Platelet morphology: reliable technology doesn’t require duplication

Right now, more than ever, the economic landscape in healthcare requires optimal utilization of existing resources to control costs. In this reality, newly developed technologies should not only be “new” to the market, but must also offer real-life clinical improvements to patients and providers and also be more cost-effective than existing technologies.

This is especially true in the laboratory. While new tests are constantly being created, often little research and attention is being paid to tests that already exist and may not be used to their full potential. This inadequate utilization not only leads to the unintended practice of requiring more—not better, just more—testing; it also reduces the return on investment for existing technologies: a direct contradiction to the current needs of healthcare systems worldwide.

One classic example of underutilization is the Complete Blood Count (CBC), one of the most affordable and most commonly used tests in medicine. The CBC is composed of several parameters relating to the cellular composition of the blood, yet only a few of these parameters are used routinely by clinicians in their diagnostic and therapeutic decisions. For the most part, clinicians use the CBC to count cells: red blood cells (RBC), white blood cells (WBC), platelet, and differential counts. But counting is only part of the equation—only a portion of the total value that can be offered by the CBC.

CBC and cell morphology

Today certain automated hematology analyzers are capable of analyzing full blood cell morphology. Instead of simply counting cells, these instruments are capable of looking at multiple morphologic features of all blood cell types, such as their size, the nuclear/cytoplasmic ratio, the granularity of their cytoplasm, the complexity of the nucleus, and even the internal density of the cell. Of the morphologic parameters generated by these instruments, some can be used in the laboratory for setting up decision rules that can flag the presence of clinically important morphologic changes (e.g., neutrophil hypogranularity),¹ while others are reportable parameters that we are all familiar with.

Some good examples of reportable morphologic parameters are the RBC Mean Corpuscular Volume (MCV) and Red Blood Cell Distribution Width (RDW), CBC parameters commonly used to determine anemia sub-classification. Examining the morphology of RBCs as part of the CBC thus provides clinicians with the information they need to determine the most likely root cause of the anemia and direct the complete diagnostic work-up effectively.

On the other hand, platelet morphology is vastly underused—despite numerous diagnostic applications for the Mean Platelet Volume (MPV). The MPV can be used to improve the detection of platelet morphologic abnormalities such as platelet clumps and giant platelets, provide information for thrombotic risk,^2-6 predict the likelihood of bone marrow metastasis in cancer patients,⁷ provide prognostic information for patients with myelodysplasia,⁸ and help identify the origin of thrombocytopenia (e.g., whether caused by decreased bone marrow platelet production or increased peripheral destruction).^9,10

MPV and thrombocytopenia

A healthy bone marrow produces sufficient platelets, and a healthy patient will have a normal platelet count. When the platelet count decreases, a healthy bone marrow will respond by increasing platelet production.^11-13 If platelets are being destroyed in the periphery, then platelet production by the healthy bone marrow increases in kind, resulting in high levels of immature platelets circulating in the blood. Alternatively, a diseased bone marrow cannot produce sufficient platelets; thus thrombocytopenia caused by an unhealthy bone marrow will result in a very low number of immature platelets.

Immature platelets can be differentiated from their mature counterparts by distinct morphologic features: they tend to be larger in size and to have reticulated cytoplasm due to remnants of RNA. Since immature platelets are typically larger, their presence in increased numbers leads to higher MPV. Newer technologies have also been developed that can measure the fraction of immature platelets by recognizing them, thanks to their reticulated cytoplasm. The MPV, however, has distinct advantages: its measurement is directly correlated to platelet size and the MPV is routinely reported as part of all CBCs at no additional cost. Therefore the MPV results are available at exactly the same time the platelet count is reported and thrombocytopenia is identified, without the need for any additional testing to be ordered by the clinician.

Since increases in immature platelets are associated with higher MPV, multiple studies have clearly demonstrated the clinical value of this parameter to discriminate low production from increased destruction of platelets as causes of thrombocytopenia.^7,9,10

Reticulated platelets and thrombocytopenia

New technologies have been introduced recently which rely on fluorescence methods to recognize reticulated immature platelets and thus determine the cause of thrombocytopenia.

