nm0260: can i just say a few words before i actually formally start lecturing 
er i want to explain just a little bit about what we're trying to do er in 
these half-dozen or so lectures er my aim in a way with these lectures has been 
to try and illustrate a s-, er a chunk of medical practice er today now er 
rather than have sort of all a lot of theoretical stuff about the epidemiology 
of disease and genetics of disease and so on just trying to show you how 
actually er clinicians cope with disease and try and relate that to the science 
to the biology underlying the pathology of the disease and the nature of the 
treatments they're using so sometimes people find this these few lectures a 
little bit bitty which i'm afraid they are er now you had Dr namex er was it 
this morning er talking to you about the physiology of kidney er kidneys er and 
of kidney disease the way kidney diseases develop and talking you to you i 
think in general terms about the nature of treatment of kidney 
disease now Dr namex is not er the usual lecturer i have to say that the usual 
lecturer that gives those lectures is Dr er Dr namex whom i know very well and 
i don't know Dr namex and i i'm er told that he did a good job and i sincerely 
hope he did er but the point about that is to give you background in-, into the 
understanding of renal disease now my colleague Dr er golly my colleague Dr his 
name drops out of my mi-, namex er on Thursday will be talking to you about 
care and maintenance looking after of er patients who've been transplanted for 
kidney disease and he'll be talking to you a little bit about the ethics of 
transplantation and inevitably he will go over some of the ground that i'm 
going to present now what i want to talk about as you can see is the science of 
transplantation in other words i'm going to present to you the biology 
underlying transplantation of organs and of bone marrow er and how the 
physicians er can cope with that and make it work okay so i'm providing the 
scientific backup er as it were to the clinical side and a and as i say the aim 
is overall to get you some give you some feel for how modern clinical medicine 
works in a rather highly technical biological area now we can't hope to cover 
the whole of medicine so we've honed in homed in i'm sorry on this little area 
of renal disease and renal transplantation okay now those of you and i know 
some of you have already looked at the web may have looked at my lecture notes 
er and i'm sorry i but i've actually revised the lectures and what i'm 
presenting this afternoon is related to what's on the web already but is in 
fact presented in quite a different way what i've deliberately done is actually 
cut down on the detail and try i'm trying to bring out a bit more clearly the 
principles because in my experience if i try and teach too 
much about histocompatibility and about how er T-cells respond to 
histocompatibility antigens in detail er er people don't really cope with that 
er in the short period that i have to present it so i'm giving it to you in 
rather more general terms today er and i know some people may not like that 
some people i know prefer to have much more detail well you can find detail in 
your textbooks i always say these lecture notes will go up onto the web er 
within the very next few days what i haven't done yet either is revised er the 
reading that i have on the web site i will do that before the end of term which 
i'm very much aware is Friday okay so i hope by Friday i won't absolutely 
guarantee but i hope by Friday to have some reading some more up to date 
reading some of the reading material as i say is still perfectly valid so it 
will remain there but i will try to put some new stuff on as well okay 
nm0260: we've got a 
two hour slot now the material i'm going to present i suspect will take more 
than an hour but i'm hoping very much that it won't take two hours but in the 
past i've tried to present it in one hour and i've had to rush it so i've 
extended it to two hours but i don't think i need two hours right so let's 
actually get on with things the title of these lectures is The Science of 
Transplantation and that's very much what i'm going to be talking about er 
rather than the medical clinical aspect of transplantation which as i've said 
i'm leaving to namex er namex so the first point the first thing i want to do 
is just show you the aims a little bit about the aims of these lectures what i 
want to cover er is the range of transplant medicine the sorts of areas where 
transplantation is done and why it's done okay secondly i want to give you er 
an understanding of the immunological basis of graft rejection i'll define 
these terms presently er then i shall go on 
to talk to you about immunosuppressive therapy and how that works to present 
prevent grant rejection and finally a few words on what might be the future er 
the way forward for transplantation science and its application to medicine so 
i shall start off with a few simple definitions er couple of them are pretty 
obvious we talk about the host that is the person who receives the 
transplantation whatever it is although in fact more commonly the term we use 
is recipient somebody who receives the graft and then secondly the partner in 
this business of course is the donor and that is the one who provides the graft 
now donors are going to fall into two main categories those that are alive and 
those that are dead now living donors er would may be either unrelated to the 
recipient or related a sibling or or parent or whatever obviously living donors 
can donate things like blood bone marrow and one of your two kidneys but 
they're going to 
be hard-pressed indeed to donate lungs or something like that so more often in 
fact much more often er in transplantation medicine the s-, donor is dead and 
is a cadaver cadaveric donor and obviously under those circumstances the whole 
range of organs potentially is available for transplantation now just in 
passing when i say d-, dead cadaveric er r-, rather ghoulishly w-, er it ha-, 
the the the the person has to be somewhat freshly dead because of course er 
very shortly after death the tissues and the organs of the cadaver start to 
decay and and disintegrate and so if somebody dies in a traffic accident or is 
in the intensive therapy unit or something like that and is on a heart lung 
machine which they're about to turn off okay then the surgeons come in and whip 
out the bits the kidney the liver the lungs and so on now i'm not going to go 
into the ethical aspects of that but obviously there's a lot of ethics er which 
well namex will deal with tomorrow 
Thursday rather a lot of ethical issues are around about organ donation in both 
living and cadaveric donors okay more simple definitions the word transplant is 
er synonymous we use th-, them synonymously use it synonymously with graft the 
transplanted graft may be actually tissue of some sort such as blood or marrow 
or indeed skin or may be er an intact whole organ such as kidney all right now 
there are three other simple terms that i need to define one is autograft that 
means that you're grafting from one part of the body to another part of the 
body and that's very commonly done for skin in the case of burns obviously 
autografting we er the equivalent term is autologous graft autografting 
presents no immunological differences because you're just moving a bit of 
tissue from one part of the body to another part of a body autografting is 
actually very important and i was talking to a surgical friend of mine 
and she was describing to me the sort of facial reconstructions that happen in 
severe burn patients er i-, or severe or in situations where there's been a 
severe d-, dec-, severe accident and there's been extensive damage say to the 
face they transplant bits of bone er from the femur or a humerus or something 
like that to reconstruct the bone and muscle they get from somewhere else and 
skin from somewhere else and they reconstruct er the person's face and that is 
of course autografting in the case of things like kidneys what's much more 
often done is an allograft now people you will recollect the use of the world 
word allele which means different an allograft comes from a different 
individual of the same species okay er allogeneic graft is a term which is 
often used simultaneously s-, synonymously i'm sorry and finally in this sort 
of er hierarchy of grafting we 
have a xenograft now xeno means foreign and xenografts come from or or 
xenogeneic graft comes from a member of a different species for example pig to 
man yeah so let's briefly look at the range of transplant medicine what is done 
and why i'm going to talk briefly about currently successful grafts that is 
grafting transplanting which in which is w-, which is in use in medical 
practice today right at the end i shall refer to perhaps some possible future 
types of graft now blood transfusion is obviously a kind of graft which has 
been in use for a very long time er bone marrow or stem cell transplantation is 
much more recently used er and i'll describe that in a moment and then finally 
er we have solid organ transplantation things like kidneys and livers so there 
are these three broad areas of transplantation bone sorry blood 
bone marrow and solid organs and they are actually all of them really somewhat 
different in their characteristics now blood transfusions er have been 
attempted were attempted in th-, in in the nineteenth century and th-, there 
were a number of problems immediately discovered the major problem comes from 
the A-B-O blood group system er which i guess many of you will be familiar with 
there are on the surface there is on the surface of red cells an antigen which 
occurs in three er allelic forms there's the A form the B form and the null or 
zero form okay er obviously you can get heterozygotes so you get A-A A-B B-B A-
BO and B-O and whatever now obviously if you transfuse blood of one er blood 
group type into an individual of another blood group type the immune system of 
the recipient will recognize the blood as foreign 
and you will get an immune response typically you will get an antibody response 
to the A-B-O antigen and that will lyse the red cells with the aid of 
complement and do all sorts of serious damage to the individual er the 
consequence o-, of the damage that's done is often very severe illness er 
indeed death in some cases if you transfuse the wrong blood once the A-B-O 
group system blood system is properly was properly worked out m-, early in the 
last century er er s-, b-, blood transfusion is obviously a simple 
