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What I want to talk to you today
about is the creation of

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3-dimensional structures.
And I want to emphasize their

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connection to the function of the
organisms and the organ.

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Let's look, for example,
at the kidney.  This is a fantastic

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example of structure- function
relationships.

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The kidney comprises thousand of
tubules that are involved in

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filtering the blood and collecting
urine for excretion.

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Now, not only do these tubules
function to transport the urine as

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it is being processed,
they also are connected very closely

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to various cell types that are
involved in this filtration

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process.
So the 3-dimensional structure of

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the kidney is integral,
is required for its normal function.

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And this is true in essentially
every organ.  So it's very important

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to think about the relationship of
different cells to one another in a

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particular organ.
And if you're thinking about,

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for example, using tissue
engineering to develop an artificial

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kidney and making some kind of
pastiche of cells and polymers and

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putting that together in
some functional way --

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-- it's important to understand
these structure-function

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relationships in the normal organ
before you try to engineer something

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that is a surrogate.
Individual cells also have

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particular shapes,
and this is integral to their

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function.  We'll talk a bunch about
neurons, cells that have very long

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processes that transmit nervous
impulses or transmit electrical

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impulses and communicate
in that way.

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The shape of the neuron,
the structure, the 3-dimensional

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structure is absolutely required for
its function.  Tom,

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I think we could have a little more
light at the back there.

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It looks really dark if you guys
are trying to take notes.

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Thank you.  OK.  We have,
you've seen this movie previously.

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OK.  This is the zebra fish.  The
zebra fish embryo.

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Isn't it dark at the back?
Yeah.  Tom?  Oh, it takes a while.

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It takes a while.  OK.
I want to show you this movie again

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that you've seen a while.
You have it on your website.

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And I want to show it to you now in
a different way.

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We're actually going to first show
that during the early development of

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the fish, these two cells on top
initially give rise to a ball of

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cells sitting on top of this yoke
cell that is just a ball of cells.

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And then suddenly these cells start
to move during the gastrula and then

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the neurula phases of development.
And it's this cell movement that

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builds structure.
And I want to try to address with

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you how this cell movement occurs
and how it builds structure.

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So let's look at this movie again.
Here we go.  Cell division,

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building up the raw materials to
make the embryo.

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A group of cells sitting on top of
the yolk.  And now watch here.

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It starts to move.  Those cells
have made the decision to move,

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and they going to move towards this
one dorsal side of the embryo.

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And they are going to move and move
some more to build the eye,

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the brain and the various somites
along the length of the axis.

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OK.  So that process, the
generation of 3-dimensional

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structure is a very large part of
the process of building the embryo.

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And it's interdigitated with making
the cell types.

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A muscle cell is not a muscle cell
and it's not functional unless it is

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a fused myotube that has the
appropriate 3-dimensional

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structure.
So cell type and 3-dimensional

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structure are very closely related.
OK.  So let's talk about a toolkit

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that is involved in this process.
And the first thing that we can

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discuss is what is in this toolkit.
Well, actually, you don't really

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have much in the toolkit
to build an organism.

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I can think of one thing.
What's in your toolkit to build

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your 3-dimensional organism?
Yes.  Did I hear cells?  Yeah.

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I hope I heard cells.  Well, there
are cells.  And then if you want to

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look for something else to build the
organism with there are cells and

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then there are cells.
That's what you've got.

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In your toolkit you've got cells.
And the challenge of the organism is

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to use that singular building
material to build the various and

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the huge array of structures that
there are.  So what

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about these cells?

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And the first thing about these
cells is that they come in two forms.

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They come as so-called epithelia or
epithelial sheets.

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And they come as single cells which
are called mesenchyme.

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And these two types of cells
interconvert with one another.

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And it's from these two groups of

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cells, that can be different cell
types, but they're either sheets or

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they're single cells,
that one builds the organism.

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So the epithelia are sheets, the
mesenchyme are single cells.

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And let me look at the next diagram
that I drew for you.

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And you have it in front of you.
If you don't have the handout, does

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anyone not have the handout?
Could I have, Lesley, could you

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[UNINTELLIGIBLE] please?
Thanks a lot.  OK.  If you look at

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the handout, you have this except
for one thing I added this morning.

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Here is a sheet of cells.  So I'm
going to do minimal board work today

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because you have a lot of the
information right in front of you.

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You have a sheet of cells here.
And this sheet of cells called an

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epithelium is a sheet of cells
because it is joined together very

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tightly.  And the cells are joined
together by various junctions that

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I've shown here in yellow or that
I've shown in blue.

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And this sheet of cells,
as Professor Jacks told you,

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has two part, has two sides, an
apical side and a basal side.

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OK?  Remember from cell biology?
And it touches something called the

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basement membrane or the basement
lamina, which is part of the

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extracellular matrix that I'll
mention again later.

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Now, this epithelium,
this sheet of cells can transform

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into single cells.
And these single cells have the

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property of being non-adherent.
And they don't attach very tightly

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to the extracellular matrix,
although they are in contact with it.

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And in order to go from an
epithelium to a mesenchymal state

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there are changes in gene expression
that take place,

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we know, at the transcriptional
level.  And these result in changes

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in cell adhesion and changes in the
organization of the cytoskeleton.

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This is a reversible process,
and mesenchyme can go and turn back

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into epithelium.
Now, the epithelial sheet is a very

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important part of the body,
both as a building block for various

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structures, but also as a barrier.
So your skin is a barrier because

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the cells in it are very tightly
joined together and they form an

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impermeable barrier.
But that's true of essentially every

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organ you have.
Every organ you have is surrounded

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by an epithelium,
or one or more epithelia,

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and these surface barriers so that
stuff in the organ doesn't get out,

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stuff outside the organ doesn't get
in.  And if there is a lesion,

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a break in this epithelium it is a
big deal.  And that is wound.

