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COMPLETE IMMUNOLOGY & PATHOLOGY NOTES

Full Module Coverage - All 46 ILOs

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PART 1: FUNDAMENTALS OF IMMUNOLOGY

ILO 1 - Key Definitions: Antigen, Epitope, Paratope

Memory hook: Epitope = part of antigen. Paratope = part of antibody. They are complementary to each other.

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ILO 2 - Two Arms of the Immune System

IMMUNE SYSTEM
│
├── INNATE IMMUNITY (non-specific)
│   ├── Present from birth
│   ├── Responds IMMEDIATELY (minutes-hours)
│   ├── No memory
│   ├── Recognizes PAMPs and DAMPs via PRRs
│   └── Components: barriers, phagocytes, NK cells, complement, cytokines
│
└── ADAPTIVE IMMUNITY (specific)
    ├── Develops over days-weeks
    ├── Highly specific to individual antigens
    ├── Has MEMORY (basis of vaccination)
    ├── Two arms: Humoral (B cells → antibodies) + Cellular (T cells)
    └── Properties: Specificity, Diversity, Memory, Self-tolerance

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ILO 3 - Innate Immune System: Cells, Tissues, Mechanisms

Barriers (First Line)

• Physical: Skin (keratin layer), mucous membranes, cilia (respiratory tract)
• Chemical: Stomach acid (pH 2), lysozyme in tears/saliva, defensins in skin/gut
• Microbiological: Normal flora compete with pathogens

Pattern Recognition

• PAMPs = Pathogen-Associated Molecular Patterns (on microbes, e.g., LPS)
• DAMPs = Damage-Associated Molecular Patterns (from dying cells, e.g., HMGB1, uric acid)

Cellular Components of Innate Immunity

Neutrophils

• First cells to arrive at infection (within 30-60 min)
• Short-lived (circulate 6-8 hours, survive 1-2 days in tissue)
• Functions:
  • Phagocytosis - engulf and destroy bacteria
  • Degranulation - release antimicrobial enzymes (elastase, myeloperoxidase)
  • NET formation - Neutrophil Extracellular Traps = chromatin + antimicrobial proteins released to trap pathogens
  • Respiratory burst - NADPH oxidase generates superoxide → H₂O₂ → hypochlorous acid (bleach)
• Granule contents: Primary (azurophilic) = myeloperoxidase, lysozyme, defensins. Secondary (specific) = lactoferrin, collagenase

Macrophages

• Arise from circulating monocytes; tissue-resident variants include Kupffer cells (liver), microglia (brain), alveolar macrophages (lung), osteoclasts (bone)
• Two activation states:
  • M1 (classical activation): Triggered by IFN-γ + LPS → kill intracellular pathogens, produce TNF, IL-1, IL-6, IL-12; present antigens
  • M2 (alternative activation): Triggered by IL-4, IL-13 → tissue repair, fibrosis, anti-inflammatory
• Functions: Phagocytosis, antigen presentation (MHC II), cytokine production, cleanup of apoptotic cells

Dendritic Cells (DCs)

• KEY ANTIGEN-PRESENTING CELLS (APCs) - bridge innate and adaptive immunity
• Immature DCs: reside in tissues, phagocytose antigens
• Mature DCs: migrate to lymph nodes, present antigen to naïve T cells via MHC-II
• Plasmacytoid DCs produce large amounts of Type I interferons (IFN-α/β) - critical in antiviral defense
• Express high levels of costimulatory molecules (CD80, CD86)

Natural Killer (NK) Cells

• Large granular lymphocytes; part of innate system (no antigen-specific receptors)
• Kill two categories of targets:
  1. Cells that LACK MHC-I (virally infected, cancer cells often downregulate MHC-I)
  2. Cells coated with IgG antibodies (Antibody-Dependent Cellular Cytotoxicity = ADCC)
• Mechanism: Release perforin (punches holes) and granzymes (activate caspase cascade → apoptosis)
• "Missing self" hypothesis: NK cells are inhibited by MHC-I on normal cells. Loss of MHC-I → NK activation

Mast Cells and Basophils

• Mast cells: tissue-resident; basophils: circulating
• Both contain large granules with histamine, heparin, proteases, leukotrienes
• Activated by: IgE cross-linking (Type I hypersensitivity), complement fragments (C3a, C5a), pathogens
• Key role in allergy and anaphylaxis

Eosinophils

• Granules contain: major basic protein (MBP), eosinophil cationic protein (ECP), eosinophil peroxidase
• Main function: Defense against parasites (too large to phagocytose → degranulate onto them)
• Also involved in Type I hypersensitivity (late phase reaction)
• Recruited by IL-5

Monocytes

• Circulating precursors of macrophages and dendritic cells
• 2 populations: Classical monocytes (CD14++CD16-) - phagocytic; Non-classical monocytes (CD14+CD16++) - patrolling

Phagocytosis - Mechanism

1. Recognition → Fc receptors, complement receptors, mannose receptors bind opsonized bacteria
2. Engulfment → pseudopods extend around particle, form phagosome
3. Phagolysosome formation → lysosomes fuse, lower pH
4. Killing → Reactive oxygen species (ROS), NO, defensins, lysozyme
5. Antigen processing → peptides loaded onto MHC-II

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ILO 4 - Complement System

Overview

• Family of ~30 plasma proteins produced mainly by the liver
• Circulate as inactive precursors (zymogens), activated by sequential cleavage
• Three activation pathways converge at C3 convertase

THREE PATHWAYS - FLOWCHART

CLASSICAL PATHWAY          LECTIN PATHWAY           ALTERNATIVE PATHWAY
(Antibody-dependent)       (Pattern recognition)     (Spontaneous)
        │                          │                        │
 C1q binds Ag-Ab            MBL binds mannose          C3 hydrolysis (C3 tickover)
 complex (IgG or IgM)       on microbial surfaces       → C3(H2O) on microbial
        │                          │                    surfaces (no Ab needed)
   C1r, C1s activated        MASP-1, MASP-2 activated        │
        │                          │                        │
   C4 → C4a + C4b            C4 → C4a + C4b            Factor B + Factor D
   C2 → C2a + C2b            C2 → C2a + C2b            → C3 convertase (C3bBb)
        │                          │                  (Stabilized by Properdin)
        └──────────────────────────┘
                           ↓
                  C3 CONVERTASE (C4b2a)
                           ↓
                  C3 → C3a + C3b
                           ↓
                  C5 CONVERTASE (C4b2a3b)
                           ↓
                  C5 → C5a + C5b
                           ↓
              C5b + C6 + C7 + C8 + C9 polymerize
                           ↓
              MEMBRANE ATTACK COMPLEX (MAC = C5b-9)
                           ↓
              Pores in bacterial membrane → osmotic lysis

Biological Effects (EXAM FAVORITE)

Complement Regulatory Proteins (prevent self-attack)

• C1-inhibitor (C1-INH) - inhibits C1r, C1s (deficiency → hereditary angioedema)
• DAF (Decay Accelerating Factor / CD55) - accelerates decay of C3 convertase
• CD59 (Protectin) - prevents MAC formation on host cells
• Factor H, Factor I - regulate alternative pathway
• Deficiency of CD55/CD59 → Paroxysmal Nocturnal Hemoglobinuria (PNH)

