Q&A with Dr. Aaron Gardner

[Arashi]

Aaron, some time ago Desmond posted this link: http://online.liebertpub.com/doi/abs....TEA.2013.0547 and we thought that hairloss was cured since it seemed these guys could now culture cells while keeping all genes expressed. But re-reading that abstract they dont specifically say that 100% genes were expressed. What do you think ? Cause if I understand correctly, once we can culture dp cells (enough of them) while retaining 100% gene expression, then we've overcome a major hurdle and you guys can focus on things like hair growth direction etc, right ?

[Armandein]

the first important question, which often appears in our discussions but nobody is able to really answer it scientifically:

what is it all about with the sebaceous gland? Is it required during the hair cloning process to also create a seb. gland along with the whole follicle or can it be assumed that a lab-grown follicle would attach to one of the many exisiting seb. glands in the bald scalp instead?
Because we never heard of researchers that they are also trying to create the seb. glands in lab, and many people state a follicle without the gland is no real follicle. What is your opinion about it?

a dump question: is the seb. gland even required for hair at all? What would happen if you implant a lab-grown hair into the scalp, and the follicle wouldn't attach to a seb. gland?
it is often said that the seb. gland keeps the hair moist and soft, but isn't it more harmful than useful =D it creates a lot of sebum which is always bad for the skin, and probably also for the follicle, as the gland also does DHT conversion, where the DHT then moves down to the follicle.
in other words: isn't the seb. gland just a bad byproduct of the nature? (like the blind gut?) =D

Joaquim you are my hero
IMHO hair shaft and dermal papilla are only a part of the pilosebaceous unit. I am wondering why sebaceous gland is so important, even in multipili hairs have the same number of sebaceous glands? It is possible that SG are part of interactions between the DP and hair follicle epithelium gives rise to the hair shaft.

[Artha]

Thanks alot Mr. Gardner!

[HairlossAt15]

Dr. Gardner,

Have you read much about the PGD2 research (eg http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3319975/ and http://www.ncbi.nlm.nih.gov/pubmed/24521203 ) and if so may we have your opinion on the potential of this pathway?

Thanks a lot.

[Arashi]

Wow, we've got more interesting info in the last 2 days then I got in the last 2 years that I've been member here So you don't accept donations, then we should at least collect money to erect a statue for you Aaron Thanks a thousand !!

[joachim]

A mature follicle is made up of tens/hundreds of thousands of cells. We don't need this many cells to make a follicle, we are trying to replicate the early induction events by implanting a relatively small number of cells ~2-8,000 into the skin, that can induce the surrounding tissues to form a follicle. If you think of hair follicle formation in the embryo then it is coming from a tiny number of cells.

with what cell types are currently working in the lab? (of which cells does the follicle consist?)

DP cells
DSC cells
Epithelial cells
ECM? we heard the extracellular matrix is build up automatically when the DP cells come and interact together (according to team Lauster)

are you able (and other researches too) to isolate EVERY different cell type out of the follicle? or is there still a cell type which makes problems and can't be isolated for multiplitication?

[agardner]

This sounds quite 'automated' in the sense that, when compared to traditional hair transplants a doctor needs to spend several hours harvesting donor grafts and then several more hours implanting them into balding areas.

Would the culturing and implanting of the DP cells into the scalp be rather quick in comparison to a hair transplant? If so, would this method be costly for patients?

• Isolation is relatively quick, I can do ~20 follicles an hour.
• Expansion in 2D takes ~4-6 weeks depending on desired end number.
• 3D culture as we currently do it is relatively laborious. For example creating ~500 spheres would take about an hour. These need to be fed manually every 3 days which would take several hours. There are ways of automating this though, we just don't do this.
• Implantation technique, this I don't know. I would assume it would take a similar amount of time to follicle implantation. We currently separate the dermis and the epidermis enzymatically, then place the spheres between them, this isn't clinically viable.
• Cost, I honestly have no idea, a similar cost to the surgeries I would assume.

[agardner]

Would additional funding directed to your university's endowment would help speed up your research?

If yes, how much do you need?

I would be very, very careful about contributing money directly to a research project, and be wary of someone requesting funding in this manner. I personally wouldn't accept money in this way.

If you are interested in contributing money then I can suggest several means of doing so:
1) Form a charity. Pool any money, have a structure, recruit experts to your board. Then you can analyse the field and decide where you want to apply your funding. By forming a charity you gain numerous tax benefits, especially in the UK not sure about elsewhere. From this position you could then also ask for representation on the various hair research societies, request conference attendance/filming etc...

