Brainscape's Knowledge GenomeTM


Top Flashcards (Certified)
All Flashcards

4.1 Transfusion and Transplant Immunology > Practical 3: Immunohistochemistry for Rejection > Flashcards

Practical 3: Immunohistochemistry for Rejection Flashcards

(76 cards)

Start Studying
1
Q

What is the renal corpuscle and what does it do?

A

Its the Bowman’s capsule and the glomerulus

It filters plasma from capillaries into the renal tubules

Glomerulus at the centre (a meshwork of capillaries), surrounded by the Bowman’s capsule, the space in between is known as the Bowman’s space

This is where the ultra-filtrate of plasma is first collected

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
2
Q

Talk about CD45 as a rejection marker, what is it, were is it found

A

CD45 is a transmembrane glycoprotein

Its a protein tyrosine phosphatase receptor type C

It was originally calle lucocyte common antigen (LCA)

The gene is located on chromosome 1 in humans

It can activate SRC tyrosine kinases e.g. p56Lck

Spice variants result in various isoforms

CD45RA is expressed by naive T cells while CD45RO is experessed by activated and memory T cells

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
3
Q

Talk about CD68 as a marker

A

It is a transmembrane glycoprotein highly expressed by cells of the monocyte lineage e.g. monocytes, osteoclasts and macrophages

Scavenger receptor found on monocytes in blood and macrophages in tissue

Gene is located on chromosome 17 in humans

A scavenger receptor binds

Binds oxLDL, phosphatidylserine on apoptotic cells

Mainly located on the endosomal/lysosomal membrane but can rapidly translocate to surface membrane

May have a negative effect on MHC presentation of exogenous antigen

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
4
Q

Talk about CD19

A

A transmembrane glycoprotein
Gene located on chromosome 16
Expressed on all B cell stages except plasma cells
Functions as an adaptor protein that recreuits cytoplasmic signalling proteins and reduces the threshold for signal transduction from the BCR for B cell activation

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
5
Q

Talk about CD20

A

A B cell marker, a non-glycosylated phosphoprotein member of the MS4A family which forms a tetraspan membrane-bound protein
Encoded by a gene on chromosome 11
The extracellular portion provided the docking site for anti-CD20 mAbs binding
CD200 is expressed on the surface of all B cells beginning at the pro-B phase and progressively increasing in concentration until maturity
It is thought to funtion as a modulator of cell growth and differentiation, and to initiate intracellular signals

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
6
Q

Talk about CD3

A

Expressed only by T cells
Always associated with the TCR-produces the TCR complex
Responsible for signal transduction
Composed of 2 heterodimers and, in90% of human and murine T cells, a homodimer of zeta chain

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
7
Q

What are some signs of acute rejection in the kidney

A

Neutrophil infiltration of the glomerulus and interstitium - also seen in ischamic injury

Lymphocytic infiltrate of the renal tubules (not so much the glomeruli)

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
8
Q

What is the Banff Classification of Allograft Pathology?

A

A standardised working classification system developed in 1991 which has contributed to the standardisation of definitions for histologic injuries resulting from renal allograft rejections and provided a universal grading system for assessing these injuries

It is used to clarify the mechanism of renal allograft injury, thus providing a guide to clinical therapy

Its how pathologists report to transplantation surgeons on rejection

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
9
Q

What is the Banff Classification based on?

A

It dinstinguishes between immune and non-immune injury as well as classified the type of rejection as antibody-mediated rejection (AMR) or T cell mediated rejection (TCMR), both of which may be acute or chronic based on specific pathological findings

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
10
Q

What IHC marker does the Banff classification use?

A

It uses C4d as a marker of renal pathology associated with active AMR

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
11
Q

What are PTCs?

A

Peritubular capillaries - tiny blood vessel in the kidney

They surround the cortical parts of the proximal and distal tubules -> not the same as the vasa recta which goes into the medulla to approach the loop of Henle i.e. PTCs are capillaries of the cortex while vasa recta are capillaries of the medulla

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
12
Q

Explain how a Banff score is assigned

A

Look at histology of peritubular capillaries - scored according to the number of inflammatory cells present within the capillary lumina

Level of C4d staining in the same tissue also contributes to the score

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
13
Q

What exactly is C4d?

