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Monday, January 25, 2010

How to be a science geek on YouTube, Part I

by Jennifer Phillips, Ph.D.

Studying a disease like Usher syndrome has required me to become intimately familiar with the inner workings of the sensory systems involved. The days I don’t spend actually discovering new things about eyes and ears in my own lab are spent reading everything I can get my hands on about what other people know about them. Some people (*cough* Mark *cough*) might consider this reading of dry scientific papers an onerous task, but I actually love it. While most of the science I’ve discussed on this blog so far has been fairly clinically oriented I think there’s some value in spending some time getting back to the basics of the system. I was faffing about on YouTube recently and rediscovered a very cool little video that seemed appropriate to share on this blog. The video is very short, but I feel compelled to burn up a few more minutes of your time to give a little background and explain what it is and why it’s cool—at least in my humble opinion.

The cochlea is an extraordinary structure. The image below, taken from an article in the journal Nature Reviews Neuroscience, shows a cutaway section of cochlea to reveal the key organ within it: the organ of Corti. The organ of Corti contains the sensory cells–mechanosensory hair cells, to be precise—responsible for hearing:


From Fettiplace R and Hackney CM (2006) The sensory and motor roles of auditory hair cells Nat. Rev. Neuro. 7: 19–29 doi:10.1038/nrn1828

Note that there are two different types of hair cells present here: A representative inner hair cell on the left, shaped like a teardrop, and the outer hair cells on the right, shaped like cylinders. As you may recall from earlier discussions of inner ear anatomy, the tips of all mechanosensory hair cells, both inner and outer, feature specialized structures called stereocilia. A number of stereocilia stick up from each of these cells like stiff little bristles protruding into the hollow, air-filled part of the cochlea and are collectively known as the hair bundle. Hair bundles are the ‘receivers’ of the cell, responsible for intercepting sound waves in the environment and converting them into electrical energy. In Usher syndrome patients, the organization of the hair bundles is perturbed, so the cell’s ability to receive sound signals is impaired. On the plus side, the fact that the hearing problems in Usher syndrome stem from defects in the ‘receivers’ (a.k.a. ‘sensorineural deafness’) is the reason that cochlear implants can successfully substitute in this capacity.

At the other end of the hair cell there are synapses, which are responsible for converting the electrical message into a chemical message that adjacent nerves (stringy yellow or red lines in the figure above) can receive and relay to the brain.

In addition to both of these signal processing regions, outer hair cells, the ones that resemble cylinders on the right side of the above figure, exhibit the unique property of electromotility in which the electrical signal is converted into a mechanical signal, such that the sides of these cells can actually shorten or lengthen in response to sound. Functionally, the electromotility of the outer hair cells provides improved auditory acuity, enabling the outer hair cells to discern subtle differences in frequencies. The complex and nuanced physiology of the inner ear is a marvel to behold, and as wonderful and life changing as cochlear implants have been for Usher patients, specializations like electromotile hair cells demonstrated that our technology is still quite limited in its ability to emulate the intricacies of our sensory systems.

Whoops, I promised you a movie, didn’t I? Ok, here it is. When you push ‘play’, at long last, you’ll see a recording of a cultured outer hair cell taken from a mammalian cochlea. The sound to which the cell is responding is being played through a thin glass pipette, which you can see, faintly, in the frame, and the electromotility response should be pretty apparent. Enjoy.

Friday, January 15, 2010

One Opinion on How To Form an Opinion

by Mark Dunning

My wife is a fan of the sniff test. The expiration date on a carton of milk doesn’t matter. She still wants to stick it under my nose and ask “What do you think? Is this OK?” I usually offer my opinion by gagging or shrugging. The interesting thing is that she doesn’t always act on my opinion. Sometimes I’ll say it’s good and she’ll throw it out, sometimes I’ll run screaming from the room and she’ll pour it on her Cocoa Puffs. That’s OK. I’m just offering my opinion. I don’t have any particular proof that it’s good or it’s bad. The checkbook is a different story. I know what’s in the bank and I know our weekly expenses. So when my wife asks if we can afford something, she (usually) heeds my advice because I have the facts.

