Healing Damaged Lungs with Stem Cells

Emphysema, bronchitis, asthma and cystic fibrosis are all diseases of the airways and they are the second leading cause of death worldwide. Over 35 million Americans alone suffer from chronic respiratory disease.

Scientists from the Weizmann Institute have now proposed a new direction for treating these diseases that might lead to a new method for alleviating some of their suffering. The study’s findings, which were published in Nature Medicine, show that it might be possible to use fetal stem cells to repair damaged lung tissue.

Particular stem cells normally found in the lungs are highly similar to those in the bone marrow. Organ-specific stem cells tend to be concentrated in special compartments rather than being distributed throughout the tissue. This insight prompted Prof. Yair Reisner of the Weizmann Institute’s Immunology Department to suggest: “That understanding suggested to us that we might be able to apply our knowledge of techniques for transplanting bone marrow stem cells to repairing lung tissue.”

Bone marrow transplants are based on two main principles: the ability of stem cells to navigate through the blood to the appropriate compartment and the prior clearing out of the compartment to make room for the transplanted stem cells. Dr. Reisner and his group thought it might possible to apply these principles to introducing new stem cells into the lungs. However, before they could do this, they needed to find a source of lung stem cells suitable for transplanting.

Reisner and his co-workers used fetal lung tissue from mice and humans (20–22 weeks of gestation for humans, and embryonic day 15–16 or E15–E16 for mice). Cells from these stages have differentiated into lung progenitor cells and are fully capable of lung regeneration. Reisner and his colleagues conducted a series of experiments in which they cleared the lung’s stem cell compartments with a new method developed on their own laboratory, and then were injected these new lung progenitor cells into mouse models of lung damage. The fetal lung stem cells found their way through the blood to the lungs and settled into the proper compartment. By six weeks, these cells were well on their way to differentiating into normal lung tissue. In these mice, their damaged lungs healed, and their breathing improved significantly.

Next, Reisner intends to determine the correct dosage of drugs that are needed to prevent rejection of the transplanted cells, which will be needed following such procedures. “But our real vision, bolstered by this success,” says Reisner, “is to create a bank of lung tissue that will be a resource for embryonic lung stem cells.” This bank could mean that there is a ready source of cells for repairing the damage in those with severe respiratory disease.

Reisner’s work shows that fetal lung progenitors can repopulate lungs and heal them. If Reisner can find a way to generate early lung progenitors from pluripotent stem cells, then such cells can be used to heal damaged lungs.

One Type of Lung Cell Can Regenerate Another

A collaboration between the Perelman School of Medicine at the University of Pennsylvania and Duke University has found that lung tissue has a much great ability to regenerate than previously thought.

Lungs contain thousands of tiny clusters of sacs called alveoli. Gas-exchange between the air and our blood stream occurs across the thin lining of the alveoli, which are lined with extensive networks for diminutive blood vessels called capillaries. The cells that form the paper-thin lining of the alveoli are called type 1 cells. Within the alveoli are cells called type 2 cells, which secrete surfactant; a soapy substance that prevents the alveoli from collapsing upon themselves when we exhale. Some premature babies do not make enough surfactant and must be treated with surfactant to help them breathe.

Work in mice demonstrated that both type 1 and type 2 cells descend from a common embryonic precursor during lung development. When mice had bits of their lungs removed, labeling studies established that the newly re-established type 2 cells were made from type 1 cells and that some of the newly made type 1 cells were formed from type 2 cells. These results were confirmed by cell culture experiments that grew single type 1 or type 2 lung cells in culture; in both cases, the cultures gave rise to mixed cultures consisting of both type 1 and type 2 lung cells. These data demonstrate that type 1 lung cells can give rise to type 2 lung cells and visa versa.

Previously, the Duke University term had demonstrated that type 2 lung cells in mice not only produce surfactant, but also function as progenitors for other lung cells in adult mice. This shows that type 2 lung cells can definitely differentiate into type 1 lung cells. However, there was no evidence that type 1 lung cells could give rise to other types of lung cells.

