Airway Contractility in the Precision-Cut Lung Slice after Cryopreservation

Rosner, Sonia R.; Ram-Mohan, Sumati; Paez-Cortez, Jesus; Lavoie, Tera; Dowell, Maria; Yuan, Lei; Ai, Xingbin; Fine, Alan; Aird, William C.; Solway, Julian; Fredberg, Jeffrey J.; Krishnan, Ramaswamy

doi:10.1165/rcmb.2013-0166ma

Cited by 41 publications

(54 citation statements)

References 46 publications

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“…There also appears to be a general decrease in cytokine production during cultivation indicating a change in the cell population or cell function within PCLS over time in culture. One potential way to overcome this physiological change would be to explore the use of cryopreservation as described in Rosner et al 27 Their results showed minimal cell death, mainly along the edges, but also some reduction in the metabolic activity of the tissue following a freeze-thaw cycle. However, further optimization of such a cryopreservation technique could help to practically address the short window of viability of PCLS in culture.…”

Section: Discussionmentioning

confidence: 99%

Evaluation of inflammatory and immune responses in long-term cultured human precision-cut lung slices

Temann

Golovina

Neuhaus

et al. 2016

Human Vaccines & Immunotherapeutics

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The development of systems that are more accurate and time-efficient in predicting safety and efficacy of target products in humans are critically important in reducing the cost and duration of pharmaceutical development. To circumvent some of the limitations imposed by the use of animal models, ex vivo systems, such as precision-cut lung slices (PCLS), have been proposed as an alternative for evaluating safety, immunogenicity and efficacy of vaccines and pharmaceuticals. In this study, we have established a human PCLS system and methodology for PCLS cultivation that can provide long-term viability and functionality in culture. Using these techniques, we found that cultured PCLS remained viable for at least 14 d in culture and maintained normal metabolic activity, tissue homeostasis and structural integrity. To investigate whether cultured PCLS remained functional, lipopolysaccharide (LPS) was used as a target stimulating compound. We observed that after an 18-hour incubation with LPS, cultured PCLS produced a set of pro-inflammatory cytokines, including TNF-a, IL-1b, IL-6 and IL-10 as well as the enzyme COX-2. Furthermore, cultured PCLS were shown to be capable of generating re-call immune responses, characterized by cytokine production, against antigens commonly found in routine vaccinations against influenza virus and tetanus toxoid. Taken together, these results suggest that human PCLS have the potential to be used as an alternative, high-throughput, ex vivo system for evaluating the safety, and potentially immunogenicity, of vaccines and pharmaceuticals.

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Section: Discussionmentioning

confidence: 99%

Evaluation of inflammatory and immune responses in long-term cultured human precision-cut lung slices

Temann

Golovina

Neuhaus

et al. 2016

Human Vaccines & Immunotherapeutics

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“…PCLSs have been widely used for studies of airway contractility and mechanisms of airway hyperreactivity in asthma (4,5,9,15,22,23). Because airway contraction requires the coordinated activity of ASMCs, this multicellular response requires high rates of ASMC viability after cryopreservation.…”

Section: Contractile and Relaxant Responses Of Airways In Cryopreservmentioning

confidence: 99%

“…Cryopreservation has been reported to store live biological samples with varying success (7,8), and its application to mouse PCLSs (mPCLSs) was previously assessed (9). This study employed histology and phase-contrast imaging to show that the lung tissue integrity and agonist-induced airway contraction were retained in cryopreserved mPCLSs.…”

mentioning

confidence: 99%

Cryopreserved Human Precision-Cut Lung Slices as a Bioassay for Live Tissue Banking. A Viability Study of Bronchodilation with Bitter-Taste Receptor Agonists

