Labs

Let’s begin with the patient’s complete blood count (CBC):

WBCHbHctMCVMCHCRDW-SDPLT
19.08.123.482 34.648.111

What’s what: WBC, white blood cell count; Hb, hemoglobin; MCV, mean cell volume; MCHC, mean cellular hemoglobin concentration; RDW-SD, red cell distribution width-standard deviation; platelets, PLT; Normal values: WBC 5-10 x 109/L, RBC 4-6 x 1012/L, Hb 12-16 g/dL, Hct 35-47%, MCV 80-100 fL, MCHC 32-36 g/dL, RDW-SD < 45%, platelets (PLT) 150-450 x 109/L

The patient has leukocytosis, anemia and thrombocytopenia.

A white cell differential was unremarkable, revealing an isolated neutrophilia (not shown) without evidence of a left shift. These findings are relatively non-specific, possibly secondary to stress or bleeding.

In terms of making a diagnosis, our best bet is to focus on the anemia and thrombocytopenia. Let’s begin with a differential diagnosis for anemia (though we could just as easily begin with causes of thrombocytopenia) and then see how the concomitant thrombocytopenia helps to narrow the differential.

Assuming that the cytopenias are causally related in this patient, the goal is to identify those conditions that are associated with both anemia and thrombocytopenia.

We will begin at the top of the algorithmic tree by dividing anemia into those causes associated with an appropriate reticulocyte count (we will come back to what that means in a bit) and those associated with an inappropriately low reticulocyte response. Let’s consider the side of the ledger with an appropriate reticulocyte response.

Active bleeding as a cause of anemia is usually clinically apparent from the history and/or a change in the vital signs. Hemolytic anemia can be further divided into causes that are positive for direct antiglobulin test (DAT, otherwise known as the Coombs test) and those that are DAT negative. Let’s consider DAT-positive (DAT+) vs. DAT-negative (DAT-) “buckets”.

Let’s stick with the hemolysis branch for now.

DAT, direct antiglobulin test

In the following sorting exercise, drag the card on the top with a hemolytic disease/condition into the appropriate diagnostic bucket on the bottom.

Valve hemolysis
Delayed transfusion reaction
Chronic kidney disease
B12 deficiency
Cold agglutinin disease
ABO incompatibility
TMA
Hemolytic disease of the newborn
Hereditary spherocytosis
G6PD deficiency
Sickle cell disease
Babesiosis
Clostridial sepsis
Thermal injury
Spur cell anemia
Warm autoimmune hemolytic anemia
Hypothyroidism
Alcohol toxicity
Immune HA
Non-immune HA
Non-hemolytic

You’re doing great! In this schematic, we have filled in the various causes of immune and non-immune hemolytic anemia. Immune-mediated (DAT-positive) hemolytic anemia may be divided into allo- and autoimmune causes. Non-immune causes are further separated into extracorpuscular and intracorpuscular etiologies. In extracorpuscular non-immune hemolytic anemia, the red bloods cell are intrinsically normal, but suffer collateral damage from their interactions with the environment, for example by passing over fibrin strands in microvessels (as occurs in thrombotic microangiopathy), by undergoing turbulent flow through paravalvular leaks (causing valve hemolysis), or by being exposed to toxins related to chronic liver disease or to certain pathogens. Intracorpuscular causes include those conditions in which the primary defect is in the red cell, either in its hemoglobin molecule, in its membrane structure or in its intracellular enzymes. Intracorpuscular non-immune hemolytic anemia is almost always hereditary in nature. One exception is paroxysmal nocturnal hemoglobinuria, which is an acquired red cell membrane disorder.

DAT, direct antiglobulin test; HA, hemolytic anemia; TMA, thrombotic microangiopathy; MAHA, macroangiopathic hemolytic anemia; SCD, sickle cell disease; HS, hereditary spherocytosis; HE, heredity elliptocytosis; PHN, paroxysmal nocturnal hemoglobinuria; Def, deficiency; PKD, pyruvate kinase deficiency.