For an example of an extremely effective charity that directly funds research in the UK, http://www.cysticfibrosis.org.uk. I am sure there are equivalents outside of the UK.

2) Donate to an existing funding body/charity. For example in the UK the MRC accepts donations and you can specify areas of interest that you would like you money to go towards. http://www.medicalresearchfoundation.org.uk, again I'm sure equivalents exist outside of the UK. But again, pooling money would be the best option.

3) Sponsor early career researcher attendance to conferences. This is vital, encouraging young scientists to build networks in the field, present their data and see other groups present their data will only help. Early career researchers also often don't have much provision in their funding for travel so these grants are vital.

4) There will be investment opportunities, but I'm not a financial whiz so can't really comment on this.

But just to reiterate be very, very careful of directly giving money. Snake-oil salesmen will abound.

[agardner]

what is known about the trials in Taiwan with over 400 people, which should start soon? What kind of team is it? Why are they starting trials soon, if there are still some missing puzzles? What company is behind it, and what else do we know about them?

I mean, to start a trial with 400 people requires some confidence.
They are hopefully not going to just randomly inject some cells into bald people and see what happens then.

Are you aware of what's going on in Taiwan?

Yes I am aware of the clinical trial, but I don't know anything more than you guys until they present their findings. It will be very interesting to see what comes out of the study, especially as the implants will be in for a lot longer time period than in the models used by Dr Higgins. But I'm not aware of what their outputs will be.

http://clinicaltrials.gov/ct2/show/record/NCT00506636

[agardner]

This:

"A:
I'm not sure. The majority of studies use epithelial cells that are very receptive to follicle induction i.e. mouse or human neonatal epithelial cells. We are attempting to use adult human only cells, but this is "healthy" tissue. As far as I know there are no groups using alopecia scalp tissue to test for inductivity. Speculating, I would assume if an inductive enough construct was created that the initial follicle would form. However, as this isn't treating the underlying causes of the various alopecias I would assume the follicle would then degrade as the previously, perhaps even at a faster rate due to the loss of fatty tissue in the scalp."

We're in it for the long haul boys. Especially if the roots of baldness do not lie with instructions given by the hair follicle cells. If it's the rest of the body dictating what hair is lost then it's going to require a replacement of your entire scalp to cure this thing. which I suppose is possible too with tissue engineering, but we're looking at many decades and very invasive procedures. you would probably have to not only get all new follicles, but also remove all the old ones that are signalling your scalp to degrade.

The effectiveness of any treatment is going to vary case by case, but making new follicles does not fix any underlying issues that caused hair loss in the first instance. I envisage that complementary therapies will have to be used as well in the more severe cases. Beyond this I could see full scalp regeneration technologies being developed, as it would be much easier to control in the lab, but that's not something that will come after reliable follicle induction.

[agardner]

another question:

Has anyone of the researchers around the world ever created a cosmetically acceptable lab-grown copy of a follicle which is sprouting hair now on someone's head (a real human)? maybe by chance, and maybe not consistently, but did it ever happen?

Not that I'm aware of. It's possible that someone's tried it but I'm not aware of it.

[agardner]

a general question:

Are you, Dr. Gardner, and Jahoda fully aware of the other researchers work?
Do you even exchange information with Tsuji Labs, Team Lauster, or others on a regular basis?
Or is there too much competitive thinking?

We are aware of the work that they present at conferences, in papers etc... and where they are planning to take their research.

But as a whole groups are very unlikely to exchange information before publication. Jobs, especially for early career researchers like myself are dependant on papers and grants, if you give your ideas and data away before you get a paper/grants then you might not have a job. It's not right, I really don't like the system (I can't think of anyone who does) but unfortunately that is how the system is.

If you want to read more:

http://www.theguardian.com/higher-education-network/blog/2014/may/01/academic-anonymous-leaving-academia

http://www.theguardian.com/higher-education-network/blog/2014/mar/22/postdoc-research-universities-publish-academics

http://www.reddit.com/r/IAmA/comments/1sq4vd/im_randy_schekman_corecipient_of_the_2013_nobel

http://en.wikipedia.org/wiki/Publish_or_perish

I am very pro-open access, and I want to talk about my work with other scientists and the people who my research will impact on but it's very difficult.

[agardner]

another scientific question:

how many cells are required to form a follicle? are we talking about hundrets, thousands, millions, or billions?

