Association of Hydroxychloroquine Dose With Adverse Cardiac Events in Patients With Systemic Lupus Erythematosus.

OBJECTIVE: To determine whether hydroxychloroquine (HCQ) dose is associated with adverse cardiac outcomes in patients with systemic lupus erythematosus (SLE). METHODS: Patients with SLE taking HCQ and with ≥1 echocardiogram followed at a tertiary care center in the Bronx, New York between 2005 and 2021 were included. The HCQ weight-based dose at the HCQ start date was the main exposure of interest. The outcome was incident all-cause heart failure with reduced ejection fraction (HFrEF), life-threatening arrhythmia, or cardiac death. We used Fine-Gray regression models with death as a competing event to study the association of HCQ dose with the outcome. Due to a significant interaction between smoking and HCQ exposure, models were stratified by smoking status. Propensity score analysis was performed as a secondary analysis. RESULTS: Of 294 patients, 37 (13%) developed the outcome over a median follow-up time of 7.9 years (interquartile range [IQR] 4.2-12.3 years). In nonsmokers (n = 226), multivariable analysis adjusted for age, body mass index, hypertension, chronic kidney disease, diabetes mellitus, and thromboembolism showed that higher HCQ weight-based doses were not associated with an increased risk of the outcome (subdistribution hazard ratio [HR] 0.62 [IQR 0.41-0.92], P = 0.02). Similarly, higher baseline HCQ doses were not associated with a higher risk of the outcome among smokers (n = 68) (subdistribution HR 0.85 [IQR 0.53-1.34] per mg/kg, P = 0.48). Propensity score analysis showed comparable results. CONCLUSION: Higher HCQ doses were not associated with an increased risk of HFrEF, life-threatening arrhythmia, or cardiac death among patients with SLE and may decrease the risk among nonsmokers.

Characterization of the SRD5A3-CDG Clinical Spectrum

Abstract We aimed to characterize the clinical spectrum of patients diagnosed with SRD5A3-CDG, a subtype of congenital disorders of glycosylation (CDG) due to variants in the steroid 5a-reductase type 3 (SRD5A3) gene. It presents with multi-systemic involvement including neurological disability, dermatologic abnormalities, and ophthalmological defects. We conducted a cross-sectional study of children (n=6, ages 4-16 years) with a confirmed diagnosis of SRD5A3-CDG (c.57G>A, p.W19X). Families completed a detailed medical history questionnaire, two quality of life measures, and an adaptive behavior scale. Prevalent clinical features in our cohort included visual impairment (6/6), developmental delay (6/6), nystagmus (5/6), retinal dystrophy (4/6), and hypotonia (3/6). The Vineland Adaptive Behavior Scales demonstrated deficits across all functional domains (Composite Mean 36.17 ± 26.88), although one child did not show significant deficits. The QI-Disability Form demonstrated a mean total score of 64.8 (±12.7), and the PedsQL-Family Impact Module demonstrated a mean total score of 56.5 (±31.5). Vineland composite scores did not correlate with levels of disability captured by the QI-Disability Form (Pearson Correlation range -0.63 to +0.69, p>0.05 on all subscales). Ultimately, despite genotypic homogeneity, there is notable variability in adaptive functioning and quality of life among affected children that does not correlate with age.

Neutralizing antibody activity against 21 SARS-CoV-2 variants in older adults vaccinated with BNT162b2.