Let’s return to the first branch point in the flow chart for anemia and consider the causes of anemia associated with an inappropriate reticulocyte response (the so-called hypoproliferative anemias; upper right in above schematic).

The first branch point in considering hypoproliferative anemias is the mean cell volume (MCV). This represents the time-honored morphological classification of anemia and is very helpful when sorting through the differential diagnosis of anemia.

DAT, direct antiglobulin test; HA, hemolytic anemia; TMA, thrombotic microangiopathy; MAHA, macroangiopathic hemolytic anemia; SCD, sickle cell disease; HS, hereditary spherocytosis; HE, heredity elliptocytosis; PHN, paroxysmal nocturnal hemoglobinuria; Def, deficiency; PKD, pyruvate kinase deficiency.

Let’s do another sorting exercise. In this case, drag the disease or condition into the appropriate red cell size bucket.

Thalassemia
Excess schistocytes
Chronic kidney disease
Primary bone marrow process
Medications, e.g., hydroxyurea
Hypothyroidism
Inflammation
Chronic liver disease
Iron deficiency
Alcohol
Lead poisoning
B12 deficiency
Hypogonadism
Folate deficiency
Hypoadrenalism
Microcytic
Normocytic
Macrocytic

Here is the classification of anemia, based on kinetics (reticulocyte count) and morphology (mean cell volume). A second component of the morphological classification, which we have chosen not to include in order to keep things simple, is the mean corpuscular hemoglobin concentration (MCHC), which may be used to separate microcytic anemia into hypochromic (low MCHC) causes, as seen in iron deficiency anemia, and normochromic (normal MCHC) causes, which is more typical of thalassemia.

DAT, direct antiglobulin test; HA, hemolytic anemia; TMA, thrombotic microangiopathy; MAHA, macroangiopathic hemolytic anemia; SCD, sickle cell disease; HS, hereditary spherocytosis; HE, heredity elliptocytosis; PHN, paroxysmal nocturnal hemoglobinuria; Def, deficiency; PKD, pyruvate kinase deficiency. primary BM, primary bone marrow condition.

Time for another sorting exercise! This time, we are going to identify conditions that are associated with concomitant anemia and thrombocytopenia. Drag and drop the cause of anemia on the top cards into the appropriate bucket (associated with concomitant thrombocytopenia or not).

Sickle cell disease
PK deficiency
Hereditary spherocytosis
Hypothyroidism
Chronic liver disease
Immune-mediated hemolytic anemia
Chronic kidney disease
Iron deficiency
Thalassemia
Primary bone marrow process
Thrombotic thrombocytopenia (TMA)
Alcohol toxicity
Severe B12 deficiency
G6PD deficiency
Severe infection, e.g., Clostridial sepsis
Valve hemolysis
Thrombocytopenia
No thrombocytopenia

The following is the differential diagnosis for anemia, with those conditions associated with concomitant thrombocytopenia indicated in dark blue.

DAT, direct antiglobulin test; HA, hemolytic anemia; TMA, thrombotic microangiopathy; MAHA, macroangiopathic hemolytic anemia; SCD, sickle cell disease; HS, hereditary spherocytosis; HE, heredity elliptocytosis; PHN, paroxysmal nocturnal hemoglobinuria; Def, deficiency; PKD, pyruvate kinase deficiency. primary BM, primary bone marrow condition.

Below, we have reorganized the diagnostic algorithm for anemia in table format according to those conditions associated with concomitant thrombocytopenia (TP):

