Synergy between type 1 fimbriae expression and C3 opsonisation increases internalisation of E. coli by human tubular epithelial cells

<p>Abstract</p> <p>Background</p> <p>Bacterial infection of the urinary tract is a common clinical problem with <it>E. coli </it>being the most common urinary pathogen. Bacterial uptake into epithelial cells is increasingly recognised as an important feature of infection. Bacterial virulence factors, especially fimbrial adhesins, have been conclusively shown to promote host cell invasion. Our recent study reported that C3 opsonisation markedly increases the ability of <it>E. coli </it>strain J96 to internalise into human proximal tubular epithelial cells via CD46, a complement regulatory protein expressed on host cell membrane. In this study, we further assessed whether C3-dependent internalisation by human tubular epithelial cells is a general feature of uropathogenic <it>E. coli </it>and investigated features of the bacterial phenotype that may account for any heterogeneity.</p> <p>Results</p> <p>In 31 clinical isolates of <it>E. coli </it>tested, C3-dependent internalisation was evident in 10 isolates. Type 1 fimbriae mediated-binding is essential for C3-dependent internalisation as shown by phenotypic association, type 1 fimbrial blockade with soluble ligand (mannose) and by assessment of a type 1 fimbrial mutant.</p> <p>Conclusion</p> <p>we propose that efficient internalisation of uropathogenic <it>E. coli </it>by the human urinary tract depends on co-operation between type 1 fimbriae-mediated adhesion and C3 receptor -ligand interaction.</p

CCL2 nitration is a negative regulator of chemokine-mediated inflammation.

Chemokines promote leukocyte recruitment during inflammation. The oxidative burst is an important effector mechanism, this leads to the generation of reactive nitrogen species (RNS), including peroxynitrite (ONOO). The current study was performed to determine the potential for nitration to alter the chemical and biological properties of the prototypical CC chemokine, CCL2. Immunofluorescence was performed to assess the presence of RNS in kidney biopsies. Co-localisation was observed between RNS-modified tyrosine residues and the chemokine CCL2 in diseased kidneys. Nitration reduced the potential of CCL2 to stimulate monocyte migration in diffusion gradient chemotaxis assays (p < 0.05). This was consistent with a trend towards reduced affinity of the nitrated chemokine for its cognate receptor CCR2b. The nitrated chemokine was unable to induce transendothelial monocyte migration in vitro and failed to promote leukocyte recruitment when added to murine air pouches (p < 0.05). This could potentially be attributed to reduced glycosaminoglycan binding ability, as surface plasmon resonance spectroscopy showed that nitration reduced heparan sulphate binding by CCL2. Importantly, intravenous administration of nitrated CCL2 also inhibited the normal recruitment of leukocytes to murine air pouches filled with unmodified CCL2. Together these data suggest that nitration of CCL2 during inflammation provides a mechanism to limit and resolve acute inflammation

Complement factor I deficiency: a potentially treatable cause of fulminant cerebral inflammation

Objective To raise awareness of complement factor I (CFI) deficiency as a potentially treatable cause of severe cerebral inflammation. Methods Case report with neuroradiology, neuropathology, and functional data describing the mutation with review of literature. Results We present a case of acute, fulminant, destructive cerebral edema in a previously well 11-year-old, demonstrating massive activation of complement pathways on neuropathology and compound heterozygote status for 2 pathogenic mutations in CFI which result in normal levels but completely abrogate function. Conclusions Our case adds to a very small number of extant reports of this phenomenon associated with a spectrum of inflammatory histopathologies including hemorrhagic leukoencephalopathy and clinical presentations resembling severe acute disseminated encephalomyelitis. CFI deficiency can result in uncontrolled activation of the complement pathways in the brain resulting in devastating cerebral inflammation. The deficit is latent, but the catastrophic dysregulation of the complement system may be the result of a C3 acute phase response. Diagnoses to date have been retrospective. Diagnosis requires a high index of suspicion and clinician awareness of the limitations of first-line clinical tests of complement activity and activation. Simple measurement of circulating CFI levels, as here, may fail to diagnose functional deficiency with absent CFI activity. These diagnostic challenges may mean that the CFI deficiency is being systematically under-recognized as a cause of fulminant cerebral inflammation. Complement inhibitory therapies (such as eculizumab) offer new potential treatment, underlining the importance of prompt recognition, and real-time whole exome sequencing may play an important future role

Comparison of the outcome of kidney transplantation after pulsatile or continuous ex vivo hypothermic machine perfusion of kidneys donated after cardiac death: analysis of kidney pairs

