In the future, induced pluripotent stem cells could be used to generate red blood cells at scale, reducing the reliance on blood donation.

© Zhong Ri

Scaling up universal blood production

25 Aug 2021

A scalable cell culture platform can generate universal red blood cells from stem cells, paving the way for manufacturing an abundant supply of blood for transfusions.

As doctors bustle through surgical procedures and emergency cases, 325 units of blood are needed every day in Singapore—equivalent to over 170 liters containing 650 trillion red blood cells (RBCs). While hospitals build up stockpiles for emergency transfusions, rare blood types may be in limited supply and relying solely on donors is unsustainable in the long run.

Supply shortages may someday be resolved by using human induced pluripotent stem cells (hiPSCs)—unspecialized cells that can generate different cell types, including RBCs. The challenge, however, is two-fold: first, creating the right environment to trigger stem cell differentiation into RBCs, and second, reproducibly scaling up the process. At a minimum, ultrahigh-density cultures of 100 million cells/mL would be needed at the industrial or clinical level.

“If you develop a protocol that only works at the lab scale but cannot translate into suspension bioreactors, that method will not make any impact to patients,” explained Steve Oh, Director of the Stem Cell Bioprocessing group at A*STAR’s Bioprocessing Technology Institute (BTI).

Unlike the traditional monolayers where cells are typically grown, large-scale bioreactors involve the continuous agitation of suspended cells. To elevate RBC generation to the bioreactor scale, Oh and a team of scientists has developed a culture platform to do just that.

By culturing hiPSC lines on Laminin-521-coated microcarriers (MCs)—small beads that provide cells a surface to attach to—the team carried out the entire differentiation process via the agitation suspension platform. In just over a month, the researchers produced type O-negative RBCs, the universal donor blood type, while the standard monolayer protocol failed to produce RBCs at all.

From six-well plates, the hiPSC-MC aggregates were also tested out in larger 500-mL spinner flasks, generating high-density cell cultures of up to 17 million cells/mL. “This is a 5,000-fold improvement from where we started with 100-microliter 96-well plates. We need to improve by another 1,000-fold in the next few years to bring this application into human clinical trials,” Oh said.

Despite some differences with adult RBCs, the hiPSC-derived RBCs possess the key components to be functional. “Genetically, both sources of RBCs are almost identical and there are no obvious red flags like oncogenes expressed,” Oh noted. The next steps, he said, entail further testing in mice models, including evaluating the RBCs’ capacity to carry and dispatch oxygen to surrounding tissues.

By integrating CRISPR technologies and additional molecular components, the team is upgrading its suspension platform to increase cell density and accelerate differentiation. Simultaneously, they are looking at founding an RBC manufacturing company with a vision to bring off-the-shelf blood to Singapore—and the world.

The A*STAR-affiliated researchers contributing to this research are from the Bioprocessing Technology Institute (BTI), the Institute of Molecular and Cell Biology (IMCB) and the Singapore Immunology Network (SIgN).

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Sivalingam, J., Yu, S., Lim, Z.R., Lam, A.T.L., Lee, A.P., et al. A scalable suspension platform for generating high-density cultures of universal red blood cells from human induced pluripotent stem cells. Stem Cell Reports 16, 1-16 (2020) | article

About the Researcher

Steve Oh is the Director of the Stem Cell Bioprocessing group at A*STAR's Bioprocessing Technology Institute. His research interests include: microcarrier cultures of pluripotent stem cells, bioreactor expansion of fetal MSC and hESC, engineered surfaces for hESC cultures, differentiation of hESC derived cardiomyocytes with small molecule inhibitors, and imaging and monitoring of pluripotent stem cells and cardiomyocytes.

This article was made for A*STAR Research by Wildtype Media Group