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Year : 2021  |  Volume : 64  |  Issue : 4  |  Page : 850-851
Critical inclusions in neutrophils—Cryptic and elusive entities


1 Department of Laboratory Medicine, Sir H.N. Reliance Foundation Hospital and Research Centre, Mumbai, Maharashtra, India
2 Department of Internal Medicine, Sir H.N. Reliance Foundation Hospital and Research Centre, Mumbai, Maharashtra, India
3 Department of Hematology, Sir H.N. Reliance Foundation Hospital and Research Centre, Mumbai, Maharashtra, India

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Date of Submission24-Nov-2020
Date of Decision09-Dec-2020
Date of Acceptance18-Feb-2021
Date of Web Publication20-Oct-2021
 

How to cite this article:
Ray M, Sharma S, Vazifdar A, Thacker HP, Shah SS. Critical inclusions in neutrophils—Cryptic and elusive entities. Indian J Pathol Microbiol 2021;64:850-1

How to cite this URL:
Ray M, Sharma S, Vazifdar A, Thacker HP, Shah SS. Critical inclusions in neutrophils—Cryptic and elusive entities. Indian J Pathol Microbiol [serial online] 2021 [cited 2021 Nov 27];64:850-1. Available from: https://www.ijpmonline.org/text.asp?2021/64/4/850/328542




Blue–green inclusions in the cytoplasm of neutrophils is a rare finding, associated with poor prognosis in critical patients. These inclusions tend to be missed on peripheral blood smear examination owing to their extreme scarcity. They are associated with high short-term mortality rate.[1],[2] There is no standardized term for these inclusions, which makes reporting perplexing for pathologists and interpretation confusing for clinicians. Thorough search of medical literature revealed only 81 cases reported thus far.[2],[3],[4],[5],[6]

A 52-year-old female presented with gradually increasing swelling of left forearm, associated with patchy purpura, of 5 days duration. She was prescribed anti-inflammatories and antibiotics by her local physician for cellulitis. She developed altered sensorium of 24-h duration. Her comorbidities included hypertension, obsessive compulsive disorder, malnutrition, and osteoporosis. Examination revealed undernourishment, hypotension, tachypnoea, tachycardia, and severe hypoglycemia, with random blood glucose of 10 mg/dl. She was admitted in the intensive care unit in view of hypoglycemic encephalopathy. Her hemogram revealed anemia with hemoglobin of 11.2 g/dl, relative neutrophilia, and adequate platelets. Liver profile showed raised serum (S.) aspartate and alanine transferases (ALT) of 338 U/L and 349 U/L, respectively. S.Creatinine was low (0.32 mg/dl). S.Lactate and S.Procalcitonin were elevated at 45.4 mg/dl and 9.1 ng/ml, respectively. High-resolution computed tomography-chest indicated bilateral pneumonia. Ultrasound-abdomen was unremarkable. Cultures and polymerase chain reaction for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) were negative. She was administered injection (inj.) dextrose, piperacillin-tazobactum, clindamycin, and vasopressor infusion. Over the next few days, she developed worsening acute kidney injury and acute liver injury with raised S.ALT, peaking at 1470 U/L on day 6. Vasopressors were up-titrated and antibiotics escalated to inj.meropenem. Dialysis was initiated. She developed disseminated intravascular coagulopathy on day 5. Her blood picture showed progressively increasing neutrophilic leukocytosis with toxic changes. [Figure 1]. She had worsening anemia, with presence of normocytic red blood cells (RBCs), acanthocytes, and nucleated RBCs, accompanied by thrombocytopenia. She was transfused with relevant blood products. Her condition deteriorated and she was intubated. On day 9, her white cell counts declined, when neutrophilic inclusions (~2.0%) became apparent. On Leishman stain, these inclusions were coarse, greenish-blue, refractile, of variable sizes, ranging in number from one to several [Figure 2]. They were also noted and reported (~5.0%) on the morning of day 10, shortly after which she succumbed. Her peripheral blood smears were reviewed retrospectively for these critical inclusions. We found that they had been present from the day of admission, seen extremely rarely in less than 1.0% of neutrophils (ranging from 0.1% to 0.7%), and gradually increasing during hospitalization.
Figure 1: Peripheral blood smear (Leishman stain, ×1000) - Neutrophils showing toxic granules. Subtle critical inclusions can also be noted (arrow). Normocytic, normochromic RBCs and many acanthocytes are seen in the background

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Figure 2: Peripheral blood smear (Leishman stain, ×1000) - Greenish-blue, coarse, refractile inclusions of varying sizes, shapes and numbers are seen within the cytoplasm of neutrophils