CategoryCause of anemiaCause of TPComments
BleedingBlood lossAny cause of TPTP increases risk of bleeding.
Hemolysis
AIHAImmune-mediated red cell destructionITP – immune mediated platelet destructionCombined AIHA and ITP is called Evans syndrome.
TMAMicroangiopathic hemolysisConsumptionTP is part of the disease spectrum.
Clostridial sepsisIntravascular hemolysisConsumptionTP is common in all types of sepsis.
BabesiosisIntravascular hemolysisUnclear mechanismTP is a common finding in babesiosis.
CirrhosisSpur cell anemia, hypersplenismHypersplenismTP is the most common cytopenia in cirrhosis.
PNH (membranopathy)Intravascular hemolysis, marrow suppressionMarrow suppressionTP is common in PNH.
Normocytic anemia
Primary bone marrow processMarrow suppressionMarrow suppressionMyelodysplasia, aplastic anemia, plasma cell dyscrasia, and leukemia may all cause TP.
Macrocytic anemia
AlcoholMarrow suppressionMarrow suppressionTP is more common than anemia with alcohol toxicity.
Vitamin B12 deficiencyIneffective erythropoiesisMarrow suppressionTP and leukopenia may both occur.
TP, thrombocytopenia; AIHA, autoimmune hemolytic anemia; ITP, immune thrombocytopenia; TMA, thrombotic microangiopathy; PNH, paroxysmal hemoglobinuria.

Recall from the history that the patient’s complete blood count was normal just three months ago. That piece of information helps us narrow the differential diagnosis. For example, it is unlikely that vitamin B12 deficiency and many of the more chronic causes of primary bone marrow failure would present in such a precipitous manner. In fact, the timing of the anemia and thrombocytopenia helps us narrow the likely diagnosis to:

  • Acute primary bone marrow process (for example, acute leukemia)
  • Autoimmune hemolytic anemia and ITP (Evans syndrome)
  • Thrombotic microangiopathy:
    • Thrombotic thrombocytopenic purpura
    • Hemolytic uremic syndrome
    • Disseminated intravascular coagulation
  • Infection
    • Babesiosis
    • Clostridial sepsis (though the presentation is usually more dramatic)

What conditions are associated with an elevated reticulocyte count?

a
Anemia of inflammation
b
Hemolytic anemia
Hemolysis typically involves the destruction of mature red cells, leaving the bone marrow unscathed and poised to compensate by increasing the production and release of reticulocytes into the blood.
c
Immune thrombocytopenia
d
Bleed
Bleeding, like hemolysis, does not impair erythropoiesis, allowing the bone marrow to respond by producing and releasing more reticulocytes. Acute bleeds result in reticulocytosis.

Let’s consider the differential diagnosis of recent/new onset thrombocytopenia and anemia. What would you expect the reticulocyte count to be in each case – appropriately increased or inappropriately low (answer on next slide)?

DiseaseReticulocyte count
ITP with major bleed or Evans syndrome
Thrombotic thrombocytopenia purpura
Primary bone marrow conditions
Hemolytic uremic syndrome
Disseminated intravascular coagulation
Babesiosis
ITP, immune thrombocytopenia

Let’s return to our differential diagnosis of recent/new onset thrombocytopenia and anemia. What would you expect the reticulocyte count to be in each case – appropriately increased or inappropriately low (answer on next slide)?

DiseaseReticulocyte count
ITP with major bleed or Evans syndrome Appropriate
Thrombotic thrombocytopenia purpuraAppropriate
Primary bone marrow conditions Inappropriate
Hemolytic uremic syndromeAppropriate
Disseminated intravascular coagulationAppropriate
Babesiosis Appropriate
ITP, immune thrombocytopenia

Keep in mind that an appropriate reticulocyte response assumes that the bone marrow is able to make more red cells. If there is a concomitant condition, for example iron deficiency or inflammation, that suppresses erythropoiesis, the patient may not be able to mount an appropriate reticulocyte count even in the face of severe hemolysis or bleeding.

Let’s look at the reticulocyte count in our patient:

ParameterVAlue
Percentage reticulocytes4.5%
Absolute reticulocyte count0.13 x 1012/L

What is the patient’s red cell count in trillions (1012)/L?

a
1.2 trillion
b
2.9 trillion

Correct! 4.5% x [2.89 x 1012/L] = 0.13 x 1012/L

c
6.7 trillion

Returning to our differential diagnosis of newly diagnosed thrombocytopenia and anemia in an acutely ill patient, we can eliminate primary bone marrow disorders on account of the patient’s reticulocyte response.

