The Past, Present, and Potential of Mesenchymal Stem Cells

The mesenchymal stem or stromal cells (MSCs) are integral parts of research history. Herein, we cover how therapeutic research has evolved alongside the timeline of MSCs research development.

In 1966, Prof. Friedenstein discovered that transplanted bone marrow cells could differentiate into osteoblasts in vivo. In 1968, Friedenstein proposed the existence of osteoblastic and hematopoietic progenitor cells, which was further confirmed by experimentation on animal models in 1971. Fibroblast-like cells were isolated from bone marrow in 1974, cell colonies of which differentiated into osteoblasts and supported formation of hematopoietic clones. In 1988, the bone marrow stromal cells were designated as osteogenic stem cells.

Osteogenic stem cells remain capable of cell division even after 20-30 multiplication and proliferation cycles, and can still form bone tissue after implantation in the peritoneum of rats. Notably, in 1987, Dr. Charbord discovered that bone marrow fibroblasts and bone marrow stromal cells (osteogenic stem cells) are two separate entities with similar morphological characteristics. Dr. Charbord found that bone marrow stromal cells differed from bone marrow fibroblasts in the uptake of serum proteins, thus making it clear that MSCs were not fibroblasts, although the cell morphology and fibroblasts are very similar in vitro culture. The key difference is that the MSC-derived osteogenic stem cells can differentiate into both osteoblasts and chondroblasts.

Osteoprogenitors were also identified in 1984 based on the size of the nucleus, which was later identified as stromal cells in 1990. In 1991, cultured MSCs obtained from Wharton jelly of umbilical cord were demonstrated to pass through six generations and release collagen and cytokeratin. In the same year, Dr. Charbord assessed the phenotypes of MSCs and identified cell surface markers, including HLA-DR, CD11c, vWF, CD68, CD45, CD51, and fibronectin. Dr. Charbord and Dr. Simmons were the first two scientists who focused on cell surface markers of MSCs.Although Friedenstein recognized that MSCs were involved in a component of the hematopoietic microenvironment to support hematopoiesis in 1974, it was not until 1992 that MSCs were clearly proposed as trophoblast cells shown to support the differentiation of hematopoietic stem cells into megakaryocytes, granulocytes, and macrophages by secreting CSF-1, G-CSF, GM-CSF, IL-6, IL-3, and other growth factors.   

In 1992, Dr. Owen demonstrated the ability of MSCs to differentiate into osteoblasts and adipocytes. Differentiation of both osteoblasts and adipocytes was inhibited when dexamethasone was added to the culture medium. When dexamethasone and 1, 25-dihydroxyvitamin D3 are added to the culture medium, the growth of adipocytes is inhibited while differentiation of osteoblasts is enhanced. It was first demonstrated in a 1999 Science article that MSCs can simultaneously differentiate into chondrocytes, adipocytes, and osteoblasts. It is this article that ignited everyone’s enthusiasm for MSCs research, focusing on the various differentiation functions of MSCs, expecting MSCs to play a huge role in regenerative medicine.

In 1999, it was discovered that the number of bone marrow MCSs declined with age in humans. However, the same phenomenon was observed in rats in 1992. The number of MSCs in the bone marrow of older rats was less than that of younger rats, that is, older rats had fewer fibroblast colony-forming units (FCF-U) and fewer adherent cells.

Unlike other researchers, Prof. Caplan studied MSCs that were derived from the bone marrow of chicken embryos. Owing to their origin from the embryo, the cells were named “mesenchymal stem cells“. He published 3 consecutive articles in 1994, demonstrating the therapeutic prospects of autologous MSCs in bone and cartilage tissue regenerative medicine.

Initial Clinical Research Trials

In 1995, the first clinical research results of MSCs were reported.  These adherent stromal cells were extracted from the bone marrow of patients suffering from malignant hematologic diseases, isolated and cultured, then reinfused into these patients to observe the clinical effect and evaluate the safety of these cells. This historical  milestone highlighted the first attempt at understanding the clinical implications of bone marrow MSCs.

Dr. Darwin proposed MSCs to be bone marrow stromal cells in 1997, followed by his suggestion that these cells could serve as vectors in gene therapy in 1998. In 2000, MSCs were obtained from umbilical cord blood with subsequent isolation of these cells in different tissues, including adipose, thymus, placenta, gingiva, fetal blood, and fetal lung. The multiple differentiation potential of these cells was demonstrated by a study conducted in 2000. In this study, the human MSCs were injected into the sheep fetuses. Within the sheep fetus, these human MSCs differentiated into different cell lines such as chondrocytes, thymic stromal cells, adipocytes, bone marrow stromal cells, and cardiomyocytes.

In 2002, it was suggested that the size of MSCs determines the quality of these cells. This can be elaborated by the fact that MSCs with smaller cytosomes tend to have the highest adipogenic potential and are capable of generating single-cell colonies. Additionally, the immunosuppressive capability of MSCs was discovered in 2002. Subsequently, in 2003 Dr. Le Blanc, demonstrated that the MSCs are hypoimmunogenic and do not elicit immune rejection in either allogeneic or cross-species applications. In a clinical study (Lancet, 2004), Dr. Le Blanc demonstrated the use of MSCs for treating graft resistance host disease (GVHD). This is the first clinical research article using the immunosuppressive ability of MSCs to exert therapeutic effects. Extensive immunosuppressive function can be said to be a unique function of MSCs, which has the function of inhibiting activation of various immune cells. Treg cells with similar functions are restricted to T lymphocytes. The discovery of the immunosuppressive function of MSCs has also greatly promoted the clinical research of MSCs for the treatment of immunoreactive diseases.

