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Serological Tests for Hepatitis B

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The Key Markers

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Individual Marker Details

HBsAg

• The principal screening test for HBV infection
• Appears approximately 1 month after infection (~4 weeks), detected as early as 1-2 weeks and as late as 11-12 weeks
• HBV DNA can actually be detected 2 weeks before HBsAg appears
• Persistence beyond 6 months = chronic infection
• Positive in both acute and chronic infection

Anti-HBs

• Indicates immunity - either from natural recovery or vaccination
• Appears after HBsAg disappears (following a gap called the "window period")
• The only anti-HBV antibody that is protective/neutralizing
• After vaccination: anti-HBs is present but anti-HBc is absent

Anti-HBc (IgM and IgG)

• IgM anti-HBc: The first antibody detected; present during acute infection; appears at onset of symptoms; may be the only marker detectable during the window period
• IgG anti-HBc: Replaces IgM with recovery; persists for life as a marker of prior exposure
• Anti-HBc is not neutralizing and does not confer immunity
• (Note: IgM anti-HBc may reappear during acute reactivation of chronic hepatitis B)

HBeAg

• Signifies active viral replication
• Presence = high HBV DNA levels (up to >10⁹ IU/mL)
• Absence + anti-HBe = lower replication (but can still be high in precore/BCP mutants)

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Serological Course: Acute HBV Infection with Resolution

1. HBV DNA rises first (detectable ~2 weeks before HBsAg)
2. HBsAg and HBeAg appear (weeks 4-8)
3. ALT and jaundice peak after HBV DNA peaks (~week 12)
4. IgM anti-HBc appears at symptom onset - may be the only marker in the window period
5. HBsAg and HBeAg disappear (~week 20)
6. Anti-HBe appears
7. Window period: HBsAg gone, anti-HBs not yet present - only anti-HBc detectable
8. Anti-HBs appears (~week 32+) - signals recovery and immunity
9. IgM anti-HBc replaced by IgG anti-HBc (persists for life)

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Serological Course: Progression to Chronic Infection

• HBsAg persists beyond 6 months
• HBV DNA remains detectable
• IgM anti-HBc wanes; total anti-HBc persists
• Anti-HBs never develops in chronic infection

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Pattern-Based Interpretation Table

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Isolated Anti-HBc (Anti-HBc Alone)

1. Resolved infection (most common) - anti-HBs has waned over time
2. False-positive anti-HBc - patient is actually susceptible
3. Low-level chronic carrier
4. Resolving acute infection (window period)

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HBV DNA (Virological Marker)

• Provides the most direct measure of viral replication
• Measured in IU/mL; levels fluctuate in chronic HBV
• Threshold of 2,000 IU/mL is the key cut-off used in most chronic HBV guidelines
• HCC risk increases dramatically when baseline HBV DNA exceeds ~2,000 IU/mL
• Serial testing is important for management decisions and treatment monitoring

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Newer Markers

Quantitative HBsAg

• HBV produces >1,000-fold more surface proteins than intact virions
• Reflects transcriptional activity of covalently closed circular DNA (cccDNA)
• Cut-off of 1,000 IU/mL proposed to differentiate HBeAg-negative chronic hepatitis from inactive carrier state
• Better predictor of HCC and liver-related mortality than HBV DNA in HBeAg-negative patients with low viremia (<2,000 IU/mL)

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Quick Summary: Clinical Status vs. Test Results

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Pathogenesis of Atherosclerosis (Robbins & Cotran)

The "Response-to-Injury" Hypothesis

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Step-by-Step Pathogenesis

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Step 1 - Endothelial Injury and Dysfunction

• Increased permeability
• Enhanced leukocyte adhesion
• Altered gene expression

Laminar flow normally upregulates Krüppel-like factor-2 (KLF2) which switches on atheroprotective genes. Statins also exert some of their benefit through KLF2 upregulation.