DiseaseReticulocyte count
ITP with major bleed or Evans syndrome Appropriate
Thrombotic thrombocytopenia purpuraAppropriate
Primary bone marrow conditions Inappropriate
Hemolytic uremic syndromeAppropriate
Disseminated intravascular coagulationAppropriate
Babesiosis Appropriate
ITP, immune thrombocytopenia

Let’s look at the various markers of hemolysis when the patient first presented to the emergency room:

ParameterValueNormal Range
LDH2112 IU/L 94-250 IU/L
AST/ALT71/30 IU/L 0-40 IU/L for both
Haptoglobin< 10 mg/dL 30-200 mg/dL
Bilirubin (total)3.6 mg/dL 0-1.5 mg/dL

In summary, the above results are classic for hemolysis!

Interim summary

This is a 56 year-old man with recent/new onset anemia and thrombocytopenia associated with increased reticulocytes and positive hemolytic markers. Based on these data, we are thinking about Evans syndrome, thrombotic thrombocytopenia purpura (TTP), Babesiosis or other thrombotic microangiopathies such as hemolytic uremic syndrome and disseminated intravascular coagulation. The elephant in the room, which we are ignoring for the moment, is the change in mental status. This drives TTP to the top of the differential diagnosis. But let’s play along and put the neurological findings aside for a moment.

Now it’s time to consider the peripheral smear. In addition to low platelet numbers, what findings would you expect to see in the 4 conditions under consideration (answer on next slide)?

ConditionPeripheral smear findings
Evans syndrome
TTP
HUS
DIC
Babesiosis
TTP, thrombotic thrombocytopenia purpura; HUS, hemolytic uremic syndrome; DIC, disseminated intravascular coagulation.

Now it’s time to consider the peripheral smear. In addition to low platelet numbers, what findings would you expect to see in the 4 conditions under consideration (answer on next slide)?

ConditionPeripheral smear findings
Evans syndrome Polychromatophilia, spherocytosis
TTP Polychromatophilia, schistocytes
HUSPolychromatophilia, schistocytes
DICPolychromatophilia, schistocytes
BabesiosisPolychromatophilia, intraerythrocytic inclusions
TTP, thrombotic thrombocytopenia purpura; HUS, hemolytic uremic syndrome; DIC, disseminated intravascular coagulation.

The peripheral smear from the patient is shown here:

The smear shows reduced platelets and presence of schistocytes (horn cell, blue arrow; helmet cell, asterisk).

For more information about schistocytes, click here.

The patient’s direct antiglobulin test (DAT) was negative. The combination of appropriately elevated reticulocytes, positive hemolytic makers and negative DAT places him in the extracorpuscular hemolytic anemia “bucket”. The presence of schistocytes on the peripheral smear further narrows the diagnosis to thrombotic microangiopathy (TMA), which may be caused by thrombotic thrombocytopenia purpura, hemolytic uremic syndrome, disseminated intravascular coagulation or a handful of other secondary causes (see next slide),

Let’s consider the thrombotic microangiopathies (TMAs) for a moment.

Definition: TMAs are a group of rare disorders characterized by thrombocytopenia, microangiopathic hemolytic anemia (MAHA), and microvascular thrombus formation ((fibrin/platelet rich), leading to tissue injury. MAHA, in turn, is characterized by red blood cell destruction within the microvasculature accompanied by thrombocytopenia due to platelet activation and consumption.

Classification: TMAs may be classified as primary or secondary. Primary TMAs occur spontaneously with no associated underlying cause, and include TTP and HUS. Secondary TMAs are associated with an underlying condition.

TTP, thrombotic thrombocytopenic purpura; STEC-HUS, Shiga toxin-related hemolytic uremic syndrome; aHUS, atypical hemolytic uremic syndrome; HTN, hypertension; CTD, connective tissue disease; DIC, disseminated intravascular coagulation.