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That's what a wound is.  It's a
break in the epithelium.

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And there is a very rapid and
profound system of wound healing to

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repair these epithelial sheets.
Now, during development there are

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many transitions from epithelium to
mesenchyme.  So the term that you

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really need to know is abbreviated
EMT.  Not to be confused

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with other EMTs.
This refers to the epithelial

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mesenchymal transition,
which I did not write on your

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handout so I will write it here.

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The epithelial mesenchymal
transition.  And it's always said

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that way, even if it's actually a
mesenchymal epithelial transition.

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One of the most profound examples of
epithelial mesenchymal transitions

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is during the formation of the
nervous system.

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Your central nervous system,
as we'll discuss in a few lectures,

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forms from a tube that rolls up
during development.

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And this tube is an epithelial
sheet.  As the tube is rolling up,

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a group of cells, shown here in
yellow, moves away,

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breaks away from the tube and
undergoes an epithelial to

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mesenchymal transition.
This group of cells is called the

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neural crest cell population,
and these neural crest cells then

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migrate away from the neural tube
and go and set up the entire

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peripheral nervous system.
Not eripheral nerves, peripheral

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nerves.  They also make all the
pigment cells in the body and the

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adrenal medulla.
This epithelial mesenchymal

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transition is not only crucial
during development.

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It's also believed now to be crucial
for metastasis of tumors during

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progression of cancer.
And Professor Jacks will address

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that later in the course.
This is a movie demonstrating the

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migration of the neural crest cells
out from the neural tube which is in

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the middle here,
this white tube.  And these little

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dots migrating out are shown over a
period of about 12 hours.

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The single cells migrating out from
the neural tube as they have

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undergone that epithelial
mesenchymal transition.

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All right.  So I wasn't wrong in
facetiously saying what's in your

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toolkit is cells.
That's what's in your toolkit.

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But clearly the cells are slightly
different from one another or

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profoundly different from one
another in their disposition.

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And we'll talk about what
mesenchyme and what epithelial can

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do in a moment.
The other thing that's different or

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the other thing that's important
that makes cells actually very good

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for building is that they are
plastic.  So let's go through a few

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things that they have going for them.
I want to talk about adhesion.

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I'm going to mention junctions,
I'm going to mention cell sorting,

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and I'm going to mention the
extracellular matrix.

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And some of this stuff you've had
before so I'm going to go through it

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quickly.  This is a diagram that's
on your handout,

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it's from your book,
to indicate that there are many ways

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epithelial sheets are stuck together
that involve very tight apposition

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of the cell membranes,
or in the case of tight junctions or

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slightly less tight apposition in
the case of these things called

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desmosomes.  You can go back and
remind yourselves.

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We mentioned these previously.
The most important thing is you

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understand that cells
are joined together.

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In this micrograph,
to demonstrate how tightly cells are

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joined together,
this is a sheet of cells where the

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nuclei are stained green and the red
is staining for a particular

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specific protein that is found in a
kind of junction called a tight

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junction.  And these tight junctions
outline the cells.

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In other words, the cells are glued
together very tightly.

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Here's another one.
Cell sorting.  There is another kind

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of cell adhesion interaction that's
very important for cell adhesion.

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I've indicated here something
called cadherins.

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Cadherins are interesting.
They're calcium-dependent adhesion

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molecules.  And if you have sharp
eyes you'll see up here something

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that's got a beta and then there's a
word here catenin.

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Remember beta-catenin from
dorsal-ventral axis formation?

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This is the same beta-catenin.
Not only is it a transcription

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factor, it is also involved in cell
adhesion.  There's an interesting

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complication of biology for you,
but I don't want to dwell on that.

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Cadherins are essential for
sticking cells together.

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These are pictures of frog embryos.
This is a normal embryo that's been

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cut open and the cells remain a
tight mass.  However,

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if you take an embryo and you inject
it with inhibitors of cadherin

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function the cells become completely
loose from one another and you can

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actually see the outlines of the
cells because they are no

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longer stuck together.
Now, this is fascinating and very

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important for the animal because it
turns out there are lots of

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different adhesion molecules.
And different cells types sort out

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according to the adhesion molecules
they are expressing on their cell

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surfaces.  So this is a rendition of
a fantastic old experiment that was

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done in the 1950s to demonstrate how
cells sort out.

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What was done,
these are two frog embryos,

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and a piece of the future skin or
epidermis was removed from one,

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and then a piece of the future
neural plate, the nervous system was

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removed from another.
And these had different colors so

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you could tell which cells were
which.  Now, if you take those cells

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and you put them in a medium that
does not contain calcium all the

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cadherins and other adhesion
molecules cannot work.

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And these cells fall into a pile of
single cells.  And you can take your

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pipette and mix them up in your
little dish, and you get this salt

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and pepper arrangement of the two
kinds of cells.

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And then you can add a little bit
of calcium back to the medium,

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and the cells will form this big
ball of cells.

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And it's salt and pepper again.
The two neural plate cells and the

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epidermal cells are mixed up.
But if you go away and have dinner

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or have a good night's sleep and
come back the next day and look at

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your ball of cells,
you see that amazingly the cells

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have sorted themselves out.
The epidermal cells have gone back

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together and the neural cells have
gone back together with one another.

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And you can do this with cells from
almost any organ.

