Correct answer: Adequate for degree of anemia

Discussion

This patient’s RPI is 4.6. An RPI below 2 indicates that the bone marrow is inadequately responding to the patient’s anemia. An RPI of 2 or more indicates that the bone marrow is appropriately responding to either blood loss or to red blood cell destruction (hemolysis).

The RPI is a two-step calculation obtained from the reticulocyte percent and the patient’s hematocrit (or hemoglobin).

Step 1: Absolute reticulocyte count = reticulocyte count multiplied by patient’s hematocrit (or hemoglobin) divided by the normal hematocrit (or hemoglobin)

Step 2: Reticulocyte index = absolute reticulocyte count divided by the maturation factor

Maturation factors are found in the following table:

Question 2 of 10

Correct answer: Hyperproliferative and macrocytic

Discussion

The kinetic description of anemia refers to the patient’s bone marrow response to anemia. In this case, her RPI is above 2, which represents a hyperproliferative response from the bone marrow. The morphologic description of anemia is described using the mean corpuscular volume (MCV). Microcytic anemia refers to MCV below 80 femtoliters (fL). In normocytic anemia, the MCV ranges from 80-100 fL. In macrocytic anemia, the MCV is above 100 fL. Describing anemia in both kinetic and morphologic terms helps narrow down a differential diagnosis for anemia. Microcytic anemia is seen most often in cases of iron deficiency or thalassemia. Macrocytic anemia is seen most commonly in cases of vitamin B12 or folate deficiency, liver disease, drug effect, myelodysplastic syndrome, hypothyroid, alcohol use, and hemolytic anemia. Macrocytosis can be seen in the setting of hemolytic anemia because reticulocytes are released from the intramedullary space early and have a larger diameter compared with fully mature red blood cells. As noted above, a hyperproliferative response from the bone marrow is indicative of bleeding or hemolysis, whereas a hypoproliferatve response from the bone marrow has a much larger differential diagnosis.¹

Question 3 of 10

Correct answer: Direct Coombs test, LDH, and haptoglobin

Discussion

This patient’s anemia is macrocytic and hyperproliferative. She does not have any clinical evidence of bleeding, so hemolysis should be high on your differential diagnosis. The first step to evaluate for hemolysis would be to obtain a direct Coombs test along with LDH and haptoglobin tests. In the setting of hemolytic anemia, the LDH is typically elevated and the haptoglobin (a scavenger protein that binds to free hemoglobin in the plasma) is very low, often undetectable. Another clue for hemolysis in this case is the elevated unconjugated bilirubin and scleral icterus.

The direct Coombs test incubates the patient’s red blood cells with the Coombs reagent, which is a polyspecific anti-human immunoglobulin. If a patient’s red blood cells are coated with immunoglobulin G autoantibody or complement, the Coombs reagent will cause agglutination in the test tube. A second step is done to determine whether the red blood cells are coated with immunoglobulin G or complement. The indirect Coombs is not used to evaluate hemolysis and is often ordered in error. The indirect Coombs is used by blood banks to see if a patient’s plasma contains alloantibodies against a known panel of red blood cell antigens. ²

Case continued

Test results show the patients LDH level is 1500 U/L and haptoglobin is less than 10 mg/dL. The direct Coombs test is positive for immunoglobulin G and negative for complement.

Question 4 of 10

Correct answer: 

Discussion

The peripheral blood smear in option D shows several microspherocytes (black arrows). Microspherocytes are present in warm autoimmune hemolytic anemia due to splenic macrophage engulfing a portion of the red blood cell membrane as they phagocytose the Fc receptor of the immunoglobulin attached to the surface of the red blood cell membrane. If the red blood cell escapes total engulfment, the membrane changes from the normal bi-concave disk to a spherocyte, due to decreased cell surface membrane.³ Option A represents iron deficiency anemia, with microcytic and hypochromic red blood cells, and target cells. Option B depicts cold agglutinin disease, in which the red blood cells are coated with complement (+/- IgG) and agglutinate together in the periphery at cool temperatures of the body, such as distal fingers, nose, and ears. Option C shows a normal peripheral blood smear.

Question 5 of 10

Correct answer: Systemic lupus erythematosus

Discussion

This patient is a 28-year-old female diagnosed with warm autoimmune hemolytic anemia (WAIHA). WAIHA can be primary or secondary. Secondary WAIHA is often seen in the setting of SLE.⁴ The patient is having joint pain and facial rash, which are clues to underlying SLE. Although lymphoproliferative disorders such as CLL/SLL can also present with secondary WAIHA, our patient is in her 20s, making a non-Hodgkin lymphoma such as CLL/SLL less likely. Neither osteogenesis imperfecta nor platelet delta storage pool deficiencies are associated with WAIHA.

