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Publikation 3D printed microfluidic modules. Passive mixers and cells encapsulation in alginate(De Gruyter, 02.09.2022) Dalcanale, Federico; Caj, Michaela; Schuler, Felix; Ganeshanathan, Kireedan; Suter-Dick, LauraPassive mixers and droplet generation microfluidic chip modules were designed and manufactured using a commercial SLA 3D-printer. The mixing modules were designed specifically for 3D-printing and evaluated using FEM modeling. The co-flow droplet generator was used for cancer cells encapsulation and drug potency evaluation.01A - Beitrag in wissenschaftlicher ZeitschriftPublikation Single Cell Gene Expression analysis in a 3D microtissue liver model reveals cell type-specific responses to pro-fibrotic TGF-β1 stimulation(MDPI, 22.04.2021) Messner, Catherine; Babrak, Lmar; Titolo, Gaia; Caj, Michaela; Miho, Enkelejda; Suter-Dick, Laura3D cell culture systems are widely used to study disease mechanisms and therapeutic interventions. Multicellular liver microtissues (MTs) comprising HepaRG, hTERT-HSC and THP-1 maintain multicellular interactions and physiological properties required to mimic liver fibrosis. However, the inherent complexity of multicellular 3D-systems often hinders the discrimination of cell type specific responses. Here, we aimed at applying single cell sequencing (scRNA-seq) to discern the molecular responses of cells involved in the development of fibrosis elicited by TGF-β1. To obtain single cell suspensions from the MTs, an enzymatic dissociation method was optimized. Isolated cells showed good viability, could be re-plated and cultured in 2D, and expressed specific markers determined by scRNA-seq, qRT-PCR, ELISA and immunostaining. The three cell populations were successfully clustered using supervised and unsupervised methods based on scRNA-seq data. TGF-β1 led to a fibrotic phenotype in the MTs, detected as decreased albumin and increased αSMA expression. Cell-type specific responses to the treatment were identified for each of the three cell types. They included HepaRG damage characterized by a decrease in cellular metabolism, prototypical inflammatory responses in THP-1s and extracellular matrix remodeling in hTERT-HSCs. Furthermore, we identified novel cell-specific putative fibrosis markers in hTERT-HSC (COL15A1), and THP-1 (ALOX5AP and LAPTM5).01A - Beitrag in wissenschaftlicher ZeitschriftPublikation Implementation of a human renal proximal tubule on a chip for nephrotoxicity and drug interaction studies(Elsevier, 04.04.2021) Suter-Dick, Laura; Caj, Michaela; Hutter, Simon; Vormann, Marianne; Vriend, Jelle; Lanz, Henriette; Gijzen, Linda; van den Heuvel, Angelique; Joore, Jos; Trietsch, Sebastian; Stuut, Christaan; Nieskens, Tom T.G.; Peters, Janny; Ramp, Daniela; Russel, Frans; Roth, Adrian; Lu, Shuyan; Polli, Joseph; Jacobsen, BjörnProximal tubule epithelial cells (PTEC) are susceptible to drug-induced kidney injury (DIKI). Cell-based, two-dimensional (2D) in vitro PTEC models are often poor predictors of DIKI, probably due to the lack of physiological architecture and flow. Here, we assessed a high throughput, 3D microfluidic platform (Nephroscreen) for the detection of DIKI in pharmaceutical development. This system was established with four model nephrotoxic drugs (cisplatin, tenofovir, tobramycin and cyclosporin A) and tested with eight pharmaceutical compounds. Measured parameters included cell viability, release of lactate dehydrogenase (LDH) and N-acetyl-β-d-glucosaminidase (NAG), barrier integrity, release of specific miRNAs, and gene expression of toxicity markers. Drug-transporter interactions for P-gp and MRP2/4 were also determined. The most predictive read outs for DIKI were a combination of cell viability, LDH and miRNA release. In conclusion, Nephroscreen detected DIKI in a robust manner, is compatible with automated pipetting, proved to be amenable to long-term experiments, and was easily transferred between laboratories. This proof-of-concept-study demonstrated the usability and reproducibility of Nephroscreen for the detection of DIKI and drug-transporter interactions. Nephroscreen it represents a valuable tool towards replacing animal testing and supporting the 3Rs (Reduce, Refine and Replace animal experimentation).01A - Beitrag in wissenschaftlicher Zeitschrift