Short-term changes observed in multiparametric liver MRI following therapy with direct-acting antivirals in chronic hepatitis C virus patients.

METHODS: We applied multiparametric MRI to assess changes in liver composition, perfusion and blood flow in 17 patients before direct-acting antiviral (DAA) therapy and after treatment completion (within 12 weeks of last DAA tablet swallowed). RESULTS: We observed changes in hepatic composition indicated by a reduction in both liver longitudinal relaxation time (T1, 35 ± 4 ms), transverse relaxation time (T2, 2.5 ± 0.8 ms; T2* 3.0 ± 0.7 ms), and liver perfusion (28.1 ± 19.7 ml/100 g/min) which we suggest are linked to reduced pro-inflammatory milieu, including interstitial oedema, within the liver. No changes were observed in liver or spleen blood flow, splenic perfusion, or superior mesenteric artery blood flow. CONCLUSION: For the first time, our study has shown that treatment of HCV with DAAs in patients with cirrhosis leads to an acute reduction in liver T1, T2 and T2* and an increase in liver perfusion measured using MR parameters. The ability of MRI to characterise changes in the angio-architecture of patients with cirrhosis after intervention in the short term will enhance our understanding of the natural history of regression of liver disease and potentially influence clinical decision algorithms. KEY POINTS: • DAAs have revolutionised the treatment of hepatitis C and achieve sustained virological response in over 95% of patients, even with liver cirrhosis. • Currently available non-invasive measures of liver fibrosis are not accurate after HCV treatment with DAAs, this prospective single-centre study has shown that MRI can sensitively measure changes within the liver, which could reflect the reduction in inflammation with viral clearance. • The ability of MRI to characterise changes in structural and haemodynamic MRI measures in the liver after intervention will enhance our understanding of the progression/regression of liver disease and could potentially influence clinical decision algorithms.

Mannan binding lectin-associated serine protease 1 is induced by hepatitis C virus infection and activates human hepatic stellate cells.

Mannan binding lectin (MBL)-associated serine protease type 1 (MASP-1) has a central role in the lectin pathway of complement activation and is required for the formation of C3 convertase. The activity of MASP-1 in the peripheral blood has been identified previously as a highly significant predictor of the severity of liver fibrosis in hepatitis C virus (HCV) infection, but not in liver disease of other aetiologies. In this study we tested the hypotheses that expression of MASP-1 may promote disease progression in HCV disease by direct activation of hepatic stellate cells (HSCs) and may additionally be up-regulated by HCV. In order to do so, we utilized a model for the maintenance of primary human HSC in the quiescent state by culture on basement membrane substrate prior to stimulation. In comparison to controls, recombinant MASP-1 stimulated quiescent human HSCs to differentiate to the activated state as assessed by both morphology and up-regulation of HSC activation markers α-smooth muscle actin and tissue inhibitor of metalloproteinase 1. Further, the expression of MASP-1 was up-regulated significantly by HCV infection in hepatocyte cell lines. These observations suggest a new role for MASP-1 and provide a possible mechanistic link between high levels of MASP-1 and the severity of disease in HCV infection. Taken together with previous clinical observations, our new findings suggest that the balance of MASP-1 activity may be proinflammatory and act to accelerate fibrosis progression in HCV liver disease.

Platelet function testing in patients with post-operative tonsillectomy bleeding may be a useful early identifier of inherited platelet function disorders.

BACKGROUND: Post-tonsillectomy bleeding is the most frequent complication of tonsillectomy. Inherited platelet function disorders have an estimated prevalence of 1 per cent. Any association between post-tonsillectomy bleeds and undiagnosed inherited platelet function disorders has not been investigated before. OBJECTIVES: To assess the prevalence of inherited platelet function disorders in a cohort of post-tonsillectomy bleed patients. METHODS: An observational cohort study was conducted using hospital digital records. Platelet function analyser 100 ('PFA-100') closure time was tested on post-tonsillectomy bleed patients who presented to hospital. RESULTS: Between 2013 and 2017, 9 of 91 post-tonsillectomy bleed patients who underwent platelet function analyser 100 testing (9.89 per cent) had positive results. Five patients (5.49 per cent) had undiagnosed inherited platelet function disorders. Four patients had false positive results secondary to a non-steroidal anti-inflammatory drug effect (specificity of 95.3 per cent) proven by repeat testing six weeks later, off medication. The false negative rate was 0 per cent. CONCLUSION: The prevalence of inherited platelet function disorders in our post-tonsillectomy bleed cohort is five-fold higher than in the general population. Platelet function analyser 100 testing when patients present with a post-tonsillectomy bleed allows management of their inherited platelet function disorder.