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You can mix cells from two organs
particularly when they're embryonic

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organs.  You can mix them together,
but also in the adults to some

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extent.
And these cells from different

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organs will sort out.
And they sort out because of

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specific adhesive molecules that
they have.  And the term that one

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uses for this is homotypic binding
where cells will interact with one

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another through membrane-bound
receptors.  Again,

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I'm assuming you remember this is a
lipid bilayer with a protein

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sticking through.
Two receptors,

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proteins sticking through the lipid
bilayer interacting with one another

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in calcium-dependent or independent
ways.  And there may be more than

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00:17:10,000 --> 00:17:14,000
one receptor that mediates
cell-specific interactions.

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00:17:14,000 --> 00:17:18,000
But these interactions keep
different cells separate from one

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00:17:18,000 --> 00:17:22,000
another and facilitate development
and building structure.

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00:17:22,000 --> 00:17:26,000
OK.  Here's another one.
The extracellular matrix.

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00:17:26,000 --> 00:17:30,000
What is the extracellular matrix?
We mentioned this at the beginning

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00:17:30,000 --> 00:17:34,000
of the course.
Professor Jacks threw it at you in

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00:17:34,000 --> 00:17:38,000
cell biology.  The extracellular
matrix refers to the stuff on which

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00:17:38,000 --> 00:17:42,000
the cells sit,
so cells in your body are not

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00:17:42,000 --> 00:17:47,000
sitting on nothing.
An epithelium is not just floating

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00:17:47,000 --> 00:17:51,000
free or a tube is not floating free
in liquid.  It is actually

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00:17:51,000 --> 00:17:55,000
surrounded by a bunch of proteins
and carbohydrates that are secreted

233
00:17:55,000 --> 00:18:00,000
by other cells and form the
extracellular matrix.

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00:18:00,000 --> 00:18:04,000
Or, in the case of epithelia,
the basement membrane.  OK?  That's

235
00:18:04,000 --> 00:18:09,000
what the extracellular matrix is
called.  It's highly organized in

236
00:18:09,000 --> 00:18:14,000
the case of the epithelium.
And it consists of proteins and

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00:18:14,000 --> 00:18:19,000
various things called proteoglycans
which are sugars bound to proteins.

238
00:18:19,000 --> 00:18:24,000
Now, one of the proteins in the
extracellular matrix is collagen.

239
00:18:24,000 --> 00:18:29,000
And collagen is the most abundant
protein in the Animal Kingdom.  OK?

240
00:18:29,000 --> 00:18:33,000
It comprises a very high percentage
of your body mass.

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00:18:33,000 --> 00:18:37,000
And if collagen, if there are
mutations in collagen many things

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00:18:37,000 --> 00:18:41,000
can go wrong.  For example,
there is a disorder called

243
00:18:41,000 --> 00:18:45,000
osteogenesis imperfecta where your
bones don't form properly.

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00:18:45,000 --> 00:18:49,000
That's a mutation in one of the
collagen genes.

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00:18:49,000 --> 00:18:53,000
There are a lot of collagen genes,
and the protein mass of collagen is

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00:18:53,000 --> 00:18:57,000
enormous.  Now,
the cells are sitting on the

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00:18:57,000 --> 00:19:01,000
extracellular matrix.
But there is also a fantastic,

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00:19:01,000 --> 00:19:05,000
and then that's good, that's helpful
for the cells in a support sense.

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00:19:05,000 --> 00:19:09,000
It gives them some support.  But
the extracellular matrix does a lot

250
00:19:09,000 --> 00:19:12,000
more than that.
It actually communicates to the

251
00:19:12,000 --> 00:19:16,000
cells.  And it does so by means of
adapter proteins.

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00:19:16,000 --> 00:19:20,000
You have this as a handout,
if you didn't realize.  This is one

253
00:19:20,000 --> 00:19:23,000
of your handout diagrams.
OK?  These adaptor proteins have

254
00:19:23,000 --> 00:19:27,000
the property of being transmembrane
or at least integral membrane

255
00:19:27,000 --> 00:19:31,000
proteins that are attached
to the cells.

256
00:19:31,000 --> 00:19:35,000
But they also attach to proteins in
the extracellular matrix.

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00:19:35,000 --> 00:19:40,000
And this allows them to sense what
the extracellular matrix contains

258
00:19:40,000 --> 00:19:45,000
and to transmit that information to
the cells.  Because the thing,

259
00:19:45,000 --> 00:19:50,000
of course, about living organisms
and the 3-dimensional structures in

260
00:19:50,000 --> 00:19:55,000
them is that the process of both
building the structure and

261
00:19:55,000 --> 00:20:00,000
maintaining the structure
is a dynamic one.

262
00:20:00,000 --> 00:20:04,000
It's not equivalent to building
Building 10 or Stata.

263
00:20:04,000 --> 00:20:08,000
It's not equivalent to putting
components together and getting a

264
00:20:08,000 --> 00:20:12,000
structure.  You get the structure,
and the structure is maintained

265
00:20:12,000 --> 00:20:16,000
because the structure senses how
it's doing, whether it's intact or

266
00:20:16,000 --> 00:20:20,000
not.  If one of the walls in this
building fell down,

267
00:20:20,000 --> 00:20:24,000
it would fall down until someone
repaired it.  If one of the tubes in

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00:20:24,000 --> 00:20:28,000
your body gets a hole in it,
your body will sense it and try to

269
00:20:28,000 --> 00:20:33,000
repair it.
That same thing is true when the

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00:20:33,000 --> 00:20:38,000
epithelia, and I'll tell you in a
moment, when mesenchymal cells are

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00:20:38,000 --> 00:20:43,000
actually doing their building
process, they are sensing what is

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00:20:43,000 --> 00:20:48,000
around them.  And they do it through
these adapter proteins.