Question 6 of 10

Correct answer: Hereditary spherocytosis

Discussion

All of the options are types of non-immune hemolytic anemia. Hereditary spherocytosis is the most common inherited red blood cell cytoskeleton defect, which changes the shape of the red blood cell from bi-concave disk to a spherocyte. The spherocytes are not able to navigate as well through splenic sinusoids and get trapped and removed from the circulation by macrophages. Hereditary spherocytosis is often inherited in an autosomal dominant pattern.⁵ Hereditary spherocytosis looks very similar to WAIHA on peripheral smear.

Paroxysmal nocturnal hemoglobinuria (PNH) is an acquired defect in hematopoietic stem cells, which results in complement mediated hemolysis. PNH does not often present with microspherocytes because red blood cells in PNH are killed by the terminal complement system in the form of intravascular hemolysis, rather than macrophage engulfment of Fc receptors from immunoglobulin-coated antibodies. G6PD deficiency is the most common enzyme deficiency of red blood cells that is inherited in X-linked fashion, thus males are more commonly affected than females. G6PD deficiency can lead to hemolytic episodes in settings of increased oxidative stress (infections/certain medications). Pyruvate kinase deficiency is the second most common red blood cell enzyme defect, but the mechanism of hemolysis is less well understood. Neither G6PD nor pyruvate kinase deficiency present with microspherocytes on peripheral blood smear.

Case continued

Our patient sees a hematologist as well as a rheumatologist, and she is ultimately diagnosed with SLE and secondary WAIHA. She is treated with high-dose prednisone (1 mg/kg) for 3 weeks, and she has an excellent hemoglobin level response. She is also started on mycophenolate mofetil (CellCept) at 500 mg twice daily, which marginally controls her SLE symptoms. Her WAIHA remains in remission with a gradual taper of steroids.

About 6 months later, her anemia returns, and you order blood tests. Results show the following:

WBC: 2.9 × 109/L (reference range, 3.7-11 × 109/L)

Hemoglobin: 11 g/dL (11.5-15.5)

Hematocrit: 34.3% (36-46)

MCV: 84.1 fL (80-100)

Platelet count: 205 × 109/L (150-400)

Absolute neutrophil count: 1.7 k/uL (1.45-7.50)

Absolute lymphocyte count: 0.6 k/uL (1.0-4.00)

Haptoglobin: 297 mg/dL (31-238)

LDH: 190 U/L (135-214)

Ferritin: 375 ng/mL (14.7-205.1)

Serum iron: 35 µg/dL (41-186)

Transferrin iron-binding capacity: 375 µg/dL (232-386)

Transferrin saturation: 9% (15-57)

Reticulocyte %: 1.5% (0.4-2.0)

Question 7 of 10

Correct answer: Anemia of chronic disease

Discussion

Our patient has been diagnosed with an autoimmune disease — SLE — that can be chronic. This disease can lead to anemia of chronic disease, which is largely caused by the upregulation of hepcidin, an acute phase reactant. Increased levels of hepcidin lead to downregulation of the ferroportin receptors on enterocytes, which downregulates the absorption of iron from the gut.⁶ Hepcidin also inhibits the release of iron from macrophages, causing an essential state of functional iron deficiency even though the body has plenty of iron stores (thus not anemia due to iron deficiency). Iron tests can help discern between anemia of chronic disease and anemia of iron deficiency (see Table 1). The reticulocyte production index will be low in both iron-deficient anemia and anemia of chronic disease, whereas the reticulocyte production index is elevated in the setting of acute blood loss. The reticulocyte production index, LDH, and haptoglobin are not suggestive of hemolysis.

Table 1. Laboratory tests to differentiate between iron-deficient anemia and anemia of chronic disease.

