Preserving Airway Smooth Muscle Contraction in Precision-Cut Lung Slices

Guang Li,Jonathan A Cohen,Carolina Martines,Yan Bai,Xingbin Ai,Joseph D Brain,Ramaswamy Krishnan,Sumati Ram-Mohan

doi:10.1038/s41598-020-63225-y

Abstract

Precision-cut lung slices (PCLS) are ideal for measuring small airway contraction. However, these measurements are currently limited to acute exposure scenarios that typically last a few minutes to a few hours. Using an insulin-supplemented culture medium, we prolong the small airway contractility in mouse PCLS for up to two weeks. Compared to conventional culture medium, insulin-supplemented culture medium provides no additional benefit in preserving cellular viability or airway structure. However, it protects the airway smooth muscle (ASM) against a loss of smooth muscle myosin heavy chain (SMMHC) expression. We elucidate the significance of this new culture medium for chronic disease modeling of IL-13-induced airway hyper-responsiveness.

Highlights

A powerful tool for studying airway reactivity is the preparation of precision-cut lung slices (PCLS)
These results show that insulin has a beneficial effect on the maintenance of airway contractility of Precision-cut lung slices (PCLS)
The structure of the airway was intact for at least 15 days in both PCLS cultures (Fig. 2a). These findings indicate that the preservation of airway responsiveness in PCLS conferred by the DMF12-Ins culture medium is not due to an improvement of cell viability, maintaining cell viability is critical for airway smooth muscle (ASM) contraction

Summary

Introduction

A powerful tool for studying airway reactivity is the preparation of precision-cut lung slices (PCLS). PCLS can be prepared from naïve, genetically modified, or diseased animals as well as from human surgical samples and lung explants, and even cryopreserved for future use[9,10] They are ideally suited to bridge the translational gap between cell culture models and whole animal or human subject studies. To preserve airway contraction in PCLS, we test insulin supplementation to the conventional DMEM/F12 (DMF12) medium We show that this newly formulated insulin supplemented DMF12 (DMF12-Ins) medium preserves contractile proteins in airway smooth muscle cells and prolongs the airway contractile function in the mouse PCLS for at least two weeks. This improved PCLS culture facilitates chronic disease modeling, such as Interleukin (IL)-13-induced airway hyper-reactivity

Methods

Results

Conclusion