Utilizing a Human Lung Chip that reproduces the structures and functions of the lung air sac, or “alveolus,” the research group found that applying mechanical forces that mimic breathing motions suppresses influenza virus replication by activating protective inherent immune reactions.” This research study demonstrates the value of breathing movements for human lung function, including immune responses to infection, and reveals that our Human Alveolus Chip can be used to design these actions in the deep portions of the lung, where infections are often more serious and lead to hospitalization and death,” stated co-first author Haiqing Bai, Ph.D., a Wyss Technology Development Fellow at the Institute. Bai and his team first lined the two parallel microfluidic channels of an Organ Chip with different types of living human cells– alveolar lung cells in the upper channel and lung blood vessel cells in the lower channel– to recreate the interface in between human air sacs and their blood-transporting blood vessels. To imitate the conditions that alveoli experience in the human lung, the channel lined by alveolar cells was filled with air while the blood vessel channel was perfused with a flowing culture medium including nutrients that are typically delivered via the blood. RAGE is more highly revealed in the lung than in any other organ in the human body, and has actually been linked as a major inflammatory arbitrator in numerous lung diseases.
” This research study demonstrates the significance of breathing movements for human lung function, including immune responses to infection, and shows that our Human Alveolus Chip can be utilized to design these reactions in the deep portions of the lung, where infections are frequently more extreme and result in hospitalization and death,” said co-first author Haiqing Bai, Ph.D., a Wyss Technology Development Fellow at the Institute. “This design can also be utilized for preclinical drug testing to ensure that prospect drugs in fact minimize infection and inflammation in functional human lung tissue.” The outcomes are published today (April 8, 2022) in Nature Communications.
Producing a flu-on-a-chip
As the early phases of the COVID-19 pandemic made painfully clear, the lung is a vulnerable organ where inflammation in action to infection can generate a “cytokine storm” that can have deadly consequences. The lungs are also extremely intricate, and it is tough to replicate their distinct features in the lab. This complexity has actually impeded sciences understanding of how the lungs function at the cell and tissue levels, in both healthy and diseased states.
The Wyss Institutes Human Organ Chips were developed to resolve this problem, and have been revealed to faithfully duplicate the functions of numerous various human organs in the laboratory, including the lung. As part of jobs funded by the NIH and DARPA because 2017, Wyss researchers have actually been working on replicating numerous illness in Lung Airway and Alveolus Chips to study how lung tissues respond to breathing viruses that have pandemic potential and test prospective treatments.
In both the upper alveolar channel (top) and lower blood vessel channel (bottom) of the Alveolus Chips, the cells formed intact tissues held together by proteins in between the cells called ZO1 and VE-cadherin, which are stained green and red, respectively. Credit: Wyss Institute at Harvard University
Throughout his Ph.D. training, Bai studied diseases that impact the small air sacs deep inside the lungs where oxygen is quickly exchanged for carbon dioxide. That structure prepared him to tackle the difficulty of recreating a flu infection in an Alveolus Chip so that the group might study how these deep lung spaces install immune responses against viral intruders.
Bai and his group initially lined the 2 parallel microfluidic channels of an Organ Chip with various types of living human cells– alveolar lung cells in the upper channel and lung blood vessel cells in the lower channel– to recreate the interface between human air sacs and their blood-transporting blood vessels. To simulate the conditions that alveoli experience in the human lung, the channel lined by alveolar cells was filled with air while the capillary channel was perfused with a flowing culture medium containing nutrients that are typically delivered via the blood. The channels were separated by a porous membrane that permitted particles to flow in between them.
Previous studies at the Wyss Institute have developed that using cyclical stretching to Alveolus Chips to mimic breathing motions produces biological responses that imitate those observed in vivo. This is accomplished by applying suction to hollow side chambers nearby to the cell-lined fluidic channels to rhythmically stretch and unwind the lung tissues by 5%, which is what human lungs typically experience with every breath
When the team infected these “breathing” Alveolus Chips with H3N2 influenza by introducing the infection into the air channel, they observed the development of numerous known trademarks of influenza infection, including the breakdown of junctions in between cells, a 25% boost in cell death, and the initiation of cellular repair programs. Infection also led to much greater levels of numerous inflammatory cytokines in the blood vessel channel consisting of type III interferon (IFN-III), a natural defense against viral infection that is likewise activated in vivo flu infection studies.