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00:20:48,000 --> 00:20:53,000
The adaptor proteins are receptors,
by definition, they're binding to

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00:20:53,000 --> 00:20:58,000
something in the ECM.
And a large number of them comprise

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00:20:58,000 --> 00:21:04,000
of class of proteins called
integrins, as an example of a name.

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00:21:04,000 --> 00:21:10,000
All right.  So let's move on here
and let's mention --

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00:21:10,000 --> 00:21:21,000
-- the cytoskeleton as being

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00:21:21,000 --> 00:21:32,000
required for shape and movement.

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00:21:32,000 --> 00:21:35,000
We've mentioned the cytoskeleton
before.  I'll talk about it more in

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00:21:35,000 --> 00:21:39,000
a moment.  This is a diagram from
your book that you had before.

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00:21:39,000 --> 00:21:42,000
The cell is not a floppy bag of
liquid.  It contains many filaments

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00:21:42,000 --> 00:21:46,000
that keep it rigid,
that allow it to have particular

283
00:21:46,000 --> 00:21:49,000
shapes and that allow it to change
its shape.  We're going to be

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00:21:49,000 --> 00:21:53,000
talking most about microfilaments
which comprise polymers of actin,

285
00:21:53,000 --> 00:21:56,000
the protein actin.  There are also
intermediate filaments and

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00:21:56,000 --> 00:22:00,000
microtubules which we mentioned last
lecture when we talked about

287
00:22:00,000 --> 00:22:04,000
the sperm flagellum.
This is a micrograph of tube stained,

288
00:22:04,000 --> 00:22:08,000
of cells stained for the microtubule
network.  The nucleus is yellow.

289
00:22:08,000 --> 00:22:12,000
Stained for one of the major
proteins in the microtubules,

290
00:22:12,000 --> 00:22:17,000
which is tubulin.  And you can see
this very extensive meshwork

291
00:22:17,000 --> 00:22:21,000
throughout these cells.
And the same is true for actin.

292
00:22:21,000 --> 00:22:25,000
The same is true for intermediate
filaments.  So cells are very well

293
00:22:25,000 --> 00:22:30,000
supported by these filaments.  OK.
And, finally, let me mention cell

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00:22:30,000 --> 00:22:35,000
division and cell death as being
part of the toolkit that allows one

295
00:22:35,000 --> 00:22:39,000
to build structure.
And I'll show you one experiment

296
00:22:39,000 --> 00:22:44,000
which involves the inhibition of
cell death in the embryo.

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00:22:44,000 --> 00:22:49,000
This is a mouse, normal mouse
embryo, and this is a mouse embryo

298
00:22:49,000 --> 00:22:53,000
in which a protein was removed.
This protein is called caspase-9.

299
00:22:53,000 --> 00:22:58,000
And caspases are involved in
killing cells during development,

300
00:22:58,000 --> 00:23:03,000
as Professor Jacks mentioned to you.
In this mutant animal,

301
00:23:03,000 --> 00:23:07,000
one of the things you should notice
is that the brain is hugely

302
00:23:07,000 --> 00:23:11,000
overgrown.  And this is an
indication of the amount of cell

303
00:23:11,000 --> 00:23:15,000
death that has to happen during
normal brain development.

304
00:23:15,000 --> 00:23:19,000
OK?  And the flip side I'm not
going to dwell on.

305
00:23:19,000 --> 00:23:23,000
You need to get not only the normal
amount of cell proliferation,

306
00:23:23,000 --> 00:23:27,000
but it needs to be in the right
place.  All right.

307
00:23:27,000 --> 00:23:31,000
So let's talk about the behavior of
cells and how this works together to

308
00:23:31,000 --> 00:23:36,000
give 3-dimensional structures.
And I want to talk very briefly

309
00:23:36,000 --> 00:23:40,000
about, actually,
not so briefly, about the behavior

310
00:23:40,000 --> 00:23:48,000
of single cells --

311
00:23:48,000 --> 00:23:53,000
-- and the property that is most
important to them,

312
00:23:53,000 --> 00:23:58,000
which is the ability to move.
So the point about an epithelial

313
00:23:58,000 --> 00:24:03,000
mesenchymal transition is that what
comes out of it are single cells.

314
00:24:03,000 --> 00:24:08,000
And this is important in metastasis
and in normal development.

315
00:24:08,000 --> 00:24:13,000
The difference between this
epithelium and these single cells is

316
00:24:13,000 --> 00:24:18,000
that the single cells are free to
move because they are not attached

317
00:24:18,000 --> 00:24:23,000
to a sheet.  And that is how cells
get from one place to another in the

318
00:24:23,000 --> 00:24:29,000
body.  OK?  So let's dwell on this
in more detail.

319
00:24:29,000 --> 00:24:33,000
So how do cells know where they're
going?  Well, this is a very

320
00:24:33,000 --> 00:24:37,000
interesting question.
They know where they're going

321
00:24:37,000 --> 00:24:41,000
because they're told to go somewhere.
So I told you the neural crest

322
00:24:41,000 --> 00:24:45,000
migrating out of the neural tube.
This is a group of cells from an

323
00:24:45,000 --> 00:24:49,000
organism called dictyostelium.
And these are single cells.

324
00:24:49,000 --> 00:24:53,000
They've been fluorescently labeled.
And up here in the corner someone

325
00:24:53,000 --> 00:24:58,000
has put invisibly a drop of the
second messenger cyclic AMP.

