Researchers at the Wyss Institute at Harvard University have developed an animal-free science innovation platform by combining a robotic smoking machine with a lung-on-a-chip technology to fully and realistically mimic human smoking patterns.
HUMAN HEALTH ISSUE
Evaluating the health impact of airborne compounds is critical to determining pathogenesis and mechanisms of injury. Cigarette smoking is the primary risk factor for the development of chronic obstructive pulmonary disease (COPD) and other lung disorders. In addition, tobacco-related products, such as e-cigarettes are gaining popularity; however, the biological impact of their emissions on the lung is poorly understood. Previous work by the Wyss Institute for Biologically Inspired Engineering at Harvard University reported how human organ-on-a-chip (organ chip) microfluidic cell culture technology could be leveraged to develop a breathing lung small airway-on-a-chip (small airway chip) lined with living human bronchiolar epithelium cultured at an air–liquid interface (ALI) that also permits smoke exposure. Although that work demonstrated the integration of physiologically accurate mechanical cues (i.e., fluidic shear and cyclic strain mimicking breathing in alveoli), there was still a need to develop both a small airway organ chip and a way of inducing physiological shear and dosing with smoke and other aerosols in the airway compartment to enable investigations of airway pathologies beyond the alveolus.
ANIMAL FREE SCIENCE INNOVATION POTENTIAL
This work builds on an earlier protocol used to develop microfluidic organ chips by incorporating a polyester track-etched membrane in combination with larger cross-section fluidic channels and a more streamlined fabrication process. Although the preparation of small airway chips is detailed in the protocol by Benam et al., here we connected the small airway chip microfluidically to a biomimetic smoking robot (BSR). This combination of respiration driven by a smoking robot—which inhales and exhales whole smoke from burning cigarettes in and out of the epithelium-lined microchannel under dynamic conditions—recapitulated human smoking behaviour. The BSR recapitulates the mechanics and dynamic inhalation behavior of a smoker to facilitate study of smoking styles or topographies under realistic conditions with dynamic smoke generation. The system consists of three components: (i) a smoke generator, (ii) a microrespirator and (iii) LabVIEW control software that couples the two hardware components to create programmable smoking behavior. The compact smoke generator holds up to ten cigarettes or e-cigarettes inside a revolving holder. The LabVIEW control software and interface not only link the two hardware components, they also enable programming of an arbitrary smoke topography. Ignition times, inhalation times, puff duration, inter-puff intervals and other parameters can be incorporated into the program to model the smoking typography of real-world smokers. The interface and hardware also leave room for the
automation of other components, such as additional valves or switches, which can be used to expand the current capability.
Exposure of lung tissues to cigarette smoke is a major cause of human disease and death worldwide. Unfortunately, adequate model systems that can reliably recapitulate disease biogenesis in vitro, including exposure of the human lung airway to fresh whole cigarette smoke (WCS) under physiological breathing airflow, are lacking. This protocol extension builds upon, and can be used with, our earlier protocol for microfabrication of human organs-on-chips. Here, we describe the engineering, assembly and operation of a microfluidically coupled, multi-compartment platform that bidirectionally ‘breathes’ WCS through microchannels of a human lung small airway microfluidic culture device, mimicking how lung cells may experience smoke in vivo. Several WCS-exposure systems have been developed, but they introduce smoke directly from above the cell cultures, rather than tangentially as naturally occurs in the lung due to lateral airflow. We detail the development of an organ chip-compatible microrespirator and a smoke machine to simulate breathing behavior and smoking topography parameters such as puff time, inter-puff interval and puffs per cigarette. Detailed design files, assembly instructions and control software are provided. This novel platform can be fabricated and assembled in days and can be used repeatedly. Moderate to advanced engineering and programming skills are required to successfully implement this protocol. When coupled with the small airway chip, this protocol can enable prediction of patient-specific biological responses in a matched-comparative manner. We also demonstrate how to adapt the protocol to expose living ciliated airway epithelial cells to smoke generated by electronic cigarettes (e-cigarettes) on-chip.
KH. Benam et al. Biomimetic smoking robot for in vitro inhalation exposure compatible with microfluidic organ chips. Nature Protocols. January 10, 2020. Epub ahead of print. https://doi.org/10.1038/s41596-019-0230-y.
D. Huh et al. Microfabrication of human organs-on-chips. Nature Protocols. November 8(11):2135-57. https://doi.org/10.1038/nprot.2013.137.