Publications



The choice of ultra-low attachment plates impacts primary human and primary canine hepatocyte spheroid formation, phenotypes, and function
2024 · Biotechnology Journal · Volume 19, Issue 2
Chen Xing, Aurino Kemas, Evgeniya Mickols, Kathrin Klein, Per Artursson, Volker M. Lauschke

Organotypic three-dimensional liver spheroid cultures in which hepatic cells retain their molecular phenotype and functionality have emerged as powerful tools for preclinical drug development. In recent years a multitude of culture systems have been developed; however, a thorough side-by-side benchmarking of the different methods is lacking. Here, we compared the performance of ten different 96- and 384-well microplate types to support spheroid formation and long-term culture. Specifically, we evaluated differences in spheroid formation kinetics, viability, functionality, expression patterns, and their utility for hepatotoxicity assessments using primary human hepatocytes (PHH) and primary canine hepatocytes (PCH). All 96-well plates enabled formation of PHH liver spheroids, albeit with differences between plates in spheroid size, geometry, and reproducibility. Performance of different 384-wells was less consistent. Only 6/10 microplates supported the formation of PCH aggregates. Interestingly, even if PCH aggregates in these six microplates were more loosely packed than PHH spheroids, they maintained their function and were compatible with long-term pharmacological and toxicological assays. Overall, Corning and Biofloat plates showed the best performance in the formation of both human and canine liver spheroids with highest viability, most physiologically relevant phenotypes, superior CYP activity and lowest coefficient of variation in toxicity assays. The presented data constitutes a valuable resource that demonstrates the impacts of current ultra-low attachment plates on liver spheroid metrics and can guide evidence-based plate selection. Combined, these results have important implications for the cross-comparison of different studies and can facilitate the standardization and reproducibility of three-dimensional liver culture experiments.

Effects of Piper aduncum (Piperales: Piperaceae) Essential Oil and Its Main Component Dillapiole on Detoxifying Enzymes and Acetylcholinesterase Activity of Amblyomma sculptum
2024. Int. J. Mol. Sci. 2024, 25(10)
Adalberto Alves Pereira Filho, Vladimir Fazito do Vale, Caio Marcio de Oliveira Monteiro, Mayara Macedo Barrozo, Mariana Alves Stanton, Lydia Fumiko Yamaguchi, Massuo Jorge Kate, Ricardo Nascimento Araújo

Amblyomma sculptum is a species of tick in the family Ixodidae, with equids and capybaras among its preferred hosts. In this study, the acaricidal activity of the essential oil (EO) from Piper aduncum and its main component, Dillapiole, were evaluated against larvae of A. sculptum to establish lethal concentration values and assess the effects of these compounds on tick enzymes. Dillapiole exhibited slightly greater activity (LC50 = 3.38 mg/mL; 95% CI = 3.24 to 3.54) than P. aduncum EO (LC50 = 3.49 mg/mL; 95% CI = 3.36 to 3.62) against ticks. The activities of α-esterase (α-EST), β-esterase (β-EST), and glutathione-S-transferase (GST) enzymes in A. sculptum larvae treated with Dillapiole showed a significant increase compared to the control at all concentrations (LC5, LC25, LC50 and LC75), similar results were obtained with P. aduncum EO, except for α-EST, which did not differ from the control at the highest concentration (LC75). The results of the acetylcholinesterase (AChE) activity show an increase in enzyme activity at the two lower concentrations (LC5 and LC25) and a reduction in activity at the two higher, lethal concentrations (LC50 and LC75) compared to the control. These results suggest potential mechanisms of action for these natural acaricides and can provide guidance for the future development of potential plant-derived formulations. Keywords: Amblyomma cajennense complex; α-esterase; β-esterase; glutathione-S-transferase; natural products

Standardized conditions for growth of lung cancer tumoroids cultured reproducibly in 3D-cultures
2024 Cancer Res
Karina Malmros, Daniel Carlsén, Hans Brunnström