326
00:24:58,000 --> 00:25:03,000
And you can see these cells that had
been milling around randomly start

327
00:25:03,000 --> 00:25:08,000
to realize this and move as a
consorted group towards the source

328
00:25:08,000 --> 00:25:13,000
of cyclic AMP.
OK?  And this is one of the ways a

329
00:25:13,000 --> 00:25:18,000
dictyostelium organizes itself.
The cells chemotax, they follow a

330
00:25:18,000 --> 00:25:23,000
particular chemical stimulus.
How do they move?  How do cells

331
00:25:23,000 --> 00:25:28,000
move?
Well, they move because the

332
00:25:28,000 --> 00:25:32,000
reorganize their cytoskeleton,
particularly their actin

333
00:25:32,000 --> 00:25:36,000
cytoskeleton in a concerted way.
So I've drawn for you a kind of

334
00:25:36,000 --> 00:25:40,000
animated cartoon.
You have the whole thing in front

335
00:25:40,000 --> 00:25:45,000
of you.  And I'll animate it and see
how it goes.  We start off here with

336
00:25:45,000 --> 00:25:49,000
our green cell.
This is a mesenchymal cell.

337
00:25:49,000 --> 00:25:53,000
And it is sitting on some kind of
extracellular matrix,

338
00:25:53,000 --> 00:25:57,000
which I've shown as kind of
organized, but actually it may not

339
00:25:57,000 --> 00:26:02,000
be.  But that doesn't matter.
It is loosely attached to this

340
00:26:02,000 --> 00:26:06,000
extracellular matrix.
It's got these receptors sticking

341
00:26:06,000 --> 00:26:11,000
out of its membrane that can sense
extracellular matrix,

342
00:26:11,000 --> 00:26:15,000
but only at the rear of the cell is
it actually attached by these red

343
00:26:15,000 --> 00:26:20,000
triangles to the ECM.
OK?  That's what my red triangles

344
00:26:20,000 --> 00:26:24,000
indicate.  Now,
the cell suddenly as it's sitting

345
00:26:24,000 --> 00:26:29,000
there or moving randomly along comes
across these black circles --

346
00:26:29,000 --> 00:26:34,000
-- which are some kind of molecule
that can interact with these

347
00:26:34,000 --> 00:26:40,000
receptors and tell it that it ought
to go in a particular direction.

348
00:26:40,000 --> 00:26:46,000
So how does it do this?  Well, the
first thing it does it to elicit

349
00:26:46,000 --> 00:26:52,000
this interaction with a substrate or
with these black molecules in the

350
00:26:52,000 --> 00:26:58,000
substrate.  This is a receptor
ligand interaction.

351
00:26:58,000 --> 00:27:02,000
It's exactly the same kind of
interaction that you talked about in

352
00:27:02,000 --> 00:27:07,000
cell biology, that we talked about
in the formation,

353
00:27:07,000 --> 00:27:11,000
earlier formation lectures,
cell receptor ligand interaction.

354
00:27:11,000 --> 00:27:16,000
And it does the same thing that
other receptor ligand interactions

355
00:27:16,000 --> 00:27:20,000
do.  It activates some kind of
signal transduction process,

356
00:27:20,000 --> 00:27:25,000
but this signal transduction process
has the property of telling the

357
00:27:25,000 --> 00:27:29,000
actin filaments in this local region
of the cell that they

358
00:27:29,000 --> 00:27:33,000
should polymerize.
OK?  So this is cell signaling in a

359
00:27:33,000 --> 00:27:37,000
very local region of the cell.
And I'll show you the movie in a

360
00:27:37,000 --> 00:27:41,000
moment.  You'll see it's
extraordinary.

361
00:27:41,000 --> 00:27:45,000
So here are the actin filaments
forming in this region of the cell.

362
00:27:45,000 --> 00:27:49,000
And I've called this the front of
the cell. OK?  It's an arbitrary

363
00:27:49,000 --> 00:27:53,000
term.  But the front of the cell is
arbitrary but it's going to refer to

364
00:27:53,000 --> 00:27:57,000
the direction that the cell is
moving.  So there's actin

365
00:27:57,000 --> 00:28:01,000
polymerization of the front.
And the cell also sends out some

366
00:28:01,000 --> 00:28:06,000
protrusions called filopodia or
lamellipodia.  And it sends out

367
00:28:06,000 --> 00:28:11,000
these protrusions to try to sense
where it should move next.

368
00:28:11,000 --> 00:28:16,000
Now, once it's done that, once it's
made these polymerized actin

369
00:28:16,000 --> 00:28:21,000
filaments it contracts.
However, it doesn't go anywhere

370
00:28:21,000 --> 00:28:26,000
because it's still attached at the
rear.  So it's kind of like pushing

371
00:28:26,000 --> 00:28:32,000
down on your accelerator when your
hand break is still engaged.

372
00:28:32,000 --> 00:28:36,000
OK?  You don't really go very far.
So the next thing it has to do is

373
00:28:36,000 --> 00:28:40,000
to lose the adhesion at the rear.
I couldn't get this to work but it

374
00:28:40,000 --> 00:28:44,000
will work in a moment.
OK.  And it also depolymerizes the

375
00:28:44,000 --> 00:28:48,000
actin at the rear.
So there.  There we go.

376
00:28:48,000 --> 00:28:52,000
There's loss of the adhesion and
there's loss of the actin at the

377
00:28:52,000 --> 00:28:56,000
rear.  And now it's in a position,
it's disengaged the hand break and

378
00:28:56,000 --> 00:29:00,000
it can move forward, so
it goes forward.  OK.

379
00:29:00,000 --> 00:29:04,000
This is my cartoon.
This is the real thing.

380
00:29:04,000 --> 00:29:08,000
This is a cell where actin has been
labeled fluorescently.