SARS-CoV-2 variants may threaten the effectiveness of vaccines and antivirals to mitigate serious COVID-19 disease. This is of most concern in clinically vulnerable groups such as older adults. We analysed 72 sera samples from 37 individuals, aged 70-89 years, vaccinated with two doses of BNT162b2 (Pfizer-BioNTech) 3 weeks apart, for neutralizing antibody responses to wildtype SARS-CoV-2. Between 3 and 20 weeks after the second vaccine dose, neutralizing antibody titres fell 4.9-fold to a median titre of 21.3 (neutralization dose 80%), with 21.6% of individuals having no detectable neutralizing antibodies at the later time point. Next, we examined neutralization of 21 distinct SARS-CoV-2 variant spike proteins with these sera, and confirmed substantial antigenic escape, especially for the Omicron (B.1.1.529, BA.1/BA.2), Beta (B.1.351), Delta (B.1.617.2), Theta (P.3), C.1.2 and B.1.638 spike variants. By combining pseudotype neutralization with specific receptor-binding domain (RBD) enzyme-linked immunosorbent assays, we showed that changes to position 484 in the spike RBD were mainly responsible for SARS-CoV-2 neutralizing antibody escape. Nineteen sera from the same individuals boosted with a third dose of BNT162b2 contained higher neutralizing antibody titres, providing cross-protection against Omicron BA.1 and BA.2. Despite SARS-CoV-2 immunity waning over time in older adults, booster vaccines can elicit broad neutralizing antibodies against a large number of SARS-CoV-2 variants in this clinically vulnerable cohort.

Comparative Risk of Alzheimer Disease and Related Dementia Among Medicare Beneficiaries With Rheumatoid Arthritis Treated With Targeted Disease-Modifying Antirheumatic Agents.

Importance: Cytokine signaling, including tumor necrosis factor (TNF) and interleukin (IL)-6, through the Janus-kinase (JAK)-signal transducer and activator of transcription pathway, was hypothesized to attenuate the risk of Alzheimer disease and related dementia (ADRD) in the Drug Repurposing for Effective Alzheimer Medicines (DREAM) initiative based on multiomics phenotyping. Objective: To evaluate the association between treatment with tofacitinib, tocilizumab, or TNF inhibitors compared with abatacept and risk of incident ADRD. Design, Setting, and Participants: This cohort study was conducted among US Medicare fee-for-service patients with rheumatoid arthritis aged 65 years and older from 2007 to 2017. Patients were categorized into 3 cohorts based on initiation of tofacitinib (a JAK inhibitor), tocilizumab (an IL-6 inhibitor), or TNF inhibitors compared with a common comparator abatacept (a T-cell activation inhibitor). Analyses were conducted from August 2020 to August 2021. Main Outcomes and Measures: The main outcome was onset of ADRD based on diagnosis codes evaluated in 4 alternative analysis schemes: (1) an as-treated follow-up approach, (2) an as-started follow-up approach incorporating a 6-month induction period, (3) incorporating a 6-month symptom to diagnosis period to account for misclassification of ADRD onset, and (4) identifying ADRD through symptomatic prescriptions and diagnosis codes. Hazard ratios (HRs) with 95% CIs were calculated from Cox proportional hazard regression after adjustment for 79 preexposure characteristics through propensity score matching. Results: After 1:1 propensity score matching to patients using abatacept, a total of 22 569 propensity score-matched patient pairs, including 4224 tofacitinib pairs (mean [SD] age 72.19 [5.65] years; 6945 [82.2%] women), 6369 tocilizumab pairs (mean [SD] age 72.01 [5.46] years; 10 105 [79.4%] women), and 11 976 TNF inhibitor pairs (mean [SD] age 72.67 [5.91] years; 19 710 [82.3%] women), were assessed. Incidence rates of ADRD varied from 2 to 18 per 1000 person-years across analyses schemes. There were no statistically significant associations of ADRD with tofacitinib (analysis 1: HR, 0.90 [95% CI, 0.55-1.51]; analysis 2: HR, 0.78 [95% CI, 0.53-1.13]; analysis 3: HR, 1.29 [95% CI, 0.72-2.33]; analysis 4: HR, 0.50 [95% CI, 0.21-1.20]), tocilizumab (analysis 1: HR, 0.82 [95% CI, 0.55-1.21]; analysis 2: HR, 1.05 [95% CI, 0.81-1.35]; analysis 3: HR, 1.21 [95% CI, 0.75-1.96]; analysis 4: HR, 0.78 [95% CI, 0.44-1.39]), or TNF inhibitors (analysis 1: HR, 0.93 [95% CI, 0.72-1.20]; analysis 2: HR, 1.02 [95% CI, 0.86-1.20]; analysis 3: HR, 1.13 [95% CI, 0.86-1.48]; analysis 4: 0.90 [95% CI, 0.60-1.37]) compared with abatacept. Results from prespecified subgroup analysis by age, sex, and baseline cardiovascular disease were consistent except in patients with cardiovascular disease, for whom there was a potentially lower risk of ADRD with TNF inhibitors vs abatacept, but only in analyses 2 and 4 (analysis 1: HR, 0.76 [95% CI, 0.50-1.16]; analysis 2: HR, 0.74 [95% CI, 0.56-0.99]; analysis 3: HR, 1.03 [95% CI, 0.65-1.61]; analysis 4: HR, 0.45 [95% CI, 0.21-0.98]). Conclusions and Relevance: This cohort study did not find any association of risk of ADRD in patients treated with tofacitinib, tocilizumab, or TNF inhibitors compared with abatacept.