Case continued

The rheumatologist adjusts our patient’s SLE medications, and her hemoglobin recovers to the normal range. She goes on to become pregnant, and she breast feeds for 1 year after the baby is born. She then returns to your clinic with recurrent anemia. Laboratory tests show the following values:

WBC 4.5 × 109/L

Hemoglobin: 9.1 g/dL

Hematocrit: 30.4%

Platelet count: 205 × 109/L

MCV: 78 FL

Serum iron: 42 µg/dL

Transferrin iron-binding capacity: 398 µg/dL

Transferrin saturation: 11%

Ferritin: 9.6 ng/mL

Reticulocyte %: 1.1%

Question 8 of 10

Correct answer: Iron-deficient anemia

Discussion

Her lab values now are consistent with iron-deficient anemia. She has low storage of iron seen by decreased ferritin, which is specific for iron deficiency. Iron deficiency is further supported by her low serum iron level, low transferrin saturation, and increased transferrin iron-binding capacity.

Question 9 of 10

Correct answer: Increased iron utilization

Discussion

Iron-deficient anemia can occur due to increased utilization state, such as with pregnancy and lactation.⁷ It is important to look for the cause of iron deficiency. In young females, menstrual blood losses or pregnancy are the most common causes of iron deficiency. Our patient did not have a history of heavy menstrual periods, but she had been pregnant and breast fed the child. In older individuals, it is important to look for gastrointestinal sources of blood losses, as you do not want to miss any underlying malignancy. Inflammatory bowel disease, Helicobacter Pylori infection, and celiac disease are other causes of iron deficiency, but our patient did not have any symptoms suggestive of these disorders.

Case continued

Our patient does well for the next 20 years, and her SLE is well controlled with weekly methotrexate. She moved to another state but did not re-establish with a rheumatologist and her methotrexate supply ran out. About 3 months later, she suddenly felt unwell, with headache and vague abdominal pain and feelings of confusion. She called 911 and was taken to the emergency department. Her initial exam was non-focal and her vital signs were in the normal range. Results from the lab tests and a peripheral blood smear are shown below.

WBC: 6.7 × 109/L

Hemoglobin: 5.6 g/dL

Hematocrit: 16.5%

Platelet count: 8 × 109/L

MCV: 80.5 fL

Reticulocyte count: 14%

Prothrombin time: 11.9 seconds (reference range, 9.7-13.0 seconds)

International normalized ratio: 1.1 (0.9-1.3)

Fibrinogen activity level: 294 mg/dL (200-400)

Haptoglobin: < 10 mg/dL

LDH: 3,009 U/L

Direct Coombs test: Negative

A peripheral blood smear showed the following:

Question 10 of 10

Correct answer: Order ADAMTS13 activity assay, and immediately start high dose steroids, caplacizumab, and initiate plasma exchange

Discussion

Patients with one type of autoimmune disorder can have additional types of autoimmune disease. In this case, our patient has SLE, a history of WAIHA, and now presents with a clinical picture concerning for thrombotic thrombocytopenic purpura (TTP). TTP is most commonly caused by an autoantibody to ADAMTS13, which is an important enzyme that cleaves ultra-large von Willebrand multimers. Without clipping of these large multimers, the large von Willebrand proteins are very hemostatic and are responsible for microthrombosis in small arterial vasculature. These microthrombi consume platelets (leading to thrombocytopenia) and shear red blood cells as they pass through the microthrombi (leading to schistocytes).

TTP is a medical emergency and requires prompt recognition and treatment. Undetectable ADAMTS13 is pathognomonic for this disorder, but treatment should not wait on results in the setting of intermediate or high clinical suspicion of TTP. There is a validated score, the PLASMIC score,⁸ that can be used to predict the presence of TTP. The standard of care for TTP is currently triple therapy, with urgent plasma exchange, high-dose glucocorticoids, and caplacizumab, which is a nano-antibody that interferes with platelets binding to von Willebrand protein.⁹ It is important to recognize concurrent anemia and thrombocytopenia as potentially dangerous disorders that fall under the microangiopathic hemolytic anemia umbrella.

KEY POINTS

• Anemia is a symptom of an underlying disease, and the cause of anemia should be determined.
• Using the kinetic (reticulocyte production index) and morphologic description (using MCV) of anemia can help narrow the differential diagnosis and guide the next steps toward determining an underlying cause.
• Hemolytic anemia can have several causes, and a helpful first step is to determine whether the hemolytic anemia is immune (warm or cold autoimmune hemolytic anemia) or nonimmune hemolytic anemia.
• Iron-deficiency anemia and anemia of chronic disease are very common entities. Results from iron studies can help differentiate between the two disorders; however, overlap between the lab values does occur.
• The presence of acute anemia and thrombocytopenia should prompt an urgent evaluation for a microangiopathic hemolytic anemia disorder, such as thrombotic thrombocytopenic purpura, which requires timely recognition and treatment.