Periodic Reporting for period 3 - BIRDIE (Bioprinting on-chip microphysiological models of humanized kidney tubulointerstitium)
Reporting period: 2023-09-01 to 2025-03-31
Chronic kidney disease (CKD) is the 11th leading cause of mortality worldwide. With the progress of CKD, patients with a renal filtration rate below 10% of the normal filtration capacity undergo dialysis while waiting for a transplant. Kidney transplant (KT) is the first choice for the treatment but organ shortage is one limitation. After organ transplant, patients have to take immunosuppressive drugs (ISD) to ensure organ survival. However, ISD can promote microbial infections in transplant patients. Kidneys from donors are often infected with BK polyoma virus (BKPyV) which require strict follow-up. No suitable treatment has been identified for BKPyV infection and reactivation can occur which leads to graft loss.
Renal nephrotoxicity, is a recognized problem where patients develop CKD while being treated for other pathologies. The incidence of drug-induced nephrotoxicity varies up to 26% in adult and up to 16% in pediatric patients. Furthermore, nephrotoxicity is also frequently associated to novel therapeutic treatments where the drugs have shown no toxicity in preclinical studies but prove less safe during clinical trials stages. In some cases, drugs cleared during the clinical trials have been withdrawn from the market at a later stage due to AKI events.
In BIRDIE project, we propose to:
Develop and implement the dual organ-chip-model to investigate BKPyV infection and drug induced AKI. We will use the HUMIMIC chip to better mimic the complexity of the renal cellular and extracellular environment, which are thought to participate to early pathophysiology of the BKPyV infection. A “personalized” version of the model will be aimed by applying iPSCs-derived renal progenitor cells;
Develop strategy allowing bioprinting of renal tubulointerstitium 3D model. Parallel peritubular capillary and proximal tubule will be bioprinted mimicking the cellular and extracellular composition of the native kidney. The interstitial cellular and extracellular matrix compositions will be manufactured with novel bioprinting technique. This model will be initially manufactured with cell lines and once the manufacturing strategy is optimized iPSCs-derived cells will be used;
Combine the above mentioned objectives to achieve a physiologically relevant bioprinted on-chip 3D model to study viral- and drug-induced AKI. To obtain a further maturation of the bioprinted tissue and to perform long-term cultures with perfusion, the bioprinted model will be combined with the novel on-chip platform.
During BIRDIE we concluded that:
- Spatial omics allowed the characterization of tissue biopsies of patients with BKPyV. Single-cell RNA transcriptomics data allowed to capture the dynamics of in vitro BKPyV-infected tubules on a chip, leading to the identification of biomarkers that can be used in the future to develop early diagnosis and further antiviral strategies.
- Metanephric mesenchyme, ureteric bud progenitors and proximal tubule-like cells were successfully obtained differentiating various hiPSC lines.
- The new bioprinted system developed enabled the printing of multilayered tissues that can be integrated into organ-on-a-chip platforms. The hybrid Biopen–Biopixlar system allowed for selective exposure to nephrotoxic compounds as well as nanoparticles. The integrated label-free method allowed to tracking cellular changes using the built-in phase microscope. Core-shell bioprinting allowed to create fibers with epithelium and endothelium emulating the tubulointerstitium.
- iPSCs-derived renal cells integrated in the BIRDIE microfluid chip responded to known nephrotoxic compounds proving that this platform is suitable to investigate nephrotoxicants. The dual organ-on-chip also demonstrated its suitability to test compounds affecting multiple organs.
- Perfusable on-chip BIRDIE models open new possibilities for modelling complex physiological conditions.
Renal nephrotoxicity, is a recognized problem where patients develop CKD while being treated for other pathologies. The incidence of drug-induced nephrotoxicity varies up to 26% in adult and up to 16% in pediatric patients. Furthermore, nephrotoxicity is also frequently associated to novel therapeutic treatments where the drugs have shown no toxicity in preclinical studies but prove less safe during clinical trials stages. In some cases, drugs cleared during the clinical trials have been withdrawn from the market at a later stage due to AKI events.