Medications for Multiple Sclerosis and Risk of Malignancy: What Next?

Many autoimmune diseases confer a higher risk of cancer on patients compared to the general population. A controversial factor tying autoimmune diseases to malignancy is harm from immunosuppressive treatment. Nonetheless, multiple sclerosis is different from other autoimmune diseases, and findings from other disease populations may not apply. In this issue of Neurotherapeutics, Dolladile and colleagues from France present new evidence about the risks of cancers in patients with multiple sclerosis treated with disease-modifying therapies based on analyses of spontaneous reporting data. This commentary discusses the context, limitations, and implications of these findings.

Serological Cross-Reactions between Expressed VP2 Proteins from Different Bluetongue Virus Serotypes.

Bluetongue (BT) is a severe and economically important disease of ruminants that is widely distributed around the world, caused by the bluetongue virus (BTV). More than 28 different BTV serotypes have been identified in serum neutralisation tests (SNT), which, along with geographic variants (topotypes) within each serotype, reflect differences in BTV outer-capsid protein VP2. VP2 is the primary target for neutralising antibodies, although the basis for cross-reactions and serological variations between and within BTV serotypes is poorly understood. Recombinant BTV VP2 proteins (rVP2) were expressed in Nicotiana benthamiana, based on sequence data for isolates of thirteen BTV serotypes (primarily from Europe), including three 'novel' serotypes (BTV-25, -26 and -27) and alternative topotypes of four serotypes. Cross-reactions within and between these viruses were explored using rabbit anti-rVP2 sera and post BTV-infection sheep reference-antisera, in I-ELISA (with rVP2 target antigens) and SNT (with reference strains of BTV-1 to -24, -26 and -27). Strong reactions were generally detected with homologous rVP2 proteins or virus strains/serotypes. The sheep antisera were largely serotype-specific in SNT, but more cross-reactive by ELISA. Rabbit antisera were more cross-reactive in SNT, and showed widespread, high titre cross-reactions against homologous and heterologous rVP2 proteins in ELISA. Results were analysed and visualised by antigenic cartography, showing closer relationships in some, but not all cases, between VP2 topotypes within the same serotype, and between serotypes belonging to the same 'VP2 nucleotype'.

Biologic Switching Among Nonsystemic Juvenile Idiopathic Arthritis Patients: A Cohort Study in the Childhood Arthritis and Rheumatology Research Alliance Registry.

OBJECTIVE: Biologic medications have significantly improved disease control and outcomes of patients with juvenile idiopathic arthritis (JIA). Current treatment recommendations suggest escalating therapy, including changing biologics if needed, when inactive or low disease activity is not attained. The patterns and reasons for switching biologics in clinical practice in North America are not well described. METHODS: We used the Childhood Arthritis and Rheumatology Research Alliance Registry and included individuals with JIA if they newly started a biologic after January 1, 2008, and had at least 12 months of subsequent observable time. Subjects with systemic JIA were excluded. We compared characteristics of switchers and nonswitchers using chi-square for categorical variables and Wilcoxon rank-sum test for continuous variables, and used linear regression for time analysis. RESULTS: Of the eligible children, 1361 with JIA in the registry started a biologic (94% tumor necrosis factor inhibitors [TNFi]). Median followup time was 30 months and 349 (26%) switched biologics. Among biologic switchers, ineffectiveness/disease flare was the most common reason for switch (202, 58%). The most common documented switch was from etanercept to another TNFi (221, 63%). The median time to switch to a second biologic decreased substantially from 55.2 months in 2008 to 7.2 months in 2016. CONCLUSION: In a multicenter cohort of patients with JIA starting a biologic, one-quarter switched to a second biologic, and the time to switching decreased in recent years. Additional studies should evaluate the outcomes and optimal timing of switching and preferred sequence of biologic use.