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Neutrophilic blue-green cytoplasmic inclusions have been reported in acute hepatic failure, lactic acidosis, multiorgan failure, septic shock, trauma, and SARS-CoV-2.[1],[2],[3],[4],[5],[6],[7],[8] They occur over a wide range of ages, in both sexes.[1],[7] They can be found, albeit less commonly, in monocytes too.[1],[2],[4],[8] These inclusions are ill-defined, coarse, refractile, refringent, and vibrant.[4],[9] Their color has been described as “blue-green,” “green,” and “bright green,” This subtle variation in color perception may be because of specific Romanowsky stain utilized and the illumination source of the viewing microscope. The composition and pathogenesis of these granules are not clearly understood. They are thought to be composed of lipofuscin-like material, containing approximately of two-third protein, one-third lipid, and <2.0% metal, like iron.[1],[8] They show positivity with oil red O and Ziehl-Neelsen stains, and presence of lipid-rich cytoplasmic inclusions by transmission electron microscopy. They are negative on Perls', Periodic acid-Schiff, myeloperoxidase, Warthin-Starry, Gomori methenamine silver, and Gram stains.[1] These inclusions may be produced secondary to acute hypoxic hepatic injury or other tissue injury. They are therefore commonly, but not exclusively, associated with high liver transaminases, lactate dehydrogenase, and lactic acid levels.[1],[7] Similar findings were seen in our case too. Few cases are not critical, not associated with liver insult, or infections. They may be because of sub-clinical tissue injury.[7] These inclusions are associated with high mortality rates (65-100%) within short periods of detection in critically ill patients.[1],[2] One retrospective study reported a lower short-term mortality rate of 31%.[7] Our patient expired within 48 h of detection of the critical inclusions. Our review of smears revealed their evidence extremely sparsely at an earlier stage, signifying importance of knowledge of these entities and vigilant screening of blood smears. Seventy-two hours following detection is considered the “critical window” for patient survival.[5]

Due to the above-mentioned mortality factor, the inclusions have been termed “crystals of death,” “critical green inclusions.” “green crystals of death,” “death cells,” etc.[1],[4],[5],[8] Labelling them with color (?blue/?green) is dependent on the subjectivity of the reporting pathologist, and as such does not convey the significance to the physicians. On the other hand, implying a consequence, such as death, to deliver impact seems rather harsh. Besides, not all cases are associated with death.[1],[5],[6],[7],[8],[9] For these reasons, we propose to refer to these inclusions concisely as “CRITICAL INCLUSIONS.”

Critical inclusions can be utilized as prognostic markers in severely ill patients. Increasing awareness among pathologists and clinicians can forewarn and direct investigations towards possible undetected tissue damage or lactic acidosis, and thereby prevent further clinical deterioration. A good clinic-pathological correlation will serve in better understanding of these enigmatic molecules.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.



 
   References Top

1.
Hodgson TO, Ruskova A, Shugg CJ, McCallum VJ, Morison IM. Green neutrophil and monocyte inclusions-time to acknowledge and report. Br J Haematol 2015;170:229-35.  Back to cited text no. 1
    
2.
Cantu MD, Towne WS, Emmons FN, Mostyka M, Borczuk A, Salvatore SP, et al. Clinical significance of blue-green neutrophil and monocyte cytoplasmic inclusions in SARS-CoV-2 positive critically ill patients. Br J Haematol 2020;190:e89-92.  Back to cited text no. 2
    
3.
Vicente-Steijn R, Tomé A, Maduell F, Xipell M, Castro P, Molina A. Green inclusions in neutrophils: A critical finding that must be reported. Int J Lab Hem 2020;42:e101-4.  Back to cited text no. 3
    
4.
Dienstmann G, Comar SR, Souza MLR, Ruaro G, Leite LAC. Critical blue-green inclusions in neutrophil and monocyte cytoplasm in a healthy patient affected by COVID-19. Hematol Transfus Cell Ther 2020;42:318-19.  Back to cited text no. 4
    
5.
Marshall M, Rios I. Green neutrophilic inclusions and a novel association with gangrenous ischemic colitis. J Case Rep Images Pathol 2020;6. Available from: https://www.idoriums.com/edpanel/media/Z11_Journal%20of%20Case%20Reports%20and%20Images%20in%20Pathology/2020/pdf/100041Z11MM2020.pdf. [Last accessed on 2020 Nov 23].  Back to cited text no. 5
    
6.
Bowen RK, Koutsavlis I. The rare green-blue neutrophil inclusion bodies. Worth reporting in blood films? A case of a patient with polytrauma. Clin Case Rep 2020;08:3549-50.  Back to cited text no. 6
    
7.
Courville EL, Crisman S, Linden MA, Yohe S. Green neutrophilic inclusions are frequently associated with liver injury and may portend short-term mortality in critically ill patients. Lab Med 2017;48:18-23.  Back to cited text no. 7
    
8.
Gorup T, Cohen AT, Sybenga AB, Rappaport ES. Significance of green granules in neutrophils and monocytes. Proc (Bayl Univ Med Cent) 2017;31:94-6.  Back to cited text no. 8
    
9.
Haberichter KL, Crisan D. Green neutrophilic inclusions and acute hepatic failure: Clinical significance and brief review of the literature. Ann Clin Lab Sci 2017;47:58-61.  Back to cited text no. 9
    

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Correspondence Address:
Manjusha Ray
Department of Laboratory Medicine, Sir H.N. Reliance Foundation Hospital and Research Centre, Mumbai - 400 004, Maharashtra
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/IJPM.IJPM_1351_20

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