Complement Deficiency Diseases

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PART 2: ADAPTIVE IMMUNITY

ILO 5 - Types of Adaptive Immunity

ADAPTIVE IMMUNITY
│
├── HUMORAL IMMUNITY (B cells, antibodies)
│   ├── Effective against: Extracellular bacteria, viruses, toxins
│   ├── Mechanism: Antibody production, complement activation, opsonization, ADCC
│   ├── Active humoral: Vaccination, natural infection → long-lasting
│   └── Passive humoral: Mother → fetus (IgG), breast milk (IgA), antiserum
│
└── CELL-MEDIATED IMMUNITY (T cells)
    ├── Effective against: Intracellular pathogens (viruses, TB, fungi, parasites)
    ├── Mechanism: Direct cytotoxicity (CD8+), macrophage activation (CD4+ Th1)
    ├── Active: Natural infection/vaccination
    └── Passive: Transfer of sensitized T cells (experimental)

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ILO 6 - Structure and Function of Antibody (Immunoglobulin)

Basic Structure

                    Fab (antigen-binding)
                   /                    \
    V  [H]   V                    V   [H]  V
    │    │    │                    │    │    │
    C  [H]   C ——— S-S ———————— C  [H]  C
         │                              │
         └──────── Fc REGION ───────────┘
                   (Crystallizable)
                   (Effector functions)

• 4 chains: 2 heavy chains + 2 light chains (κ or λ), held by disulfide bonds
• Fab region = Fragment Antigen Binding = 2 Fab arms = antigen binding
• Fc region = Fragment Crystallizable = effector functions (complement activation, FcR binding)
• Hinge region = between Fab and Fc, gives flexibility, contains disulfide bonds
• Variable regions (V): Contain CDRs (Complementarity-Determining Regions = actual paratope)
• Constant regions (C): Determine antibody class (isotype)
• Papain cleaves at hinge → 2 Fab + 1 Fc
• Pepsin cleaves below hinge → 1 F(ab')₂ + pFc'

Functions of Antibodies

1. Neutralization - block binding of virus/toxin to cell receptors
2. Opsonization - Fc portion binds FcγR on macrophages/neutrophils → phagocytosis
3. Complement activation - IgG and IgM fix complement via classical pathway
4. ADCC (Antibody-Dependent Cellular Cytotoxicity) - IgG-coated target cells killed by NK cells
5. Agglutination - clump bacteria/viruses → easier phagocytosis
6. Neonatal immunity - IgG crosses placenta (FcRn receptor)
7. Mucosal protection - Secretory IgA in breast milk, saliva, tears

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ILO 7 - Five Classes of Human Antibodies

Exam trick: "GAMED" = IgG, IgA, IgM, IgE, IgD

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ILO 8 - Active vs. Passive Humoral Immunity

Serum sickness can occur with passive immunity (heterologous serum proteins act as antigens → Type III hypersensitivity)

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ILO 9 - Cells Involved in Cell-Mediated Immunity

CD4+ T Helper Cells (Th)

• Recognize antigen on MHC Class II (found on APCs)
• Th1 cells: Activated by IL-12 + IFN-γ → secrete IFN-γ, TNF → activate macrophages, promote CTL responses → defense against intracellular bacteria (TB, Listeria), fungi
• Th2 cells: Activated by IL-4 → secrete IL-4, IL-5, IL-13 → promote IgE production, eosinophil recruitment → defense against helminths; involved in allergy
• Th17 cells: Activated by TGF-β + IL-6, IL-23 → secrete IL-17, IL-22 → neutrophil recruitment → defense against extracellular bacteria and fungi; involved in autoimmunity
• T follicular helper (Tfh) cells: Reside in germinal centers → help B cells produce high-affinity antibodies, class switching, somatic hypermutation

CD8+ Cytotoxic T Lymphocytes (CTLs)

• Recognize antigen on MHC Class I (found on all nucleated cells)
• Kill cells displaying foreign peptides (virus-infected, cancer cells)
• Killing mechanisms:
  1. Perforin-granzyme pathway - perforin creates pores, granzymes enter and activate caspases → apoptosis
  2. Fas-FasL pathway - CTL FasL binds target cell Fas → apoptosis
  3. Cytokines (TNF, lymphotoxin) - can kill directly

Regulatory T Cells (Tregs)

• CD4+CD25+FoxP3+ cells
• Suppress immune responses → prevent autoimmunity, limit inflammation
• Mechanisms: IL-10, TGF-β secretion; cell contact via CTLA-4

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ILO 10 - Key Surface Molecules of T and B Cells

T Cell Surface Molecules

B Cell Surface Molecules

Two Signals Required for Lymphocyte Activation

Signal 1: TCR/BCR binding to antigen (specificity)
Signal 2: Costimulation (CD28-B7 for T cells; CD40-CD40L for B cells)
         ↓
  Both signals → activation, proliferation, effector functions
  Signal 1 alone → ANERGY (unresponsiveness) → tolerance

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ILO 11 - MHC Molecules (Major Histocompatibility Complex)

MHC Class I

MHC Class II

Class III MHC

• Encodes complement proteins (C2, C4, factor B), TNF, heat shock proteins - not classical antigen-presenting molecules

Exam mnemonic: "8 is great" = CD8 recognizes MHC Class I (endogenous, 8-10 aa peptides). "4×2=8" = CD4 × 2 = MHC Class II (note: Class 1+1=2 → extracellular); the extrinsic/exogenous go on II.

HLA Features

• Highly polymorphic: >10,000 alleles across all HLA genes; >3500 for HLA-B alone
• Codominant expression: Both maternal and paternal alleles expressed
• HLA haplotype: Block of HLA genes inherited together en bloc (25% chance siblings share both haplotypes)
• HLA typing: Critical for transplantation matching

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ILO 12 - Antigen Processing and Presentation

Endogenous Pathway (MHC Class I)

Intracellular protein (viral/tumor)
        ↓ Ubiquitination
   Proteasome (cytosol) → Peptide fragments (8-10 aa)
        ↓ TAP1/TAP2 transporter
   Endoplasmic Reticulum
        ↓ Loaded onto MHC-I (with help of tapasin, calreticulin)
   Golgi → Cell surface
        ↓ Recognized by CD8+ CTL

Exogenous Pathway (MHC Class II)

Extracellular protein (bacteria, viruses outside cell)
        ↓ Phagocytosis/endocytosis
   Endosome → Lysosome (acid pH, cathepsins) → Peptide fragments (13-25 aa)
        ↓
   MHC-II synthesized in ER, CLIP (invariant chain peptide) blocks peptide binding
        ↓ CLIP removed by HLA-DM in MHC class II compartment
   MHC-II loaded with exogenous peptide → Cell surface
        ↓ Recognized by CD4+ T helper cells

Cross-Presentation

• Dendritic cells can present exogenous antigens on MHC Class I to CD8+ T cells
• Critical for priming CTL responses against viruses and tumors (dendritic cell doesn't need to be infected)

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ILO 13 - HLA-Associated Diseases

Exam tip: HLA-B27 + Ankylosing Spondylitis is the MOST TESTED association. Know B27 = seronegative spondyloarthropathies (Ankylosing spondylitis, Reactive arthritis, Psoriatic arthritis, IBD-associated arthritis = "ABRI").