Background Hypothermic machine perfusion is used to improve renal perfusion and reduce the rate of early and late graft dysfunction. It has been used in our unit since 2001. It has two modes of flow: continuous or pulsatile. The aim of this study is to compare the modes of perfusion in terms of perfusion-related parameters, graft survival and estimated glomerular filtration rate. Methods All donation after cardiac death kidneys between 2002 and 2014 were reviewed. Sixty-four pairs of kidneys were identified of which one kidney underwent pulsatile and the other continuous perfusion. Machine parameters including resistance and perfusion flow index levels at 0, 1, 2, 3, 4 hours were recorded and glutathione S-transferase measured in perfusate. Delayed graft function frequency, estimated glomerular filtration rate from the 1st week of transplantation until 5th year and graft survival rates were determined. Results Machine parameters were similar at all time points. Delayed graft function frequency, estimated glomerular filtration rates and graft survival were equivalent irrespective of perfusion mode. Conclusion Pulsatile perfusion may be regarded as more physiological. However, we could not identify differences in short or long term outcomes following transplantation of kidneys from the same donor that had been perfused under pulsatile or continuous conditions

Outcomes of patients with atypical haemolytic uraemic syndrome with native and transplanted kidneys treated with eculizumab: a pooled post hoc analysis

Atypical hemolytic uremic syndrome (aHUS) often leads to end-stage renal disease (ESRD) and kidney transplantation; graft loss rates are high due to disease recurrence. A post hoc analysis of four prospective clinical trials in aHUS was performed to evaluate eculizumab, a terminal complement inhibitor, in patients with native or transplanted kidneys. The trials included 26-week treatment and extension periods. Dialysis, transplant, and graft loss were evaluated. Study endpoints included complete thrombotic microangiopathy (TMA) response, TMA event-free status, hematologic and renal parameters, and adverse events. Of 100 patients, 74 had native kidneys and 26 in the transplant subgroup had a collective history of 38 grafts. No patients lost grafts and only one with preexisting ESRD received a transplant on treatment. Efficacy endpoints were achieved similarly in both subgroups. After 26 weeks, mean absolute estimated glomerular filtration rate increased from baseline to 61 and 37 mL/min/1.73 m2 in native (n=71; P<0.0001) and transplanted kidney (n=25; P=0.0092) subgroups. Two patients (one/subgroup) developed meningococcal infections; both recovered, one continued therapy. Eculizumab was well tolerated. Eculizumab improved hematologic and renal outcomes in both subgroups. In patients with histories of multiple graft losses, eculizumab protected kidney function. (ClinicalTrials. gov numbers : NCT00844545, NCT00844844, NCT00838513, NCT00844428, NCT01193348, and NCT01194973) This article is protected by copyright. All rights reserved

A urinary microRNA panel that is an early predictive biomarker of delayed graft function following kidney transplant

Predicting immediate and subsequent graft function is important in clinical decision-making around kidney transplantation, but is difficult using available approaches. Here we have evaluated urinary microRNAs as biomarkers in this context. Profiling of 377 microRNAs in the first urine passed post-transplantation identified 6 microRNAs, confirmed to be upregulated by RT-qPCR in an expanded cohort (miR-9, -10a, -21, -29a, -221, and -429, n = 33, P < 0.05 for each). Receiver operating characteristic analysis showed Area Under the Curve 0.94 for this panel. To establish whether this early signal was sustained, miR-21 was measured daily for 5 days post-transplant, and was consistently elevated in those developing Delayed Graft Function (n = 165 samples from 33 patients, p < 0.05). The biomarker panel was then evaluated in an independent cohort, sampled at varying times in the first week post-transplantation in a separate transplant center. When considered individually, all miRs in the panel showed a trend to increase or a significant increase in those developing delayed Graft Function (miR-9: P = 0.068, mIR-10a: P = 0.397, miR-21: P = 0.003, miR-29a: P = 0.019, miR-221: P = 0.1, and miR-429: P = 0.013, n = 47) with Area Under the Curve 0.75 for the panel. In conclusion, combined measurement of six microRNAs had predictive value for delayed graft function following kidney transplantation. Introductio

A genome-wide association study of anorexia nervosa.

