Driving discoveries: Connecting our fellows and students
Abstract presentation details for our series of virtual seminars.
Seminar 4: Building a community
Invited 15-minute presentations:
Short, selected abstracts:
Authors
Miguel Barroso
Affiliations
Translational and Clinical Research Institute, Faculty of Medical Sciences, Newcastle University, Central Parkway, Newcastle upon Tyne, NE1 3BZ, United Kingdom.
Abstract
Autosomal dominant polycystic kidney disease (ADPKD) is a cystic kidney disease and ciliopathy. Genetically, over 90% of cases are caused by mutations in PKD1 and PKD2. Recently, additional genes such as DNAJB11, GANAB, ALG8, ALG9 and ALG6 have been associated with phenotypes of polycystic kidney disease and polycystic liver disease.
We analysed whole genome sequencing (WGS) data from patients with renal and/or liver cysts within the Genomic England research environment of the100,000 Genomes Project. Using the tiering data, we analysed heterozygous protein-truncating variants in each of the ALG genes and used online tools to determine pathogenicity and allele frequency of these variants. We also investigated the burden of variants in the each of the ALG genes in a case population of patients with a cystic phenotype in kidney or liver (n=1486) and compared it with the burden of variants in the same genes in a control population of patients without a cystic phenotype in kidney or liver (n=28778).
We identified 7 “unsolved” patients with an ADPKD-like phenotype: in 5 of them we identified disease-causing variants in ALG8 and in 2 of them we identified disease-causing variants in ALG9.
Comparing the case and control populations: the burden of truncating alleles is higher in cases than in controls in ALG1, ALG3, ALG5, ALG8, ALG9, ALG11, ALG12 and ALG14. Applying a χ2 test (with Yates's correction) there is a relationship between the type of variant (truncating variant vs non-truncating) in ALG8 (p-value: 0.046) and being a case or a control. ALG9 (p-value: 0.078) is being close to the significant threshold p-value of 0.05. Additionally, we repeated the analysis removing the patients already solved by Genomics England and observed that this relationship is met in ALG8 (p-value: 0.01) and ALG9 (p-value: 0.008). ALG12 (p-value: 0.07) is being close to the significant threshold p-value of 0.05.
WGS allowed the identification of rare genetic variants associated with ADPKD, reinforcing the importance of analysing the pathogenic variants in ALG genes, in particular ALG8 and ALG9, when PKD1 and PKD2 genes cannot provide a genetic diagnosis of ADPKD.
Authors
Nida Ziauddeen1,2, Simon DS Fraser1,2, Nisreen A Alwan1,2,3, Ho M Yuen1, Robin F Jeffrey4, Rafaq Azad4, Dagmar Waiblinger4, Dan Mason4, John Wright4, Richard J.M. Coward5, Paul J Roderick1
Affiliations
1School of Primary Care, Population Sciences and Medical Education, Faculty of Medicine, University of Southampton, Southampton, UK; 2NIHR Applied Research Collaboration Wessex, Southampton, UK; 3NIHR Southampton Biomedical Research Centre, University of Southampton and University Hospital Southampton NHS Foundation Trust, Southampton, UK; 4Bradford Institute for Health Research, Bradford Royal Infirmary, Bradford, UK; 5Bristol Renal, Bristol Medical School, University of Bristol, Bristol, UK
Abstract
Foetal and early childhood development contributes to the risk of non-communicable diseases such as obesity and cardiovascular disease. Similarly, in utero events could affect kidney development potentially increasing risk of future chronic kidney disease (CKD). Severe CKD in adults is more common in South Asians and in lower socio-economic (SE) groups. Using data from the Born in Bradford (BiB) cohort, we have shown that South Asian babies have smaller kidneys compared with white British babies. We aimed to investigate whether kidney size at birth is associated with markers of kidney function at 7-11 years and if there were ethnic and SE differences.
Foetal kidney dimensions were measured using ultrasound scans at 34 weeks gestation and used to derive kidney volume (cm3) in 1802 participants in the BiB cohort. At 7-11 years, we assessed estimated Glomerular Filtration Rate (eGFR) using serum creatinine and cystatin C concentrations with the Schwartz combined formula1 and Zappitelli cystatin C formula2 in 457 (25%) of the 1802 participants. Child height was standardised to time of sample using centile charts. Linear regression was used to examine the association between renal volume and eGFR adjusted for ethnicity, maternal characteristics at pregnancy (age, educational attainment, housing tenure, employment status, BMI, height, alcohol consumption, smoking, gestational diabetes), birthweight, gestational age at birth, gender, child BMI and age at kidney function measurement.
Mean eGFR was 91.9 (standard deviation (SD) 8.8) using Schwartz and 95.3 (SD 10.2) using Zappitelli. Renal volume was positively associated with eGFR calculated using Schwartz (0.62ml/min diff per unit increase in volume, 95% CI 0.24 to 1.00) and Zappitelli (0.71, 95% CI 0.31 to 1.12). There was no association between ethnicity or SE status (using maternal education) and eGFR at 7-11 years.
Foetal renal volume was associated with small increases in eGFR in early childhood but there was no evidence of ethnic or SE differences in kidney function. To increase power, analysis of 1275 BiB participants who provided blood for renal assessment but did not have foetal renal ultrasounds, to investigate ethnicity and SES will be presented. Longitudinal follow-up is required to investigate the relationship between renal volume and markers of kidney function as children go through puberty.
Authors
Robert J. P. Pope, Gavin I. Welsh, Craig A. McArdle and Richard J. M. Coward
Affiliations
Bristol Renal, University of Bristol
Abstract
Introduction
The podocyte is a key cell in maintaining kidney function as evidenced by more than 50 human genetic mutations described that code for proteins with podocyte-specific effects, all of which result in albuminuria, a risk factor for renal failure. Podocytes are insulin sensitive, and the importance of this has been demonstrated through the generation of a mouse lacking podocyte insulin receptors, that develops significant kidney disease1. Furthermore, recent studies of large diabetic patient cohorts have revealed that individuals with severe insulin resistance are more likely to develop kidney disease and progress to end stage renal failure.
Given the importance of insulin sensitivity in kidney health, we have developed high throughput, semiautomated assays to assess insulin signalling in human podocytes. These assays have been used to screen for compounds that can modulate insulin sensitivity as such molecules could have huge therapeutic benefits.
Methods
Podocytes were cultured in “non-diabetic” (RPMI) and “diabetic” (high-glucose, high insulin, high pro-inflammatory cytokines; IL-6 and TNF-alpha) environments. We performed a primary screen assessing each compound, 1,280 in total, in the Sigma LOPAC under four different conditions: non-diabetic, non-diabetic + insulin stimulation, diabetic and diabetic + insulin stimulation. IN Cell 2200 was used to measure podocyte activity (phosphorylation of Ak strain transforming at serine 473 (pAkt 473) and nuclear to cytoplasmic translocation of the transcription factor FOXO1 via a Clover reporter construct), within the same cell. Cell number was also considered in every captured field meaning that three parameters were measured in four conditions for each library compound.
Results
The screen revealed a number of “hits” as defined by a set percentage (between 20 and 40% dependent on measure) increase or decrease, above the test plate mean. Several previously described compounds were identified as modulating insulin sensitivity within our screen conditions, helping to validate that the assays are both highly robust and sensitive. Several novel compound hits were also identified and are under further validation and investigation.
Discussion
The LOPAC library includes 1,280 pharmacologically active compounds and covers most drug classes currently described with many in clinical use. Using the library in this unbiased screen of human podocytes has identified a number of previously undescribed modulators of insulin signalling.
Authors
Katherine Clark, Hayley Martin, Henry Kibble, Kieran Palmer, Emmanouil Kountouris, Amelia Holloway, Anna Naito, Kate Bramham.
Affiliations
King’s College Hospital NHS Foundation Trust, London, United Kingdom; King’s College London School of Life Course Sciences, London, United Kingdom .
Abstract
Pregnancies affords an opportunity to diagnose asymptomatic chronic kidney disease (CKD) which may be masked by gestational change in serum creatinine. In addition, kidney injury may occur in pregnancy leading to CKD. Postpartum assessment enables detection of on-going kidney dysfunction. We aimed to determine the prevalence of kidney disease in postpartum women with chronic hypertension, pregnancy induced hypertension (PIH) or preeclampsia in a previous or current pregnancy.
Women with singleton pregnancies with estimated GFR (CKD-EPI) below 90mls/min/1.732m and/or proteinuria at six-weeks after delivery were offered specialist renal midwifery clinic follow-up at approximately 6 months postpartum.
175 (42.4%) out of 413 women attended their appointment at a median of 185 days (IQR 246.25) after delivery. Over half of the women with proteinuria and/or reduced eGFR at six weeks postpartum had CKD (97/175 (55.2%)): 52/175 (29.7%) Stage 1 (52/52 (100%) A2); 43/175 (18.3%) Stage 2 (18/43 (41.9%) A1, 10/43 (23.3%) A2, 15/43 (34.9%) A3); and 2/175 (1.1%) had CKD Stage 3 A2. There was no relationship between hypertensive diagnosis or its severity in pregnancy and postpartum CKD.
Postpartum assessment may afford an opportunity to detect early CKD, in all women with previous hypertension in pregnancy regardless of hypertensive diagnosis. Assessment of renal function pre-conception or in early pregnancy according to creatinine centile could provide a useful baseline to facilitate diagnosis and enable risk stratification for postpartum follow up. More work is needed to engage women postpartum and facilitate testing.
