Cathy Schechterly

An assessment of hepatitis E virus (HEV) in US blood donors and recipients: No detectable HEV RNA in 1939 donors tested and no evidence for HEV transmission to 362 prospectively followed recipients.

Hepatitis E virus (HEV) infection has become relevant to blood transfusion practice because isolated cases of blood transmission have been reported and because HEV has been found to cause chronic infection and severe liver disease in immunocompromised patients.

Parvovirus B19 infection transmitted by transfusion of red blood cells confirmed by molecular analysis of linked donor and recipient samples.

BACKGROUND: Extremely high viremic levels of parvovirus B19 (B19V) can be found in acutely infected, but asymptomatic donors. However, reports of transmission by single‐donor blood components are rare. In this prospective study, paired donor‐recipient samples were used to investigate the transfusion risk.

Investigation of residual hepatitis C virus in presumed recovered subjects

Recent studies have found hepatitis C virus (HCV) RNA in peripheral blood mononuclear cells (PBMCs) of the majority of presumed recovered subjects. We investigated this unexpected finding using samples from patients whose HCV RNA and anti‐HCV status had been serially confirmed. HCV RNA was detected in PBMCs from 66 of 67 chronic HCV carriers. Subpopulation analysis revealed that the viral load (log copies/106 cells) in B cells (4.14 ± 0.71) was higher than in total PBMCs (3.62 ± 0.71; P < 0.05), T cells (1.67 ± 0.88; P < 0.05), and non‐B/T cells (2.48 ± 1.15; P < 0.05). HCV negative‐strand RNA was not detected in PBMCs from any of 25 chronically infected patients. No residual viral RNA was detected in total PBMCs or plasma of 59 presumed recovered subjects (11 spontaneous and 48 treatment induced) using nested real‐time polymerase chain reaction with a detection limit of 2 copies/μg RNA (from ∼1 × 106 cells). PBMCs from 2 healthy HCV‐negative blood donors became HCV RNA positive, with B‐cell predominance, when mixed in vitro with HCV RNA–positive plasma, thus passively mimicking cells from chronic HCV carriers. No residual HCV was detected in liver or other tissues from 2 spontaneously recovered chimpanzees. Conclusion: (1) HCV RNA was detected in PBMCs of most chronic HCV carriers and was predominant in the B‐cell subpopulation; (2) HCV detected in PBMCs was in a nonreplicative form; (3) HCV passively adsorbed to PBMCs of healthy controls in vitro, becoming indistinguishable from PBMCs of chronic HCV carriers; and (4) residual HCV was not detected in plasma or PBMCs of any spontaneous or treatment‐recovered subjects or in chimpanzee liver, suggesting that the classic pattern of recovery from HCV infection is generally equivalent to viral eradication. (HEPATOLOGY 2013)

Quantitative analysis of anti–hepatitis C virus antibody–secreting B cells in patients with chronic hepatitis C

To investigate the quantitative characteristics of humoral immunity in patients with hepatitis C, we established an enzyme‐linked immunosorbent spot (ELISpot) assay for detection of anti–hepatitis C virus (HCV)‐secreting B cells. Receiver operating characteristic curve analysis demonstrated 100% specificity and 58% to 92% sensitivity for detecting B‐cell responses to NS5b, NS3, E2, and core antigens. The median sum of anti‐HCV–secreting B cells to all HCV antigens tested was significantly higher in 39 patients with chronic hepatitis C (47.3 spot forming cells [SFCs]/106 peripheral blood mononuclear cells [PBMCs]) than in 9 recovered subjects (15.3 SFCs/106 PBMCs; P = .05) or 11 uninfected controls (5.3 SFCs/106 PBMCs; P < .001); the significant difference (P = .018) in chronic versus recovered patients was in reactivity to nonstructural antigens NS3 and NS5b. Anti‐HCV immunoglubulin M (IgM)–secreting B cells were also readily detected and persisted decades into HCV infection; there was no difference in IgM‐positive cells between chronic and recovered patients. ELISpot reactivity to genotype 1–derived antigens was equivalent in patients of genotypes 1, 2, and 3. There was significant correlation between the numbers of anti‐HCV IgG‐secreting B cells and serum aminotransferase and to the level of circulating antibody. In conclusion, ELISpot assays can be adapted to study B‐cell as well as T‐cell responses to HCV. Measurement at the single‐cell level suggests that humoral immunity plays a minor role in recovery from HCV infection and that B‐cell immunity is strongest in those with persistent infection. (HEPATOLOGY 2005.)

