Outcome measures analysis following total knee arthroplasty in patients with severe haemophilic arthropathy of the knee.

Total knee arthroplasty (TKA) has been the gold standard for treating severe haemophilic arthropathy of the knee when all conservative measures fail. However, performing a TKA in patients with haemophilic arthropathy is difficult due to severe joint deformity and destruction, and poor bone quality. The aim of the present study was to evaluate the short-term results of TKA in the treatment of knee haemophilic arthropathy in a tertiary referral centre, with an emphasis on health-related quality of life and knee function. A prospective study was conducted that included 19 male patients with end-stage haemophilic knee arthropathy who underwent TKA in a tertiary referral centre. Clinical outcome and health-related quality of life were assessed by the Western Ontario and McMaster Universities Arthritis (WOMAC) index and the Short Form-36 (SF-36) both pre-operatively and at 1-year post-operatively. The mean age of the patients was 50.37±7.63 years (range, 40-65 years). Pre-operative health-related quality of life was impaired in all patients in all SF-36 domains but was markedly improved after TKA. Knee function in all dimensions (pain, stiffness and physical function), as measured by the WOMAC questionnaire, significantly improved after TKA. Pre-operative pain, stiffness and function, along with total WOMAC score, were strongly and negatively correlated with pre-operative SF-36. Overall, the present study indicated a significant improvement in quality of life and clinical outcome after TKA in patients with advanced haemophilic arthropathy. More studies with longer follow-up periods in a larger population are needed to fully elucidate the mid- and long-term values of TKA in haemophilic patients.

Quality of Life among Patients with Retinal Vein Occlusion: A Case-Control Study.

Purpose: The purpose of this study was to evaluate health-related quality of life in patients with retinal vein occlusion (RVO) and investigate the possible risk factors for poor quality of life in patients with RVO.Methods: Participants in the study were 67 patients with RVO, 42 male and 25 female, mean-aged 73.1 ± 10.9 years, and 70 sex- and age-matched controls. Demographic data, lifestyle factors and medical history were recorded. All patients underwent best-corrected visual acuity measurement, dilated fundoscopy and optical coherence tomography. All participants completed two questionnaires assessing quality of life (EQ-5D, NEI VFQ-25). Risk factors for health-related quality of life in RVO patients were investigated.Results: Patients with RVO exhibited significantly lower composite score for VFQ-25 compared to controls (74.1 ± 3.8 vs. 91.7 ± 3.9 for patients and controls, respectively, p < .001). In addition, RVO patients had significantly lower EQ-5D Index score compared to controls (0.88 ± 0.15 vs. 0.92 ± 0.12 for patients and controls, respectively, p = .043). Risk factors associated with quality of life in patients with RVO were found the alcohol consumption, the presence of thyroidopathy, coagulation disorders, visual acuity in the eye with RVO, central retinal thickness, the type of edema, the presence of ischemia and the condition of external limiting membrane. In multivariate analysis, only alcohol consumption and visual acuity in the eye with RVO were found to be independent risk factors, affecting quality of life in RVO patients.Conclusions: Patients with RVO presented lower quality of life in comparison with controls. Potential risk factors should be taken into account and their early detection may improve quality of life in such patients and lead to targeted health policies.

Early caloric deprivation in preterm infants affects Bayley-III scales performance at 18-24 months of corrected age.

BACKGROUND: Adequate nutrition is essential for optimal neurodevelopment to preterm infants. Our aim was to evaluate the impact of caloric deprivation on Bayley-III scales performance at 18-24 months of corrected age, in a cohort of preterm infants. METHODS: We prospectively enrolled infants with gestational age <30 weeks and birth weight <1500 g. Apart from a whole cohort analysis, we performed a subgroup analysis between infants received inadequate calories (<85 Kcal/kg/day) during the first two weeks of age, compared to a standard nutrition group. All infants underwent a Bayley-III assessment at 18-24 months of corrected age. RESULTS: From the 63 preterm infants analysed, 25% had caloric deprivation compared to 75% with adequate nutrition. Caloric deprived infants were of lower gestational age and birth weight, and received a lower amount of enteral feeding during the first 14 days of age. There were no differences between the two groups regarding the common neonatal co-morbidities. Caloric deprived infants had significantly lower composite index scores at 18-24 months of corrected age. Caloric deprivation, late onset sepsis, necrotizing enterocolitis, and bronchopulmonary dysplasia were significant risk factors of neurodevelopmental impairment. CONCLUSIONS: Several neonatal factors affect the neurodevelopmental outcome of preterm infants, and nutrition may pose an important role.