Authors
Anam Asad 1, Daniel S March 1,2, James O Burton 1,2,3
Affiliations
1 University of Leicester Department of Cardiovascular Sciences, Leicester; 2 John Walls Renal Unit , Leicester; 3 School of Sport Exercise and Health Sciences Loughborough University, Loughborough
Abstract
Background: Acute kidney injury (AKI) is common, occurring in 7-18% of patients admitted to hospital. AKI has high levels of associated morbidity and mortality and frequently results in chronic kidney disease (CKD). There are currently no effective therapies promoting renal recovery post-AKI hence reducing the incidence of long-term renal damage. Animal studies demonstrate the potential for reno-protective effects of physical activity. There are no human studies corroborating these data.
Aim: This study aimed to investigate associations between physical activity levels and recovery in renal function, following stage 3 AKI.
Methods: This study enrolled 41 hospitalised participants with stage 3 AKI. Participants completed two questionnaires: the Duke Activity Status Index (DASI), and the General Practice Activity Questionnaire (GPPAQ). Both questionnaires provided a subjective measure of physical activity levels. Eleven participants also wore an accelerometer for one week following discharge and 6-months post discharge to ascertain an average daily step count. Renal function (estimated by creatinine) was extracted from medical records. Creatinine 12-months prior to AKI provided a baseline measurement and further values were taken during hospitalisation, at discharge and 1, 3 and 6-months post-discharge to allow assessment of renal recovery.
Results: The average baseline eGFR (CKD-EPI) and median creatinine was 71±20ml/min/1.73m2 and 85 (49) μmol/L . During hospitalisation, all participants experienced a threefold increase in creatinine, 436(265) μmol/L, consistent with stage 3 AKI. Patients’ renal function continued to recover 3 months post discharge, with the greatest recovery occurring during hospitalisation, prior to discharge. A higher daily step count immediately following discharge was significantly associated with better baseline renal function (r=-0.6636, p=0.03). At 3 and 6-months following discharge, increased activity levels shown through better DASI scores, were associated with improved renal recovery (r= -0.6109, p= 0.0805; r= -0.6394, p= 0.0186). Those with greater step counts at 6-months also displayed better renal recovery at 6 months (r=-0.600, p=0.2080).
Conclusions: Individuals with higher activity levels had better renal function at 3 and 6- months following stage 3 AKI. These data suggest that higher levels of physical activity may be protective and promote renal recovery. Future large prospective trials should aim to trial the effect of an exercise programme on renal function in AKI patients.
Authors
Isaac Chung1, Mukunthan Srikantharajah, Robin Ramphul, Juan Carlos Kaski, Debasish Banerjee1,2
Affiliations
1Renal and Transplantation Unit, St George’s University Hospitals NHS Foundation Trust; 2Cardiology Clinical Academic Group, Molecular and Clinical Sciences Research Institute, St George’s, University of London
Abstract
Chronic kidney disease patients are at higher risk of cardiovascular disease and mortality, which partially improves with kidney transplant but remain elevated compared to general population. This study aims to investigate the progression of vascular structure and function in CKD and transplant patients.
Vascular markers including carotid intima-medial thickness [c-IMT], ankle-brachial pressure index [ABPI], brachial artery flow mediated dilatation [ba-FMD], brachial artery nitroglycerin mediated dilation [ba-NMD], augmentation index [AIx] and carotid-femoral pulse-wave velocity [cf-PWV] were measured at baseline and at 3-year follow-up.
There was no significant difference between CKD and kidney transplant patients at baseline in ba-FMD (6.67 ± 1.47 vs 5.75 ± 0.80; p-value=0.91), ba-NMD (11.1 ± 5.52 vs 11.95 ± 5.25; p-value=0.76), c-IMT (0.67 ± 0.15 vs 0.58 ± 0.80; p-value=0.20), AIx (24.67 ± 7.58 vs 17.50 ± 14.60; p-value=0.21), and cf-PWV (9.13 ± 2.26 vs 8.50 ± 1.84; p-value=0.58).
In the CKD cohort, there was no difference between baseline and follow-up in ba-FMD (4.70 ± 2.76 vs 3.57 ± 2.41, p-value=0.47), ba-NMD (11.06 ± 5.51 vs 11.61 ± 4.37, p-value=0.85), c-IMT (0.67 ± 0.15 vs 0.63 ± 0.11, p-value=0.68), Aix (25.60 ± 8.09 vs 33.00 ± 15.70, p-value=0.72), and cf-PWV (9.13 ± 2.27 vs 9.81 ± 1.42, p-value=0.55). In the transplant cohort, there was no difference between baseline and follow-up in ba-FMD (4.90 ± 4.03 vs 5.58 ± 2.69, p- value =0.66), c-IMT (0.58 ± 0.08 vs 0.57 ± 0.10, p-value =0.97), Aix (17.50 ± 14.07 vs 23.20 ± 19.11, p-value=0.46), and cf-PWV (8.50 ± 1.84 vs 8.92 ± 2.68, p-value=0.69). Ba-NMD worsened in 3-years follow-up compared to baseline in the transplant cohort (11.94 ± 5.25 vs 16.65 ± 3.21, p-value=0.03).
CKD and kidney transplant patients have similar measures of vascular structure and function at baseline; after 3-years, CKD patients remain similar to baseline in vascular health, while renal transplant patients remain similar to baseline apart from decreased Ba-NMD when compared to baseline.
Authors
Rebecca Preston1, Qing-jun Meng1, Rachel Lennon1,2
Affiliations
1Wellcome Centre for Cell-Matrix Research, Division of Cell-Matrix Biology and Regenerative Medicine, School of Biological Sciences, Faculty of Biology Medicine and Health, The University of Manchester, UK; 2Department of Paediatric Nephrology, Royal Manchester Children’s Hospital, Manchester University Hospitals NHS Foundation Trust, Manchester Academic Health Science Centre, Manchester, UK.
Abstract
The circadian network in mammals is responsible for the temporal coordination of numerous physiological processes that are necessary for homeostasis. Peripheral tissues demonstrate circadian rhythmicity and dysfunction of core clock components has been implicated in the pathogenesis of many disease processes, including renal tubular dysfunction. However, the role of the circadian clock in the regulation of glomerular function is lacking.
Glomerular disease is characterised by pathological proteinuria, which along with the rate of glomerular filtration, display robust circadian oscillation. It is therefore likely that an intra-glomerular clock is responsible for circadian oscillation in glomerular function. Using real-time bioluminescence recordings from circadian reporter mice and human podocytes transfected with clock reporters, we demonstrate robust autonomous circadian oscillation in glomeruli and live cultured podocyte cells for the first time.
To confirm the presence of intra-glomerular clock machinery, we employed a mouse model expressing a knock-in fluorescent fusion of endogenous BMAL1, a core component of the molecular clock. Quantitative live imaging revealed the circadian expression and nuclear-cytoplasmic mobility of endogenous BMAL1, confirming high molecular abundance in glomeruli and podocyte cells. Global patterns of glomerular circadian gene expression were defined using transcriptional profiling of glomeruli isolated from wildtype mice over a circadian time-course.
Transcriptional profiling identified 260 glomerular rhythmic genes under circadian control. Subsequent bioinformatic analysis revealed genes and pathways implicated in glomerular physiology and pathology, including regulation of the extracellular matrix. Thus, here we confirm the presence of intra-glomerular clock machinery and have identified the first glomerular circadian transcriptome. Structural changes in the glomerular basement membrane, a specialised extracellular matrix which crucially controls glomerular filtration, is the hallmark of glomerular disease.
Establishing a temporally resolved glomerular matrisome may provide a useful tool for studying the two-way interactions between the extracellular matrix and the intracellular time-keeping mechanisms in this critical niche tissue. These results provide a firm basis for future studies that aim to elucidate the functional implication and therapeutic potential of circadian rhythm in glomerular health and disease.
Authors
R Harwood, T Borges Da Silva, A Muniz Garcia, M Mousavinejad, S Kenny, B Wilm, P Murray
Affiliations
Institute of Systems, Molecular and Integrative Biology, University of Liverpool, UK.
Abstract
Regenerative medicine therapies have shown exciting potential for the amelioration of acute kidney injury (AKI). Pre-clinical results have not been replicated in clinical trials and safety concerns have been raised, leading to questions about the best dose, route of administration of therapies and timing of administration of therapies. We aimed to assess the impact of these variables in pre-clinical models of AKI to determine the optimal method of administering human umbilical cord derived mesenchymal stromal cells (hUC-MSCs) and their derived extra-cellular vesicles (hUC-ECVs).
We performed a bilateral renal ischaemia reperfusion injury (r-IRI) to BALB/c mice. Animals received hUC-MSCs or hUC-ECVs as therapies and phosphate buffered saline (PBS) or human dermal fibroblast derived extracellular vesicles (hDF-ECVs) as control. Therapies were given by intracardiac or intravenous injection, at different doses (hUC-MSCs: 5x105 and 2.5x105, hUC-ECVs: 1x107, 1x108, 2x108 and 1x109) and at different times. Kidney function was measured by transdermal glomerular filtration rate (GFR) on days 1 and 3. Serum and kidneys were sampled on day 3, the endpoint of the experiment. Results were analysed by a two-way mixed effects model with Dunnett’s correction for multiple comparisons or a one-way ANOVA with Tukey correction. Significance is taken at p <0.05.