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ILO 14 - Humoral vs. Cellular Immunity Comparison

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ILO 15-16 - Cytokines

Definition

Major Cytokine Classes

Pro-inflammatory Cytokines (Innate response)

Adaptive Immune Cytokines

How Cytokines Collectively Provide Immunity (ILO 16)

PATHOGEN ENTRY
     ↓
Innate cells (macrophages, DCs) detect PAMPs
     ↓
IL-1, TNF, IL-6 → INFLAMMATION (fever, vascular changes, acute phase proteins)
IL-12 → NK cell activation → IFN-γ → macrophage killing of intracellular organisms
Type I IFNs → antiviral defense
     ↓
APCs present antigen to T cells + costimulation
     ↓
IL-2 → T cell proliferation
IL-12, IFN-γ → Th1 → IFN-γ → macrophage activation → kill intracellular bugs
IL-4 → Th2 → IL-4/IL-5/IL-13 → B cell class switching to IgE, IgG1; eosinophil recruitment
TGF-β + IL-6 → Th17 → IL-17 → neutrophil recruitment
     ↓
B cells with T help → Plasma cells → Antibodies → neutralize, opsonize, fix complement
     ↓
IL-10, TGF-β → RESOLUTION (Tregs dampen the response, prevent autoimmunity)

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PART 3: IMMUNOLOGICAL DISORDERS

ILO 17-19 - Hypersensitivity Reactions

Definition (ILO 17)

Gell and Coombs Classification

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TYPE I - Immediate Hypersensitivity (Allergy / Anaphylaxis)

SENSITIZATION PHASE (1st exposure):
Allergen → Th2 response → IL-4 → B cell class switch to IgE
IgE produced → binds Fc epsilon RI receptor on MAST CELLS and BASOPHILS
(IgE-armed mast cells distributed throughout tissues)

EFFECTOR PHASE (2nd exposure):
Same allergen cross-links IgE on mast cells
→ MAST CELL DEGRANULATION
→ Release of mediators:

• Preformed (early phase, seconds-minutes): Histamine, heparin, tryptase, TNF-α, proteases
• Newly synthesized (minutes-hours): Prostaglandin D2, Leukotrienes (C4, D4, E4 = SRS-A), PAF, cytokines (IL-4, IL-5, IL-13)
• Late phase (4-24 hrs): Eosinophil and neutrophil recruitment, further tissue damage

• Local: Hay fever (rhinitis), asthma, eczema, urticaria, food allergy
• Systemic: ANAPHYLAXIS (life-threatening)
  • Urticaria, angioedema
  • Bronchospasm (from leukotrienes)
  • Vasodilation → hypotension → anaphylactic shock
  • Laryngeal edema (can cause death by asphyxia)
  • Triggers: bee sting, peanuts, shellfish, drugs (penicillin, NSAIDs), latex, radiocontrast

1. Epinephrine (adrenaline) IM - first and most important treatment
2. Antihistamines (H1 blocker) - block histamine receptors
3. Corticosteroids - reduce late-phase reaction
4. Bronchodilators (salbutamol) - for bronchospasm
5. IV fluids for hypotension

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TYPE II - Antibody-Mediated (Cytotoxic) Hypersensitivity

1. Complement activation: Ab + cell surface Ag → complement → MAC + opsonization → phagocytosis
2. Opsonization/Phagocytosis: Fc receptors on macrophages bind IgG-coated cells → phagocytosis
3. ADCC: NK cells bind IgG on target cells → release perforins/granzymes
4. Receptor dysfunction: Ab binds receptor → abnormal signaling (no direct killing)
   • Stimulatory: Graves disease (anti-TSH-R antibody activates thyroid)
   • Inhibitory: Myasthenia gravis (anti-AChR antibody blocks acetylcholine)

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TYPE III - Immune Complex-Mediated Hypersensitivity

Antigen (soluble) + Antibody → IMMUNE COMPLEXES (Ag-Ab)
                                      ↓
              Normally cleared by macrophages/complement
              If EXCESS antigen or defective clearance:
                                      ↓
         Immune complexes DEPOSIT in small vessels, glomeruli, joints, skin
                                      ↓
         Complement activation → C3a, C5a → Mast cell degranulation → inflammation
         Neutrophil recruitment → lysosomal enzymes → TISSUE DAMAGE

• NOT directed at cell surface antigens (soluble Ag-Ab complexes)
• Affects multiple organs (wherever complexes deposit)
• Low complement levels (consumed) - diagnostic clue
• Detected by: immunofluorescence (granular pattern), C3 deposition

• Occurs 7-10 days after injection of foreign protein (heterologous serum)
• Symptoms: fever, urticaria, arthralgias, lymphadenopathy, glomerulonephritis, vasculitis
• Self-limiting once antigen eliminated
• Mechanism: Type III (immune complex) - excess antigen, complexes deposit systemically
• Low serum C3, C4 levels

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TYPE IV - Delayed-Type Hypersensitivity (Cell-Mediated)

1. Classic DTH (CD4+ Th1-mediated):
   • Sensitized CD4+ Th1 cells encounter antigen
   • Secrete IFN-γ → macrophage activation
   • Macrophages release enzymes, reactive oxygen species → TISSUE DAMAGE
   • Granuloma formation if antigen persists
   • Examples: PPD (tuberculin) skin test, contact dermatitis (nickel, latex, urushiol in poison ivy)
2. CTL-mediated (CD8+):
   • Sensitized CD8+ CTLs kill antigen-bearing cells
   • Examples: Graft rejection (acute cellular), viral infection of tissue, some drug reactions

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PART 4: TRANSPLANTATION IMMUNOLOGY

ILO 20-25 - Transplant Rejection

ILO 20 - Immune Recognition of Allografts

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ILO 21-22 - Classification of Graft Rejection

HYPERACUTE REJECTION

• Timing: Minutes to hours after transplant
• Mechanism: Pre-formed antibodies (anti-donor HLA, anti-ABO blood group antibodies) in recipient react with graft endothelium → immediate complement activation → thrombosis
• Morphology: Diffuse thrombosis, ischemia, infarction of graft; vessel walls necrosis (fibrinoid necrosis)
• Why: Previous blood transfusion, pregnancy, or failed transplant → pre-sensitized
• Prevention: ABO compatibility testing, cross-match test (mix recipient serum with donor cells before transplant)

ACUTE REJECTION

• Timing: Days to weeks post-transplant
• Mechanism: Host T cells (CD4+ and CD8+) react against donor MHC antigens → Th1 activates macrophages; CD8+ CTLs kill graft cells
• Morphology: Interstitial lymphocytic infiltrate, tubulitis (in renal transplant - lymphocytes invade tubules), endothelialitis, parenchymal cell necrosis
• Treatment: High-dose corticosteroids ± ATG (anti-thymocyte globulin)

• Timing: Days to weeks (may also be late)
• Mechanism: De novo (new) donor-specific antibodies (DSA) against HLA antigens → complement activation + ADCC → endothelial damage
• Morphology: Microvascular inflammation, C4d deposition in peritubular capillaries (diagnostic marker), arteritis
• Treatment: Plasmapheresis, IVIG, anti-CD20 (rituximab)

CHRONIC REJECTION

• Timing: Months to years
• Mechanism: Indirect T cell pathway + chronic antibody-mediated damage → progressive loss of graft function
• Morphology:
  • Vessels: Fibrous intimal thickening (transplant arteriopathy/arteriosclerosis) → vessel lumen obliterated
  • Parenchyma: Interstitial fibrosis, tubular atrophy (kidney), bronchiolitis obliterans (lung), accelerated atherosclerosis (heart)
  • Hallmark: Fibrous intimal thickening of vessels (smooth muscle proliferation)
• Outcome: Slowly progressive graft loss; not responsive to immunosuppression
• Often caused by indirect pathway allorecognition and chronic DSA

