Morphology Case Study. Presented by Niamh O Donnell, BSc, MSc. Medical Scientist Haematology Laboratory Cork University Hospital
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1 Morphology Case Study Presented by Niamh O Donnell, BSc, MSc. Medical Scientist Haematology Laboratory Cork University Hospital
2 41 year old male presented to GP for routine check-up in May FBC Results: WBC x10^9/l RBC 4.71 x10^12/l HB 14.9 g/dl HCT l/l MCV 93.9 fl MCH 31.7 pg Neutrophils 7.93 Lymphocytes Monocytes 2.81 Eosinophils 0.26 Basophils 0.00 MCHC 33.8 g/dl PLTS 203 x10^9/l
3 FBC in UHK showed leucocytosis with a lymphocytosis Blood film displayed lymphocytosis and smear cells Flow cytometry Phenotype: CD
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5 Diagnosed with B-Cell Chronic Lymphocytic Leukaemia (CLL) in Slow progressing CLL April 2016: Patient symptoms: Lymphadenopathy night sweats, loss of appetite weight loss Flow cytometry: B-CLL Phenotype Cytogenetics: CD38+, TP53 wildtype IGHV unmutated
6 IGHV Antigen binding site Antigen binding site
7 FCR (Fludarabine, cyclophosphamide rituximab) chemotherapy started on July 2016 Following 3 cycles of FCR chemotherapy, patient presents with fever end of August Pt admitted to UHK in September 2016 Query sepsis due to chemotherapy
8 06/07/2016 Pre Chemo1 15/07/ /08/16 Pre Chemo2 31/08/16 Pre Chemo3 WBC /09/16 RBC HB HCT MCV MCH MCHC PLT NEUTS LYMPHS MONOS EOSINS BASOS
9 No source of infection identified, patient transferred to CUH on 14th September 2016 Chemotherapy discontinued, Initial Blood results on admission to CUH: WBC RBC HB PLT NEUTS LYMPHS 3.5 x10^9/l 3.01 x10^12/l 10.0 g/dl 193 x10^9/l 2.65 x10^9/l 0.32 x10^9/l
10 Patient continued to have intermittent pyrexia ( Temperature 40-41⁰C), had generalised skin shedding, continue trying to identify source of sepsis: Negative Results: Enterovirus/Rubella/ Varicella Zoster/CMV/EBV/ HepB/HepC /CSF/Aerobic and anaerobic blood cultures GCSF Given no response Antivirals, antifungals, antibiotics administered
11 Blood Counts September /9/16 18/9/16 22/9/16 25/9/16 27/9/16 28/9/16 29/9/16 3/10/16 Wbc x10^9/l Neuts x10^9/l Hb g/dl Plts x10^9/l Ferritin ng/ml CRP mg/ml <10 <
12 Blood Film
13 Bone Marrow aspirate
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18 BMA (Sept 2016): 14% macrophages with prominent phagocytic activity. Treatment commenced Flow cytometry (Sept 2016): No phenotypic evidence of CLL BMA (Oct 2016) : macrophages with phagocytic activity 3/10/16 7/10/16 9/10/16 10/10/16 14/10/16 Fibrinogen g/l
19 Diagnosis
20 Recap year old male, history CLL 2011 continuing pyrexia of unknown origin Pancytopenic: Ferritin Platelets Hb Wbc BMA: Macrophages with phagocytic activity Fibrinogen
21 Diagnosis
22 Diagnosis...HLH HLH = Hemophagocytic Lymphohistiocytsosis Rare, aggressive syndrome of uncontrolled activation of the cellular immune system Incidence 1.2/1,000,000 M:F 1:1, Infants more commonly affected Classification: - primary - secondary
23 Classification 1. Primary (familial): Typically presents in infants, caused by a gene mutation involved in cytotoxic killing 2. Secondary (acquired): Absence of known genetic cause, can occur in children and adults, typically in the setting of an underlying infection, malignancy, immunodeficiency, autoimmune disorder
24 HLH: Pathophysiology Two types of cells involved in HLH: Macrophages: antigen presenting cells derived from circulating monocytes. Present foreign antigens to lymphocytes. In HLH macrophages become activated and secrete excessive amount of cytokines, ultimately causing severe tissue damage that can lead to organ failure. Natural Killer Cells (NK) and Cytotoxic Lymphocytes (CTL): NK cells eliminate damaged, stressed or infected host cells such as macrophages, typically in response to viral infection or malignancy. CTLs are activated lymphocytes that lyse cells such as macrophages bearing foreign antigen. In HLH NK and/or CTLs fail to eliminate activated macrophages, leading to excessive macrophage activity
25 Hemophagocytosis: macrophages engulf host cells excessive marker of macrophage activation.
26
27 Results in accumulation of activated T-lymphocytes and activated histiocytes (macrophages) with increasingly high levels of cytokines (cytokine storm) Key cytokines found at extremely high levels in the plasma of patients with HLH include interferon gamma, tumor necrosis factor (TNF)-alpha, interleukins IL-6, IL-8, IL-10, IL- 12, IL-18 and soluble IL-2 receptor (CD25)
28 Case Study Cytogenetics sent to Great Ormond Street Hospital: Soluble CD25 >20,000 pg/ml (normal range: <2,500 pg/ml) Normal cytotoxic granule release assay as detected by CD107a expression B-CLL Minimal Residual Disease detected Diagnosis: Secondary HLH
29 Diagnosis Diagnosis is challenging, symptoms non-specific, and many features overlap with other causes of severe illness including sepsis. In 2004 The Histocyte Society proposed an updated set of diagnostic criteria to aid in identification of patients with HLH.
30 The diagnosis of HLH may be established by: A. A molecular diagnosis consistent with HLH OR B. Fulfillment of five out of the eight criteria listed below: 1) Fever >38.5 ⁰C 2) Splenomegaly 3) Cytopenias (affecting at least 2 of 3 lineages in the peripheral blood): Hemoglobin <9 g/dl (in infants <4 weeks: hemoglobin <10 g/dl) Platelets <100 x10 9 /L Neutrophils <1 x10 9 /L 4) Hypertriglyceridemia (fasting, 265 mg/dl) and/or hypofibrinogenemia ( <1.50 g/l) 5) Hemophagocytosis in BM, spleen, lymph nodes or liver 6) Low or absent NK cell activity 7) Ferritin >500 ng/ml 8) Elevated scd25 (sil-2 receptor)
31 Treatment and Prognosis HLH-2004: International consensus treatment 8-week induction therapy with corticosteriods, etopside and cyclosproine. Stem cell Transplant Prognosis: without treatment 1-2 months
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38 Conclusion HLH rapidly progressive and fatal syndrome of uncontrolled immune activation Heightened awareness of HLH : significance of lab tests
39 Special Thanks to Professor Mary Cahill, Dr. Vitaly Mykytiv, Dr. Rachel Brodie Dr. Norma Reidy, Ms. Shelia Carmody, Ms Trish Hyland
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