straightforward and life saving treatment okay so having overcome the A-B-O 
problem that's fine but still you will recollect you will understand er that 
there are residual problems in blood transfusion particularly this issue of the 
transmission of blood borne pathogens er and virologists will recognize all 
those acronyms H-I-V er er human immunodeficiency virus and H-B-V and H-C-V too 
hepatitis viruses and of course C-J-D now these are 
blood borne pathogens which of course if you transfuse blood contaminated with 
those pathogens you pass the er transmit er the disease obviously er the viral 
problems can be resolved by screening you check the blood for the presence of 
the er viruses either antigenically by serotype se-, serological assays or by 
some sort of molecular assay and if the blood is clear then you're happy to use 
it C-J-D Creutzfeldt-Jakob Disease is much more problematic er because there's 
no simple test to detect C-J-D so you can't screen this er problem if it is a 
problem er has been solved largely by separating the leukocytes from the red 
blood cells when you're doing a regular blood transfusion what's needed is not 
the leukocytes but of course the red cells for carrying oxygen around the body 
you can eliminate the leukocytes fairly straightforwardly by a a kind of er 
gradient centrifugation so that 
you can prepare blood which is essentially completely free of leukocytes now 
prions which are the C-J-D organism er are in leukocytes if they're in blood at 
all not in the red cells so if you get rid of the leukocytes then you can 
transfuse the blood safely okay so blood transfusion is a form of 
transplantation which is very straightforward er and sorted out go on to bone 
marrow transplantation now everybody is familiar with what bone marrow is bone 
marrow is the er or is the tissue which generates all the haema-, 
haematopoietic tissues the blood and the white blood cells er and o-, other 
parts of the haematopoietic system er individuals who have failed bone marrow 
er are going to be very sick er and a life saving treatment is the trans 
plantation of bone marrow into those individuals now when i say bone marrow 
transplantation actually i mean it in a slightly wider sense it can be the 
actual bone marrow which is taken from 
the donor's bone and i have had it i have done it and i tell you it is very 
painful don't recommend it except for very close friends er so you can take the 
actual bone marrow or you can take what are called C-D-thirty-four cells now 
those of you who are familiar with immunology will recollect that C-D-thirty-
four is the antigen which defines the er pluripotent stem cell which generates 
all cells of the haematopoietic ser-, system now the C-D-thirty-four cells are 
normally within the bone marrow that's why you go for the bone marrow but you 
can get C-D-thirty-four cells out of the bone marrow into the peripheral 
circulation by treating people with certain cytokines which cause the C-D-
thoity-fo thirty-four cell to proliferate and move from the bone marrow into 
the periphery into the circulation so that's why we speak about mobilization of 
C-D-thirty-four cells into the periphery and then of course you can get these 
cells by simply taking blood and 
that's much less painful than bone marrow donation okay so you can get the stem 
cells of the haematopoietic system either from the bone marrow or from the 
blood if you treated patients people with a cert-, with the cytokines now bone 
marrow transplantation is used broadly in these two situations the one major 
condition er where the bone marrow completely is non-functional is a condition 
called SCID severe combined immunodeficiency okay and the other situation er 
where bone marrow fails completely is in certain therapeutic manoeuvres carried 
out in the treatment of some cancers and i'll talk about both these in a moment 
so let's go on briefly and talk about SCID just for a moment some of you 
writing the essay on on gene therapy will have heard about SCID in the context 
of gene therapy so you will recollect that SCID is an inherited failure of the 
ontogeny of T-cells individuals who've inherited this condition 
fail to produce functional T-cells what that means is not having T-cells is 
that essentially you have no immune responsiveness at all you have you have the 
non-adaptive immune system things like natural killer cells but your T-cell 
system has failed and so the adaptive immune system is essentially absent you 
can neither generate cell-mediated immunity nor can you make antibodies so you 
are very sick extremely susceptible to infection okay quite simply in this 
situation bone marrow transplantation provides the patient with normal T-cells 
which are of course derived from the donor okay so the transplanted r-, the r-, 
r-, recipient's T-cells after the transplant are of course of donor origin not 
his own that goes without saying now in certain cancers er the therapy of 
cancer often depends on the use of so-called cytotoxic drugs now these 
cytotoxic drugs are 
designed to kill the cancer cells but they are never completely specific well 
rarely are they even slightly sic-, specific in fact er to the cancer cells and 
so if you treat patients with cancer with these drugs a consequence is damage 
to all those organs and tissues which are rapidly proliferating all right so if 
you give a large dose of this sort of therapy you will damage er the bone 
marrow of the patient to such an extent that the bone marrow will no longer 
function is no longer able to produce haematopoietic cells under those 
circumstances the patient will die okay so what we do what is done rather is 
that the patients who are treated with very large doses of therapy to destroy 
the cancer are what what is called rescued by bone marrow transplantation now 
what that simply obviously means is that the bone the patient is in a se-, es-, 
essentially brought back to life by the transplantation of bone marrow now 
equally transplanted marrow in this 
in some situations also seems to have an anti-cancer effect which is a by-
product a bonus if you like in other words the transplanted marrow seems to 
develop some sort of immune response to the cancer and that actually helps 
survival in some circumstances but i'm not going to talk about that in detail 
now talking about SCID the graft obviously has to be an allograft because the 
patient has no bone marrow of his own no no no adequate bone marrow at least so 
the situation there it has to be an allograft in the case of the cancer therapy 
you can actually autograft in other words you can take the patient's own bone 
marrow treat the guy with cytotoxic therapy and then replace his own bone 
marrow okay you can also do an allograft in that situation and both these 
techniques have their ad-, advantages and disadvantages the advantage of the 
autograft is you have er you no 
immunological problems again the disadvantage of the autograft is that 
potentially the engrafted marrow carries with it cancer cells okay which of 
course is self-defeating because the cancer cells will then start growing again 
'cause they haven't been subjected to the therapy so the autograft has the 
disadvantage that in fact you may be grafting cancer cells at the same time as 
you're grafting back the patient's marrow in the case of the allograft you have 
immunological problems which i'll describe presently but the allograft seems 
surprisingly to have a better er anti-cancer effect than the autograft so there 
are advantages both ways from a clinical point of view er i think the autograft 
generally is preferred because it's easier to manage because the immunological 
consequences of the allograft can be very severe which we'll get on to in a 
moment okay pause a moment to let you catch up so this 
is showing it diagrammatically this is the sick patient who has a cancer say 
originating in the stomach which has spread about the body now this patient is 
going to die unless you can treat the cancer that's spread about the body so 
you give him very large dose of anti-cancer drugs sufficient to kill all the 
cancer cells but at the cost of destroying the bone marrow so that's what that 
bomb is so this guy is flat out and you put back in stem cells by stem cells i 
mean either bone marrow or the C-D-thirty-four cells er and who from either i 
autograft or allograft to replace the haematopoietic system and then with luck 
the patient's cured and there is actually good evidence that cures do occur 
under these circumstances so this is a sort of therapy important therapy so 
let's go on to solid organ grafts and the idea here is very straightforward and 
simple er if an 
organ fails replace it all right straightforward but we do there are a number 
of problems as you imagine the first problem er er and this was and continues 
to be a very serious problem is that the success of the graft depends on the 
feasibility of the surgery required to transfer the solid organ from the donor 
to the recipient now in the case of things like kidney and liver er to a large 
extent lung and heart the plumbing if you like the blood vessels and the 
various er er nervous connections are relatively straightforward and so the 
surgeon can transfer an organ from one individual to another without too much 
difficulty with other organs and the arch-example is the pancreas it would be 
marvellous to transplant the pancreas to treat diabetes remember the pancreas 
the plumbing is so complex that surgically it's not feasible it's simply too 
complex for the surgeon to be able successfully to transfer the organ it's just 
not possible take too long 
too detailed too many tubes too many arteries too many veins too many this and 
that and the other so it's not practical to transplant things like pancreas so 
this is the first consideration is the surgery practical so assuming that we 
have got through to the situation where we have er practical surgery for the 
transplantation of a tissue now i want to move along to talking about the 
immunological basis of graft rejection now obviously this is the scientific 
aspect that i really need to talk about to dwell on in some detail right do 
recommend er that you review your second year immunology notes and think about 
er immunology i should have should have said at the beginning that some of the 
material that i'm presenting now obviously is related to what i presented to 
you in the second year so some of it will be familiar i hope it'll be familiar 
and you should also look 
at your second year notes so what's the problem manifestly the problem is graft 
rejection graft rejection is the er phenomenon in which the transplanted organ 
