CLINICAL PHATHOLOGY INDIVIDUAL ASSIGNMENT

TITLE:band neutrophils,toxic neutrophils,hypersegmented

neutrophils,hyposegmented neutrophils,giant toxic neutrophils

SUBMMITED BY:TEMELASH TEREFE TESHKOLA

ID……………………………….R3267/09

SUBMITTED TO: Dr.JIRATA

SUBMITTION DATE:17/8 2013 E.C

BISHOFTU,SHEWA,ETHIOPIA
Band neutrophils
Band neutrophils are slightly less mature than segmented neutrophils and have indented,
unsegmented "C" or "S" shaped nuclei.  Band neutrophils normally account for approximately 5-
10% of peripheral blood leukocytes.  An increased proportion of band neutrophils can be seen in
infectious and inflammatory conditions.

An elevated concentration of band neutrophils in the blood is always the result of infection or
inflammation. In the instance of infection, the source is likely bacterial. The causes of
inflammation are varied.  Physical stressors like exercise, seizures, anxiety, tobacco use, burn
injuries, heart attack, appendicitis, and splenectomy can result in inflammation. There are a few
chronic conditions that lead to inflammation, such as inflammatory bowel disease and arthritis.
Certain medications (corticosteroids or epinephrine) are known to elevate neutrophils in the
blood.  Most severely, an inflated level of band neutrophils may be due to chronic myelogenous
leukemia—a cancer that starts inside the bones where white blood cells are produced. 
Toxic neutrophils
A common and important morphologic abnormality of neutrophils is so-called “toxic change“.
Contrary to what has been written in many books, toxic change in neutrophils is not necessarily
associated with “toxemia”. The term derives from the fact that these abnormalities were first
noticed in human patients with gram negative sepsis and endotoxemia. However, toxic change
in neutrophils do not reflect a “toxic effect” of bacteria on neutrophils but are morphologic
abnormalities acquired during maturation under conditions that intensely stimulate neutrophil
sproduction and shorten the maturation time in the bone marrow. This accelerated maturation
occurs secondary to cytokine stimulation, which is usually in reponse to inflammation. Animals
recovering from bone marrow injury or who are administered hematopoietic cytokines (e.g.
granulocyte-colony stimulating factor or G-CSF) can also show accelerated maturation in
neutrophils or toxic change. On the standard hemogram, the presence of toxic change is
reported under the WBC exam  section and is subjectively graded as mild, moderate and
marked.

Toxic changes in neutrophils include toxic granulation, toxic vacuolization, and Döhle bodies.
Toxic granulation and Döhle bodies each may be present in an individual cell without the other
finding and either change alone is sufficient to designate a neutrophil as toxic. Toxic granulation
is defined by the presence of large, purple or dark blue cytoplasmic granules in neutrophils,
bands, and metamyelocytes. Vacuoles within the cytoplasm of these same cells define toxic
vacuolization. The vacuoles are variable in size and may coalesce, sometimes distorting the
neutrophil cytoplasm to form pseudopodia. Ethylenediaminetetraacetic acid (EDTA) blood
collection may produce degenerative vacuolization; in this context, only a few, small, punched-
out appearing vacuoles may be found. However, as it may be difficult to distinguish toxic from
degenerative vacuoles, neutrophil vacuoles should not be labeled as toxic vacuoles unless
accompanied by other toxic changes. Döhle bodies appear as single or multiple blue or gray-blue
inclusions of variable size (0.1 to 5.0 μm) and shape (round, or elongated or crescent shaped) in
the cytoplasm of neutrophils, bands, or metamyelocytes. They are often found at the periphery of
the cytoplasm, near the cell membrane. These inclusions represent parallel strands of rough
endoplasmic reticulum. Toxic changes result from the action of cytokines released in response to
infection, burns, trauma, and granulocyte colony stimulating factor (G-CSF), and they indicate a
shortened maturation time and activation of post-mitotic neutrophil precursors. In the May-
Hegglin anomaly, inclusions that resemble Döhle bodies are seen. Unlike Döhle bodies,
however, the May-Hegglin inclusion is due to aggregates of non-muscle myosin heavy chain
IIA. Also seen in concert with neutrophil abnormalities are thrombocytopenia and giant platelets.
The May-Hegglin anomaly is inherited in an autosomal dominant fashion, owing to mutations in
MYH9.