Circulating let‐7 levels in plasma and extracellular vesicles correlate with hepatic fibrosis progression in chronic hepatitis C

The goal of this study was to determine whether an association exists between circulating microRNA (miRNA) levels and disease progression in chronic hepatitis C (CHC), whether plasma or extracellular vesicles (EVs) were optimal for miRNA measurement and their correlation with hepatic miRNA expression, and the mechanistic plausibility of this association. We studied 130 CHC patients prospectively followed over several decades. A comprehensive miRNA profile in plasma using microarray with 2578 probe sets showed 323 miRNAs differentially expressed between healthy individuals and CHC patients, but only six that distinguished patients with mild versus severe chronic hepatitis. Eventually, let‐7a/7c/7d‐5p and miR‐122‐5p were identified as candidate predictors of disease progression. Cross‐sectional analyses at the time of initial liver biopsy showed that reduced levels of let‐7a/7c/7d‐5p (let‐7s) in plasma were correlated with advanced histological hepatic fibrosis stage and other fibrotic markers, whereas miR‐122‐5p levels in plasma were positively correlated with inflammatory activity, but not fibrosis. Measuring let‐7s levels in EVs was not superior to intact plasma for discriminating significant hepatic fibrosis. Longitudinal analyses in 60 patients with paired liver biopsies showed that let‐7s levels in plasma markedly declined over time in parallel with fibrosis progression. However, circulating let‐7s levels did not parallel those in the liver. Conclusion: Of all miRNAs screened, the let‐7 family showed the best correlation with hepatic fibrosis in CHC. A single determination of let‐7s levels in plasma did not have superior predictive value for significant hepatic fibrosis compared with that of fibrosis‐4 index, but the rate of let‐7s decline in paired longitudinal samples correlated well with fibrosis progression. Pathway analysis suggested that low levels of let‐7 may influence hepatic fibrogenesis through activation of transforming growth factor β signaling in hepatic stellate cells. (Hepatology 2016;64:732‐745)

Absence of transfusion-associated microchimerism in pediatric and adult recipients of leukoreduced and gamma-irradiated blood components.

BACKGROUND: Transfusion‐associated microchimerism (TA‐MC), the persistence of significant levels of donor white blood cells (WBCs) in blood recipients for prolonged periods, has been demonstrated after nonleukoreduced and leukoreduced transfusion to patients with severe traumatic injury. Development of TA‐MC has not been rigorously studied in settings that do not involve massive trauma where the blood is leukoreduced and irradiated.

Leukoreduction and ultraviolet treatment reduce both the magnitude and the duration of the HLA antibody response

Both leukoreduction and ultraviolet (UV) light treatment of blood products have been shown to reduce the incidence of HLA antibody development in recipients, but the impact of these treatments on the magnitude and persistence of the antibody response is less clear.

Preferential association of hepatitis C virus with CD19+ B cells is mediated by complement system