In this present work, however, lung injury in mice stimulated the type 1 cells to divide and differentiate into type 2 cells over a period of three weeks while the lung regenerated. According to Jonathan Epstein from the University of Pennsylvania, It’s as if the lung cells can regenerate from one another as needed to repair missing tissue, suggesting that there is much more flexibility in the system than we have previously appreciated. These aren’t classic stem cells that we see regenerating the lung. They are mature lung cells that awaken in response to injury. We want to learn how the lung regenerates so that we can stimulate this process in situations where it is insufficient, such as in patients with COPD (chronic obstructive pulmonary disease).”

This is one of the first studies to demonstrate that mature cells that were thought to be completely at the end of their growth and differentiation capabilities can revert to an earlier state under the right conditions without the use of transcription factors, but by responding to damage.

These two research teams are also applying the approaches outlined in this publication to cells from other tissues, such as the intestine and skin, in order to study the mechanisms of cell maintenance and differentiation, and then relate these same mechanisms back to the heart. They also hope to apply these findings in clinical settings for patients who suffer from idiopathic pulmonary fibrosis, acute respiratory distress syndrome and other such conditions where the alveoli cannot supply sufficient amounts of oxygen to the blood.

Scientists Use Stem Cells to Grow Three-Dimensional Mini Lungs.

In research done in several laboratories, lung tissue was derived from flat cell culture systems or by growing cells on scaffolds made from donated organs.

Now in a new study published in the online journal eLife, a multi-institution team has defined a culture system for generating self-organizing human lung organoids, which are three-dimensional structures that mimic the structure and complexity of human lungs.

“These mini lungs can mimic the responses of real tissues and will be a good model to study how organs form, change with disease, and how they might respond to new drugs,” said study senior author Jason R. Spence, Ph.D., an assistant professor of internal medicine and cell and developmental biology at the University of Michigan Medical School.

Spence and his colleagues successfully grew structures that resembled both the large airways or bronchi and small lung sacs, known as alveoli.

These mini lung structures were developed in a cell culture system. Therefore, they lack several components of the human lung, including blood vessels, which are a critical component of gas exchange during breathing.

Despite that, these cultured organoids can serve as a unique research model system for researchers as they grind out basic science ideas that are turned into clinical innovations. These three-dimensional mini-lungs should be an excellent complement to research in liver laboratory animals.

Traditionally, the behavior of cells has been investigated in the laboratory in two-dimensional culture systems where cells are grown in thin layers on cell-culture dishes. Most cells in the body, however, exist in a three-dimensional environment as part of complex tissues and organs. Tissue engineered have been trying to re-create these environments in the laboratory by successfully generating small version of particular organs known as organoids, which serve as models of the stomach, brain, liver and human intestine. The advantage of growing three-dimensional structures of lung tissue, according to Dr. Spence, is that the organization of organoids bears greater similarity to the human lung.

To make these lung organoids, researchers at the U-M’s Spence Lab and colleagues from the University of California, San Francisco; Cincinnati Children’s Hospital Medical Center; Seattle Children’s Hospital and University of Washington, Seattle manipulated several of the cell signaling pathways that control the formation of organs.

First, stem cells were induced to form a type of tissue called endoderm, which is found in early embryos and gives rise to the lung, liver and several other internal organs. Second, the group activated two important development pathways (FGF and WNT signaling ) that are stimulate endoderm to form three-dimensional tissue. By inhibiting two other key development pathways at the same time (BMP and TGFβ signaling), the endoderm became tissue that resembles the early lung found in embryos.

In the laboratory, this early culture-derived lung-like tissue spontaneously formed three-dimensional spherical structures as it developed. Afterwards, they had to expand these structures and develop them into lung tissue. In order to do this, Spence and his colleagues and collaborators exposed the cells to additional proteins involved in lung development (FGF and Hedgehog).

After all this manipulation, the resulting lung organoids survived in the laboratory for over 100 days.