Background: Lung cancer is the most common cause of cancer-related death in Sweden with an estimated mortality of 3500 persons in 2021. Late diagnosis and heterogeneity of the disease lead to difficulty in predicting optimal treatment regime for each case. Further development of personalized therapy would be facilitated by reliable and efficient cultivation of cancer cell samples from patients in a clinical setting. A promising method is to grow the cells as 3D structures known as tumoroids, though the choice of extracellular matrix substrate or solid support most suitable for this purpose should be elucidated. Aim: To assess which extracellular matrix substrates or solid supports which enable lung cancer tumoroid generation reproducibly without affecting the status or detection of genetic alterations and protein expression, using lung cancer cell lines. Methods: Eight lung cancer cell lines, where of three adenocarcinoma and five squamous cell carcinomas, were included in this project. Each cell line was characterized with immunohistochemistry (IHC) for commonly expressed antigens in the two lung cancer types. Also, protein detection with Western Blots was performed. Cell lines found not expressing relevant antigens associated with the specific lung cancer variant were discarded. The remaining cell lines were cultivated in BIOFLOAT TC-Platte 96 well plate, Nunclon Sphera flasks, or GrowDex-T hydrogel, later transferred to a Corning Elplasia 6 well plate, and monitored for tumoroid generation. Results: The lung squamous cell carcinomas HCC-95 and HCC-1588 were both positive for CK5 and p40, while H1703 was negative for both markers. The adenocarcinoma H1975 was positive for TTF-1, also, unexpectedly, seen for NCI-H1703, earlier defined as squamous cell carcinoma. The tested cell lines were successfully cultured as tumoroids in the BIOFLOAT TC- 96 well plate, Nunclon Sphera flasks, and GrowDex-T hydrogel. The mutational status of the tumoroids as compared to the original tumors is currently under evaluation. Conclusion: We here show successful 3D culturing of cell lines with alternative substrates. Further fine-tuning of the chosen reproducible system’s simplicity and cost is required before culturing tumoroids from lung cancer patients can be launched.

Chalcones induce apoptosis, autophagy and reduce spreading in osteosarcoma 3D models
2024 Biomedicine & Pharmacotherapy Volume 179
M. Rossi, C. Pellegrino, M.M. Rydzyk, G. Farruggia, D. de Biase, S. Cetrullo, S.D. Adamo, A. Bisi, P. Blasi, E. Malucelli

Osteosarcoma is the most common primary bone malignancy with a challenging prognosis marked by a high rate of metastasis. The limited success of current treatments may be partially attributed to an incomplete understanding of osteosarcoma pathophysiology and to the absence of reliable in vitro models to select the best molecules for in vivo studies. Among the natural compounds relevant for osteosarcoma treatment, Licochalcone A (Lic-A) and chalcone derivatives are particularly interesting. Here, Lic-A and selected derivatives have been evaluated for their anticancer effect on multicellular tumor spheroids from MG63 and 143B osteosarcoma cell lines. A metabolic activity assay revealed Lic-A, 1i, and 1k derivatives as the most promising candidates. To delve into their mechanism of action, caspase activity assay was conducted in 2D and 3D in vitro models. Notably, apoptosis and autophagic induction was generally observed for Lic-A and 1k. The invasion assay demonstrated that Lic-A and 1k possess the ability to mitigate the spread of osteosarcoma cells within a matrix. The effectiveness of chalcone as a natural scaffold for generating potential antiproliferative agents against osteosarcoma has been demonstrated. In particular, chalcones exert their antiproliferative activity by inducing apoptosis and autophagy, and in addition they are capable of reducing cell invasion. These findings suggest Lic-A and 1k as promising antitumor agents against osteosarcoma cells.