What tests would be most helpful in distinguishing between thrombotic thrombocytopenia purpura (TTP) and hemolytic uremic syndrome (HUS)?

a
Fibrinogen
Fibrinogen is typically normal in both conditions.
b
Prothrombin time
Prothrombin time is typically normal in both conditions.
c
Creatinine
Patients with HUS, but not TPP, typically present with renal dysfunction.
d
Blood culture

The patient’s BUN and creatinine were normal. Thus, a diagnosis of hemolytic uremic syndrome is unlikely.

What tests would be most helpful in distinguishing between thrombotic thrombocytopenia purpura (TTP) disseminated intravascular coagulation (DIC)?

a
Fibrinogen
Reduced in many cases of DIC, typically normal in TTP.
b
Prothrombin time
Increased in many cases of DIC, typically normal in TTP.
c
Ferritin
Not normally helpful in distinguishing between DIC and TTP.
d
C-reactive protein
Not normally helpful in distinguishing between DIC and TTP.

The fibrinogen and prothrombin times were normal in this case. The ISTH has developed a scoring system for the diagnosis of disseminated intravascular coagulation (DIC). For starters, it is necessary to identify an underlying condition that causes DIC, and that is not apparent in this patient, making a diagnosis of DIC unlikely to begin with. Next, the following lab parameters are considered:

ParameterScoreComment
Fibrinogen (g/L)>1 (0)
<1 (1)
Overall sensitivity of low fibrinogen level reported to be about 30%.
Prothrombin time (PT) (seconds)<3 (0)
3-6 (+1)
>6 (+2)
Elevated in 50%-60% of patients with DIC at some point during course of illness.
Platelet count (x 109/L)>100 (0)
50-100 (+1)
<50 (+2)
Thrombocytopenia reported to occur in up to 98% of DIC cases.
D-dimers or FDPsNo increase (0)
Moderate increase (+2)
Severe increase (+3)
Elevated FDPs and D-dimers are sensitive, but not specific for DIC.

Data are presented as values with number of points in parentheses.

A total score of ≥5 is compatible with diagnosis of DIC

This patient did not have D-dimers or FDPs measured. So, he could technically have made the cut for DIC if he had a severe increase in D-dimers or FDPs (for a total of 5 points). However, the absence of an identifiable underlying condition makes this all but a moot point.

By a process of elimination – even in the absence of a history or physical exam – we arrive at thrombotic thrombocytopenia purpura (TTP) as the most likely diagnosis (red color indicates the results found in the patient):

DiseaseReticulocyte countSchistocytesRenal functionFibrinogen/PT
ITP with major bleed or Evans syndrome AppropriateNoNormal Normal
TTP Appropriate YesNormal Normal
Primary bone marrow conditions Inappropriate NoNormal Normal
HUSAppropriate YesAbnormal Normal
DIC Appropriate YesNormalLow
BabesiosisAppropriateNoNormalNormal
TTP, thrombotic thrombocytopenia purpura; HUS, hemolytic uremic syndrome; DIC, disseminated intravascular coagulation

… add in the neurological changes, and the pretest probability of TTP further increases.

Is there a single diagnostic test for thrombotic thrombocytopenic purpura (TTP)?

a
Yes
ADAMTS13 activity level (level < 10% is diagnostic of TTP).
b
No

What is ADAMTS13?

a
A clotting factor that stabilizes fibrin clot
b
An enzyme that cleaves von Willebrand factor
Correct. ADAMTS13 is a metalloprotease that regulates the platelet-aggregating activity of von Willebrand factor (vWF) by cleaving it at specific sites. In thrombotic thrombocytopenia purpura (TTP), ADAMTS13 activity is deficient due to presence of autoantibodies (acquired TTP) or inherited mutations in ADAMTS13 gene.
c
An antibody that targets platelets

The ADMATS13 is a send-out test. The initial diagnosis of thrombotic thrombocytopenia purpura (TTP) must be made without the benefit the ADAMTS13 level, which typically takes several days to come back. One can either use their clinical experience to determine who is at high risk for the condition or use a clinical prediction score for TTP. One of these is called the PLASMIC score. Let’s see how it works on the next slide.

