By isolating the cellular source of the regenerative signals, which are produced by blood vessels in the lungs, the scientists were able to trigger regeneration in mice that had had one lung surgically removed.
The lungs of an average person contain about 700 million alveoli, each one wrapped in a mesh of capillaries. Smoking, infection, toxic chemicals, and other insults can damage or destroy alveoli. Millions of people are short of breath or seriously disabled by chronic bronchitis, emphysema, interstitial fibrosis, cancer, and other diseases that damage the lungs. There are no definitive treatments, and Rafii said that supportive care costs the United States some $150 billion a year.
“We have shown that the blood vessels are not just passive conduits for conveying nutrients and oxygen.”
Patients who have a lung surgically removed due to cancer or respiratory diseases have been known to regain some of the lost lung function, presumably as a result of regeneration. In the laboratory, scientists have found that mice can regenerate alveolar lung tissue under certain conditions. But Rafii’s report in the October 28, 2011, issue of the journal Cell, in which he and postdoctoral fellow Bi-Sen Ding at Weill Cornell Medical College identified key molecular signals that jump-start the formation of new alveoli, provides the first detailed mechanism to explain the process.
Rafii said it may eventually be possible to trigger repair of damaged lung tissues in human patients by activating the healthy endothelial cells that line their alveolar capillaries. Another strategy would be to transplant vascular cells from the lungs of genetically-matched donors as a source of regeneration-stimulating factors.
Moreover, the scientists demonstrated they could switch on the regenerative process in mice whose left lungs had been surgically removed. Regeneration was associated with an increase in the number of alveoli of the intact right lungs and improved respiratory function, restoring the animals’ breathing capacity almost to normal.
The spark that sets the process in motion, the scientists said, is an activated enzyme, MMP14, secreted by lung-specific endothelial cells that line the fine capillaries of the air sacs. MMP14 increases the availability of epidermal growth factors to stimulate alveolar regeneration. These specialized vascular cells, called pulmonary capillary endothelial cells (PCECs), are in direct cellular contact with alveolar epithelial cells. If opened out flat, the PCECs encompassing alveoli in a healthy human would cover an area roughly equal to that of a tennis court.
“We have shown that the blood vessels are not just passive conduits for conveying nutrients and oxygen,” Rafii said. “When you remove the left lung, the PCECs get stimulated in the remaining right lung, producing angiocrine growth factors. These angiocrine factors cause epithelial cells positioned in the vicinity of PCECs to regenerate new alveolar sacs. This is a transformative finding.”
It hasn’t been determined precisely how removal of one lung leads to regeneration in human patients, but the researchers speculate that the abrupt increase in blood flow and biomechanical forces in the remaining lung could be the trigger for the PCECs. The Rafii laboratory, which has previously characterized specialized endothelial cells in bone marrow and liver that can induce regeneration, designed the current experiments to search for equivalent vascular cells and signals in the lung.
To begin to investigate the specific signals that trigger lung regrowth, Rafii recruited Ding, who was skilled at the challenging task of safely removing the left lung from mice, a surgical procedure called a pneumonectomy. Performing the procedure in mice allows scientists to model the process and look for the mechanisms that drive regeneration.
Using gene-knockout methods and screening models, the scientists showed that lung regeneration begins with the activation of the angiocrine factor MMP14 enzyme, which is generated exclusively by PCEC cells. Activated MMP14 liberates growth-stimulating molecules, including EGFs from the extra-cellular matrix components in which they are normally trapped. Once released, those growth factors induce alveolar epithelial cells to proliferate and form new alveolar sacs.
“MMP14 is probably the tip of the iceberg,” said Rafii. “Activated endothelial cells produce many other angiocrine factors to promote lung and organ regeneration, as well as orchestrating tumor growth.”
In fact, the process can be modeled in a laboratory dish. Rafii said that when PCEC cells are co-cultured with epithelial cells (they form the thin inside lining of the alveoli), the latter respond by expanding and forming minuscule “angiospheres” – three-dimensional units closely resembling the alveoli sac of the lung.
When the researchers transplanted activated PCECs from pneumonectomized mice into mice that lacked the angiocrine growth factors that they had found were needed for lung regeneration, the recipient mice regained the ability to regenerate additional lung tissue, and their normal lung function was restored.
Rafii is optimistic that his team’s findings may help researchers develop therapeutic strategies to trigger alveolar regeneration in patients who suffer from debilitating pulmonary disorders.