Possible ancient oceans on Mars: evidence from Mars Orbiter Laser Altimeter data.

High-resolution altimetric data define the detailed topography of the northern lowlands of Mars, and a range of data is consistent with the hypothesis that a lowland-encircling geologic contact represents the ancient shoreline of a large standing body of water present in middle Mars history. The contact altitude is close to an equipotential line, the topography is smoother at all scales below the contact than above it, the volume enclosed by this contact is within the range of estimates of available water on Mars, and a series of extensive terraces parallel the contact in many places.

Response rates to combination therapy for chronic HCV infection in a clinical setting and derivation of probability tables for individual patient management.

Evidence for efficacy of established treatment guidelines for chronic hepatitis C virus (HCV) disease is based on multinational randomized controlled trials (RCTs). Strategies for managing HCV, however, require an assessment of the effectiveness of intervention in routine clinical practice. We report the outcomes of combination therapy in a large cohort of HCV-infected individuals in the UK. A total of 347 (113 genotype 1, 234 genotype non-1) patients were treated with pegylated interferon and ribavirin according to current guidelines. Forty-two (37.2%) of those with genotype 1 infection and 164 (70.1%) with genotype non-1 infection achieved sustained viral response (SVR). Thirty-nine (11%) patients withdrew from treatment. In addition to viral genotype, factors predictive of a response to therapy were age at start of treatment and disease stage on pretreatment liver biopsy. Multivariate regression analysis demonstrated that the effects of age [odds ratio 0.5; 95% confidence interval (0.31-0.82) per 10-year increment (P = 0.006)] were confined to genotype 1 disease. In order to further inform the management of the individual patient, a multivariate logistic model was used to predict the probability of SVR for subgroups defined by disease stage, genotype and age at commencement of therapy. This model revealed striking differences in predicted response rates between subgroups and provided a strong rationale for early treatment, particularly for those with genotype 1 disease. Our study demonstrates that results comparable with those of RCTs can be achieved in clinical practice, and suggests that prediction of response rates based on probability modelling will provide a valuable adjunct to individual patient management.

Hepatitis C virus infection.

Hepatitis C virus (HCV) infection is a major public health problem. Up to 3% of the world's population is infected with HCV, and at least 200 000 adults in the UK carry the virus. Of those exposed to HCV, 80% become chronically infected, and at least 30% of carriers develop chronic liver disease, including cirrhosis and hepatocellular carcinoma. This review provides an overview of selected features of the molecular biology and pathogenesis of HCV infection, and thereafter discusses in detail the epidemiology of HCV, the hepatic and extra-hepatic diseases caused by the virus, and the current treatment options for both acute and chronic virus infection. The special cases of healthcare workers, prison inmates and individuals coinfected with human immunodeficiency virus and HCV are considered in detail.

Human retroviruses.

After many unsuccessful years of searching, the first pathogenic human retrovirus, the human T-cell leukaemia lymphoma virus (HTLV-I), was reported as recently as 1980 and since that time has been causally linked to the adult T-cell leukaemia lymphoma syndrome. A second HTLV (HTLV-II) isolated shortly afterwards is less clearly linked to some leukaemic and chronic lymphoid malignancies. The second major family of human retroviruses are the human immunodeficiency viruses (HIV) the first group of isolates (HIV-I) of which cause the acquired deficiency syndrome (AIDS). A second group of these viruses (HIV-II), have recently been identified in West Africa. They appear to be less clearly associated with disease and more similar in molecular structure to the Simian immunodeficiency viruses. AIDS has now become a major global pandemic, and vaccine and therapeutic strategies are urgently being investigated in an effort to control the disease. Unfortunately, current results are not very encouraging. In the meantime, preventative and educational measures are of utmost priority in order to prevent further spread. It is not unlikely that new human retroviruses will be discovered over the next few years.

Low incidence of human herpesvirus 8 in stem cell collections from myeloma patients.

Human herpesvirus 8 is a gammaherpesvirus which may be implicated in the pathogenesis of multiple myeloma. Viral DNA sequences have been found in the bone marrow, peripheral blood and leukapheresis products of myeloma patients. These findings have significant implications for the use of leukapheresed cells in the transplantation and immunotherapy of myeloma. The studies suggest the cell which harbours the virus may be dendritic in origin. We have previously reported that dendritic cells cultured for use in the clinical setting do not harbour HHV-8. In this study, we examined the leukapheresis products of a larger cohort of myeloma patients for the presence of HHV-8 using a highly sensitive PCR technique. A strong association between HHV-8 and myeloma was not confirmed, with only 4% of the patient samples positive for viral sequences. While further study is needed, the current use of apheresis cells and their cultured progeny in the treatment of myeloma should not be compromised.