Epidermal Neurite Density in Skin Biopsies From Patients With Juvenile Fibromyalgia.

OBJECTIVE: Fibromyalgia (FM) is defined by idiopathic, chronic, widespread musculoskeletal pain. In adults with FM, a metaanalysis of lower-leg skin biopsy demonstrated 45% pooled prevalence of abnormally low epidermal neurite density (END). END < 5th centile of the normal distribution is the consensus diagnostic threshold for small-fiber neuropathy. However, the clinical significance of END findings in FM is unknown. Here, we examine the prevalence of small-fiber pathology in juvenile FM, which has not been studied previously. METHODS: We screened 21 patients aged 13-20 years with FM diagnosed by pediatric rheumatologists. Fifteen meeting the American College of Rheumatology criteria (modified for juvenile FM) underwent lower-leg measurements of END and completed validated questionnaires assessing pain, functional disability, and dysautonomia symptoms. The primary outcome was proportion of FM patients with END < 5th centile of age/sex/race-based laboratory norms. Cases were systematically matched by ethnicity, race, sex, and age to a group of previously biopsied healthy adolescents with selection blinded to biopsy results. All 23 controls matching demographic criteria were included. RESULTS: Among biopsied juvenile FM patients, 53% (8/15) had END < 5th centile vs 4% (1/23) of healthy controls (P < 0.001). Mean patient END was 273/mm2 skin surface (95% CI 198-389) vs 413/mm2 (95% CI 359-467, P < 0.001). As expected, patients with FM reported more functional disability, dysautonomia, and pain than healthy controls. CONCLUSION: Abnormal END reduction is common in adolescents with FM, with similar prevalence in adults with FM. More studies are needed to fully characterize the significance of low END in FM and to elucidate the clinical implications of these findings.

Trends in Off-Label Drug Use in Ambulatory Settings: 2006-2015.

BACKGROUND: Off-label drug use in children is common and potentially harmful. In most previous off-label use research, authors studied hospitalized children, specific drug classes, or non-US settings. We characterized frequencies, trends, and reasons for off-label systemic drug orders for children in ambulatory US settings. METHODS: Using nationally representative surveys of office-based physicians (National Ambulatory Medical Care Surveys, 2006-2015), we studied off-label orders of systemic drugs for children age <18 based on US Food and Drug Administration-approved labeling for age, weight, and indication. We characterized the top classes and diagnoses with off-label orders and analyzed factors and trends of off-label orders using logistic regression. RESULTS: Physicians ordered ≥1 off-label systemic drug at 18.5% (95% confidence interval: 17.7%-19.3%) of visits, usually (74.6%) because of unapproved conditions. Off-label ordering was most common proportionally in neonates (83%) and in absolute terms among adolescents (322 orders out of 1000 visits). Off-label ordering was associated with female sex, subspecialists, polypharmacy, and chronic conditions. Rates and reasons for off-label orders varied considerably by age. Relative and absolute rates of off-label orders rose over time. Among common classes, off-label orders for antihistamines and several psychotropics increased over time, whereas off-label orders for several classes of antibiotics were stable or declined. CONCLUSIONS: US office-based physicians have ordered systemic drugs off label for children at increasing rates, most often for unapproved conditions, despite recent efforts to increase evidence and drug approvals for children. These findings can help inform education, research, and policies around effective, safe use of medications in children.

Telomere elongation through hTERT immortalization leads to chromosome repositioning in control cells and genomic instability in Hutchinson-Gilford progeria syndrome fibroblasts, expressing a novel SUN1 isoform.