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Step 2 - Lipid Accumulation (Hypercholesterolemia)

1. The dominant lipids in atheromatous plaques are cholesterol and cholesterol esters
2. Genetic defects in lipoprotein uptake (e.g., familial hypercholesterolemia - loss-of-function LDL receptor mutations) cause accelerated atherosclerosis and MI before age 20 in homozygotes
3. Epidemiologic correlation between total plasma cholesterol/LDL and atherosclerosis severity
4. Lowering serum cholesterol slows progression, causes regression of some plaques, and reduces cardiovascular events

• Chronic hyperlipidemia → lipoproteins accumulate in the intima
• Lipoproteins aggregate or become oxidized by free radicals from inflammatory cells
• Modified (oxidized) LDL is taken up by macrophages via scavenger receptors (not the normal LDL receptor)
• Because scavenger receptor-mediated uptake cannot be downregulated, macrophages become overloaded → foam cells (lipid-filled macrophages)
• Smooth muscle cells (SMCs) similarly become foam cells via LDL receptor-related proteins
• Modified lipoproteins are also directly toxic to ECs, SMCs, and macrophages
• This stimulates release of growth factors, cytokines, and chemokines → vicious cycle of inflammation

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Step 3 - Inflammation and the Inflammasome

1. Cholesterol crystals and free fatty acids accumulate in macrophages
2. Sensed by cytosolic innate immune receptors → NLRP3 inflammasome assembly
3. Inflammasome activation → production of IL-1β and IL-18
4. IL-1 activates ECs and recruits/activates mononuclear cells (macrophages + T lymphocytes)
5. Activated macrophages produce ROS → further LDL oxidation, plus growth factors driving SMC proliferation
6. Activated T lymphocytes elaborate interferon-γ → activates macrophages, ECs, and SMCs
7. This creates a self-amplifying loop of inflammatory cell recruitment and vessel wall injury

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Step 4 - Monocyte Recruitment and Foam Cell Formation (Fatty Streak)

• Dysfunctional ECs express adhesion molecules (VCAM-1, ICAM-1, selectins) → monocytes and T lymphocytes adhere
• Monocytes migrate into the intima → differentiate into macrophages
• Macrophages and SMCs engulf oxidized LDL via scavenger receptors → transform into foam cells
• Foam cells accumulate in the intima → first visible lesion = fatty streak (appears as yellow streaks on intimal surface, near vessel ostia)
• Fatty streaks are present even in children and adolescents

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Step 5 - Smooth Muscle Cell Proliferation and ECM Synthesis (Plaque Progression)

• Growth factors (PDGF from platelets, macrophages, ECs, SMCs; FGF; TGF-α) recruit medial SMCs into the intima
• SMCs proliferate and synthesize ECM proteins (especially collagen) → forms the fibrous cap
• This converts a fatty streak into a mature fibrofatty atheroma
• The fibrous cap stabilizes the plaque
• Conversely, activated inflammatory cells release matrix metalloproteinases (MMPs) that degrade ECM → destabilizes plaques → prone to rupture

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Plaque Morphology (Mature Atheroma)

• Fibrous cap - SMCs embedded in dense collagen/ECM; surface covered by (often dysfunctional) ECs
• Necrotic/lipid core - extracellular lipid, cholesterol crystals, necrotic cell debris, foam cells
• Shoulders - region at plaque edges containing macrophages, T lymphocytes, SMCs
• Calcification - common in advanced plaques
• Neovascularization - new small vessels from adventitia (vasa vasorum) may supply the plaque

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Stable vs. Vulnerable (Unstable) Plaques

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Consequences of Atherosclerosis

1. Luminal narrowing - chronic ischemia (stable angina, claudication)
2. Plaque rupture/thrombosis - acute MI, stroke (most feared)
3. Atheroembolism - debris discharged into bloodstream causes microemboli
4. Aneurysm formation - ischemic atrophy of media with elastic tissue loss → wall weakening → dilation/rupture

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KEY CONCEPTS (Robbins Summary)

• Atherosclerosis is an intimal-based lesion with a fibrous cap and atheromatous core; constituents include SMCs, ECM, inflammatory cells, calcifications, lipids, and necrotic debris
• Atherogenesis is driven by interplay of vessel wall injury and inflammation
• All major risk factors converge on EC dysfunction and influence inflammatory cell and SMC recruitment
• Plaques develop slowly over decades; stable plaques cause chronic ischemia; unstable plaques cause acute catastrophic ischemic events

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Morphology of Cutaneous Squamous Cell Carcinoma (SCC)

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The Big Picture First

Master memory peg: "SCC = Squamous = Keratin-making cells gone rogue. The better they still make keratin, the more well-differentiated they are."