In addition, the capillary cells of contaminated chips revealed greater levels of adhesion molecules, which enabled immune cells including B cells, T cells, and monocytes in the perfusion medium to connect to the capillary walls to help fight the infection. These outcomes verified that the Alveolus Chip was installing an immune action against H3N2 that recapitulated what happens in the lung of human clients contaminated with influenza virus.
Focus on your breath.
The team then carried out the same experiment without mechanical breathing motions. To their surprise, chips exposed to breathing movements had 50% less viral mRNA in their alveolar channels and a significant reduction in inflammatory cytokine levels compared to static chips. Genetic analysis exposed that the mechanical strain had actually activated molecular paths connected to immune defense and numerous antiviral genes, and these activations were reversed when the cyclical stretching was stopped.
Haiqing Bai– Co-first author, Haiqing Bai, Ph.D., brings his experience studying illness that impact the human lungs air sacs, or “alveoli” to his research on Organ Chips at the Wyss Institute. Credit: Wyss Institute at Harvard University
” This was our most unforeseen finding– that mechanical tensions alone can produce an innate immune action in the lung,” said co-first author Longlong Si, Ph.D., a previous Wyss Technology Development Fellow who is now a Professor at the Shenzhen Institute of Advanced Technology in China.
Understanding that often the lungs experience greater than 5% strain, such as in chronic obstructive lung disorder (COPD) or when clients are placed on mechanical ventilators, the researchers increased the stress to 10% to see what would take place. The higher strain caused a boost in inherent immune reaction genes and procedures, including a number of inflammatory cytokines.
” Because the greater strain level resulted in greater cytokine production, it might describe why patients with lung conditions like COPD suffer from persistent inflammation, and why clients who are placed on high-volume ventilators in some cases experience ventilator-induced lung injury,” Si discussed.
From a chip to medical trials
The researchers then went an action even more, comparing the RNA molecules present in cells within strained vs. fixed Alveolus Chips to see if they might identify how the breathing motions were producing an immune reaction. They determined a calcium-binding protein, called S100A7, that was not discovered in static chips but highly expressed in strained chips, recommending that its production was induced by mechanical stretching. They likewise found that increased expression of S100A7 upregulated numerous other genes involved in the inherent immune action, consisting of multiple inflammatory cytokines.
RAGE is more highly expressed in the lung than in any other organ in the human body, and has actually been linked as a significant inflammatory mediator in a number of lung diseases. The drug azeliragon is a recognized inhibitor of RAGE, so the scientists perfused azeliragon through the blood vessel channel of strained Alveolus Chips for 48 hours, then contaminated the chips with H3N2 infection.
Based on this promising outcome, the group then infected stretched Alveolus Chips with H3N2 and administered azeliragon at its healing dose 2 hours after infection. This technique substantially blocked the production of inflammatory cytokines– a result that was further improved when they added the antiviral drug molnupiravir (which was recently authorized for clients with COVID-19) to the treatment regimen.
These outcomes stood out of Cantex Pharmaceuticals, which owns patent rights to azeliragon and had an interest in using it to deal with inflammatory illness. Based in part on the Wyss groups operate in Alveolus Chips, Cantex accredited azeliragon for the treatment of COVID-19 and other inflammatory lung illness in early 2022. Given the drugs exceptional safety record in previous Phase 3 scientific trials, the company has made an application for FDA approval to start a Phase 2 trial in patients with COVID-19 patients, and plans to follow with additional Phase 2 trials for other illness including COPD and steroid-resistant asthma.
” Thanks to the fantastic work of the researchers at the Wyss Institute, we now have compelling evidence that azeliragon may have the prospective to prevent severe COVID-19 health problem in the kind of a once-a-day pill. Were delighted to have the chance to conduct medical trials of azeliragon for this illness, looking for to bring this innovative therapy to patients to prevent the life-threatening inflammation that is a major reason for hospitalization and death,” said Stephen Marcus, M.D., CEO of Cantex.