We found that neither hUC-MSCs nor hUC-ECVs showed consistent evidence of efficacy after bilateral r-IRI (Figure 1.1). Animals receiving 5x105 hUC-MSCs by intracardiac injection had a significantly higher mortality rate than animals receiving it intravenously (4/6 vs 0/6 (p 0.01)). The same effect was not seen after intracardiac injection of hUC-ECVs or PBS. Altering the dose of hUC-ECVs and hUC-MSCs did not improve the kidney function of animals compared to those receiving PBS and neither did additional hUC-MSCs or hUC-ECVs 24 hours after injury.
These rigorously performed and repeated experiments cast doubt on the potential for regenerative medicine therapies to be efficacious at treating severe AKI after rIRI and demonstrate the need for animal experiments to be reported completely and rigorously. Further work using different strains of mice may enable the discrepancies between this work and the published literature to be better understood.
Seminar 3: Accelerating discovery
Invited 15-minute presentations:
Authors
Jane E. Carré, Patricia E. Russell, Charles Affourtit
Affiliations
School of Biomedical Sciences, Faculty of Health, University of Plymouth, PL6 8BU, Devon UK
Abstract
A significant debilitating factor reported by patients with chronic kidney disease is the muscle weakness and wasting (sarcopenia) By contributing to frailty, sarcopenia contributes to patient morbidity and mortality, and is a significant predictor of transplant success. Wasting occurs as muscle protein balance shifts towards net degradation, accompanying early-onset insensitivity to the muscle anabolic hormone insulin. A bioenergetic component to sarcopenia is implicated since mitochondrial changes in muscle of CKD patients are consistently reported. Importantly, it remains unclear whether these mitochondrial changes represent bioenergetic dysfunction or occur as an adaptation to altered energy demand.
We address this issue using advanced extracellular flux analysis to assess how effectors of atrophy impact ATP supply and demand processes in differentiated L6 myotube models. We show that the atrophy-inducing glucocorticoid dexamethasone boosts muscle energy (ATP) cost of protein turnover, enhancing proteasomal degradation and driving energy metabolism through increased ATP demand. An observed lack of appropriate biogenesis response could lead to subsequent mitochondrial insufficiency in the face of this increased bioenergetic load. On the other hand, myotube exposure to the uraemic toxin indoxyl sulfate has little, or even a suppressive, effect on ATP demand flux but confers resistance to insulin (in terms of glucose uptake, energy cost of protein turnover, and glycolytic ATP supply) and decreases protein synthesis. Increased reliance of myotubes on mitochondrial energy supply potentially explains previous reports of indoxyl sulfate-enhanced ROS production.
This ongoing work highlights that muscle weakness and wasting in CKD likely occurs by multiple mechanisms. Effects occur on both ATP supply and ATP demand, but mitochondrial/bioenergetic changes do not necessarily reflect mitochondrial dysfunction. With growing interest in therapeutic approaches that enhance or develop the mainstay nutritional and exercise therapies for sarcopenia in CKD, there is a need understand the interaction between ATP supply and demand to determine the potential and focus for therapeutic bioenergetic regulation. Notably, inappropriately targeting muscle bioenergetics in uraemic sarcopenia has potential to worsen bioenergetic imbalance and exacerbate the condition.
Authors
Rhys D R Evans1, Marilina Antonelou1, Sanchutha Sathiananthamoorthy1, Marilena Rega2, Scott Henderson1, Lourdes Ceron-Gutierrez3, Gabriela Barcenas-Morales4, Christoph A. Müller5,6, Rainer Doffinger3,7, Stephen B Walsh1, Alan D Salama1
Affiliations
1Department of Renal Medicine, University College London, Royal Free Hospital, London, UK; 2Institute of Nuclear Medicine, University College London, University College London Hospital, London, UK; 3Department of Clinical Biochemistry and Immunology, Addenbrookes's Hospital, Cambridge, UK; 4Laboratorio de Inmunologia, FES-Cuautitlan, UNAM, Mexico; 5Department of Radiology, Medical Physics, Medical Center University of Freiburg, Faculty of Medicine, University of Freiburg, 79106 Freiburg, Germany; 6German Center for Cancer Research (DKFZ), 69120 Heidelberg, Germany;7National Institute of Health Research (NIHR), Cambridge Biomedical Research Centre, Cambridge, UK
Abstract
Increased extracellular sodium activates Th17 cells, which provide protection from bacterial and fungal infections. Whilst high salt diets have been shown to worsen autoimmune disease, the immunological consequences of clinical salt depletion are unknown. Here we investigate immunity in patients with inherited salt-losing tubulopathies (SLT). Forty-seven genotyped SLT patients (with Bartter, Gitelman, or EAST Syndromes) are recruited. Clinical features of dysregulated immunity are recorded with a standardised questionnaire and immunological investigations of IL-17 responsiveness undertaken. The effect of altering extracellular ionic concentrations on immune responses is then assessed. Patients are hypokalaemic and hypomagnesaemic, with reduced interstitial sodium stores determined by 23Na-magnetic resonance imaging. SLT patients report increased mucosal infections and allergic disease compared to age-matched controls. Aligned with their clinical phenotype, SLT patients have an increased ratio of Th2:Th17 cells. SLT Th17 and Tc17 polarisation is reduced in vitro, yet STAT1 and STAT3 phosphorylation and calcium flux following T cell activation are unaffected. In control cells, the addition of extracellular sodium (+40mM), potassium (+2mM), or magnesium (+1mM) reduces Th2:Th17 ratio and augments Th17 polarisation. Our results thus show that the ionic environment typical in SLT impairs IL-17 immunity, but the intracellular pathways that mediate salt-driven Th17 polarisation are intact and in vitro IL-17 responses can be reinvigorated by increasing extracellular sodium concentration. Whether better correction of extracellular ions can rescue the immunophenotype in vivo in SLT patients remains unknown.
Authors
Eoin D O’Sullivan1, Katie J Mylonas1, Rachel Bell1, Cyril Carvalho1, David P Baird1, Marie Docherty1, Carolynn Cairns1, Kevin M Gallagher1, Ross Campbell1, Alexander Laird1, Neil C Henderson1, Tamir Chandra1, Kristina Kirschner2, Bryan Conway1, Gry H. Dihazi7, Michael Zeisberg3, Laura Denby1, Jeremy Hughes1, Hassan Dihazi3, David A Ferenbach1.
Affiliations
1 University of Edinburgh, Edinburgh, UK. 2 University of Glasgow, Glasgow, UK. 3 University Medical Centre Göttingen, Göttingen, Germany
Abstract
Progressive fibrosis and maladaptive organ repair result in significant morbidity and millions of premature deaths annually. Senescent cells accumulate with ageing and after injury and are implicated in organ fibrosis, but the mechanisms by which senescence influences repair are poorly understood. Using a relevant murine model of injury and repair we show that obstructive injury generates senescent epithelia which persist after resolution of the original injury, promote ongoing fibrosis and impede adaptive repair. We validate these findings in humans, showing that senescence and fibrosis persist after relieved renal obstruction. We next characterise senescent epithelia in murine renal injury using single cell RNA-Seq. We extend our classification to human kidney and liver disease and identify conserved pro-fibrotic proteins which we validate in vitro and in human disease. We demonstrate a link between senescent epithelia and fibrosis by showing one such molecule, Protein Disulfide Isomerase Family A Member 3 (PDIA3), is essential for TGF-beta mediated fibroblast activation. Inhibition of PDIA3 in vivo significantly reduces kidney fibrosis after injury and is a new potential therapeutic pathway. Analysis of the signalling pathways of senescent epithelia connects senescence to organ fibrosis, permitting rational design of anti-fibrotic therapies.
Short, selected presentations:
Authors
Amelia RMJ Abbott1, Dr Thomas J Wilkinson2,3, Dr Emma L Watson2,4, Professor Alice C Smith2,5 & Dr Luke A Baker2,5.
Affiliations
1 College of Life Sciences, University of Leicester, University Rd, Leicester, LE1 7RH; 2 Leicester Kidney Lifestyle Team, Department of Health Sciences, College of Life Sciences, University of Leicester, UK; 3 NIHR Applied Research Collaboration East Midlands, Leicester Diabetes Centre, UK; 4 Department of Cardiovascular Sciences, University of Leicester, UK; 5 Leicester Biomedical Research Centre, Leicester, UK.
Abstract
Skeletal muscle wasting and dysfunction, termed uraemic sarcopenia, is a debilitating co-morbidity for those suffering from chronic kidney disease (CKD), contributing to a reduced quality of life and elevated risk of mortality. Mechanisms for this dysfunction have been linked to systemic inflammation, however, specific mechanistic targets are yet to be isolated. Chemerin, a novel adipokine, is known to play a role in inflammatory processes, but its potential role in uraemic sarcopenia and muscle physiology more widely is yet to be fully explored.
Our aims were: 1) To investigate the relationship between chemerin and skeletal muscle strength and function; and 2) to investigate the effect of chemerin on myotubes to develop early rationale for its use as a therapeutic target.
In-vivo: Blood and urine samples from 129 CKD patients and 40 controls were analysed by ELISA to quantify chemerin concentrations. These were correlated against eGFR, muscle strength, measured as handgrip strength, and muscle function, measured using a sit-to-stand-60 test. In-vitro: Skeletal muscle biopsies from 4 CKD patients and 6 controls were cultured and exposed to chemerin (12.5nM, 50nM). Analysis was conducted to quantify the mRNA gene expression of pro-inflammatory indicators in response to differing chemerin concentrations. Immunocytochemistry analysis was conducted to determine the effect of chemerin on myotube morphology as an indicator of muscle mass and maturity.