Extrahepatic disease manifestations are common in chronic hepatitis C virus (HCV) infection. The mechanism of HCV‐related lymphoproliferative disorders is not fully understood. Recent studies have found that HCV in peripheral blood mononuclear cells from chronically infected patients is mainly associated with cluster of differentiation 19‐positive (CD19+) B cells. To further elucidate this preferential association of HCV with B cells, we used in vitro cultured virus and uninfected peripheral blood mononuclear cells from healthy blood donors to investigate the necessary serum components that activate the binding of HCV to B cells. First, we found that the active serum components were present not only in HCV carriers but also in HCV recovered patients and HCV‐negative, healthy blood donors and that the serum components were heat‐labile. Second, the preferential binding activity of HCV to B cells could be blocked by anti‐complement C3 antibodies. In experiments with complement‐depleted serum and purified complement proteins, we demonstrated that complement proteins C1, C2, and C3 were required to activate such binding activity. Complement protein C4 was partially involved in this process. Third, using antibodies against cell surface markers, we showed that the binding complex mainly involved CD21 (complement receptor 2), CD19, CD20, and CD81; CD35 (complement receptor 1) was involved but had lower binding activity. Fourth, both anti‐CD21 and anti‐CD35 antibodies could block the binding of patient‐derived HCV to B cells. Fifth, complement also mediated HCV binding to Raji cells, a cultured B‐cell line derived from Burkitts lymphoma. Conclusion: In chronic HCV infection, the preferential association of HCV with B cells is mediated by the complement system, mainly through complement receptor 2 (CD21), in conjunction with the CD19 and CD81 complex. (Hepatology 2016;64:1900‐1910).

An Epidemiologic Investigation of a Case of Acute Hepatitis E

ABSTRACT Hepatitis E virus (HEV) is considered a zoonotic infection in developed nations. A case of acute hepatitis E in a researcher following a scalpel injury while working on a pig prompted a seroepidemiologic study to identify potential modes of transmission and determine the seroprevalence of HEV among animal handlers at the institute. Sera from personnel (n = 64) in two animal facilities and age/sex-matched blood donors (n = 63) as controls were tested for IgG anti-HEV and, if positive, for IgM anti-HEV and HEV RNA. Sera and stool from pigs aged 6 to 12 weeks from the breeding farm and older pigs from animal facilities were tested similarly. The median age of personnel was 36 years, 74% were white, 56% were male, and 74% had direct exposure to pigs. The prevalence of anti-HEV was 3.1% among personnel compared to 3.2% among blood donors; none were positive for IgM anti-HEV or HEV RNA. IgG anti-HEV was detected in sera from 10% of pigs aged 6 to 8 weeks, 80% aged 10 weeks, 100% aged 12 weeks, and 76% aged >12 weeks. HEV RNA was detected in stool but not sera from three 12-week-old pigs. Sequencing revealed HEV genotype 3 with ∼10% difference between the patient and pig sequences. Parenteral transmission is a potential mode of acute HEV infection. The low and similar seroprevalence of anti-HEV between the at-risk group and age-matched blood donors suggests low transmission risk with universal precautions among animal handlers.

Inherent transcriptional signatures of NK cells are associated with response to IFNα + rivabirin therapy in patients with Hepatitis C Virus

BackgroundDifferences in the expression of Natural Killer cell receptors have been reported to reflect divergent clinical courses in patients with chronic infections or tumors. However, extensive molecular characterization at the transcriptional level to support this view is lacking. The aim of this work was to characterize baseline differences in purified NK cell transcriptional activity stratified by response to treatment with PEG-IFNα/RBV in patients chronically infected with HCV.MethodsTo this end we here studied by flow cytometer and gene expression profile, phenotypic and transcriptional characteristics of purified NK cells in patients chronically infected with HCV genotype-1 virus who were subsequently treated with PEG-IFNα/RBV. Results were further correlated with divergent clinical response obtained after treatment.ResultsThe pre-treatment transcriptional patterns of purified NK cells from patients subsequently undergoing a sustained virologic response (SVR) clearly segregated from those of non-responder (NR) patients. A set of 476 transcripts, including molecules involved in RNA processing, ubiquitination pathways as well as HLA class II signalling were differently expressed among divergent patients. In addition, treatment outcome was associated with differences in surface expression of NKp30 and NKG2D. A complex relationship was observed that suggested for extensive post-transcriptional editing. Only a small number of the NK cell transcripts identified were correlated with chronic HCV infection/replication indicating that inherent transcriptional activity prevails over environment effects such as viral infection.ConclusionsCollectively, inherent/genetic modulation of NK cell transcription is involved in setting the path to divergent treatment outcomes and could become useful to therapeutic advantage.