So B-cells, as their name suggests,
‘B’, develop in the bone marrow.
So the bone marrow is
where B-cells develop.
And in fact, virtually all the cells of the
immune system start of
from multipotent hematopoietic stem cells in the bone marrow.
And B-cells can become fully
mature within the bone marrow.
They don’t need to go anywhere else to become
mature B-cells, they develop within the bone marrow.
Once they are mature, they can leave the
bone marrow and travel to the secondary
lymphoid tissues; the mucosa-associated lymphoid
tissue, the lymph nodes and the spleen.
So let’s have a look at how B-cell
development in the bone marrow
occurs and the stages in B-cell maturation in the bone marrow.
I’ve already mentioned, they start
off from hematopoietic stem cells.
These develop into pro-B cells.
Those hematopoietic stem cells, that instead
of becoming a mast cell, or instead of becoming
a dendritic cell, those that are going to
become B-cells initially become pro-B cells.
The next stage in their development
is to become a pre-B cell.
Then they develop into immature B-cells and
of course eventually into mature B-cells.
There are two stages in this
development process where these B-cells
undergo very, very extensive cell
division, in other words proliferation.
That’s at a very early stage in their
maturation, at the stem cell, pro-B cell stage.
And then later on, at the pre-B, immature B-cell stage,
they undergo further massive rounds of proliferation.
And they’re dividing all the time, but
these two stages in their development
are characterized by particularly extensive cell proliferation.
There are two genes that are called RAG1 and
RAG2, stands for Recombination Activating Genes.
We already mentioned that the immunoglobulin genes in
the B-cell undergo recombination to create diversity.
These Recombination Activating Genes obviously
play a role in it as their name suggests.
And these genes are also expressed
at two different points in the
development and maturation of B-cells, at the pro-B pre-B stage.
And then they can become re-expressed later on in immature
B-cells, to fine tune the antigen receptor on the B-cell.
There’s another molecule which we’ll hear about shortly
called TdT - Terminal Deoxynucleotidal Transferase.
Bit of a mouthful that isn’t it?
We normally just refer to it as TdT because Terminal
Deoxynucleotidal Transferase is
quite a long word to say, so TdT.
And that’s expressed again at the stage
between pro-B cells and pre-B cells.
If we look at the immunoglobulin gene DNA in the B-cell
and the RNA that’s produced from that DNA. In the stem
cell stage and the pro-B cell stage, these immunoglobulin
genes are what we call, in the germ line configuration.
That means how these genes are inherited through the sperm
and the egg, the germline, and every single cell in the
body will have this DNA, just like you have genes for… all
the different genes you have in all the different cells.
And in all the cells in the body, they will be
in the germline orientational configuration.
And it’s only once cells develop to become B-cells
that these immunoglobulin genes recombine.
And this recombination initially starts off at the
pre-B cell stage, and some segments within the heavy
chain genes for the immunoglobulins… because immunoglobulins
consist of a heavy chain and a light chain.
The first event that says, “Mmm, that cell, that
cell’s going to become a B-cell”, is when the
B-cell recombines its heavy chain genes, and it
recombines three gene segments as we’ll see shortly.
They’re called V, D and J -
Variable, Diversity and Joining.
Once that recombination process has occurred in
the pre-B cell, messenger RNA is produced for
the heavy chain and that includes a segment
that is from the constant region called mu (µ).
So you get µ mRNA in the pre-B cell.
As that cell then develops to the final stage before full
maturity, in other words the immature B-cell, the recombined
heavy chain gene associates the product of that gene,
associates with the product of a recombined light chain gene.
Because in the light chains, the V gene segments and J
gene segments that recombine produce a light chain protein.
So now we have Ig for the DNA that’s recombined
in the heavy chain and the light chain.
We have a µ heavy chain protein
and either a kappa (k) or lambda (λ)
light chain protein produced from the k or λ messenger RNA.
And then finally we end up via alternative
splicing of the V, D and J with the constant region
of the IgM antibody which is encoded by a Cµ
gene or the IgD antibody encoded by a Cδ gene.
So the stage of maturation; early on the immunoglobulin
genes are not recombined, so there’s no expression.
In order to express these genes
they need to be recombined.
They only recombine in B-cells, and they
only recombine at the pre-B cell stage.
So you start off at the pre-B cell stage with the first
indication that this cell is going to become a B-cell.
It has cytoplasmic µ heavy chain
and on the surface it has what is
called the pre-B cell receptor,
which is a surrogate light chain.