TIMELINE OF REJECTION:
Minutes → Hours: HYPERACUTE (pre-formed Ab, ABO mismatch)
Days → Weeks: ACUTE CELLULAR (T cells) or ACUTE HUMORAL (new DSA)
Months → Years: CHRONIC (obliterative vasculopathy, fibrosis)

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ILO 23-24 - Improving Graft Survival

Pre-transplant Methods

1. ABO blood group compatibility - mandatory
2. HLA matching - match as many loci as possible (HLA-A, B, C, DR, DQ, DP); best match = 6/6
3. Crossmatch testing - mix recipient serum with donor T cells; if positive = hyperacute rejection risk
4. Panel Reactive Antibody (PRA) - measures % of donor HLA panel recipient reacts to; high PRA = highly sensitized

Post-transplant Immunosuppression (ILO 33)

INDUCTION (at time of transplant):
- Depleting Ab: ATG (anti-thymocyte globulin), Basiliximab (anti-IL-2R/CD25)

MAINTENANCE (lifelong):
- Calcineurin inhibitors: Cyclosporine, Tacrolimus (FK506) → block IL-2 transcription → T cell suppression
- mTOR inhibitors: Sirolimus, Everolimus → block IL-2 signaling → T cell proliferation blocked
- Antimetabolites: Mycophenolate mofetil (MMF) → block purine synthesis → lymphocyte proliferation blocked
  Azathioprine → purine analog → suppress rapidly dividing cells
- Corticosteroids → broad immunosuppression, anti-inflammatory

REJECTION TREATMENT:
- Pulse steroids (high-dose methylprednisolone)
- ATG for steroid-resistant rejection
- Plasmapheresis + IVIG for antibody-mediated rejection

Recent Developments (ILO 24)

• Belatacept - fusion protein blocking CD28-B7 costimulation → selective T cell tolerance
• Tocilizumab - anti-IL-6R; used in antibody-mediated rejection
• Eculizumab - anti-C5 complement inhibitor; used in antibody-mediated rejection
• Kidney paired donation - incompatible pairs swap donors
• Xenotransplantation - genetically engineered pig organs (CRISPR to delete pig Gal antigens) - recent breakthroughs
• Tolerance induction protocols - combined bone marrow + organ transplant → chimerism → graft tolerance without lifelong immunosuppression

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ILO 25 - Graft-versus-Host Disease (GvHD)

• Immunocompetent donor T cells recognize host tissues as foreign
• Donor T cells attack host tissues
• Opposite of normal rejection (here the graft attacks the host)

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PART 5: IMMUNOLOGICAL TOLERANCE AND AUTOIMMUNITY

ILO 26 - Immunological Tolerance

Central Tolerance (Generative Organs)

• Positive selection: T cells that recognize self-MHC survive (those that don't = die by neglect)
• Negative selection (clonal deletion): T cells with HIGH affinity for self-MHC + self-peptide → apoptosis (elimination of autoreactive clones)
• AIRE (AutoImmune REgulator): Expressed by thymic medullary epithelial cells; drives expression of peripheral tissue antigens in thymus → delete self-reactive T cells
• AIRE deficiency → APS-1 (Autoimmune Polyendocrinopathy Syndrome type 1)

• Receptor editing: Change light chain gene to avoid self-reactivity
• Clonal deletion: Strongly self-reactive B cells deleted

Peripheral Tolerance

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ILO 27-28 - Autoimmunity

Mechanisms of Breakdown of Self-Tolerance (ILO 27)

1. Molecular mimicry: Foreign antigen resembles self-antigen → antibodies/T cells cross-react. Example: Group A Strep M protein mimics cardiac myosin → rheumatic fever
2. Polyclonal activation: Mitogens (EBV, LPS) activate many lymphocyte clones including autoreactive ones
3. Loss of Treg function: Decreased IL-10/TGF-β → uncontrolled autoimmune responses
4. Epitope spreading: Initial response to self-antigen exposes new epitopes → broadening autoimmune response
5. Release of sequestered antigens: Trauma releasing lens protein (sympathetic ophthalmia), myelin
6. Aberrant MHC-II expression: IFN-γ induces MHC-II on cells that normally don't express it → present self-antigens
7. Genetic susceptibility: HLA associations (see ILO 13), PTPN22 mutations, FOXP3 mutations

Common Autoimmune Diseases and Their Autoantibodies (ILO 28)

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ILO 29 - Systemic Lupus Erythematosus (SLE)

Epidemiology

• F:M = 10:1; peak age 20-40 years
• Prevalence: ~1 in 700 women (20-60 yrs); 1 in 250 Black women
• Most common serious autoimmune disease

Pathogenesis

1. Genetic factors: HLA-DR2, HLA-DR3 (RR 2-3, combined ~5); complement deficiencies (C1q, C2, C4) → poor immune complex clearance; PTPN22 polymorphism
2. Environmental trigger: UV light → apoptosis of skin cells → release of nuclear antigens (dsDNA, histones, snRNPs)
3. Failure of clearance: Defective C1q, DNase I activity → nuclear debris accumulates
4. Autoantibody formation: Plasmacytoid DCs produce excess IFN-α → TLR9/TLR7 activation → B cells produce anti-nuclear Abs
5. Immune complex deposition: Anti-dsDNA + dsDNA complexes → deposit in glomeruli, vessels, joints → complement activation → inflammation → tissue damage
6. IFN-α signature: Blood cells show striking Type I IFN gene expression pattern

Clinical Manifestations (SOAP BRAIN MD mnemonic)

2019 EULAR/ACR Classification Criteria

• ANA ≥1:80 = obligatory entry criterion
• Score-based system: ≥10 points = SLE

Lupus Nephritis (Most Serious Complication)

• Class I: Minimal mesangial - normal light microscopy
• Class II: Mesangial proliferative - mesangial deposits
• Class III: Focal proliferative (<50% glomeruli)
• Class IV: Diffuse proliferative (>50% glomeruli) - MOST SEVERE; "wire-loop" lesions = subendothelial deposits
• Class V: Membranous - subepithelial deposits; nephrotic syndrome
• Class VI: Sclerosing

Morphological Features (Exam Favorite)

• "Wire-loop" glomerular lesions in Class IV (LM: thickened capillary walls from massive subendothelial deposits)
• Libman-Sacks endocarditis - non-infective endocarditis; small vegetations on BOTH surfaces of valves (especially mitral)
• Onion-skin lesion - concentric periarteriolar fibrosis in spleen
• Hematoxylin bodies - in tissue, represent engulfed nuclei of necrotic cells

Investigations

• ANA (most sensitive, 95-99%)
• Anti-dsDNA (most specific, 60-70% sensitivity, titer correlates with disease activity)
• Anti-Sm (highly specific, 25-30%)
• Complement: Low C3, C4 (consumed by immune complexes) - correlates with activity
• CBC: Anemia (hemolytic or anemia of chronic disease), leukopenia, thrombocytopenia
• Urinalysis: Proteinuria, hematuria, RBC casts (active nephritis)
• Renal biopsy: For classification and treatment decisions

Management

• Mild disease: Hydroxychloroquine ± NSAIDs
• Moderate disease: Low-dose corticosteroids
• Severe/organ-threatening: High-dose corticosteroids + cyclophosphamide or mycophenolate
• Biologic: Belimumab (anti-BLyS/BAFF) - reduces B cell survival
• Anifrolumab (anti-IFN-α receptor) - newer biologic
• Lupus nephritis: MMF or cyclophosphamide + steroids
• Antiphospholipid syndrome: Anticoagulation (warfarin)