Immortalizing primary cells with human telomerase reverse transcriptase (hTERT) has been common practice to enable primary cells to be of extended use in the laboratory because they avoid replicative senescence. Studying exogenously expressed hTERT in cells also affords scientists models of early carcinogenesis and telomere behavior. Control and the premature ageing disease-Hutchinson-Gilford progeria syndrome (HGPS) primary dermal fibroblasts, with and without the classical G608G mutation have been immortalized with exogenous hTERT. However, hTERT immortalization surprisingly elicits genome reorganization not only in disease cells but also in the normal control cells, such that whole chromosome territories normally located at the nuclear periphery in proliferating fibroblasts become mislocalized in the nuclear interior. This includes chromosome 18 in the control fibroblasts and both chromosomes 18 and X in HGPS cells, which physically express an isoform of the LINC complex protein SUN1 that has previously only been theoretical. Additionally, this HGPS cell line has also become genomically unstable and has a tetraploid karyotype, which could be due to the novel SUN1 isoform. Long-term treatment with the hTERT inhibitor BIBR1532 enabled the reduction of telomere length in the immortalized cells and resulted that these mislocalized internal chromosomes to be located at the nuclear periphery, as assessed in actively proliferating cells. Taken together, these findings reveal that elongated telomeres lead to dramatic chromosome mislocalization, which can be restored with a drug treatment that results in telomere reshortening and that a novel SUN1 isoform combined with elongated telomeres leads to genomic instability. Thus, care should be taken when interpreting data from genomic studies in hTERT-immortalized cell lines.

Risk of malignancy associated with paediatric use of tumour necrosis factor inhibitors.

OBJECTIVE: To determine whether tumour necrosis factor inhibitor (TNFi) use is associated with an increased rate of incident malignancy compared with no TNFi use in the treatment of juvenile idiopathic arthritis (JIA), paediatric inflammatory bowel disease (pIBD) and paediatric plaque psoriasis (pPsO). METHODS: We performed a retrospective cohort study of administrative claims data from the USA from 2000 to 2014. Exposure to TNFi was considered permanent from the first observed exposure onward. The malignancy outcome was defined by diagnosis codes with evidence of cancer treatment. We calculated standardised incidence ratios (SIRs) comparing the observed number of malignancies to the expected numbers according to cancer surveillance data. We used multivariable Cox proportional hazards models to estimate adjusted HRs (aHRs) for incident malignancy. RESULTS: We identified 15 598 children with TNFi use and 73 839 children with no TNFi use (30 703 and 121 801 person-years of follow-up, respectively). We identified 15 malignancies among children with TNFi use (SIR 2.9 (1.6 to 4.9)) and 42 malignancies among children without TNFi use (SIR 2.1 (1.5 to 2.9)). The aHR was 1.58 (0.88 to 2.85) for TNFi use versus no TNFi use. In pIBD, TNFi use with thiopurine use was associated with a higher SIR (6.0 (1.2 to 17.5)) compared with TNFi use without thiopurine use (2.5 (0.7 to 6.4)). CONCLUSION: Children diagnosed with JIA, pIBD and pPsO had an increased rate of malignancy compared with the general population, but treatment with TNFi did not appear to significantly further increase the risk compared with no TNFi use. More data are needed about the long-term risks of TNFi use.

Oral glucocorticoid use and osteonecrosis in children and adults with chronic inflammatory diseases: a population-based cohort study.