er is damaged fails through large-scale inflammation er and then literally 
starts to fall apart under immunological attack okay now the reason for this is 
that in an allograft not an autograft i should have included with autografts 
grafting from identical twins of course that's equivalent to an autograft but 
in the case of allografts the immune system of the recipient sees the graft as 
non-self somebody else manifestly and this is because there are alloantigens on 
the in the and on the graft which are different from the self-antigens these 
alloantigens you see the word allo it's related to allele again different 
alleles of the same gene okay related to allogeneic and allograft the 
alloantigens are recognized by the host immune system and adaptive 
immune mechanisms develop in the host which will eliminate the transplanted the 
non-self organ non-self tissue now what we have to recognize what you have to 
understand is that the situations with bone marrow transplantation and solid 
organ transplantation is that in the bone marrow transplantation the recipient 
the host doesn't have an immune system so obviously the host is not going to 
recognize the engrafted marrow as foreign got no ho-, go-, the host has got no 
immune system but of course the engrafted marrow is a source of immune cells 
and those immune cells as they encounter the host's tissues will recognize the 
host the recipient as foreign so in bone marrow transplantation and this is 
what makes this such a hazardous situation er the the bone marrow transplant 
actually rejects the host a rather unpleasant thought obviously in the 
situation of the solid organ allograft or indeed xenograft er the 
host er which has an intact immune system is rejecting the graft okay so we 
have these two different situations in the bone marrow and the solid organ gr-, 
er graft so in sort of clinical terms what's happening in graft rejection 
basically you're getting an immune response to the host tissue in G-V-H-D i 
should have defined graft versus host disease is abbreviated as G-V-H or G-V-D 
or G-V-H-D okay in this situation you get severe damage to the host epithelial 
er tissues the skin er and the lining of the gut but perhaps more seriously you 
get damage to the endothelium now you remember that the endothelium er is the 
lining of blood vessels and in the case of capillaries the very fine blood 
vessels the endothelial cells are the entire wall of the blood vessel so if 
you're getting an immunological reaction er to the endothelium you're going to 
get extensive bleeding into the tissues and remember 
that particularly in the kidney say the endothelium provides a very has a very 
important functional role 'cause this the endothelium which performs the 
filtration in the glomerulus and the various other bits and pieces of the 
kidney so if you're damaging the glomerulus with immune response then the 
kidney function's going to fail so in the case of graft versus host disease the 
patient has severe er generalized organ failure and if it's not treated that 
will inevitably result er in death of the patient now in the case of solid 
organ rejection of course essentially the same thing is happening and you're 
getting damage again particularly to the endothelium but also again to the 
actual cells within the organ okay leading to failure of the organ so that 
simply is the nature of rejection it's the destruction of the cells of the 
organ right now 
i'm going to specialize now and talk particularly about kidney transplantation 
because that's the subject of these lectures and it happens to be what i know 
about most so we'll get on now and talk specifically about kidneys now as you 
will have heard from Dr namex er earlier er kidney transplantation without any 
question is the best er treatment for what is called end-stage renal failure 
end-stage renal failure is when you have irrespo-, irreversible and total 
failure of the kidney okay er because transplantation allows the patient to 
have essentially normal function normal functioning they can walk around as 
usual go swimming doesn't have to have the dialysis bag er doesn't have to 
worry about things like haematopoietin injections his diet can be fairly free 
he can drink reasonably modest amounts of alcohol and so on so a kidney 
transplant patient really has a more or less normal function 
whereas somebody on dialysis is severely limited in lifestyle and in health and 
i think the point has been made quite clearly that transplantation despite the 
initial costs of the actual operation is in the longer run much cheaper than 
dialysis dialysis i forget what it costs five-thousand a year something like 
that whereas once you've done the transplant which maybe costs ten-thousand the 
cost of maintaining the patient is much less so over a period of ten years the 
transplant is teacher and in our modern new up to date N-H-S it cost that 
matters very much indeed the main limitation as i suspect you are all familiar 
is er the availability of donors now again namex will talk about this more 
tomorrow so i won't go into that in detail but the lack of donors the few 
number of small number of donors means that there are serious scientific 
consideration to other ways and means of 
restoring organ function replacing organ function i should say so as i was 
saying just now kidney transplantation is surgically straightforward surgically 
quite feasible there's the ureter to reconnect there's a couple of arteries a 
couple of veins to reconnect but basically surgeons find this fairly 
straightfoward er so the main problem is as i've been saying rejection of the 
graft and you've got to control that rejection now experience with patients 
who've been transplanted implies has told us taught us that there are actually 
three forms of rejection three different kinds of rejection er which are 
described as hyperacute and acute and chronic now the hyperacute rejection 
happens more or less immediately as soon as the kidney is plumbed into the 
recipient and the veins and arteries are reconnected and blood starts to flow 
through the kidney the host's blood starts the flow through the kidney what 
happens is that if there is hyperacute rejection going on immediately or very 
very quickly before the cessation of the operation er it will be seen that the 
kidney starts to swell become oedematous okay starts to swell and goes much 
darker in colour er and the consequence of the rejection is that the organ very 
quickly will fail and immediately has to be replaced and they have to rejoin 
the various bits and pieces close up the patient and put him back onto dialysis 
until a more suitable kidney comes along okay so that happens immediately 
during the operation we'll talk about the causes in a moment acute rejection 
despite the term acute meaning again more or less immediate acute rejection 
tends to occur some weeks after the actual transplantation six to twelve weeks 
typically what happens in this situation of course you can't see what's 
happening er [laugh] obviously er but clinically what's happening is that 
kidney function starts rapidly to decline so it's all the symptoms of end-stage 
renal failure that you will have heard from Dr namex this morning er where you 
start to get protein in the urea er urine er and you start to get large amounts 
of creatinine in the circulation because of damage to tissues and things like 
that okay now i should have said that hyperacute rejection is actually 
irreversible once it's set in there's nothing can be done okay and that's why 
you have to remove it and start again in the case of acute rejection usually er 
it can be reversed we'll go on to why and how if there's time presently okay 
sf0261: does that if the if the hyperacute one does that kill the kidney 
nm0260: yes 
sf0261: so you can't use it 
nm0260: can't use it it's dead finished and i'll tell you why in a moment okay 
now the acute rejection as 
i say usually can be reversed by w-, adjusting the therapy that's being carried 
out chronic rejection happens years after the transplantation five years ten 
years longer what it is it's a gradual slow but sadly irreversible decline in 
kidney function over a period of years and u-, ultimately the kidney fails 
completely and the patient is back er in square one okay so let's go on er and 
actually talk about the causes the immunological causes of these different 
kinds of rejection now hyperacute rejection is actually pretty straightforward 
it's due to the presence in the patient of antibodies circulating antibodies 
which are specific for the transplanted tissue which recognize antigens on a 
transplanted tissue especially by recognizing alloantigens on the endothelium 
of the transplanted tissue okay having recognized antigens on the endothelium 
er you remember complement okay and how antigen-antibody 
complexes activate complement and the co-, activated complement destroys cell 
membranes locally so you've got a complement-mediated lysis of the endothelium 
of the organ now i've mentioned endothelium as a target for attack er in 
rejection phenomena the result is loss massive loss of fluid into the organ and 
that causes the swelling okay the organ becomes oedematous and of course the 
damage to the glomerulus in this situaton completely er destroys the function 
of the kidney so that kidney is dead now you might ask where did these 
antibodies arise how do they arise rather er and the answer is er often through 
pregnancy the fetus is in effect in effect an allograft because immunologically 
the fetus is a hybrid between the father and the mother so the mother will 
potentially become sensitized to the paternal antigens of the fetus now there 
is a little bit of exchange of blood across the placenta there's no qu-, it 
it's quite clear 
that there will be fetal cells in the maternal circulation so the mother will 
develop antibodies to the fetus okay now suppose that the transplant for 
whatever reason maybe it comes from the husband or father perhaps i should say 
partner maybe that transplant comes from the husband you wouldn't do this in 
fact for these reasons er but obviously er under those circumstances the 
transplant will share antigens with the fetus and the mother's antibodies will 
destroy it er the other situation where you are likely to get these antigens 
antibodies sorry arising is where there's been blood tranfusions now if you've 
had blood transfusion in the past that meant you will have had the leukocytes 
as well from the donor er and you obviously you had the blood the red blood 
cells but the leukocytes will bear antigens that are shared by the endothelium 
okay so if by chance you've been transplanted with blood that shares antigens 
