Advanced pulmonary models

Abstract

The present invention relates to microfluidic fluidic systems and methods for the in vitro modeling diseases of the lung and small airway. In one embodiment, the invention relates to a system for testing responses of a microfluidic Small Airway-on-Chip infected with one or more infectious agents (e.g. respiratory viruses) as a model of respiratory disease exacerbation (e.g. asthma exacerbation). In one embodiment, this disease model on a microfluidic chip allows for a) the testing of anti-inflammatory and/or anti-viral compounds introduced into the system, as well as b) the monitoring of the participation, recruitment and/or movement of immune cells, including the transmigration of cells. In particular, this system provides, in one embodiment, an in-vitro platform for modeling severe asthma as “Severe Asthma-on-Chip.” In some embodiments, this invention provides a model of viral-induced asthma in humans for use in identifying potentially effective treatments.

What is claimed is: 
     
         1 . A method of treating cells, comprising:
 a) providing a microfluidic device and respiratory cells;   b) stimulating said respiratory cells in said microfluidic device, or stimulating said respiratory cells outside said microfluidic device, so as to create stimulated cells, said stimulated cells exhibiting a respiratory disease phenotype, wherein when said respiratory cells are stimulated outside said microfluidic device they are thereafter introduced into said microfluidic device; and   c) contacting said stimulated cells with a microorganism or virus to create stimulated cells with a severe respiratory disease phenotype.   
     
     
         2 . The method of  claim 1 , wherein said respiratory disease phenotype is an asthma phenotype. 
     
     
         3 . The method of  claim 1 , wherein said respiratory disease phenotype is a COPD phenotype or a CF phenotype. 
     
     
         4 . The method of  claim 1 , wherein said stimulated cells in said microfluidic device are subject to flow conditions. 
     
     
         5 . The method of  claim 4 , wherein said flow conditions comprise flowing air and/or flowing culture media. 
     
     
         6 . The method of  claim 4 , wherein said flow conditions comprise flowing blood. 
     
     
         7 . The method of  claim 1 , wherein said stimulating comprises exposing said respiratory cells to a cytokine. 
     
     
         8 . The method of  claim 7 , wherein said cytokine is an interleukin. 
     
     
         9 . The method of  claim 8 , wherein said interleukin is IL-13. 
     
     
         10 . The method of  claim 1 , wherein said respiratory cells are in a layer on a surface of said microfluidic device. 
     
     
         11 . The method of  claim 10 , wherein said surface is part of a channel. 
     
     
         12 . The method of  claim 10 , wherein said surface is a membrane. 
     
     
         13 . The method of  claim 1 , wherein said respiratory cells are lung epithelial cells. 
     
     
         14 . The method of  claim 1 , wherein said microfluidic device comprises a channel and said respiratory cells are disposed in said channel. 
     
     
         15 . The method of  claim 1 , further comprising the step of d) imaging the stimulated cells after step c). 
     
     
         16 . The method of  claim 1 , further comprising the step of d) measuring the concentration of at least one cytokine. 
     
     
         17 . The method of  claim 1 , further comprising the step of d) detecting hyperplasia. 
     
     
         18 . The method of  claim 17 , wherein said hyperplasia is goblet cell hyperplasia. 
     
     
         19 . The method of  claim 13 , wherein said lung epithelial cells are on a first surface of a membrane and endothelial cells are on a second surface of a membrane. 
     
     
         20 . The method of  claim 19 , further comprising the step of adding immune cells to said microfluidic device. 
     
     
         21 . The method of  claim 20 , further comprising the step of measuring the extent of recruitment of said immune cells into the endothelium and/or epithelium. 
     
     
         22 . The method of  claim 20 , wherein said immune cells comprise neutrophils. 
     
     
         23 . The method of  claim 1 , wherein said stimulated cells of step b) are contacted by bacteria in step c). 
     
     
         24 . The method of  claim 1 , wherein said stimulated cells of step b) are contacted by fungi in step c). 
     
     
         25 . The method of  claim 1 , wherein said stimulated cells of step b) are contacted by a virus in step c). 
     
     
         26 . The method of  claim 25 , wherein said stimulated cells of step b) are contacted by a respiratory virus in step c). 
     
     
         27 . The method of  claim 26 , wherein said respiratory virus is selected from the group consisting of parainfluenza virus, influenza virus, rhinovirus, coronaviruses, human respiratory syncytial virus, and adenoviruses. 
     
     
         28 . The method of  claim 1 , where said respiratory cells are bronchial cells. 
     
     
         29 . The method of  claim 28 , wherein said bronchial cells are mucociliary bronchiolar airway epithelial cells. 
     
     
         30 . The method of  claim 1 , further comprising measuring cell size and/or number for at least one population of cells present in the microfluidic device. 
     
     
         31 . A method of treating cells, comprising:
 a) providing a microfluidic device and respiratory cells derived from a patient with a respiratory disease; and   b) contacting said respiratory cells with a microorganism or virus in said microfluidic device, or contacting said respiratory cells with a microorganism outside said microfluidic device, so as to create a severe respiratory disease phenotype wherein when said respiratory cells are contacted outside said microfluidic device they are thereafter introduced into said microfluidic device.   
     