“We expected different cells types to form, but their organization into structures resembling human airways was a very exciting result,” said author Briana Dye, a graduate student in the U-M Department of Cell and Developmental Biology.

While this type of experiment is remarkable, this is only the beginning of lung tissue engineering.  These mini-lungs  will hopefully serve and new model systems for drug testing and researching genetic diseases that affect the lungs, such as cystic fibrosis, sarcoidosis, or inherited forms of emphysema.  It will be a while before scientists can make replacement lungs for human patients, but these experiments by Spence and others are a remarkable start.

Rare Stem Cell Heals Damaged Lungs; Notch Signaling May Hold the Key to Lung Fibrosis

Patients who survive an acute lung injury are able to recover their lung function, which suggests that adult lungs regenerate to a certain extent. Depending on the cause and severity of the injury, multiple progenitor cells, including alveolar type II cells and distal airway stem cells, have been shown to drive lung tissue regeneration in mice. Now, Andrew Vaughan and others have described another cell type in the lungs involved in the repair process in mice when mouse lungs are damaged from influenza virus infection or inhalation of the anticancer drug bleomycin.  This cell type is called the rare lineage-negative epithelial progenitor (LNEP).

LNEP cells are quiescently present within normal distal mouse lung and do not express mature lineage markers (for example, a protein called club cell 10 or CC10 or surfactant protein C, otherwise known as SPC).  However, Vaughan and others demonstrate that LNEPs are activated to proliferate and migrate to damaged sites and mediate lung remodeling following major injury.

Vaughan and others used lineage tracing approaches and cell transplantation strategies and showed that LNEP cells, but not mature epithelial lineage cells, are multipotent in their ability to give rise to both club cells and alveolar cells.  Interestingly, activation of the Notch signaling pathway in LNEP cells initially activated them, but persistent Notch activation inhibited subsequent alveolar differentiation, resulting in failed tissue regeneration (characterized by the formation of abnormal honeycomb cysts in the mouse lung).  Thus Notch signaling is only required at the beginning of their activation, and then must be down-regulated if the LNEP cells are to reconstruct normal lung tissue.  Interestingly, scarred over or fibrotic lungs from patients with idiopathic pulmonary fibrosis or a disease called scleroderma show evidence of hyperactive Notch signaling and their lungs also contain very similar-looking honeycomb cysts.  This strongly suggests that dynamic Notch signaling also regulates the function and differentiation of LNEP-analogous human lung progenitor cells.  Thus designing treatments that properly regulate Notch signaling and, consequently, LNEP activity may potentially halt the development of lung fibrosis in humans.

Lung Stem Cells Heal Lungs and Point to Possible New Treatments

Frank McKeon, Ph.D., and Wa Xian, Ph.D. from Jackson Laboratory and their colleagues have identified the a certain lung stem cell, and the role it plays in regenerating lungs.

This work, which appeared in the Nov. 12 issue of the journal Nature, provides some much-needed clarification of the nuts and bolts of lung regeneration and provides a way forward for possible therapeutic strategies that harness these lung stem cells.

“The idea that the lung can regenerate has been slow to take hold in the biomedical research community,” McKeon says, “in part because of the steady decline that is seen in patients with severe lung diseases like chronic obstructive pulmonary disease (known as COPD) and pulmonary fibrosis.”

McKeon noted that there is ample evidence of a robust system for lung regeneration. “Some survivors of acute respiratory distress syndrome, or ARDS, for example, are able to recover near-normal lung function following significant destruction of lung tissue.”

This is a capacity that humans share with mice. Mice infected with the H1N1 influenza virus show progressive inflammation in the lung followed by the death and loss of important lung cell types. However, over the course of several weeks, the lungs of these mice recover and show no signs of previous lung injury.

Because of the presence of such robust lung regeneration in mice, these organisms provide a fine model system to study lung regeneration.

