ABSTRACT
Hypermutations in hepatitis B virus (HBV) DNA by APOBEC3 cytidine deaminases have been detected in vitro and in vivo, and APOBEC3G (A3G) and APOBEC3F (A3F) have been shown to inhibit the replication of HBV in vitro, but the presumably low or even absent hepatic expression of these enzymes has raised the question as to their physiological impact on HBV replication. We show that normal human liver expresses the mRNAs of APOBEC3B (A3B), APOBEC3C (A3C), A3F, and A3G. In primary human hepatocytes, interferon alpha (IFN-alpha) stimulated the expression of these cytidine deaminases up to 14-fold, and the mRNAs of A3G, A3F, and A3B reached expression levels of 10%, 3%, and 3%, respectively, relative to GAPDH mRNA abundance. On transfection, the full-length protein A3B(L) inhibited HBV replication in vitro as efficiently as A3G or A3F, whereas the truncated splice variant A3B(S) and A3C had no effect. A3B(L) and A3B(S) were detected predominantly in the nucleus of uninfected cells; however, in HBV-expressing cells both proteins were found also in the cytoplasm and were associated with HBV viral particles, similarly to A3G and A3F. Moreover, A3G, A3F, and A3B(L), but not A3B(S), induced extensive G-to-A hypermutations in a fraction of the replicated HBV genomes. In conclusion, the editing enzymes A3B(L), A3F, and most markedly A3G, which are expressed in liver and up-regulated by IFN-alpha in hepatocytes, are candidates to contribute to the noncytolytic clearance of HBV.
Subject(s)
Cytidine Deaminase/biosynthesis , DNA Replication/genetics , Gene Expression Regulation, Viral , Hepatitis B virus/genetics , Base Sequence , Carcinoma, Hepatocellular/pathology , Cell Line, Tumor , Cells, Cultured , Cytidine Deaminase/genetics , Fluorescent Antibody Technique , Hepatocytes , Humans , Interferons/pharmacology , Minor Histocompatibility Antigens , Molecular Sequence Data , Mutagenesis , RNA, Messenger/analysis , Reference Values , Reverse Transcriptase Polymerase Chain Reaction , Sensitivity and SpecificityABSTRACT
While pathophysiology of elevated cytokines is well delineated, reference values for children are unknown, although they may vary physiologically with age and differ from those of adults. Between June and November 2001, interleukin (IL)-6, IL-10 and tumor necrosis factor-alpha (TNF-alpha) concentrations from blood samples of 79 healthy children in six different age groups (group I: 0-3 months; group II: 4-12 months; group III: 13-24 months; group IV: 25-36 months; group V: 37-48 months; group VI: 49-60 months) were measured with ELISA. TNF-alpha was within 2.2-3.5 pg/ml in all groups with a trend toward higher values in groups II and III (p = ns). IL-6 was significantly lower in group III than in groups IV (p = 0.0165) and VI (p = 0.0147). IL-10 was within 3.3-5.5 pg/ml in all groups (p = ns). In regression analysis no correlation between age and cytokine concentrations was found. Although not statistically significant, IL-6 was lower and TNF-alpha higher than the adult reference values provided by the kit manufacturer. Although reference cytokine levels seem not age-related during early infancy, IL-6 is significantly lower during the second year of life than later. In infants aged 5 years or younger, reference levels of IL-6 should be chosen lower, and those of TNF-alpha higher, than the adult reference values.
Subject(s)
Interleukin-10/blood , Interleukin-6/blood , Tumor Necrosis Factor-alpha/metabolism , Adult , Child, Preschool , Humans , Infant , Reference ValuesABSTRACT
BACKGROUND: In contrast to RBC transfusion, where ABO mismatch is potentially lethal, immunologic ABO matching has been considered less critical for PLTs. Nonetheless, PLTs bear ABO blood group antigens, some of them expressing very high levels. STUDY DESIGN AND METHODS: The expression of A antigen was investigated by flow cytometry on resting and stimulated human PLTs of 100 A and 10 O group donors, as well as on 17 PLT concentrates (PCs) after apheresis and daily during a 6-day storage, to determine possible changes in expression of A antigen on PLT surface. RESULTS: Considerable variation of A antigen expression on PLT surface of A1 group individuals was observed; A2 group PLTs could not be distinguished from O group PLTs. The variability of A antigen on A group PLTs also became evident on investigating PLT lysates by ELISA. A1 group PCs showed a significant increase of A antigen expression on their surface owing to apheresis (p = 0.001) and to storage (p = 0.0091). CONCLUSION: Apheresis and prolonged storage of A1 group PCs independently led to overexpression of A antigen on the PLT surface. This may make such PCs more susceptible to destruction by anti-A of O or B group recipients.