I get asked my opinion on Usher syndrome treatments a lot. I find this surprising because as any regular reader of this blog knows, I’m generally light on facts. But that’s the problem when it comes to Usher syndrome treatments; we’re all light on facts. You see there are no widely accepted, scientifically proven treatments for the vision component of Usher syndrome. So just about everything at this point is an opinion. There are few statements of debits and credits and cold hard facts. That leaves families, like my wife with her spoiled milk, to decide for themselves if a treatment is good or bad. My advice? Line up a lot of noses before pouring anything on your Cocoa Puffs.

In my last post, I urged families to fully research any proposed treatment with a level head before embarking on it. All too often desperate families are willing to jump on the first positive advice they get without fully vetting it first. It’s human nature to run to the light when you’re stumbling around in the dark.

I find families take chances with unproven treatments for Usher syndrome that they would never even take with a proven treatment for other conditions. We’re all familiar with the concept of getting a second opinion. A doctor says you need hernia surgery, you goes ask another doctor if he or she agrees with the first. Yet families with Usher are often so relieved that someone is offering something that might help, that they just jump in without question. In fact, families are often careful NOT to ask questions because that might expose some reason why the treatment is not appropriate causing the physician to withdraw the treatment offer and stifling that glimmer of hope. It’s the exact opposite of how they should act.

Now no one sniffs around for Usher syndrome treatments more than I do. So here’s what I do to determine if something passes the sniff test:

Governmental approval

Say what you will about our governmental bureaucracy, but it works pretty well when it comes to ensuring treatments are safe. There are no Usher syndrome specific treatments that are currently approved by the Food and Drug Administration or recommended by the National Eye Institute that I know (again, I might be light on the facts). There are approved treatments that might be appropriate for Usher syndrome such as Cochlear Implants and Vitamin A but these are not Usher specific treatments.

Still, that’s the first thing I look for: Is this treatment or this study approved by the NEI or the FDA? If not, I’m skeptical. As a result, I tend to be more skeptical of any treatments outside the US. That may not be correct thinking since there are many reputable and talented researchers outside this country. In fact, my wife’s cousin, who is British, is the first call we make when we need information. So while I’m more skeptical of international efforts, I obviously don’t discount them completely.

Clinical trials are different from treatments. A clinical trial is a test of a potential yet unproven treatment. These are enticing to many families who have already heard over and over that there is no cure for Usher. So if there is no cure presently, something that holds promise might be worth trying.

I’ll talk more about clinical trials in a later posting, but like treatments, I would want to know that a clinical trial met governmental requirements before I participated. You can find a registry of clinical trials maintained by the US government at this site developed by the FDA and NIH. The nice thing about this site is that it also includes most internationally and privately funded clinical trials as well as those funded in part by the US government. It’s a pretty comprehensive list. I personally would be very wary of participating in any that is not on this site.

Get the Facts

Just because the government approved a treatment or a trial doesn’t mean it’s right for you or your family. So the next thing I do is judge the science for myself. Now I’m not a physician or a research doctorate. The details do go over my head at times. Jennifer wrote recently about how to spot good science which is a good lesson to follow. Personally, I look for several things in particular.

How long has this treatment been studied?
How many people have been part of the study?

Who are the people that participated?

How long have those individuals participated in the study?

I’ll give you an example. I know of one particular treatment study that’s been going on for nearly 20 years with hundreds of participants, many of whom have been participants for the entire life of the study. That’s good science right there. Lots of data to back up the findings of the study. If it’s been going on that long with that many people, they have probably seen most of the common side effects which makes me more confident it’s not going to harm my daughter.

The caveat in this one particular study is that while Usher patients were included, they weren’t the focus. So then the same questions come up. Exactly how many Usher patients are we talking about? What type were they?

I personally don’t want to go first. If a bunch of people with the same type of Usher have tried it with no ill effects, then I’ll consider it. You might be less cautious. That’s your prerogative. As I said, this is just my opinion.

For a great list of questions to ask before participating in any clinical trial, check out The Center for Information and Study on Clinical Research Participation. They have a great brochure with a list of questions to ask.

Deciding Who To Trust

Traditional hernia surgery is FDA approved and has been performed on thousands of patients with great success. However, if I’m due to have that surgery and the doctor comes in smelling of bourbon and slurring his speech, I’m not having it. Why? Because I don’t trust him. Trust is the most important factor in deciding if a treatment is right for you.