with your tran-, sorry so if by 
chance you've been transfused with blood that shares antigens with the donor 
tissue then you will have antibodies to it okay so how to avoid hyperacute 
rejection it is actually very straightforward as i've said there's obviously no 
point in transplanting an organ if it's going to be immediately rejected so 
quite simply you check in advance that the recipient does not have any anti-
donor antibodies and that's actually very simply done by the so-called 
crossmatch test so you take donor leukocytes all right and you incubate those 
leukocytes with serum from the recipient if the ser-, s-, recipient a-, a-, a-, 
and sorry and a source of complement so if the donor cells lyse under those 
circumstances you know there are antibodies in the recipient serum which with 
complement will lyse the target leukocytes okay so under those 
circumstances if the crossmatch test fails you would not transplant that 
particular organ into that particular recipient okay you may ask where the 
leukocytes come from to do this test in the case of a live related live donor 
it's perfectly obvious you take a little bit of blood in the case of the 
cadaveric donor you use usually the spleen as a sort of l-, source of 
leukocytes for this test okay very straightforward simple test and if a-, and 
if if it if you fail then you don't do the transplant but acute rejection is a 
very different situation er and this is due to the development over a period of 
weeks cell-mediated responses to donor alloantigens okay er these responses 
particularly are C-D-eight-positive cytotoxic T-cells now you've all come 
across cytotoxic cells in immunology before now er if you generate anti- donor 
C-D-eight cells those will obviously damage 
the donor tissues and in particular again it's the endothelium which is damaged 
and a little bit of techno-, technical terminology here if the endothelium is 
being damaged this is r-, er described as vascular rejection for obvious 
reasons 'cause the endothelium is the vasculature and may also as well or 
alternatively er damage the actual cells of the organ and that is described as 
cellular rejection after infiltrating into the organ now i make this 
distinction er because vascular rejection is much more severe than 
straightforward cellular rejection er celluclar rejection is much more easily 
coped with than is vascular rejection for the obvious reason that if you're 
getting vascular damage damage to the endothelium that is going to do much more 
severe damage to the organ than a l-, than damage to the er parenchymal cells 
of the organ okay few more words about the acute rejection as a part of the 
diagnostic process for 
acute rejection er perhaps i should say if a if a patient is er well i've 
already said that you get er protein in the urea and you get an increase of 
creatinine during acute rejection but the patient also becomes feverish now you 
remember fever is a symptom of immunological activations and the patient is 
feverish has these other problems er now there are various reasons why the 
patient may become feverish and have these sorts of problems not all of them 
are rec-, are rejection so as a part of the diagnosis of rejection what is done 
is that a biopsy is taken to do that er the physician inserts a needle er into 
the kidney under ultrasonic er direction and takes in the needle hollow needle 
obviously a hypodermic needle takes a piece of core a core of tissue and 
removes that and you can stain this and examine it histologically and if you do 
that and there's rejection occurring then you see all the classical hallmarks 
of inflammation which i hope you recollect are 
things like leukocytic infiltration into the organ things like activation of 
adhesion molecules remember release of cytokines remember and chemokines so if 
the biopsy the histological appearance of the biopsy is that then you've got a 
clear er diagnosis of rejection and if it's vascular involvement it's vascular 
rejection if it's just cellular involvement it's cellular rejection i've 
already said about damage to the endothelium that will lead to kidney failure 
so the question that arises is how to avoid graft loss due to acute rejection 
now since the point that i have been making is that the immunological response 
is due to allo-, alloantigenic differences between the donor and the recipient 
manifestly one of the things that is going to be done is to try and minimize 
the differences the antigenic differences between the donor and the host okay 
and this you may understand is called tissue type matching tissue typing 
for short so you match as closely as possible the antigenic properties of the 
donor tissue to the antigenic properties of the recipient i'll go into this in 
more detail presently and quite obviously the other thing that's done is that 
in the case of these patients the immune response will be suppressed by drug 
treatment okay so you do have two er two strategies one is reduce the antigenic 
differences by tissue typing tissue matching and suppress the immune response 
so that the immune response when it occurs and it will occur er is minimized 
doesn't do too much damage is the hope so we go on to chronic rejection now 
this is actually a somewhat different situation er histologically er that is 
microscopically er the a-, appearance of the tissue undergoing chronic 
rejection is quite different from the t-, appearance of tissue undergoing acute 
rejection there is no clear-cut evidence of inflammatory responses there may be 
some 
evidence of minor inflammation but not much okay so it's not really an 
immunologically-mediated rejection but what is found is that in these 
situations that you get deposition of quantities of fibrous scar tissue er 
collagen and and fibres er fibroblastic cells in large numbers and suchlike 
obviously if you have a lot of fibrous scar tissue deposited in in in the organ 
er in the kidney whose function is crucially dependent on the delicate 
structure of the glomerulus and the other bits and pieces then er you'll get 
literally quite literally clogging up of these parts of the organ and gradual 
loss of function okay as i've already said this is irreversible cannot be 
reversed and once the organ has failed it has to be removed and you start again 
though i've said that it is probably not an immunological response 
immunological er effect because you see no real signs of inflammation and 
inflammation is 
the hallmark of an immune response almost certainly this chronic rejection is 
due to a long period of chronic but low-level inflammation there are there is a 
little bit of inflammation going on so there is continuous minor damage to the 
tissues okay in order to repair this minor damage you have what amounts to 
wound healing responses occurring now wound healing is a very interesting 
biological phenomenon you've all of you experienced wound healing you cut 
yourself and a scar develops to close up the tissue the scar is the deposition 
of fibrous tissue particularly collagen collagen fibrals and obviously that 
happens in the kidney or the liver of whatever you have the consequence of the 
failure of the organ so almost certainly er we've got a wound healing response 
generating fibrous tissue in response in as a consequence of the chronic 
inflammation so what can we do about that now as i've said there is no 
effective therapy and it is supposed that in fact probably all grafts 
eventually are lost through chronic rejection but er it might be such a slow 
process that the patient may actually in fact outlive the graft and there is 
quite good evidence that this does happen people die from other causes before 
the organ fails now if as i say the chronic rejection is due to chronic 
inflammation or is due to the inflammation which was generated perhaps in the 
acute rejection then one supposes that chronic rejection is less likely to 
develop if you can minimize the acute rejection in other words if you manage 
the patient carefully so that the acute response to the tissues is minimized so 
if you want to reduce chronic infection and this very much appears to be the 
situation the way to do that is to reduce the probability of acute infec-, 
acute rejection at the early stage of the transplantation okay i will have a 
break 
in a few minutes because Natalie has to change her er tape in any case so i'll 
stop in a few minutes and we'll have a short break but i want to talk first 
before i stop a little bit about this issue of tissue matching and this is 
where we could get into some pretty heavy immunology i'm not going to animal in 
animal models when you graft from one mouse to another it's long been 
established that there are two antigenic differences which matter and we talk 
about major histocompatibility antigens okay major histocompatibility 
alloantigens strictly speaking you've heard of these they are H-L-A in the 
human and H-two in the mouse if two individuals differ in their H-L-A in their 
major histocompatibility antigens what happens is severe and rapid rejection 
occurs that's why these things are called major histocompatibility antigens but 
they aren't the only histocompatibility antigens they aren't the 
only alloantigens which mediate er tissue rejection and there's actually a 
whole bundle of minor histocompatibility antigens two examples in the human are 
H-Y and H-A-one and in the mouse it's everything that's not H-two H-one H-three 
H-four H-five and so on okay if anybody had ever wondered why the M-H-C of the 
mouse was H-two it's because there are multiple loci controlling tissue 
rejection which were numbered one to N and two happened to be the major complex 
okay now if you have if you have identity for the major histocompatibility 
antigens for differences in the minor histocompatibility antigens you still get 
rejection it still occurs but as the minor implies er the rejection is less 
severe and rather slower but nonetheless quite short now just very briefly when 
we take two individuals from a population if we ask the question can we match 
absolutely er tissues from those two individuals taken at random they're not 
twins two individuals taken at random and the answer is that total matching of 
tissues is in fact completely impossible for two reasons one is i hope you 
recollect that the H-L-A system is extremely polymorphic what that means is 
that there are many alleles dozens of alleles at any one locus and they are not 
randomly distributed amongst the population but m-, er fairly randomly 
distributed amongst the population okay and there are six ma-, six major loci 
on each chromosome and there are two chromosomes so an in any in any individual 
