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Bioengineering approaches to blood cell production
blood platelets in many aspects, including morphology, expression of lineage-specific antigens, and ability to aggregate in response to agonists.18,57 A major advantage of hiPSC is the possibility of using human leukocyte antigen- matched hiPSC or cells genetically engineered to lack cer- tain antigens in order to produce platelets that would not be rejected by the recipient.58 Alternatively, as hiPSC can be generated from any donor, they are theoretically suit- able for generating a bank of phenotypically matched ery- throcytes. Various research groups have published meth- ods for generating red blood cells from hiPSC59,60 albeit with some concerns about the expression of hemoglobin and enucleation potential, which could not be achieved in the context of in vitro differentiation, unless in the presence of feeder cells or after injection into mice recipients. Therefore, although very attractive, both hESC and hiPSC might not represent the best choice of stem cells for pro- ducing blood components. In most cases the differentia- tion protocols are long and expensive. Indeed, high costs for generating, validating, and maintaining these cell lines should be considered.61 Finally, there is still the concern that any cellular product derived from them could be oncogenic or teratogenic,62 given the potential genomic instability of these stem cell lines.
To overcome these limitations, the latest stem cell source to be investigated has been adipose tissue, which has the advantage of being ubiquitously available and easily acces- sible in large quantities with minimally invasive harvesting procedures. Ono-Uruga et al. recently demonstrated that
the production of endogenous thrombopoietin is involved in megakaryocyte differentiation and platelet production from adipose-derived mesenchymal stromal/stem cell lines.63 The lack of specific markers for the identification and isolation of this subset of stem cells and the absence of standardized isolation and culture protocols make it diffi- cult to translate this approach into different laboratories.64
Finding the route towards differentiation: cytokines instruct but are not enough
It is conceivable that a regular, non-donor-derived sup- ply of erythrocytes and platelets could be achieved in the near future, thus making it necessary to define the best culture conditions to maximize the yield of platelets gen- erated per single megakaryocyte, as well as the number of enucleated erythrocytes obtained in vitro (Figure 3).
Recombinant human thrombopoietin is commonly used to produce megakaryocytes.65-67 Thrombopoietin analogs, such as eltrombopag and romiplostim, have been tested in culture but their use for ex vivo platelet manufac- turing is still limited because they can induce proliferation of immature progenitors.68,69 Thrombopoietin and its analogs are combined with a wide variety of cytokines including stem cell factor and interleukins (e.g., IL-3, IL-6, IL-11) which synergize to produce very pure populations of mature megakaryocytes forming proplatelets without the need for serum supplementation or co-culture with
Figure 3. Overview of soluble factors used in the production of platelets and erythrocytes in vitro. Different cocktails of cytokines, pharmacological agents and/or co-culture with feeder cells have been used to generate platelets and erythrocytes in vitro. HSC: hematopoietic stem cell; FLT3: Fms related tyrosine kinase 3; SCF: stem cell factor; IL: interleukin; IGF-1: insulin-like growth factor-1; TPO: thrombopoietin; EPO: erythropoietin; SR1: stemRegenin 1; AhR: aryl hydrocarbon receptor. The figure was created using Servier Medical Art templates licensed under a Creative Commons Attribution 3.0 Unported License (https://smart.servier.com).
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