Definition/General
Introduction:
Hemophagocytosis represents a reactive phenomenon characterized by phagocytosis of hematopoietic cells by activated macrophages in bone marrow
It is a hallmark feature of hemophagocytic lymphohistiocytosis (HLH)
The condition results from immune dysregulation with excessive cytokine production
It can be primary (genetic) or secondary (acquired).
Origin:
Results from abnormal immune activation leading to excessive macrophage proliferation and activation
Genetic mutations in cytotoxic pathways (PRF1, UNC13D, STX11, STXBP2) cause primary forms
Secondary forms triggered by infections, malignancies, or autoimmune disorders
Leads to tissue infiltration by activated macrophages showing hemophagocytosis.
Classification:
Primary HLH: Familial hemophagocytic lymphohistiocytosis (FHL) types 1-5
Primary HLH: Immune deficiency syndromes (Griscelli syndrome, Chediak-Higashi syndrome)
Secondary HLH: Infection-associated (EBV, CMV, bacterial, fungal)
Secondary HLH: Malignancy-associated (lymphomas, leukemias)
Secondary HLH: Autoimmune-associated (macrophage activation syndrome).
Epidemiology:
Primary HLH: Peak incidence in infancy and early childhood
Secondary HLH: Can occur at any age
EBV-associated HLH common in Asian populations
Overall incidence: 1.2 cases per million children annually
Male-to-female ratio: 1.2:1
High mortality rate: 50-60% without treatment.
Clinical Features
Presentation:
Fever (>38.5°C for >7 days)
Hepatosplenomegaly (90% of cases)
Cytopenia affecting ≥2 cell lines
Neurological symptoms (seizures, altered consciousness)
Skin rash and lymphadenopathy
Coagulopathy and bleeding manifestations.
Symptoms:
Persistent high fever unresponsive to antibiotics
Progressive weakness and fatigue
Easy bruising and bleeding tendency
Abdominal distension due to organomegaly
Respiratory distress and cough
Neurological deterioration including seizures
Weight loss and failure to thrive in children.
Risk Factors:
Family history of HLH or unexplained infant deaths
Consanguinity in parents
Immunocompromised states
EBV infection especially in Asian populations
Malignancy particularly T-cell lymphomas
Autoimmune disorders (systemic lupus erythematosus, rheumatoid arthritis)
Recent infections or vaccinations.
Screening:
HLH-2004 diagnostic criteria (fever, splenomegaly, cytopenia, hypertriglyceridemia)
Laboratory workup: ferritin, fibrinogen, triglycerides
Flow cytometry for NK cell function and degranulation assays
Genetic testing for primary HLH genes
Infectious disease screening: EBV, CMV, bacterial cultures.
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Gross Description
Appearance:
Bone marrow aspirate shows increased cellularity with prominent macrophages
Hypercellular bone marrow with normal or increased megakaryocytes
Scattered large macrophages containing engulfed hematopoietic cells
Reduced fat spaces due to increased cellularity.
Characteristics:
Macrophages with abundant cytoplasm containing phagocytosed cells
Well-preserved cellular morphology of engulfed cells initially
Various stages of cellular digestion within macrophage cytoplasm
Increased reticulin fibrosis may be present in advanced cases.
Size Location:
Diffuse involvement of bone marrow spaces
Paratrabecular and intertrabecular distribution of hemophagocytic macrophages
Perivascular aggregation of activated macrophages
Focal to diffuse pattern depending on disease severity.
Multifocality:
Multifocal bone marrow involvement in most cases
Extramedullary hemophagocytosis in spleen, liver, lymph nodes
CNS involvement with hemophagocytic macrophages in cerebrospinal fluid
Systemic distribution affecting multiple organ systems.
Microscopic Description
Histological Features:
Activated macrophages with enlarged, vesicular nuclei and abundant cytoplasm
Hemophagocytosis: macrophages containing intact or partially digested blood cells
Erythrophagocytosis: red blood cells within macrophage cytoplasm
Leukophagocytosis: neutrophils and lymphocytes being phagocytosed
Thrombophagocytosis: platelets within macrophages.
Cellular Characteristics:
Large macrophages with kidney-shaped or multilobated nuclei
Abundant eosinophilic cytoplasm with foamy appearance
Prominent nucleoli and open chromatin pattern
Phagocytosed cells show various degrees of digestion
Hemosiderin deposits within macrophage cytoplasm.
Architectural Patterns:
Scattered distribution of hemophagocytic macrophages throughout marrow
Clustering pattern around blood vessels in some cases
Normal trilineage hematopoiesis with maturation preserved
Increased macrophage-to-other cell ratio
Reactive lymphocytosis may be present.
Grading Criteria:
Quantitative assessment: >2% hemophagocytic macrophages is significant
Mild hemophagocytosis: 2-5% of nucleated cells
Moderate hemophagocytosis: 5-10% of nucleated cells
Severe hemophagocytosis: >10% of nucleated cells
Clinical correlation essential as hemophagocytosis can be seen in other conditions.