Chemerin levels were elevated in CKD patients (urine: p=0.0292, plasma: p<0.001), negatively correlated with eGFR (p<0.001) and correlated with worsening muscle function (p=0.026). In-vitro, chemerin induced increases in pro-inflammatory gene expression in a dose dependent fashion, with increases in IL-6 and TNFα in response to 12.5nM (IL-6: p=0.0182, TNFα: p=0.0297) and 50nM (IL-6: p=0.0076, TNFα: p=0.0051) in comparison with control conditions. No significant differences were noted in myotube morphology in response to either concentration.
To conclude, this preliminary data suggests that chemerin may play a role in the development of uraemic sarcopenia, through inflammatory related mechanisms. Future work will continue developing this hypothesis and uncover the specific mechanisms by which chemerin influences skeletal muscle inflammation and how that relates to uraemic sarcopenia, to uncover potential therapeutic opportunities for the benefit of those suffering from CKD.
Authors
Katie J Mylonas, Ross A Campbell, Jeremy Hughes and David A Ferenbach.
Affiliations
Centre for Inflammation Research, Queens Medical Research Institute, University of Edinburgh, 47 Little France Crescent, Edinburgh EH16 4TJ, Scotland, United Kingdom.
Abstract
Senescent cells (SCs) accumulate in the kidney with age/injury. They are metabolically active, promoting inflammation/fibrosis via release of senescence associated secretory phenotype (SASP) cytokines. We have shown that both natural ageing and sublethal total body irradiation (TBI) induces renal epithelial senescence, with administration of the senolytic drug ABT- 263 prior to renal ischaemia reperfusion injury (IRI) being protective; reducing tissue loss, fibrosis and inflammation, whilst promoting structural integrity/regeneration.
I hypothesise that SCs compromise renal repair/regeneration after IRI in mice by driving excessive monocyte/macrophage recruitment and polarisation towards pro-inflammatory, rather than reparative, phenotypes.
In order to test this in vitro, human proximal tubular epithelial cells (PTECs) were irradiated with 10 gy radiation, to induce senescence. Senescent PTECs produced significantly increased levels of monocyte chemoattractant protein (MCP)-1 (5x fold change vs control, p<0.01), indicating that epithelial senescence drives monocyte recruitment. Macrophages were cultured from healthy human volunteer blood and exposed to the supernatant (conditioned medium) of senescent or control PTECs. Markers of pro-inflammatory macrophages including CD80 (x2 fold increase, p<0.05) and IL-1B (x 3 fold increase, p<0.05) were significantly increased after exposure to senescent conditioned medium vs control.
TBI was carried out to produce premature ageing/senescence in murine kidneys in vivo. Mice were subsequently treated with senolytic or vehicle before IRI. One day post-IRI, macrophages were depleted using clodronate liposomes. Flow cytometry at d7 and d35 post-IRI indicated that the presence of SCs drove persistent monocyte/macrophage recruitment (CD45+F480+Ly6C+ cells). Treatment with ABT-263 and/or clodronate liposomes significantly decreased CD45+ leukocytes and F480lowCD80+ monocyte-derived inflammatory macrophages recruitment to the kidney by d35. This correlated with an increase in kidney mass in ABT-263 and/or clodronate-treated mice indicating better structural integrity of the kidney in the absence of SCs and/or excessive monocyte/macrophage recruitment. qPCR of the renal tissue indicated less Mcp-1 expression with ABT-263 and/or clodronate vs complete untreated, as well as less senescence and fibrosis.
Collectively, these results support the hypothesis that senescence drives pathogenic monocyte/macrophage recruitment and a proinflammatory, profibrotic phenotype in the kidney. Importantly, selective targeting of either SCs or macrophages reduced renal fibrosis and thus represent candidates for future clinical translation.
Authors
Pinyuan Tian1, Mychel Raony Paiva Teixeira Morais1,Craig Lawless1, Syed Murtaza-Baker2, Louise Hopkinson1, Steven Woods1, Aleksandr Mironov1, David Andrew Long2, Daniel Gale3, Telma Maria Tenorio Zorn4, Susan Kimber1, Roy Zent5, Rachel Lennon*1.
Affiliations
1Faculty of Biology, Medicine and Health, University of Manchester, Manchester, UK, 2Developmental Biology and Cancer Programme, UCL Great Ormond Street Institute of Child Health, London, UK, 3Centre for Nephrology, University College London, London, UK, 4Institute of Biomedical Sciences, University of São Paulo, São Paulo, Brazil, 5Department of Medicine, Vanderbilt University Medical Center, Nashville, USA
Abstract
Basement membranes (BMs) are condensed sheets of extracellular beneath continuous layers of cells matrix composed primarily of type IV collagen and laminins. In the kidney, the glomerular basement membrane (GBM) between the podocytes and the glomerular endothelial cells is a critical component of the glomerular filtration barrier. Previous studies revealed that the GBM composition undergoes profound changes throughout kidney development, but little is known about mechanisms that regulate BM assembly and isoform transitions due to lack of proper models. We investigated the dynamics of BM assembly in human pluripotent stem cell derived kidney organoids. Whole mount immunofluorescence demonstrated that kidney organoids differentiate into glomerular structures containing podocytes (WT1+, NPHS1+), proximal tubule (LTL+), distal tubule (ECAD+), and endothelial cells (CD31+). Notably, immunostaining of the key BM markers showed that organoids form BM during differentiation, and more importantly, recapitulate a sequence of assembly events with initial deposition of laminin followed by incorporation of type IV collagen, nidogen and perlecan. Single-cell RNA sequencing analysis from day 25 kidney organoids (Combes et al., 2019) confirmed NPHS2+/PODXL+ podocytes were the main source of COL4A3, COL4A4. These findings fit with the current understanding of kidney development in vivo and demonstrate that kidney organoids can efficiently recapitulate the spatiotemporal emergence of GBM components during glomerulogenesis.
A scaffold of matrix proteins supports the cells of the glomerular filters. The glomerular cells produce the proteins that make up the scaffold, however we do not fully understand how this process is regulated or altered in disease. Here we use a new model called kidney organoids, which are an aggregation of human cells cultured in a dish to mimic kidney growth in the body. We investigate changes in the molecules that make up the scaffold when the organoids grow. This study has great potential to gain insight into the mechanism, and find new targets and treatment for glomerular disease.
Authors
E Bowen, J Hurcombe, F Barrington, L Farmer, M Saleem and R Coward.
Affiliations
University of Bristol.
Abstract
Haemolytic uraemic syndrome (HUS) is a thrombotic microangiopathy (TMA) that has a predilection to present in the kidney. It is a triad of a microangiopathic haemolytic anaemia, thrombocytopenia and acute kidney injury. In 90% of cases, HUS follows gastroenteritis secondary to infection with Shiga toxin (Stx) producing enteric pathogens such as Escherichia coli. Stx HUS is the leading cause of acute kidney injury in children with an associated mortality of 5%. The precise pathophysiological mechanisms following Stx infection leading to TMA are poorly understood.
The objective of our work was to determine whether Stx acts via the podocyte globotriaosylceramide (Gb3) receptor to cause HUS. To illustrate that Gb3 is the receptor for Stx in-vivo, we generated a constitutive whole body Gb3 knockout mouse (Gb3KO). These mice were given intraperitoneal (IP) Stx alongside wild type control mice. Gb3KO mice were completely protected from the cytotoxic effects of Stx even at x400 the lethal dose. Gb3KO mice showed no changes in their creatinine, urine ACR or renal histology. All wild type mice died within 4 days of receiving Stx from extra-glomerular dehydration effects as demonstrated by ATN on renal histology.
We then generated an inducible podocyte expressing Gb3 mouse on a Gb3 null background (PodGb3). This mouse only expresses Gb3 in its podocyte in the presence of doxycycline due to tetracycline-controlled transcription of Gb3 synthase; allowing specific study of the role of the podocyte in HUS. These mice were given IP Stx alongside age-matched controls. PodGb3 mice developed HUS (thrombocytopenia, haemolytic anaemia and uraemia p<0.05) at day 10 following IP Stx. Renal histology demonstrated glomerular TMA; with intracapillary thrombus formation seen on electron microscopy. Immunofluorescence demonstrated an increase in glomerular fibrinogen deposition and C3b vs. controls. Additionally, glomerular expression of complement regulator Factor H, was significantly reduced in PodGb3 mice, rendering them more susceptible to complement attack.
These data support the role of the podocyte as a key initiator in Stx HUS. This could explain why the glomerulus is the prime target of systemic Stx driven infections and lead to the development of novel therapies for this devastating disease.
Authors
Aneesha Bhandari1, Katherine Bull1,2, Zouchun Shang3, Yanan Xing3, Yanru An3, Tanya Crockford1, Richard Cornall1,2
Affiliations
1Nuffield Department of Medicine, University of Oxford, 2Oxford Kidney Unit, Oxford University Hospitals Trust, 3Beijing Genomics Institute, Qingdao
Abstract
Lupus nephritis (LN) is characterised by renal immune-complex (IC) deposition, yet how these deposits recruit inflammatory mediators and the signalling between resident and recruited cells remains unclear. To develop targeted therapies, we need to understand cellular responses within glomeruli. Interpretation of renal single cell RNA sequencing (scRNA-Seq) is limited by relative depletion of glomerular cells and interglomerular pathological heterogeneity. We applied cellular resolution spatial transcriptomics to a murine model of LN to interrogate in situ the response to glomerular IC deposition. #
6-week-old female BALB/c mice were treated for 8 weeks with topical Imiquimod (IMQ) or Vaseline control. Kidneys were processed for scRNA-seq (10x Genomics) or spatial transcriptomics (BGI Stereo-seq), generating 10mm2 of 600nm resolution data, with a median 1021 transcripts per 14um bin.