While azeliragon is an appealing anti-inflammatory drug, the researchers alert that more research studies are needed to figure out a reliable and safe treatment program in humans. RAGE is a vital player in initiating helpful inflammation versus pathogens in the early stages of an infection, and preventing it too soon might prevent a patient from installing an enough immune reaction.
Given the Alveolus Chips lots of advantages over conventional preclinical designs, the Wyss group is exploring the incorporation of extra cell types such as macrophages into the chips to increase their intricacy and design more biological procedures, such as adaptive immunity. They are also utilizing their existing design to study the effectiveness of new compounds, drugs, and biologics (such as mRNA rehabs) against influenza, SARS-CoV-2, and other illness.
” This important paper resulted in the discovery of RAGE inhibitors pledge for treating inflammatory lung illness, which was the basis for the recent license of azeliragon to Cantex and its motion towards human clinical trials for COVID-19. I am very happy of this group and how quickly this clinical finding was translated into commercialization that will hopefully result in lifesaving treatment for patients. This is what the Wyss Institute is all about,” said senior author Donald Ingber, M.D., Ph.D., who is the Wyss Institutes Founding Director along with the Judah Folkman Professor of Vascular Biology at Harvard Medical School (HMS) and Boston Childrens Hospital, and Hansjörg Wyss Professor of Bioinspired Engineering at the Harvard John A. Paulson School of Engineering and Applied Sciences.
Recommendation: “Mechanical control of natural immune reactions versus viral infection exposed in a human Lung Alveolus Chip” by Haiqing Bai, Longlong Si, Amanda Jiang, Chaitra Belgur, Yunhao Zhai, Roberto Plebani, Crystal Yuri Oh, Melissa Rodas, Aditya Patil, Atiq Nurani, Sarah E. Gilpin, Rani K. Powers, Girija Goyal, Rachelle Prantil-Baun & & Donald E. Ingber, 8 April 2022, Nature Communications.DOI: 10.1038/ s41467-022-29562-4.
Additional authors of the study include Amanda Jiang, Chaitra Belgur, M.S., Yunhao Zhai, Ph.D., Melissa Rodas, and Aditya Patil and Girija Goyal, Ph.D. from the Wyss Institute, and previous Wyss Institute members Roberto Plebani, Ph.D., Crystal Oh, Atiq Nurani, M.S., Sarah Gilpin, Ph.D., Rani Powers, Ph.D. and Rachelle Prantil-Baun, Ph.D
. This research study was supported by the Wyss Institute for Biologically Inspired Engineering at Harvard University, the United States Defense Advanced Research Projects Agency (DARPA) under Cooperative Agreement HR0011-20-2-0040, and the National Institutes of Health under grants UG3-HL-141797 and UH3-HL-141797.
The Human Alveolus Chip contains hollow side channels that enable suction to be used to the chips, applying cyclic pressure that imitates the movements of normal human breathing (left). A permeable membrane separates human alevolus cells in the upper channel from human blood vessel cells in the lower channel, allowing them to exchange molecular signals (right). Credit: Wyss Institute at Harvard University
Human Lung Chip exposes the impacts of breathing movements on lung immune responses and causes repurposing of prospective therapeutics for breathing diseases, consisting of COVID-19.
Every breath extends the lungs tissues with each inhale and relaxes them with each exhale. The mere movements of breathing are understood to influence important functions of the lungs, including their development in children, the production of air-exchange-enhancing fluid on their inner surfaces, and maintenance of healthy tissue structure.
These immunofluorescence micrographs (at different zooms) reveal the 3D cellular structure that establishes within the alveolar channel and mimics the microstructure of human alveoli. Credit: Wyss Institute at Harvard University
Using a Human Lung Chip that duplicates the structures and functions of the lung air sac, or “alveolus,” the research study team found that using mechanical forces that imitate breathing motions reduces influenza virus replication by triggering protective natural immune actions. They likewise recognized numerous drugs that decreased the production of inflammatory cytokines in infected Alveolus Chips, which could be beneficial in dealing with excessive inflammation in the lung. Based upon these research studies, among those drugs was licensed to Cantex Pharmaceuticals for the treatment of COVID-19 and other inflammatory lung illness. Data from the research were just recently included in the companys Investigational New Drug (IND) application to the FDA to start a Phase 2 medical trial for COVID-19.