By Bryan Peacker, ’18

Introduction

In the past fifteen years, stem cells have risen to the forefront of the development of promising new therapies, but not without controversy. From news of illegal stem cell clinics spuriously claiming to be able to cure countless diseases through unproven transplants to the notorious 2014 paper in Nature that falsified data and purported to have created a trailblazing and simple method of deriving stem cells, the attention that stem cells have garnered in the past few years is well-deserved (Okobata et al., 2014). Despite numerous ongoing clinical trials, stem cell therapies are far from becoming pervasive due to both ethical and technical considerations. In 2008, scientists at University of Louisville claimed to have found new stem cell population in adults called very small embryonic-like stem cells (VSELs), which could act as an alternative to the controversial use of human embryonic stem cells for therapy (Ratajczak et al., 2008). However, their very existence has been called into question, and the lack of consensus in the scientific community about these tiny cells remains to be resolved.

What Are Stem Cells?

In mammals, most cells in the body are specialized, and can only perform a specific role. For instance, if liver cells were to be transplanted to the pancreas with the intent of replacing pancreatic beta cells, the transplanted cells would not be able to produce insulin. Although this may sound trivial, it turns out that some cells can specialize into cells from any tissue. These cells are called pluripotent, for their potential to change into any cell type, and the process of becoming specialized is called differentiation (National Institutes of Health, 2015). A cell can only be considered a stem cell, however, if it can both become tissue- or organ-specific and replenish itself indefinitely through cell division, according to the National Institutes of Health (2015). Thus, a stem cell, when kept under the right conditions, can potentially keep dividing and make more of itself forever. This makes stem cells attractive candidates for regenerative therapies and for developing models of diseases.

Where can stem cells be obtained? Although there are very small numbers of minimally differentiated stem cells in adults, most stem cells are either obtained from embryos or induced into pluripotency from adult cells. When a mammalian egg is fertilized and becomes an embryo, all cells in the very early stages of that embryo are pluripotent, meaning that they can differentiate into any cell type that is eventually found in the adult. This was the first source of stem cells discovered, and the first pluripotent stem cells to ever be isolated were taken from mouse embryos in 1981 (Martin, 1981). These cells are called embryonic stem cells (ES cells), but the embryo must be destroyed in order to obtain them, which brings about serious ethical considerations, particularly in humans. A second way to obtain stem cells is to use viruses to introduce DNA into the cell, so that the cell produces factors that have been shown to induce pluripotency. This procedure effectively reprograms adult differentiated cells into pluripotent cells, and cells derived in this manner are appropriately called induced pluripotent stem cells (Takahashi & Yamanaka, 2006). This Nobel Prize-winning technique was lauded for circumventing the ethical repercussions of destroying embryos, but the viruses used for the induction can sometimes lead to cancer, making induced pluripotent stem (iPS) cells difficult to utilize for therapies (National Institutes of Health, 2015).

A Surprising Discovery

For several decades, it has been widely-known that there are stem cells present in very small numbers in adult mammals, most notably the bone marrow (Maximow, 1909). However, the stem cells of the bone marrow and all adult stem cells are multipotent as opposed to pluripotent, meaning that they can differentiate into only cells within a specific tissue, rather than into any cell lineage in the body (National Institutes of Health, 2015). However, in 2008, researchers purported to have discovered a pluripotent stem cell line called very small embryonic like stem cells (VSELs) isolated from bone marrow (Kucia et al., 2006). Subsequent studies found these same cells in the pancreas, testes, and heart (Kucia et al., 2008). These cells make up about 0.01% of cells in the bone marrow, but can be isolated and expanded in cell culture, and can differentiate into the three germ layers of the early embryo, a promising initial sign of pluripotency (Ratajczak et al., 2008).

Most surprisingly, the same laboratory found VSELs circulating in the bloodstream of mice, which led the authors to hypothesize that this is the body’s way to relocate stem cells to different tissues when needed (Kucia et al., 2008). In addition, levels of VSELs increase following organ injury, suggesting that these cells play a role in repair in mice (Kucia et al., 2008). This finding indicates that VSELs are ideal for regenerative therapy, since the in vivo purpose of these cells appears to be for autologous repair. In support of these findings, in 2015 scientists in France claimed to have isolated VSELs in young, middle-aged, and aged human subjects, thus extending the implications to humans (Solovat et al., 2015).

The advantages of VSELs over both embryonic stem cells and induced pluripotent stem cells are clear. While embryonic stem cells face both ethical dilemmas and the risk immune rejection upon transplantation, VSELs can be obtained from the patient without ethical problems. Although iPS cells also do not face either of these issues, reprogramming iPS cells involves the random integration of viral DNA into the target cell genome, which can lead to irreversible and deleterious consequences such as cancer or genetic diseases (Medvedev et al., 2010). VSELs don’t face any of these problems. In practice, VSELs could be isolated from a patient, expanded in the laboratory, and then re-injected into the patient for regenerative therapy without immune rejection and with minimal manipulation (Solovat et al., 2015).

Controversy

An important consideration that appears to have been ignored in the papers that Ratajczak’s group published about VSELs is that pluripotency has a very strict definition in developmental biology, and only if a cell fulfills the most stringent tests can it be considered to be pluripotent. Although VSELs can differentiate into all three germ layers in vitro and although they express pluripotency markers, they cannot contribute to a developing embryo (Ratajczak et al., 2008). This is an example of a stringent test of pluripotency, and failing it would normally mean that the cells cannot be called pluripotent, but nonetheless Ratajczak and others continue to call them pluripotent.