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ILO 30 - Rheumatoid Arthritis (RA)

Aetiopathogenesis

1. Genetic: HLA-DR4 (DRB1*0401, *0404) - shared epitope hypothesis; PTPN22 polymorphism
2. Environmental trigger: Smoking (most important modifiable risk factor) → citrullination of proteins
3. Citrullination: PAD4 enzyme converts arginine → citrulline in proteins → neo-antigens
4. Anti-CCP antibodies form against citrullinated proteins (earliest marker, can appear years before symptoms)
5. T cell activation: CD4+ Th1 and Th17 cells activated against citrullinated antigens in synovium
6. Synovitis: TNF-α, IL-1, IL-6, IL-17 → synovial inflammation, angiogenesis, pannus formation
7. Pannus: Aggressive granulation tissue from inflamed synovium → invades cartilage and bone
8. Destruction: Osteoclasts (activated by RANKL) erode bone; matrix metalloproteinases destroy cartilage

Clinical Manifestations

• Symmetric, additive polyarthritis - small joints (MCPs, PIPs, wrists) preferentially affected
• Morning stiffness >1 hour (key feature)
• Soft tissue swelling, warmth, tenderness
• Late deformities:
  • Swan neck deformity (PIP hyperextension, DIP flexion)
  • Boutonniere deformity (PIP flexion, DIP hyperextension)
  • Ulnar deviation at MCPs
  • Z-thumb deformity
  • Subluxation of atlanto-axial joint (C1-C2) → can compress spinal cord

Morphological Features

• Synovium: Villous hypertrophy, lining cell hyperplasia (2→20 cell layers), lymphocytic infiltrate (T cells, B cells), germinal center formation, increased vascularity
• Pannus: Fibrovascular tissue that erodes cartilage and bone
• Rheumatoid nodule: Center = fibrinoid necrosis surrounded by palisading macrophages/histiocytes, outer = chronic inflammatory cells

Investigations

• RF (Rheumatoid Factor): IgM anti-IgG; positive in 70-80%; NOT specific (also in SLE, Sjögren's, healthy elderly)
• Anti-CCP: Most specific (95-98%); early marker; predicts erosive disease
• ESR/CRP: Elevated (disease activity markers)
• X-ray: Soft tissue swelling, periarticular osteopenia (early), joint space narrowing, erosions (late), subluxation
• Synovial fluid: Turbid, WBC 2000-100,000 cells/μL (predominantly neutrophils), low glucose

2010 ACR/EULAR Criteria (score ≥6 = RA)

• Joint involvement (1-4 small joints = 2 pts; ≥10 joints = 5 pts)
• Serology: High-positive RF or Anti-CCP = 3 pts
• Acute phase: Abnormal CRP or ESR = 1 pt
• Duration ≥6 weeks = 1 pt

Management

• NSAIDs - symptom control
• DMARDs (Disease-Modifying Anti-Rheumatic Drugs):
  • Methotrexate - first-line DMARD; folic acid supplementation needed; regular LFTs
  • Hydroxychloroquine, sulfasalazine, leflunomide
• Biologics (for failed cDMARDs):
  • Anti-TNF: Infliximab, etanercept, adalimumab (most widely used class)
  • Anti-IL-6R: Tocilizumab, sarilumab
  • Anti-CD20: Rituximab
  • Abatacept (CTLA-4-Ig, blocks CD28 costimulation)
  • JAK inhibitors: Tofacitinib, baricitinib (small molecules)
• Corticosteroids: Bridge therapy, joint injections
• Surgery: Joint replacement for damaged joints

---

ILO 31 - Ankylosing Spondylitis (AS)

Key Features

• Prototype seronegative spondyloarthropathy
• HLA-B27 positive in ~90% (strongest HLA-disease association known)
• M:F = 2-3:1; onset typically 15-35 years

Aetiopathogenesis

• HLA-B27 presents arthritogenic peptide to CD8+ T cells
• OR misfolded HLA-B27 triggers ER stress → UPR → IL-23 induction
• Gut dysbiosis → microbial translocation → immune activation
• IL-17 and IL-23 pathway prominent → new therapeutic target

Clinical Manifestations

• Inflammatory back pain: Insidious onset, worse in morning, improves with exercise, does not improve with rest
• Sacroiliitis (bilateral) - pathognomonic; butt/groin pain, worse at night
• Spinal involvement: Progressive - lumbar → thoracic → cervical
• Bamboo spine: Late finding on X-ray - complete fusion of vertebrae by syndesmophytes (bone bridges)
• Peripheral arthritis: Large joints (hip, shoulder)
• Enthesitis: Inflammation at tendon/ligament insertions (heel pain = Achilles enthesitis; plantar fasciitis)
• Uveitis (iritis): Most common extra-articular feature (25-30%); acute, unilateral, painful red eye
• Others: Aortitis (aortic regurgitation), cardiac conduction defects, apical pulmonary fibrosis, cauda equina syndrome (rare)

Assessment Criteria

• BASDAI (Bath AS Disease Activity Index) - patient-reported
• Modified New York Criteria (1984):
  • Definite AS = bilateral sacroiliitis Grade ≥2 + one clinical criterion
  • OR unilateral sacroiliitis Grade 3-4 + one clinical criterion
  • Clinical criteria: Low back pain >3 months; limited lumbar motion; limited chest expansion

Associated Conditions

• Reactive arthritis (Reiter syndrome = AS + urethritis + conjunctivitis + triggered by infection)
• Psoriatic arthritis, IBD-associated arthritis
• All seronegative (RF negative)

Management

• NSAIDs (mainstay of treatment; actually slow radiographic progression)
• Exercise and physiotherapy (maintain posture/flexibility)
• Anti-TNF biologics (infliximab, etanercept, adalimumab) - for inadequate NSAID response
• IL-17 inhibitors (secukinumab, ixekizumab) - highly effective
• IL-23 inhibitors (risankizumab)
• Sulfasalazine - only for peripheral arthritis; NO benefit for axial disease
• Corticosteroids NOT effective for axial disease

---

ILO 32 - Systemic Sclerosis (Scleroderma)

Key Concept

• Autoimmune disease characterized by: (1) Vasculopathy, (2) Autoimmunity, (3) Fibrosis
• Fibroblast activation → excessive collagen deposition throughout body

Two Main Subtypes

Pathogenesis

1. Raynaud's phenomenon (earliest feature) - episodic vasospasm of small vessels → white → blue → red fingers/toes on cold or stress
2. Vascular injury → endothelial activation → platelet aggregation → intimal proliferation → vessel narrowing
3. Immune activation → T cell infiltration of skin → cytokines (TGF-β, IL-4, IL-13) → fibroblast activation
4. Fibrosis → collagen deposition → skin thickening, organ fibrosis

Clinical Features

• Skin: Tight, thickened skin; sclerodactyly; Raynaud's; calcinosis; telangiectasia; mat telangiectasias
• GI: Esophageal dysmotility (most common GI manifestation; GERD, dysphagia); small bowel malabsorption; wide-mouthed diverticula
• Lung: ILD (interstitial lung disease) - most common cause of death in diffuse SSc; PAH in limited SSc
• Kidney: Scleroderma renal crisis - acute renal failure + malignant hypertension; treat with ACE inhibitors
• Heart: Myocardial fibrosis, pericarditis, conduction defects
• MSK: Arthralgias, myopathy, tendon friction rubs (pathognomonic)