381
00:29:08,000 --> 00:29:12,000
And I want you to watch two things.
It's on a repeat loop so we'll

382
00:29:12,000 --> 00:29:16,000
watch it a number of times.
At the part of the cell that is

383
00:29:16,000 --> 00:29:20,000
sticking out, you should be able to
see these bright cables or these

384
00:29:20,000 --> 00:29:24,000
bright lines.  These are cables of
polymerized actin.

385
00:29:24,000 --> 00:29:28,000
OK?  And the cell is sticking out
these protrusions and

386
00:29:28,000 --> 00:29:32,000
polymerizing actin.
And it's moving down in this plane

387
00:29:32,000 --> 00:29:36,000
of the board, or the screen as it's
making these polymers.

388
00:29:36,000 --> 00:29:40,000
OK?  So this is the front of the
cell.  And then if you look at the

389
00:29:40,000 --> 00:29:44,000
back of the cell,
let's wait for the loop again to

390
00:29:44,000 --> 00:29:48,000
start, and you will see,
here's the back of the cell with

391
00:29:48,000 --> 00:29:52,000
initially polymerized actin.
And if you watch individual lines

392
00:29:52,000 --> 00:29:56,000
you will see them go away as the
actin is depolymerized and the cell

393
00:29:56,000 --> 00:30:00,000
loses its adhesiveness
at the back.  OK?

394
00:30:00,000 --> 00:30:04,000
So this process involves selective
adhesion, loss of adhesion mediated

395
00:30:04,000 --> 00:30:09,000
by actin polymerization through the
cytoskeleton.  There's a signal

396
00:30:09,000 --> 00:30:14,000
transduction pathway involved.
It involves things called GTPases

397
00:30:14,000 --> 00:30:19,000
that you've talked about previously.
And it's really an extraordinary

398
00:30:19,000 --> 00:30:24,000
process.  All right.
But let's move on and let's talk

399
00:30:24,000 --> 00:30:32,000
about cell sheets.

400
00:30:32,000 --> 00:30:36,000
So cell migration is crucial to get
from one place to another,

401
00:30:36,000 --> 00:30:40,000
but you cannot really build much
with single cells because they're

402
00:30:40,000 --> 00:30:44,000
single cells.  They don't really
give you any kind of integral

403
00:30:44,000 --> 00:30:48,000
structure, any kind of cohesive
structure.  And so you have to turn

404
00:30:48,000 --> 00:30:52,000
these single cells back into sheets
if you're going to use them then for

405
00:30:52,000 --> 00:30:56,000
building.  So what do cell sheets do?
Ah, OK.  So let me go through a

406
00:30:56,000 --> 00:31:00,000
couple of things that
cell sheets do.

407
00:31:00,000 --> 00:31:05,000
Cell sheets can get longer.
They can change the number of

408
00:31:05,000 --> 00:31:10,000
layers that they have.
They can change the number of

409
00:31:10,000 --> 00:31:15,000
length, the number of layers that
they have, and they can bend.

410
00:31:15,000 --> 00:31:20,000
Here are some cartoons that I drew
for you.  Here's a sheet of cells.

411
00:31:20,000 --> 00:31:25,000
And you can lengthen it in two ways.
You can stretch it out so that the

412
00:31:25,000 --> 00:31:30,000
surface area to volume
ratio changes.

413
00:31:30,000 --> 00:31:34,000
And you get a longer,
thinner sheet of cells.

414
00:31:34,000 --> 00:31:39,000
You can also take this initially
bilayered sheet of cells and fit the

415
00:31:39,000 --> 00:31:43,000
cells in between one another by a
process called intercalation,

416
00:31:43,000 --> 00:31:48,000
or the term that's usually used is
convergent extension.

417
00:31:48,000 --> 00:31:53,000
And that lengthens a sheet of cells.
And the reason that a small embryo

418
00:31:53,000 --> 00:31:57,000
becomes longer is because of this
process of intercalation where the

419
00:31:57,000 --> 00:32:02,000
embryonic cells interdigitate with
one another and make the whole

420
00:32:02,000 --> 00:32:07,000
embryo stretch out.
You can also change the number of

421
00:32:07,000 --> 00:32:12,000
cell layers to get a more complex
tissue, a more complex structure.

422
00:32:12,000 --> 00:32:17,000
You can break two cell layers into
two individual layers.

423
00:32:17,000 --> 00:32:22,000
That's called delamination.
Or you can turn two cell layers

424
00:32:22,000 --> 00:32:27,000
into one cell layer by another
process of intercalation so that you

425
00:32:27,000 --> 00:32:32,000
get a single sheet from what was
initially a two cell thick sheet.

426
00:32:32,000 --> 00:32:36,000
Again, this is in front of you and I
don't want you to dwell on it.

427
00:32:36,000 --> 00:32:40,000
Let's talk about bending cell
sheets.  This is really interesting

428
00:32:40,000 --> 00:32:44,000
because one of the things that comes
out of bending cell sheets are tubes.

429
00:32:44,000 --> 00:32:49,000
Now, tubes are pervasive.
There is not an organ in your body

430
00:32:49,000 --> 00:32:53,000
that does not have some kind of
tubular structure.

431
00:32:53,000 --> 00:32:57,000
And if you think about it,
if I were to give you a pile of

432
00:32:57,000 --> 00:33:02,000
cells and say, here,
build me a tube.

433
00:33:02,000 --> 00:33:07,000
You could probably come up with
several ways that you build a tube.

434
00:33:07,000 --> 00:33:12,000
And, in fact, the organism has come
up with several ways.

435
00:33:12,000 --> 00:33:17,000
One of the ways is to roll an
epithelial sheet into a tube.

436
00:33:17,000 --> 00:33:22,000
So here is the sheet.  And you roll
it up to form a tube.