This approach, however, has certain important challenges. First and foremost, because reticulated platelet measurement is not part of the routine CBC, a new test must be ordered separately. Therefore, once a patient is diagnosed with thrombocytopenia, a sample may need to be run again, or the patient may even need to return to the laboratory for a new draw, and the clinician has to wait for new test results before he can evaluate likely causes for the low platelet count.¹⁴

Second, tests examining reticulated cytoplasm require intra-cellular staining to identify RNA remnants. There are strict storage temperature and duration requirements that must be met for optimal measurement of reticulated platelets.^14,15

Reasons for under-utilization of MPV

The MPV is a perfect example of underutilization of a valuable existing clinical parameter in favor of “newer” testing. One of the reasons may be that “newer” implies advancement, but upon closer analysis one can realize that often the most beneficial solution for patients and laboratories may rest in improving the clinical utilization of already existing tests. This approach is also better suited for the economic pressures of modern day laboratory medicine.

Another reason for the underutilization of the MPV may be the variable performance of instruments in reporting this parameter.¹⁶ Hematology analyzers rely on direct current impedance to identify cellular size, and since platelets are the smallest particles in the blood, their volume measurement requires very complex data analysis algorithms and a high level of expertise dealing with impedance technology. Also, since platelets are not perfectly round, volume information based on optical measurements may be directly impacted by the direction the platelet was facing when analyzed. As a result, some cell counting technologies fail to consistently report the MPV as part of their CBC results.

MPV—an old/new parameter with extensive value

For those technologies capable of reliably measuring the MPV, its clinical applications are multiple. In addition to identifying whether thrombocytopenia is being caused by a diseased bone marrow, clinical applications of the MPV include:

• Effective marker of thrombotic risk^2-6
• Accurate prognosticator in myelodysplasia (MDS)⁸
• Improved detection of platelet morphologic abnormalities within the laboratory.

As a marker of thrombotic risk, multiple studies have shown that patients with higher MPV are at increased risk for a thromboembolic event. This is most likely due to the fact that immature platelets (typically larger in size) are more reactive than their older counterparts. This has been demonstrated in different clinical scenarios:

• In one study with more than 25,000 patients and 12 years of follow-up, patients with higher MPV had higher incidence of venous thromboembolism. There was a 1.5-fold increase in risk when the thromboembolism was unprovoked (e.g., absence of risk factors such as smoking, cancer, prothrombotic mutations), and 1.3-fold increase in risk when provoked.²
• Higher MPVs were observed in patients undergoing acute myocardial infarction. In addition, when a myocardial infarction occurred, the risk of death was higher in patients with a higher MPV.^3-6
• Among myocardial infarction patients who underwent coronary angioplasty, those with higher MPV had a higher risk of restenosis.³

As a marker of prognosis in MDS, the MPV can be used alone or in combination with the platelet count. Since MDS leads to thrombocytopenia, in those cases when the bone marrow is not yet severely affected and is still able to respond to increased levels of thrombopoietin, there will be an increased number of immature platelets—and thus a higher MPV. Conversely, if the MPV is low in spite of the decreased platelet count, this indicates a more advanced or aggressive case of MDS, where the bone marrow is completely unable to respond to the thrombopoietic stimulus. If used in isolation, both the MPV and the platelet count have comparable performance in discriminating patients with better or worse prognosis at the time of diagnosis of MDS. Used together, studies have shown that the combination of these parameters has an even better prognostic performance.⁸

Finally, the MPV can be used in the laboratory for improved detection of platelet morphologic abnormalities such as giant platelets and platelet clumps—critical abnormalities that can cause platelets to be misinterpreted as WBCs and lead to erroneous results in the CBC. While most instruments have built-in flags for these abnormalities, flags are never perfect; false negatives can lead to the laboratory reporting out inaccurate results, and false positives can create an unnecessary added workload.

These types of abnormalities are reflected by morphologic parameters such as the MPV and the platelet distribution width (PDW). In certain hematology instruments, these parameters can be used for writing user-derived decision rules that trigger specific alarms indicating the need for a microscopic review. For example, laboratories that wish to improve their sensitivity in detecting platelet clumps can write a decision rule where cases with low platelets and increased MPV and PDW are reviewed even when a platelet clump flag was not present. This allows for the detection of even smaller clumps that typically would not have been flagged by the instrument. And unlike the built-in flags for these abnormalities that can only be turned on or off and do not give the operator any flexibility to adjust the detection sensitivity and specificity, the use of these morphologic decision rules is totally flexible, because the operator chooses the exact cut-off points for triggering the flag. This is a great advance since users can now tailor the performance of their instruments to meet the exact needs of their laboratory, instead of being forced to accept the flag performance as established by the manufacturer.