The MSCs were termed skeletal stem cells by professor Paolo Bianco in 2004, as it was proven through transgenic mice that the continuously elevated parathyroid hormone promotes the differentiation of MSCs into osteocytes and osteogenesis through the parathyroid hormone-related peptide receptor. Bianco elaborated on the idea, emphasizing that the concept of ‘skeletal stem cells’ is closely related to the correct clinical application.

As of 2005, the International Society for Cellular Therapy proposed that MSCs should be called multipotent mesenchymal stromal cells, as the organization proposed that these cells do not have the characteristic properties of stem cells (being able to differentiate in vivo). In 2006, the criteria for classifying multipotent mesenchymal stromal cells were proposed and given as follows:

1.Walled (adherent) structure

2.Specific cell phenotype or surface antigen expression

3. Potential of the cell to differentiate into chondrocytes and adipogenic osteoblasts

In 2005, a decline in the immunosuppressive potential of bone marrow MSCs (BMSCs) was observed in patients with aplastic anemia, followed by the discovery of a similar trend in individuals suffering from ITP, rheumatoid arthritis, multiple myeloma, and systemic lupus erythematosus. Alongside impaired immunosuppressive ability of BMSCs, changes such as slowed proliferation and decreased secretion of cytokines were also observed.

In July 2011, The Korean Food and Drug Administration approved the marketing of an MSC drug known as Herticellgram-AMI for the indication of acute myocardial infarction. This became the first stem cell therapy created from autologous BMSCs to treat acute myocardial infarction. Pharmaceutical companies including Medi-post began the production of Cartistem in January 2012, which is a cartilage regeneration drug. At the same time, another company named Anterogen, received approval for the production of Cuepistem for the treatment of anal fistula. The MSCs were derived from bone marrow and adipose tissue in Cartistem and Cuepistem treatment, respectively.  

In 2012, Canada approved the use of Prochymal, an injectable MSC preparation formulated by Osiris Therapeutics in the 1990s. However, the company announced the failure of the phase III clinical trial in children with GVHD treated with Prochymal. The reason behind this failure is attributed to the use of liquid nitrogen frozen transport, which had negative effects on the immune modulation potential of MSCs.

The results of a second phase III of clinical trials that used adipose MSCs, CX601 (Alofisel), for treating complex perianal fistula clonorchiasis were reported in The Lancet in 2016. Results reporting the long-term efficacy of this treatment were published in the year 2018.

Pericytes were discovered in 1990 by Dr. Diaz Flores as cells present in periskeletal and perivascular regions. These cells promote tissue repair and are capable of differentiation into chondrocytes and osteoblasts. Dr. Bianco used a laser confocal technique to identify the bone marrow stromal cells that were localized around the vessels.

In 2006, it was found that MSCs are present in the stroma of organs and tissues around the blood vessels. In 2008, Dr. Bruno suggested that the MSCs derived from pericytes have similar cell surface antigens and pluripotency. The same finding was proposed by Dr. Caplan in the same year. The two ideas, ‘Hit and Run’ and ‘Touch and Go,’ emphasize that the body rapidly clears MSCs via migration into damaged organs and the secretion of therapeutic molecules.

What is the Association Between the Pericytes and MSCs in Developmental Embryology? 

In 2009, researchers proposed that the pericytes in the umbilical cord are a subpopulation of MSCs in the umbilical cord. The researchers also suggested that the MSCs transform into fibroblasts after a long duration of culture and ultimately lose their ability to divide into other cell types such as adipocytes and osteoblasts. The MSC’s heterogeneity remained unclear. In the following year, Prof. Caplan proposed a new name for MSCs, calling them Medical Signaling Cells, as these cells release immunomodulatory and nutrition-related bioactive factors.

In 2017, it was proposed that the MSCs protect the bone marrow against melanoma metastasis. Regardless of the theoretical and clinical advancements, the dispute concerning the nomenclature of MSCs remains.

Summary

The MSCs can be found on the external surface of blood sinusoids, a typical type of blood vessel in the bone marrow. These cells originate as osteoblasts and ultimately form the bone marrow niche or microenvironment for hematopoietic stem cells (HSCs), which have a longer research history than MSCs. The hematopoietic stem cells and MSCs vary in their functional characteristics and treatment methods. This comparison can be further elaborated by an example that hematopoietic stem cell transplantation involves the removal of diseased cells before the procedure, a process known as myeloablation. However, there is no established method for the removal of diseased MSCs to make room for new cells.

Given that MSCs do not express MHC-II antigens, have low immunogenicity, and can suppress immune responses, some experts believe that MSCs are immune-privileged cells. However, a large number of animal experiments and limited clinical studies have shown that the imported allogeneic MSCs cannot persist in vivo for a long time, according to data from animals and humans after childbirth. Since MSCs cannot exist for a long time in vivo, MSCs cannot differentiate into progeny cells to play a cell replacement role, which means that differentiation is not the therapeutic mechanism of MSCs. This is why Prof. Caplan emphasized that MSCs are not stem cells. Even if differentiation is not the therapeutic mechanism of MSCs, the stemness (self-renewal and differentiation in vivo) of MSCs should not be denied, because MSCs have the functions of self-renewal and differentiation in vivo as discussed above.

For over 15 years, Cyagen has been involved in the field of stem cell research, providing products with remarkable proliferation and differentiation potential that have assisted researchers and investigators in achieving better results.

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