A

C4d is the end product of C4 regulation by factor 1

C4 -> C4b -> iC4b (by factor 1) -> C4d (by factor 1)

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
14
Q

What is AMR, how does it occur, how does it affect the tissue

A

Acute Antibody Mediated Rejection

Abs against donor Ags bind to Ags expressed on endothelial cells in the graft vessesl
The subsequent compplement activation and cell adhesion results in endothelial cell necrosis, followed by platelet deposition and coagulation
Mononuclear cells adhere to the endothelium of the glomeruli and the peritubular capillaries
This process is accompanied by C4d deposition in both tissues

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
15
Q

How do we use C4d to investigate AMR

A

C4d negative staining in peritubular capillaries seen in early development of antibody-mediated rejection

CD45 showing glomerular and peritubular infiltration 2 days post transplant = rejection - treatment needed

Persistant peritubulat capillaritis with Cd4+ cells - 12 days post-transplant

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
16
Q

What are some histologic signs of chronic rejection

A

Intimal damage to blood vessels
Less obvious mononuclear cell infiltrate
Progressive deposition of material in the interstitial space (IS)
Thickening of the IS which causes damage to the tubules, blood vessels and glomeruli

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
17
Q

c4d positive

A

Marker of the activation of the classical pathway of the comlpement system

Diagnosis of AMR by 4d depotision in PTCs

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
18
Q

CD3, CD4, CD8 positivity

A

CD3 in T cells
CD4 in helper T cells
CD8 in cytotoxic T cells

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
19
Q

CD68 positivity

A

Positive in monocytes and tissue macrophages

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
20
Q

CD68 positivity

A

Positive in monocytes and tissue macrophages

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
21
Q

CD56 positive

A

Positive in NK cells and cytototoxic T cells

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
22
Q

CD25 positive

A

Receptor for interleukin 2 and a maker of T cell activation

To identify the activated T cells in TCMR

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
23
Q

MIB-1 positive

A

A nuclear protein related to cell proliferation or cycle

To examine the proliferation of T cells (used with CD3 marker)

