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Stefan Heller, Ph.D., and his team grew cells that not only look but also behave like mouse inner-ear hair cells.Their success points to exciting developments for managing hearing loss and beyond.
Stefan Heller, Ph.D., is a professor of otolaryngology and a professor of molecular and cellular biology at the Stanford University School of Medicine.He is also the principle investigator of Heller Lab in Stanford's Department of Otolaryngology. Roughly a decade ago, Heller proposed that stem cells could be used to create the specialized inner-ear hair cells that are critical for our ability to hear. Unlike birds, fish, and amphibians,mammals cannot regenerate hair cells, so once they are lost or damaged, the hearing loss is permanent. Last May Heller reported in the journal Cell that he and his team had successfully created mouse cells that not only resemble but also behave like mouse inner-ear hair cells. In an interview with Hearing Health, Heller reviewed the advances his lab has made since that significant study.
why mouse cells?
We explored mouse embryonic stem cells mainly because they are much easier to grow than human embryonic stem cells. Other labs have established protocols describing how to coax mouse embryonic stem cells into neurons and other cell types, including photo receptors of the eye, so we thought mouse embryonic stem cells would be a good starting point for figuring out ways to make inner ear cell types, first from mouse cells and in the future from human stem cells.
In fact, in 2003 we published a protocol of how to create these cells, but it was very inefficient and highly time consuming. We realized we needed to make the cells more efficiently but we also wanted to make sure they functioned as hair cells. They should really look like hair cells and work like hair cells.
It only took us seven years. I'm being a little sarcastic because we thought it would take perhaps a year. This illustrates that it wasn't an easy thing to do.
how did you create the mouse inner-ear hair cells?
There were people in the field who have been working on inner-ear development. People may wonder why the study of very basic inner-ear development is important for the development of therapies.I think it is very important because we learned what developmental biologists had discovered about how the ear is formed in the embryo, and particularly which growth factors and signaling pathways are involved. We applied that knowledge to guide embryonic stem cells in the test tube.
so you used as a blueprint what was discovered about inner-ear cell development?
Yes, we mimicked nature. In 2003 we didn't know how nature worked. But in the last five or 10 years there were pretty significant findings made by our colleagues about inner-ear cell development and we were able to apply that knowledge. Although unfortunately we are not further with the development of therapies, we have learned a lot. This is important because it helps to translate our findings toward human cells.
what are the next steps for your research?
There are two directions. One direction is to move everything to human embryonic stem cells. We are currently doing this, and we've made pretty good progress in the last two years. I'm quite optimistic that we can reach the same stage with human cells as we have with mouse cells in the near future.There may be another roadblock but I hope what took us seven years with mice may take us only four years with humans.
I believe this direction is a very important one because you can apply this technology to make inner-ear hair cell types from another cell called the iPS cell, the induced pluripotent stem cell.We plan to use iPS cells to generate inner-ear cells from patients who have a specific type of hearing loss.
You can generate iPS cells from a simple skin biopsy from patients. They are ethically perfectly fine there is no discussion about these types of cells as there is with embryonic stem cells.
They are powerful new tools because,as with embryonic stem cells, you can turn them into any cell type in the body.
Let's assume we have a human patient who has defective hair cells or who has lost hair cells because of a genetic defect. We would be able to convert skin cells from this patient into iPS cells, turn them into hair cells again, and perhaps repair the genetic defector look for drugs that suppress the genetic defect. There is a huge potential for new discoveries and very likely also for personalized medicine.
can you give an example of this kind of medicine using ips cells?
There was a paper in Nature recently by a Stanford researcher who generated iPS cells from a patient who has a mutation in a specific gene, causing heart defects. He was able to use the heart cells generated from this patient to find drugs that were able to repair or to reduce the bad effect of the faulty gene. In effect he discovered the cure for that heart defect. This is really powerful stuff.
In the hearing field I think we are far off with this kind of specialized cure because heart cells are very easy to make from iPS cells. Pluripotent cells like stem cells have a default pathway,the direction they will go in even if you don't do anything to them. If you don't take care of them they will eventually either convert into neurons orinto heart cells.
To make an ear cell is really a very different process. We need to find ways to derail these cells from their default pathway and convince them to do something different. It's not only once but multiple times that you have to keep pushing them in the right direction,and you lose a lot of cells, so the efficiency is low. Basically we need to figure it out based on trial and error or by using developmental approaches.