For me, trust in a physician comes down to one thing (aside from sobriety, of course): Will he or she take the time to answer, thoughtfully, every one of my questions? In my case, this is my daughter we’re talking about. I don’t care how many degrees or plaudits a person has, they are not touching my daughter until I understand every single detail of what is going to happen. If they don’t have the time to spend to make sure that I am completely comfortable with a treatment, then they don’t care enough about my daughter and they aren’t going near her.

Of course, there are other factors I consider as well. Degrees help. Obviously I weigh the opinion of someone who is medically trained more than the average guy on the street. But I have found that many physicians don’t know the first thing about Usher syndrome and in those cases, I actually value the opinion of families with Usher over those of a particular physician. So degrees are important, but expertise in the field matters more to me.

Finally there is family. I mentioned earlier my wife’s cousin in London. She’s at the top of our trust list because she is medically trained, an expert in the field, family, and always willing to spend an hour explaining a treatment to us. Bella’s godfather is also a physician. Nobody cares more about her well being than him, but he is not an expert on Usher syndrome. We talk to him about proposed treatments to see if they at least are logical medically, but I don’t think I’d trust a treatment he proposed even if his heart was in the right place (not that he would do that). And of course there is Grandma and Grandpa and aunts and uncles and cousins and other people who care deeply about Bella. We consult them for the sniff test, but like my wife with the milk, they don’t have a big influence on the final decision.

Finding Experts

One of the reasons we created the Coalition for Usher Syndrome Research was due to the frustration many of us, families and researchers alike, felt in trying to find purported experts. Families, physicians, and researchers alike wanted to be able to quickly find experts in a particular treatment or Usher sub-type who were willing to spend the time to speak to them. Or they wanted to find that expert to provide a second opinion. It was incredibly difficult.

I’d love to say it’s easy now, but though it’s gotten better, it’s still pretty opaque. The best places to check are with the Coalition for Usher Syndrome Research or with the Foundation Fighting Blindness. The Coalition is aware of just about every researcher working on Usher focused projects. The Foundation Fighting Blindness knows just about every retinal related project going on in the world, including those that focus on Usher or might be appropriate for Usher. If neither of those organizations are aware of a particular researcher, I’d be skeptical. I wouldn’t dismiss him or her, but I’d be more wary.

Decision Time

At the end of the day, however, there is only one opinion that matters. It’s yours. Use the resources and process above to educate yourself then look in the mirror and ask yourself these questions:

Have I learned all that I can about this particular treatment?

Have I fully vetted that information with enough people that I trust?

Do I trust the folks administering the treatment?

Do I feel confident it won’t harm me or my family in any way?

Do I truly believe it has the potential to help me or my family?

Then make your decision and trust it. No one is more expert on what is best for you and your family than you.

Thursday, January 7, 2010

Fresh Science for a Fresh New Year

By Jennifer Phillips, Ph.D.

Happy New Year, Dear Readers! I hope 2010 is off to a wonderful start for everyone. I took some welcome time away from the lab over the holidays, and although I couldn’t completely unplug from correspondence and submission deadlines, it was enough of a break to enable me to refresh and recharge sufficiently. As such, my well-rested brain is ready to churn out a hearty helping of science for your reading pleasure.
Figure 1: Top: My brain on vacation; Bottom: My brain while writing this post.

Just before the break, a new paper came across my desk that I thought would be great fodder for discussion on this blog. The paper, entitled “Cell Transplantation to Arrest Early Changes in an USH2A animal model”, was published online last month in the journal of Investigative Ophthalmology and Visual Science, and there are two key findings reported:

1. The paper provides a nice characterization of the retinal defects in a mouse model for Usher Syndrome type 2A, showing late-onset and progressive vision loss quite similar to the human disease pathology.
2. As the paper’s title suggests, it presents evidence that transplanting a particular type of cultured cells into the retinas of these Ush2a mice improved visual function.

The results could have an impact on developing Usher treatments in the future. The research describing the disease progression in the mouse model adds a great deal to the understanding of what might be going on at the cellular and molecular level in the retinas of human USH2A patients. As previously discussed, such information could have long term benefits for developing treatments targeted to specific areas or functions of retinal cells. The transplant data, clearly, provide one of the necessary precursors to a clinical study of the rescuing effects these cultured cells could have on human USH2A patients—and perhaps other types of Usher syndrome as well.