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GROSS MORPHOLOGY (What you see clinically)

SCC In Situ (Carcinoma in Situ / Bowen's disease)

• Sharply defined hyperkeratotic plaques or nodules
• Color ranges from erythematous to skin-colored
• Scaly, rough surface
• Often arise on top of prior actinic keratoses (sun-damaged precursors)

Invasive SCC

• Nodular, often scaly lesion
• Commonly ulcerates (classic "ulcerated nodule" appearance)
• Firm, indurated base
• Surrounding skin shows signs of sun damage (solar elastosis)
• Common sites: face, ears, lips, scalp, dorsum of hands - all sun-exposed

Memory trick for gross features: "NUSH" - Nodule → Ulcerates → Scaly surface → Hyperkeratotic (crusty top)

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MICROSCOPIC MORPHOLOGY

The Defining Feature

"Atypical keratinocytes originating in the epidermis and breaching the basement membrane to infiltrate the dermis."

Key Microscopic Features (the "must-know" list)

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THE GRADING SYSTEM (Broder's Grading)

Memory trick for grading: Think of it as "How much keratin is left?"

• Grade 1 = Lots of keratin (horn pearls everywhere) = "Keratinizing like crazy"
• Grade 4 = Almost no keratin (just single dying cells trying) = "Barely remembers it used to make keratin"

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THE HORN PEARL - The Star Feature ⭐

Memory trick: "A pearl forms by wrapping layers around a core - just like SCC cells wrap keratin around themselves in the dermis. The more pearl-forming, the more 'precious' (well-differentiated) the tumor."

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HISTOLOGIC VARIANTS (High-Yield)

Memory trick for variants - "A SAD V":

• A = Acantholytic (fake glands)
• S = Spindle cell (looks like sarcoma)
• A = Anaplastic/undifferentiated
• D = Desmoplastic (lots of stroma)
• V = Verrucous (wart-like, no pearls)

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THE COMPLETE MICROSCOPIC PICTURE - One Paragraph to Memorize

"SCC in situ shows highly atypical cells at ALL levels of the epidermis with nuclear crowding and disorganization - but still above the basement membrane. Invasive SCC shows atypical keratinocytes breaching the basement membrane and infiltrating the dermis in nests or cords. Well-differentiated tumors form prominent horn pearls with individual cell dyskeratosis and parakeratosis. Poorly differentiated tumors show marked nuclear pleomorphism, frequent mitoses, foci of necrosis, and only abortive single-cell keratinization."

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MASTER MEMORY FRAMEWORK

"KAPID" for the 5 core micro features:

• K = Keratin pearls (horn pearls) - the signature
• A = Atypical keratinocytes (pleomorphic nuclei, mitoses)
• P = Parakeratosis + individual cell dyskeratosis
• I = Invasion through basement membrane into dermis
• D = Differentiation determines grade (lots of K → Grade 1; no K → Grade 4)

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CLINICAL BEHAVIOR POINTS (for context)

• ~4% have already metastasized to regional lymph nodes at diagnosis
• Higher metastatic risk: lesions >2 cm, deep invasion below subcutaneous fat, perineural invasion, lesions near ear or lip
• SCC from actinic keratoses: locally aggressive but slow to metastasize
• SCC from burn scars, chronic ulcers, or non-sun-exposed skin: more aggressive
• Metastatic cutaneous SCC responds well to immune checkpoint inhibitors (due to high mutational burden from UV damage)