there's going to be twelve loci with dozens of alleles at each loc-, locus so 
you can quickly imagine that the probability of finding 
two identical individuals is going to be very small indeed er i won't well i 
will remind you class one and class two A B C D-R D-P D-Q all right now even 
supposing by chance you're lucky enough to match all the majors the probability 
of matching all the minors is nil because a minor antigen essentially is the 
product of any gene for which you have different alleles okay now when you 
think about the n-, the way genetics works it's going to be impossible to find 
two individuals that have no allelic differences at all except twins okay so 
total matching is impossible so that is why in transplantation you have to have 
immunosuppression okay we'll make a short break there er if you want to nip out 
and get a cup of coffee i will restart in ten minutes okay 
nm0260: 
can we settle down as i was saying at the end of the previous hour tissue 
matching total tissue matching is obviously impossible er for the reasons i've 
given you that doesn't mean to say that partial matching is not impo-, not is 
not possible er there's a couple of phenomena which make partial matching quite 
possible one is the the phenomenon of linkage disequilibrium what that means is 
that s-, er within a population some sets of alleles tend to stick together 
okay and occur at a higher frequency than others what i mean by the term of 
haplotype i think you'll probably intuitively understand is you have a series 
of loci along a chromosome linked along a chromosome then a haplotype is a set 
of alleles on along those particular along that along those p-, of those 
particular loci and we'll talk about haplotypes er with namex next t-, on on 
Thursday so linkage diseq-, equilibrium 
ensures that some haplotypes are more common than others okay so the chance of 
getting a set of histocompatibility antigens together in two individuals is 
rather better than y-, than you'd expect if there really was random segregation 
now you remember that Mendel said that traits are randomly segregated that's 
not true as you know they are linked and so what we're talking about is linkage 
keeping together a chunk of the chromosome so that the chances of having a set 
of alleles together is actually much higher than you'd expect by chance the 
second point that matters that's important is although i said there are six 
loci er some H-L-A antigens i'm talking exclusively about humans of course 
that's why i use the terminology H-L-A and not M-H-C okay some H-L-A antigens 
seem to matter much more in transplantation in particular it seems that the D-R 
of class two is very important and A and B of 
class one is also very important implying that D-P and D-Q and C don't matter 
so much which it seems to be the case now another point er which course you'll 
immediately understand is that if you're working with live related donors er 
those individuals will have at least one haplotype in common er er er perhaps i 
should be a little bit careful what i say here but there's been one or two 
unfortunate situations where fathers and children are found not to be related 
and that's always very embarrassing for the transplant physician to explain the 
important essential thing is that if we can partially match and that does 
actually improve graft survival again namex will talk about this on Thursday 
many other factors matter a-, as well m-, some factors perhaps matter more er 
than matching but matching is not unimportant so that is an end an aim now how 
is it done how do you tissue match the old-fashioned way is to use 
antibodies specific for particular H-L-A types in much the same sort of way as 
in the crossmatch test if you have an antibody say to H-L-A B-twenty-seven and 
that will lyse the target cells from the donor then those target cells must be 
H-L-A B-twenty-seven so there's a simple serological test the disadvantage of 
serological tests comes from the very polymorphism that they use to detect 
there are so many different possible t-, antigenic types that you have to have 
very large batteries of er antibodies and a worse problem is that antibodies do 
not cannot distinguish between some closely related allelic types okay so you 
have an antibody that says two types are the same whereas in fact they're 
different w-, don't want to go into that in any detail but because er what 
happens what happens now is that P-C-R techniques are used for typing er using 
allele specific primers okay so that primer will only prime that particular 
allele so you can only detect that 
particular a-, allele and you set up the reactions in a multiplex fashion so 
you can use use multiple primers in the same reaction and run just one track on 
a gel and you see various bands depending on which one is is there which 
haplotype which which antigenic type is there and of course if you run 
multiplex reactions er and multiple gel tracks you can very easily and quickly 
analyse dozens of different H-L-A types in the same in in the same er test so 
it's done by multiplex P-C-R which with which now makes er antibody typing 
quite obsolete okay so there we are we match as closely as possible er but we 
can never completely match so let's go on now and talk about how the immune 
system actually does recognize the foreign tissue er this is where we perhaps 
get into slightly heavy immunology i hope that you recogni-, r-, remember 
recollect the classical paradigm of T-cell recognition of antigen okay cast 
your mind back to the second year immunology lectures where you were taught 
that the T-cell receptor for antigen recognizes on the surface of the target 
cell or the antigen presenting cell call it what you will not the antigen not 
the foreign antigen i should say but a fragment of the pep-, of the foreign 
antigen a peptide fragment of the foreign antigen in association with the H-L-A 
antigen and the essential point with that is that the T-cell will only 
recognize the foreign peptide in association with self H-L-A this is the 
phenomenon of H-L-A restriction that the T-cell will only recognize foreign 
antigenic peptides in the context of its own H-L-A own H-L-A self H-L-A that's 
what happens for example in virus infections now in transplantation there's a 
rather different situation and on the face of it it contradicts this paradigm 
because what is being recognized and this is very well established is not self 
plus foreign peptide but instead is the 
foreign M-H-C antigen itself the foreign H-L-A itself which is recognized by 
the T-cell so this is different from the classical paradigm of antigenic 
recognition by T-cells okay now i know i talked about this a little bit in the 
second year we distinguished t-, actually two forms of antigenic recognition 
and transplantation one is so-called direct recognition where what is happening 
is what i've just said is if the host and the donor differ in H-L-A and that 
the T-C-R of the T-cell T-cell receptor recollect of the T-cell is binding to 
the foreign H-L-A now i showed you this diagram and i probably didn't explain 
it very well but this is the classical situation with viral antigens this is 
the T-cell receptor of the T-cell this is the is the self H-L-A antigen with 
which is associated a peptide a foreign peptide and these double lines are 
supposed to imply recognition by the T-cell receptor of 
this target structure okay so that's what normally happens that's what the T-
cell receptor's for now what we are saying is where we have er foreign H-L-A 
being recognized essentially this same T-cell receptor is cross-reacting 
remember antigenic cross-reaction this is an equivalent situation it's cross-
reacting with the foreign H-L-A which as you can see has a different shape from 
the safe s-, self H-L-A in conjunction with a different peptide but the point 
is by chance these two structures are similar by chance so that the same T-cell 
receptor recognizes both and that is the basis of so-called allorecognition of 
foreign H-L-A so that's direct recognition direct because it's the H-L-A which 
is being recognized should say in passing that H-L-A antigens on the cell 
surface are never empty they always have a peptide associated with them okay so 
again the structure is always peptide plus H-L-A 'cause you never get empty H-L-
A on cell surface so it's a 
rather complicated situation but there is good experimental evidence plenty of 
good experimental evidence that this actually is what's going on and i don't 
want to go into the experimental evidence because it really is too complicated 
don't have time now having said that there's direct recognition that implies 
also that there is indirect recognition now this indirect recognition is much 
more like the classical paradigm of self H-L-A plus foreign peptide 'cause the 
situation here is where the host and the donor have at least one matched H-L-A 
antigen okay now i've been saying that you make an effort to match H-L-A 
antigens for a transplant so normally you transplant there will be matched H-L-
A antigens between donor and recipient now the matched H-L-A antigen will be 
seen as self by the host T-cell receptors okay and if you have shall we say a 
foreign peptide in 
the H-L-A groove in the peptide groove that complex of foreign peptide plus 
matched H-L-A in effect is equivalent to a say a virus peptide and self H-L-A 
and potentially will activate the T-cell now this is exactly the same situation 
as with virus recognition exactly the same the question that arises is where 
then does the foreign peptide come from okay to sit in the peptide groove of 
the matched H-L-A where does that foreign peptide come from and the answer is 
that it is potentially derived from any alloantigen in the donor cell now i've 
talked about minor antigens remember so if the minor antigen provides a peptide 
which can associate with the matched H-L-A that can drive the T-cell response 
in exactly the same way as a viral antigen will there's no difference so the 
indirect recognition pathway is actually equivalent to the classical virus 
peptide activation pathway so it's that's the simple one the complex one is the 
direct because you 
have to think about er cross-reaction okay so i say the foreign peptide can be 
derived from any alloantigen i've said minor antigens or i'll mention a a minor 
antigen in a moment but obviously peptides derive from mismatched H-L-A 
antigens in the donor tissue could also provide er peptides which would 
function in this way okay so we have direct recognition and indirect 
recognition two different pathways of T-cell activation which essentially 