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
24
Q

PCNA positive

A

Proliferating cell nuclear antigen

marker of proliferation

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
25
Perforin positive
A glycoprotein for pore formation in cell membranes of target cells Positive in NK cells and cytotoxic T cells used to identify cytotoxic T cells in TCMR
26
Granzyme B positive
Expressed in cytoplasmic granules of activated cytotoxic T cells and NKs
27
TIA-1 positive
A cytotoxic granule-associated protein expressed in NK cells and cytotoxic T cells
28
CD38/CD138 (syndecan-1)
Positive in plasma cells To detect plasma cell-rich acute rejection
29
CD20/CD79a
Positive in B cells To detect posttransplantation lymphoproliferative disorder
30
Myeloperoxidase (MPO)
Positive in polymorphonuclear leukocytes To clairfy the type of infiltrating cells in allograft rejection
31
FOXP3
Positive in Treg cells (Controversial findings in allograft rejection)
32
IL-17
Positive in Th17 cells The expression of IL-17 increases in allograft rejection
33
alphaSMA (aSMA)
Positive in vascular smooth muscle cells and interstitial myofibroblasts Used to examine the thickening of the initma in chronic rejection or the degree of interstitial fibrosis
34
type IV collagen
Positive in glomerular and tubular basement membrane
35
CD31
Positive in endothelial cells To identify and evaluate the PTC and glomerular capillaries
36
D2-40
Positive in lymphatic endothelium To distinguish lymph vessels from PTC
37
Give a general note on immunosuppression in transplantation
A majority of patients receive organs mismatched at one or more HLA loci To prevent rejection, immunosuppressant drugs are administered These drugs leave patients highly susceptible to infection especially when at greatests levels pre and post transplantation The dose of drugs is gradually reduced to maintenance levels which prevent acute rejection while sustaining active defennces against infection - likelihood for chronic rejection increases Each drug also has various side effects so they are often used in combination to increase immunosuppressive effects but not toxic effects There is a x3 times higher risk of malignant disease when on these drugs for long periods of time e.g carcinomas, lymphomas, kaposi's sarcoma
38
What are the three classes of immunosuprresive drugs?
Non-specific depletion of the majority of lymphocytes and monocytes and the inhibition of the responsiveness of the reamining few e.g. Abs and corticosteroids given prior to transplantation Interference with 3 signals needed for T cell activation Cytotoxic drugs which have their affect after naive T cells have begun to proliferate
39
Give three examples of interfering with 3 signals needed for T cell activation
The signal from the TCR (TCR/CD3) - NFkB and NFAT The costimulatory signal (B7/CD28) The signal generated by IL-2 binding to its receptor
40
What are class 1 immunosuppressive drugs, give two examples?
Antibodies that react broadly with WBCS They are given prior to and post transplantation to deplete these cells - rabbit antithymocyte globulin - Alemtuzumab (rat mIgG ant-CD52)
41
What is rabbit antithymocyte globulin?
A polyclonal mixture of high affinity antibodies that binds T, B, NK, dendriticand endothelial clls, fixes complement and acts as opsonins
42
Wht is Alemtuzumab?
Humanised rat mIgG that is specific for CD52 CD52 is expressed on almost all lymphocytes, monocytes and macrophages Therefore it results in a profound, long-kasting lymphopenia
43
How does Alemtuzumab work?
The cell surface complex of CD52 and anti-CD52 (alemtuzumab) is unusually efficient at fixing complement -> complement mediated cell lysis Antibodies that bind CD52 are very close to the cell membrane increasing the likelihood that C3b will bind covalently to the leukocte suface
44
What are corticosteroids, give two examples?
Drugs such as hydrocortisone (cortisol) and prednisone Drugs which do not act on cell-surface receptors but instead diffuse across the plasma membrane and bind cytoplasmic receptor They are most effective when administered prior to transplantation They alone insufficiently immunosupress to prevent graft rejction but work well in combination with cytotoxic drugs
45
What is hydrocortisone (cortisol)?
A steroid made by the adrenal crotex It has been made clinically for more than 50 years to reduce inflammation
46
What is prednisone?
A synthetic derivative of hydrocortisone, it is a pro-drug When enzymatically converted into prednisolone it is about 4 times more potent in reducing inflammation
47
How do corticosteroids immunosuppress?
The prevent the action of NFkB by increasing the production of IkBa (inhibitor)
48
Explain the pathway of how steroids work
In the cytosol of the cell, steroid receptors form complexes with heat shock protein Hsp30 Steroids diffuse across the cell membrane and bins to this complex, releasing Hsp30 The steriod receptor now crosses the nuclear membrane and enters the nucleus Here is binds to gene regulatory sequences and activates transcription where they increase the production of IkBa (inhibitor)
49
What does IkB normally do, how is this affected by steroids?
IkB normally binds NF-KB keeping it in the cytosol steroids increase IKB production thus keeping NF-KB in the cytosol, preventing gene regulation e.g. IL2 production
50
How is NFKB normally activated
DAG activates PKCtheta which leadds to the assembly of a membrane-bund coplex - CARMA1, BCL-10, MALT1 This complex activate a multiprotein enzyme complex (IKKa, IKKB, IKKy), inhibitor of KB kinase (IKK) IKK phosphorylates the inhibitor of IKB IKB now releases NF-KB allowing it to move into the nuclease
51
What are some of the affects of corticosteroid therapy?
Decrease in inflammatory cytokine production e.g. IL1, TNF-alpha etc Decrease in adhesion moleules Prednisolone alters lymphpocyte homing, lymphocytes are barred from entering the SLT, instead the congregate in the BM Induction of apoptosis in lymphocytes and eosinophils
52
What are some of the side effects of corticosteroid therapy?
Fluid retention Weight gain Daibetes Loss of bone mineral Thinning of skin
53
Talk about blocking signals from the TCR-NFAT activation by calcineurin, give two examples
Introduced in the 1970s - used throughout 80s and 90s (cyclosporin era of drugs) Cyclosporin A Tacrolimus