Topical treatment with toll-like receptor 7 agonist IMQ for 8 weeks results in increased numbers of activated B and T cells, and glomerular mesangial expansion and endothelial proliferation consistent with Class-II LN, representing a model of early active LN.
Sequential clustering of scRNA-Seq data led to the identification of renal and immune cell clusters, however some key glomerular populations remained scarce. B and T-cells and myeloid cells were predominantly from IMQ mice and included adherent and infiltrating neutrophils, and resident macrophages expressing C1q. Clustering spatial data identified key renal populations including glomerular cells. Within the podocyte spatial cluster, the complement C3 convertase regulator Decay-accelerating factor (DAF) was differentially expressed (DE) with IMQ treatment, and comparison with single nuclei RNA-Seq confirmed podocyte specific upregulation. Systemic DAF deficiency is known to exacerbate MRL/Lpr LN, and podocyte specific loss of DAF exacerbates FSGS. We found that podocyte DAF expression co-localised with signals of activated B cell glomerular infiltration.
By mapping single cell data onto spatial transcriptomics, we can visualise infiltrating cells and stromal cells in context and begin to dissect cellular crosstalk in glomerular disease. For example, our data show that in LN podocyte DAF expression is be upregulated, which may be in response to B cell signalling, as a protective response. Application of this methodology to human kidney tissue has potential to transform renal histology.
Authors
Melanie MY Chan1, Omid Sadeghi-Alavijeh1, Horia C Stanescu1, Catalin D Voinescu1, Glenda M Beaman2,3, Marcin Zaniew4, Stefanie Weber5, Alina C Hilger6,7, William G Newman2,3, Adrian S Woolf8,9, John O Connolly1,10, Dan Wood10, Alexander Stuckey11, Athanasios Kousathanas11, Genomics England Research Consortium11, Robert Kleta1,12, Detlef Bockenhauer1,12, Adam P Levine1,13, and Daniel P Gale1
Affiliations
1Department of Renal Medicine, University College London, London, UK. 2Manchester Centre for Genomic Medicine, Manchester University NHS Foundation Trust, Manchester, UK. 3Evolution and Genomic Sciences, School of Biological Sciences, University of Manchester, Manchester, UK. 4Department of Pediatrics, University of Zielona Góra, Poland. 5Department of Pediatric Nephrology, University of Marburg, Marburg, Germany. 6Children’s Hospital, University of Bonn, Bonn, Germany. 7Institute of Human Genetics, University of Bonn, Bonn, Germany. 8Division of Cell Matrix Biology & Regenerative Medicine, School of Biological Sciences, Faculty of Biology, Medicine and Health, University of Manchester, Manchester, UK. 9Royal Manchester Children’s Hospital, Manchester University NHS Foundation Trust, Manchester Academic Health Science Centre, Manchester, UK. 10Department of Adolescent Urology, University College London Hospitals NHS Foundation Trust, London, UK. 11Genomics England, Queen Mary University of London, London, UK. 12Nephrology Department, Great Ormond Street Hospital for Children NHS Foundation Trust, London, UK.13Research Department of Pathology, University College London, London, UK.
Abstract
Background: Posterior urethral valves (PUV) are the commonest cause of childhood kidney failure and a major unmet clinical problem in paediatric nephrology. While usually sporadic, familial clustering and twin studies suggest a genetic component that is as yet unidentified. Using whole genome sequencing (WGS) we sought to understand the genetic architecture of PUV and identify key contributing genes.
Methods: We analysed WGS data from 132 unrelated male PUV patients and 23,727 ancestry-matched unaffected controls from the 100,000 Genomes Project, seeking enrichment of common and rare single-nucleotide and structural variation (SV) on a genome-wide, per-gene, and cis-regulatory element basis.
Results: Exome-wide there was no significant enrichment of rare coding variation in any one gene. SV analysis identified an increased burden of rare inversions affecting CTCF-only cis-regulatory elements (P=3.1x10-5; OR 2.1), but these did not affect any single genomic locus recurrently. GWAS of 17 million variants with minor allele frequency [MAF] >0.001 revealed significant (P<5x10-8) associations at two loci: 12q24.21 (P=7.8x10-12; OR 0.4; MAF 0.37) and 6p21.1 (P=2.0x10-8; OR 7.2; MAF=0.007), both of which replicated in an independent cohort of 398 European PUV patients. Bayesian fine mapping and in silico functional annotation mapped these loci to the transcription factor TBX5 and planar cell polarity gene PTK7, respectively. Both are highly expressed in the embryonic mouse urethra and known to regulate development.
Conclusion: This work demonstrates that non-specific perturbations of broad regulatory networks and chromatin looping may be important in the pathogenesis of PUV. Furthermore, for the first time, two genetic loci for PUV are identified which are mapped to the genes TBX5 and PTK7, providing novel insights into the biological mechanisms underlying this complex disorder.
Authors
Mazzarino M*,1, Cetin E*,1, Hughes T1, Ramji D2, Fraser D1, Meran S1, Labéta MO1, Raby A-C1
Affiliations
1Division of Infection and Immunity, School of Medicine, Cardiff University; 2Biomedicine Division, School of Biosciences, Cardiff University.
*These authors contributed equally to this work
Abstract
Cardiovascular disease (CVD) mortality is greatly increased in people with CKD. Traditional risk factors (e.g. diabetes, hypertension, obesity, ageing) cannot fully explain this high incidence. Therefore, factors specific to CKD, such as chronic inflammation or dialysis-associated infections, are believed to drive CVD. Given the importance of Toll-like receptors (TLRs) in mediating sterile and pathogen-induced inflammation, our recent research has focused on investigating their role in driving increased CVD risk in CKD. We hypothesise that CKD itself, the dialysis treatment, or dialysis-associated infections, lead to increased systemic concentrations of TLR endogenous ligands, called Damage-Associated Molecular Patterns (DAMPs). In turn, we envisage that these DAMPs will induce vascular inflammation and promote atherosclerosis development, an inflammatory disease of the arterial wall that underlies most cardiovascular complications.
Our recent studies identified 4 DAMPs (Hsp70, Hyaluronic acid, HMGB-1 and Calprotectin), significantly elevated in CKD plasma. Follow-up in vitro studies demonstrated that some of these DAMPs promote pro-atherogenic responses by macrophages, including pro-inflammatory mediator production (e.g. MCP-1, IL-8, IL-6), chemoattractant-driven migration, foam cell formation and expression of a large number of genes involved in most aspects of the atherosclerotic process (e.g., leukocyte recruitment, macrophage differentiation, cholesterol metabolism, cell death). In vivo experiments are ongoing to assess the potential of these CKD-associated DAMPs to mediate vascular inflammation and promote atherosclerosis.
In parallel in vivo work, we observed that a peritoneal infection episode - a common occurrence in patients on PD - leads to lasting changes to the systemic inflammatory state, which can be expected to promote atherosclerosis (e.g. increased pro-inflammatory monocytes proportion, expression of adhesion molecules and enhanced monocyte migratory capacity). Using patients and mouse plasma samples, we observed that a peritoneal infection also leads to a temporary increase in Calprotectin and Histone H3, another TLR DAMP. Further in vivo and in vitro experiments are ongoing to determine the potential role that these DAMPs may be playing in mediating the pro-atherosclerotic changes observed following peritonitis.
By investigating a potential role for TLR DAMPs in mediating increased vascular inflammation and promoting atherosclerosis in CKD and PD, our ultimate aim is to design therapeutic strategies to reduce long-term CV risk in CKD patients.
Authors
Michael Crompton1, Joanne K. Ferguson1, Raina D. Ramnath1, Karen L. Onions1, Anna S. Ogier1, Monica Gamez1, Colin J. Down1, Laura Skinner1, Lauren K. Dixon1, Judit Sutak2, Steven J. Harper3, Paola Pontrelli4, Loreto Gesualdo4, Hiddo L. Heerspink5, Robert D. Toto6, Gavin I. Welsh1, Rebecca R. Foster1, Simon C. Satchell1, Matthew J. Butler1
Affiliations
1 Bristol Renal, Bristol Medical School, University of Bristol, UK; 2 Pathology Department, Southmead Hospital, Bristol, UK; 3 School of Physiology, Pharmacology & Neuroscience, University of Bristol, UK; 4 Division of Nephrology, Department of Emergency and Organ Transplantation, Aldo Moro University of Bari, Italy; 5 Department of Clinical Pharmacology, University Medical Center Groningen, University of Groningen, The Netherlands; 6 Department of Clinical Sciences, The University of Texas Southwestern Medical Center, USA
Abstract
The glomerular endothelial glycocalyx (GEnGlx), a luminal proteoglycan layer, forms the first part of the filtration barrier. We have previously shown that excess mineralocorticoid receptor (MR) activation induces albuminuria via matrix metalloproteinase (MMP)-dependent damage of the GEnGlx. In rodent models, damage to the GEnGlx occurs early in the pathogenesis of diabetic nephropathy (DN). Here we sought to determine whether MR antagonism with spironolactone could prevent the development of albuminuria in diabetes, by preserving the GEnGlx.