Anorexia nervosa (AN) is a complex and heritable eating disorder characterized by dangerously low body weight. Neither candidate gene studies nor an initial genome-wide association study (GWAS) have yielded significant and replicated results. We performed a GWAS in 2907 cases with AN from 14 countries (15 sites) and 14 860 ancestrally matched controls as part of the Genetic Consortium for AN (GCAN) and the Wellcome Trust Case Control Consortium 3 (WTCCC3). Individual association analyses were conducted in each stratum and meta-analyzed across all 15 discovery data sets. Seventy-six (72 independent) single nucleotide polymorphisms were taken forward for in silico (two data sets) or de novo (13 data sets) replication genotyping in 2677 independent AN cases and 8629 European ancestry controls along with 458 AN cases and 421 controls from Japan. The final global meta-analysis across discovery and replication data sets comprised 5551 AN cases and 21 080 controls. AN subtype analyses (1606 AN restricting; 1445 AN binge-purge) were performed. No findings reached genome-wide significance. Two intronic variants were suggestively associated: rs9839776 (P=3.01 × 10(-7)) in SOX2OT and rs17030795 (P=5.84 × 10(-6)) in PPP3CA. Two additional signals were specific to Europeans: rs1523921 (P=5.76 × 10(-)(6)) between CUL3 and FAM124B and rs1886797 (P=8.05 × 10(-)(6)) near SPATA13. Comparing discovery with replication results, 76% of the effects were in the same direction, an observation highly unlikely to be due to chance (P=4 × 10(-6)), strongly suggesting that true findings exist but our sample, the largest yet reported, was underpowered for their detection. The accrual of large genotyped AN case-control samples should be an immediate priority for the field

Type I interferon causes thrombotic microangiopathy by a dose-dependent toxic effect on the microvasculature

Many drugs have been reported to cause thrombotic microangiopathy (TMA), yet evidence supporting a direct association is often weak. In particular, TMA has been reported in association with recombinant type I interferon (IFN) therapies, with recent concern regarding the use of IFN in multiple sclerosis patients. However, a causal association has yet to be demonstrated. Here, we adopt a combined clinical and experimental approach to provide evidence of such an association between type I IFN and TMA. We show that the clinical phenotype of cases referred to a national center is uniformly consistent with a direct dose-dependent drug-induced TMA. We then show that dose-dependent microvascular disease is seen in a transgenic mouse model of IFN toxicity. This includes specific microvascular pathological changes seen in patient biopsies and is dependent on transcriptional activation of the IFN response through the type I interferon α/β receptor (IFNAR). Together our clinical and experimental findings provide evidence of a causal link between type I IFN and TMA. As such, recombinant type I IFN therapies should be stopped at the earliest stage in patients who develop this complication, with implications for risk mitigation

Efficacy and safety of baricitinib or ravulizumab in adult patients with severe COVID-19 (TACTIC-R): a randomised, parallel-arm, open-label, phase 4 trial

Background From early in the COVID-19 pandemic, evidence suggested a role for cytokine dysregulation and complement activation in severe disease. In the TACTIC-R trial, we evaluated the efficacy and safety of baricitinib, an inhibitor of Janus kinase 1 (JAK1) and JAK2, and ravulizumab, a monoclonal inhibitor of complement C5 activation, as an adjunct to standard of care for the treatment of adult patients hospitalised with COVID-19. Methods TACTIC-R was a phase 4, randomised, parallel-arm, open-label platform trial that was undertaken in the UK with urgent public health designation to assess the potential of repurposing immunosuppressants for the treatment of severe COVID-19, stratified by a risk score. Adult participants (aged ≥18 years) were enrolled from 22 hospitals across the UK. Patients with a risk score indicating a 40% risk of admission to an intensive care unit or death were randomly assigned 1:1:1 to standard of care alone, standard of care with baricitinib, or standard of care with ravulizumab. The composite primary outcome was the time from randomisation to incidence (up to and including day 14) of the first event of death, invasive mechanical ventilation, extracorporeal membrane oxygenation, cardiovascular organ support, or renal failure. The primary interim analysis was triggered when 125 patient datasets were available up to day 14 in each study group and we included in the analysis all participants who were randomly assigned. The trial was registered on ClinicalTrials.gov (NCT04390464). Findings Between May 8, 2020, and May 7, 2021, 417 participants were recruited and randomly assigned to standard of care alone (145 patients), baricitinib (137 patients), or ravulizumab (135 patients). Only 54 (39%) of 137 patients in the baricitinib group received the maximum 14-day course, whereas 132 (98%) of 135 patients in the ravulizumab group received the intended dose. The trial was stopped after the primary interim analysis on grounds of futility. The estimated hazard ratio (HR) for reaching the composite primary endpoint was 1·11 (95% CI 0·62–1·99) for patients on baricitinib compared with standard of care alone, and 1·53 (0·88–2·67) for ravulizumab compared with standard of care alone. 45 serious adverse events (21 deaths) were reported in the standard-of-care group, 57 (24 deaths) in the baricitinib group, and 60 (18 deaths) in the ravulizumab group. Interpretation Neither baricitinib nor ravulizumab, as administered in this study, was effective in reducing disease severity in patients selected for severe COVID-19. Safety was similar between treatments and standard of care. The short period of dosing with baricitinib might explain the discrepancy between our findings and those of other trials. The therapeutic potential of targeting complement C5 activation product C5a, rather than the cleavage of C5, warrants further evaluation