Development of a strand-specific RT-PCR based assay to detect the replicative form of hepatitis C virus RNA.

The recent development of tagged RT-PCR and rTth RT-PCR has greatly improved strand-specific detection of hepatitis C virus (HCV) RNA but these assays are still prone to some false detection of the incorrect strand of RNA. In this study we aimed to address additional factors which contribute towards false detection of HCV RNA. Firstly the benefits of both tagged primers and the thermostable reverse transcriptase rTth during cDNA synthesis were combined and it was found that strand specificity was greatly improved without compromising sensitivity. The reliability of the assay was then optimised by addressing the following issues: control synthetic transcripts should be free of contaminating plasmid DNA, residual RT activity should be minimised in the presence of PCR primers and cDNA should be free of unincorporated tagged RT primer prior to PCR amplification. The alterations made to the assay eliminated completely false detection of the incorrect strand of RNA in the control assay whilst the correct strand was consistently detected at a cDNA dilution of 10(-3)-10(-4). Negative strand was not detected in RNA isolated from serum but was detected, at a ten-fold lower level than positive strand, in RNA isolated from liver tissue.

Q fever in pregnancy.

We describe a case of acute symptomatic infection with Coxiella burnetii acquired between the 16th and 28th week of pregnancy. Oral ciprofloxacin therapy was started on diagnosis, at the 28th week of pregnancy, but symptoms were unabated after 3 weeks treatment, suggesting persisting infection of the products of conception. Caesarean section was therefore performed at 32 weeks gestation when a healthy infant was delivered, and subsequent investigations showed no evidence of transplacental spread of infection. Infection control measures were applied at the time of delivery to minimize the risk of infection to obstetricians and midwives from potentially infectious products of conception.

Erythema nodosum following typhoid vaccination.

Erythema nodosum has recently been recognised in association with salmonella infection (1). We report a case of classical erythema nodosum which developed following a typhoid vaccination.

Clinical trial: a primary-care-based model for the delivery of anti-viral treatment to injecting drug users infected with hepatitis C.

BACKGROUND: Injecting drug use is the main risk factor for hepatitis C virus (HCV) infection. Secondary-care-based strategies for the management of HCV do not effectively target this vulnerable population. AIMS: To evaluate the feasibility, safety and efficacy of a primary-care-based model for the delivery of HCV services including anti-viral therapy to injecting drug users. METHODS: A partnership between a clinical nurse specialist employed by, and working under the supervision of, a secondary-care-based hepatitis service and drug workers and general practitioners. Three hundred and fifty-three clients attending opiate substitution clinics in primary care were evaluated. Outcomes were: number of new diagnoses of HCV infection, number of clients assessed as suitable for anti-viral treatment, and number of patients treated. RESULTS: 174 HCV antibody positive clients were identified. Of these, 124 were chronically infected with HCV of whom only six had been previously identified. Of 118 new chronically-infected individuals, 86 entered the care pathway, 43 were assessed as suitable for anti-viral treatment and 30 have so far been treated. Outcomes of anti-viral treatment are comparable with those obtained in secondary care settings. CONCLUSION: A primary-care-based model offers a new paradigm for the treatment of HCV in injecting drug users.

A closer look at water-related geologic activity on Mars.

Water has supposedly marked the surface of Mars and produced characteristic landforms. To understand the history of water on Mars, we take a close look at key locations with the High-Resolution Imaging Science Experiment on board the Mars Reconnaissance Orbiter, reaching fine spatial scales of 25 to 32 centimeters per pixel. Boulders ranging up to approximately 2 meters in diameter are ubiquitous in the middle to high latitudes, which include deposits previously interpreted as finegrained ocean sediments or dusty snow. Bright gully deposits identify six locations with very recent activity, but these lie on steep (20 degrees to 35 degrees) slopes where dry mass wasting could occur. Thus, we cannot confirm the reality of ancient oceans or water in active gullies but do see evidence of fluvial modification of geologically recent mid-latitude gullies and equatorial impact craters.

Caspase-independent killing of Burkitt lymphoma cell lines by rituximab.

