Unmasking insulin resistance triggering mechanisms using microphysiological two-organ systems as in vitro disease models of metabolic syndrome and non-alcoholic fatty liver disease

Project Information

LIV-AD-ON-CHIP

Grant agreement ID: 845147

DOI

10.3030/845147

Project closed

EC signature date 2 May 2019

Start date 1 July 2020

End date 26 September 2023

Funded under

• EXCELLENT SCIENCE - Marie Skłodowska-Curie Actions

Total cost

€ 162 806,41

EU contribution

€ 162 806,40

Coordinated by

EBERHARD KARLS UNIVERSITAET TUEBINGEN
Germany

Periodic Reporting for period 1 - LIV-AD-ON-CHIP (Unmasking insulin resistance triggering mechanisms using microphysiological two-organ systems as in vitro disease models of metabolic syndrome and non-alcoholic fatty liver disease)

Reporting period: 2020-07-01 to 2022-06-30

Summary of the context and overall objectives of the project

The proposed project represents an integrative and multidisciplinary endeavor aimed at developing and utilizing a microphysiological organ-on-a-chip model to create a human physiologically and physio-pathologically relevant in vitro insulin signaling model. Recognizing the persistent challenge of insulin resistance, despite decades of study using traditional clinical and cell culture approaches, the adoption of organ-on-a-chip technology promises novel insights into this field. While human organ-on-a-chip models have been established for drug screening using transformed cell lines, the limitations of such models underscore the necessity of employing more physiologically relevant human primary cells, particularly hepatocytes, over extended periods.
To address this, the project outlines four specific objectives:
1. Establish methodologies for adipocyte and hepatocyte bioenergetic characterization on-chip (O1).
2. Differentiate stem cells into adipocyte-like and hepatocyte-like cells focusing on their bioenergetic competence (O2).
3. Develop a physiological model for liver-adipose tissue crosstalk (O3).
4. Utilize a physio-pathological model for insulin resistance studies (O4).
Distinguishing itself from previous work, this project human primary mature adipocytes characterized for their bioenergetic metabolism. Additionally, innovative flexible multi-organ-interconnection concepts enable molecular studies of multi-organ diseases such as metabolic syndrome and non-alcoholic fatty liver disease (NAFLD). These approaches pave the way for the introduction of other cell types, such as immune cells, facilitating a deeper understanding of disease mechanisms and enabling the creation of patient-specific disease models.
The biomedical applications of this work are wide-ranging: (i) Biomarker discovery for early non-symptomatic manifestations of insulin resistance, potentially preventing the development of diseases like NAFLD and metabolic syndrome. (ii) Food science applications, including testing nutritional schemes using personalized long-term cultures. (iii) Identification of future pharmacologic targets to modulate or reverse the progression of insulin resistance at an early non-symptomatic state.
In summary, this project offers a promising modular multi-organ platform for advancing our understanding of insulin resistance and related metabolic disorders, with implications for both basic research and practical applications in healthcare and nutrition.

Work performed from the beginning of the project to the end of the period covered by the report and main results achieved so far

The project report outlines work conducted across multiple work packages (WP) focusing on characterizing bioenergetic profiles of adipocytes and hepatocytes, differentiating stem cells into adipocyte-like and hepatocyte-like cells, and establishing physiological models of liver-adipose tissue interaction.
In WP1, efforts were directed towards characterizing hepatocyte bioenergetic profiles using on-chip culture techniques. Challenges in fabrication and tissue generation led to downscaled approaches using alternative materials. Sensor integration for monitoring oxygen and glucose levels facilitated metabolic activity assessment. Similar efforts were made to characterize adipose tissue bioenergetics both off-chip and on-chip, demonstrating insulin responsiveness and immunocompetence.
WP2, focusing on stem cell differentiation, faced delays due to the pandemic and resource allocation challenges. Instead, emphasis was placed on utilizing human primary hepatocytes.
WP3 involved on-chip culture and characterization of stem cell-derived adipocyte-like cells, showcasing successful differentiation and functionality assessments including response to β-adrenergic stimulation.
In WP4, efforts were made to establish a liver-adipose physiological model using multi-organ chip platforms. Media optimization and connection strategies were explored, demonstrating improved viability and functionality of cultured cells.
WP5 demonstrated the functional demonstration of a liver-adipose tissue connection, showcasing increased intracellular fatty acid accumulation in the liver tissue when connected downstream of white adipose tissue, indicating a physiological interaction between the two tissues.
Throughout the report, challenges such as fabrication difficulties, inter-donor differences, and media optimization were highlighted, alongside advancements in sensor integration, stem cell differentiation, and multi-organ chip platform development.
Overall, the project aimed to advance understanding of adipocyte and hepatocyte bioenergetics, stem cell differentiation techniques, and physiological interactions between liver and adipose tissues, with implications for metabolic research and drug development.

The work funded under LIV-AD-ON-Chip has been published in reputable journals such as Advanced Science, Education Sciences, and Open Biology. Additionally, efforts towards education and outreach resulted in unexpected publications, including a first last-author publication on training needs in organ-on-a-chip and a publication in Frontiers for Young Minds aimed at explaining the concept of organ-on-a-chip to an audience aged 8-15 years old.

Furthermore, the project's findings have been presented internationally, highlighting its significance and impact. Notably, the research was selected for an oral presentation at the MPS Summit 2023 in Berlin, emphasizing its recognition within the scientific community. Additionally, the project was recognized with a guest lecture invitations in Germany, Spain and Portugal. These presentations showcase the project's contributions to advancing research in the field of metabolic disorders and its recognition among peers in the scientific community.

Progress beyond the state of the art and expected potential impact (including the socio-economic impact and the wider societal implications of the project so far)

LIV-AD-ON-A-HIP allowed me to leverage my interdisciplinary training to bridge engineering and life sciences in addressing the societal challenge of quantifying organ-specific human insulin resistance in health and disease, thereby advancing our understanding of disease mechanisms. This approach was the development of modular tailored Multi-Organ-on-Chip (OoC) models as a New Approach Method (NAM) and animal testing-free approach, filling a research niche where traditional animal data has not provided satisfactory answers, given the differences in glucose metabolism regulation between humans and rodents.
By identifying insulin resistance thresholds in an OoC platform, this work will facilitate the development of new pharmacological approaches for metabolic syndrome, non-alcoholic fatty liver disease (NAFLD), type 2 diabetes, and elucidate the obesogenic mechanisms of EDC in both environmental and food toxicology. Moreover, this initiative will support the creation of an insulin resistance adverse outcome pathway (AOP), addressing a critical need expressed by regulatory bodies such as the European Food Safety Agency (EFSA), European Chemical Agency (ECHA), and European Commission.

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