Male Wistar rats injected with streptozotocin (50mg/kg I.P.) were used to study if spironolactone (50mg/kg/day S.C.), an MR inhibitor, could preserve the GEnGlx and limit the development of DN. MMP2 and MMP9 activities were quantified using a specific activity assay. Our glomerular permeability assay was used to directly measure the albumin permeability (Ps’alb), in isolation from haemodynamic changes. Our novel fluorescent profile peak-to-peak confocal imaging technique was validated against electron microscopy GEnGlx thickness measurements. We applied our validated peak-to-peak method to assess GEnGlx damage on renal biopsies from patients with DN and healthy controls. To confirm MR-mediated MMP inhibition in human DN we quantified MMP activity in urine from patients randomised to receive spironolactone or placebo.
Diabetic rats developed increased MMP activity, increased Ps’alb and albuminuria, with associated GEnGlx loss. MR antagonism reduced glomerular active MMP9, urine active MMP2 and -9, preserved the GEnGlx, restored Ps’alb and prevented diabetes-induced albuminuria progression. Enzymatic degradation of the GEnGlx, with hyaluronidase, reversed the effect of MR antagonism in diabetic rats, confirming the importance of GEnGlx preservation in this model. In humans, GEnGlx thickness was significantly reduced in DN compared to healthy controls. These findings were replicated in both centres, confirming GEnGlx damage in human diabetes. MR antagonism in patients with DN significantly reduced urine active MMP2 and albuminuria compared with placebo and baseline. Taken together our work suggests GEnGlx preservation is an important novel mechanism of reno-protection by MR inhibitors in diabetes.
Seminar 2: Transforming treatments
Abstracts:
Authors:
Syazrah Salam, 1,2 Orla Gallagher,2 Fatma Gossiel,2 Margaret Paggiosi,2 Arif Khwaja,1 Richard Eastell 2
Affiliations:
1Sheffield Kidney Institute, Sheffield Teaching Hospitals NHS Foundation Trust and 2Department of Oncology and Metabolism and Mellanby Centre for Musculoskeletal Research, University of Sheffield.
Abstract:
Patients with advanced chronic kidney disease (CKD) have high fracture risk due to high prevalence of both osteoporosis and renal osteodystrophy (ROD). ROD is characterised by abnormalities in bone turnover and mineralisation, resulting from biochemical and hormonal changes associated with CKD. The use of anti-resorptive or anabolic bone treatment for osteoporosis to reduce fracture risk in advanced CKD is limited by the requirement for a bone biopsy to exclude pre-existing low or high bone turnover respectively. Therefore developing validated non-invasive measures of bone turnover using biomarkers and imaging has the potential to transform the treatment of osteoporosis in advanced CKD.
In a study involving 43 CKD stages 4-5D patients, I assessed the diagnostic accuracy of circulating bone turnover markers (BTMs) and high resolution bone imaging to identify patients with low or high bone turnover ROD confirmed on trans-iliac bone biopsy.
BTMs such as bone alkaline phosphatase, intact procollagen type 1 N-terminal propeptide and tartrate-resistant acid phosphatase 5b were able to identify low bone turnover patients with good diagnostic accuracy. Area under the receiver operating characteristic curves (AUCs, 95% CI) were 0.82 (0.67 to 0.93), 0.79 (0.64 to 0.90) and 0.80 (0.64 to 0.91) respectively. High resolution bone imaging of distal radius also had similar diagnostic accuracy to the BTMs for low bone turnover ROD. The BTMs had similar diagnostic accuracy to intact parathyroid hormone [AUC 0.76 (0.60 to 0.88)] for the diagnosis of high bone turnover ROD.
Bone biomarkers and high resolution bone imaging can be used to identify patients who may be suitable for bone-specific treatment to reduce fracture risk without performing bone biopsy.
Authors
Thompson ER1,2,3Sewpaul A1,2,3,Figuereido R1,2,3, Bates L1,3, Ferdinand JR 3,Tingle SJ1,2,3,Connelly CM1, Hosgood SA3,4, Nicholson ML3,4, Clatworthy M R3,4, Ali S1,3, Sheerin NS1,2,3, Wilson CH1,2,3
Affiliations
1Translational and Clinical Research Institute, Newcastle University, 2 Institute of Transplantation, Freeman Hospital, Newcastle upon Tyne, 3 NIHR Blood and Transplant Research Unit in Organ Donation & Transplantation, 4 Department of Surgery, University of Cambridge
Abstract
Normothermic machine perfusion (NMP) is a novel clinical approach to overcome the limitations of traditional hypothermic organ preservation. NMP can be used to assess and recondition organs prior to transplant. In addition, NMP provides an opportunity to deliver therapeutic agents directly to the organ thus avoiding many limitations associated with systemic treatment of the recipient. We have investigated the potential reconditioning effects of two novel therapeutics in this setting. Firstly, a cell therapy, Multipotent Adult Progenitor Cells (MAPCs) that possesses potent immunomodulatory properties that could minimize ischemia reperfusion injury. Data demonstrated that NMP-MAPC treated kidneys had improved urine output, decreased expression of injury biomarker NGAL and improved microvascular perfusion on contrast-enhanced ultrasound. This was associated with an anti-inflammatory secretome that resulted in decreased neutrophil chemotaxis and improved endothelial integrity. Immunofluorescence revealed prelabelled MAPCs were found resident in the perivascular space of kidneys during NMP. The second therapy investigated in this setting was an RNA interference therapeutic, an antisense oligonucleotide (ASO) designed to block detrimental microRNA-24-3p function. Data demonstrated t ha t endosomal uptake during NMP conditions facilitated ASO co- localisation with proteins involved in the RNA-induced silencing complex (RISC) and demonstrated engagement of the miRNA target. This pattern of uptake was not seen during cold perfusion. RNAseq analysis revealed a protective decrease in inflammatory pathways in NMP-ASO treated kidneys and upregulation of microRNA-24-3p targets. A DCD pig model of kidney NMP was also developed and demonstrated no adverse toxicity associated with NMP-ASO therapy. In summary, NMP of donor kidneys prior to transplantation provides a platform for direct delivery of cellular and RNAi therapeutics to optimize organ quality prior to transplantation.
Authors
E. M. Castle1*, G.Dijk2, S.Shah1, E.Asgari3, R.Phillips4, J.Greenwood5, K. Bramham6, J. Chilcot7 S.Greenwood1
Affiliations
1 King’s Kidney Care, 2 Hammersmith renal unit, 3 GSTT renal unit, 4 CTU ICL, 5 Department of neurosurgery UCL, 6 Department of women and children health KCL, 7 Department of Health Psychology, KCL. *[email protected]
Abstract
Weight gain within the first year of receiving a kidney transplant is a critical health issue, with over half of kidney transplant recipients (KTRs) gaining > 5% of their body weight. Post-transplant weight gain is largely an increase in fat mass, which is associated with insulin resistance. KTRs have requested support with physical activity (PA) and weight gain prevention, however there is no routine care offered. A systematic review and meta-analysis (Castle et al., 2021) revealed that there is no recommended treatment to prevent weight gain for KTRs within the first year of receiving a transplant.
A Kidney Research UK funded PhD project delivered the design, revision and early testing of a digital health intervention (DHI)- the exercise and weight prevention in renal transplant online resource (ExeRTiOn). This intervention, co-designed with people who had received a kidney transplant and healthcare professional experts, provides people who have received a new kidney transplant with essential weight prevention guidance. Mixed-methods research studies allowed for the assessment of usability, acceptability and feasibility of the ExeRTiOn DHI, and iterative user-driven refinements.
The 12-week ExeRTiOn DHI was shown to be acceptable, and user-friendly resource to deliver weight prevention guidance to new KTRs in a South London kidney transplant centre (Castle et al., 2020). The intervention was revised based on participant feedback, for a feasibility RCT. The ExeRTiOn DHI was feasible to deliver, and secondary outcome data revealed that the ExeRTiOn DHI group had maintained their body weight at 12 months, whereas the median weight in the standard care group increased by 12kg. KTRs in the intervention arm also increased their six-minute walk distance, whereas this declined in the standard care group.
This ExeRTiOn DHI was shown to be feasible, acceptable, and user-friendly. Infrastructure to support digital healthcare research is warranted. Future research is required to explore the effectiveness and cost-effectiveness of weight gain prevention interventions for new KTRs.
Authors
B.R. Fletcher1, S. Damery2, O. Aiyegbusi1, N. Anderson1, M. Calvert1, P. Cockwell3, J. Ferguson3, M. Horton4, M.C.S. Paap5, C. Sidey-Gibbons6, A. Slade1, N. Turner7, D. Kyte1,8
Affiliations
1Centre for Patient Reported Outcomes Research (CPROR), Institute for Applied Health Research (IAHR), University of Birmingham, Birmingham, UK, 2Applied Research Collaboration West Midlands (ARCWM), Institute for Applied Health Research (IAHR), University of Birmingham, Birmingham, UK, 3Department of Renal Medicine, Queen Elizabeth Hospital Birmingham, University Hospitals Birmingham, Birmingham, UK, 4Leeds Psychometric Laboratory for Health Sciences, University of Leeds, Leeds, UK, 5Department of Child and Family Welfare, Faculty of Behavioural and Social Sciences, University of Groningen, Groningen, The Netherlands, 6MD Anderson Center for INSPiRED Cancer Care, University of Texas, Texas, USA, 7Centre for Inflammation Research, University of Edinburgh, Edinburgh, UK, 8School of Allied Health and Community, University of Worcester, Worcester, UK
Abstract
The importance of patient-reported outcome measurement in chronic kidney disease (CKD) populations has been established. However, there remains a lack of research that has synthesised data around CKD-specific symptom and health related quality of life (HRQOL) burden to inform capture of the most relevent, patient-important, information in a way that minimises patient burden.