Caspase-independent cell death may have a critical role to play in the therapeutic destruction of tumours. Recently it has been suggested that one of the mechanisms by which rituximab, a therapeutic anti-CD20 antibody, kills B cells is caspase-independent. In this study we show that rituximab can induce death in a variety of Burkitt lymphoma derived cell lines. Rituximab-treated cells show leakage of adenylate kinase, surface expression of phosphatidylserine, upregulation of the cellular stress protein HSP70, phosphorylation of the survival protein Akt, and depolarisation of the mitochondrial membrane but no loss of cytochrome c or apoptosis inducing factor. Caspase inhibitors do not block these events. In support of these data there is no cleavage of caspases 3, 8 and 9, poly(ADP-ribose) polymerase, BH3 interacting domain death agonist or genomic DNA. Morphologically, cells show nuclear enlargement and cytoplasmic vacuolisation. Triggering of receptor mediated death in CD95 responsive lines results in "classical" apoptosis indicating that the internal machinery necessary for apoptosis is intact in these lines. The results suggest that rituximab can kill human B cells via a caspase-independent form of programmed cell death that shares features of apoptosis and necrosis. This pathway may be relevant to the clinical efficacy of rituximab.

Clinical pathways for patients with newly diagnosed hepatitis C - what actually happens.

Management of hepatitis C virus (HCV)-infected individuals requires referral to specialist care. To determine whether patients newly diagnosed as anti-HCV positive are appropriately referred for further investigation and management, and if not, to determine why not. We studied patients tested for antibodies to HCV by Nottingham Public Health Laboratory in a 2-year period (2000-2002). The progress of newly diagnosed anti-HCV positive patients into specialist clinics for further management was documented. For patients not referred for specialist care, a questionnaire was sent to the clinician requesting the initial anti-HCV test, to identify reasons for nonreferral. Eleven thousand one hundred and seventy-seven patients were tested for anti-HCV. Two hundred and fifty-six (2.3%) were newly diagnosed as being anti-HCV positive. Two per cent of samples sent from primary care were anti-HCV positive, compared to 18.8, 18.9 and 1.3% sent from prison, drug and alcohol units, and secondary care, respectively. About 64.3% of positive patients diagnosed in primary care were referred to specialist care, compared to 18.4, 42.4 and 62.6% of patients diagnosed in the other three settings. One hundred and twenty-five (49%) newly diagnosed patients were referred appropriately for further management. 68 of these attended clinic, 45 underwent liver biopsy and 26 (10%) began treatment. One hundred and thirty-one patients (51%) were not referred. In 54 cases, there was no evidence that the anti-HCV positive result reached the patient. In 15, referral was considered but rejected, and 20 patients were referred to non-HCV-specialists (their general practitioners or to genito-urinary medicine). Hence less than 50% of newly diagnosed anti-HCV positive patients are referred to an appropriate clinic for further investigation and management. Reasons for this are multifarious and complex, reflecting both systems failure and patient choice. Unless these are understood and addressed, the Department of Health Hepatitis C Strategy (2002) and Action Plan for England (2004) will fail to achieve their intended objectives.

Viruses and apoptosis.

Apoptosis, or programmed cell death, is essential in development and homeostasis in multi-cellular organisms. It is also an important component of the cellular response to injury. Many cells undergo apoptosis in response to viral infection, with a consequent reduction in the release of progeny virus. Viruses have therefore evolved multiple distinct mechanisms for modulating host cell apoptosis. Viruses may interfere with either the highly conserved 'effector' mechanisms of programmed cell death or regulatory mechanisms specific to mammalian cells. In addition to conferring a selective advantage to the virus, the capacity to prevent apoptosis has an essential role in the transformation of the host cell by oncogenic viruses. This article provides a focussed review of apoptosis and illustrates how the study of viruses has informed our understanding of this process. Selected mechanisms by which viral gene products interfere with cell death are discussed in detail and used to illustrate the general principles of the interactions between viruses and apoptosis.

The right end of the unique region of the genome of human herpesvirus 6 U1102 contains a candidate immediate early gene enhancer and a homologue of the human cytomegalovirus US22 gene family.

The nucleotide sequence of a 12 kbp HindIII fragment (HindIII C) from the right end of the unique component of the genome of human herpesvirus 6 (HHV-6) (strain U1102) was determined. The sequence has a mean G + C content of 42% and contains approximately 28 copies of a tandemly repeated 104 to 107 bp element, which, with a single exception, contain a cleavage site for KpnI (the KpnI repeats). Each of these elements contains potential binding sites for transcription factors NF-kappa B and AP2. The KpnI repeats lie immediately upstream of a region previously identified as a candidate immediate early (IE) gene locus and therefore may constitute an IE gene enhancer element. One incomplete and six complete open reading frames (ORFs) were identified in the unique sequence of the HindIII C fragment. The predicted products of these ORFs do not include homologues of proteins encoded by members of the alpha- or gamma-herpesvirus sub-family. However, the HindIII C fragment does contain a homologue of the US22 gene family, previously found only in the beta-herpesvirus human cytomegalovirus (HCMV). These findings provide evidence that the close phylogenetic relationship between HHV-6 and HCMV is not confined to the beta-herpesvirus-specific arrangement of conserved replicative and structural genes which has been demonstrated previously.