Management

• Raynaud's: CCBs (nifedipine), phosphodiesterase inhibitors
• ILD: Mycophenolate, cyclophosphamide, nintedanib
• PAH: Endothelin antagonists (bosentan), PDE-5 inhibitors, prostacyclins
• Renal crisis: ACE inhibitors (captopril) - life-saving
• GERD: PPIs
• No treatment definitively reverses fibrosis

---

PART 6: IMMUNODEFICIENCY DISORDERS

ILO 33 - Immunosuppressant Drug Classification

IMMUNOSUPPRESSANTS
│
├── CORTICOSTEROIDS
│   └── Prednisone, methylprednisolone, dexamethasone
│       → Broad anti-inflammatory; inhibit IL-2, NF-κB, cytokine production; lyse lymphocytes
│
├── CALCINEURIN INHIBITORS
│   ├── Cyclosporine (Ciclosporin A) → blocks calcineurin → no IL-2 transcription
│   └── Tacrolimus (FK506) → same mechanism; 100× more potent
│   → Side effects: Nephrotoxicity, hypertension, neurotoxicity, diabetes (tacrolimus)
│
├── mTOR INHIBITORS
│   ├── Sirolimus (Rapamycin) → blocks mTOR → T cell not proliferating even with IL-2
│   └── Everolimus
│   → Side effects: Hyperlipidemia, poor wound healing, thrombocytopenia
│
├── ANTIMETABOLITES
│   ├── Mycophenolate mofetil (MMF) → inhibits IMPDH → blocks guanosine synthesis → lymphocytes can't proliferate
│   ├── Azathioprine → metabolized to 6-MP → purine analog → blocks DNA synthesis
│   └── Methotrexate → inhibits DHFR → blocks folate → reduces lymphocyte and other rapidly dividing cells
│
├── ALKYLATING AGENTS
│   └── Cyclophosphamide → cross-link DNA → kill dividing lymphocytes
│       → Used in severe autoimmune disease (lupus nephritis, vasculitis, GPA)
│
├── BIOLOGIC AGENTS
│   ├── Anti-CD20: Rituximab → deplete B cells
│   ├── Anti-TNF: Infliximab, etanercept, adalimumab → RA, AS, IBD, psoriasis
│   ├── Anti-IL-6R: Tocilizumab → RA, cytokine storm
│   ├── CTLA-4-Ig (Abatacept/Belatacept) → block T cell costimulation
│   ├── Anti-CD25 (Basiliximab) → block IL-2 receptor → transplant induction
│   └── ATG (Anti-thymocyte globulin) → deplete T cells → transplant induction, aplastic anemia
│
└── NEWER SMALL MOLECULES
    └── JAK inhibitors: Tofacitinib, baricitinib, ruxolitinib → block cytokine signaling pathways

---

ILO 34-35 - Immunodeficiency Disorders

Classification Diagram (ILO 34)

IMMUNODEFICIENCY DISORDERS
│
├── PRIMARY (Congenital/Genetic)
│   │
│   ├── COMBINED (B + T cell defects)
│   │   ├── Severe Combined Immunodeficiency (SCID)
│   │   │   ├── X-linked (most common) - γc chain mutation (affects IL-2R, IL-4R, etc.)
│   │   │   ├── Adenosine deaminase (ADA) deficiency
│   │   │   └── Purine nucleoside phosphorylase (PNP) deficiency
│   │   └── Wiskott-Aldrich Syndrome (WASp mutation; triad: eczema + thrombocytopenia + immunodeficiency)
│   │
│   ├── PREDOMINANTLY T CELL DEFECTS
│   │   └── DiGeorge Syndrome (22q11.2 deletion; thymic aplasia; hypoparathyroidism, conotruncal heart defects)
│   │
│   ├── PREDOMINANTLY B CELL/ANTIBODY DEFECTS
│   │   ├── X-linked Agammaglobulinemia (XLA / Bruton's disease) - BTK mutation; no B cells after 6 months
│   │   ├── Common Variable Immunodeficiency (CVID) - defective B cell differentiation; low all Ig
│   │   ├── Selective IgA deficiency - most common primary immunodeficiency; mucosal infections
│   │   └── Hyper-IgM syndrome - CD40L mutation; no class switching; elevated IgM, low IgG/A/E
│   │
│   ├── PHAGOCYTE DEFECTS
│   │   ├── Chronic Granulomatous Disease (CGD) - NADPH oxidase deficiency; can't kill catalase+ organisms
│   │   ├── Leukocyte Adhesion Deficiency (LAD) - CD18 mutation; neutrophils can't migrate; delayed cord separation
│   │   └── Chediak-Higashi - LYST mutation; defective granule formation; partial albinism, neuropathy
│   │
│   └── COMPLEMENT DEFICIENCIES
│       ├── C1q/C1r/C1s/C2/C4 → SLE-like, poor immune complex clearance
│       ├── C3 → Recurrent pyogenic infections (most severe)
│       ├── C5-C9 → Recurrent Neisseria infections
│       └── C1-inhibitor → Hereditary angioedema
│
└── SECONDARY (Acquired)
    ├── HIV/AIDS (most important)
    ├── Malnutrition (most common cause worldwide)
    ├── Malignancy (lymphoma, leukemia, myeloma)
    ├── Drugs (immunosuppressants, chemotherapy, steroids)
    ├── Irradiation
    ├── Splenectomy (↑ risk encapsulated organisms: S. pneumoniae, H. influenzae, N. meningitidis)
    └── Chronic diseases (DM, renal failure, liver cirrhosis)

Primary Immunodeficiency - Pathogenesis and Clinical Consequences (ILO 35)

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PART 7: HIV/AIDS

ILOs 36-41 - HIV Infection and AIDS

Epidemiology and Risk Factors (ILO 36)

• Global: ~39 million people living with HIV worldwide (UNAIDS 2023)
• Sub-Saharan Africa most affected (most prevalent)
• Risk factors:
  • Unprotected sexual intercourse (most common in heterosexual transmission globally; MSM in developed countries)
  • Intravenous drug use (needle sharing)
  • Blood transfusions/clotting factors (now rare with testing)
  • Mother-to-child (vertical) transmission
  • Healthcare worker needlestick (low risk ~0.3%)
• Agent: HIV-1 (worldwide) and HIV-2 (West Africa, less pathogenic)
• HIV is a lentivirus (slow retrovirus) belonging to Retroviridae

Structure of HIV

• Envelope: Lipid bilayer derived from host cell membrane
• gp120: Surface glycoprotein; binds CD4 + CCR5/CXCR4
• gp41: Transmembrane; mediates membrane fusion
• Inside: Cone-shaped capsid (p24) containing:
  • 2 copies of single-stranded RNA genome
  • Reverse transcriptase, integrase, protease

Mode of Transmission (ILO 37)

1. Sexual contact - vaginal and anal intercourse (anal receptive = highest risk)
2. Blood/Blood products - IV drug use, transfusion, occupational
3. Vertical (Mother-to-child):
   • Transplacental (in utero)
   • During delivery (intrapartum) - most common
   • Breastfeeding (postnatal)
   • Prevention: Antiretroviral therapy during pregnancy; caesarean section; avoid breastfeeding; neonatal ART prophylaxis

• Acute seroconversion: 2-4 weeks after infection
• Clinical latency (asymptomatic phase): Median 10 years (untreated)
• AIDS: Median 10-11 years after infection (untreated)