HCV Defective Genomes Promote Persistent Infection by Modulating the Viral Life Cycle.

Defective interfering (DI) RNAs have been detected in several human viruses. HCV in-frame deletions mutants (IFDMs), missing mainly the envelope proteins, have been found in patient sera and liver tissues. IFDMs replicate independently and can be trans-packaged into infectious virions in the presence of full length viral genome. So far, their biological role is unclear. In this study, we have isolated and cloned IFDMs from sera samples and liver tissues of patients infected with HCV genotypes 1b, 2a, and 3a. IFDMs were present in up to 26% of samples tested. Using the in vitro HCV cell culture system, co-expression of the wild type (wt) HCV replicon with HCV IFDMs RNA resulted in increased HCV replication. Additionally, co-transfection of the HCV full length genome RNA and a defective mutant missing the envelope region led to increased viral release, collectively suggesting an important biological role for IFDMs in the virus life cycle. Recently, exosomes, masters of intercellular communication, have been implicated in the transport of HCV viral genomes. We report for the first time that exosomal RNA isolated from HCV sera samples contains HCV defective genomes. We also demonstrate that inhibition of exosomal biogenesis and release influences HCV viral replication. Overall, we provide evidence that the presence of HCV IFDMs affects both viral replication and release. IFDMs exploit exosomes as means of transport, a way to evade the immune system, to spread more efficiently and possibly maintain persistent infection.

Impact of Radiofrequency Ablation and Antiarrhythmic Medications on the Quality of Life of Patients with Supraventricular Tachycardias: Preliminary Validation of the Greek Version of the Umea22 (U22) Questionnaire.

OBJECTIVE: This study aims to (i) translate, culturally adapt, and preliminarily validate the arrhythmia-specific Umea22 (U22) questionnaire and (ii) assess the impact of radiofrequency (RF) ablation and medical treatment on the quality of life of patients with supraventricular tachycardias (SVTs). METHODS: A total of 140 patients with atrioventricular nodal re-entry tachycardia (AVNRT) and atrioventricular re-entry tachycardia (AVRT) were enrolled in the study. Of these, 100 patients underwent RF ablation (group A) and 40 patients were managed with antiarrhythmic medications (group B). Health-related quality of life (HRQoL) was assessed for both groups using the Short Form-36 Health Survey (SF-36) and the arrhythmia-specific Umea22 (U22) questionnaire at baseline and 3-month follow-up. Exploratory and confirmatory factor analyses were performed to assess the validity of the U22 questionnaire. Univariate comparisons of HRQoL scores between study timepoints and multivariate regression analyses adjusting for baseline confounders were conducted. RESULTS: The factor analysis of the U22 questionnaire yielded a six-factor model ("burden of spells"; "heart contractility"; "character of spells"; "general/non-specific feeling"; "other specific somatic symptoms"; "fear") with acceptable fit results. Patients of group A showed significant improvement in all SF-36 and U22 scores at 3 months' follow-up compared to baseline (all p<0.05). Patients of group B presented deterioration of the total SF-36 score (p=0.001) and improvement of certain U22 measures, namely, well-being (p=0.004), heartbeat speed, and intensity during arrhythmia spells (p<0.0001 for both measures) at 3 months' follow-up, compared to baseline. Employment status, male sex, and urban residence emerged as important predictors. CONCLUSION: The Greek version of the U22 questionnaire is a valid tool to assess SVT-related symptoms. RF ablation appears to exert more pronounced beneficial outcomes on HRQoL of patients with SVTs compared to medical treatment. Prompt referral of patients with SVTs to specialist centers may favorably affect their quality of life and should be encouraged.

Health-Related Quality of Life and Association With Arthropathy in Greek Patients with Hemophilia.