A mature follicle is made up of tens/hundreds of thousands of cells. We don't need this many cells to make a follicle, we are trying to replicate the early induction events by implanting a relatively small number of cells ~2-8,000 into the skin, that can induce the surrounding tissues to form a follicle. If you think of hair follicle formation in the embryo then it is coming from a tiny number of cells.

[agardner]

* Aaron, thank you for coming here and talking to us.

* Are you saying that when you do manage to encapsulate DP spheres with epithelial cells you get total (100%) preservation of hair inductivity despite mass pass culture?

* I think you're saying is that the problem is getting the epithelial cells to stick to the DP cells. I gather that you're saying that sometimes the epithelial cells do stick to the DP cells but other times they don't. Since sometimes the epithelial cells do stick to the DP cells I'm wondering why you can't simply isolate the method that results in the epithelial cells sticking to the DP cells and only use that method? In other words, since you are able to make the epithelial cells stick to DP cells sometimes then just use the method that results in the epithelial cells sticking to the DP cells and the problem is solved, right?

* When you do get the epithelial cells to stick to the DP cells what do you differently from when epithelial cells don't stick to the DP cells?

* What are you going to try to do to get the epithelial cells to stick to the DP cells? Do you have any ideas?

1) We rapidly expand our DP cells in 2D culture, during this step they lose their inductive capacity. After expanding the cells we transition to a 3D model. Just by placing the DP cells in a 3D system they form tightly packed spheres. Dr Higgins in her paper showed that this act of forming a sphere restores ~40% of the genetic signature of a functional human DP that is lost during 2D expansion. We're attempting to coat with epithelial cells, when they work we see markers of inductivity turned on that aren't present even in the dermal only spheres. We haven't repeated Dr Higgins work looking at the genetic signature, but we are going to, so I can't give a numerical figure for restoration. It will be higher than ~40% restoration, but I would think it unlikely to be 100%.

2) We use the same culture and coating techniques however sometimes it works and sometimes it doesn't. We've looked to see if there are any correlations e.g. donor age, donor tissue location, cell passage number etc. but nothing has stood out so far. So there is patient variability, we need to understand what is causing this variability. We are attempting several things:

• Differnt epithelial cell populations, e.g. hair specific populations
• Different 2D epithelial expansion techniques
• Different coating techniques
• and several other things

[agardner]

Question about 3D printing:

There are currently 2 or 3 serious companys working on 3D printed organs with some good breakthroughs already. However, it's still much work to do for creating complete working livers, kidneys including blood vessels etc.
blood vessel printing with the help of some special support material just achieved the first success some weeks ago.

But let's mention one company specialized to cell-by-cell 3D printing:

http://www.organovo.com/

They are really able to print a bunch of different cells in an exact 3D shape using Hydrogel as support material for the cells. Printing cell by cell, accurate and reliable.

so when the key in follicle formation and achieving 100% gene expression is to guide the cells in a way that they mimic the natural form of a follicle to enable the different cells interacting with each other, then why not use Organovo's 3D printer to print all the required cells (DP, epithelial, dermal sheath cup, ...) in the exact shape of a follicle? The process probably only takes a few minutes per follicle, and after that they can interact and bind together (takes some hours probably). The hydrogel dissolves after sometime, I think. At least as proof-of-concept this should really be looked into in detail. Organovo would of course be interested to give their printer the chance to try this out. This could be tested within a few months to have proof-of-concept.

When the 3D printing process turns out to be successful then the printer just have to be extended by multiple nozzles and chambers (reservoires) for the different cells to allow printing of multiple follicles simultaneously.
but first, the proof of concept is important!

And we always hear that the right 3D culturing technique is important (hanging drops, PVA tubes, Matrigel culturing, etc.) but the 3D printer is the easiest way to make a perfectly shaped follicle with different cell types, and the technology is already available.

Maybe the biochip from Team Lauster can be combined with it: first, 3D print the cells, then supply the dishes with further nutrients and proteins so that they have the right signaling to develop a hair.

What do you think? do we have a real chance with 3D printing or is it simply infeasible and we should all forget about 3D printing follicles?
If it's infeasible, what's the reason for it? Has anyone of the hair researches tried it out ever?

3D printing is going to be big, there are lots of people looking into this and I know there are people talking about doing it with the follicle (which makes me think they are already attempting it). As you say it would seem to be the ideal way to organise complicated structures in the lab prior to implantation.

It is one of the methods that we have thought about for improving dermal/epidermal interactions in our 3D models. But the technology isn't there yet. But it is something that I am keeping an eye on and the majority of other groups will be as well (if they are not already experimenting themselves).