Research Projects of the Biotechnology Working Group
Supervised by Prof. Judit Pongrácz, a special area of 3D printing is bioprinting, in which living tissue (mixture of cells and biogel) is printed. This technology applies technical advances of conventional 3D printing to meet current medical needs. The utilization of artificial human drug testing systems is important for both basic and applied research. Bioprinted organoids are very helpful in studying human diseases at the molecular level.
We plan to develop a workflow suitable for the expansion of human primary cells (3D bioreactor) followed by their printing (3D bioprinting) and completed by their maturation (3D bioreactor). The method may be applicable for the regeneration of joints, teeth, bones, or the reconstruction of ureters/urethras. Our research, focusing on bioprinters and bioreactors, will provide the necessary innovation. We plan to develop transplantation-ready tissues using bioprinters and bioreactors. It is expected that in vitro engineered tissues will significantly reduce the therapeutic tissue regeneration period.
3D lung tissue
Supervised by Prof. Judit E Pongrácz, (Department of Pharmaceutical Biotechnology), bioprinting of a complex 3D lung tissue will commence using various technologies. The challenges are numerous, therefore, apart from the printing process itself, molecular biotechnology-based mechanisms will also be applied to direct cellular growth, blood vessel formation and innervation of the in vitro prepared tissues relying on existing and excellent clinical collaborations.
Primary pulmonary epithelial, microvascular epithelial cells, smooth muscle and pulmonary fibroblasts, including iPCS, will be used to create the tissues. Expertise in all the necessary steps including scaffold 3D printing, genetic engineering, cellular trans-differentiation, cell and tissue maturation are present and in abundance within the research group.
One scaffold material is bioPCL (poly-caprolactone), which is a sterile-printed, antigen and LPS-free, both biocompatible and biodegradable, suitable for the initial tissue formation. In order to create blood vessels, VEGF-A will be cloned, purified and bound to the scaffold. Cells or prepared tissue aggregates will be seeded on 3D bioprinted scaffolds providing the necessary niche to support cellular differentiation. Bioreactors will be used for steady flow tissue culturing of the prepared tissues. Applications are numerous. Lung diseases including lung cancer, pulmonary inflammation, fibrosis and asthma, etc., are the major cause of morbidity and mortality world-wide. Using such lung model tissues, researchers can now gain a unique insight into regenerative and disease processes, and such knowledge will aid novel drug target discovery including the potential for future personalized therapy.
3D thymus engineering
Supervised by Dr. Krisztián Kvell, Department of Pharmaceutical Biotechnology, this sub-project outlines a complex 3D printing and biotechnology-based process in support of creating functional thymus tissue using an artificial scaffold. Cells are prepared from a byproduct of blood-transfusion (buffy coat) which is person-specific, and thus, has the potential for personalized therapy in specific cases of acute immune deficiency. Our research group has expertise in all the necessary steps and protocols including scaffold 3D printing, genetic engineering, cellular trans-differentiation, and cell and tissue maturation.
One scaffold material is bioPCL (poly-caprolactone), which is sterile-printed, antigen and LPS-free, both biocompatible and biodegradable, ideally suitable for the suggested immunological application. In order to create cellular components of the thymus, two cell types will be purified and further processed from Buffy-coat. Peripheral blood monocytes are purified by magnetic beads. A special trans-differentiation protocol allows for the production of blood-borne fibroblasts (fibrocytes). Fibroblasts can then be efficiently further differentiated into thymic epithelial cells (by over-expressing FoxN1). These will be seeded on 3D bioprinted scaffolds providing the necessary niche to support thymocyte development. Buffy coat preparations also contain hemopoietic stem cells (HSCs), albeit at very low frequencies. Yet, they can be efficiently enriched by magnetic separation. Once HSCs are provided with the proper micro-environment (3D scaffold with epithelial and fibroblast cells from above), they readily begin T-cell lineage commitment to develop into thymocytes. T-cell lineage commitment is a well characterized, precisely orchestrated process which requires a tightly regulated micro-environment and also, a sufficient amount of time. Functional thymus tissue may require dynamic (rather than static) culture conditions. Bioreactors required for steady flow tissue culturing will soon be available to our research group. Applications are numerous. On one hand, it provides a unique research tool for regenerative immunology for basic research. On the other hand, it also holds promise for future personalized therapy of certain acute immune deficiencies as an outcome of applied research.
3D lymphoid organoid and arthritis model
The researchers of the Department of Immunology and Biotechnology will perform three parallel studies applying 3D technology. By combining various stromal constituents (fibroblasts, blood and lymphatic endothelial cells) and hematopoietic (lymphoid, dendritic and other myeloid) cells, Péter Balogh and his associates develop 3D lymphoid organoids which will allow the dynamic modelling of lymphoid tissues to be investigated. Within this field, they will analyze the clustering of lymphoid cells and connective components complexed through various approaches (aggregation culture or bioprinting), followed by the evaluation of immunological potency of such aggregates by implantation into immunodeficient mice lacking a RAG enzyme.
In addition to the development of in vitro lymphoid organoids, they will develop a cell separation device requiring 3D printing, for the rapid and efficient selection of cells. Within this scope of effort, plastic matrices with optimized surface geometry will be designed for coating with monoclonal antibodies developed earlier for the selective removal of leukocyte subsets.
Ferenc Boldizsár and his associates investigate the course of rheumatoid arthritis (RA, one of the most frequent autoimmune diseases), characterized by the severe destruction of joints with surfaces covered by hyaline cartilage resulting from the abnormal activation of the immune system against normal constituents of cartilage tissue. In this study, the destruction mechanisms of cartilage tissue created by 3D printing can be investigated under controlled conditions, using leukocytes from human patients or RA mouse models. The goal of this work is to develop and analyze in vitro organoid models to complement ongoing in vivo research at the Department of Immunology and Biotechnology, focusing on RA.