We carried out a systematic review to produce a comprehensive and consolidated synthesis of symptom prevalence/severity and HRQOL across CKD stage and treatment groups.
We included all studies that included adult patients with CKD stage 1-5 not on dialysis, or receiving dialysis, or in receipt of a kidney transplant, and measured symptom prevalence/severity and/or HRQOL using a patient reported outcome measure.
Random effects meta-analyses were used to pool data on symptom prevalence and severity, and HRQOL scores. We stratified meta-analyses by CKD status: Stage 1-5 not on dialysis (predialysis), on dialysis, and post renal transplantation.
Our search identified 1,291 studies, of which 415 studies with 177,539 participants from 57 countries were included in the review and meta-analyses. Studies utilised 67 different symptom and HRQOL outcome measures. A total of 68 symptoms were reported, with predialysis and dialysis populations sharing a similar profile of the most prevalent symptoms.
Fatigue was particularly prevalent in predialysis and dialysis, and especially severe for dialysis patients. The evidence suggests that HRQOL is consistently worse in dialysis than predialysis, and that depression is more prevalent and severe. A number of symptoms were less prevalent/severe within the post-transplantation population, which may suggest attribution to CKD.
This systematic review highlights the considerable symptom and HRQOL burden associated with CKD. These data provide a basis for understanding the true burden for patients of one of the most common long-term conditions (affecting up to 1 in 7 adults) and will underpin the accurate development of tools to deliver virtual and remote care.
Authors
Dr Amarpreet Thind 1,2 ,Dr Annabel Rule 1, Dawn Goodall 2, Dr Shuli Levy 2, Dr Sarah Brice 2, Mr Frank Dor 1,2, Nicola Evans 2, David Ospalla, Professor Nicola Thomas 3, Dr David Wellsted 4, Dr Lina Johannsson 1,2, Dr Michelle Willicombe 1,2, Professor Edwina Brown 1,2
Affiliations
1Imperial College London, 2Imperial College Healthcare NHS Trust, 3London South Bank University, 4University of Hertfordshire
Abstract
Patient demand for and actual rates of transplantation in older people are both increasing in line with ageing of the dialysis population. Transplantation in older people, however, has mixed outcomes; increases in life expectancy are met with increased risks of hospitalisation, infections and peri-operative morbidity. Consequently, the impact on quality of life for older people is highly variable. With increasing awareness of the higher risk of frailty and cognitive impairment in people with end-stage kidney disease, information is needed on how these factors affect wait list progress and outcomes following kidney transplantation.
The Kidney Transplantation in Older People (KTOP) study is a prospective, observational study being conducted at the Imperial College Renal and Transplant centre. From October 2019 all patients aged 60 or above who are activated on the wait list or undergoing living kidney transplantation, are eligible for recruitment. Each participant completes a combination of questionnaires assessing frailty, cognitive function, and quality of life. These questionnaires are repeated at defined time points whilst on the wait list or following kidney transplantation. Clinical data is concurrently collected. A small number of patients will take part in a simultaneous qualitative study to assess how expectations meet the reality of transplantation.
Thus far 203 patients have been recruited, 75 of whom have received a kidney transplant. Current baseline frailty rates are 16.7% in the waitlist group and 16.1% in the patients transplanted (based on pre-transplant assessment with the Edmonton Frail Scale). A further 20.6% and 14.3% of patients in the wait list and transplant groups respectively, are considered ‘vulnerable’ to frailty. Baseline rates of cognitive impairment were 35.9% and 32.7% in the wait list and transplant patients respectively, based on Montreal Cognitive Assessments completed pre-transplant. The study is currently continuing with follow up activities and will complete in November 2022. The study will report on progression in these parameters over time, their association with clinical outcomes, and the impact on quality of life.
Ultimately, the KTOP study will enrich our understanding of transplantation in older people and help to tailor the assessment and care of this vulnerable population in order to maximise outcomes.
Authors
Natalie Hall*2, Christina Joanne Pearce*1, Joanna Hudson1; Ken Farrington2; David Wellsted2; Julia Jones3; Shivani Sharma2; Sam Norton1; Paula Ormandy4, Nick Palmer5 and Joe Chilcot1
*joint first authors
Affiliations
1Health Psychology Section, Psychology Department, Institute of Psychiatry, Psychology and Neuroscience, King's College London, 5th floor Bermondsey Wing, Guy's Campus, London Bridge, London SE1 9RT, UK; 2Department of Psychology, School of Life and Medical Sciences, University of Hertfordshire, College Lane Campus, Hatfield AL10 9AB, UK; 3School of Health and Social Work; University of Hertfordshire; Hatfield, AL10 9AB, UK; 4School of Health and Society, University of Salford, Salford, M6 6PU, UK; 5Kidney Care, Alton, GU34 1EF, UK.
Abstract
Depression is highly prevalent across the spectrum of Chronic Kidney Disease (CKD) and related to poor clinical outcomes. There is limited evidence regarding the most effective and acceptable interventions to manage depression in people living with kidney disease and little is known about the care pathways for depression in CKD. This review aimed to investigate how depression is identified and managed in adult patients with CKD, since the publication of the NICE clinical guidelines for managing depression in chronic physical health problems published in 2009.
Eight relevant databases were utilised to identify appropriate literature, using a systematic search with pre-defined inclusion criteria. Data were extracted which were of relevance to the identification and/or management of depression in CKD.
Of 63 articles identified, 40 were included in the sample as relevant records for extraction (spanning 2010-2020). Of these records there were: nineteen cross-sectional questionnaire studies (48%); thirteen review articles (33%); three cohort studies (8%); two RCTs (5%) and one audit. Only two studies referred to the NICE 2009 guidelines.
The most frequently utilised depression screening tools included: The Beck Depression Inventory (n=18 studies, 46%); the Patient Health Questionnaire (PHQ-9 and 2 (n=5 studies, 13%) and the Hospital Anxiety and Depression Scale (n=3 studies, 8%).
Evidence regarding effective treatments for depression in CKD included: Cognitive Behavioural Therapy (n=4 studies, 10%); psychotherapy (n=1 study, 3%); and counselling (n=1 study, 3% weak evidence).
There was limited methodologically strong evidence for pharmacological interventions and adverse events appear common following SSRI initiation. Two RCTs included in this review found no effect of Sertraline on depression and reported more adverse events in this group. Despite this, data from the USA suggests that over forty percent of CKD patients are prescribed antidepressants.
In conclusion the identification of depression is commonly evaluated using validated screening tools with appropriate renal specific cut-off scores. The most rigorous RCT studies show no beneficial effect of the use of antidepressants for use in patients with CKD. Psychological therapies have been more widely studied with moderate evidence regarding efficacy and acceptability. Understanding national clinical practice patterns may help facilitate the development of renal specific guidelines for the identification and management of depression in CKD.
Authors
Tegwen Elliott, Nina Jordan, Sarah Hosgood, Michael Nicholson
Affiliations
Department of Surgery, University of Cambridge
Abstract
Normothermic machine perfusion (NMP) is a method of kidney preservation designed to restore cellular metabolism after cold ischaemia. Kidneys are perfused with an oxygenated red-cell based solution for 1h at 36°C. During NMP red-cells can become damaged releasing free haem into the perfusate. This can act as a damage associated molecular pattern (DAMP) activating inflammatory signaling pathways. The aim of this study was to measure levels of haem and inflammatory mediators during NMP and to determine their association with early graft function.
Levels of free haem and inflammatory cytokines were measured in perfusate samples from 43 donation after cardiac death (DCD) kidneys undergoing NMP. Levels were correlated with the duration of delayed graft function (DGF). Changes in transcriptional gene expression were analysis in 4 NMP kidneys declined for transplant using the NanoString nCounter Organ Transplant Panel.
In the transplanted kidneys, levels of haem and IL-6 increased significantly during NMP (p<0.0001, <0.0001). Higher levels of haem were associated with older units of packed red-cells (p=0.022). Twenty-seven kidneys (63%) had DGF with a median duration of 4 days (range 1-24 days). There were no significant correlations with levels of haem or IL-6 in the duration of DGF (p=0.471. IL-6 p=0.217). In the declined kidneys, levels of haem also increased significantly during NMP (P=0.0001). Forty-two differentially expressed genes were significantly upregulated after NMP and 1 downregulated. Genes associated with apoptosis (FOS and JUNE), inflammatory cytokines (IL-6, SOCS3, ATF3), chemokines (CXCL8, CXCL2, CC3/L1) and oxidative stress (KLF4) were in the top 20 significantly upregulated differentially expressed genes. Several genes associated with anti-inflammatory properties (IL-10) and endothelial cell recovery; vascular endothelial growth factor (VEGF) were also significantly upregulated. Collagen type 1alpha 1chain (COL1A1), a gene involved with fibrosis was significantly downregulated (fold change -3.31; P=0.047).
A significant amount of haem can be detected in the perfusate after NMP particularly in when older units of red-cells are used. There was no direct association with levels of haem or inflammation on early graft function. Transcriptional analysis demonstrated some beneficial effects of NMP but there was significant upregulation of genes involved with apoptotic, inflammatory and oxidative pathways that may be stimulated by the harmful effects of haem.