Three-dimensional co-culturing of stem cell-derived cardiomyocytes and cardiac fibroblasts reveals a role for both cell types in Marfan-related cardiomyopathy
2024
Jeffrey Aalders, Laurens Léger, Louis Van der Meeren, Sanjay Sinha, Andre G Skirtach, Julie De Backer, Jolanda van Hengel

Live Microscopy of Multicellular Spheroids with the Multimodal Near-Infrared Nanoparticles Reveals Differences in Oxygenation Gradients
2024
Angela C. Debruyne, Irina A. Okkelman, Nina Heymans, Cláudio Pinheiro, An Hendrix, Max Nobis, Sergey M. Borisov, Ruslan I. Dmitriev

The correlation between TRIM28 expression and immune checkpoints in CRPC
2024
Dun Xue, Qian Zuo, Jie Chang, Xinghui Wu

The protective effect of silver nanoparticles’ on epithelial cornea cells against ultraviolet is accompanied by changes in calcium homeostasis and a decrease of the P2X7 and P2Y2 receptors
2024
Patryk Krzeminski, Irena Misiewicz Krzeminska, Marta Grodzik, Irena Padzinka Pruszynska, Paulina Kaucharzewska, Agnieszka Ostrowska, Ewa Sawosz, Pawel Pomorski

Extracellular vesicles from II trimester human amniotic fluid as paracrine conveyors counteracting oxidative stress
2024
Giorgia Senesi, Laura Guerricchio, Maddalena Ghelardoni, Nadia Bertola, Stefano Rebellato, Nicole Grinovero, Martina Bartolucci, Ambra Costa, Andrea Raimondi

A Novel ceRNET Relying on the lncRNA JPX, miR-378a-3p, and Its mRNA Targets in Lung Cancer
2024
Nicola Mosca, Mariaceleste Pezzullo, Ilenia De Leo, Anna Truda, Giovanna Marchese, Aniello Russo, Nicoletta Potenza

Induced pluripotent stem cell-derived human macrophages as an infection model for Leishmania donovani
2024
Lore Baert, Serena Rudy, Mélanie Pellisson, Thierry Doll, Romina Rocchetti, Marcel Kaiser, Pascal Mäser, Matthias Müller

AVJ16 inhibits the RNA binding protein IGF2BP1 in lung adenocarcinomas and prevents tumor growth in mice
2024
Nadav Wallis, Tehila Gershon, Sojod Shaaby, Froma Oberman, Myriam Grunewald, Deborah Duran, Amandeep Singh, Gilad Vainer, Vladimir S. Spiegelman, Arun K. Sharma

Dexamethasone Selectively Inhibits Detachment of Metastatic Thyroid Cancer Cells during Random Positioning
2023
Daniela Melnik, José Luis Cortés-Sánchez, Viviann Sandt, Stefan Kahlert, Sascha Kopp, Daniela Grimm, Marcus Krüger

A Biomimetic Polynucleotides–Hyaluronic Acid Hydrogel Promotes the Growth of 3D Spheroid Cultures of Gingival Fibroblasts
2023
Maria Teresa Colangelo, Maria Luisa Vicedomini, Silvana Belletti, Paolo Govoni, Stefano Guizzardi, Carlo Galli

Single-cell transcriptomics of NRAS-mutated melanoma transitioning to drug resistance reveals P2RX7 as an indicator of early drug response
2023
Tijana Randic, Stefano Magni, Demetra Philippidou, Christiane Margue, Kamil Grzyb, Jasmin Renate Preis, Joanna Patrycja Wroblewska

Expression and role of the immune checkpoint regulator PD-L1 in the tumor-stroma interplay of pancreatic ductal adenocarcinoma
2023
Tina Daunke, Silje Beckinger, Sascha Rahn, Sandra Krüger, Steffen Heckl, Heiner Schäfer, Daniela Wesch, Christian Pilarsky, Markus Eckstein, Arndt Hartmann

deCLUTTER2+ – a pipeline to analyze calcium traces in a stem cell model for ventral midbrain patterned astrocytes
2023
Martyna M. Grochowska, Federico Ferraro, Ana Carreras Mascaro, Domenico Natale, Amber Winkelaar, Valerie Boumeester, Guido J. Breedveld, Vincenzo Bonifati

