The effect of smear cells on the classification of white blood cells and their role in morphological diagnosis

01

Preface

Smeared cells, also known as degenerative cells or basket cells, are often caused by mechanical force during slide pushing, or can be caused by the degeneration of cell aging, usually a swollen nucleus with a vague structure and no cytoplasm, which is uniformly lilac-red

02

Case 1

Case after

(i)

The patient, 72 years old, was admitted to the hospital with "poor tolerance, fatigue, and yellow face for 1 week", anemia, yellowing of the skin and sclera of the whole body, multiple lymphadenopathy that can be touched on both sides of the neck and armpits, and untouched under the hepatic and spleen ribs

Figure 1 Blood count results report

The blood routine showed that the patient had severe anemia, a significant increase in the white blood cell count and lymphatic ratio, and a part of the scatter point with high lateral fluorescence values above the lymphocyte pellet and the neutrophil pleucum in the scatter chart, and the instrument alarmed that "leukocytosis, lymphocytosis, immature granulocytes, blast cells/abnormal lymphocytes? ", after the automatic tablet pusher pushing the film, staining microscopy, found that a large number of smeared cells appeared in the field of view, referring to the relevant literature [1] and the guidelines for the normalization of blood cell analysis reports [2], 20% albumin and EDTA-K2 anticoagulant whole blood were mixed in a 1:5 ratio, and then the same method was pushed and stained again (Figure 2), and the white blood cell classification and morphological analysis

Fig.

After albumin protection treatment, the smear cells are significantly reduced, microscopic mature small lymphocytes are more common, leukocyte classification and morphological analysis report: neutral neutrin 0.

On the same day, the clinical department further sent a bone marrow specimen for examination, and the reported results were as shown in Figure 3:

Figure 3 Bone marrow cell morphology report sheet

Flow cytometry results: CONFORMED TO CD5+CD10-SMALLB CELL LYMPHOMA PHENOTYPE, WITH HIGH

Case studies

(i)

Chronic lymphocytic leukemia (CLL) blood picture is mainly manifested by increased white blood cells, increased lymphatic ratio, mainly morphologically mature small lymphocyte proliferation, more common smear cells, these smear cells are actually degenerate lymphocytes: because there is a wave protein in lymphocytes related to cell hardness, its expression is abnormal can increase the fragility of lymphocytes [3], making it easy to break during the pushing process, which is manifested as smeared cells

In view of the blood picture characteristics of this case, we tried several classification methods and compared

Table 1 Comparison of leukocyte classification results for different counting methods

As can be seen from the table, the results obtained by the traditional classification method of blood push tablets that are not counted in the application of cells show that the lymphatic ratio is lower than that of the instrument, but the difference is not large, perhaps due to the significant increase in the lymphatic ratio in the peripheral blood of the patient, the ratio of neutrophils and monocytes other than lymph is not more than 30%, even if a part of the lymph is destroyed in the process of extracorporeal tablets and becomes a smeared cell, it cannot change the trend of

The other two classification methods, namely the classification results before albumin treatment (counting the smear cells) and after the albumin addition (not included in the smeared cells), are similar to the instrument classification results, which to a certain extent reflect the actual situation of the patient's peripheral blood leukocyte classification, and can be used as an attempt or reference

Of course, the manual classification results will also be affected by factors such as the classification area, blood cell recognition skills, etc.

brief summary

(i)

In this case, the instrument classification lymphatic ratio is significantly increased and the appearance of a large number of smeared cells in the peripheral blood smear gives us a more intuitive prompt, from the blood picture, bone marrow image characteristics are not difficult to point to the diagnosis of lymphocyte proliferative diseases, flow immunotype results confirm the morphological judgment

03

Case 2

Case after

(i)

The patient, a 64-year-old male, was admitted to the hospital

Routine blood showed: WBC 12.

Fig.

Figure 5 Blood smear cell morphology (10×100-fold)

The cells labeled in the figure are larger in body 1 and 2, and there are more coarse particles in the plasma, which need to be distinguished from the neutrophils with more particles; Cell 3 and 4 cytoplasm are abundant, gray-blue, with no or only a small number of particles in the plasma, and the nuclear chromatin is coarse, which needs to be distinguished from monocytes; Cells 5 and 6 are late red and late larocytes, respectively; Cell 7 is large and the nuclear chromatin is more delicate, which needs to be distinguished from

Clinically, in order to exclude hematologic diseases, a bone marrow cell smear was sent at the same time, but the material was not ideal, the bone marrow had nucleated cell proliferation was reduced, and degenerative cells
could be seen.
Careful reading of the film revealed that some of the larger, easily destroyed abnormal lymphocytes of the cell body were found in the thick area of the smear (shown by the arrow in Figure 6), and particles were visible in some of the plasma, and particles were still scattered around the smeared cells
.

