Workshop Report

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The importance of mimicry of physiological aspects in vitro has incrementally came into focus within various disciplines of life sciences in recent years. To discuss recent advances and exchange scientific experiences and the know-how the “Advanced Cell Culture Technologies” workshop was organised by the ÖGMBT Working Group "Cell-based assays, therapies and products" that took place at BOKU on 17.03.2023.

In addition to insightful and exciting keynote lectures by by PD Dr. Lavrentieva (Leibniz Universität Hannover) and Prof. Dr. Hansmann (Fraunhofer ISC, Würzburg), who shared their views on advanced 3D cell culture technologies and potential application of automation and artificial intelligence in research, the workshop was complemented by a variety of short talks that covered a wide range of relevant topics, including 3D tumor models, 3D encapsulation strategies and employment of mechanical cues in tissue engineering applications.

The unique highlight of the workshop were roundtable discussions, which were dedicated to a specific aspect of advanced cell culture technologies. The participants were invited to share their interests, expertise as well as discuss potential limitations and potential solutions within each field. In the following a short summary of discussions is given:

1) Hydrogels in 3D cell culture (Moderator: Farhad Chariyev-Prinz)

“At this roundtable we discussed how and what hydrogel systems are employed to mimic physiological aspects in vitro. In this context, applications in the fields of tissue engineering, EV engineering as well as development of in vitro model systems have been most prevalent. Despite this broad applicability, some important limitations could also be identified. Although application of 3D systems is a significant step up compared to commonly used 2D systems, complete mimicry of physiological properties is yet to be achieved; this includes availability of specific binding sites, employment of xeno-free components, development of strategies for cell release and realization of tissue specific mechanical properties. During a lively discussion several potential solutions have been proposed, however a suggestion to further “enhance the collaboration between biology, chemistry and engineering” was identified as the most straightforward and necessary approach to facilitate the development of the field.”

2) Automation and AI in cell culture technologies (Moderator: Jan Hansmann)

“The roundtable evaluated the current state of AI and automation in cell culture labs. It was found that methods of AI are already applied, e.g. in image analysis. Considering the use automation technology in cell culture, there are various systems from partial to full automation. Examples for partial automation such as pipetting robots or FACS systems were identified, whereas holistic approached, which can perform a complete cell culture process starting from cell isolation and finally producing a cell culture product, are limited. Reasons might be initial costs and the need of highly trained staff. As limitations for AI applications, data availability, lab-specific constraints, cyber security, and reservations from researchers against this technology were listed. Reservations could arise from the fact that AI technologies are very complex and in some cases such as neural networks, they seem to be a black box. A common challenge for AI and automation is the legal framework, which is difficult to apply for automation and AI in specific points. Examples such as cyber security or automated data management were brought up. To overcome these hurdles, expert from AI and automation should be included in the refinement of the legal framework. Then, automation and AI could increase standardization and robustness and decrease cost in cell culture labs. An additional strong argument for automation is the release of capacities, when the assessment of results is supported by AI and routine work is done by a robotic system. “

3) Dynamic Culture Systems and Processes (Moderator: Cornelia Kasper)

“The discussion at this table focused on the evaluation of on existing culture systems and trends in the field. Early in the discussion the lack of “common” components and designs for a “one fits all” application systems became obvious. A considerable number of different bioreactors exist and well-established ones are being adapted to fulfil the need of “cell-based therapy product” manufacturing (e.g. Tissue Engineering, stem cell expansion, extracellular vesicle production), but only few approaches offer several operational modes and a more broad applicability; in this context only few systems are currently being developed to address this issue. Another critical point on all discussed levels was the use of disposable components: the amount of plastic waste is immense and the single use technology makes processes expensive. At the same time safety issues need to be considered as well as robustness and easy handling and scalability. One big challenge still obviously persits in the transfer of established know-how and instrumentation from biopharmaceutical production field to (stem) cell based therapy production (including implementation of sensors systems, feeding strategies etc). A closer collaboration, exchange of knowledge and adaptive conceptualization for cell therapy manufacturing would be tremendously advantageous to the field towards realization of modular platforms.”

4) Primary cells of the future (Moderator: Dominik Egger)

“At this roundtable we have discussed the necessity to use primary cells more frequently and what is missing to do so. We found out that most attendees would like to use primary cells more frequently in their research, especially in the context of studies concerning tissue engineering for in vitro models. The limited availability, limited expandability, and the high variability in reproducibility were identified as main limiting factors for using primary cells more frequently. After a lively discussion it was proposed to use more progenitor cells that can be expanded ahead of terminal differentiation. Regarding the problem of reproducibility, the group discussed the pros and cons of using either a pooled cell bank of many donors against many biological replicates. The most important outcome was the idea of developing a network of clinicians and scientists that would allow for reporting available donor tissue for the isolation of cells more easily and swiftly. This could allow to use available tissue for the isolation of more than one cell type by more than one group and increase the efficiency of tissue donations.“

5) Monitoring and imaging in 3D cell cultures (Moderator: Antonina Lavrentieva)

“The current state of the art and existing challenges were discussed at the round table “Monitoring and imaging in 3D cell cultures”. Participants emphasized that more complex 3D cultivation systems require more complex analysis tools. High-throughput screening in 3D systems also remains difficult. CLSM is currently a gold standard, but this technique is semi-quantitative and requires labeling that can affect cell function and intracellular processes. Raman technology lacks tools for easy analysis and interpretation of spectral data that focus on biologically relevant information. WF fluorescence microscopy is available in every lab but lacks Z resolution. Spatial heterogeneity of 3D samples, maintenance of cell viability, and optical tissue transparency remain major issues. Most of the analysis is endpoint and normalization needs to be addressed. There are also still no clear definitions of 3D constructs and their size and geometry. Tissue clearing can be used as possible solutions (but this requires tissue fixation). Non-optical methods and lens-free microscopy are under development and may help to better monitor complex 3D constructs in the near future. The use of genetically encoded biosensors and small sensor molecules provides spatio-temporal information for non-invasive monitoring of 3D cultures.”

In the second half of the event the programm was rounded up by a lively poster session, which allowed for further scientific exchange. We warmly congratulate the two winners of the poster award Sonya Ciulean (Fraunhofer IZI, Leipzig) and Julia Moldaschl (BOKU).

Prof. Dr. Cornelia Kasper, Dr. Dominik Egger and Dr. Farhad Chariyev-Prinz, who organized the event on the part of BOKU with the support of ÖGMBT were very satisfied. "We received a lot of positive feedback about this format and we are very happy that we could offer a “live” workshop again after the long Corona break," said Prof. Kasper.

Joining and actively participating in the Working Group “Cell based assays, therapies and products” of the ÖGMBT is possible at any time via the website: https://oegmbt.at/ueber-uns/working-groups/cbatp.