I-CBP112 declines overexpression of ATP-binding cassette transporters and sensitized drug-resistant MDA-MB-231 and A549 cell lines to chemotherapy drugs
2023
Magdalena Strachowska, Karolina Gronkowska, Maciej Sobczak, Marika Grodzicka, Sylwia Michlewska, Kinga Kołacz, Tuhin Sarkar, Joanna Korszun, Maksim Ionov, Agnieszka Robaszkiewicz

Propranolol, Promising Chemosensitizer and Candidate for the Combined Therapy through Disruption of Tumor Microenvironment Homeostasis by Decreasing the Level of Carbonic Anhydrase IX
2023
Barbora Puzderova, Petra Belvoncikova, Katarina Grossmannova, Lucia Csaderova, Martina Labudova, Silvia Fecikova, Jaromir Pastorek

In Silico Identification and In Vitro Evaluation of New ABCG2 Transporter Inhibitors as Potential Anticancer Agents
2023
Simone Di Micco, Veronica Di Sarno, Martina Rossi, Vincenzo Vestute, Takumi Konno, Sara Novi, Mario Felice Tecce, Valeria Napolitano, Tania Ciaglia

Thyroid-Friendly Soft Materials as 3D Cell Culture Tool for Stimulating Thyroid Cell Function
2023
Indong Jun, Hyunki Cho, Sebastian E. Amos, Youngjun Choi, Yu Suk Choi, Chang Seon Ryu, Sang-Ah Lee, Dong-Wook Han, Hyung-Seop Han, Ji Hun Yang, Hyun-Woo Jeong, Honghyun Park

Redox double-switch cancer theranostics through Pt(IV) functionalised manganese dioxide nanostructures
2023
Beatriz Brito, Maria Rosaria Ruggiero, Thomas W. Price, Milene da Costa Silva, Núria Genicio, Annah J. Wilson, Olga Tyurina, Veronika Rosecker, Thomas R. Eykyn, Manuel Bañobre-López, Graeme J. Stasiuk

Mechanistic, Functional, and Clinical Aspects of Pro-inflammatory Cytokine Mediated Regulation of ADME Gene Expression in 3D Human Liver Spheroids
2023
Katharina Klöditz, Eida Tewolde, Åsa Nordling, Magnus Ingelman-Sundberg

Butein-instigated miR-186-5p-dependent modulation of TWIST1 affects resistance to cisplatin and bioenergetics of Malignant Pleural Mesothelioma cells
2022
Mario Cioce, Daniela Rutigliano, Vito Michele Fazio

Acanthamoeba spp. aggregate and encyst on contact lens material increasing resistance to disinfection
2022
Allison Campolo, Reed Pifer, Rhonda Walters, Megan Thomas, Elise Miller, Valerie Harris, Jamie King, Christopher A. Rice, Paul Shannon, Brian Patterson, Monica Crary

Novel artemisinin derivative FO8643 with anti-angiogenic activity inhibits growth and migration of cancer cells via VEGFR2 signaling
2022
Xiaohua Lu, Sebastian Blatt, Mona Dawood, Sabine M. Klauck, Edmond Fleischer

Integration of Bioinspired Fibrous Strands with 3D Spheroids for Environmental Hazard Monitoring
2022
Chang Gyun Park, Indong Jun, Sangmin Lee, Chang Seon Ryu, Sang-Ah Lee, Jaeho Park, Hyung-Seop Han, Honghyun Park, Andreas Manz, Heungsoo Shin

Artemisinin derivative FO-ARS-123 as a novel VEGFR2 inhibitor suppresses angiogenesis, cell migration, and invasion
2022
Xiaohu Lu, Mohamed Elbadawi, Sebastian Blatt, Mohamed E.M. Saeed, Xiaolin Xiao, Xiao Ma, Edmond Fleischer, Peer W. Kammerer