PLASMIC score

Plasmic score from MdCal

Notes:

  • The higher the score, the greater the likelihood of TTP.
  • Many of the parameters included in the PLASMIC score are negative predictors of thrombotic thrombocytopenic purpura (TTP). For example, the score takes into consideration non-TTP thrombotic microangiopathy caused by transplantation and cancer (based on history), disseminated intravascular coagulation (based on an elevated INR) and hemolytic uremic syndrome (based on elevated creatinine).
  • Schistocytes are not included in the score because they are considered a precondition for applying the score.
  • The PLASMIC score was developed using a cohort of 214 patients with suspected thrombotic microangiopathy and has been validated in 2 additional cohorts of patients.

Our patient’s platelet count of 11 x 109/L qualifies for one point. He also gets one point for the unmistakable finding of hemolysis, and another point each for not having a history of cancer or solid organ or hematopoietic stem cell transplantation. That’s four points so far. Add another point for the normal PT/INR and we are up to a total of five points. The patient’s mean cell volume (MCV) was < 90 fL for another point. Finally, the creatinine was normal, bringing the tally to seven points for a pretest probability of about 70%.

What is the explanation for a low mean cell volume (one of the PLASMIC criteria) in thrombotic thrombocytopenic purpura (TTP)?

a
Patients lose hemoglobin in their urine and develop iron deficiency
TTP presents acutely, while the development of iron deficiency from hemoglobinuria is a chronic process.
b
Patients with thalassemia are more likely to develop TTP
This statement is not true.
c
The presence of schistocytes causes a reduction in mean cell volume (MCV)
Correct. Schistocytes are by definition smaller than red cells, so if there are enough of them, they may reduce the MCV.

Some final thoughts about diagnosing thrombotic microangiopathy (TMA), including thrombotic thrombocytopenia purpura (TTP):

Any time a patient presents to urgent care with new onset of anemia and thrombocytopenia, we must consider thrombotic microangiopathy, especially TTP and atypical hemolytic uremia syndrome, because these conditions, if left undiagnosed and untreated, have high mortality. As hematologists, we are trained to use pattern recognition to identify patients with this condition, and the patient in this case would be no exception: new anemia, thrombocytopenia and mental status changes = TMA until proven otherwise. A quick look at the reticulocyte count, the DAT, the hemolysis indices and the peripheral smear, and the diagnosis is clinched within an hour of seeing the patient. That being said, it is always important to cross-check real time data with conceptually sound diagnostic frameworks – like the flow charts and algorithms in this case study – and to constantly update your assessment accordingly. The purpose of this exercises in this case study is not to slow you down at the bedside, but rather to help you build enduring conceptual frameworks with which to approach diagnosis.

Before moving to treatment, let’s consider the full panel of tests we might order in a patient with suspected thrombotic thrombocytopenic purpura (TTP). The following are recommendations by the British Committee for Standards in Haematology (see clinical practice guideline) in such patients:

TestComments
Complete blood countAnaemia, thrombocytopenia
Peripheral smearSchistocytes
Reticulocyte countIncreased
HaptoglobinReduced
Clotting screen including fibrinogenNormal
Urea and electrolytes+/- Renal impairment
Troponin T/Troponin IFor cardiac involvement
Liver function testsUsually normal
CalciumMay reduce with TPE
Lactate dehydrogenase (LDH)Increased
Direct antiglobulin testNegative
Blood group and antibody screenTo allow provision of blood products
Pregnancy testIn women of child-bearing age
ADAMTS 13 assay ((activity/antigen and inhibitor/antibody
in specialized laboratory)
Do not wait for result before starting treatment in suspected
TTP
Electrocardiogram/EchocardiogramTo document/monitor cardiac damage
CT/MRI brainTo determine neurological involvement
TPE, therapeutic plasma exchange

Should the patient be admitted to the hospital?

a
Yes
Thrombotic thrombocytopenic purpura has a mortality of up to 90% if left untreated.
b
No

The patient is admitted directly to the hospital