Microglia originate in the yolk sac (embryonic origin)

59:50yolk sack. Back when we're an embryo, um, they colonize the brain act before the bloodb brain barrier forms. >> Oh, okay. >> Okay. So, they get inside and then the bloodb brain barrier forms and then they maintain themselves within the bloodb brain barrier. So, they they don't come from the bone marrow like other immune cells do. Okay. They're sort of innate in the brain and then they stay in the brain. Okay. >> Okay. It's another big reason why it's hard to access these things is they're inside the bloodb brain barrier, right? So you can't really like poke it with drugs and things like that. Um, >> and a lot of times they're basically in homeostasis, so they're doing their normal thing where they're moving around, they're cleaning up um byproducts. They have these things

1:00:31called um processes which are um if you've ever seen immune cells like in in real time, they'll like protrude out. They'll have these arms that stick out and they're basically just surveying, right? They don't have eyes. So, they're they're using these >> protrusions to increase their surface area and sense the chemical environment. And if they sense something that's bad, >> then they go into a state of response where they try to eat whatever thing is bad or they try to deal with the the plaques and and stuff like that. But if you have too much of that, then that's going to destroy the neurons around you as well,

1:01:11>> right? So if you have too much upregulation of that state, that's pretty bad, >> right? >> And what ends up happening is you've got this aggressive amoid deposition that happens in let's say mice models, right? >> Yep. >> This is the this is something that mimics the Alzheimer's. So you've got a neuron in the middle and all of these plaques that are sort of getting in the way of the neuron trying to do everything. Yes. >> And if your micro gleal cell like actually senses some of these plaques, senses too much of the plaques or just has this upregulation and can't downregulate, then it's the a runaway reaction of immune stuff that's happening in our

1:01:52brain. So what what we're sort of getting at here is this idea that these immune cells might be overactivated >> uh based on some stimuli and that overactivation >> um is actually the driver for the neurodeeneration. Yeah, it could be one of the drivers >> one one of the drivers >> um of of the the decline in in cog in cognition because now >> it's these arms that are going to pick up stuff. >> Yeah. >> Are are are just everywhere doing too much. Doing too much picking up too many people off the street. >> Yeah. And when they're doing too much,

1:02:34they're not doing their original job >> job >> of like cleaning >> cleaning the plaque and like compacting it and all this other kind of stuff. It's like if the janitor [snorts] uh became a vigilante at night at the building and so instead of doing the normal janitor stuff. >> Yeah. Now the building the building's falling apart while the janitor is going to be a vigilante. >> Yeah. Yeah. And he's not good at vigilanteism either. It's like bro, [laughter] >> you know. Um so so they had a bunch of different methodologies that they used um which was which was quite cool. They used a mouse model and the mouse model um mimics the the adult Alzheimer's. They also have um this crelocks recombination system which what you can do is you can induce gene deletion four

1:03:15to six weeks after they're born right usually you imagine with gene deletion the deletion needs to happen and then you create the life form >> but then you have problems because like what if the development is perturbed >> right then you've created this defective life form you're making conclusions and you don't know if it's the development or the actual gene that's happening in the adult population. So here this is a way to delay um deleting a gene and then it ensures that any phenotypes that are observed and anything that we observe in the experiment is due to a loss of gene function in the adult deceased brain because Alzheimer's is the adult >> it's not it's not something that you

1:03:56know it's this gene is doing something bad once you're getting old right and so that's what we want to mimic. So, so they were able to do that. They also have this um they the other really cool thing that they could do was [snorts] something called trap which is translating ribosome affinity purification. The idea is I want to prove that the the transcripts the mRNA that's that's getting translated in the microg GA like so the instructions from the DNA into making protein those instructions that the microg ga is paying attention to it's coming from the actual microg ga and not because the microg ga is like let's say eating

1:04:37another immune cell right because if the microglea eats another immune cell well that immune cell is going to have its own >> mRNA And when I sequence, I don't know if this mRNA came from me doing my own thing or me eating something else and then that food is what's getting transcribed, right? So, how do we how do we make that distinction? There's this incredible um way of doing it called GFP tagging where you take this green fluorescent protein, you tag a ribosome with it. Okay? So the ribosome has attached this green fluorescent protein and now the ribosome is going along and it's transcribing your mRNA. You take that you take those cells in that culture

1:05:18and you have another part that attaches to the GFP that green fluorescent protein but that thing is attached to a magnetic bead. >> Okay. >> Okay. And now with a magnetic field, I can rip >> only the ribosomes from the from the microglea, right? Cuz the magnetic bead is something I can use a force on. >> Right. Right. Right. Right. [laughter] >> And then and and then and then I have and the ribosome is going to have an MRA NA attached, right? Cuz it's in the middle of doing some job. Now all of a sudden you like pick it up. So it's like dragging the mRNA with it. And then you're like, "Okay, now I'm going to I'm going to sequence this mRNA." That gives

1:05:58me an insight into the actual transcriptto of the of the immune cell rather than all of the mRNA in the immune cell. That makes sense. Right. I'm actively looking at only the mRNA that is being transcribed into protein that's actually being >> put >> Yes. >> through this machinery. >> Yes. Yes. >> Right. I thought that's a really cool technique. I I I I just [laughter] >> who the the way that folks think about this >> um as like the solution to the problem >> because like it it makes sense you want to be able to know what is actively being translated. >> Yeah. >> Yes. >> And so you need to be able to track like in order to be able to validate like

1:06:40what is your process by which you can actually do validation and it's a very clever way. >> It's very clever. It's like just like yeah basically tag the factory >> right >> that's making the stuff >> right and [snorts] so any problem >> and then and then drag the factory out right >> and then see what's inside the factory >> right so if you want only the cars in that particular >> like let's say city and you don't want all the cars that are >> right >> being ridden around by the by the general population you just tag the factory drag out the factory >> factory no that yeah that's that's really good >> you know >> that's really good >> it's it it It was It was very cool. So, um, now let's get into >> It's funny they call it trap, too.

