Chronic hepatitis C virus (HCV) infection is widely recognized as a cause of extrahepatic manifestations, including autoimmune disorders. ^{1 – 4} Autoimmune hemolytic anemia (AIHA) is rare. Warm AIHA with HCV infection in treatment-naïve patients has been described either during interferon treatment or in treatment-naive patients. ^{4 – 7} However, the association between AIHA due to cold agglutinin and untreated hepatitis C seems to be a rare situation, to the best of our knowledge only two cases have been reported. ^{8 , 9} Herein, we present a case of HVC infection complicated by cold AIHA.

A 49-year-old man with a history of HVC diagnosed two months ago, presented with dyspnea and pallor. HCV belonged to the type 1a genotype and the viral load was 1.6 10 ⁶ UI/ml. Fibroscan showed F2 hepatic fibrosis stage. No evidence of recent drug use was found. On admission, the patient was pale, polypneic, and tachycardic with mild hepatomegaly. The patient did not have splenomegaly or lymphadenopathy. Laboratory tests showed severe anemia with a low hemoglobin (Hb) level (4.2 g/dl), macrocytosis (MCV 120 fL), reticulocytosis (5.2%), and normal white blood cell and platelet counts. A peripheral blood smear showed anisocytosis and polychromasia with rare schizocytes. Hemolysis workup revealed elevated indirect bilirubin (84 μmol/l), high lactate dehydrogenase (LDH) (580 U/L), and low haptoglobin concentration (0.15 g/l). These findings were consistent with the diagnosis of hemolytic anemia.

The presence of anti-complement C3d++ was confirmed using a monospecific direct antiglobulin test (DAT- Low Ionic Strength Solution). Anti-immunoglobulin G antibodies were absent. Screening of the eluate was negative, and the cold agglutinin titer was 1/16 at 4°C. Hemoglobin electrophoresis results were normal. A bone marrow biopsy was performed to elucidate the underlying etiology of the patient’s AIHA and showed erythroid hyperplasia with no evidence of tumor lymphoma. Further investigations, including anti-smooth muscle, antinuclear antibodies, and anti-DNA antibodies, were negative. However, type III mixed cryoglobulinemia was also observed. Thoracic-abdominal computed tomography (CT) indicated that the underlying malignancy was ruled out. Serology results for hepatitis B and human immunodeficiency virus (HIV) were also negative. Based on the above findings, a diagnosis of AIHA due to cold agglutinin complicating untreated chronic hepatitis C was made. The patient required repeated packed RBC transfusions on admission. When the diagnosis of AIHA was confirmed, the patient received corticosteroid therapy (1.5 mg/kg/day) for six weeks. No additional blood transfusion was needed because a remarkable improvement in the Hb level was obtained (9.8 g/dl). After three months, the patient started HCV treatment with oral antiviral therapy. During the follow-up, the patient’s hematologic disorders were completely renormalized.

HCV-related immune disorders are frequent secondary to the activation of the immune system.

HCV lymphotropism determines dysregulation of the immune system, facilitating clonal B-lymphocyte expansion and autoantibody production. ¹⁰

This state represents a trigger in the pathogenesis of virus-related immune disorders both in systemic autoimmune diseases (Sjögren syndrome, rheumatoid arthritis, etc.) and in organ-specific autoimmune diseases (diabetes mellitus, thyroid disorders, etc.). ¹¹

A wide variety of hematologic disorders associated with HCV, such as anemia, neutropenia, and thrombocytopenia, are well documented and are usually associated with interferon (IFN) and ribavarin therapy. ^{2 , 12 – 14} It has been suggested that AIHA observed in untreated HCV infection occurs especially in cirrhotic patients, who have a higher prevalence of autoimmune diseases and respond well to corticosteroid therapy. ¹² However, HCV-related AIHA is uncommon in treatment-naive patients. ⁴

Most cases previously reported in the literature describe warm AIHA with no history of autoimmune disorders. ^{14 – 19} Anemia was not associated with any other hematological disorders with a good response to steroid therapy.

However, the association between AIHA due to cold agglutinin and untreated hepatitis C seems rare. To the best of our knowledge, only two cases have been reported. ^{8 , 9} In the latter, cold agglutinin-mediated hemolytic anemia occurred in 72- and 74-year-old men who were positive for type III mixed cryoglobulinemia, as in our patient. Mixed cryoglobulinemia is the most documented extrahepatic manifestation of HCV infection. ²⁰

A common hypothesis for HCV-related cryoglobulinemia is chronic antigen stimulation of the humoral immune system, with clonal b-lymphocyte expansion producing circulating immunocomplexes. It may also facilitate the development of autoimmune diseases and lymphoproliferative disorders. However, in our patient, as well as in the two published cases, there was no evidence of lymphoproliferative disorder. ⁹

Our patient was diagnosed with cold agglutinins AIHA, including hemolytic anemia, reticulocytosis, elevated lactate dehydrogenase, hyperbilirubinemia, positive direct antiglobulin test (DAT), monospecific DAT positive for C3d, and negative for IgG and cold agglutinin titers ≥ 64. In our patient, the thermal amplitude, which is the highest temperature at which the antibody reacts with the antigen, at 4°C was useful for diagnosis, considering the cold agglutinin titer < 64. ²¹

Cold agglutinins can adhere to red blood cells at low temperatures (complement activation). C3b-coated erythrocytes are removed by macrophages. ²²

Subsequently, it is imperative to rule out potential etiologies of cold agglutinin-induced AIHA, including medication side effects, systemic lupus erythematosus, and malignancy. In our patient and in two similar published cases, no etiology was found, except for HCV infection.

Pathogenesis of AIHA induced by infectious agents is still not well understood. Inflammatory state may explain this rare condition. ^{23 , 24}

Different treatment modalities are available for AIHA: corticosteroids, anti-CD20 (rituximab), Intravenous immunoglobulin (IVIG), and splenectomy. However, in cold AIHA, steroids are less effect in patients with cold AIHA compared with patients with warm AIHA. ^{14 , 19 , 25} Our patient was a good responder to corticosteroids.

Steroids do not seem to show good results in all cases. ⁸ In this situation, anti-CD20 or IVIG may be considered. ²⁶ However, it is important to note that the concomitant use of HCV antiretroviral drugs could explain the good response to corticosteroid therapy observed in our patient.

AIHA due to cold agglutinin is a very uncommon hematologic manifestation of HCV infection, particularly in treatment-naive patients. HCV seems to be mysterious, and further research is needed to elucidate the mechanisms of some related HCV autoimmune manifestations.

Patient gave an informed written consent to publish details regarding the case summary and its use in medical publications.