54
How does cyclosporin A wor
Its a cyclic decapeptide derived from the soil fungus Tolypocladium inflatum It inhibits T cell activation by disrupting transduction signals from the TCR Cyosporin A binds to cyclophilin which binds calcineurin preventing its activation by Ca2+ and thus blocking nFAT activation
55
How does Tacrolimus work
Tacrolimus is isolated from a soil actinomycete Its a macrolide, structurally distinct from cyclosporin A but works in similar way Tacrolimus binds FK-binding protein which then binds clacineurin, preventing its activation by Ca2+ and thus blocking NFAT activation
56
What are some side effects of Cyclosporin and Tacrolimus
Since these drugs dont effect proliferatin cells, haematopoiesis and intestinal damage do not occur Nephrotoxicity may occur in patients who become sensitised, these can no longer tolerate the drug
57
What affects do tacrlimus and cyclosporin A have
The reduce expression of T cytokines such as IL-2 and TNF-a reduced cell division of T cells (lack of IL2) No cytokines to act on B cells -> cells cant divide, cells will apoptose once activated without these cytokines
58
Give an example of an antibody specific for CD3
Introduced in 1986, CD3-specific mouse mAB OKT3
59
How is OKT3 used? (3)
Treatment involved a 5-15 day course of daily injections of mouse-CD3 antibodis combined with prednisolone Anti-CD3 caused the TCRs to be internalised and therefore they were unable to recognise antigens Attempts to convert therapeutic mouse anti-CD3 antibodies into either chimeric or humanised forms were unsuccessful - hence taken off market
60
What was OKT3 replaced with?
Anti-CD52 and rATG
61
Talk about the use of a combination of immunosuppressants to prevent rejection
After transplantation patients are maintained on a combination of immunouppressive drugs that because of their cytotoxicity and the immunodeficiency they cause, are gradually reduced in dosage As dosage is increased the risk of chronic rejection increases Sometimes early symotoms of rejection appear
62
Why was OKT3 taken of the market?
Its an anti human CD3 mouse mAb but we were unable to humanise it/make a monoclonal antibody It ended up causing serum sickness, an extreme form of type III hypersensitivity which causes skin haemorrhages and an urticarial rash Immune complexes are deposited in small blood vessels - activates complement and phagocytosis - induces fever Immune reaction to the mouse antigens
63
Give four examples of drugs which block signals
Belatocept Basiliximab Daclizumab Rapamycin (sirolimus)
64
What is Belatocept?
A soluble, synthetic chimeric protein that combines the extracellular domain of CTLA-4 with the hings and fc domains of the IgG1 heavy chain Acts to prevent alloreactive T cells receiving their co-stimulatory signal from activated DCs expressing B7 and presenting alloantigen It works as well as the older drugs but preserves kidney function more, it is associated with increased incidence of episodes of acute rejection
65
How does IL-2 act normally?
IL-2 acts in both an autocrine and paracrine fashion by binding to the IL-2 receptor expressed by T cells Naive alloreactive T cells express a low-affinity receptor for IL-2 (consists of B and y chains) On recognition of alloag the alpha chain (CD25) is synthesised - high affinity receptor
66
What are two IL-2 signal blocking drugs?
Chimeric basiliximab Humanised daclizumab Both are IgG mAbs specific for CD25
67
How do basiliximab and daclizumab work?
They are both mAbs specific for CD25 Thes are first given just before the transplant is performed Subsequent infusions are gien during the first 2 months after transplantation Benefit is that this treatment targets T cells embarking on activation - does not induce significant immunodeficiency
68
How does basiliximab and daclizumab only work on partially activated T cells
Anti-CD25 anitbodies dont bind to the low affinity IL-2 receptor on naive alloreactive T cells Anti-CD25 will bind to the high affinity IL2 receptor on activated T cells and prevents the generation of signal 3
69
What are cytotoxic drugs?
These kill alloantigen-activated proliferatin T cells -> hence they are given after transplantation
70
List the cytotoxic drugs
Azathioprine Mycophenalate mofetil Cyclophosphamide Methotrexate
71
What is azathioprine
A pro-drug much used in kidney transplantation, administered after transplantation Its eventually converted to 6-thioinosinic acid which inhibits the production of inosinic acid, an intermediate in the production of adenine and guanne - this inhibits DNA replication Like all cytotoxic drugs it damages all tissues normally active in cell division e.g. bone marrow, intestinal epithelium, hair follcles etc Same side effects as chemo: anaemia, leukopenia, thromboytopenia, intestinal damage, hair loss, hepatotoxic
72
What is mycophenalate mofetil?
A product of penicillium stoloniferum Its metabolise in the liver to mycophenolic acid It inhibits inosine monophosphate dehydrogenase, necessary for guanine synthesis
73
What is cyclophosphamide?
N2 mustard compound developed as a chemical weapon Used heavily during WWI Its a pro-drug converted to phosphoramide mustard which alkylates and cross-links DNA molecules This affects normal cell division and transcription Equally immunosuppressive pre and post antigen stimulation In addition to side-effects shared with other cytotoxic drugs it specifically damages the bladder (cancer or haemorrhagic cystitis) Not particularly toxic to the liver - useful alternative -used in short courses
74
What is methotrexate?
A drug found to be very effective in cancer treatment It prevents DNA replication by inhibiting dihydrofolate reductase which is essential for the synthesis of thymidine (thymine + deoxyribose) Its the drug of chouce for inhibiting GVHD in bone marrow transplant receipients
75
Sirolimus
Binds FK-binding protein like Tacrolimus but prevents T cell activation by inhibiting signal transduction from the IL-2R It also induces Treg development It impedes progression through the proliferation cycle by inhibiting p70 S6 kinase and activity of cdk2-cyclin E complex
76
Sirolimus
In notes

4.1 Transfusion and Transplant Immunology (24 decks)

Brainscape helps you reach your goals faster, through stronger study habits.
© 2025 Bold Learning Solutions. Terms and Conditions