Why do the INR and PTT measure different pathways?

test tubeCoagulation questions seem to come up all the time! Here’s a good one from one of our readers.

Q. In both the PT and PTT we add thromboplastin, right? So how come the PT measures the extrinsic pathway and the PTT measures the intrinsic pathway”

A. This is a great question because it really gets at the underlying concepts of the PT (INR) and PTT. When I was a medical student, I never really thought about why the INR only measured the extrinsic pathway and the PTT measured only the intrinsic pathway. I just memorized the substance added to the test tube in each test, and the pathway the test measured. Later on, though, I realized I didn’t have a clue as to why the tests measured the pathways they did.

Before we get into the reasoning behind the tests, a quick correction is in order. We don’t add thromboplastin in both the INR and PTT. In the INR, you add thromboplastin, and in the PTT you add phospholipids (not thromboplastin). It turns out thromboplastin is a substance that contains both phospholipids AND a tissue-factor-like substance. That’s why they call the assay the “partial thromboplastin time” – because you only need to add part of the thromboplastin reagent (the phospholipid part) to get this test to run.

To understand why the PT measures just the extrinsic pathway and the PTT measures just the intrinsic pathway, you need to know what activates these pathways in the body. The extrinsic pathway is activated by tissue factor. The intrinsic pathway can be activated by a bunch of things, the most important of which is thrombin.

Why the INR measures the extrinsic pathway
To get blood in a test tube to form fibrin along the extrinsic pathway, you need to add some tissue-factor-like substance. Also, since you removed the platelets and calcium before running the test, you need to add those things back into the test tube (the coagulation system needs a phospholipid surface, normally provided by platelets, and calcium to run). Thromboplastin is a substance that contains both phospholipids and a tissue-factor-like substance. Add thromboplastin and some calcium, and the blood in the test tube will form fibrin via the extrinsic pathway.

Why the PTT measures intrinsic pathway
To get blood in a test tube to form fibrin along the intrinsic pathway, you don’t need to add any tissue-factor-like substance (if you do, the extrinsic pathway will be activated!). All you need to do is add back what you took out of the blood (phospholipids and calcium), as well as something like silica or kaolin to activate the intrinsic pathway (normally, thrombin does this job in vivo), and you’ll form fibrin along the intrinsic pathway. This is actually why the intrinsic pathway was named the way it was: everything you need to get the pathway to run is “intrinsic” to the blood. The extrinsic pathway requires something “extrinsic” to the blood (tissue factor) for it to run.

Bottom line
The INR activates the extrinsic pathway because in this test you add thromboplastin (which contains both a tissue-factor-like substance and phospholipids) to the test tube. The PTT activates the intrinsic pathway because in this test you add just phospholipids to the test tube – and without tissue factor around, fibrin is formed along the intrinsic pathway.

Hematopathology Quiz Part 2

pencils

Here is the second of three installments of a really nice quiz one of our readers sent (the first installment is here, and the third one is coming up). This is a quiz he recently gave to his residents (from medicine, pediatrics and pathology) rotating through hematopathology. Give it a try and see how many you get right! Make sure you check out the answers and nice explanations (if I do say so myself) at the end.

 

1. Smudge cells may be seen on the peripheral blood smear in which of the following?

A. Chronic lymphocytic leukemia
B. Chronic myeloid leukemia
C. Acute myeloid leukemia
D. Thalassemia

 

2. The erythroblasts in acute erythroleukemia (AML- M6) are usually positive for which of the following?

A. PAS
B. CD71
C. CD117
D. All of the above

 

3. The rare Bombay phenotype is seen in people who have blood type:

A. O
B. A
C. B
D. AB

 

4. Cryoprecipitate contains which of the following factors?

A. Factor VIII
B. Factor IX
C. Factor II
D. Factor VII

 

5. The major hematopoietic site in the human embryo is:

A. Spleen
B. Liver
C. Kidney
D. Yolk sac

 

6. POEMS syndrome includes all of the following EXCEPT:

A. Polyneuropathy
B. Ophthalmopathy
C. M-protein
D. Skin abnormalities

 

7. The most common class of immunoglobulin produced by the malignant cells in multiple myeloma is:

A. IgA
B. IgD
C. IgG
D. IgM

 

8. Which of the following is true with respect to hemophilia A?

A. Inheritance is autosomal dominant
B. It is more common in females
C. It never manifests before puberty
D. Patients are deficient in factor VIII

 

9. The cells of which of the following have a “fried-egg” appearance on bone marrow sections?

A. Hairy cell leukemia
B. Follicular lymphoma
C. Acute promyelocytic leukemia
D. Burkitt’s lymphoma

 

10. Which of the following immunophenotypic markers is a pan-B-cell marker?

A. CD3
B. CD19
C. CD33
D. CD56

 

Scroll down for the answers…

 

 

1. A. Smudge cells (also called ghost cells or basket cells) are actually damaged lymphocytes rather than a unique type of cell. Lymphocytes are more fragile than myeloid cells (like neutrophils) – so when you make a blood smear in a patient with a lymphoid disorder (like chronic lymphocytic leukemia), there are likely to be a bunch of smudge cells.

2. D. CD71 and CD117 are both erythroid markers. Erythroblasts in AML-M6 will often stain positively with the PAS (periodic acid schiff) stain (which stains glycogen and mucopolysaccharides). Normal erythroblasts are generally not PAS positive though.

