A Novel Approach for Expansion of High Quality Mesenchymal Stem Cells

By on April 18, 2017

A Guest Blog by Iris Flores, Business Development Manager, StemBioSys, Inc.

Human mesenchymal stem cells (hMSCs) are a promising tool for therapeutic applications in cell-based therapy and regenerative medicine. Increasing evidence has shown that MSCs represent new hope for treating, and in some cases, perhaps even curing human diseases like diabetes, Alzheimer’s, Parkinson’s, Muscular Dystrophy , inflammatory disorders, osteoarthritis and damaged cartilage, to name a few.   And although stem cells have emerged as the future of health care there are still a number of unmet needs, which must be addressed before widespread clinical and therapeutic applications become a reality. One of the major challenges in cell therapy is obtaining sufficient numbers of quality stem cells while maintaining their differentiation potential. BM-HPME® (Bone Marrow – High Performance Micro Environment) overcomes key obstacles to capturing and growing stem cells without losing potency.

Mesenchymal stem cells (MSCs) have been investigated extensively for their considerable immunomodulatory potential and in vitro studies typically access cell behavior on coatings primarily consisting of single purified proteins, protein components, or even directly on tissue culture treated plastic (TCP). Culturing cells on TCP not only introduces cells to a foreign environment, but requires the cells to expend precious energy on producing a provisional cell matrix on which they can grow more naturally.

The gene expression maps in the figure below (Figure. 1) clearly show that cells are behaving much differently on a natural substrate, the BM-HPME®, than when they are grown on plastic.

Figure 1. Gene expression map comparing MSCs grown on BM-HPME® versus TCP

 

Why BM-HPME®?

Multiple studies have shown that BM-HPME® provides a higher quantity of usable stem cells, that are small, more proliferative, and less differentiated when compared to cells grown on TCP and other comparable substrates. What makes BM-HPME® so original and unique is that it is the only stem cell substrate to closely mimic the full biochemical and structural microenvironment that stem cells normally see in their native environment. The microenvironment plays a critical role in stem cell behavior and lineage differentiation. In general, cells tend to maintain better function when grown on cell-derived matrices such as BM-HPME®, thus providing support for a more in vivo like approach. Placing cells onto a foreign or biosynthetic matrix elicits a rapid response from the cells, which is not consistent with attempting to expand cell numbers without inducing differentiation.

BM-HPME® at its core is a naturally produced extracellular matrix (ECM) of proteins synthesized in vitro by bone marrow stromal cells. This product is composed of more than 150 proteins all of which were secreted and assembled by the bone marrow cells during production. The final product is cell free with only the ECM attached to the surface of the culture vessel. This cell culture substrate provides a native three-dimensional microenvironment that can be used for rapid expansion of high quality (MSCs). Figure. 2 depicts the BM-HPME® process.

Figure 2. BM-HPME® Production Process

Stem cells thrive in this biological substrate. The naturally produced BM-HPME® allows for rapid and efficient expansion of stem cells which express biochemical and functional characteristics of potent cells and a gene expression profile highlighted by increased expression of anti-inflammatory cytokines.

Supporting Evidence

BM-HPME® promotes cell adhesion

As described above, BM-HPME® is unlike any other tissue culture surface. StemBioSys’ BM-HPME® is a natural 3-dimensional microenvironment produced by stem cells for stem cells. Multiple studies performed using BM-HPME® have exhibited efficient growth of stem cells opening up potential for research, diagnostic and clinical use. In the figures below (Figure. 3 and 4), mononuclear cells were isolated from human umbilical cord blood and seeded on both TCP (left) and BM-HPME® (right). Cells were incubated at 37°, 5% CO2, and 95% O2. Representative images were taken at 96 hours.

Figure 3. Mononuclear cells isolated from human umbilical cord on TCP Figure 4. Mononuclear cells isolated from human umbilical cord on BM-HPME®

BM-HPME® enhances MSC colony formation and response to chemical induction

Additional derterminants of stem cell potency are colony forming efficiency and the capability of multilineage differentiation. MSCs that were grown on TCP and BM-HPME® were subsequently tested for these determinants of stem cell potency. Cells grown on BM-HPME® were smaller in size and had much greater colony-forming efficiency compared to cells grown on TCP. Evaluation of the differentiation potential of MSCs for adipogenic and osteogenic lineages was also performed using Oil Red and von Kossa staining methods respecively. The images below reflect the differences in CFU efficiency and multilineage differentiation potential between MSCs grown on TCP compared to those grown on BM-HPME®. In Figure. 5, it is shown that cells grown on a more natural microenvironment such as BM-HPME® clearly exhibit better differentiation potential.