amount to the same end result which is activation of the T-cell and damage to 
the tissues i won't show you that again but i will give you an example of a 
minor antigen now again from your genetics you know that a major genetic 
difference between males and females is that males in addition to the X 
chromosome possess a Y chromosome now Y chro-, chromosomes don't don't encode 
very much they encode maleness they do encode what's called the male specific 
antigen 
okay which is not surprising isn't it the H-, the Y chromosome is going to have 
pro-, going to encode proteins which are not present in the female so you've 
got w-, male specific antigen which er is called the H- Y antigen H for 
histocompatibility Y for the Y chromosome encoded by the Y chromosome it's 
expressed in all male tissues so you your kidney your liver or my kidney my 
liver er bear Y antigen and fragments of that Y antigen can be presented by H-L-
A A-one or B-seven or B-eight what does that point-two mean i can't remember 
what i mean by that is obviously different er one fragment is presented by H-L-
A A-one which is a particular H-L-A A type another fragment of the H-Y antigen 
is presented by B-seven another fractio-, fragment of the Y antigen is 
presented by B-eight so what happens when you transplant male tissues into a 
female these H-Y antigens will activate T-cell responses in the female okay and 
you can demonstrate these T-cell responses get a 
C-D-eight T-cell responses to the female to the male tissue to the male tissue 
in the female again for reasons that i'm not going to go into at the present 
but b-, that namex will go into er that actually doesn't matter terribly much 
so when you have the choice of a if you're a a woman and you have the choice of 
a male or a female kidney it actually doesn't matter but in bone marrow 
transplantation it does 'cause bone marrow transplantation the matching 
actually is much more critical and it is clear that if you have Y or other 
minor antigen mismatches in bone marrow transplantations that can cause 
distinct problems immunological problems but in solid organ transplantation it 
doesn't really matter as things are clinically i should say but the point that 
i want to make there that i'm leading up to is that you have a in tran-, tissue 
matching there is not not so much a double whammy as the catch-twenty-two 
situation if you match 
H-L-A completely which is not impossible but unlikely er there'll be no direct 
recognition 'cause the H-L-As are all the same but there will still be indirect 
recognition of minor antigens on the other hand if you completely mismatch so 
there are no shared H-L-A antigens there'll be no indirecognition because 
there'll be no inverted commas self H-L-A to present the peptides but there 
will be massive direct rejec-, er recognition okay so this just is to reinforce 
give you a bit of background to the science of immunology of transplantation a 
bit of background to why how it what are the causes for rejection and lead me 
to this point that under any circumstances immunosuppression is going to be 
always necessary to suppress the immune responses which are inevitably going to 
arise to the transplanted tissue so that logically brings me on to the next 
section which is how does 
immunosuppressive therapy work quite simply immunosuppression is the use of 
drugs to prevent the development of the immune response obviously these drugs 
had to be chosen so that they're not too toxic so they don't damage the 
individual too much which they do they have side effects but they suppress 
immune responses so that you do not reject the tissue okay there's actually 
lots and lots and lots of immunosuppressive drugs available now many many many 
er dozens some are fantastically expensive er none is completely effective all 
have more or less severe side effects some less severe some very some 
essentially trivial some really unpleasant and the worst thing about them is 
that all of them are non-specific by non-specific i mean they suppress all 
immune responses not just the immune response of the grafted tissue which is 
what you want you don't want to suppress immune response to other things 
obviously so none of 
these drugs is ideal and just very quickly er we talk about first line drugs 
now first line drugs are those that are the first choice of drugs in clinical 
treatment and there are three types of drugs which are in in common use er 
you'll get a bit more fr-, about this from namex and there's plenty to read 
about about these drugs there are the so-called calcineurin inhibitors what 
these do is intervere interfere with the activation of the I-L-two gene okay so 
I-L-two gene I-L-two production is blocked okay remember I-L-two is an 
essential T-cell factor T-cell growth factor so in the absence of I-L-two T-
cells will not proliferate so that's why the calcineurin inhibitors are good 
immunosuppressive drugs they prevent T-cell proliferation the second class are 
the steroids things like prednisolone er which fairly broadly suppress 
production of a range of cytokines now you remember the importance of cytokines 
in immune responses so if you've suppressed cytokine production 
you're going to generally depress er inflammatory responses across the board 
immune responses across the board so steroids are often used or are used 
they're a front line drug first line drug and then the third class are 
inhibitors of D-N-A synthesis which block cell proliferation in a non-specific 
way now remember in an immune response a major part of the immune response the 
development of the immune response is clonal proliferation clonal expansion so 
if you have drugs which prevent cell proliferation you'll block clonal 
expansion so these three classes of drugs ah other drugs that are also used 
tend to be the expensive drugs which are good drugs very effective drugs but 
they're so expensive er that they're only used if the other drugs aren't 
working okay and they tend to be used transiently examples of those are 
antibodies to T-cells you imagine perfectly well if you treat an individual 
with antibody to C-D-three antigen that will lyse with complement all the T-
cells 
so this anti-C-D-three or for that matter I-L-two-R is going to be a very 
potent immunosuppressive drug but it's so expensive er monoclonal antibodies er 
cost if we have to use them in gram quantities and in gram quantities they are 
very expensive and there's more drugs which are coming into clinical practice 
now rapamycin looks like a favourite which seems to be another inhibitor of 
self-proliferation and this one works by blocking growth factor er signalling 
within the cell okay so these are second line drugs how are they used now the 
general principi for many kinds of chemotherapy for all sorts of disease now is 
to use combinations of drugs not one drug alone but two or three or four or 
five drugs together it's found that these tend to be more effective the 
combinations are more effective than single drugs now let's for example suppose 
that drug A inhibits ninety per cent of the immune response drug B 
ditto drug C ditto supposing that these three drugs work in different ways you 
might expect that the combination will inhibit ninety-nine-point-nine per cent 
of the immune reponse okay which is much better so they're used in combination 
and it's also found er that using them in combination you can actually reduce 
the doses that are used and that reduces the s-, undesirable side effects of 
the drugs which i'll come on to so actually this a-, tells us er some important 
things about drug therapy generally in medicine and i can mention drug therapy 
for cancer for example drug therapy for H-I-V infections the tendency is to use 
multiple drugs because in combination they're more effective than the single 
drugs alone perhaps it's not so surprising now initially the drugs are used in 
high concentrations high doses er er er and this is because when the patient 
has just been transplanted obviously that's a situation where the immune system 
of the patient is getting 
this tremendous jolt half a kilo or so of foreign tissue being put in and so 
you get tremendous activation potentially of the immune response so that's the 
point where you the s-, physicians come in with very large amounts of these 
drugs in order to suppress the very strong immune response which will occur 
initially the happy thing is that in fact the drug doses can be tapered down 
and this tells us actually something quite important you can reduce over a 
period of months the amount of drugs that are used and this is telling us that 
in fact that the patient is becoming acclimatized if you like become adapted to 
the graft okay getting used to the graft immunologically speaking that's an 
important observation it's never reduce to zero because generally if the drugs 
are abandoned then the tissue the the organ is rejected so you er is what it 
means is that the patients who have transplants are going to be treated by im-, 
with 
immunosuppression forever or well the continue the rest the rest of the their 
life or at least the rest of the graft's life now i've mentioned this issue of 
acute rejection one wants to minimize acute rejection so as soon as patients 
start to show symptoms of acute rejection the dosage of the drugs is increased 
and if that doesn't work to prevent the rejection then the second line drugs 
like anti-C-D-three are used as well so the acute rejection is a sort of 
emergency situation where the drug dosages are put up in an attempt to prevent 
rejection and the fact is that the physicians have got so used to using these 
drugs and so accustomed to their patients that in fact er acute rejection is 
now quite rare er i can't remember the figures but the actual so-called 
episodes where acute rejection occurs when i say acute rejection i don't mean 
that the tissue is lost i mean that symptoms of r-, rejection develop which 
have to be treated successful treatment of course prevents 
the acute rejection so you don't lose the tissue must make that clear er and 
the physicians have got this so well under control that graft loss to acute 
rejection almost never happens now it's very rare well not very rare but it's 
it's rare okay so that summarizes these points really in the absence of 
immunosuppression grafts are inevitably rejected acute rejection occurs after a 
delay i suppose because in the presence of immunosuppression it takes a long 
time for the immune responses to develop something that w-, should normally 
take a few days takes a few weeks because you're reducing the proliferation of 
the cells and that's what i'm saying in this last point that anti-graft T-cells 