     
         32 . The method of  claim 31 , wherein said respiratory disease phenotype is asthma. 
     
     
         33 . The method of  claim 31 , wherein said respiratory disease is CF or COPD. 
     
     
         34 . The method of  claim 31 , wherein said respiratory cells in said microfluidic device are subject to flow conditions. 
     
     
         35 . The method of  claim 33 , wherein said flow conditions comprise flowing air and/or flowing culture media. 
     
     
         36 . The method of  claim 31 , wherein said respiratory cells are contacted by bacteria in step b). 
     
     
         37 . The method of  claim 31 , wherein said respiratory cells are contacted by fungi in step b). 
     
     
         38 . The method of  claim 31 , wherein said respiratory cells are contacted by a virus in step b). 
     
     
         39 . The method of  claim 38 , wherein said respiratory cells are contacted by a respiratory virus in step b). 
     
     
         40 . The method of  claim 39 , wherein said respiratory virus is selected from the group consisting of parainfluenza virus, influenza virus, rhinovirus, coronaviruses, human respiratory syncytial virus, and adenoviruses. 
     
     
         41 . The method of  claim 31 , where said respiratory cells are bronchial cells. 
     
     
         42 . The method of  claim 31 , further comprising c) exposing said respiratory cells to a test agent. 
     
     
         43 . The method of  claim 31 , further comprising the step of c) imaging the cells after step b). 
     
     
         44 . The method of  claim 31 , further comprising the step of c) measuring the concentration of at least one cytokine. 
     
     
         45 . The method of  claim 31 , further comprising the step of c) detecting hyperplasia. 
     
     
         46 . The method of  claim 45 , wherein said hyperplasia is goblet cell hyperplasia. 
     
     
         47 . The method of  claim 31 , further comprising measuring cell size and/or number for at least one population of cells present in the microfluidic device. 
     
     
         48 . A method of treating cells, comprising:
 a) providing a microfluidic device comprising a first channel and a second channel, said first channel comprising respiratory epithelial cells, said second channel comprising endothelial cells;   b) stimulating said respiratory epithelial cells to create stimulated epithelial cells with a respiratory disease phenotype; and   c) contacting said stimulated epithelial cells with a microorganism or virus to create stimulated epithelial cells with a severe respiratory disease phenotype.   
     
     
         49 . The method of  claim 48 , wherein said first channel is an upper channel and said second channel is a lower channel separated from said upper channel by a membrane. 
     
     
         50 . The method of  claim 48 , further comprising exposing said respiratory epithelial cells to an air interface. 
     
     
         51 . The method of  claim 48 , further comprising exposing said endothelial cells to culture media at a flow rate. 
     
     
         52 . The method of  claim 48 , wherein said respiratory epithelial cells are bronchial cells. 
     
     
         53 . The method of  claim 52 , wherein said bronchial cells are mucociliary bronchiolar airway epithelial cells. 
     
     
         54 . The method of  claim 48 , wherein said endothelial cells are microvascular endothelial cells. 
     
     
         55 . The method of  claim 48 , further comprising exposing said stimulated cells to a test agent. 
     
     
         56 . The method of  claim 55 , wherein said test agent comprises a candidate drug. 
     
     
         57 . The method of  claim 55 , wherein said test agent comprises a CXCR2 inhibitor. 
     
     
         58 . The method of  claim 56 , wherein said candidate drug reduces or prevents a severe respiratory disease phenotype. 
     
     
         59 . The method of  claim 55 , wherein said test agent is a steroid. 
     
     
         60 . The method of  claim 48 , wherein said respiratory disease phenotype is an asthma phenotype. 
     
     
         61 . The method of  claim 48 , wherein said respiratory disease phenotype is a COPD phenotype or CF phenotype. 
     
     
         62 . The method of  claim 48 , further comprising the step of detecting hyperplasia. 
     
     
         63 . The method of  claim 62 , wherein said hyperplasia is goblet cell hyperplasia. 
     
     
         64 . The method of  claim 62 , wherein said hyperplasia is detected by counting cells. 
     
     
         65 . The method of  claim 62 , wherein said hyperplasia is detected by measuring cell size. 
     
     
         66 . The method of  claim 48 , further comprising measuring cell size and/or number for at least one population of cells present in the microfluidic device. 
     
     
         67 . A method of treating cells, comprising:
 a) providing respiratory cells derived from a patient with a respiratory disease and a microfluidic device, said microfluidic device comprising a first channel and a second channel, said first channel comprising respiratory epithelial cells, said second channel comprising endothelial cells; and   b) contacting said respiratory cells with a microorganism or virus in said microfluidic device, or contacting said respiratory cells with a microorganism outside said microfluidic device, so as to create a severe respiratory disease phenotype wherein when said respiratory cells are contacted outside said microfluidic device they are thereafter introduced into said microfluidic device.   
     
     
         68 . The method of  claim 67 , further comprising measuring cell size and/or number for at least one population of cells present in the microfluidic device. 
     
     
         69 . The method of  claim 67 , wherein said microorganism is  P. aeruginosa.