Health-related quality of life (HRQoL) is increasingly implicated in contemporary hemophilia management. This study focuses on the assessment of HRQoL in Greek patients with hemophilia and the comparison with normative data from the general population, as well as on the extent arthropathy may affect the patients' HRQoL. One hundred and nine adult patients completed the Greek social functioning (SF-36) and Haem-A-QoL questionnaires. Arthropathy was assessed by both the World Federation of Hemophilia clinical score and Pettersson radiological score. The most impaired domains of Haem-A-QoL were sports/leisure (SL) and physical health (PH; mean scores 61.2 and 42.2, respectively). The patients experienced statistically significant lower mean scores in all SF-36 domains than the normative sample, especially in role physical (RPH), bodily pain (BP), and general health (GH) subscales. Among Haem-A-QoL subscales, SL and PH were found strongly associated with severity of arthropathy using both orthopedic scores ( P < .001), and maintained the statistical significance after adjustment for age ( P < .05). A poor orthopedic status was also negatively associated with certain SF-36 subscales. However, none of these correlations remained after adjustment with age. Compared to normative data from Greece, patients with hemophilia showed deterioration in all HRQoL subscales, with a more pronounced effect in RPH, BP, and GH subscales. Health-related quality of life was strongly influenced by arthropathy, mainly in the physical aspects of HRQoL. The use of the disease-specific Haem-A-QoL tool can capture additional associations between HRQoL and hemophilic arthropathy.

Occult hepatitis B virus infection in Greek patients with congenital bleeding disorders.

Occult Hepatitis B Infection (OBI) is a form of chronic HBV infection characterized by low level HBV DNA, without detectable HBV surface antigen (HBsAg). OBI is frequently associated with the presence of anti-HBc and in some cases also with anti-HBs. Patients, who formerly received non-inactivated factor concentrates, can potentially be considered at high risk for OBI, especially since these patients usually are HIV or HCV co-infected. This study aimed to assess the prevalence of occult HBV infection in Greek patients with hereditary bleeding disorders. The study sample comprised of 114 patients from a single haemophilia center. All patients were screened for HBV serum markers and individually tested for HBV DNA using a qualitative PCR. Presence of HBV DNA was further confirmed by quantification of viral load with an ultrasensitive in-house real time PCR. 88 and 21 patients with haemophilia A and B, respectively, 4 patients with von Willebrand Disease and 1 patient with severe factor VII deficiency were screened for the presence of OBI. Anti-HBc were detected in 53 (46.5%) subjects; 18 of them were anti-HBs(-) and 35 anti-HBs(+). Anti-HBe were present in 26 subjects. Two out of 114 patients were HBsAg(+). Of the remaining 112 HBsAg(-) patients tested, two (1.8%) were found HBsAg(-), HBV DNA(+), anti-HBc(+) and anti-HBs(-) and were identified as potential OBI cases. Both cases exhibited very low DNA levels; 38.2IU/mL in patient A and 14.2IU/mL in patient B. Both patients were HBeAg(-), but patient A had HBe antibodies. Patient B was also HIV/HCV co-infected. In conclusion, two cases of OBI with low HBV viraemia were identified among patients with congenital bleeding disorders. Although the incidence in our sample is moderately low (1.8%), close monitoring of these infections is of great clinical significance, especially in patients with co-infections and concomitant immunosuppression.

Alterations in the iron homeostasis network: A driving force for macrophage-mediated hepatitis C virus persistency.

Mechanisms that favor Hepatitis C virus (HCV) persistence over clearance are unclear, but involve defective innate immunity. Chronic infection is characterized by hepatic iron overload, hyperferraemia and hyperferittinaemia. Hepcidin modulates iron egress via ferroportin and its storage in ferritin. Chronic HCV patients have decreased hepcidin, while HCV replication is modified by HAMP silencing. We aimed to investigate interactions between HCV and hepcidin, during acute and chronic disease, and putative alterations in cellular iron homeostasis that enhance HCV propagation and promote viral persistence. Thus, we used HCV JFH-1-infected co-cultures of Huh7.5 hepatoma and THP-1 macrophage cells, HCV patients' sera and Huh7 hepcidin-expressing cells transfected with HCV replicons. Hepcidin levels were elevated in acutely infected patients, but correlated with viral load in chronic patients. HAMP expression was up-regulated early in HCV infection in vitro, with corresponding changes in ferritin and FPN. Hepcidin overexpression enhanced both viral translation and replication. In HCV-infected co-cultures, we observed increased hepcidin, reduced hepatoma ferritin and a concurrent rise in macrophaghic ferritin over time. Altered iron levels complemented amplified replication in hepatoma cells and one replication round in macrophages. Iron-loading of macrophages led to enhancement of hepatic HCV replication through reversed ferritin "flow." Viral transmissibility from infected macrophages to naïve hepatoma cells was induced by iron. We propose that HCV control over iron occurs both by intracellular iron sequestration, through hepcidin, and intercellular iron mobilisation via ferritin, as means toward enhanced replication. Persistence could be achieved through HCV-induced changes in macrophagic iron that enhances viral replication in these cells.

Psychometric properties of the Greek Haem-A-QoL for measuring quality of life in Greek haemophilia patients.