grow more slowly under immuno-, immunosuppression now this brings us to another 
point i mentioned that there looks as if there's some sort of adaptive 
phenomenon which occurs in the patient because you can taper off the drug 
dosage without losing the graft studies 
with mice and other rodents have shown have suggested that if you graft foreign 
tissues into the animal whilst that animal is strongly immunosuppressed okay 
you indru-, induce a condition called tolerance which i hope you're familiar 
with in which you do not get an immune response with the foreign antigen in 
that situation you can stop all the immune suppression and the graft does not 
reject it because the animal is tolerant okay and this has been demonstrated 
repeatedly in mice i will say that mouse mice are really very different from 
humans and you cannot transfer er experimental data from murine models directly 
to human so it is not the case er that tolerance is sure to arise and there are 
many differences again i'm not got time to go into the differences between the 
mouse models and the human situation but the differences are significant enough 
to persuade one that the two situations are not the same but still the question 
arises does some sort of tolerance phenomenon arise in humans now i'm going to 
dip into a little bit of my own research er which we've done here er in 
collaboration with the local hospitals the local transplant unit er in 
principle you can d-, detect the development of tolerance in transplanted 
humans because what you can do is measure the numbers of T-cells in the 
recipient who are able which are able to respond to the donor's antigens okay 
so if you can count the numbers of er T-cells that respond to the patient to to 
the donor tissues enumerate them er then if you can show that these numbers are 
declining with time okay you can then argue that tolerance is developing but 
unfortunately in experimental terms this is not an easy thing to do and i just 
want to give you a little bit of the technology er c-, we are talking about 
science after all okay the way this has 
been done is by the so-called lymphocyte dilution assay lymphocyte dilution 
i've forgotten to write that down oh there it is lymphocyte dilution assay i'm 
sorry what we do here those of you that have done some tissue culture may be 
familiar with ninety-six well plates you will have done those that are done by 
others you will have used them for the for the er what's that red blood cell 
assay i've forgotten haemagglutination assay er so you can you take these 
ninety-six well plates er and you put in each well a fixed number of the cell 
of cells from the donor these we call the stimulator cells fixed numbers same 
across the plate every well gets the same number okay what you do then is 
titrate across the plate er successively reducing numbers of the recipient 
cells which we describe as the responder cells okay 'cause they're going to 
respond to the antigens on the donor cell and you will have noticed i've made a 
bog-up here of these numbers but what i'm trying to 
show is doubling dilutions as you go across the plate okay now if you score 
some measure of T-cell activation in these wells and i should say all this 
particular column has got the same number of responder cells all this column 
have got the same number so you've got twelve sorry eight replicates for each 
column okay so you do this in eight replicates across the plate if the 
responder T-cell does in fact respond if there is a responder T-cell and it 
does respond then if you have some sort of readout for the response you can 
score the well as positive or negative okay so if you go across the plate at 
this very high number of responder cells all the wells show response the next 
all the wells show response again the next all the wells show response then you 
find a column on your plate where not all the wells show response okay so X is 
response blank is no response so along here you've got one two three four five 
six out of eight has my arithmetic 
gone wrong again however many a number of responding wells and in this column 
you've got a smaller number of responding wells and in this column you've got 
no responding wells now what you can argue is where there's a response there's 
at least one responding T-cell at least one okay so by the application of 
simple statistical methods you can then calculate back what is the proportion 
of responding T-cells in the original starting population now this is a 
classical dilution analysis assay now you've done dilution analysis you did 
dilution analysis or the virologists did anyway er in your haemagglutination 
titrations which is exactly the same principle you dilute across a plate an 
indicator and you score positive or negative and then you have an end point 
it's exactly what we're doing here but what we're doing in essence is counting 
the number of responding T-cells in the patient okay and i'll show you some 
typical er data slightly difficult data to understand the c-, the rows indicate 
individual patients patients one to five for ethical reasons i can't tell you 
who they were this is the response to the donor cells as i've just described 
and as a control we have what we call the anti-third party response let's not 
worry too much about that er let's in fact leave the third party response out 
because what i want to concentrate on is the anti-donor response prior to 
transplantation and sometime post-transplantation in some patients you find 
that the number of responding cells declines these are cells per million C-D-
three cells going down roughly tenfold there and roughly tenfold there in these 
other individuals they've stayed pretty much the same or gone down a little bit 
okay so there clearly are differences between the individual patients and in 
some patients the numbers of responding cells go down quite 
dramatically implying some sort of tolerance and if we put together the data 
generally er again i don't want er haven't got the time to go into detail here 
but these are the responses that i've just been describing in terms of cells 
per million prior to the transplantation and in the same patients that's why 
they're joined up okay sometime after the transplantation now in these patients 
the numbers clearly have gone down in these patients they've stayed static or 
clearly gone up now again i'm not going to go into the clinical details of this 
but these patients actually are doing less well than those patients okay so in 
experimental terms one can demonstrate that some sort of tolerance does indeed 
develop in some patients who have transplanted so in theory these guys do not 
need full immune respon-, er er immune suppression and indeed these patients er 
there 
were steroids were no longer used in their treatment and they got no worse so 
they were able to do without steroids so y-, they only needed two of the three 
drugs that are normally used so these patients who appear tolerant clearly are 
better than those patients that that don't okay so you can do it you can 
demonstrate tolerance so the implications of the development of tolerance in in 
patients then er is that at least in principle you can discontinue the drug use 
or perhaps drastically reduce the drug use you want to perhaps have a little 
bit of immunosuppression but very much less than you would normally er and in 
fact very occasionally patients have abandoned the use of their drugs they get 
fed up taking all these drugs all their life and say bugger it i'm not going to 
take them any more usually what happens then is that the graft is rejected but 
in a few lucky people er the graft is not rejected and they are genuinely 
obviously tolerant okay but er if you talk to namex on Thursday he will say 
this will be a very brave physician that will say okay we've got laboratory 
information that tells us the patients are tolerant therefore we shall stop the 
drugs so there are ethical issues here again all right so that's a little bit 
of er a sideway sideline why would we want to discontinue drugs it's because of 
the side effects of drugs you don't want to use these nasty things you can 
avoid th-, avoid it side effects in increasing severity in fact are hairiness 
er it wouldn't bother me so much but i daresay there are some ladies that would 
object to growing beards well they do don't they er also some of the drugs are 
actually toxic to the kidney they damage the kidney that's not an ideal 
situation no there's even some argument that the development of chronic 
rejection is due to the nephrotoxicity of the drugs that are used to prevent 
rejection 
i think that's not the case obviously a considerable problem is infection okay 
er patients on immunosuppression by definition are susceptible to infection in 
fact that's not too bad because by and large er the guys who are transplanted 
people that are transplanted are a little bit older in their fifties sixties 
usually these guys have experienced most common infections and so they have 
lots of antibodies circulating so they are essentially immune to common 
infections like common colds or perhaps more severe things like streptococcus 
sore throats and so on one of the few compensations for growing old is that you 
tend not to get colds because you're immune to all the cold viruses that 
circulate so by and large infection normally is not too much of a problem but 
what is a very severe problem is cancer er and it turns out that in in 
transplanted immunosuppressed individuals can't forget can't remember the exact 
numbers the exact risk of 
cancer but it's a significantly elevated risk of cancer in patients who are 
immunosuppressed okay er and the cancers that arise the most common ones are 
skin cancers those don't matter too much because they're easily visible i'm not 
talking about melanoma if anybody has heard of melanoma there are other skin 
cancers i'm talking about non-melanoma skin cancers these are alarming but not 
dangerous because they're quite well treated and obviously they're easily 
recognized quickly and easiy recognized so skin cancers are not too much of a 
problem but what is a problem are B-cell lymphomas er and now those of you that 
are virologists will recollect Epstein-Barr virus and that this can transform B-
cells and make them into malignant cells normally the immune system prevents 
the transform the Epstein-Barr virus transform cells from cour-, developing 
into a cancer but if you immunosuppress then there is probability er that the 
transformed 
B-cells will develop into a lymphoma and that is a very serious situation very 
serious indeed for all sorts of reasons so these side effects are all very 
