Discovery and Correction of Spurious Low Platelet Counts due to EDTA-Dependent Pseudothrombocytopenia.

BACKGROUND: Ethylene diamine tetraacetic acid dependent pseudothrombocytopenia (EDTA-PTCP) is a laboratory artifact that may lead to unnecessary evaluation and treatment of patients. The purpose of this article is to discuss how to identify EDTA-PTCP and correct spurious low platelet counts in clinical laboratories. METHODS: We use two criteria to screen for platelet aggregation: (1) an abnormal platelet count in EDTA-treated blood from a patient lacking clinical signs of a platelet disorder, and (2) an instrument flag for platelet clumps. EDTA-PTCP was confirmed by microscopic examination for platelet agglutination and by platelet counts that corrected with citrate sample. In addition, the time course of EDTA-PTCP was investigated in samples from 26 patients anticoagulated with EDTA-K2 and sodium citrate. Amikacin (5 mg/ml) was added to tubes with EDTA-K2 or sodium citrate from seven additional cases in order to confirm its dissociative effect on platelet aggregation. RESULTS: In our laboratory, the overall incidence of EDTA-PTCP was approximately 0.09%; and the duration was between 2 weeks and 6 months. EDTA-PTCP was time-dependent and occurred as early as 10 min after sample collection. Weaker agglutination could also occur in most corresponding citrate-treated samples. The dissociative effect of amikacin on platelet agglutination was case-specific and not concentration-dependent. CONCLUSIONS: The method of screening for platelet clumping with the help of XE5000 images is convenient. The decline in the platelet count is related to the length of time and the intensity of chelation. Amikacin supplement is not always effective for correcting platelet counts in vitro.

[Value of percentage of highly fluorescent lymphocytic cells for rapidly assessing septicemia in tumor patients].

OBJECTIVE: To evaluate the value of percentage of highly fluorescent lymphocytic cells (HFLC%) for rapidly assessing septicemia in tumor patients. METHODS: Blood samples were collected from 130 patients with tumors (60 septicemia patients and 70 non-septicemia patients) and 80 healthy controls. HFLC% was analyzed with Sysmex XE-5000, the level of C-reactive protein (CRP) measured with a commercially available turbidimetric immunoassay kit and the level of procalcitonin (PCT) determined with a semiquantitative chromatographic immunoassay kit. The diagnostic values of HFLC% and CRP in septicemia were evaluated with ROC analysis. RESULTS: The values of HFLC% and CRP were significantly higher in the septicemia group than those in the non-septicemia and healthy groups (0.30% (0.10%-0.70%) vs 0.10% (0-0.20%), 0.10% (0-0.20%) ; 80.3 (28.5-129.5) vs 3.3 (1.4-41.4) , 1.4 (0.6-2.5) mg/L, all P < 0.01) . The ROC-AUCs for HFLC% and CRP for a diagnosis of septicemia were 0.72 (sensitivity 71.7%, specificity 58.7%) and 0.92 (sensitivity 96.7%, specificity 82.0%). Both of them could judge septicemia better. Additionally, HFLC% was correlated with the levels of PCT and CRP (r = 0.637, 0.241, both P < 0.01). CONCLUSIONS: HFLC% may be used as a rapid and simple auxiliary indicator in the diagnosis of septicemia in patients with tumors. And it is conducive to make an early diagnosis of septicemia and avoid unnecessary use of antibiotics.

[Comparison of testing methods for reticulated platelets].

OBJECTIVE: To compare the accuracy, stability and sample cross-contamination of two independent methods of detecting the percentage of reticulated platelets in peripheral blood and establish a local normal reference range so as to provide methodological rationales in clinical laboratory. METHODS: The percentages of reticulated platelets in peripheral blood of a healthy population were measured by Sysmex XE-5000 blood cell analyzer with polymethyl oxazine staining and flow cytometer with thiazole orange staining respectively. The correlation between the results of two methods was analyzed by Spearman's nonparametric correlation. Information about stability was obtained from measurements of the percentages of reticulated platelets in peripheral blood at designated time points. The analyses of accuracy, sample cross contamination and local normal reference range were performed routinely. RESULTS: The coefficient of variation (CV) of data was lower (16.2%) than that from flow cytometer (35.1%). The sample cross-contaminations of two methods were the same at around 5%. The percentage of reticulated platelets in peripheral blood was stable and consistent whereas the results of flow cytometer fluctuated at different time points within 4 h after blood sampling. The correlation of results obtained from two methods was significant (P < 0.01, r(2) = 0.923). The local normal reference range was 1.0% - 7.5% for Sysmex XE-5000 versus 3.0% - 10.5% for flow cytometer. CONCLUSIONS: Fully automatic blood cell analyze is more advanced than flow cytometer for its simple operation and stable data. And the former is an ideal first-choice for detecting the percentage of reticulated platelets in peripheral blood.