Now that we know how to do this with mouse cells, we will be able to apply our technology to human cells in the near future. This will open the door for very specific studies of certain causes of hearing loss and hopefully how they can be cured.
what is the other direction for your research?
The other direction that is evolving is something we found when we tried cell transplantation in animal experiments.In these experiments we observed that the transplanted cells were causing tumors. This has been observed before with cells that were derived from embryonic stem cells. Embryonic stem cells are very potent, they grow well and that is the problem. We are currently finding ways to eliminate tumors from growing. We are figuring out how we can purify the inner-ear cells that we generate in the culture dish and leave behind the non-inner ear cells that apparently have tumorigenic potential.
Let's say we start out with 100cellsit's actually millions of cells at a time, but let's say it's 100. Out of these 100 cells, maybe 2 cells have the capacity to turn into an inner-ear hair cell. This is how inefficient it is now. Some of the 98 other cells that do not turn into hair cells still have the proliferative capacity that the original embryonic stem cells had, and this can become dangerous.
Personally, I am not happy doing animal experiments when I know that the outcome is very bad. We have currently stopped all animal experiments regarding transplantation because of the tumors.
how are you trying to eliminate the bad cells and purify the good cells?
Luckily this issue has been solved in other fields. People have found with hematopoietic or blood-forming cells that the good cells and the bad cells present different kinds of proteins on their surface. You can use antibodies to recognize these different protein markers, and as a result sort the cells. This is just one example of how we apply cell sorting to our research to make it more efficient. In this instance we are using sorting to find cells that are the most likely to turn into hair cells. Our goal is not only to eliminate bad cells but also to find the best inner-ear cells by optimizing at every stage of the protocol.
The protocol has multiple steps. As an example, say after the first step we have 30 percent efficiency, meaning that 30 percent of the cells we generate are correct. The second step has 50percent efficiency. So overall that is only 15 percent. All of these intermediate steps need individual optimization so that the result at the end is better than 2 percent.
what other plans are on the horizon?
My goal is to develop a treatment that is not transplantation. I am more interested in creating a drug that can be applied without surgery. To that end, another line of research that we are currently pursuing is to use mouse and human inner-ear cells that we generate from stem cells in order to test potential drug candidates in other words, to test on inner ear tissue that was generated in a test tube.
For instance, you can test a drug for its potential to prevent aminoglycoside toxicity, or its potential to induce hair-cell regeneration. You would not need to use many animals because by generating cells in a culture dish, you can split them up into hundreds or thousands of culture dishes. You can do thousands of tests in an afternoon. I find that very powerful. I think this line of research can contribute to novel drug discoveries in the future.
I have two dogs I love animals and as I mentioned before, it breaks my heart to do animal experiments when we're no tat the point where it makes sense. Of course, animal experiments are critical when you are at the right point, since cells and computer models can only take us so far. However, I think it's important to do them when it comes to showing that what we have developed is working, but we should avoid doing them if there's another way.
"I'm quite optimistic that we can reach the same stage with human cells as we have with mouse cells in the near future. I hope what took us seven years with mice may take us only four years with humans."
A view of hair cell like cells made from mouse embryonic stem cells. The maintenance of embryonic stem cells in the lab.
taking the long view, what are the applications for your research with regard to genetic testing?
Novel genetic technologies are revolutionizing medicine. This is happening right now and it will result in better health care for patients with hearing loss.
For example, it is already possible to test a patient for all known hearing loss genes without the tremendous effort that was needed a couple of years ago. I presume that these novel tests will be soon on the market and available to the general population.
This means a doctor will be able to tell a patient or a parent of a child with hearing loss not only why there is hearing loss but also better predict the progression of the hearing loss. In some situations, it will help in making decisions as to whether a hearing aid or a cochlear implant is the most appropriate treatment.
We are just at the beginning of using these novel technologies. I believe that our future work will play a role in the discovery of the underlying principles causing human hearing loss that have not been researched in the past. About half of the hearing loss genes are currently undiscovered and it is necessary to know what the problems are before we can seriously think about curing them.
Hearing loss is a very complex disorder much more complex than we originally thought. But giving up is not an option. We will use these new technologies and collaborate with researchers and clinicians so that one day, a child with hearing loss will have better treatment options.
Yishane Lee, Editor
"We will be able to apply our technology to human cells in the near future. This will open the door for very specific studies of certain causes of hearing loss and hopefully how they can be cured."