undesirable so that is why drug treatment is tapered off luckily that can be 
done and ideally drug treatment would be reduced to nothing okay so i'm close 
to winding up now we will l-, end a little before five o'clock what may be the 
future better drugs okay more effective less toxic ideally graft specific 
that's the ideal there are problems in the development of new drugs and again 
from a pharmaceutical those of you who are interested in the pharmaceutical 
industry this strikes me as actually really quite a problem for the 
pharmaceutical industry present immunosuppressive regimes are very good they 
work very well as i've said you almost never lose transplants through acute 
rejection so to demonstrate better drugs is going to take years of 
clinical study years v-, very long clinical trials because i forget what is the 
average survival of a graft nowadays but it's many years several years five 
eight years something like that so if you set up a clinical trial to compare 
two different er drug regimes immunosuppressive regimes it's going to be years 
before you're going to get a result and drug companies are not happy with that 
and the second thing which actually in some ways is more difficult is that 
testing new drugs may well be unethical now you will not have been exposed to 
much ethics sadly i think you ought to be but the point is that if you have a 
drug regime which is an effective therapy there is no way that you can say to a 
patient we are not going to treat you because we want to test a new drug so 
inevitably in immunotherapy and other kinds o-, sorry immunosuppression and in 
other kinds of therapy if you want to test a new drug that is usually 
going to be by adding it in to the previous set of drugs or at best exchanging 
it for one of the currently used drugs for ethical reasons now unless the new 
drug is significantly better clearly better than the old drug and remember the 
old drugs are very good then it's going to be extremely difficult to 
demonstrate a real difference because you can't leave the patient untreated 
except for your new drug in other words you can't and you cannot possibly do 
placebo trials people will have h-, have heard of what a place-, or heard of 
the term placebo that's when you use a dummy drug when you want to test a new 
drug you couldn't possibly use dummy drugs in this situation so there are 
ethical problems in testing new drugs as well as straightforward practical 
problems now another severe problem which i pointed out right at the beginning 
is that actually transplantation er is presently limited not by the clinical 
skills but by the numbers of donors i forget what the numbers of people on the 
waiting lists are locally it's large numbers and after the Alder Hey thing that 
you may have heard of er numbers of donors declined dramatically in this 
country people decided they weren't going to give up their or-, organs even 
after death and [laugh] another er significant thing is that with the reduction 
of the violence in Northern Ireland which is an excellent thing there are fewer 
freshly dead cadavers to provide transplants so the numbers of organs from 
Northern Ireland has dried up in fact i think namex will be telling you on 
Thursday that one of the main sources of organs is actually Spain er where they 
drive so badly that there are large numbers of traffic accidents which are a 
source of transplant tissues okay so there are things like that anyway w-, we 
talk about xenografting to overcome shortage of donors pig kidneys primates are 
obviously a potential source 
of grafts because they're so closely related to us but they're ruled out for 
all sorts of reasons pig kidneys are about the same size as ours and their 
biochemistry is similar but there is a severe problem of hyperacute er 
rejection for reasons which aren't very clear to er to me all of us or very 
nearly all of us have large amounts of circulating antibodies which will react 
with pig tissues and cause hyperacute rejection so this problem actually as you 
know has been solved and i don't know if i showed you this by humanizing 
kidneys basically what is done is that the kidney tissues are genet-, kidney 
the sorry the pig is genetically engineered so that the the tissues are not 
susceptible to complement lysis and again the details don't matter but the t-, 
tissues are not lysed by complement so humanization of pigs prevents hyperacute 
rejection 
now this has never been tested in human er transplantation it has been tested 
in primate transplantation so you take humanized pig kidney and transplant it 
into a chimpanzee this has been done on a smaller scale half a dozen or so 
chimpanzees or whatever and you find that the hyperacute rejection problem is 
overcome but er the the acute rejection which occurs is violent and cannot be 
controlled with the immunosuppressive drugs so any human trial of xenografting 
irrespective of any other problems ethical or otherwise any sort of xenograft 
trial in humans is a long way off if ever we have talked about therapeutic stem 
cell cloning and i for one have got serious ethical reservations about stem 
cell cloning er sorry embryonic stem cells it depends how you go about it 
perhaps but i have reservations give it a little bit of thought you might be 
able to generate 
stem cells we could regenerate say T-cells which as you appreciate will 
function in suspension you do not need to have an organized tissue for stem 
cells T-cells to work well that's not quite true because you know know i i hope 
that the main immunological interactions occur in lymphoid tissues so the T-
cells do function in an organized tissue environment but T-cells are as you 
know able to circulate and put themselves into the tissues where they want to 
be so you can well imagine er that T-cells which function in this way could be 
used could be derived from embryonic stem cells and effectively used but i for 
one i cannot see that you would be able to generate in vitro notice in vitro an 
organ such as a kidney which is a complex assembly of many different cell types 
put together in a very elaborate manner okay so i do not see that an organ can 
be made in vitro even if er we could genuinely m-, generate stem cells which 
could potentially generate a kidney it's 
just too complicated so if you want a kidney that means you and you want it by 
embryonic technology that means you've got to grow the fetus to a point at 
which it has intact kidneys and that is to my mind ethically obviously 
completely impossible so therapeutic stem cell cloning there's a big question 
mark over that except in perhaps in simple situations now if you think i've 
talked about basically bone marrow blood bone marrow kidneys liver er heart and 
lung and i've said that some kinds of transplants are impractical because the 
surgery is just too difficult now people have talked about limb transplantation 
and and rather horrifying to me thought is face transplantation has been talked 
about the surgeons say that this is technically possible and indeed a hand has 
been transplanted but i don't know if anybody recollects the patient got very 
unhappy and had it amputated he was very unhappy with having somebody else's 
hand could see it all 
the time 'cause a kidney okay you can forget about it but not a hand so the guy 
had the hand amputated face transplants it has been said they are technically 
possible the technology the surgical technology there is very complicated 
because your face has lots of different muscles which work in a very complex 
way so we are have very expressive faces er most other animals do not have 
expressive faces we do and that's because of the complicated nervous er er er 
and and vasculature n-, nerves and vasculature in the face okay but the 
surgeons say that it's technically possible but again there's a massive ethical 
problem here which doesn't seem to have been recognized is that what if 
subsequently that patient er acute rejection or chronic rejection occurred and 
the face was rejected what do you do you nip out and find another corpse or 
what so for ethical reason although it's been discussed in the press in the 
media a face transplant is not really on pancreas 
that was the example that i used at the beginning o-, of something which is 
surgically impractical but highly desirable because of diabetes if you could 
transplant the pancreas into a into a diabetic perhaps that might er give you 
er a cure for diabetes remember diabetes is treatable by insulin but is not 
curable but if you can transplant a pancreas that would cure it now what is 
being attempted is to generate beta cells the islet cells which make insulin 
remember from the pancreas prepare those in vitro from a pancreas and invue 
infuse them through the portal vein which takes them into the liver er and with 
luck the beta cells will lodge in the liver tissue and there be functional er 
and secrete insulin perhaps in brackets this is this sort of technology is just 
coming into clinical testing so that's a possibility but to bring my me to the 
s-, final conclusion er plainly spare part surgery and bone 
marrow transplantation and things like that does work whereas technically a-, 
and ethically possible it does work er and it works so well that in fact in 
many cases the graft as it were outlives the patient patient dies from other 
some other cause er and the kidney liver whatever is still functioning at the 
time of death the other cause may be due to the underlying disease perhaps give 
you an example er d-, diabete-, diabetics sadly er often have kidney failure er 
and they can be treated obviously with kidney transplantation but sadly they 
tend to die er from other complications of diabetes such as heart disease and 
if they die of heart disease their kidney may be still perfectly all right so 
spare part surgery as we have it now is actually pretty good but you might 
might think me that i'm a think of me as a bit of a sceptic er er er and 
perhaps i am but in fact for very good reasons i don't myself see any radical 
improvements likely in the near future and i think pretty well all the 
clinicians i talk to and indeed the scientists the rational scientists that i 
talk to about this tend to agree that in in in in in transplantation and indeed 
in a number of areas of modern medicine we have reached some sort of er limit 
which we seem to be unable to get across all the simple sort of problems have 
been solved the hard problems are too hard okay well i'll leave it there er and 
as i say namex will be talking to you about more clinical aspects of how to 
care for transplant patients on Thursday