Duke University | Howard Hughes Undergraduate Program

 "Enzymes are things invented by biologists that explain things which otherwise require harder thinking." – Jerry Lettvin

Everyday I slice and dice tissue, use innumerable pipette tips, apply chemicals to slides, and spend hours on amazingly expensive lab equipment. Inevitably, what I do costs money – money that is largely provided by your wallet! Like many biomedical research laboratories, we here at the Stripp Lab receive a lot of our funding from government agencies like the National Institute of Health. Those departments are financially supported by your gracious yet compulsory donations as taxpayers. So, thank you! Whether you’ve realized it or not, you play a huge role in keeping our research afloat, and you deserve to know what we’re doing. You wouldn’t just hand someone your money without knowing what that money’s going to be used for, especially during times of economic crisis. Let me enlighten you with what you are allowing me to do during my time as a Howard Hughes Research Fellow.

It’s quite likely that you or someone you know has or had some sort of lung disease, whether that’s asthma, emphysema, or a kind of chronic obstructive pulmonary disease. If you do, you know how debilitating and life-altering these conditions can be. A person with lung disease may need to be provided with an inhaler, home oxygen therapy, a cocktail of prescription medication, and receive periodic vaccinations. Some of these diseases can be genetically influenced, triggered by pollutants in the air you breath, or caused by smoking.

Lung disease can have devastating effects on the anatomy of the lung. Once injured, the epithelial cells that line bronchioles begin to die, leaving the airway open to infection. The cells in the terminal bronchioles near the alveoli, where gas exchange occurs, can also be destroyed by lung disease; this can make oxygen intake and carbon dioxide release difficult, resulting in struggled breathing among other serious problems.

What if these damaged airways, compromised by cell death, could be restored? That is one of the long-term aims of Dr. Stripp’s research. One solution may come in the form of stem cells.

Stem cells have received a lot of attention in the news media and scientific community over the last few decades. Stem cells are those cells that have not been assigned a specific duty in the cell (e.g. red blood cells have a job; their job is to transport oxygen to your tissues), can self-replicate, and can give rise to differentiated cells (specialized, often localized cells with a “job”). There are basically two different categories of stem cells: adult (those found in deve loped systems like bone marrow, intestine, lungs) and embryonic (those found in the inner cell mass of an early stage embryo). The latter is quite controversial, because many scientists hold that they have a greater ability to differentiate into a larger number of different cell types (pluripotent) compared to an adult stem cell (multipotent) and some religious and ethical groups do not approve of killing an embryo to harvest its stem cells. Regardless, if a stem cell can create other types of cells, why couldn’t one be made to regenerate lung tissue? That is one idea we are exploring.

Lung stem cells are also quite controversial in the biology world. Stem cells are often very active, replenishing depleted supplies of cells at great rates. Lung stem cells are different, because, in the adult, they are not extremely active. They are most active during the embryonic development of the lung, and then they begin to quiet as the necessity to build lung tissue decreases. Other less stem cell-like cells are able to upkeep the lung in the adult; these Clara cells protect the bronchiolar epithelium by secreting Clara cell secretory protein (CCSP) and a component of the lung surfactant and by detoxifying harmful inhaled substances. cells also multiply and differentiate into the ciliated cells that line the epithelium.

Okay, so Clara cells can regenerate the bronchiolar epithelium, but what happens when Clara cells are wiped out by pollution? This question was recently addressed, and it turns out that there exists a population of rare pollution-resistant Clara cells (vCE or variant CCSP-expressing) that inhabit specific locations of the lung. vCE cells can replenish the depleted Clara and ciliated cell populations, ablated by pollutants like naphthalene and ozone. However, if the lung injury sustained is severe enough, some of the vCE cells can be wiped out. If enough are ablated, the lung will be compromised.

What if more vCE cells could be present and assist in epithelial restoration of Clara and ciliated cells? That is what my research focuses on. By increasing the concentration of a protein called beta-catenin in adult mice, I may be able to reprogram Clara cells to the vCE status. An accumulation of a particular form of beta-catenin has been shown to activate the transcription factors that are normally turned on during embryonic development, when the population of vCE cells is relatively high.

I spend most of my time in the Stripp Lab preparing mice tissue, staining it for proteins like CCSP and beta-catenin forms, and analyzing the immunofluorescence under a microscope.

Take a look at one of the cool photos that I have taken of one of my samples!

This is an image of the conducting airway epithelium of a mouse bronchiole. The blue staining shows where cell nuclei are located. The red/orange between and around the epithelial cells shows one form of beta-catenin. The green shows the form of beta-catenin we have tried to genetically introduce in our transgenic mice. What are important to notice here are those epithelial cells that show only green, indicating an increase in the beta-catenin that has been shown to induce stem cell (vCE in this case) reprogramming.

Remarkably, this one image has taken months to cultivate. I only took over when the lung tissue was embedded in wax. From that point on, I needed to spend about a week cutting the wax, putting the samples on microscope slides, staining with immunofluorescent antibodies, and analyzing with the microscope. Before all that work, the experiment needed to be set up, the mice injected with certain chemicals, their progeny screened, and the lungs dissected, fixed, and embedded in wax. All the work and number of people involved in the generation of just one image like the one above are testaments to the cooperation, patience, and ingenuity required in research.

I know that this information is a lot to take in! It was certainly difficult for me to wrap my head around the details during my first few days in the lab, but stay tuned to my blog updates and everything will start to become clearer!

- Tyler