In BIRDIE project, we propose to:
Develop and implement the dual organ-chip-model to investigate BKPyV infection and drug induced AKI. We will use the HUMIMIC chip to better mimic the complexity of the renal cellular and extracellular environment, which are thought to participate to early pathophysiology of the BKPyV infection. A “personalized” version of the model will be aimed by applying iPSCs-derived renal progenitor cells;
Develop strategy allowing bioprinting of renal tubulointerstitium 3D model. Parallel peritubular capillary and proximal tubule will be bioprinted mimicking the cellular and extracellular composition of the native kidney. The interstitial cellular and extracellular matrix compositions will be manufactured with novel bioprinting technique. This model will be initially manufactured with cell lines and once the manufacturing strategy is optimized iPSCs-derived cells will be used;
Combine the above mentioned objectives to achieve a physiologically relevant bioprinted on-chip 3D model to study viral- and drug-induced AKI. To obtain a further maturation of the bioprinted tissue and to perform long-term cultures with perfusion, the bioprinted model will be combined with the novel on-chip platform.
During BIRDIE we concluded that:
- Spatial omics allowed the characterization of tissue biopsies of patients with BKPyV. Single-cell RNA transcriptomics data allowed to capture the dynamics of in vitro BKPyV-infected tubules on a chip, leading to the identification of biomarkers that can be used in the future to develop early diagnosis and further antiviral strategies.
- Metanephric mesenchyme, ureteric bud progenitors and proximal tubule-like cells were successfully obtained differentiating various hiPSC lines.
- The new bioprinted system developed enabled the printing of multilayered tissues that can be integrated into organ-on-a-chip platforms. The hybrid Biopen–Biopixlar system allowed for selective exposure to nephrotoxic compounds as well as nanoparticles. The integrated label-free method allowed to tracking cellular changes using the built-in phase microscope. Core-shell bioprinting allowed to create fibers with epithelium and endothelium emulating the tubulointerstitium.
- iPSCs-derived renal cells integrated in the BIRDIE microfluid chip responded to known nephrotoxic compounds proving that this platform is suitable to investigate nephrotoxicants. The dual organ-on-chip also demonstrated its suitability to test compounds affecting multiple organs.
- Perfusable on-chip BIRDIE models open new possibilities for modelling complex physiological conditions.
All partners in BIRDIE project have collaborated and exchanged information, knowledge and technology. TissUse (TU) has transferred their HUMIMIC technology to Nantes Université (NU) who have optimized culture conditions of primary renal proximal tubule epithelial cells (hRPTEC) in HUMIMIC chips for development of improved in vitro model. The Biopen technology from FluiCell (FC) was transferred to NU and researchers were trained. The protocols of nanoparticles (virus) delivery to the cells was developed and tested. We perfomred RNAseq and spatial transcriptomic pipeline and acquired data for native renal cells/tissue biopsies (control, BKPyV and T-cell rejection). Moreover, at University of Maastricht (UM) we have optimized hiPSC differentiation to obtain metanephric mesenchyme, ureteric bud progenitors and iPSCs-derived proximal tubule-like cells for 3D bioprinting. The Biopixlar bioprinter from FC was transferred, and testing was performed. We have worked on developing new coating strategies to allow further support for cells. We transferred human induced pluripotent stem cell lines from TU to UM, and we have validated the differentiation protocol with these lines. Moreover, TU optimized the cryopreservation of organoids and tested the shipment to NU. FC redesigned the optical path of the bioprinter and developed an integrated Biopen and Biopixlar fluidic heads into the same platform. The methods for bioprinting core-shell cellularized fibers were developed, and the fiber integration into the BIRDIE chip was investigated. Due to the limited integration of bioprinted constructs observed during the final reporting periods, additional strategies were investigated and implemented. Viral infection and nephrotoxicity have been validated on chips cultured with iPSCs-derived proximal tubule-like cells. In terms of exploitable results, some of the project developments are available as new products or add-ons to the technology available before the project. The examples of exploitable results identified during the project were: a) BK virus infection diagnostics platform, b) Kidney analysis assay, c) Disposable kidney chip, d) Combined bioprinting and viral exposure assay printhead and integrated imaging module for bioprinting QC, e) Renal tubulointerstitium model. Dissemination of project results was made through participation in national and international conferences and through scientific publications. Furthermore, three workshops were organized during the project, and different stakeholders were involved.
In vitro models hold significant potential for advancing the development of novel therapies for patients undergoing organ transplantation. Due to the combined influence of immunosuppressive therapy and the presence of BK polyomavirus (BKPyV), treatment strategies for these patients must ensure effective control or clearance of viral infection. The in vitro models established within the BIRDIE project are designed to assess new therapeutic strategies, both during early drug development and through the generation of patient-specific models derived from induced pluripotent stem cells (iPSCs). Such personalized systems will facilitate tailored treatment approaches. By producing human-relevant data, these models are expected to enhance our understanding of kidney disease and its management while contributing to a reduction in the reliance on in vivo studies.