Quantification of Schistocytes from the Peripheral Blood Smear in Thrombotic Thrombocytopenic Purpura (TTP) Compared to Non-TTP Thrombocytopenic Hospitalized Patients

Introduction

The diagnosis of TTP requires thrombocytopenia without an underlying cause and microangiopathic hemolytic anemia (MAHA)¹. TTP is usually confirmed with a low ADAMTS level. However, the turnaround time for an ADAMTS level is several days and treatment for this rare disease cannot be delayed, since it is uniformly fatal if left untreated but highly responsive to plasma exchange therapy. A rapid diagnosis is required and still must be made on clinical findings and peripheral smear morphology. The degree of MAHA that is seen with TTP, though, has not been well defined, or correlated with ADAMTS results. Automated fragmented red cell analyzers are not yet widely available or accepted, so the determination of MAHA still relies on manual smear reviews. Presenting smears from pts with TTP over a 1 ½ year period were reviewed, with manual counting of schistocytes, and were compared to smears from in-patient non-TTP cases of thrombocytopenia where hematology was consulted to rule out TTP.

Methods

29 cases were identified and peripheral smears from date of presentation were reviewed, initially by a hematologist and experienced special hematology technician, then by 2 medicine residents after they were given a 1 hour tutorial on smear interpretation. All smear reviewers were instructed on schistocyte identification as defined by the International Council for Standardization in Hematology (ICSH)². The number of schistocytes per 1000 red cells was counted at 1000 power magnification. The blood smears were coded/de-identified and then read "blindly" by the 4 observers. Degree of agreement between the 1^st 2 observers was measured (using Pearson correlation coefficient), as was the their average as compared to the average #schistocytes counted between the 2 med residents. Time taken to review smears was also measured.

Results

Ave age 54 (19-83); 59% female, 41% male; 62% Caucasian; 35% AA; 3% Hispanic. 9 had TTP and 20 did not. All cases of TTP had a low ADAMTS level (med <3%, range <3-13%) and all but one had a +inhibitor (one had congenital TTP, confirmed by ADAMTS mutation analysis). The non-TTP cases included sepsis (8/20), DIC (3/20), metastatic cancer (2/20), scleroderma crisis (2/20) and single cases of HIT, EtOH, gestational, MPN/MDS and unknown causes for thrombocytopenia. For TTP cases: med plt count was 22k (4-52k); LDH 760 (309-1175); hgb 9.4 (7.1-10.9); creat 1.59mg/dL (0.6-4.85). For non-TTP cases: med plt count was 44k (3-126k); LDH 300 (155-1111); hgb 8.1 (5.9-14.7); creat 1.36mg/dL (0.9-9.1). 3 non-TTP pts had ADAMTS levels sent (med= 69%, 57-100%).

The med #schistocytes in TTP cases was 6% (ave 6.3, 1.85-11.95%) compared to 0.625% (ave 1.52%, 0.25-7.5%) in the non-TTP group (p= 0.00002). The R-coefficient for the 1^st 2 observers was 0.93 (95% CI 0.85-0.96) and for their average compared to the 2 medicine residents it was 0.82. The average time to read smears overall was 10.5 minutes.

Conclusions:

This retrospective study showed that manual quantification of schistocytes is feasible, may be done in a timely fashion and with excellent interobserver reproducibility. All pts had some degree of MAHA but the TTP cases, not surprisingly, had a significantly higher percentage of schistocytes/1000 RBCs than non-TTP thrombocytopenic in-pt consult cases (6% vs 0.625%, respectively, p= 0.00002). All cases of low ADAMTS, positive inhibitor confirmed TTP had >2.5% schistocytes, compared to 4/20 non-TTP cases. A larger cohort of acquired TTP should be studied to determine if there is a cut-off % schistocytes which, in the appropriate clinical setting, should warrant urgent plasma exchange while waiting for ADAMTS levels to return.

¹George JN and Al-Noun ZL. Diagnostic and therapeutic challenges in the TTP and HUS syndromes. ASH Education Book. 2012, p. 604-609.

²Zini G et al. ICSH recommendations for identification, diagnostic value, and quantitation of schistocytes. International J of Lab Hematology. 2011; 34: 107-116.