Most of the toxic changes reflect asynchrony of maturation between the nucleus and cytoplasm.
During normal granulocytopoiesis, the lengthening and pinching of the nucleus are coordinated
with progressive condensation of the chromatin and loss of cytoplasmic protein synthetic
machinery (RNA in the form of ribosomes and rough endoplasmic reticulum – which imparts a
blue color to the cytoplasm with hematologic stains). With accelerated maturation, nuclear
divisions may be skipped (resulting in larger cells than normal) and the cells retain immature
features, including increased amounts of rough endoplasmic reticulum or ribosomes in the
cytoplasm and lighter chromatin than normal. Specific granules may be less visible than in
normally mature cells and, in some species, primary granules (normally inapparent in
neutrophils) retain their affinity for the stains used for peripheral blood. The cells can also have
frothy or vacuolated cytoplasm.

Normal neutrophils and neutrophils with toxic change

In a blood smear there are five main features of toxic change:

 Cytoplasmic basophilia: A streaky diffuse irregular blue appearanceto the cytoplasm. It

is due to the presence of polyribosomes and rough endoplasmic reticulum.
  Döhle bodies: These are pale round to linear blue aggregates in the cytoplasm, caused by
whorls of rough endoplasmic reticulum (see image to the right). This is often the earliest
and first indication of toxic change. However, note that healthy cats can have low
numbers of small Döhle bodies and we have observed that Döhle bodies form in
neutrophils with storage – this was by comparing freshly made blood smears to blood
smears made from the submitted stored blood in our laboratory (storage-related artifact,
for images of this, see blood artifacts under the hematology album). Thus, Döhle bodies
alone do not always indicate toxic change.
 Cytoplasmic vacuolation: These are indistinct vacuoles in the cytoplasm, giving it a
frothy appearance and is due to degranulation of lysosomes. Note that clear punctate
vacuoles are usually not attributed to toxic change, but are frequently another storage-
related artifact (these vacuoles can occur within 4 hours of blood collection).
 Nuclear immaturity: The nuclear chromatin is lighter (finer) and less coarsely clumped
than normal. This is the hardest and most subtle change to see.
 Toxic granulation: These are distinct red granules in the cytoplasm due to the primary
granules taking up stain. This is a rarely seen feature in domestic animals, but is most
commonly seen in large animals (horses, ruminants, camelids – see image below).

Toxic granulation in a camelid with enteritis.

Because toxic change usually indicates an inflammatory leukogram, it frequently accompanies a

left shift, i.e. presence of immature neutrophils (bands, metamyelocytes, myelocytes), in animals.
However, some animals can have a left shift without toxic change (e.g. some dogs with immune-
mediated hemolytic anemia) and toxic change can be present (albeit rarely) in animals without a
left shift. In the latter circumstances, we look for reasons other than inflammation for the toxic
change (e.g. marrow dysplasia, storage artifact). The degree of toxic change can also be a helpful
finding. The more severe the toxic change, the more intense the inflammatory stimulus. In one
prospective study of 105 horses presenting to an emergency clinic with medical or surgical colic,
the presence of bands and moderate to severe toxic change were associated with decreased
survival .

Hypersegmented neutrophil

Neutrophil hypersegmentation can be defined as the presence of neutrophils whose nuclei have
six or more lobes or the presence of more than 3% of neutrophils with at least five nuclear
lobes.This is a clinical laboratory finding. It is visualized by drawing blood from a patient and
viewing the blood smeared on a slide under a microscope. Normal neutrophils are uniform in
size, with an apparent diameter of about 13 μm in a film. When stained, neutrophils have a
segmented nucleus and pink/orange cytoplasm under light microscope. The majority of
neutrophils have three nuclear segments (lobes) connected by tapering chromatin strands. A
small percentage have four lobes, and occasionally five lobes may be seen. Up to 8% of
circulating neutrophils are unsegmented (‘band’ forms).

To be considered a neutrophil with a hypersegmented nucleus, the neutrophil should demonstrate

six or more lobes. Hypersegmented neutrophils are uncommon unless there is megaloblastic
hematopoiesis. Hypersegmented neutrophils may also be seen in sepsis, renal disease, and
myeloproliferative neoplasms. Megaloblastic hematopoiesis occurs when DNA synthesis is
impaired. Such conditions include deficiency of cofactors for nucleotide synthesis, such as
vitamin B12 and folate, and cases in which patients are receiving a nucleotide analog drug (such
as 6-mercaptopurine) or nuclear cofactor blocking agents (such as methotrexate) for the
treatment of neoplastic or rheumatologic conditions. Neutrophil with Pelger-Huët Nucleus
(Acquired or Congenital)

The presence of hypersegmented neutrophils is an important diagnostic feature of megaloblastic

anaemias. Hypersegmentation can also be seen in many other condition but with relatively less
diagnostic significance.
Hypersegmentation can sometimes be difficult to assert since interobserver variation is high and
segmentation may vary with race. Neutrophil nuclear hypersegmentation (more than five lobes)
is observed in megaloblastic anemia, chronic infection, myelodysplastic syndrome, iron

deficiency, hypogonadism, and chronic myelogenous leukemia .