Involvement of indoleamine 2,3-dioxygenase in impairing tumor-infiltrating CD8 T-cell functions in esophageal squamous cell carcinoma.

The indoleamine 2,3-dioxygenase-(IDO-) mediated microenvironment plays an important role in tumor immune escape. However, the inhibitory effects of IDO on the CD8(+) tumour-infiltrating lymphocytes (CD8(+) TILs) in esophageal squamous cell carcinoma (ESCC) have not been clarified yet. Here, we found that the level of IDO expression in ESCC tumor specimens correlated with a reduction in the number of CD8(+) TILs. Patients with high IDO expression and a low number of CD8(+) TILs had significantly impaired overall survival time. IDO expression and functional enzyme activity in ESCC cell lines could be induced by IFNγ. When exposed to the milieu generated by IDO-expressing Eca109 cells, the CD8(+) TILs were suppressed in proliferation, and their cytolytic functions against target tumor cells were lost. These results suggested that impairing CD8(+) TIL functions by IDO expressed in ESCC possibly contributed to the finding that patients with higher IDO expression have more aggressive disease progression and shorter overall survival time.

[Correlation factors of ethylenediaminetetraacetic acid-dependent pseudothrombocytopenia in cancer patients].

OBJECTIVE: To investigate the correlation factors of ethylenediaminetetraacetic acid-dependent pseudothrombocytopenia (EDTA-PTCP) in cancer patients. METHODS: The potential correlation factors of EDTA-PTCP such as gender, age, case history, tumor types, therapeutic drugs and duration of EDTA-PTCP from cancer patients were analyzed based on the patient records from October 2007 to September 2009 at our cancer center. RESULTS: A total of 49 EDTA-PTCP cases from a pool of 55 000 cancer patients were collected. No correlation was found with gender (male 49.0%, female 51.0%), concurrent hypertension (20.4%)/diabetes (10.2%) or cancer types (1 - 11 cases each type). EDTA-PTCP appeared at pre-therapy (n = 13) and post-therapy (n = 36). Eleven cases (30.6%) were chemotherapy, 5 cases (13.9%) were radiotherapy plus chemotherapy, 15 case (41.7%) were tumor resection, 5 cases (13.9%) were interventional therapy in 36 patients whose EDTA-PTCP appeared post-therapy. The most frequency use in chemotherapy patients was dexamethasone (87.5%, 14/16), and in surgery patients was penicillin antibiotics (75.0%, 15/20). And its frequency was once (n = 18) and more than twice (n = 31). If the subjects were divided into 2 groups of non-treatment plus surgery and chemotherapy plus intervention on the basis of treatment course, there was a significant difference between two groups in proportion of patients whose duration of EDTA-PTCP ≤ 2 weeks (89.3% vs 47.6%, χ(2) = 10.22, P < 0.01). CONCLUSIONS: The incidence of EDTA-PTCP in cancer patients may be associated with therapeutic drugs, but not probably with gender, concurrent hypertension/diabetes, tumor types or therapeutic regimens. Duration of EDTA-PTCP may be associated with the treatment course.

Prediction of Poor Outcomes in Patients with Colorectal Cancer: Elevated Preoperative Prothrombin Time (PT) and Activated Partial Thromboplastin Time (APTT).