437
00:33:22,000 --> 00:33:27,000
This is how your neural tube is
made.  And also part of this is that

438
00:33:27,000 --> 00:33:33,000
the cells change shape as
they are rolling up.

439
00:33:33,000 --> 00:33:37,000
So if you have a sheet of cells that
is either cuboidal or columnar,

440
00:33:37,000 --> 00:33:42,000
to get that sheet of cells to bend
you have to make wedge-shaped cells

441
00:33:42,000 --> 00:33:47,000
out of them.  You have to give them
a constriction,

442
00:33:47,000 --> 00:33:51,000
a so-called apical constriction.
So you go from a sheet of columnar

443
00:33:51,000 --> 00:33:56,000
cells to a bench sheet of cells
because you have apically

444
00:33:56,000 --> 00:34:01,000
constricted those particular cells.
And you can do that through an actin

445
00:34:01,000 --> 00:34:05,000
process or through our old friend
beta-catenin here acting as a cell

446
00:34:05,000 --> 00:34:10,000
adhesion molecule.
All right.  So here's the process

447
00:34:10,000 --> 00:34:14,000
of rolling up the neural tube in
frogs.  It's rolling up by apical

448
00:34:14,000 --> 00:34:18,000
constriction and by the epithelial
sheets bending.

449
00:34:18,000 --> 00:34:23,000
OK?  You've seen this movie
previously, and it is on your

450
00:34:23,000 --> 00:34:27,000
website so you can look at it again.
Here's another way.  You can

451
00:34:27,000 --> 00:34:32,000
balloon out an epithelium.
You can imagine an epithelium,

452
00:34:32,000 --> 00:34:36,000
and you make a little, you blow it
and you get a little balloon.

453
00:34:36,000 --> 00:34:40,000
And then you blow it some more,
you pull it some more and you can

454
00:34:40,000 --> 00:34:44,000
turn an indentation or a very small
vesicle into a long tube.

455
00:34:44,000 --> 00:34:48,000
OK?  And I'll show you.  This is
not the best diagram,

456
00:34:48,000 --> 00:34:52,000
I've got to work on this one,
but the idea is that you pull this.

457
00:34:52,000 --> 00:34:56,000
Dr. Gardel is agreeing with me,
I've got to work on

458
00:34:56,000 --> 00:35:00,000
this diagram.  OK.
But you pull out this sheet of cells

459
00:35:00,000 --> 00:35:04,000
into a long tube.
I'll talk more about this when we

460
00:35:04,000 --> 00:35:08,000
talk about lung formation in a
moment.  Here's another way.

461
00:35:08,000 --> 00:35:12,000
You can take your single
mesenchymal cells that have migrated

462
00:35:12,000 --> 00:35:16,000
to wherever they're going and you
can get them to condense into an

463
00:35:16,000 --> 00:35:20,000
epithelial sheet.
And this epithelial sheet can then

464
00:35:20,000 --> 00:35:24,000
form a tube.  This is exactly what
happens in formation of the blood

465
00:35:24,000 --> 00:35:29,000
vessels.  Your blood vessels have
got lots of layers of epithelia.

466
00:35:29,000 --> 00:35:33,000
The first one to form is the
endothelium.  It's the inner most

467
00:35:33,000 --> 00:35:38,000
layer of the blood vessel.
And it forms by condensation of

468
00:35:38,000 --> 00:35:43,000
mesenchymal cells into an epithelial
sheet.  OK.  Excellent.

469
00:35:43,000 --> 00:35:48,000
So I want to end by giving you a
specific example and talking about

470
00:35:48,000 --> 00:35:52,000
the genes involved in a particular
aspect of 3-dimensional structure

471
00:35:52,000 --> 00:35:57,000
generation.  And the example I've
chosen is formation of the lung and

472
00:35:57,000 --> 00:36:02,000
the tubules in the lung.
So your lungs are made up of

473
00:36:02,000 --> 00:36:06,000
multiple tubules that have got
little sacks on the end of them,

474
00:36:06,000 --> 00:36:11,000
alveoli that are the places where
oxygen is exchanged between the air

475
00:36:11,000 --> 00:36:15,000
you breath in and between the blood
vessels that feed into or that

476
00:36:15,000 --> 00:36:20,000
surround all the cells of the lung.
Formation of the lung tubules is

477
00:36:20,000 --> 00:36:24,000
very interesting because
mathematically,

478
00:36:24,000 --> 00:36:29,000
if you look at it,
you can really define it by a

479
00:36:29,000 --> 00:36:33,000
Mandelbrot set where you have some
kind of reiterative

480
00:36:33,000 --> 00:36:38,000
branching process.
And if you look at the lung tubules

481
00:36:38,000 --> 00:36:42,000
and count them and look at the
number of branching,

482
00:36:42,000 --> 00:36:46,000
you can figure out that you have to
go through 20 branching events to

483
00:36:46,000 --> 00:36:51,000
get to the set of lung tubules that
is in an adult human lung.

484
00:36:51,000 --> 00:36:55,000
OK?  If you look at the structure,
the structures during the process

485
00:36:55,000 --> 00:37:00,000
start with the trachea and this
thing called the bronchial bud --

486
00:37:00,000 --> 00:37:04,000
-- which divides into two to give
two primary bronchi.