McKeon and his colleagues had previously identified a type of adult lung stem cell known as p63+/Krt5+ in the distal airways. When grown in culture, these p63+/Krt5+ lung stem cells neatly formed alveolar-like structures that were similar to those found within the lung. Alveoli are the tiny, specialized air sacs that form at the ends of the smallest airways, where gas exchange occurs in the lung. Following infection with H1N1, these same stem cells migrated to sites of inflammation in the lung and clustered together to form pod-like structures that resemble alveoli, both visually and molecularly.

McKeon and his colleagues reported that when the lung is damaged by H1N1 infraction, p63+/Krt5+ lung stem cells proliferate and contribute to the development of new alveoli near sites of lung inflammation.

To determine if these cells are required for lung regeneration, McKeon and his coworkers developed a novel system that utilizes genetic tools to selectively remove these cells from the mouse lung. Mice that lack p63+/Krt5+ lung stem cells cannot recover normally from H1N1 infection, and instead exhibit scarring of the lung and impaired oxygen exchange. This demonstrates the key role p63+/Krt5+ lung stem cells play in regenerating lung tissue.

To carry this work one step further, McKeon and his team isolated and subsequently transplanted p63+/Krt5+ lung stem cells into a damaged lung. The transplanted p63+/Krt5+ cells readily contribute to the formation of new alveoli, which nicely illustrates the capacity of these cells to regenerate damaged lung tissue.

In the U.S. about 200,000 people have Acute Respiratory Distress Syndrome, a disease with a death rate of 40 percent, and there are 12 million patients with COPD. “These patients have few therapeutic options today,” Xian says. “We hope that our research could lead to new ways to help them.”

MSCs for Tissue Engineered Tracheas and Enhanced Fracture Healing

For all my readers who have ever broken a bone, this one’s for you.

Setting a broken bone properly can lead to the healing of a broken bone, but large fractures that generate gaps in bones are very hard to heal. Stem cell therapy in combination with small protein molecules called cytokines has the potential to improve bone repair, since cytokines summon resident stem cells to migrate and home to the injured site. Having said that, the engraftment, participation and recruitment of other cells within the regenerating tissue are equally important.

To stimulate stem cell-mediated healing, University College London scientists over-expressed the SDF-1 protein in mesenchymal stem cells. Since SDF-1 is a stem cell-recruitment protein, it seems reasonable to suspect that these engineered cells would effectively increase the migration of native cells to the site of fracture and enhance bone repair.

Once they made SDF-1-expressing mesenchymal stem cells, Chih-Yuan Ho and colleagues showed that these cells increased the migration of non-transfected cells in a cell culture system.

Once these SDF-1-expressing mesenchymal stem cells were implanted into rats with large bone defects, bone marrow mesenchymal stem cells that over-expressed SDF-1 showed significantly more new bone formation within the gap and less bone mineral loss at the areas next to the defect site during the early bone healing stage.

Thus, SDF-1 plays an important role in accelerating fracture repair and contributing to bone repair, at least in this rat model. SDF-1 does this by recruiting more host stem cells to the defect site and encouraging their differentiation into bone cells, which go on to produce good-quality bone.  This paper appeared the the journal Tissue Engineering, Part A.

In a second paper that appeared in the Annals of Biomedical Engineering, mesenchymal stem cells were used to tissue engineer tracheae. In this case a biocompatible scaffold was seeded various with various cells and this strategy could be a solution for tracheal reconstruction.

Yoo Seob Shin and colleagues seeded mesenchymal stem cells (MSCs) on a scaffold made from pig cartilage powder (PCP). The PCP was made with minced and decellularized pig joint cartilage and was molded into a 5 × 12 mm (height × diameter) scaffold. Mesenchymal stem cells from the bone marrow of young rabbits were grown in culture and then cultured with the PCP scaffold. After 7 weeks in culture, these tracheal implants were transplanted on a 5 × 10 mm tracheal defect in six rabbits, which were evaluated 6 and 10 weeks after the operation.

None of the six rabbits showed any sign of respiratory distress, and endoscopic examination of these tissue engineered tracheae showed that the a normal-looking respiratory epithelium completely covered the regenerated trachea. These trachea also displayed no signs of collapse or blockage.