Immunohistochemistry
Positive Markers:
CD68 (pan-macrophage marker)
CD163 (M2 macrophage marker, highly positive)
Lysozyme (strong cytoplasmic positivity)
Alpha-1-antitrypsin (cytoplasmic staining)
PGM1 (cytoplasmic positivity)
S-100 protein (variable positivity).
Negative Markers:
CD1a (negative, helps exclude Langerhans cell histiocytosis)
Langerin (negative)
CD21 (negative)
CD35 (negative)
Cytokeratin (negative)
Melanoma markers (negative).
Diagnostic Utility:
CD163 strongly positive in hemophagocytic macrophages
CD68/CD163 dual staining highlights activated macrophages effectively
Helps distinguish from other histiocytic disorders
Iron stain may show hemosiderin deposits
Reticulin stain may show increased fibrosis.
Molecular Subtypes:
Cytokine profile: elevated IL-1β, TNF-α, IL-18, interferon-γ
Soluble CD25 (sIL-2R) markedly elevated (>2400 U/mL)
Soluble CD163 significantly increased
NK cell activity decreased or absent
Degranulation markers (CD107a) reduced in primary HLH.
Molecular/Genetic
Genetic Mutations:
PRF1 mutations (perforin gene, FHL2, 20-50% of cases)
UNC13D mutations (Munc13-4 gene, FHL3, 20-30% of cases)
STX11 mutations (syntaxin 11 gene, FHL4, 10-20% of cases)
STXBP2 mutations (syntaxin binding protein 2, FHL5, 10-15% of cases)
RAB27A mutations (Griscelli syndrome type 2).
Molecular Markers:
Reduced perforin expression by flow cytometry or immunohistochemistry
Absent degranulation of NK cells and CD8+ T cells
Elevated ferritin levels (>500 ng/mL, often >10,000 ng/mL)
Hypofibrinogenemia (<1.5 g/L)
Hypertriglyceridemia (>3.0 mmol/L or >265 mg/dL).
Prognostic Significance:
Primary HLH has worse prognosis without hematopoietic stem cell transplantation
Secondary HLH prognosis depends on underlying trigger
CNS involvement indicates poor prognosis
Early diagnosis and treatment crucial for survival
Genetic testing important for family counseling and sibling screening.
Therapeutic Targets:
Immunosuppressive therapy: corticosteroids, cyclosporine A
Cytotoxic therapy: etoposide, doxorubicin
Anti-CD25 therapy: alemtuzumab in refractory cases
JAK inhibitors: ruxolitinib for secondary HLH
Hematopoietic stem cell transplantation for primary HLH
Supportive care: blood products, antimicrobials.
Differential Diagnosis
Similar Entities:
Langerhans cell histiocytosis (coffee-bean nuclei, CD1a+, Langerin+)
Rosai-Dorfman disease (emperipolesis, S-100+)
Infection-related reactive hemophagocytosis (less severe, self-limiting)
Malignancy-associated hemophagocytosis (underlying malignant cells present)
Storage diseases (Gaucher cells, specific enzyme deficiencies).
Distinguishing Features:
HLH hemophagocytosis: meets diagnostic criteria, severe systemic symptoms
Reactive hemophagocytosis: mild, self-limiting, no systemic criteria
Langerhans cell histiocytosis: CD1a+, Langerin+, coffee-bean nuclei
Rosai-Dorfman: S-100+, emperipolesis of lymphocytes
Storage diseases: specific cell morphology, enzyme studies.
Diagnostic Challenges:
Distinguishing primary from secondary HLH requires genetic testing and family history
Reactive hemophagocytosis vs true HLH based on clinical criteria
Identifying underlying trigger in secondary HLH
CNS involvement assessment may require CSF examination
Monitoring treatment response and disease progression.
Rare Variants:
Macrophage activation syndrome (MAS): associated with rheumatic diseases
EBV-associated HLH: common in East Asian populations
X-linked lymphoproliferative disease: SH2D1A or XIAP mutations
Periodic fever syndromes with secondary HLH
Malignancy-associated HLH: particularly with T-cell lymphomas.
Sample Pathology Report
Template Format
Sample Pathology Report
Complete Report: This is an example of how the final pathology report should be structured for this condition.
Specimen Information
Bone marrow aspirate and biopsy from [site], adequate for evaluation
Cellularity
Bone marrow cellularity: [percentage]% (age-adjusted normal: [range]%)
Hemophagocytosis
Hemophagocytic macrophages constitute [percentage]% of nucleated cells with phagocytosis of [cell types]
Morphological Features
Activated macrophages with [nuclear features] and [cytoplasmic characteristics]
Hematopoiesis
Trilineage hematopoiesis with [maturation pattern] and [blast percentage]%
Iron Stores
Iron stores: [increased/normal/decreased] with hemosiderin deposits in macrophages
Immunohistochemistry
CD68: [result], CD163: [result], supporting hemophagocytic macrophages
Final Diagnosis
Bone marrow with hemophagocytosis ([severity]), consistent with hemophagocytic lymphohistiocytosis
Recommendations
Clinical correlation with HLH diagnostic criteria. Consider genetic testing for primary HLH. Evaluate for underlying triggers