The Challenge of Long-Term Cultivation of Human Precision-Cut Lung Slices

Preuß, Eike B.; Schubert, Stephanie; Werlein, Christopher; Stark, Helge; Braubach, Peter; Höfer, Anne; Plucinski, Edith; Shah, Harshit; Geffers, Robert; Sewald, Katherina; Braun, Armin; Jonigk, Danny; Kühnel, Mark P.

doi:10.1016/j.ajpath.2021.10.020

Cited by 25 publications

(33 citation statements)

References 60 publications

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“…They reported sustained viability, reactivity and structural integrity of PCLS for the investigated time period. Recently, viability of human PCLS for up to 4 weeks could be shown, however, a continuous cell loss, decrease in ciliary beat and increasing manifestation of keratinizing squamous metaplasia over time were observed [ 26 ].…”

Section: Introductionmentioning

confidence: 99%

Functional changes in long-term incubated rat precision-cut lung slices

et al. 2022

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Background Respiratory diseases represent a global health burden. Because research on therapeutic strategies of airway diseases is essential, the technique of precision-cut lung slices (PCLS) has been developed and widely studied. PCLS are an alternative ex vivo model and have the potential to replace and reduce in vivo animal models. So far, the majority of studies was conducted with short-term cultivated PCLS (≤ 72 h). As there is large interest in research of chronic diseases and chronic toxicity, feasibility of cultivating human PCLS long-term over 2 weeks and recently over 4 weeks was investigated by another research group with successful results. Our aim was to establish a model of long-term cultivated rat PCLS over a period of 29 days. Methods Rat PCLS were cultured for 29 days and analysed regarding viability, histopathology, reactivity and gene expression at different time points during cultivation. Results Cultivation of rat PCLS over a 29-day time period was successful with sustained viability. Furthermore, the ability of bronchoconstriction was maintained between 13 and 25 days, depending on the mediator. However, reduced relaxation, altered sensitivity and increased respiratory tone were observed. Regarding transcription, alteration in gene expression pattern of the investigated target genes was ascertained during long-term cultivation with mixed results. Furthermore, the preparation of PCLS seems to influence messenger ribonucleic acid (mRNA) expression of most target genes. Moreover, the addition of fetal bovine serum (FBS) to the culture medium did not improve viability of PCLS. In contrast to medium without FBS, FBS seems to affect measurements and resulted in marked cellular changes of metaplastic and/or regenerative origin. Conclusions Overall, a model of long-term cultivated rat PCLS which stays viable for 29 days and reactive for at least 13 days could be established. Before long-term cultivated PCLS can be used for in-depth study of chronic diseases and chronic toxicity, further investigations have to be made.

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Section: Introductionmentioning

confidence: 99%

Functional changes in long-term incubated rat precision-cut lung slices

et al. 2022

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“…Unlike 2D monolayers or 3D well-differentiated air-liquid interface (ALI) cultures, PCLS preserve the native lung tissue architecture, cellular composition including endothelial, ATI and ATII epithelial cells, fibroblasts and maintain the native extracellular matrix [33][34][35] .…”

Section: Discussionmentioning

confidence: 99%

“…We utilized human precision-cut lung slices (hPCLS) as a biologically relevant ex vivo model that more faithfully represents lung tissues than either 2D monolayer cultures or 3D air-liquid-interface (ALI) cultures 33,34 . Following hPCLS preparation, they were in culture for up to 4 weeks; no gross alterations in cell type or morphology were observed and cilial beating was observed in all used hPCLS (8 donors; n=24).…”

Section: Human Pcls Support Robust Iav Infection and Confirm Inhibitory Effect Of Nrtki Treatmentmentioning

confidence: 99%

“…Tissues used for PCLS generation that were obtained from lung tumor resections were confirmed as tumor-free by an experienced pathologist. The freshly obtained lung tissues were processed into circular slices that were 300 microns thick and 8 mm in diameter as previously described 34 . All donors provided informed consent as approved by the Hannover Medical School Ethics Committee (Ethics vote #8867_BO_K_2020).…”