The tissue engineered tracheae were also scanned and modeled on a computer model (luminal contour). The reconstructed areas of the trachea were the right width and dimensions compared to normal adjacent trachea and were not narrow.

Detailed microscopic tissue examinations of the tissue engineered tracheae showed that the new cartilage was successfully produced by the seeded mesenchymal stem cells and there was only a minimal degree of inflammation or granulation tissue that forms on the surfaces of wounds during the healing process. This shows that the implants did not trigger a massive inflammatory response that damaged resident or implanted tissue.

The outer surfaces of tracheal cells are decorated with tiny beating hairs called cilia that constantly beat to clear particles from the respiratory system. There are also cells that secrete mucus, which acts like fly paper for invading pollutants, particles or microorganisms. in the tissue engineered tracheae, ciliary beating frequency of the regenerated epithelium was not significantly different from the normal adjacent mucosa.

Thus, mesenchymal stem cells from bone marrow seeded on a PCP scaffold successfully restored not only the shape but also the function of the trachea without any signs of graft rejection.

Bones and trachea – mesenchymal stem cells pack a powerful healing punch!!

FDA Approves Pneumostem Clinical Trial for Bronchopulmonary Dysplasia

MEDIPOST America Inc. has announced that the US Food and Drug Administration (USFDA) has approved their product Pneumostem for a Phase 1/2 clinical trial. This Phase 1/2 trial will assess the safety and efficacy of Pneumostem on prematurely born infants who are at high-risk of developing Bronchopulmonary Dysplasia.

Bronchopulmonary dysplasia (BPD) is a serious lung condition that affects infants. BPD usually affects premature infants who need oxygen given through nasal prongs, a mask, or a breathing tube in order to properly breathe.

Most infants who develop BPD are born more than 10 weeks before their due dates and weigh less than 2 pounds (about 1,000 grams) at birth, and have breathing problems. Respiratory infections that hit before or shortly after birth also can contribute to BPD.

Some infants who suffer from BPD may need long-term breathing support from breathing (NCPAP) machines or ventilators. BPD is the leading cause of mortality and severe complications in premature infants. Currently there is no approved therapies or drugs exist for BPD. This pneumostem trial is expected to draw global attention in the field of neonatal medicine, since it would provide a potential treatment for BPD where none presently exists.

Pneumostem is an off-the-shelf product made from human Umbilical Cord Blood-derived Mesenchymal Stem Cells (hUCB-MSCs). hUCB-MSCs show a terrific ability to grow in the laboratory and can also differentiate into multiple types of cells or tissues. They are immune-privileged and thus if they are used in patients other than from whom they are isolated, they do not cause adverse immune reactions. hUCB-MSCs harvested from cord blood show the lowest levels of immunogenicity compared to those by other types of adult stem cells. Thus, instead of provoking immunogenicity, they rather modulate the adverse immune reactions within the host, which makes hUCB-MSCs an ideal candidates for mass-producible stem cell drug for allogeneic use. These cells seem to facilitate regeneration of lung tissue and suppress the inflammatory responses in the lungs of premature infants.

Pneumostem has received Orphan Drug designation in Korea by the Ministry of Food and Drug Safety (MFDS) and the Korean Phase 2 study is 80% complete. The US FDA also granted Orphan Drug designation for Pneumostem demonstrating its medical value and commercial potential.

Presently, MEDIPOST America is rapidly moving begin this Pneumostem trial in the U.S. At the same time, Medipost will continue its licensing and technology transfer negotiations with multinational pharmaceutical companies.

The approval of this Pneumostem clinical trial by the US FDA, whose regulation of medicinal products is very strict (including stem cell products), might boost clinical trial approvals in other European and Asian countries.

Clinical development of Pneumostem was partly supported by Translational Stem Cell & Regenerative Medicine Consortium grant as a part of Public Health and Medicinal Technology R&D Project funded by the Korea Ministry of Health & Welfare and the Korea Health Industry Development Institute.