Comparison of plasma and blood cell samples in metagenomic next-generation sequencing for identification of the causative pathogens of fever.

Background: Metagenomic next-generation sequencing (mNGS) of plasma DNA has become an attractive diagnostic method for infectious diseases; however, the rate of false-positive results is high. This study aims to evaluate the diagnostic accuracy of mNGS in plasma versus blood cell samples for immunocompromised children with febrile diseases. Methods: The results of conventional microbiological test (CMT) and mNGS using plasma and blood cells in 106 patients with 128 episodes of febrile diseases from the Department of Hematology/Oncology were analyzed and described. Results: The positivity rates for CMT and mNGS of plasma and blood cells were 35.9 %, 84.4 % and 46.9 %, respectively (P < 0.001). Notably, mNGS identified multiple pathogens in a single specimen in 68.5 % of plasma samples and 38.3 % of blood cell samples (P < 0.001). Furthermore, plasma and blood cell mNGS identified causative pathogens in 58 and 46 cases, accounting for 53.7 % and 76.7 % of the mNGS-positive cases for each sample type, respectively (P = 0.002). By integrating results from both plasma and blood cell samples, causative pathogens were identified in 77 cases (60.2 %), enhancing sensitivity to 87.5 % but reducing specificity to 15.0 %, compared to plasma (65.9 % sensitivity and 20.0 % specificity) and blood cell samples (52.3 % sensitivity and 80.0 % specificity). Conclusions: mNGS of plasma is sensitive but has a high false-positive rate, while mNGS of blood cells has low sensitivity but higher specificity.

Interaction of E2F3a and CASP8AP2 Regulates Histone Expression and Chemosensitivity of Leukemic Cells.

Low expression levels of E2F3a and caspase 8-associated protein 2 (CASP8AP2) are associated with poor outcomes in children with acute lymphoblastic leukemia. Our previous study showed that a combined assessment of E2F3a and CASP8AP2 expression was more accurate in predicting relapse in children with acute lymphoblastic leukemia. However, the underlying mechanism remains unclear. In this study, the interaction between E2F3a and CASP8AP2 and its role in the regulation of histone expression, cell proliferation, the cell cycle, and chemosensitivity were investigated. Exogenous E2F3a-GST was coprecipitated with CASP8AP2-FLAG in HEK-293T cells. E2F3a was colocalized with CASP8AP2-GFP in the nucleus. The replication-dependent histones H2A and H2B were significantly upregulated when E2F3a or CASP8AP2 was overexpressed in HEK-293T or 697 cells and downregulated by E2F3a or CASP8AP2 knockdown. E2F3a and CASP8AP2 could collaboratively enhance the transcriptional activity of HIST1H2AG and HIST1H2BK . Both CASP8AP2 and E2F3a are involved in S phase progression. E2F3a and CASP8AP2 also affected the sensitivity of leukemic cells to daunorubicin. Therefore, CASP8AP2 and E2F3a collaboratively regulated replication-dependent histone expression, cell cycle progression, and chemosensitivity of leukemic cells.

Metagenomic next-generation sequencing for the diagnosis of fever of unknown origin in pediatric patients with hematological malignancy.

BACKGROUND AND AIMS: The aim of this study was to evaluate the performance of metagenomic next-generation sequencing (mNGS) in identifying microbiological etiologies in pediatric patients with hematological malignancies undergoing fever of unknown origin (FUO). METHODS: A total of 147 children with hematological malignancy suffering febrile diseases without definite microbiological etiologies under conventional tests were enrolled. The clinical record, serum inflammatory biomarkers and mNGS results were analyzed. RESULTS: At least one microorganism was identified by mNGS in 112 of 147 patients (76.2 %). Two or more types of organisms were detected simultaneously in 35.7 % (40/112) of samples. Of the 112 cases with positive mNGS results, the reported microorganisms were considered as etiologies of fever in 50 (44.6 %) cases. The initial antimicrobial regimens were adjusted according to the mNGS results in 48 cases, with 41 patients' febrile diseases resolved. Totally, 27.9 % (41/147) of patients benefit from mNGS. High IL-6 (>390 pg/mL) level was associated with bacterial infection and could help to interpret the results of mNGS. CONCLUSION: mNGS is a novel approach to determine the microbiological etiology of FUO in hematological malignancy patients, which benefits about a quarter of all patients tested. Integration of IL-6 can improve the diagnostic precision of bacterial infection.

Ceruloplasmin inhibits the proliferation, migration and invasion of nasopharyngeal carcinoma cells and is negatively regulated by miR-543.

BACKGROUND: Ceruloplasmin (CP), recognized as a member of multicopper oxidase family, is related to the progression of diverse cancers in human beings. This study is designed to clarify the expression characteristics, biological function and related mechanism of CP in nasopharyngeal carcinoma (NPC). METHODS: CP expression in NPC tissues and cells was probed by quantitative real-time polymerase chain reaction (qRT-PCR), immunohistochemistry (IHC) and Western blot. After gain-of-function and loss-of-function models were established, cell counting kit-8 (CCK-8), Transwell and BrdU assays were employed to measure cell viability, migration and invasion. The targeting relationship between microRNA-543 (miR-543) and CP was verified by dual-luciferase reporter gene assay. RESULTS: As against normal nasopharyngeal epithelial tissues, CP expression was significantly lower in NPC tissues, which was associated with higher clinical stage and the short overall survival time. Compared with the control group, CP overexpression markedly restrained the growth, migration and invasion of NPC cells; knocking down CP had the opposite effect. MiR-543 directly targeted CP and negatively modulated its expression. CONCLUSION: CP restrains the growth, migration and invasion of NPC cells and is negatively regulated by miR-543.