BACKGROUND AND OBJECTIVES: Health Related Quality of Life (HRQoL) is an important health outcome measure in haemophilia. The aim of this study was to assess the psychometric properties of the Greek version of Haem-A-QoL, a disease-specific questionnaire for haemophiliacs. METHODS: Haem-A-QoL and SF-36 were administered to 118 adult haemophilia patients. Hypothesized scale structure, internal consistency (Cronbach's α ), and test-retest reliability, as well as various types of construct validity were evaluated. RESULTS: Scale structure of Haem-A-QoL was confirmed, with good item convergence (87%) and discrimination (80.6%) rates. Cronbach's α was >0.70 for all but one dimension (dealing) and test-retest reliability was significantly high. The strength of Spearman's correlations between Haem-A-QoL and SF-36 scales ranged from 0.25 to 0.75 (P < 0.01). Multiple stepwise linear regression analysis revealed that all but one Haem-A-QoL dimensions were important predictors of SF-36 scales. Known-groups comparisons yielded consistent support of the instruments' construct validity and significant relationships were identified for age, educational level, haemophilia type, disease severity, and viral infections. CONCLUSION: Overall, the psychometric properties of the Greek version of Haem-A-QoL, resulting from this first time administration of the instrument to Greek adult haemophiliacs, confirmed it as a reliable and valid questionnaire for assessing haemophilia-specific HRQoL in Greece.

Development of a new ultra sensitive real-time PCR assay (ultra sensitive RTQ-PCR) for the quantification of HBV-DNA.

BACKGROUND: Improved sensitivity of HBV-DNA tests is of critical importance for the management of HBV infection. Our aim was to develop and assess a new ultra sensitive in-house real-time PCR assay for HBV-DNA quantification (ultra sensitive RTQ-PCR). RESULTS: Previously used HBV-DNA standards were calibrated against the WHO 1st International Standard for HBV-DNA (OptiQuant(R) HBV-DNA Quantification Panel, Accrometrix Europe B.V.). The 95% and 50% HBV-DNA detection end-point of the assay were 22.2 and 8.4 IU/mL. According to the calibration results, 1 IU/mL equals 2.8 copies/mL. Importantly the clinical performance of the ultra sensitive real-time PCR was tested similar (67%) to the Procleix Ultrio discriminatory HBV test (dHBV) (70%) in low-titer samples from patients with occult Hepatitis B. Finally, in the comparison of ultra sensitive RTQ-PCR with the commercially available COBAS TaqMan HBV Test, the in-house assay identified 94.7% of the 94 specimens as positive versus 90.4% identified by TaqMan, while the quantitative results that were positive by both assay were strongly correlated (r = 0.979). CONCLUSIONS: We report a new ultra sensitive real time PCR molecular beacon based assay with remarkable analytical and clinical sensitivity, calibrated against the WHO 1st International standard.

Green fluorescent protein - Tagged HCV non-enveloped capsid like particles: development of a new tool for tracking HCV core uptake.

Circulating 'free' non-enveloped Hepatitis C virus (HCV) core protein has been demonstrated in HCV-infected patients, and HCV subgenomes with deletions of the envelope proteins have been previously identified. Initial studies from our laboratory, previously published, indicated that expression of HCV core in insect cells can direct the formation of capsid-like particles lacking the envelope glycoproteins. These protein nanospheres, morphologically similar to natural capsids, were shown to be taken up by human hepatic cells and to produce cell-signalling events. To follow the intracellular fate of these particles we fused the core protein to eGFP. We demonstrate that the chimeric proteins core(173)-eGFP, eGFP-core(191) and eGFP-core(173) can be efficiently expressed, self-assembled, and form fluorescent non-enveloped capsid-like particles. By using confocal microscopy and FACS analysis, we provide evidence that the fluorescent nanospheres can not only enter human hepatic cells - the main target of HCV - but also human immune cells such as T and B lymphocytes, as well as human myeloid leukaemia cells differentiated along the monocyte/macrophage-like pathway. The fluorescent particles might thus be used to trace the intracellular trafficking of naked HCV capsids as showed by live microscopy and to further understand their biological significance.

Molecular characterization of occult hepatitis B cases in Greek blood donors.