OBJECTIVES: We studied oral glucocorticoids and osteonecrosis, a rare but serious bone disease, in individuals with various chronic inflammatory diseases. We hypothesised that we would find stronger associations in adults versus children and in people with autoimmune diseases. DESIGN: Retrospective cohort study. SETTING: Population-representative data (1994-2013) from general practices in the UK. PARTICIPANTS: Children and adults diagnosed with asthma; inflammatory bowel disease; juvenile, psoriatic or rheumatoid arthritis; psoriasis; or systemic lupus. EXPOSURES: Oral glucocorticoid patterns. PRIMARY AND SECONDARY OUTCOME MEASURES: Diagnosed osteonecrosis (primary) and osteonecrosis plus clinical features (eg, symptoms, pain medication, surgical repair) (secondary). Discrete time failure models estimated the adjusted hazard ratio (aHR) of incident osteonecrosis following oral glucocorticoid exposure. Hypothesis testing was one sided (with corresponding 90% CI) since glucocorticoids were unlikely protective. RESULTS: After adjusting for demographic, disease-related and health utilisation factors, glucocorticoid exposure was associated with osteonecrosis in adults (ages 18-49, aHR 2.1 (90% CI 1.5 to 2.9); ages ≥50, aHR 1.3 (90% CI 1.01 to 1.7)). However, low-dose glucocorticoids, corresponding to average doses <7.5 mg prednisolone daily and maximum doses <30 mg daily, were not associated with osteonecrosis in adults. Furthermore, even at high glucocorticoid doses, there was no evidence of increased osteonecrosis among glucocorticoid-exposed children (p=0.04 for interaction by age) (any glucocorticoid exposure, ages 2-9: aHR 1.1 (90% CI 0.7 to 1.7); ages 10-17: aHR 0.6 (90% CI 0.3 to 1.6)). Arthritis, inflammatory bowel disease and lupus were independently associated with osteonecrosis, but there was a similar dose relationship between glucocorticoids and osteonecrosis among adults with low-risk and high-risk diseases. CONCLUSIONS: Glucocorticoid use was clearly associated with osteonecrosis in a dose-related fashion in adults, especially young adults, but this risk was not detectable in children. The absolute risk of glucocorticoid-associated osteonecrosis in the general paediatric population and in adults taking low glucocorticoid doses is at most extremely small.

An Abbreviated Montreal Cognitive Assessment (MoCA) for Dementia Screening.

OBJECTIVE: The Montreal Cognitive Assessment (MoCA) is a cognitive screening instrument growing in popularity, but few studies have conducted psychometric item analyses or attempted to develop abbreviated forms. We sought to derive and validate a short-form MoCA (SF-MoCA) and compare its classification accuracy to the standard MoCA and Mini-Mental State Examination (MMSE) in mild cognitive impairment (MCI), Alzheimer disease (AD), and normal aging. METHODS: 408 subjects (MCI n = 169, AD n = 87, and normal n = 152) were randomly divided into derivation and validation samples. Item analysis in the derivation sample identified most sensitive MoCA items. Receiver Operating Characteristic (ROC) analyses were used to develop cut-off scores and evaluate the classification accuracy of the SF-MoCA, standard MoCA, and MMSE. Net Reclassification Improvement (NRI) analyses and comparison of ROC curves were used to compare classification accuracy of the three measures. RESULTS: Serial subtraction (Cramer's V = .408), delayed recall (Cramer's V = .702), and orientation items (Cramer's V = .832) were included in the SF-MoCA based on largest effect sizes in item analyses. Results revealed 72.6% classification accuracy of the SF-MoCA, compared with 71.9% for the standard MoCA and 67.4% for the MMSE. Results of NRI analyses and ROC curve comparisons revealed that classification accuracy of the SF-MoCA was comparable to the standard version and generally superior to the MMSE. CONCLUSIONS: Findings suggest the SF-MoCA could be an effective brief tool in detecting cognitive impairment.

Diagnosis, management and post-mortem findings of a human case of rabies imported into the United Kingdom from India: a case report.