Figure 5 Differences in CFU efficiency and Mulilineage differentiation potential between MSCs grown on TCP versus BM-HPME®

BM-HPME® reduces Donor-Donor variability

MSC growth can vary significantly between donors and preparations. One of the most important parameters in clinical cell culture is the population doubling time as it corrleates to loss of potency and genomic instability. Drawing from our historic data we can see that MSCs grown on BM-HPME® have a shorter doubling time and also exhibit less variation in growth potential from donor to donor. This further illustrates the advantage of placing cells in a natural enviroment. See Figure 6.

Figure. 6 Doubling Time and Donor-Donor Variability

MSCs cultured on BM-HPME® are smaller, grow faster and retain expression of SSEA-4

Multiple studies performed using BM-HPME® have been shown to allow a variety of stem cells to replicate more rapidly and with greater preservation of stem cell phenotype. As part of a sponsored research agreement we examined the cell yield, cell diameter and Stage Specific Embryonic Antigen-4 (SSEA-4) expression of human mesenchymal stem cells (hMSCs) cultured on BM-HPME® and compared them to hMSCs grown on other substrates. See Figures 7-9 for the experimental results. (B., Langer, D. Anderson Labs, MIT).

MSCs cultured on BM-HPME® are smaller, grow faster and retain expression of SSEA-4.

Figure 7. hMSC Yield on various substrates afer 48 hours
Figure 8. Cell Diameter from hMSCs grown on various substrates
Figure 9. SSEA4 Expression from hMSCs grown on various substrates
*Note: There were not enough cells from Product 2 to analyze in Figure 9

BM-HPME® vs Traditional Tissue Culture Substrates

Stem cells do not normally encounter plastic in the body so it makes sense that they would not grow normally on plastic. BM-HPME® is designed for stem cells by stem cells. It is 100% biologic and closely mimics the natural microenvironment is which stem cells reside. ECM coated dishes provide cells with only a fraction of the proteins that are normally present in tissue. The BM-HPME® advantage is that cells can grow on an environment similar to that from which they came.

BM-HPME® A Flexible Culture Option

BM-HPME® is available in a variety of sizes. The 3D microenvironment supports the attachment and proliferation of all varieties of MSCs tested to date and the cells can be detached easily for downstream analysis. Multiple published studies on BM-HPME® have demonstrated enhanced proliferation and increased expression of key stem cell characteristics when compared to cells grown on TCP (Ragelle et al., Biomaterials, 2017).   Additionally, BM-HPME® arrives easy-to-use with only a short rehydration step. Once rehydrated you can immediately begin to culture your cells in the media of your choice. For more information on BM-HPME®, visit www.stembiosys.com/sales/product-specs

If you have questions about BM-HPME, please stop by and talk with Iris, who will be at the StemBioSys booth 1357 at Experimental Bio, April 22-26.


About the author:
Iris Flores, Business Development Manager, StemBioSys, Inc.

About the company:
StemBioSys (SBS) is a life sciences company located in San Antonio, Texas. SBS is focused on developing technologies that will enhance and further the research , diagnostic, pharmaceutical and clinical application of stem cells. Producing scalable quantities of stem cells creates new hope for treating and perhaps even curing human diseases. Currently, SBS is the only company to commercialize a 3-dimenensional, bone marrow stem cell-derived extracellular matrix (BM-HPME®) which mimics the natural stem cell microenvironment providing critical biochemical and physical cues for maintaining stemness during cell culture. For additional information, please contact us at info@stembiosys.com or visit us at www.stembiosys.com


References:

  1. Comprehensive proteomic characterization of stem cell-derived extracellular matrices. Héloïse Ragelle et al., Biomaterials, 2017.
  2. Rescuing replication and osteogenesis of aged mesenchymal stem cells by exposure to a young extracellular matrix. Yun Sun et al ., FASEB, 2011.

  3. Extracellular matrix made by bone marrow cells facilitates expansion of marrow-derived mesenchymal progenitor cells and prevents their differentiation into osteoblasts. Xiao-Dong Chen et al., Journal of Bone and Mineral Research, 2007.

  4. One size does not fit all: developing a cell-specific niche for in vitro study of cell behavior. Milos Marinkovic et al., Matrix Biology, 2015.

  5. Native extracellular matrix preserves mesenchymal stem cell “stemness” and differentiation potential under serum-free culture conditions. Rakian et al., Stem Cell Research and Therapy, 2015.

  6. Reconstitution of marrow-derived extracellular matrix ex vivo: A robust culture system for expanding large-scale highly functional human mesenchymal stem cells. Stem Cells and Development, 2010.

One Comment

  1. Scott Waniger

    18 April, 2017 at 11:07 AM

    I enjoyed your article Iris and would like to have some deeper conversations regarding your organizations technology. Please reach out to me via LinkedIn at https://www.linkedin.com/in/scott-waniger-b3130012/

    Scott Waniger
    VP BioProcessing

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