Apart from the lack of rigor in the published papers about VSELs themselves, a more direct blow to the promises of VSELs came from the lab of Irving Weissman at Stanford University. After performing numerous experiments using the methods purported to have been used to isolate VSELs, his lab failed to replicate previously reported results, and showed that in some cases, the cells regenerating tissues were not VSELs, but rather a different type of stem cell population that is multipotent, and not pluripotent (Miyanishi et al., 2013). Another lab in Poland claimed to have been unable to find the markers for pluripotency that Ratajczak’s group had found in VSELs, despite attempting to replicate the protocol that Ratajczak provides in his published papers (Szade et al., 2013). Ratajczak and collaborators, however, claim that these groups lack the technical skills to isolate the correct group of cells, and remain adamant about the existence of VSELs (Abbott, 2013). This controversy over VSELs continues, and while some groups have stopped ongoing research on VSELs, others have continued. For instance, researchers at the National Institute for Research in Reproductive Health in India recently claimed that VSELs are involved in partial pancreatic regeneration, and that the loss of function in VSELs with age may lead to diabetes and cancer (Bhartiya & Patel, 2015).

Whether VSELs are pluripotent or even exist in the first place remains hotly debated, but the issue elucidates an important facet of scientific research. Even when results are published in peer-reviewed journals, they are not definitive, and what drives the community forward is the practice of being critical and maintaining rigor. If VSELs truly do have the potential to be utilized for regenerative therapies, then continued efforts to study them will not have been in vain, and a major leap in stem cell therapy will be gained. If in the near future the consensus becomes that VSELs are not stem cells, then scientific criticism will have prevailed, and scientific community will still be able to revel in success, knowing that rigor continues to drive the field forward.

References

Abbott, A. (2013, July 24). Doubt cast over tiny stem cells. Nature News. Retrieved from http://www.nature.com/news/doubt-cast-over-tiny-stem-cells-1.13435.

Bhartiya, D., & Patel, H. (2015). Very small embryonic-like stem cells are involved in pancreatic regeneration and their dysfunction with age may lead to diabetes and cancer. Stem cell research & therapy6(1), 96.

Kucia, M., Reca, R., Campbell, F. R., Zuba-Surma, E., Majka, M., Ratajczak, J., & Ratajczak, M. Z. (2006). A population of very small embryonic-like (VSEL) CXCR4+ SSEA-1+ Oct-4+ stem cells identified in adult bone marrow.Leukemia20(5), 857-869.

Kucia, M. J., Wysoczynski, M., Wu, W., ZubaSurma, E. K., Ratajczak, J., & Ratajczak, M. Z. (2008). Evidence That Very Small EmbryonicLike Stem Cells Are Mobilized into Peripheral Blood.Stem Cells26(8), 2083-2092.

Martin, G. R. (1981). Isolation of a pluripotent cell line from early mouse embryos cultured in medium conditioned by teratocarcinoma stem cells. Proceedings of the National Academy of Sciences,78(12), 7634-7638.

Maximow, A. (1909). The lymphocyte as a stem cell, common to different blood elements in embryonic development and during the post-fetal life of mammals. Folia Haematologica8, 123-134.

Medvedev, S. P., Shevchenko, A. I., & Zakian, S. M. (2010). Induced pluripotent stem cells: problems and advantages when applying them in regenerative medicine. Acta naturae2(2), 18.

Miyanishi, M., Mori, Y., Seita, J., Chen, J. Y., Karten, S., Chan, C. K., … & Weissman, I. L. (2013). Do pluripotent stem cells exist in adult mice as very small embryonic stem cells?. Stem cell reports1(2), 198-208.

National Institutes of Health, U. S. Department of Health and Human Services. (2015). Stem Cell Basics: Introduction. In Stem Cell Information. Retrieved from http://stemcells.nih.gov/info/basics/pages/basics1.aspx.

Obokata, H., Wakayama, T., Sasai, Y., Kojima, K., Vacanti, M. P., Niwa, H., … & Vacanti, C. A. (2014). Stimulus-triggered fate conversion of somatic cells into pluripotency. Nature505(7485), 641-647.

Ratajczak, M. Z., Zuba-Surma, E. K., Wysoczynski, M., Ratajczak, J., & Kucia, M. (2008). Very small embryonic-like stem cells: characterization, developmental origin, and biological significance. Experimental Hematology,36(6), 742-751.

Sovalat, H., Scrofani, M., Eidenschenk, A., & Hénon, P. (2015). Human Very Small Embryonic-Like Stem Cells Are Present in Normal Peripheral Blood of Young, Middle-Aged, and Aged Subjects.Stem cells international2016.

Szade, K., Bukowska-Strakova, K., Nowak, W. N., Szade, A., Kachamakova-Trojanowska, N., Zukowska, M., … & Dulak, J. (2013). Murine bone marrow Lin− Sca-1+ CD45− very small embryonic-like (VSEL) cells are heterogeneous population lacking Oct-4A expression. PLoS One8(5), e63329.

Takahashi, K., & Yamanaka, S. (2006). Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors.cell126(4), 663-676.

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