Setup of human liver-chips integrating 3D models, microwells and a standardized microfluidic platform as proof-of-concept study to support drug evaluation
2022
Benoit Cox, Patrick Barton, Reiner Class, Hannah Coxhead, Claude Delatour, Eric Gillent, Jamie Henshall, Emre M. Isin

A Systematic Comparative Assessment of the Response of Ovarian Cancer Cells to the Chemotherapeutic Cisplatin in 3D Models of Various Structural and Biochemical Configurations
2022
Priyanka Gupta, Aline Miller, Adedamola Olayanju, Thumuluru Kavitha Madhuri, Eirini Velliou

Hepatocyte Thorns, A Novel Drug-Induced Stress Response in Human and Mouse Liver Spheroids
2022
Chris S. Pridgeon, Dian P. Bolhuis, Filip Milosavljević, Marina Manojlović, Ákos Végvári, Massimiliano Gaetani, Marin M. Jukić, Magnus Ingelman-Sundberg

Evaluation of different methodologies for primary human dermal fibroblast spheroid formation: automation through 3D bioprinting technology
2022
Cristina Quílez, Enrique Cerdeira, Jorge González-Rico, Gonzalo de Aranda, Maria Luisa López-Donaire, José Luis Jorcano, Diego Velasco

3,4-Dihydroxyphenylethanol (DPE or Hydroxytyrosol) Counteracts ERK1/2 and mTOR Activation, Pro-Inflammatory Cytokine Release, Autophagy and Mitophagy Reduction Mediated by Benzo[a]pyrene in Primary Human Colonic Epithelial Cells
2022
Roberta Santarelli, Chiara Pompili, Maria Saveria Gilardini Montani, Lorenzo Evangelista, Roberta Gonnella, Mara Cirone

Arachidonic acid drives adaptive responses to chemotherapy-induced stress in malignant mesothelioma
2021
Mario Cioce, Claudia Canino, Harvey Pass, Giovanni Blandino, Sabrina Strano, Vito Michele Fazio

Methodology for the production of human hair follicles
2021
Cristina Quílez López

The choice of ultra-low attachment plates impacts primary human and primary canine hepatocyte spheroid formation, phenotypes, and function
2024 Biotechnology Journal. Systems & Synthetic Biology · Nanobiotech · Medicine.
Chen Xing, Aurino Kemas, Evgeniya Mickols, Kathrin Klein, Per Artursson, Volker M. Lauschke

Organotypic three-dimensional liver spheroid cultures in which hepatic cells retain their molecular phenotype and functionality have emerged as powerful tools for preclinical drug development. In recent years a multitude of culture systems have been developed; however, a thorough side-by-side benchmarking of the different methods is lacking. Here, we compared the performance of ten different 96- and 384-well microplate types to support spheroid formation and long-term culture. Specifically, we evaluated differences in spheroid formation kinetics, viability, functionality, expression patterns, and their utility for hepatotoxicity assessments using primary human hepatocytes (PHH) and primary canine hepatocytes (PCH). All 96-well plates enabled formation of PHH liver spheroids, albeit with differences between plates in spheroid size, geometry, and reproducibility. Performance of different 384-wells was less consistent. Only 6/10 microplates supported the formation of PCH aggregates. Interestingly, even if PCH aggregates in these six microplates were more loosely packed than PHH spheroids, they maintained their function and were compatible with long-term pharmacological and toxicological assays. Overall, Corning and Biofloat plates showed the best performance in the formation of both human and canine liver spheroids with highest viability, most physiologically relevant phenotypes, superior CYP activity and lowest coefficient of variation in toxicity assays. The presented data constitutes a valuable resource that demonstrates the impacts of current ultra-low attachment plates on liver spheroid metrics and can guide evidence-based plate selection. Combined, these results have important implications for the cross-comparison of different studies and can facilitate the standardization and reproducibility of three-dimensional liver culture experiments.