BACKGROUND AND OBJECTIVE: Tools for the non-invasive assessment of colorectal cancer (CRC) prognosis have profound significance. Although plasma coagulation tests have been investigated in a variety of tumours, the prognostic value of the prothrombin time (PT) and activated partial thromboplastin time (APTT) in CRC has not been discussed. Our study objective was to explore the prognostic significance of preoperative PT and APTT in CRC patients. PATIENTS AND METHODS: A retrospective analysis of preoperative coagulation indexes including PT, PTA, INR, APTT, FIB, TT, PLT, NLR and PLR in 250 patients with CRC was performed. Kaplan-Meier and multivariate Cox regression analysis were used to demonstrate the prognostic value of these preoperative coagulation indexes. RESULTS: The overall survival (OS, p<0.05) and disease-free survival (DFS, p<0.05) of CRC patients with lower PT and APTT levels were significantly prolonged. Based on univariate analysis, PT levels (p<0.001, p<0.001), PTA levels (p=0.001, p=0.001), APTT levels (p=0.001, p<0.001), INR levels (p<0.001, p<0.001), fibrinogen levels (p=0.032, p=0.036), tumour status (p=0.005, p=0.003), nodal status (p<0.001, p<0.001), metastasis status (p<0.001, p<0.001) and TNM stages (p<0.001, p<0.001) were remarkably associated with DFS and OS. Multivariate Cox regression analysis suggested that the levels of PT (HR: 2.699, p=0.006) and APTT (HR: 1.942, p=0.015), metastasis status (HR: 2.091, p= 0.015) and TNM stage (HR: 7.086, p=0.006) were independent predictors of survival in CRC. In the whole cohort, the enrolled patients were then divided into three groups according to their PT and APTT levels. The OS and DFS differed notably among the low-risk (PT<11.85 sec and APTT<25.85 sec), medium-risk (PT≥11.85 sec or APTT≥25.85 sec), and high-risk (PT≥11.85 sec and APTT≥25.85 sec) groups. CONCLUSION: Elevated levels of preoperative PT and APTT were predictors of poor outcomes in CRC patients. Moreover, the combination of preoperative PT and APTT can be a new prognostic stratification approach for more precise clinical staging of CRC.

Multivariate Analysis of Prognostic Biomarkers in Surgically Treated Endometrial Cancer.

OBJECTIVE: The aim of this study was to identify biomarkers with prognostic value in the setting of surgically treated endometrial cancer. METHODS: Medical data for 282 patients with surgically treated endometrial cancer were reviewed retrospectively. Preoperative concentrations of six serum biomarkers (CA125, CA15-3, C-reactive protein [CRP], D-dimer [D-D], platelet-to-lymphocyte ratio [PLR], and neutrophil-to-lymphocyte ratio [NLR]) were analysed to determine potential associations with clinicopathologic characteristics and to assess prognostic values separately via Kaplan-Meier method and multivariate Cox regression. RESULTS: In univariate analyses, the 5-year overall survival (OS) rate was 86.5% for a maximum follow-up period of 75 months. High concentrations of CA125, CA15-3, CRP, D-D, PLR, and NLR each proved significantly predictive of poor survival (log-rank test, P<0.01). CRP and D-D were identified as independent prognosticators, using a Cox regression model. Study patients were then stratified (based on combined independent risk factors) into three tiers (P<0.001), marked by 5-year OS rates of 92.1%, 78.4%, and 33.3%. CONCLUSIONS: All serum biomarkers assessed (CA125, CA15-3, CRP, D-D, PLR, and NLR) proved to be valid prognostic indices of surgically treated endometrial cancer. A novel prognostic grouping system, incorporating independent risk factors (CRP and D-D Concentrations), may have merit in assessing these patients preoperatively, providing a biologic basis for improved clinical staging.

Utility of neut-X, neut-Y and neut-Z parameters for rapidly assessing sepsis in tumor patients.

BACKGROUND: The Sysmex XE-5000 hematology analyzer evaluates toxic granulation and the nuclear maturity of toxic granulation neutrophils via the parameters neut-X and neut-Y. This study investigated whether neut-X and neut-Y could facilitate the auxiliary diagnosis of sepsis. METHODS: Blood samples were collected from 113 patients with tumors and 130 healthy individuals to detect neut-X, neut-Y, and C-reactive protein (CRP) values. Then, we created a new parameter, neut-Z, the vector sum of neut-X and neut-Y. Furthermore, we assessed procalcitonin (PCT) concentrations in patients with sepsis and compared the values with those for neut-X, neut-Y, and neut-Z. RESULTS: Neut-X, neut-Y, neut-Z, and CRP values were significantly higher in the sepsis group than in the non-sepsis and healthy groups. The ROC-AUCs for neut-X, neut-Y, neut-Z, and CRP for a diagnosis of sepsis were 0.87 (sensitivity 82%, specificity 79%), 0.87 (78 and 94%, respectively), 0.91 (82 and 88%, respectively), and 0.95 (96 and 89%, respectively), respectively. Additionally, neut-X, neut-Y, and neut-Z were correlated with CRP and PCT levels. CONCLUSIONS: Neut-X and neut-Y can be used as rapid and simple auxiliary indicators in the diagnosis of sepsis in patients with tumors, and neut-Z appears to display better performance than its 2 components.