Regenerative medicine, including tissue engineering (TE), cell therapy and pharmaceutical intervention represents a very promising alternative in this context. During the recent years several attempts has been made to engineer various tissue/ organs and initial evidence exists about the clinical potential of this method. Despite early clinical applications for the trachea, bladder, skin or heart valves, more solid organs can probably never be transferred to the patient regarding their demanding and complicated engineering process. For complex organs, such as heart, lung or the liver, cell therapy and pharmaceutical intervention can be an optimum alternative.
RM aims to replace and regenerate damaged tissue to restore organ function. It is very common that all three aspects of RM, such as TE, cell therapy and pharmaceutical intervention are combined at some level and should not be categorical separated from each other. The following components are necessary to be considered: a) a scaffold/ matrix seeded with b) different cell type(s) using c) the body or a device as a native or artificial bioreactor, respectively, d) bioactive molecules/Signaling and e) cell/gene therapy. We already learned that each target tissue requires specific components, and engineering processes need to be modified to distinctive needs.
a) Scaffold/ Matrix
The possibility of using TE as a potential solution in the clinic has evolved due to the improved understanding of cell-to-matrix interactions. Based on the gained knowledge from recent decades, the necessity of optimized scaffolds for cell seeding and the later in vivo outcome became a significant part in the whole concept. Notably, the target tissue and clinical propose drive the requirements for both the biological and the synthetic scaffold. Even though biological and synthetic scaffold differ completely from each other, some basic characteristics must be considered while designing them, such as the bioactivity, the capability to host seeded cells, non-immunogenicity, non-toxicity, non-carcinogenicity and non-teratogenicity.
In contrast, organ specific properties can vary from time to time e.g. for the trachea air- and liquid-tight seals, flexibility and strength to prevent the collapse of the scaffolds, and must be considered when choosing the ideal scaffold. In general, natural scaffolds are developed using decellularization methods to remove all immunogenic components. Chemical/enzymatic (e.g. DNase, Deoxycholate, TritonX, Trypsin etc.), mechanical or thermal methods can be applied to obtain a both non-immunogenic and acellular but consideration must be given to prevent the native architecture, namely, the extracellular matrix (ECM) (including collagen, elastin, etc). Resident ligands and bioactive molecules can promote self-assembly of functional cell groupings and may support homing of endogenous cell. Besides, maintained proteins, such as β-fibroblast growth factor (β-FGF) can drive different remodelling processes and provide angiogenesis. Regarding synthetic materials (e.g. Marlex mesh, polyester urethane, Polypropylene mesh) all the aforementioned native proteins are missing, which can result in i) poor vascularization ii) low level of integrity iii) migration iv) stiffness and v) easy contamination. Intensive research is ongoing to find solutions to overcome the absence of these characteristics and/or develop novel materials that can meet native characteristics. Most recently, the first transplantation of a stem cell seeded synthetic tracheal graft has demonstrated clinical evidence for a successful application of a synthetic material in human. Currently both microstructure and nanofiber composites are under investigation.
b) Cells
Seeding cells on a scaffold is associated to different issues that one has to consider, such as donor related immunogenicity (allogenic versus autologous cells), differentiated vs. undifferentiated (stem/progenitor) cells and status of the cells (freshly isolated cell or co-cultures vs. purified or expanded cells, potential pre-conditioning or differentiation of the cells). Cell type and handling result in different in vivo behavior and overall outcome. For instance, autologous mature cells are limited in their functionality and availability but de-differentiation (or tumorigenicity) is very unlikely. In contrast, allogenic mature cells are nearly always available but can easily induce immunological response in the recipients after cell transplantation/application. Adult stem and progenitor cells are multipotent and have the ability of self-renewal , besides they are only marginally ethical debated. Induced pluripotent stem cells (iPSCs) were praised as the Holy Grail of all stem cells in their early days, representing a unique, non-immunogenic, autologous alternative to embryonic stem cells (ESCs) but unfortunately, evidence exists that iPSCs appear to be prone to immune recognition and consecutive rejection. Currently the use of adult stem/ progenitor cells, such as bone marrow derived hematopoietic stem cells (HSCs) or mesenchymal stem/ stromal cells (MSCs), as well as differentiated cells, are the most readily available alternative cell source in tissue engineering. Ongoing research may achieve more efficient and imporved cellular agents for the clinical routine care in the near future.
c) Bioreactor
Both in vivo and in vitro culture is feasible and may be transferred into the clinic. The conventional in vitro cell culture is associated with several disadvantages, such as contamination and cost intensive processing. Moreover static culture conditions limits the differentiation of specific cell types into a distinct phenotype. A bioreactor should mimic a native-like environment and thus provide the optimum culture conditions for each specific cell type. For our clinical protocol of the trachea we are using a rotating bi-phase (liquid-/ air-phase) bioreactor that allows easy and sterile handling, differential cell seeding and fully-automatic cell culture. Different types of tissue require for variable seeding and culture conditions and therefore specific designed bioreactors. However, basic principles must be established. Beside the in vitro cell culture one can use the body a natural bioreactor what makes the engineering process very easy to handle and time- and cost-saving.
d) Bioactive Molecules and Pharmaceutical Intervention
As described before synthetic materials can usually not provide the properties of native tissue. Novel technologies and strategies must be developed to avoid graft failure. Recently we used en electrospun nanofiber based composite. Nanotechnology appears to be a promising tool to modify synthetic materials in terms of mimicking native architecture and physiological environment. Besides, it can be used to insert factors and proteins into the synthetic scaffold. 3-D priniting is another method to design in a fast and highly accurate manner native like structures and studies are going on to print scaffolds that actually maintain both cells and these factors, and this from the very beginning on. Currently, we focus on systemic and local administration of boosting and growth factors, such as granulocyte stimulating factor (GCSF), Erythropoietin (EPO) and transforming growth factor (TGF-β) to increase the endogenous regeneration of the body. Both experimental and initial clinical findings in trachea TE demonstrated the high potential of using these drugs/reagents 1) to mobilize stem/progenitor cells into the peripheral circulation, 2) to down-regulate apoptosis of mobilized cells and 3) to differentiate stem cells into distinct cell type. Further studies need to be performed to obtain a better and in-depth knowledge of underlying pathways.
e) Cell/Gene Therapy
Various cell types have been already applied clinical in different scenarios and data suggest the beneficial effect for patient. Particularly when complex organ structures are involved the tissue engineering approaches may fail to reconstruct the tissue and restore function. Cell and gene therapy has a high translational capacity due to its easy handling and application. However, the misuse, unknown underlying mechanism and side effects, particularly in gene therapy make this field of RM ethical debatable. Before clinical application becomes routine further in-depth investigation of associated pathways and potential complications should be performed.
Conclusion
Early clinical application demonstrated the feasibility of using regenerative approaches to replace damaged tissue and organs with rather simple architecture. Preclinical findings suggest that even higher complex organs can be successfully partly engineered. However, due to the complexness of these solid organs cell therapy might be the more promising solution for the near clinical future. Notably, pharmaceutical strategies should be widely considered to increase endogenous regeneration and self-healing capacity of the recipient. The new field of RM rises novel ethical concerns and questions – these must be addressed and answered.