Replication Cycle of HIV (ILO 37)

1. ATTACHMENT & ENTRY
   gp120 binds CD4 → conformational change → binds CCR5 or CXCR4
   gp41 fusion peptide inserts → membrane fusion → viral core enters cell

2. REVERSE TRANSCRIPTION (cytoplasm)
   RNA → DNA (by reverse transcriptase, error-prone → high mutation rate)
   Reverse transcriptase inhibitors (NRTIs, NNRTIs) act here

3. INTEGRATION
   Viral DNA transported to nucleus → INTEGRASE integrates proviral DNA into host genome
   Integrase inhibitors (INSTIs: raltegravir) act here

4. TRANSCRIPTION
   Host RNA polymerase transcribes proviral DNA → viral mRNA and genomic RNA

5. TRANSLATION
   Viral polyproteins synthesized (Gag, Pol, Env)

6. ASSEMBLY
   Viral proteins and RNA assemble at membrane → budding

7. MATURATION
   Viral PROTEASE cleaves polyproteins into functional units
   Protease inhibitors act here → immature, non-infectious virions produced

8. BUDDING
   Mature virion released → infects new cells

Pathogenesis of AIDS (ILO 38)

1. Direct killing: Viral replication → cell lysis
2. gp120 on surface of infected cell + free gp120 binds CD4 on adjacent cells → syncytia formation and death
3. CTL killing: HIV-specific CD8+ CTLs kill infected CD4+ T cells
4. Apoptosis: Activation-induced cell death in uninfected bystander T cells
5. Bone marrow suppression: HIV infects progenitor cells → reduced T cell production
6. Lymphoid tissue destruction: Progressive disruption of lymph node architecture

• Also infected (via CCR5 predominantly)
• Serve as a reservoir for HIV (can't kill them as efficiently as T cells)
• Spread HIV to brain (microglia) → HIV encephalopathy
• Impaired antigen presentation → reduced adaptive immunity

• Normal: >500 cells/μL
• At risk for opportunistic infections: <200 cells/μL (= AIDS by definition)
• Very immunocompromised: <50 cells/μL

Natural History of HIV Infection (ILO 39)

PHASE 1: ACUTE HIV SYNDROME (2-4 weeks post-infection)
- "Mononucleosis-like" illness: fever, sore throat, lymphadenopathy, rash (maculopapular on trunk), myalgias
- High viremia, transient CD4 drop
- Spontaneously resolves in 2-4 weeks

PHASE 2: SEROCONVERSION (3-7 weeks)
- HIV-specific antibodies detected (ELISA turns positive)
- HIV-specific CD8+ CTLs appear → control initial viremia
- CD4 count returns toward normal (but never completely)
- Viral set point established

PHASE 3: CLINICAL LATENCY (Asymptomatic; median 10 years untreated)
- Continuous HIV replication in lymphoid tissue
- CD4 count slowly declines (50-100 cells/μL per year)
- CD4 in peripheral blood appears stable initially
- Persistent generalized lymphadenopathy may occur
- Patient may be asymptomatic

PHASE 4: SYMPTOMATIC HIV (CD4 200-500)
- Constitutional symptoms: Weight loss, fevers, night sweats, diarrhea
- Oral candidiasis, oral hairy leukoplakia (EBV)
- Herpes zoster, recurrent HSV
- Seborrheic dermatitis, molluscum

PHASE 5: AIDS (CD4 <200 cells/μL)
- Opportunistic infections and neoplasms
- CD4 <50 → CMV, MAC, toxoplasmosis, Cryptosporidiosis

AIDS-Defining Opportunistic Infections and Neoplasms (ILO 40)

HIV Diagnosis

• ELISA (4th generation): Screen for HIV-1/2 antibodies AND p24 antigen (detects earlier); ≥2 positive ELISAs → confirm
• Western blot: Confirmatory (detects bands for p24, gp41, gp120/160)
• HIV RNA (viral load): Quantitative PCR; used to monitor treatment response and disease progression
• CD4+ T cell count: Stage disease; <200 = AIDS; guide prophylaxis
• HIV resistance testing: Genotypic or phenotypic; before starting ART

Antiretroviral Drugs (ILO 41)

ART DRUG CLASSES:
│
├── NRTIs (Nucleoside/Nucleotide Reverse Transcriptase Inhibitors)
│   → Act as chain terminators of viral DNA synthesis
│   Examples: Zidovudine (AZT), Lamivudine (3TC), Tenofovir (TDF/TAF), Abacavir, Emtricitabine (FTC)
│   Side effects: Lactic acidosis, lipoatrophy, peripheral neuropathy, bone marrow suppression (AZT)
│
├── NNRTIs (Non-Nucleoside Reverse Transcriptase Inhibitors)
│   → Bind allosteric site on RT → block conformational change
│   Examples: Efavirenz, Nevirapine, Rilpivirine
│   Side effects: Rash (Stevens-Johnson with nevirapine), CNS effects (efavirenz), teratogenicity
│
├── PIs (Protease Inhibitors)
│   → Block viral protease → non-infectious virions
│   Examples: Lopinavir/ritonavir (LPV/r), Atazanavir, Darunavir
│   Boosted with Ritonavir (blocks CYP3A4)
│   Side effects: Lipodystrophy, hyperlipidemia, hyperglycemia, GI upset
│
├── INSTIs (Integrase Strand Transfer Inhibitors) - now PREFERRED class
│   → Block viral integrase → prevent proviral DNA integration
│   Examples: Raltegravir, Elvitegravir, Dolutegravir, Bictegravir, Cabotegravir
│   Excellent tolerability, high barrier to resistance (dolutegravir, bictegravir)
│
├── Entry Inhibitors
│   ├── Fusion inhibitors: Enfuvirtide (T-20) - blocks gp41 fusion
│   └── CCR5 antagonists: Maraviroc - blocks CCR5 coreceptor (only for R5-tropic virus)
│
└── NRTI Boosters (Pharmacokinetic Enhancers)
    └── Cobicistat, Ritonavir (boost PI and INSTI levels by inhibiting CYP3A4)

• 2 NRTIs + 1 INSTI (e.g., Tenofovir + Emtricitabine + Dolutegravir = TDF/FTC/DTG)
• Goal: Undetectable viral load (<50 copies/mL) → immune reconstitution → prevent opportunistic infections

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PART 8: AMYLOIDOSIS

ILOs 42-46 - Amyloidosis

Definition (ILO 42)

Biochemistry of Amyloid

• ~95% of deposits = non-branching fibrils with β-pleated sheet conformation (all amyloid types share this structure)
• ~5% = SAP (Serum Amyloid P component) + glycosaminoglycans
• The β-pleated sheet conformation gives amyloid its characteristic staining properties

Classification and Types (ILO 43)