Figure 6 Bone marrow cell morphology (10×100)

Translucent cell analysis: LARGE T granular lymphocytes of CD3+CD4+ CD5+ CD7dim CD8- CD57Partial+ CD45RA-CD45RO+TCRab+ are visible, suggesting T-LGL
.

Case studies

(i)

In case 2, the results of the blood routine instrument classification showed that the lymphocyte ratio was in the normal range, the blood slide smear cells were not as obvious as in case 1, and the number of large granular lymphocytes with large cells was not much, and the morphology was not very typical, so at the beginning it was not considered in the direction of large granular lymphoma, but according to the characteristics of young red, larval cells and mature red blood cells with target shape and lobe red in peripheral blood, consider whether there was myelofibrosis, due to the difficulty of puncture The specimen was not ideal, and the bone marrow biopsy results could not be referred to
。 However, looking back at the peripheral blood and bone marrow smears, the smear cells in the field of vision are indeed giving us a hint of certain information, and carefully looking for abnormalities in the morphology of blood cells with the question of the cause of the smear cells can be said to be a breakthrough in the morphological diagnosis of this case
.

brief summary

(i)

Macrogravular lymphocytic leukemia is a class of disease characterized by clonal proliferation of circulating large granular (aniline blue granules) lymphocytes[4], which is divided into T-cell macrogranulocytic leukemia (T-LGL), NK-cell chronic lymphoproliferative disease, and aggressive NK-cell leukemia according to the origin of tumor cells
.

T-LGL is more common in the elderly, the median diagnosis age is 60 years old, most of them are inert, the clinical manifestations are insidious, most of them are found to be abnormal in the blood routine, often accompanied by anemia, neutrophils and thrombocytopenia, and large granular lymphocytes
can be seen in blood smears.
In normal blood, large granular lymph, composed of T and NK cells, accounts for 5% of lymphocytes, sometimes up to 10 to 15%, making the morphological diagnosis of T-LGL more difficult, and the smeared cells in this case provide ideas
for morphological diagnosis.

04 Summary

When smeared cells appear on peripheral blood smears, it is recommended to add albumin to protect cells in a certain proportion and push the slide staining microscopy classification
.
A large number of smeared cells appear on the blood smear can first refer to the percentage of lymphocytes classified by the blood routine instrument, if the proportion of lymphocytes increases significantly (generally up to 80% or more), due to the low proportion of other white blood cells such as neutrophils and monocytes, the percentage of lymphocytes will not have much impact, this situation can be treated without albumin, manual classification and review can try two methods: 1, classification of 100 white blood cells is not counted in the smear cells, and when describing the morphology, the remarks are visible / Easy to see smearing cells
.
2.
When classifying, the smear cells are directly classified as lymphocytes and recorded in 100 white blood cells, and the description indicates that "100 cells are classified to see / including X smear cells"
.

Of course, if the instrument classifies the percentage of lymphocytes in the normal range or the increase is not obvious, the influence of other cells should be taken into account, and it is recommended to add albumin protection to push the lens to verify whether the smeared cells are true lymphocytes and whether there are any abnormalities in morphology
.

In short, smearing cells, although affecting the hand-sorted lymphocyte ratio, also serves as a clue to the disease to alert us to the presence of
abnormal cells.
Especially in non-CLL cases, thinking about and tracking the causes of smeared cells can help morphology examiners find clues and abnormalities, thus avoiding missed diagnosis
of the disease.

References

[1] Matthew L， Vincent Z， Ricardo D， et al.
Albumin enhanced morphometricimage analysis in CLL.
Cytometry B Clin Cytom.
2004；57(1)：7-14.

Hematology and Humors Group of Laboratory Medicine Branch of Chinese Medical Association.
Guidelines for the Standardization of Blood Cell Analysis Reports[J], Chinese Journal of Laboratory Medicine, 2020;43(6):619-627.

[3] Szerafin L， Jakó J，Riskó F， Hevessy Z.
The prognostic value of smudge cells (Gumprechtshadows) in chronic lymphocytic leukaemia.
Orv Hetil.
2012 Nov4； 153(44)：1732-7.

[4] Lamy T， Moignet A， Loughran TP Jr.
LGL leukemia： from pathogenesis to treatment.
Blood.
2017 Mar2； 129(9)：1082-1094.