It’s not the proper light chain but it’s a sort of intermediate
light chain that’s produced to put
together with the µ heavy chain.
And then once the light chain genes
have been recombined, you get the
full mature antibody molecule on the
surface of this immature B-cell.
So it’s a mature antibody but it’s on the surface
of an immature B-cell, and this is membrane IgM.
And then as well as IgM, another class of antibody, the
IgD class is put on the surface of the mature B-cell.
As well as having antibodies on their
cell surface, B-cells have thousands
of other molecules on their cell
surface just like every cell does.
And we can use certain molecules on the surface of
B-cells to identify which stage in maturation they are.
So for example, the molecule CD34 is
present on the surface of the stem cells.
It’s also present on the
surface of the pro-B cell.
But additional molecules become
expressed, such as CD19 and CD10.
And then as those pre-B cells develop into pre-B cells,
there is low expression of a molecule called B220.
CD43 is still there.
But as that pre-B cell develops into an
immature B-cell, the expression of CD43 is lost.
So if a cell has a molecule on its surface,
we describe it as a CD-whatever positive cell.
If it doesn’t have expression, we describe
it as a CD-whatever negative cell.
So in other words, the stem cells, the pro-B
cells and the pre-B cells are CD43 positive,
whereas the immature B-cell is CD43 negative
because it has lost expression of CD43.
At that stage of the immature B-cell, there is a low level
expression of IgM antibodies on the surface of the B-cell.
And then as the B-cell further matures to become a fully
mature B-cell, the level of expression of IgM increases.
So where are these different types of B-cell,
these different stages of maturation found?
Well the stem cells, the pro-B cells, the
pre-B cells are all within the bone marrow.
And then as these cells develop into immature
B-cells, they can leave the bone marrow and
enter what immunologists refer to as the
periphery; rather strange term that, the periphery.
Simply means anywhere outside
of the primary lymphoid tissues.
So anywhere in the body that isn’t the bone marrow or
the thymus, immunologists refer to as the periphery.
And the mature B-cells are found in the
periphery, or move to the periphery.
The response to antigen; clearly if the
B-cell doesn’t have an antigen receptor
on its surface, it can’t respond, it
has no way of responding to antigen.
So because there is no surface antibody on stem cells,
pro-B cells or pre-B cells, there’s no response to antigen.
At the immature B-cell stage, when
there is low level expression of IgM,
the B-cells can undergo what is called
negative selection or deletion.
And this refers to the fact that if those B-cells
encounter molecules from our own body, what we call
self-antigen, then they become deleted, because we don’t
want B-cells that can attack our own body components.
We want B-cells that can fight
infection coming from outside the body.
So there’ll be deletion of B-cells that recognize
self-antigen and there can be receptor editing.
And this is why RAG1 and RAG2 get re-expressed at
this stage, so that the receptor can be changed.
If it reacts with self, it can be
changed to a different receptor
that reacts with a foreign pathogen
rather than reacting with self.
And then once they’re mature, of course the
B-cells are fully able to respond to antigen.
I mean that’s how it
should be obviously.
You need the B-cells
to be able to respond.
And they can become activated, they can proliferate, divide up
in number and they can differentiate and become specialized.
Particularly become specialized to produce a
progeny of a B-cell that is called a plasma cell.
And what plasma cells do is they secrete
very large amounts of antibody of the
same specificity that acts at the B-cell
receptor on the surface of the B-cell.
So, in the bone marrow we have these pre-B
cells, this early stage of development.
And they have on their cell surface not a mature form of
the antibody molecule but a preliminary version if you like
of the antibody molecule where you have the normal version
of the IgM heavy chain which is called µ heavy chain.
But it’s not a mature
version of the light chain.
They have what we refer to as a surrogate light
chain, and this is made up of two components.
One is called V pre-B and
the other is called λ5.
And then as the pre-B cell matures into an immature B-cell, it
swaps the surrogate light chain
for a proper, a real light chain.
And that can either be the k
light chain or the λ light chain.
So that the mature B-cell now has on its cell surface
a proper mature version of the antibody molecule.
And initially these mature B-cells that have not yet
encountered antigen, we call them naïve B-cells.
They’ve not yet met antigen at all.
They express on their cell surface two different
classes of antibody - IgM, surface IgM and surface IgD.
And these cells have left the bone marrow and
travelled to the secondary lymphoid tissue.
And they sit in wait in the secondary lymphoid tissue in case
their particular antigen, the
antigen they recognize comes along.