The use of sensitive nucleic acid testing for hepatitis B virus in blood donors revealed a number of HBV DNA(+) cases among HBsAg(-) donors, a status known as occult HBV infection. The purpose of this study was the serological and molecular characterization of occult HBV infection in Greek blood donors. A prospective study was undertaken in order to identify occult HBV infection cases in blood donors. As part of the routine screening of blood donations in Greece, blood units were screened individually by a multiplex HIV-1/HCV/HBV nucleic acid assay. Initially reactive samples were retested with discriminatory assays. HBV DNA(+)/HBsAg(-) samples were tested further for HBV serological markers and HBV DNA was quantified by real-time PCR. Molecular characterization was performed by sequencing the envelope and polymerase genes of HBV. Preliminary screening revealed 21 occult cases with the following patterns: anti-HBc only: 7 donors, anti-HBc/anti-HBs: 7 donors, anti-HBc/anti-HBe: 5 donors, anti-HBc/anti-HBs/anti-HBe: 2 donors. In all cases, the HBV DNA load was <351 IU/ml. Sequencing was successful in 10 donors (classified within genotype D) revealing several amino acid substitutions related to diagnostic escape and antiviral resistance. HBsAg diagnostic failure and low viral replication in occult HBV infection carriers could possibly be attributed to multiple changes in envelope and polymerase regions, respectively.

Evidence for cellular uptake of recombinant hepatitis C virus non-enveloped capsid-like particles.

Although the hepatitis C virus (HCV) is an enveloped virus, naked nucleocapsids have been reported in the serum of infected patients, and most recently novel HCV subgenomes with deletions of the envelope proteins have been identified. However the significance of these findings remains unclear. In this study, we used the baculovirus expression system to generate recombinant HCV capsid-like particles, and investigated their possible interactions with cells. We show that expression of HCV core in insect cells can sufficiently direct the formation of capsid-like particles in the absence of the HCV envelope glycoproteins and of the 5' untranslated region. By confocal microscopy analysis, we provide evidence that the naked capsid-like particles could be uptaken by human hepatoma cells. Moreover, our findings suggest that they have the potential to produce cell-signaling effects.

Modulation of the hepatitis C virus RNA-dependent RNA polymerase activity by the non-structural (NS) 3 helicase and the NS4B membrane protein.

The hepatitis C virus (HCV) nonstructural protein 5B (NS5B) is believed to be the central catalytic enzyme responsible for HCV replication but there are many unanswered questions about how its activity is controlled. In this study we reveal that two other HCV proteins, NS3 (a protease/helicase) and NS4B (a hydrophobic protein of unknown function), physically and functionally interact with the NS5B polymerase. We describe a new procedure for generating highly pure NS4B, and use this protein in biochemical studies together with NS5B and NS3. To study the functional effects of the protein-protein interactions, we have developed an in vitro replication assay using the natural noncoding 3' regions of the respective positive ((+)-3'-untranslated region) and negative ((-)-3'-terminal region) RNA strands of the HCV genome. Our studies show that NS3 dramatically modulates template recognition by NS5B and changes the synthetic products generated by this enzyme. The use of an NTPase-deficient mutant form of NS3 demonstrates that the NTPase activity (and thus helicase activity) of this protein is specifically required for these effects. Moreover, NS4B is found to be a negative regulator of the NS3-NS5B replication complex. Overall, these results reveal that NS3, NS4B, and NS5B can interact to form a regulatory complex that could feature in the process of HCV replication.

Alternate translation occurs within the core coding region of the hepatitis C viral genome.

The majority of hepatitis C virus (HCV) isolates contain an open reading frame (ORF) overlapping with the core coding sequences in the +1 frame, which was assumed to be untranslated. We present evidence supporting the expression of this ORF (designated core+1 ORF) via novel translation mechanisms. First, fusion of the luciferase gene with the HCV-1 core+1 ORF followed by in vitro translation resulted in the synthesis of a chimeric protein (core+1-luciferase) that exhibited approximately 54% luciferase activity relative to the positive control (core-luciferase). Second, antisera raised against two different synthetic core+1 peptides recognized the previously identified p16 (but not p21) core protein band expressed from HCV-1, indicating the presence of epitopes from the core+1 ORF within the p16 protein. Third, HCV-positive sera specifically recognized lysates of Escherichia coli cells expressing recombinant core+1 protein, suggesting the presence of anti-core+1 antibodies in HCV-infected patients. Finally, luciferase tagging experiments designed to assess for -1 frameshifting combined with site-directed mutagenesis experiments supported the presence of +1/-1 ribosomal frameshift translation mechanisms within the core coding region. In conclusion, our data provide evidence for novel translation mechanisms within the core coding region and demonstrate the expression of the core+1 ORF, at least for some HCV isolates.