1:07:21>> Yeah, they [laughter] they literally call it trap. Yeah. Yeah. Um, a lot of times with these acronyms, they come they I'm pretty sure they start with the acronym and then they're like, "Okay, what are the words >> that like can make it happen?" >> I like trap. Yeah. We're >> And then they put down Okay. In the lab meaning translating. Okay. Ribosome. There's an R. [laughter] >> Yeah. Exactly. So funny. Um so um let let's let's get into what they actually found. Okay. >> Okay. They focused on a transcription factor called PU1. Okay. A transcription factor is basically something that

1:08:03gets attached to the DNA and promotes RNA polymerase which is the factory that creates mRNA to actually transcribe a gene. Okay. So this is how we get from gene to mRNA to protein. That first step, you need something called a transcription factor to get where that DNA is, bring in an RNA polymerase and be like, all right, you need to start here, look for that start code on, and then and then start transcribing. >> The transcription factor is like identifying like where the things need to happen, like where the the polymerase needs to come in to do stuff. >> Yes. And each gene usually has different transcription factors. So this particular one, this gene S spi1

1:08:44has a transcription factor PU1. Okay? And different genes have different transcription factors. These transcription factors have different affinities, right? There's some that'll bind immediately. So even at low concentration, it's going to find that DN that bit of DNA and bind. Those are re those are usually the housekeeping genes that you can't live without. Okay? Those are the ones that like it's like how do I how do I make sure that I stay alive as a cell? Okay. Then there's stuff that have to do with function specific states like this PU1 gene is is the the one that is controlling whether microglea glo are chilling or if they're doing the vigilante stuff right and what they found was that when microglea were

1:09:26in the plaque environment that plaque environment actually enforces low PU1. >> Okay. It's very counterintuitive. Okay, >> you would think that, >> you know, I I want like super a lot of inflammation and a lot of policing >> around my plaques. That's not what's happening. >> What's happening is near the plaques, there's a down reggulation of PU1 of that transcription factor, which is already a first clue that perhaps we don't fully understand >> this this ecosystem of what's happening, right? Mhm. [snorts] >> So then they actually used um they used a bunch of tools to figure out what is

1:10:06the signal transduction pathway that goes from I sense a myoid, I sense my plaques to I downregulate PU1. And they used some knockout versions of mice to actually link all of these signal transduction pathways together. >> They also had they they also showed that there is an upregulation of a T- cell associated gene. >> Okay. Okay. In those low PU1 populations, >> so so when when there's low PU, right, right, there's this >> uh there's a T- cell G there's a T gene that now >> more frequently uh arises or or is um or

1:10:48pr proliferates because of that low population environment. >> Exactly. And and what what what they think is happening is when you don't have that transcription factor or when you don't have a lot of that transcription factor, the chromatin around that that that part sort of relaxes and it exposes some of the genes. And one of these genes that it exposes is CD28, which is a neuroprotective master switch. You might recognize CD28 from our Nobel Prize videos. I was literally going to say, >> okay, CD28 is the gene that sort of regulates TE-C cells from becoming normal TE- cells to T- regulatory cells, right? So >> that same gene, that same part of the

1:11:29gene is also being having a role in microglea in the brain. >> That's actually crazy. So it's like the brain is the these cells are borrowing a motif from innate immunity elsewhere in the body and saying well I can actually use this for my own gains over here >> right right for this this this function which which ultimately sort of generates this new protective master switch. So, and so, so the idea is because if we come back and because I just want to make sure I'm tracking here. Yeah. So, we have this we have this mechanism by which >> the [snorts] the micro ga is we we don't want it when

1:12:11it's close to these plaque regions. >> Uh it is less active. >> Yeah. It's the the P1 >> the P1 is less is less active. there's not enough PU1 in the cell, >> right? And that because of that that induces >> Yeah. Then the chromatin sort of relaxes >> and now parts of the DNA get exposed and one of the parts that gets exposed is the CD28 and and that and that one is sort of that this is that neuroprotective master switch which is so like we can sort it's like the Tylenol button. It's like go fix and make all my problems go away, right? And it and because of that low PU it is what helps to release more of that. Yeah. >> And so

1:12:51what you don't want to happen is that process to get >> uh prevented because that is potentially then what is causing the neurodeeneration. >> Exactly. >> Because we're not basically and you know releasing the Tylenol response. >> Yep. Yeah. And so exactly the that master switch is the switch that is controlling the sulfate of these micro GA that's turning it vigilante janitor. >> Yeah. That's okay. you know, and and so and so if we can control that and we can have therapies that now now that we understand this entire chain of of all of the stuff that's happening, we can actually, you know, maybe do something, right? >> Right. And and so these micro ga are

1:13:33now, you know, they're calling it the T-regs of the brain. >> T-Rex being again the thing that won these three