severity of GVHD related to BMT. However, its main disadvantage is limited number of HSC, leading to higher rates of graft failure and delayed time to engraftment in comparison to BMT. Mesenchymal stem cells (MSC) co-transplanted with HSC are known for supporting and enhancing hematopoietic grafts.

The aim of this study was to examine the effect of co-transplantation of human placenta mesenchymal stem cells (PMSC), on the engraftment of human HSC derived from UCB, in mice. The placenta is a relatively new source of MSC, easily available and does not involve invasive procedures or risk to the donor.

PMSC exhibited mesenchymal features, including fibroblast-like cell morphology, expression of membrane markers, common to MSC such as CD90, CD105, CD29 and CD73 and lack expression of hematopoietic membrane markers, such as CD45, CD34, CD19, CD33, CD14, HLA-DR and CD11b. They secreted hematopoietic cytokines as TPO, SCF, IL6, GM-CSF and Flt and exhibited ability for differentiation into adipocytes, chondrocytes and osteoblasts.

PMSC cells were expanded on carriers in 3D culture bioreactor, which enables growth of dense cultures for obtaining massive cell yield.

The ability of PMSC cells to support hematopoietic engraftment was examined using an animal model of non-obese diabetic severe combined immunodeficient (NOD/SCID) mice, deficient in production of T and B cells. HSC from UBC were transplanted alone or co-transplanted with PMSC, 24 hrs after treating the mice by irradiation or chemotherapy. 

Six weeks after transplantation, human HSC engraftment levels, detected by FACS as percentage of cells expressing the human leukocyte marker CD45, were significantly higher in bone marrow of mice co-transplanted with human HSC and PMSC, compared to HSC only. The human graft reconstituted all blood lineages: myeloid, lymphoid, red blood cells and platelets.

Summary: co-transplantation of PMSC expanded in 3D culture in bioreactor with UCB HSC promotes the latter engraftment. Thus, it might be an effective tool to overcome the amount limitations using UCB and allow its use as an alternative to BMT.