487
00:37:04,000 --> 00:37:08,000
Then each of those divide more,
branch more and so on.  So these are

488
00:37:08,000 --> 00:37:12,000
multi-cellular,
these are epithelial sheets that are

489
00:37:12,000 --> 00:37:16,000
branching reiteratively during lung
formation.  Here's a movie of the

490
00:37:16,000 --> 00:37:20,000
process.  You can get this to take
place in a Petri dish with the

491
00:37:20,000 --> 00:37:24,000
appropriate signals added.
And so here is the lung tubule

492
00:37:24,000 --> 00:37:28,000
branching reiteratively over a
period of about a day or

493
00:37:28,000 --> 00:37:32,000
two in tissue culture.
OK?  So this is a 3-dimensional

494
00:37:32,000 --> 00:37:36,000
tissue engineering,
or at least you can get this aspect

495
00:37:36,000 --> 00:37:40,000
of 3-dimensional tissue structure to
form in tissue culture.

496
00:37:40,000 --> 00:37:44,000
All right.  So what genes control
this?  As you know,

497
00:37:44,000 --> 00:37:49,000
as I've told you over and over and
over, everything is controlled by

498
00:37:49,000 --> 00:37:53,000
genes and the interaction of genes.
And what do we know about lung

499
00:37:53,000 --> 00:37:57,000
formation?  Well,
we know most about lung formation

500
00:37:57,000 --> 00:38:02,000
from insects.
In insects the evolutionary relic or

501
00:38:02,000 --> 00:38:06,000
the evolutionary precursor or the
evolutionary equivalent is the

502
00:38:06,000 --> 00:38:10,000
system of trachea.
These are a system of tubes that

503
00:38:10,000 --> 00:38:14,000
branch out from the spiracles,
which are holes that lead to the

504
00:38:14,000 --> 00:38:18,000
outside.  And then tubes come off
these spiracles and branch in the

505
00:38:18,000 --> 00:38:22,000
insect and actually carry air
directly to the tissues.

506
00:38:22,000 --> 00:38:26,000
There's no circulatory system
equivalent as there

507
00:38:26,000 --> 00:38:31,000
is in our cells.
But the way these tubules branch and

508
00:38:31,000 --> 00:38:35,000
form is very equivalent to ourselves.
And we know that this involves a

509
00:38:35,000 --> 00:38:40,000
receptor signaling system that you
are familiar with,

510
00:38:40,000 --> 00:38:44,000
which is a receptor tyrosine kinase
signaling system.

511
00:38:44,000 --> 00:38:48,000
And the particular one I'll tell
you about is the fibroblast growth

512
00:38:48,000 --> 00:38:53,000
factor signaling system.
You remember that growth factors

513
00:38:53,000 --> 00:38:57,000
bind to receptors,
and these growth factors can be in

514
00:38:57,000 --> 00:39:02,000
the extracellular matrix.
They usually are.

515
00:39:02,000 --> 00:39:06,000
That's one of the places where
growth factors are.

516
00:39:06,000 --> 00:39:10,000
They bind to a receptor.
And then there is a cascade of

517
00:39:10,000 --> 00:39:14,000
signal transduction that,
in this case, involves kinases.

518
00:39:14,000 --> 00:39:18,000
And eventually the final kinase
moves to the nucleus where it does

519
00:39:18,000 --> 00:39:22,000
stuff to transcription factors and
you change the transcriptional

520
00:39:22,000 --> 00:39:26,000
activity of a cell and you get stuff
happening.  OK.

521
00:39:26,000 --> 00:39:30,000
So this is a diagram of what
happens in the fruit fly where we

522
00:39:30,000 --> 00:39:35,000
know most about the process.
The epithelium that's going to give

523
00:39:35,000 --> 00:39:40,000
rise to the trachea undergoes cell
division, and at the same time it

524
00:39:40,000 --> 00:39:45,000
balloons out to form this thing
called the primary tubule.

525
00:39:45,000 --> 00:39:50,000
You have this diagram in front of
you.  It forms the primary tubule.

526
00:39:50,000 --> 00:39:55,000
After a bit, this primary tubules
branches to give secondary tubules.

527
00:39:55,000 --> 00:40:00,000
And later on it branches again to
give these tertiary tubules.

528
00:40:00,000 --> 00:40:04,000
There are a number of mutants in
drosophila that affect each of these

529
00:40:04,000 --> 00:40:08,000
processes, the primary tubule
formation secondary or tertiary

530
00:40:08,000 --> 00:40:12,000
tubule formation.
And these are members of the

531
00:40:12,000 --> 00:40:17,000
fibroblast growth factor signaling
system.  In the primary tubule the

532
00:40:17,000 --> 00:40:21,000
FGF receptor is expressed,
and the cells receive FGF which

533
00:40:21,000 --> 00:40:25,000
tells them to divide and to divide
in a particular orientation to give

534
00:40:25,000 --> 00:40:30,000
you this primary tubule
as it grows out.

535
00:40:30,000 --> 00:40:34,000
OK?  And there's a ligand sitting
around in the extracellular matrix

536
00:40:34,000 --> 00:40:39,000
that tells the cells to do this.
Now, what's really cool is that at

537
00:40:39,000 --> 00:40:44,000
the point where the secondary tubule,
I'm going to take 20 seconds,

538
00:40:44,000 --> 00:40:49,000
so I'd appreciate it if you'd listen.
At the tip of the tubule an FGF

539
00:40:49,000 --> 00:40:53,000
inhibitor called Sprouty is made.
This inhibits cell division of

540
00:40:53,000 --> 00:40:58,000
these cells right at the tip,
but the cells on either side

541
00:40:58,000 --> 00:41:03,000
continue to divide.
And so you get the branching,

542
00:41:03,000 --> 00:41:07,000
the secondary branching taking place.
And I'm going to stop there.

543
00:41:07,000 --> 00:41:12,000
And those of you, don't forget,
come along to Stata this afternoon

544
00:41:12,000 --> 00:41:15,000
if you need a review.