Cigarette Smoke Exposure as a Model of Inflammation Associated with COPD

Suffwan Eltom1, Christopher Stevenson2, Mark A. Birrell1

1 Centre for Integrative Mammalian Physiology and Pharmacology, Imperial College London, London, United Kingdom, 2 University of Southern Denmark, Institute for Medical Biology, Odense, Denmark
Publication Name:  Current Protocols in Pharmacology
Unit Number:  Unit 5.64
DOI:  10.1002/0471141755.ph0564s60
Online Posting Date:  March, 2013
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Chronic obstructive pulmonary disease (COPD) is characterized by progressive airflow limitation resulting from inflammation‐driven pathologies in the lungs that are a consequence of smoking over many years. Given that the disease is increasing globally, understanding the mechanism by which cigarette smoke (CS) causes lung inflammation and exploiting that knowledge to develop effective treatments is urgently required. Animal models of CS exposure are commonly used to examine the inflammatory processes that may be involved in the development of COPD. The protocols described in this unit detail the development of preclinical models of CS‐driven lung inflammation. These systems can be utilized to investigate the role of various biological pathways in CS‐mediated inflammation and to assess the efficacy of new therapeutic strategies for treating COPD. Curr. Protoc. Pharmacol. 60:14.24.1‐14.24.18. © 2013 by John Wiley & Sons, Inc.

Keywords: animal models; airways; COPD; emphysema; inflammation; lung

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Table of Contents

  • Introduction
  • Basic Protocol 1: Setup of the Cigarette Smoke Exposure System
  • Basic Protocol 2: Cigarette Smoke Exposure Protocol
  • Basic Protocol 3: Determination of Lung Inflammation Following CS Exposure
  • Commentary
  • Literature Cited
  • Figures
  • Tables
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Basic Protocol 1: Setup of the Cigarette Smoke Exposure System

  • 136‐liter exposure chambers (Teague Enterprises)
  • Dayton 2C820 extraction unit (Grainger Industrial Supply)
  • Electric fan (12 V DC, 6‐cm diameter, RS Components)
  • Perspex smoke extraction box (300 mm × 200 mm × 150 mm) (custom made in‐house)
  • Time‐set cigarette pinch valve (C Lee Machining)
  • Strong adhesive tape
  • AC/DC adapter (2 to 12 V; RS Components Ltd.)
  • Ashtray (100 mm × 10 mm × 40 mm)
  • Rubber bungs/stoppers, 30‐mm (VWR International)
  • Masterflex silicone tubing L/S 15 (Cole‐Parmer)
  • PVC hose 30‐mm diameter (4 m)

Basic Protocol 2: Cigarette Smoke Exposure Protocol

  • Male C57BL/6 mice (approx. 18 to 20 g), Sprague Dawley rats (approx. 200 to 225 g), or Dunkin‐Hartley guinea pigs (approx. 225 to 275 g)
  • Glycerol (Sigma‐Aldrich)
  • 70% ethanol
  • Trigene solution
  • Appropriate animal housing cages [i.e., use of individually vented cages (IVCs) reduces the smell of smoke outside of the eages]
  • Filtered research cigarettes [University of Kentucky Research Cigarettes ( #3R4F]
  • Large food container (for cigarettes)
  • Bench coat (VWR International)
  • Stainless steel animal smoking cages (400 mm × 320 mm × 150 mm; Adnor)
  • CS exposure system ( protocol 1)
  • Flow Meter, 4 liters/min (Dwyer, cat. no. VFB‐65)
  • Flashlight
  • Pocket lighter
  • Stopwatch or laboratory timer
  • Metal forceps (small size)
  • Total suspended particulate sampling unit (Teague Enterprises)
    • Air sampling pump
    • Dry gas meter
    • Filter holder
    • Tubing
  • PallFlex 25‐mm total suspended particulate (TSP) membrane filters (VWR International, Emfab #TX40H120‐WW)
  • Cotton‐tipped applicators (VWR International)

Basic Protocol 3: Determination of Lung Inflammation Following CS Exposure

  • Mice exposed to CS ( protocol 2)
  • Pentobarbitone
  • RPMI 1640 medium + GlutaMAX‐I (RPMI; Invitrogen)
  • Fetal bovine serum (FBS; Gibco, Invitrogen)
  • Liquid nitrogen (optional)
  • Collagenase (Roche Diagnostics)
  • DNase (Roche Diagnostics)
  • Penicillin/streptomycin stock (25,000 U/ml penicillin, 25 mg/ml streptomycin; Roche Diagnostics)
  • Reference blood samples
  • Wright‐Giemsa stain (Sigma)
  • 1‐ml syringes, with and without 23‐gauge needles
  • Surgical tools (forceps, small scissors, fine scissors)
  • Cannula with luer connection
  • Microcentrifuge tubes (2 ml)
  • McIlwain tissue chopper (Campden Instruments)
  • Grant OLS 200 shaking water bath (Camlab)
  • 70‐µm cell sieve
  • Sysmex cell counter (Sysmex, Milton Keynes)
  • Cytospin centrifuge (Shandon)
  • Hema‐tek 2000 automated slide stainer (Ames)
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Literature Cited

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