Optimizing MPV use in the laboratory

In summary, as a clinical diagnostic tool, the MPV offers several advantages. It does not add any cost for the patient, since it is part of a standard CBC. In cases of thrombocytopenia, there is no need for additional testing orders by the clinician and no return visit to the laboratory for the patient to evaluate the etiology of thrombocytopenia. The MPV can serve as a thromboembolic risk marker and predict prognosis in hematological neoplasia, among other diagnostic uses. It also does not have marked stability variations in different ranges of storage temperature and duration.

Finally, laboratories can use the MPV, along with the PDW and platelet count, to write decision rules where cases likely to have platelet clumps or giant platelets are reviewed even when a standard instrument flag is not raised. This allows laboratories to improve the detection of these important morphologic abnormalities, and by adjusting the cut-off points of these decision rules, this improvement can be achieved with minimal impact on the number of false positive cases.

Given the economic pressures of modern-day medicine, it is critical that laboratories make the most of existing resources. As such, the MPV, with its extensive diagnostic potential, long-term presence as part of the routine CBC, low testing burden on clinicians and patients, and potential flagging benefits for the laboratory, is a great place to start.

References

1. Miguel A, Orero M, Simon R, et al. Automated neutrophil morphology and its utility in the assessment of neutrophil dysplasia, Lab Hematol. 2007;13(3):98-102.
2. Braekkan SK, Mathiesen EB, Njolstad I, et al. Mean platelet volume is a risk factor for venous thromboembolism: the Tromso Study, Tromso, Norway. J Thromb Haemost. 2010;8(1):157-162.
3. Chu SG, Becker RC, Berger PB, et al. Mean platelet volume as a predictor of cardiovascular risk: a systematic review and meta-analysis. J Thromb Haemost. 2010;8(1):148-156.
4. Cameron HA, Phillips R, Ibbotson RM, Carson PHM. Platelet size in myocardial infarction. Br Med J. 1983;287:449.
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8. Bowles KM, Warner BA, Baglin TP. Platelet mass has prognostic value in patients with myelodysplastic syndromes. Br J Haematol. 2006;135(2):198-200.
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10. Bowles KM, Cooke LJ, Richards EM, Baglin TP. Platelet size has diagnostic predictive value in patients with thrombocytopenia. Clin Lab Haematol. 2005;27(6):370-373.
11. Bessman JD, Williams LJ, Gilmer PR Jr. Mean platelet volume. The inverse relation of platelet size and count in normal subjects, and an artifact of other particles. Am J Clin Pathol. 1981;76(3):289-293.
12. Giles C. The platelet count and mean platelet volume. Br J Haematol. 1981;48(1):31-37.
13. Thompson CB, Jakubowski JA. The pathophysiology and clinical relevance of platelet heterogeneity. Blood. 1988;72(1):1-8.
14. Osei-Bimpong A, Saleh M, Sola-Visner M, Widness J, Veng-Pedersen P. Correction for effect of cold storage on immature platelet fraction. J Clin Lab Anal. 2010;24(6):431-433.
15. Albanyan A, Murphy MF, Wilcock M, Harrison P. Changes in the immature platelet fraction within ageing platelet concentrates. J Thromb Haemost. 2008;6(12):2213-2215.
16. Briggs C, Kunka S, Machin SJ. The most accurate platelet count on the Sysmex XE-2100. Optical or impedance? Clin Lab Haematol. 2004;26(2):157-158.

Fernando Chaves, MD, is a board-certified anatomic and clinical pathologist and a board-certified hematopathologist. He is the director of global scientific affairs at Beckman Coulter, fostering the research of new clinical applications for data generated during a routine Complete Blood Count and differential and on the optimization of workflow in the hematology laboratory. While his area of major expertise is bacterial infection and sepsis, he has also researched myelodysplasia, myeloproliferative neoplasms, acute leukemias, vitamin B12 and folate deficiencies, dengue fever, and malaria, among other topics.