So, let’s fire up the old grey matter, delve a little deeper into the results and see what they found. The Ush2a ‘knockout’ mouse has been around for a few years. It is a line (or ‘family of mice’) from which the Ush2a gene has been deleted. These mice can’t make the Usherin protein, and as a consequence they exhibit the mouse version of Usher type 2a, with hair cell defects from birth and later vision problems. The original paper (linked above) from the Harvard University lab of Dr. Tiansen Li described the retinal defects in aged Ush2a mice, finding that, by 20 months of age (which is pretty old for a mouse), many photoreceptor cells had degenerated, resulting in thinning retinas and (unsurprisingly, given the extent of photoreceptor cell death) diminished ERGs. The new study, conducted by Raymond Lund and colleagues at the Oregon Health Sciences University, confirmed some of the original analyses as well as conducting new ones. The big difference was that the OHSU researchers identified some defects in younger Ush2a mice that are noticeable many months before the first signs of retinal degeneration show up, which gives us some new insights into how retinal cell behavior may be compromised in Usher patients before any of these cells actually start to die off. The researchers examined visual function with a behavioral technique known as the optomotor response (to watch a short movie of a mouse undergoing this visual function test, click here)

If you visit the above link you’ll see a mouse in the center of a rotating drum. A black and white pattern is projected onto the sides of the drum, and the pattern shifts and moves when the drum is rotated. Researchers can observe how reliably the mouse follows the moving pattern, indicating visual acuity. By varying the contrast between the color patterns, researchers can also test the sensitivity of the mouse’s ability to detect color gradients: it’s easier, visually, to detect the contrast between black and white, and more challenging to detect contrast between black and grey. The end result of this analysis is a reliable readout of the visual ability and behavior of the mice. Groups of mice with either normal copies of the Ush2a gene (control group) or the Ush2a “knockout” (experimental group) were tested with such an apparatus to gauge visual function at various ages.

These researchers found that some visual defects were detectable in the Ush2a mice quite early in life (between 2 and 3 months), many months before the documented onset of retinal cell degeneration in this line. Ush2a mice had an increasingly difficult time following the rotating patterns, and their contrast sensitivity—the ability to detect differences between shades—worsened over time as well.

The Researchers next looked for changes in the localization of retinal proteins in Ush2a mice. Even though the cell number and organization of these mice at younger ages was indistinguishable from controls, the proposed role for Usherin in helping to move proteins to different parts of the photoreceptor cell predicts that this process might be defective in animals lacking Ush2a. And indeed, that’s exactly what they found:

Figure 2: From Lu et al, 2009. Opsin protein localization in control and Ush2a mouse photoreceptors. The panels marked ‘WT’ show retinal sections from normal control mice. Those marked ‘Ush2a’ are from knockout mice, and the ‘P….’ number tells us how old the mouse in question was: “P80”= 80 days old, etc.

The top two panels (A and B) of Figure 2 show a high magnification view of photoreceptors in control mice at age 80 days and 360 days, respectively. The only part of the cell you can see in these panels is the very tip, where a chemically labeled opsin protein is present in the region known as the outer segment. The bottom four panels, C-F, show the same view of photoreceptors in Ush2a mouse eyes of various stages. Note that you can see a lot more of the cell now—long, stringy bits in the middle and chunky blobs at the bottom, because the opsin protein isn’t where it’s supposed to be. There is an apparent defect in the mechanism that transports it to the outer segment—a process that, we can infer, requires the function of Usherin protein.

Moving forward, the researchers next looked to see if the defects specific to the cellular machinery of the photoreceptors were having any effect on other populations of retinal cells. Specifically, they looked at a type of cell known as glia. In a nutshell, glial cells are sort of like parents—their job is to help specialized neurons grow and develop, keep them fed, and clean up after them . When a neuron is sick or damaged, glia often respond by producing chemicals to help stabilize the ailing neurons. In this study, the researchers looked for evidence of increased production of a protein known to be up-regulated in glia in response to retinal damage, and found, as early as 2-3 months of age, the glia in Ush2a mouse retinas began ramping up their production of this protein. This further indicated that even though the photoreceptor degeneration had not yet commenced, photoreceptor cell function impaired to the extent that the retinal environment was altered.