Section: Human Precision-cut Lung Slices (Pcls)mentioning

confidence: 99%

“…In the present study, we characterized the effect of eight currently available and FDA-approved NRTKIs on IAV replication, six of which were validated using a biologically relevant ex vivo model of human precision-cut lung slices (hPCLS). hPCLS maintain near-native structural integrity of lung tissues that allow complex and 3D cell-cell interactions of epithelial cells, mesenchymal tissue as well as the vascular compartment; thereby, offering an advantage over cultured epithelial monolayers [33][34][35] . We also identify which steps of the virus infection cycle are affected by specific NRTKIs; thereby providing insights into potential mechanisms of action in the context of influenza virus infections.…”

Section: Introductionmentioning

confidence: 99%

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Ex Vivo Validation of Six FDA-Approved Non-Receptor Tyrosine Kinase Inhibitors (NRTKIs) as Antivirals to Pandemic and Seasonal Influenza A Viruses

Meineke

Stelz

Busch

et al. 2022

Preprint

Self Cite

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Influenza viruses are important respiratory pathogens that cause substantial morbidity and mortality annually. In addition to seasonal influenza outbreaks, new emerging influenza A viruses (IAV) can cause pandemic influenza outbreaks. Apart from effective vaccines, there is a need for better treatment options to combat infections with these viruses when vaccines are not available or show reduced efficacy (e.g., in immmunocompromised patients). The limited range of licensed antiviral drugs and emergence of drug-resistance mutations highlight the need for novel intervention strategies like host-targeted antivirals. Repurposing FDA-approved kinase inhibitors may offer a fast-track for a new generation of host-targeted antivirals. Small molecule kinase inhibitors (SMKIs) can inhibit replication of viruses and improve survival in vivo; however, no SMKI has been approved for clinical use against IAV infections. In the present study, we tested eight non-receptor tyrosine kinase-inhibitors (NRTKIs) used to treat cancer and autoimmune diseases for their antiviral potential. Six of those potently inhibited virus replication (≥1,000-fold) in A549 cells infected with either A(H1N1)pdm09 or seasonal A(H3N2) strains. These compounds were validated in a biologically relevant ex vivo model of human precision-cut lung slices (hPCLS) to provide proof of principle and show efficacy against contemporary seasonal and pandemic IAVs. We identified the steps of the virus infection cycle affected by these inhibitors and assessed the effect of these NRTKIs on the host response. Considering their established safety profiles, our studies show that the use of these NRTKI shows promise and warrants further development as an alternative strategy to treat influenza virus infections.

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Hydrogel‐Embedded Precision‐Cut Lung Slices Model Lung Cancer Premalignancy Ex Vivo

Blomberg,

Sompel,

Hauer

et al. 2023

Adv Healthcare Materials

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Lung cancer is the leading global cause of cancer‐related deaths. Although smoking cessation is the best prevention, 50% of lung cancer diagnoses occur in people who have quit smoking. Research into treatment options for high‐risk patients has been constrained to rodent models, which are time‐consuming, expensive, and require large cohorts. Embedding precision‐cut lung slices (PCLS) within an engineered hydrogel and exposing this tissue to vinyl carbamate, a carcinogen from cigarette smoke, creates an in vitro model of lung cancer premalignancy. Hydrogel formulations are selected to promote early lung cancer cellular phenotypes and extend PCLS viability to six weeks. Hydrogel‐embedded PCLS are exposed to vinyl carbamate, which induces adenocarcinoma in mice. Analysis of proliferation, gene expression, histology, tissue stiffness, and cellular content after six weeks reveal that vinyl carbamate induces premalignant lesions with a mixed adenoma/squamous phenotype. Putative chemoprevention agents diffuse through the hydrogel and induce tissue‐level changes. The design parameters selected using murine tissue are validated with hydrogel‐embedded human PCLS and results show increased proliferation and premalignant lesion gene expression patterns. This tissue‐engineered model of human lung cancer premalignancy is the foundation for more sophisticated ex vivo models that enable study of carcinogenesis and chemoprevention strategies.This article is protected by copyright. All rights reserved

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The Challenge of Long-Term Cultivation of Human Precision-Cut Lung Slices

Cited by 25 publications

References 60 publications

Functional changes in long-term incubated rat precision-cut lung slices

Functional changes in long-term incubated rat precision-cut lung slices

Ex Vivo Validation of Six FDA-Approved Non-Receptor Tyrosine Kinase Inhibitors (NRTKIs) as Antivirals to Pandemic and Seasonal Influenza A Viruses

Hydrogel‐Embedded Precision‐Cut Lung Slices Model Lung Cancer Premalignancy Ex Vivo

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