Bai

Krishnamoorthy

Patel

et al. 2016

Am J Respir Cell Mol Biol

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Human precision-cut lung slices (hPCLSs) provide a unique ex vivo model for translational research. However, the limited and unpredictable availability of human lung tissue greatly impedes their use. Here, we demonstrate that cryopreservation of hPCLSs facilitates banking of live human lung tissue for routine use. Our results show that cryopreservation had little effect on overall cell viability and vital functions of immune cells, including phagocytes and T lymphocytes. In addition, airway contraction and relaxation in response to specific agonists and antagonists, respectively, were unchanged after cryopreservation. At the subcellular level, cryopreserved hPCLSs maintained Ca 21 -dependent regulatory mechanisms for the control of airway smooth muscle cell contractility. To exemplify the use of cryopreserved hPCLSs in smooth muscle research, we provide evidence that bitter-taste receptor (TAS2R) agonists relax airways by blocking Ca 21 oscillations in airway smooth muscle cells. In conclusion, the banking of cryopreserved hPCLSs provides a robust bioassay for translational research of lung physiology and disease.

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“…Dose response curves for MCh (10 −9 –10 −3 M) were recorded, after which the airways were dilated using the bitter taste receptor agonist chloroquine (10 −3 M, Sigma-Aldrich). A nylon mesh and metal washer were used to fixate the lung slice, as described previously (Rosner et al, 2014). Lung slice images were captured in time-lapse (1 frame per 2 s) using a microscope (Eclipse, TS100; Nikon).…”

Section: Methodsmentioning

confidence: 99%

Elastase-Induced Parenchymal Disruption and Airway Hyper Responsiveness in Mouse Precision Cut Lung Slices: Toward an Ex vivo COPD Model

et al. 2017

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Background: COPD is a progressive lung disease characterized by emphysema and enhanced bronchoconstriction. Current treatments focused on bronchodilation can delay disease progression to some extent, but recovery or normalization of loss of lung function is impossible. Therefore, novel therapeutic targets are needed. The importance of the parenchyma in airway narrowing is increasingly recognized. In COPD, the parenchyma and extracellular matrix are altered, possibly affecting airway mechanics and enhancing bronchoconstriction. Our aim was to set up a comprehensive ex vivo Precision Cut Lung Slice (PCLS) model with a pathophysiology resembling that of COPD and integrate multiple readouts in order to study the relationship between parenchyma, airway functionality, and lung repair processes.Methods: Lungs of C57Bl/6J mice were sliced and treated ex vivo with elastase (2.5 μg/ml) or H2O2 (200 μM) for 16 h. Following treatment, parenchymal structure, airway narrowing, and gene expression levels of alveolar Type I and II cell repair were assessed.Results: Following elastase, but not H2O2 treatment, slices showed a significant increase in mean linear intercept (Lmi), reflective of emphysema. Only elastase-treated slices showed disorganization of elastin and collagen fibers. In addition, elastase treatment lowered both alveolar Type I and II marker expression, whereas H2O2 stimulation lowered alveolar Type I marker expression only. Furthermore, elastase-treated slices showed enhanced methacholine-induced airway narrowing as reflected by increased pEC50 (5.87 at basal vs. 6.50 after elastase treatment) and Emax values (47.96 vs. 67.30%), and impaired chloroquine-induced airway opening. The increase in pEC50 correlated with an increase in mean Lmi.Conclusion: Using this model, we show that structural disruption of elastin fibers leads to impaired alveolar repair, disruption of the parenchymal compartment, and altered airway biomechanics, enhancing airway contraction. This finding may have implications for COPD, as the amount of elastin fiber and parenchymal tissue disruption is associated with disease severity. Therefore, we suggest that PCLS can be used to model certain aspects of COPD pathophysiology and that the parenchymal tissue damage observed in COPD contributes to lung function decline by disrupting airway biomechanics. Targeting the parenchymal compartment may therefore be a promising therapeutic target in the treatment of COPD.

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Airway Contractility in the Precision-Cut Lung Slice after Cryopreservation

Cited by 41 publications

References 46 publications

Evaluation of inflammatory and immune responses in long-term cultured human precision-cut lung slices

Evaluation of inflammatory and immune responses in long-term cultured human precision-cut lung slices

Cryopreserved Human Precision-Cut Lung Slices as a Bioassay for Live Tissue Banking. A Viability Study of Bronchodilation with Bitter-Taste Receptor Agonists

Elastase-Induced Parenchymal Disruption and Airway Hyper Responsiveness in Mouse Precision Cut Lung Slices: Toward an Ex vivo COPD Model

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