Thus, in addition to adding to the big picture of what we know of how retinal cell function is affected by defective or missing Usher genes, this first set of experiments also identified the earliest time point at which vision defects could be detected, thus establishing a window of opportunity for rescue therapy. With this knowledge in hand the researchers proceeded with their transplant experiments, using mice between 2 and 3 months of age (P80).

Before I proceed with the description of the transplant experiments, I want to take a moment to talk about research ethics. It’s often said that science, as an endeavor, is morally neutral, but scientists, of course, are not. The line between what is technically possible and what is morally acceptable, to us as individuals and as a society, can sometimes be blurry, and is certainly variable from person to person. The research described below might make some readers uncomfortable, and regardless of the therapeutic implications, it might not be a treatment that some of you would choose to pursue if it were available. This is a valid and important topic of discussion as we move forward into the much hoped-for realm of Usher therapies, and I plan to give it a more thorough treatment in the near future. But for the purposes of this particular blog post I will press on with my summary of the science, and leave the ethical implications thereof for next time. Here goes:

The cells these researchers selected for transplant are a kind of stem cell known as hNPCs. The ‘h’ stands for human—these cells were originally obtained from human fetal tissue and have been grown in laboratory culture dishes since that time. ‘NPC’ stands for Neural Progenitor Cells. These cells originate in the developing central nervous system and have the potential to give rise to a variety of different neurons when properly stimulated. Several labs around the country maintain hNPC lines that are requested by other labs for use in various experiments seeking to repair or forestall neural degeneration or damage of some kind.

The lead researcher on the Ush2a mouse paper under discussion has experimented previously with this cell line, rescuing retinal degeneration in a rat model and testing safety and efficacy of the surgical transplant procedure in a primate model, as a precursor to human clinical trials. With these previously established techniques, the researchers transplanted hNPCs into Ush2a mice via injection into the back of the retina, similar to the way in which the gene therapy for LCA patients was introduced. Visual function in these animals was assessed 4 weeks and 10 weeks after the transplant, and marked improvement was reported in both visual acuity and contrast sensitivity, using the optomotor response test described earlier.

They also looked at protein localization in the transplanted animals, and found significant, long-term improvement of the opsin trafficking problems observed in the photoreceptors of Ush2a mice. The cool thing about this last result is the effect of the therapy with respect to the timing of the transplant. Remember that the surgery was done at ‘P80’, the time at which the first signs of opsin mislocalization were detected in the untreated Ush2a mice. If you glance back up at panel C in figure 2, you’ll see that the opsin protein at this time point is showing up in other compartments of the photoreceptor cell, below the outer segment where it’s supposed to be. Now check this out:


Figure 3: From Lu et al, 2009. Opsin protein (green) is properly localized to the outer segments of a P150 Ush2a mouse retina when transplanted hNPC (red) cells are present. Cell nuclei are stained blue to show cell numbers and organization (I don’t know why they didn’t do this in the Figure 2 panel—it really helps, doesn’t it?).

Based on the data in this paper, the Ush2a mouse eye in this picture would have looked like the one in Panel C of Figure 2 on the day that the transplant took place, with opsin going places it shouldn’t. The presence of the transplanted cells, therefore, not only prevents opsin trafficking problems, but apparently fixes preexisting ones as well.

The real head-scratcher here, at least for me, is: How? The hNPCs, being of human origin, can be identified by an antibody that recognizes a particular human protein in these cells. This protein is labeled red in the above panel, showing you exactly where the hNPCs have ended up, ten weeks after they were transplanted. Note how they are lined up above the photoreceptors (the tips of which are stained green with the opsin protein marker). They haven’t integrated into the mouse retina, they haven’t replaced any mouse retina cells, it’s not clear that they have developed into any particular cell type, they’re just…hanging out. But they’re clearly doing something to improve protein trafficking in their neighboring cells. That blows my mind, but while I personally am dying to know HOW they are able to improve the cellular performance of their neighbors, this knowledge isn’t necessarily a prerequisite for clinical trials. The important thing (beyond safety, of course) from that standpoint is the effect itself, not the mechanism, and the effect is pretty remarkable.

Reference: Lu B. , et al: Cell Transplantation to Arrest Early Changes in an Ush2a Animal Model. IOVS Papers in Press. Published on December 3, 2009 as Manuscript iovs.09-4526