Megaloblastic anemia

Neutrophil hypersegmentation is one of the earliest, most sensitive and specific signs of
megaloblastic anemia (mainly caused by hypovitaminosis of vitamin B12 & folic acid). Nuclear
hypersegmentation of DNA in neutrophils strongly suggests megaloblastosis when associated
with macro-ovalocytosis. If megaloblastosis is suspected, a formal lobe count/neutrophil (i.e.
lobe index) above 3.5% can be obtained. Hypersegmentation persists for an average of 14 days
after institution of specific therapy.

Hypersegmented neutrophil in a a patient with vitamin B12 deficiency.  Hypersegmented

neutrophils have 6 or more nuclear lobes. They are typically seen in megaloblastic anemia due to
vitamin B12 or folic acid deficiency, but may also be present in myelodysplastic syndromes and
rare congenital conditions.
Hyposegmented Neutrophils

The presence of hyposegmented neutrophils can be an acquired phenomenon, as a result of

severe infection, burns, malignancy, chemotherapy or other drugs such as sulfonamides. When
the causative agent is removed, the cells will return to normal.
Percentages of neutrophils affected will vary in this condition. Hyposegmented neutrophils as an
aquired phenomenon are sometimes referred to as pseudo-Pelger-Huet cells.

Pseudo-Pelger-Huet Anomaly or Pelgeroid change ( PPHA) is characterized by

hyposegmentation of the neutrophil nucleus and chromatin clumping. 

Morphologic mimics of Pelger-Huet anomaly ( Psuedo-Pelger-Huet Anomaly)  are more

common than the true disorder.The changes of hypolobation can be seen primarily in three
circumstances associated with- 

1. Reactive conditions associated with severe infections,

2. Medications or drugs such as mycophenolate or valproate, 
3. Myelodysplastic syndromes or other myeloid stem cell disorders. 

Distinguishing true Pelger-Huet anomaly from mimics, though straightforward, is usually

important as it may lead to unnecessary investigations like bone marrow examination.  

In the case of myelodysplastic syndrome (MDS) or other myeloid neoplasms, the hypolobation is
often accompanied by hypogranulation of neutrophils, as well as other dysplastic changes seen in
erythroid elements, platelets, and megakaryocytes. In particular, isolated isochromosome 17q is a
cytogenetic abnormality specifically associated with these changes. 

In both reactive conditions and dysplastic-type changes, only a subset of neutrophils is usually
affected, in contrast to the entire population in Pelger-Huet anomaly. Toxic changes of
granulocytes may also be seen in cases associated with infection. 

Identification of drug-related cases requires knowledge of the clinical history and a high index of
suspicion.
In Our case, 72 years old male patient presented with pancytopenia (Haemoglobin : 10.8 g/dl,
leukocyte count : 3 × 10^9/L with and platelet counts : 29 × 10^9/L). 

Peripheral smear examination revealed 7-8% of the neutrophils having bi-lobed nuclei with
condensed chromatin and hypogranulation of cytoplasm.

Giant toxic neutrophils

Can be seen occasionally in normal peripheral blood smear

Larger than normal neutrophils and generally hyperlobulated

Found in frequency of 1 in every 20,000 neutrophils, but increase in disease state

Commonly seen in cats and dogs

REFERANCES

Willard, M.D.; Tvedten, H. and Turnwald, G.H. Small animal clinical diagnosis by laboratory
methods. 3rd edition. 1999.

 Effect of time and storage on toxic or pseudo‐toxic change in canine neutrophils. Bau-
Gaudreault L, Grimes C. Vet Clin Pathol. 48:400-405, 2019.
Toxic Neutrophils in Cats: Clinical and Clinicopathologic Features, and Disease Prevalence and
Outcome—A Retrospective Case Control Study. Segev G, Klement E, Aroch I. J Vet Int Med.
20:20-31, 2006.

Clinical, Biochemical, and Hematological Characteristics, Disease Prevalence, and Prognosis of

Dogs Presenting with Neutrophil Cytoplasmic Toxicity. Aroch I, Klement E, Segev G. J Vet Int
Med. 19: 64-73, 2005.

Association of Presence of Band Cells and Toxic Neutrophils with Systemic Inflammatory
Response Syndrome and Outcome in Horses with Acute Disease. Lambert JL, Fernandez NJ,
Roy M. J Vet Int Med. 30:1284-1292, 2016.