ABSTRACT
@#Introduction: Monocytes are essential phagocytic cells of the innate immune system as they are required for the maintenance of tissue homeostasis. However, accumulation of monocytes is implicated in various chronic inflammatory diseases like coronary heart disease, atherosclerosis and in autoimmune disorders. Therefore, the number of monocytes must be carefully regulated to avoid monocyte induced inflammatory disorders. Mesenchymal stem cells (MSCs) have shown to be effective against various inflammatory diseases due to their immunosuppressive properties. The present study was designed to evaluate the less understood immunomodulatory effect of MSCs on monocyte proliferation and survival. Method: Primary monocytes were isolated from peripheral human blood using CD14+ monocyte isolation kit. The in house produced umbilical cord MSCs were co-cultured with monocytes at different ratio and time; assessed for the monocyte viability, proliferation and cell cycle. Results: Mesenchymal stem cells suppressed monocyte proliferation in a dose-dependent manner. The antiproliferative effect of MSCs was mediated by cell cycle arrest, whereby monocytes were arrested in the G0/G1 phase of the cell cycle by preventing them from progress into S and G2/M phases. Although cell cycle arrest could potentially lead to apoptosis; however, MSCs significantly enhanced the monocytes survival and inhibited apoptosis. Conclusion: Human MSCs inhibit the stimulated monocyte proliferation without inducing cellular apoptosis at in vitro. These results reveal that MSCs can be utilised to control monocytes’ quantity during an unwanted immune response to maintain homeostasis.
ABSTRACT
Mesenchymal stem cells (MSCs) derived from human umbilical cord (UC) have been considered as an important tool for treating various malignancies, tissue repair and organ regeneration. Umbilical cord-derived mesenchymal stem cells (UC-MSCs) are better alternative to MSCs that derived from bone marrow (BM-MSCs) as they are regarded as medical waste with little ethical concern for research and easily culture-expanded. In this present study, the foetal distal end of human UC was utilised to generate MSC by explant method. Upon in vitro culture, adherent cells with fibroblastic morphology were generated with rapid growth kinetics. Under the respective inductive conditions, these cells were capable of differentiating into adipocytes and osteocytes; express an array of standard MSC’s surface markers CD29, CD73, CD90, CD106 and MHC-class I. Further assessment of immunosuppression activity revealed that MSCs generated from UC had profoundly inhibited the proliferation of mitogen-activated T lymphocytes in a dosedependent manner. The current laboratory findings have reinforced the application of explant method to generate UCMSCs thus, exploring an ideal platform to fulfil the increasing demand of MSCs for research and potential clinical use.
Subject(s)
Mesenchymal Stem CellsABSTRACT
Introduction: During the last three decades hematopoietic stem cell transplantation (HSCT) has become a well-established treatment for many hematologic malignancies. The most important limitation for HSC transplantation is the low number of hematopoietic stem cells (HSC) that can lead to delayed engraftment or graft failures. Numerous attempts have been made to improve in vitro HSC expansion via optimization of various methods such as isolation techniques, supplementing with growth factors, utilizing stromal cells as feeder layer and other culture conditions. Objective: This project is aimed to decipher the efficiency of an isolation technique and retrieval of culture expanded HSC from feeder layer using two different harvesting methods. Materials and Methods: Hematopoietic stem cells from human umbilical cord blood were isolated via MACS mediated CD34+ double sorting. Then, the cells were cultured onto MSC feeder layer for 3 and 5 days. Culture expanded cells were harvested using two different harvesting method namely cell aspiration and trypsinization methods. Hematopoietic stem cell expansion index were calculated based on harvesting methods for each time point. Results: The numbers of HSC isolated from human umbilical cord blood were 1.64 x 106 and 1.20 x106 cells at single and double sortings respectively. Although the number of sorted cells diminished at the second sorting yet the yield of CD34+ purity has increased from 43.73% at single sorting to 81.40% at double sorting. Employing the trypsinization method, the HSC harvested from feeder layer showed a significant increase in expansion index (EI) as compared to the cell aspiration harvesting method (p≤ 0.05). However, the purity of CD34+ HSC was found higher when the cells were harvested using aspiration method (82.43%) as compared to the trypsinization method (74.13%). Conclusion: A pure population of CD34+ HSC can be retrieved when the cells were double sorted using MACS and expanded in culture after being harvested using cell aspiration method.