Metabolic reprogramming driven by EZH2 inhibition depends on cell–matrix interactions
2024 Journal of Biological Chemistry (JBC).
Teresa W-M Fan, Jahid M.M. Islam, Richard M. Higashi, Penghui Lin, Christine F. Brainson

EZH2 (Enhancer of Zeste Homolog 2), a subunit of Polycomb Repressive Complex 2 (PRC2), catalyzes the trimethylation of histone H3 at lysine 27 (H3K27me3), which represses expression of genes. It also has PRC2-independent functions, including transcriptional coactivation of oncogenes, and is frequently overexpressed in lung cancers. Clinically, EZH2 inhibition can be achieved with the FDA-approved drug EPZ-6438 (tazemetostat). To realize the full potential of EZH2 blockade, it is critical to understand how cell-cell/cell-matrix interactions present in 3D tissue and cell culture systems influences this blockade in terms of growth-related metabolic functions. Here, we show that EZH2 suppression reduced growth of human lung adenocarcinoma A549 cells in 2D cultures but stimulated growth in 3D cultures. To understand the metabolic underpinnings, we employed [13C6]-glucose stable isotope-resolved metabolomics to determine the effect of EZH2 suppression on metabolic networks in 2D versus 3D A549 cultures. The Krebs cycle, neoribogenesis, γ-aminobutyrate metabolism, and salvage synthesis of purine nucleotides were activated by EZH2 suppression in 3D spheroids but not in 2D cells, consistent with the growth effect. Using simultaneous 2H7-glucose + 13C5,15N2-Gln tracers and EPZ-6438 inhibition of H3 trimethylation, we delineated the effects on the Krebs cycle, γ-aminobutyrate metabolism, gluconeogenesis, and purine salvage to be PRC2-dependent. Furthermore, the growth/metabolic effects differed for mouse Matrigel versus self-produced A549 extracellular matrix. Thus, our findings highlight the importance of the presence and nature of extracellular matrix in studying the function of EZH2 and its inhibitors in cancer cells for modeling the in vivo outcomes.

Assembling Spheroids of Rat Primary Neurons Using a Stress-Free 3D Culture System
2023
Meaghan E. Harley-Troxell, Madhu Dhar

O2-sensitive microcavity arrays: A new platform for oxygen measurements in 3D cell cultures
2023
Christoph Grun, Jana Pfeier, Gregor Liebsch, Eric Gottwald

The choice of ultra-low attachment plates impacts primary human and primary canine hepatocyte spheroid formation, phenotypes, and function
2022
Indra Van Zundert, Nina Maenhoudt, Silke De Vriendt, Hugo Vankelecom, Beatrice Fortuni, Susana Rocha

Setup of human liver-chips integrating 3D models, microwells and a standardized microfluidic platform as proof-of-concept study to support drug evaluation
2022
Benoit Cox, Patrick Barton, Reiner Clas, Hannah Coxhead, Claude Delatour, Eric Gillent, Jamie Henshall, Emre M. Isin

Self-Assembled Silk Fibroin-Based Aggregates for Delivery of Camptothecin
2022
Javier Pérez Quiñones, Cornelia Roschger, Andreas Zierer, Carlos Peniche-Covas, Oliver Brüggemann

Polyphosphazene-Based Nanocarriers for the Release of Camptothecin and Epirubicin
2022
Javier Pérez Quiñones, Cornelia Roschger, Aitziber Iturmendi, Helena Henke, Andreas Zierer, Carlos Peniche-Covas, Oliver Brüggemann

Self-Assembled Silk Fibroin-Based Aggregates for Delivery of Camptothecin
2021
Javier Pérez Quiñones, Cornelia Roschger, Andreas Zierer, Carlos Peniche-Covas, Oliver Brüggemann