What is the workup for elevated iron levels with negative HFE (Human Ferritin Encoding) testing in family medicine?

Workup for Elevated Iron with Negative HFE Testing

When HFE testing is negative despite elevated iron parameters, you must systematically evaluate for secondary iron overload disorders and non-iron causes of hyperferritinemia, prioritizing transferrin saturation interpretation and excluding common secondary causes before pursuing advanced genetic testing. 1, 2, 3

Initial Assessment and Interpretation

Confirm the Pattern of Iron Elevation

• Measure fasting transferrin saturation (TS) alongside ferritin to determine if true iron overload exists—this is the single most important distinguishing test. 1, 2

• If TS ≥45%: True iron overload is likely present despite negative HFE testing, requiring evaluation for non-HFE hemochromatosis. 1, 3

• If TS <45%: Iron overload is unlikely; the elevated ferritin represents secondary hyperferritinemia from inflammation, liver disease, or other causes. 1, 2

Critical Pitfall to Avoid

• Never rely on ferritin alone to diagnose iron overload—ferritin is an acute phase reactant that rises with inflammation, infection, liver disease, malignancy, and tissue necrosis independent of actual iron stores. 1, 2

Workup Algorithm Based on Transferrin Saturation

Pathway A: TS ≥45% (True Iron Overload Despite Negative HFE)

This pattern suggests non-HFE hemochromatosis or secondary iron overload disorders. 1, 3

Evaluate for Secondary Iron Overload Causes:

• Hematologic disorders: 3, 4

  • Thalassemia syndromes (check CBC with RBC indices, hemoglobin electrophoresis)
  • Myelodysplastic syndrome (check CBC with differential, peripheral smear)
  • Myelofibrosis (check CBC, peripheral smear, consider bone marrow biopsy)
  • Sideroblastic anemia (check peripheral smear, bone marrow if indicated)
  • Sickle cell disease (hemoglobin electrophoresis)
  • Pyruvate kinase deficiency (enzyme assay if suspected)

• Iatrogenic causes: 3

  • History of chronic transfusions
  • Excessive iron supplementation (oral or IV)
  • Long-term hemodialysis with IV iron

• Chronic liver diseases causing iron accumulation: 2, 3

  • Chronic viral hepatitis B or C (check HBsAg, anti-HCV, HCV RNA)
  • Alcoholic liver disease (detailed alcohol history, AST/ALT ratio typically >2:1)
  • Non-alcoholic fatty liver disease/metabolic syndrome (assess BMI, glucose, lipids, insulin resistance)
  • Porphyria cutanea tarda (check urine porphyrins)

Consider Non-HFE Genetic Hemochromatosis:

• If secondary causes are excluded and TS remains ≥45%, consider genetic testing for non-HFE hemochromatosis genes: 1, 3, 4
  • TFR2 mutations (Type 3 hemochromatosis)
  • SLC40A1 mutations (Type 4 hemochromatosis/ferroportin disease)
  • HAMP mutations (Type 2B hemochromatosis)
  • HJV mutations (Type 2A/juvenile hemochromatosis—especially if age <30 with cardiac or endocrine manifestations)

Quantify Hepatic Iron Content:

• MRI with quantitative assessment of hepatic iron concentration is recommended to confirm iron overload and guide management. 1, 3

• Liver biopsy should be considered if: 1, 3

  • Ferritin >1,000 μg/L with elevated liver enzymes (ALT/AST)
  • Platelet count <200 (suggests cirrhosis risk)
  • MRI results are equivocal
  • Need to assess for concurrent liver disease (e.g., autoimmune hepatitis can mimic iron overload with elevated TS)

Pathway B: TS <45% (Secondary Hyperferritinemia)

This pattern indicates the elevated ferritin is NOT due to iron overload—over 90% of cases fall into this category. 2

Systematically Exclude Common Causes:

• Chronic alcohol consumption: 2

  • Detailed quantitative alcohol history (grams/day, duration)
  • AST/ALT ratio typically >2:1 in alcoholic liver disease

• Inflammatory conditions: 2

  • Check CRP, ESR to detect occult inflammation
  • Consider rheumatologic conditions (RF, ANA, anti-CCP if joint symptoms)
  • Adult-onset Still's disease if extremely elevated ferritin (>4,000 ng/mL) with fever, rash, arthritis—check glycosylated ferritin fraction (<20% is 93% specific)

• Liver disease: 2, 3

  • Check comprehensive metabolic panel with ALT, AST, alkaline phosphatase, bilirubin, albumin
  • Viral hepatitis serologies (HBsAg, anti-HCV)
  • NAFLD/metabolic syndrome assessment (BMI, glucose, lipids, ultrasound)
  • Consider autoimmune hepatitis (ANA, ASMA, IgG levels, liver biopsy if indicated)

• Malignancy: 2

  • Age-appropriate cancer screening
  • Consider solid tumors, lymphomas, hepatocellular carcinoma (AFP, imaging)

• Infections: 2

  • Active infection causes acute ferritin rise—evaluate for occult infections

• Cell necrosis: 2

  • Muscle injury (check CK)
  • Hepatocellular necrosis (check ALT, AST)

• Chronic kidney disease: 2

  • Check creatinine, eGFR
  • In CKD, ferritin 100-700 ng/mL with TS <20% may represent functional iron deficiency

• Metabolic syndrome/NAFLD: 2

  • Ferritin elevation reflects hepatocellular injury and insulin resistance, not iron overload
  • Assess for obesity, diabetes, dyslipidemia, hypertension

Risk Stratification by Ferritin Level

• Ferritin <1,000 μg/L: Low risk of organ damage; negative predictive value of 94% for advanced liver fibrosis. 1, 2

• Ferritin 1,000-10,000 μg/L: Higher risk of advanced fibrosis/cirrhosis if true iron overload is present; requires additional evaluation with platelet count and liver enzymes. 1, 2

• Ferritin >10,000 μg/L: Rarely represents simple iron overload; requires urgent specialist referral to evaluate for life-threatening conditions (hemophagocytic lymphohistiocytosis, severe sepsis, malignancy). 2

Management Based on Findings

If Secondary Iron Overload is Confirmed:

• Treat the underlying hematologic disorder (e.g., manage transfusion-dependent anemia, consider chelation therapy if transfusion-related iron overload). 3

• Therapeutic phlebotomy may be considered if iron overload is documented by hepatic iron concentration measurement, regardless of genetic cause. 3

If Secondary Hyperferritinemia is Confirmed:

• Treat the underlying condition, not the elevated ferritin: 2

  • Weight loss and metabolic syndrome management for NAFLD
  • Alcohol cessation for alcoholic liver disease
  • Disease-specific anti-inflammatory therapy for rheumatologic conditions
  • Oncologic treatment for malignancy

• Do NOT initiate phlebotomy when TS <45%, as iron overload is not present. 1, 2

Key Clinical Pearls

• Combined interpretation of TS and ferritin is essential—never rely on a single test. 1

• In the general population, iron overload is NOT the most common cause of elevated ferritin—secondary causes account for >90% of cases. 2

• Ferritin >1,000 μg/L with elevated ALT/AST and platelet count <200 predicts cirrhosis in 80% of cases with true iron overload, warranting liver biopsy consideration. 1

• Autoimmune hepatitis can mimic iron overload syndrome with markedly elevated TS—liver biopsy can guide diagnosis. 5

• Screen first-degree relatives only if hereditary hemochromatosis (HFE or non-HFE) is confirmed. 1