Acquisition of the human adeno-associated virus type-2 rep gene by human herpesvirus type-6.

Human herpesvirus type-6 (HHV-6) is a recently isolated herpesvirus which is highly prevalent in adult populations around the world. HHV-6 was first isolated from the peripheral blood of six individuals with lymphoproliferative disorders, two of whom were also infected with human immunodeficiency virus. HHV-6, in common with other herpesviruses, transactivates the HIV long terminal repeat linked to reporter genes and has in addition been shown to accelerate HIV gene expression and CD4 cell death in cultures co-infected with both viruses. The virus is tropic for CD4+ lymphocytes and persists in the peripheral blood of most seropositive individuals. We have now identified a gene in HHV-6 encoding a 490-amino-acid polypeptide homologous to the human adeno-associated virus type-2 (AAV-2) rep gene. This gene has an essential role in AAV-2 DNA replication, can trans-regulate homologous and heterologous gene expression, and inhibits cellular transformation. The acquisition of rep by HHV-6 could be due to natural transfer of genetic information between DNA viruses of eukaryotes and is likely to have important consequences for the life-cycle of HHV-6 and for the host CD4 cell.

Structure and heterogeneity of the a sequences of human herpesvirus 6 strain variants U1102 and Z29 and identification of human telomeric repeat sequences at the genomic termini.

The unit-length genome of human herpesvirus 6 (HHV-6) consists of a single unique component (U) bounded by direct repeats DRL and DRR and forms head-to-tail concatemers during productive infection. cis-elements which mediate cleavage and packaging of progeny virions (a sequences) are found at the termini of all herpesvirus genomes. In HHV-6, DRL and DRR are identical and a sequences may therefore also occur at the U-DR junctions to give the arrangement aDRLa-U-aDRRa. We have sequenced the genomic termini, the U-DRR junction, and the DRR.DRL junction of HHV-6 strain variants U1102 and Z29. A (GGGTTA)n motif identical to the human telomeric repeat sequence (TRS) was found adjacent to, but did not form, the termini of both strain variants. The DRL terminus and U-DRR junction contained sequences closely related to that of the well-conserved herpesvirus packaging signal Cn-Gn-Nn-Gn (pac-1), followed by tandem arrays of TRSs separated by single copies of a hexanucleotide repeat. HHV-6 strain U1102 contained repeat sequences not found in HHV-6 Z29. In contrast, the DRR terminus of both variants contained a simple tandem array of TRSs and a close homolog of a herpesvirus pac-2 signal (GCn-Tn-GCn). The DRR.DRL junction was formed by simple head-to-tail linkage of the termini, yielding an intact cleavage signal, pac-2.x.pac-1, where x is the putative cleavage site. The left end of DR was the site of intrastrain size heterogeneity which mapped to the putative a sequences. These findings suggest that TRSs form part of the a sequence of HHV-6 and that the arrangement of a sequences in the genome can be represented as aDRLa-U-a-DRRa.

Human herpesvirus 6 (HHV-6) is a helper virus for adeno-associated virus type 2 (AAV-2) and the AAV-2 rep gene homologue in HHV-6 can mediate AAV-2 DNA replication and regulate gene expression.

We have previously described the apparent acquisition by human herpesvirus 6 (HHV-6) of the multifunctional rep gene of the helper-dependent human parvovirus adeno-associated virus type 2 (AAV-2). We report here that HHV-6 is a full helper virus for AAV-2 replication, suggesting a mechanism for transfer of the rep gene between the two viruses by recombination of replicative intermediates. The HHV-6 rep gene cloned under control of the human cytomegalovirus immediate early promoter complemented replication of a rep-deficient AAV-2 genome. In cotransfection experiments with heterologous promoters linked to the CAT reporter gene, HHV-6 rep activated the human immunodeficiency virus (HIV) long terminal repeat (LTR) in fibroblast cell lines but not in T-cells. In contrast, AAV-2 rep inhibited HIV LTR activity in both fibroblast and T-cell lines. The effect of HHV-6 and AAV-2 rep genes on the HIV LTR was independent of the NF-kappa B, Sp1, and TATA box elements. These results suggest that HHV-6 Rep is a multifunctional regulatory protein with properties related to, but distinct from, those of AAV-2 Rep.