3. A. People with the Bombay phenotype don’t make H antigen. What’s H antigen, you say? It is the antigen that you make first, before you create A or B antigens. To make A and B antigens, you start with a protein precursor (sticking out of the red cell membrane). You add fucose to make what’s called the H antigen. Then, if you have the A gene, you add N-acetylgalactosamine to the H antigen, which makes it an A antigen. If you have the B gene, you add galactose to the H antigen, which makes it a B antigen. If you don’t have either A or B (in other words, if you are type O), then you just leave the H antigen as is, and you don’t make either A or B antigens.

So…people with the Bombay phenotype do not make the H antigen (they stop at the protein precursor stage). They therefore don’t make A or B antigens, either (so they are type O). If a person with the Bombay phenotype needs a transfusion, you need to find Bombay blood – because if you give any other type (regular A, B, AB or O), that blood will have some H antigen in it, and the patient will bust open all those donor red cells (not a good thing to have happen!).

4. A. Cryoprecipitate is a plasma product that contains fibrinogen, von Willebrand factor, and factors VIII and XIII.

5. D. Ok, kind of a gimme.

6. B. POEMS is a rare syndrome that includes a plasma cell dyscrasia (usually myeloma) and a bunch of other stuff. The acronym covers some of the main features: polyneuropathy, organomegaly, endocrinopathy or edema, M-protein, and skin abnormalities (including hyperpigmentation and hypertrichosis). There are other potential complications, though – and not every patient with POEMS has all five features.

7. C. The most common class is IgG. The rarest is IgM. Some people argue that an IgM myeloma doesn’t exist; others say that it exists but is super super rare. Either way, if you get a plasma cell thing that expresses IgM, the first thing you should think of is Waldenstrom’s – myeloma would be way, way down on the list.

8. D. In hemophilia A, patients are deficient in factor VIII (in hemophilia B, it’s factor IX that is deficient). Inheritance is X-linked recessive (though up to 30% of cases arise spontaneously!). As any X-linked recessive disorder, it is more common in males. It often manifests in childhood.

9. A. The hairy cells in hairy cell leukemia have been described as having a “fried egg” appearance because they have these long cytoplasmic processes (“hairs”) that push the cells apart from each other. So the little cell nuclei look like the yolks of eggs, with egg whites (cytoplasm) around them.

10. B. CD19 is a marker present on most B cells (so it’s a “pan” B-cell marker). CD3 is a T-cell marker; CD33 is a myeloid cell marker, and CD56 is a natural killer cell marker.

 

Why you need to look at absolute numbers (not just percentages) of white cells

percentage

Q. I need help with this question from Robbins:

15 year old boy with fever of 10 days. Petechial hemorrhages on trunk and extremities. CBC shows Hg 13.2, Hct 38.9%, MCV, 93, plt ct 175,000 and WBC ct 1860 with 1% segs, 98% lymphs, and 1% monocytes. BM biopsy shows no abnormal cells. What’s the diagnosis?

The answer was overwhelming bacterial infection! It said you are supposed to multiply the percentages in the differential by the total WBC to get absolute values. Any tips for reading WBC counts and diffs? They are so daunting! Would it possibly be included in The Complete But Not Obsessive Hematopathology Guide? I’ve definitely gotten some questions right just from reading pathologystudent though! Such as the difference between a CML and leukomoid reaction! 🙂

A. This is a great question! Yes – there is quite a bit about the diff in The Complete Hematopathology Guide. You might just start by downloading the Top 10 Anemias if you haven’t already (it’s free) – I have a bit in there about normal CBC values. I think once you understand the diff and the CBC, it becomes a lot less overwhelming. You do need to know how to multiply the percentages to get the absolutes…but you definitely don’t have to do that on every case. And for most cases, it’s fine to just do a ballpark figure.

Why do you need to look at absolute counts?
The reason you need to look at absolute counts is because if you just look at percentages, you could mess up. Let’s say you have 60% neutrophils in a particular patient. You might take a quick look at that and go, oh, yeah, that looks fine. But if you don’t take the time to find out the absolute neutrophil count, you could be missing something. If the WBC count in that particular patient is very low, say 1.5 (normal being 4-11), then the patient would have a low number of neutrophils even though the percentage of neutrophils is normal.

Conversely, if the same patient had a very high WBC (say 120), then 60% of the total WBC would be a very high number! Higher than normal for sure. So if you just looked at the percentage of neutrophils in that case, you’d go okay, that looks normal – when in fact the number of neutrophils would be very elevated.So that’s the theory behind looking at the absolute numbers. In reality, most of the time you can just take a look at the WBC, and if it’s really high or low, then go ahead and figure out the absolute numbers of the individual white cells (by multiplying the percentage of that particular cell times the actual WBC). If the WBC is normal, or pretty close to normal, you probably don’t need to go to that amount of work.

Neutropenia and infection
The Robbins question is trying to get you to a) recognize that the patient is neutropenic (in addition to being leukopenic overall), and b) figure out that the reason the neutrophil count is so low is because the patient has a massive infection, and the neutrophils are leaving the bloodstream to go the tissues where they are needed (hence the number of neutrophils in the blood is actually low). Usually in an infection (a bacterial one anyway), the WBC is high, and the percentage (or at least the absolute number) of neutrophils is way increased. This leaving-the-bloodstream phenomenon that happened in this patient is uncommon – but it does occur.

By the way, if you just looked at the % of lymphocytes in this question, and didn’t think too much about the WBC, you might (incorrectly) conclude that the patient has a lymphocytosis (98% lymphocytes! That sounds like a lot.) In fact, the patient is actually leaning towards being lymphocytopenic (since the total number of white cells is really low). The normal absolute lymphocyte count is somewhere between 1 and 4; in this patient, it’s about 1800 (98% of 1860).

Hope that helps! I hope you can get to the point where it doesn’t feel totally overwhelming – because it’s actually pretty straightforward and doable, once you understand some general principles.