AMYLOIDOSIS - CLASSIFICATION BY PROTEIN TYPE

├── AL (Amyloid Light chain)
│   ├── Protein: Immunoglobulin LIGHT CHAINS (κ or λ) from plasma cells
│   ├── Clinical condition: Multiple myeloma, monoclonal gammopathy of unknown significance (MGUS)
│   ├── Type: PRIMARY AMYLOIDOSIS (no preceding inflammatory disease)
│   └── Distribution: Systemic; heart, kidney, liver, GI, tongue, peripheral nerves
│
├── AA (Amyloid Associated / Serum Amyloid A)
│   ├── Protein: SAA protein (acute phase protein made by liver) → cleaved to AA
│   ├── Clinical condition: CHRONIC INFLAMMATORY CONDITIONS (RA, Crohn's, TB, osteomyelitis, FMF)
│   ├── Type: REACTIVE SYSTEMIC AMYLOIDOSIS (secondary amyloidosis)
│   └── Distribution: Kidneys (most commonly), liver, spleen, adrenals
│
├── Aβ (Beta-amyloid)
│   ├── Protein: Aβ from amyloid precursor protein (APP) → cleaved by γ-secretase and β-secretase
│   ├── Clinical condition: ALZHEIMER'S DISEASE; also Down syndrome (trisomy 21 → 3 copies APP gene)
│   └── Distribution: Cerebral plaques (neuritic plaques), blood vessel walls (cerebral amyloid angiopathy)
│
├── ATTR (Transthyretin Amyloid)
│   ├── Protein: Transthyretin (TTR) = transport protein for thyroxine and retinol
│   ├── Sub-types:
│   │   ├── Familial (hATTR): Mutant TTR gene → autosomal dominant; peripheral neuropathy, cardiomyopathy
│   │   │   Example: Val122Ile (more common in African Americans, 3.4%)
│   │   └── Wild-type (wtATTR = Senile Systemic Amyloidosis): Normal TTR in elderly men → cardiac
│   └── Distribution: Heart (cardiomyopathy), peripheral nerves
│
├── AΒFP (Familial amyloid polyneuropathy)
│   └── Included in hATTR category above
│
├── Aβ2M (β2-Microglobulin Amyloid)
│   ├── Protein: β₂-microglobulin accumulates in long-term hemodialysis patients (not cleared)
│   └── Distribution: Joints, synovium, carpal tunnel → carpal tunnel syndrome
│
└── LOCALIZED AMYLOIDOSIS
    ├── Cerebral: Alzheimer's disease
    ├── Endocrine: Medullary thyroid carcinoma (calcitonin → amyloid), Type 2 diabetes (islets → IAPP)
    ├── Skin: Macular and papular amyloidosis
    └── Cardiac: Senile cardiac amyloidosis (ATTR)

Clinical Syndromes by Type

Morphological Features and Clinical Consequences by Organ (ILO 44)

Kidneys (most common site in AA; also AL)

• Grossly: Large pale "waxy" kidneys
• Microscopically: Deposits in glomerular mesangium and basement membrane → glomerular obliteration → heavy proteinuria → nephrotic syndrome → renal failure
• Congo red stain: Positive in glomeruli, vessels, tubular BM

Heart

• Grossly: Stiff, rubbery myocardium; ventricular wall thickening; "sparkling" appearance on echo
• Micro: Deposits between myocardial fibers → infiltrative cardiomyopathy
• Clinical: Restrictive cardiomyopathy (most common cardiac finding) → diastolic dysfunction, heart failure
• ECG: Low voltage despite wall thickening (classic ECG-echo discordance)
• AL and ATTR are the most common cardiac types

Liver

• Grossly: Hepatomegaly; pale, waxy ("lardaceous liver")
• Micro: Deposits in space of Disse (between hepatocytes and sinusoids), later periportal areas
• Clinical: Hepatomegaly, elevated ALP; rarely liver failure

Spleen

• "Sago spleen": Deposits in follicles → grey nodules resembling sago grains
• "Lardaceous spleen": Extensive deposits involving sinuses and red pulp → diffuse waxy appearance

GI Tract

• Deposits in vessel walls and muscularis → macroglossia (enlarged tongue - CLASSIC in AL), malabsorption, bleeding, perforation
• Macroglossia (large tongue) = PATHOGNOMONIC of systemic AL amyloidosis

Peripheral Nerves

• Sensorimotor peripheral neuropathy, autonomic neuropathy (postural hypotension, impotence, bladder dysfunction)

Brain (Alzheimer's)

• Neuritic (senile) plaques: Extracellular Aβ deposits + dying neurites + microglia/astrocytes
• Neurofibrillary tangles: Intracellular tau protein aggregates

Laboratory Diagnosis (ILO 45)

Tissue Biopsy and Staining

• Congo red stain:
  • Light microscopy: Amyloid stains pink/salmon-red
  • Polarized light: Apple-green birefringence - PATHOGNOMONIC
  • This is the gold-standard histological test
• Electron microscopy: Straight, non-branching fibrils ~8-10 nm diameter (β-pleated sheet conformation)
• Thioflavin T stain: Fluorescence under UV - yellow-green fluorescence; more sensitive but less specific than Congo red
• PAS stain: Weakly positive (glycoproteins in deposit)
• Crystal violet / Methyl violet: Metachromatic staining - amyloid stains purple/pink (instead of expected violet)

Biopsy Sites (in order of preference)

1. Abdominal fat pad aspiration - safest, easy, positive in ~80% of systemic amyloidosis
2. Rectal biopsy - positive in ~70-80%
3. Bone marrow biopsy (if myeloma suspected)
4. Renal biopsy (if renal involvement)
5. Liver/gingival biopsy

• Direct biopsy of affected organ (liver, kidney, heart) = most diagnostic but risky

Blood/Urine Tests

• Serum/urine protein electrophoresis + immunofixation → detect M-protein in AL amyloidosis
• Free light chain assay (FLC) → detect monoclonal Ig light chains (AL)
• Serum SAA level → elevated in reactive systemic amyloidosis
• Genetic testing → TTR gene mutations in familial amyloidosis
• SAP scintigraphy (123I-labeled SAP): SAP binds to amyloid deposits → whole body scan shows extent of amyloid; available in specialist centers
• Mass spectrometry on paraffin-embedded tissue: Identify exact amyloid protein type
• Echo/MRI: Restrictive cardiomyopathy features; late gadolinium enhancement (MRI) in cardiac amyloid
• Technetium-pyrophosphate (99mTc-PYP) scan: Highly specific for ATTR cardiac amyloidosis

Histology Identification (ILO 46)

1. Look for amorphous, eosinophilic, homogeneous extracellular deposits in H&E
2. Deposits appear "glassy" and push tissue aside
3. Confirm with Congo red → pink/red on light microscopy
4. Confirm with polarized light → apple-green birefringence ← KEY EXAM ANSWER
5. In kidney: deposits in glomeruli (mesangium, GBM)
6. In heart: deposits between myocardial fibers
7. In liver: deposits in space of Disse

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QUICK REFERENCE: KEY COMPARISONS AND FLOWCHARTS

Innate vs. Adaptive Immunity

CD4+ vs CD8+ T Cells

4 Types of Hypersensitivity - Quick Summary

Key Autoantibodies Summary

Transplant Rejection Summary Table

Amyloid Types Quick Reference

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Exam strategy tips:

• Morphology questions: Always associate Congo red + apple-green birefringence with amyloid; wire-loop lesions with SLE Class IV; pannus with RA; bamboo spine with AS
• Antibody questions: The "most specific" is always the higher-yield answer (anti-Sm and anti-dsDNA for SLE; anti-CCP for RA; anti-Scl-70 for diffuse SSc)
• Flowchart processes (complement, HIV replication cycle, antigen processing) are frequently asked as "which step does drug X inhibit" - know where in the pathway each drug acts
• For HIV: CD4 count thresholds and what infection occurs at each level is very testable
• For hypersensitivity: Know the Gell & Coombs classification cold - Type I = IgE/mast cells, II = cytotoxic Ab, III = immune complex, IV = T cells