BACKGROUND: Human rabies infection continues to be a significant public health burden globally, and is occasionally imported to high income settings where the Milwaukee Protocol for intensive care management has recently been employed, with limited success in improving survival. Access to molecular diagnostics, pre- and post-mortem, and documentation of pathophysiological responses while using the Milwaukee protocol, can add useful insights for the future of rabies management. CASE PRESENTATION: A 58-year-old British Asian woman was referred to a regional general hospital in the UK with hydrophobia, anxiety and confusion nine weeks after receiving a dog bite in North West India. Nuchal skin biopsy, saliva, and a skin biopsy from the site of the dog bite wound, taken on the day of admission, all demonstrated the presence of rabies virus RNA. Within 48 hours sequence analysis of viral RNA confirmed the diagnosis and demonstrated that the virus was a strain closely related to canine rabies viruses circulating in South Asia. Her condition deteriorated rapidly with increased agitation and autonomic dysfunction. She was heavily sedated and intubated on the day after admission, treated according to a modified Milwaukee protocol, and remained stable until she developed heart block and profound acidosis and died on the eighth day. Analysis of autopsy samples showed a complete absence of rabies neutralizing antibody in cerebrospinal fluid and serum, and corresponding high levels of virus antigen and nucleic acid in brain and cerebrospinal fluid. Quantitative PCR showed virus was also distributed widely in peripheral tissues despite mild or undetectable histopathological changes. Vagus nerve branches in the heart showed neuritis, a probable Negri body but no demonstrable rabies antigen. CONCLUSION: Rapid molecular diagnosis and strain typing is helpful in the management of human rabies infection. Post-mortem findings such as vagal neuritis highlight clinically important effects on the cardiovascular system which are typical for the clinical course of rabies in humans. Management guided by the Milwaukee protocol is feasible within well-resourced intensive care units, but its role in improving outcome for canine-derived rabies remains theoretical.

A robust lentiviral pseudotype neutralisation assay for in-field serosurveillance of rabies and lyssaviruses in Africa.

The inflexibility of existing serological techniques for detection of rabies in surveillance constrains the benefit to be gained from many current control strategies. We analysed 304 serum samples from Tanzanian dogs for the detection of rabies antibodies in a pseudotype assay using lentiviral vectors bearing the CVS-11 envelope glycoprotein. Compared with the widely used gold standard fluorescent antibody virus neutralisation assay, a specificity of 100% and sensitivity of 94.4% with a strong correlation of antibody titres (r=0.915) were observed with the pseudotype assay. To increase the assay's surveillance specificity in Africa we incorporated the envelope glycoprotein of local viruses, Lagos bat virus, Duvenhage virus or Mokola virus and also cloned the lacZ gene to provide a reporter element. Neutralisation assays using pseudotypes bearing these glycoproteins reveal that they provide a greater sensitivity compared to similar live virus assays and will therefore allow a more accurate determination of the distribution of these highly pathogenic infections and the threat they pose to human health. Importantly, the CVS-11 pseudotypes were highly stable during freeze-thaw cycles and storage at room temperature. These results suggest the proposed pseudotype assay is a suitable option for undertaking lyssavirus serosurveillance in areas most affected by these infections.

Oxidized low-density lipoprotein augments and 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitors limit CD40 and CD40L expression in human vascular cells.

BACKGROUND: Although CD40 signaling participates in atherosclerosis, links between lipid risk factors and this inflammatory pathway remain obscure. Cardiovascular risk reduction by 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitors (statins) may involve actions beyond lipid lowering, including reduced inflammation. Therefore, this study analyzed whether oxidized low-density lipoprotein (oxLDL) induces CD40/CD40L expression on cells implicated in atherogenesis and whether statins affect their expression in vitro as well as the expression of soluble CD40L (sCD40L) in vivo. METHODS AND RESULTS: Treatment of human vascular endothelial and smooth muscle cells and mononuclear phagocytes with oxLDL augmented the basal expression of CD40 and CD40L mRNA and protein. In contrast, cerivastatin, atorvastatin, or simvastatin concentration-dependently diminished the constitutive as well as oxLDL- or cytokine-induced expression of the receptor/ligand dyad, an effect reversed by mevalonate. Patients treated with statins had diminished sCD40L plasma levels compared with untreated control patients (8.3+/-3.1 ng/mL [n=11] versus 13.1+/-2.5 ng/mL [n=16], P<0.05), supporting the clinical relevance of the in vitro observations. Platelet-enriched plasma of mice deficient in CD40L showed markedly delayed fibrin clot formation, suggesting a role for the ligand in blood coagulation and supporting the hypothesis that statin-mediated reduction in CD40/CD40L expression might limit thrombosis. CONCLUSIONS: OxLDL may promote expression of CD40 and CD40L in human atheroma. Statins may limit the expression of the CD40 receptor/ligand dyad in two ways, directly as well as through diminished lipoprotein levels. Thus, reduced CD40 signaling may account for some of the statins' antiinflammatory action.