Authors
A.D. Lalayiannis1, N. Crabtree2, C.J. Ferro3, D.C. Wheeler4, N.D. Duncan5, C. Smith6, J. Popoola7, V. Askiti8, A. Mitsioni8, A. Kaur9, M.D. Sinha10, L. Biassoni1, S.P. McGuirk2, K.H. Mortensen1, D.V. Milford2, J. Long11, M. Fewtrell1, M.D. Leonard11, R. Shroff1
Affiliations
1 University College London Great Ormond Street Hospital Institute of Child Health, London, UK, 2Birmingham Women’s and Children’s NHS Foundation Trust, Birmingham, UK, 3University Hospitals Birmingham NHS Foundation Trust, Birmingham, UK, 4Department of Renal Medicine, University College London, London, UK, 5Imperial College Renal and Transplant Centre, Hammersmith Hospital, London, UK, 6Institute of Global Health, University College London, UK, 7 Department of Nephrology and Transplantation, St. George's University Hospital NHS Foundation Trust, London, UK, 8"P. & A. Kyriakou" Childrens' Hospital, Athens, Greece, 9Manchester University NHS Foundation Trust, Manchester, UK, 10Evelina Children’s Hospital, Guy's & St Thomas' NHS Foundation Trust, London, UK, 11 Stanford School of Medicine, Stanford University, USA
Abstract
Introduction
Patients with Chronic Kidney Disease(CKD) can have low bone mineral density(BMD) with concurrent vascular calcification. This has been demonstrated in older adults on dialysis but not in children and young adults with CKD, in whom mineral accrual by the growing skeleton may provide a ‘buffering effect’ protecting against extraosseous calcification.
Our hypothesis was that children and young adults with increasing BMD do not have evidence of vascular calcification.
Methods
This is a multicenter longitudinal study in children and young adults(5-30 years old; CKD stages 4-5 or on dialysis).
Cortical[Cort] and trabecular[Trab] BMD were assessed by peripheral quantitative computed tomography(pQCT) and lumbar spine BMD by DXA. Vascular calcification was assessed by cardiac CT for coronary artery calcification(CAC) & ultrasound for carotid intima-media thickness(cIMT); arterial stiffness by pulse wave velocity(PWV) & carotid distensibility. All bone and cardiovascular measures are presented as age- and sex-adjusted z-scores allowing for comparison of the vascular measures across all age-groups.
Results
Ninety-eight children and young adults(median age 13.82; IQR 10.69,16.46 years) were assessed at baseline and 55 followed up after a median of 1.45(1.25 to 1.81) years. Median cIMT and PWV were 2.17(1.14, 2.86) and 1.45(-0.16, 2.57) at baseline that remained static at follow up. 10% of the cohort had CAC, increasing to 56% at follow-up. TrabBMD decreased(-0.26 to -0.38,p=0.01), and there was a non-significant decrease in CortBMD(-0.47 to -1.13,p=0.09).
Baseline TrabBMD was independently associated with cIMTz (R2 0.10, β 0.34, p=0.001) on multivariable linear regression modelling. The odds of having a concurrent annualised change in cIMT(ΔcIMT) increase with annualised TrabBMD change(ΔTrabBMD) increase was 6 times higher(95% CI 1.88 to 18.35).
Young people with growth had lower ΔTrabBMD, implying ongoing skeletal mineralisation, but also lower ΔcIMT, ΔPWV and higher Δdistensibility compared to those with static growth.
Conclusion
There is a high burden of subclinical cardiovascular disease in young people with CKD 4-5 and on dialysis. An increase in cIMT was seen despite an increase in TrabBMD, with the most significant increase in cIMT in young people with static growth. The growing skeleton may offer some protection against vascular calcification.
Reconciling the calcium requirement for optimal bone mineralisation whilst simultaneously avoiding any harm to the vasculature remains challenging.
Authors
Katherine E. Memory1, Thomas J. Wilkinson1, Alice C. Smith1, Courtney J. Lightfoot1
Affiliations
1Leicester Kidney Lifestyle Team, Department of Health Sciences, University of Leicester and University Hospitals Leicester NHS Trust, Leicester, UK
Abstract
To effectively self-manage their condition, kidney transplant recipients (KTRs) need to have knowledge, skills, and confidence, termed patient activation (PA). Given that those with lower PA are more likely to be hospitalised and report poorer quality of life (QoL), there is growing interest in PA and person-centred care to encourage patients to take a more active role in their health and healthcare. Currently, limited research exists on PA and self-management (SM) experiences of KTRs. This study aimed to understand KTRs PA levels, identify characteristics associated with low PA that could be targeted in future interventions, and explore their perspectives of SM.
A mixed-methods approach was utilised: 158 KTRs (43% female, mean age 59.8 (±12.5) years, eGFR 46.9 (±18.8) ml/min/1.73m2) completed a survey on kidney disease and lifestyle, including the Patient Activation Measure (PAM). The PAM is a well-validated 13-item tool assessing an individual’s perceived activation, categorising participants into four levels (one=lowest; four=highest). 11 semi-structured interviews were conducted to explore participants’ experiences of SM, which were analysed using thematic analysis.
86/158 (54%) reported low activation (PAM levels 1 and 2). Low activated participants were older (p=.001), unemployed (p=.002), reported greater years since diagnosis of kidney disease (p=.029), more comorbidities (p=.036), more symptoms (p=.018), mental health problems (p=.026), reduced physical function (p=.002), poorer physical QoL (p=.017), and lower eGFR (p=.035). Overarching themes from the interviews were ‘SM perceptions and experiences’, ‘Facilitators and barriers to SM’, and ‘Desired SM support’.
KTRs reported low PA levels, suggesting SM support is required for KTRs. Factors associated with low PA in KTRs were identified which can be utilised to provide targeted SM resources. Tailored support should aim to minimise the effect of perceived barriers to SM, including symptoms, comorbidities, and complications. Providing collaborative consultations, identifying achievable health goals, and understanding individuals’ perceived emotional burden following transplantation could nurture effective SM in KTRs. Future SM interventions should be personalised to the individual and integrate family education and peer support.
Authors
Mark Howard1, Conrad Farrar1, Steven Sacks1
Affiliations
MRC Centre for Transplantation, Peter Gorer Department of Immunobiology, School of Immunology & Microbial Sciences, King’s College London, London, UK
Abstract
Ischaemia/reperfusion (I/R) injury is an important consequence of transplant procedures and is partially driven by complement activation. We previously identified that a fucosylated ligand exposed by ischaemia on renal tubule epithelial cells is recognised by collectin-11 (CL-11), which is known to initiate complement activation and in turn promote acute kidney injury. Consequently, we hypothesised that increasing the local tissue concentration of free L-fucose following systemic administration would compete for ligand binding of locally made CL-11 and prevent complement activation. In a mouse model of I/R injury, an increase in the renal concentration of L-fucose generated by an intra-peritoneal (IP) bolus of L-fucose and given before the ischaemic event, was equal to that which significantly blocked in vitro binding of CL-11 on hypoxia-stressed renal tubule cells. This high renal L-fucose concentration led to blockade of complement activation and reduction of post-ischaemic acute kidney injury, with additional protection gained by a second IP bolus of L-fucose given immediately after the I/R injury. This process was CL-11 dependent as shown by CL-11 deficient mice gaining no additional protection from L-fucose administration. Early data using murine renal transplants showed a preservation of renal architecture in transplants where the donor was pre-treated with L-fucose. Therefore, our results support the hypothesis that an appropriately high level of L-fucose in the kidney reduces complement-mediated damage due to ischaemic insult by obstructing the carbohydrate recognition site on CL-11, and this may have an important therapeutic role in improving the outcomes of renal transplantation.
Authors
Claudia Bruno1, Rayan Moumneh1, Lynsey Stronach3, Emilie Sauvage1, Alberto Redaelli2, Ian Simcock3, Silvia Schievano1, Claudio Capelli1 and Rukshana Shroff3
Affiliations
1University College London, Institute of Cardiovascular Science (London, UK), 2Department of Electronics, Information and Bioengineering, Politecnico di Milano (Milan, Italy), 3Great Ormond Street Hospital for Children, NHS Foundation Trust (Great Ormond Street, London, UK)
Abstract
Introduction
The majority of children have haemodialysis (HD) through central venous lines (CVLs). However, nearly half the patients require a change of line due to frequent complications such as low blood flow, thrombosis and infection. Computational fluid dynamics (CFD) can be used to characterize the haemodynamic of CVLs, evaluating quantities that cannot be measured in vivo. In this study, we modelled CVLs commonly used in children to investigate the correlation between fluid dynamics parameters and clinical outcomes.
Methods
Four models of CVLs of varying design and sizes (6.5Fr, 8Fr, 10Fr and 14Fr) were scanned using microCT in order to reconstruct accurate computational models. CFD analyses were set up to simulate CVLs working conditions in both ideal and anatomical models of superior vena cava. Velocity fields, shear stresses and Platelet Lysis Index (PLI) are some of the haemodynamic features that were analysed. Results were compared to clinical data on catheter dysfunction collected at our centre over the past two years (n= 26 patients on HD with a total of 57 CVLs). Lastly, image analyses were performed on CVLs explanted from patients.
Results
Analysis of the velocity fields showed that the arterial lumen was the most critical one in terms of stagnation areas. Presence of jets through the side holes were identified in the venous configuration (Figure 1a). Side-holes played a crucial role in fluid exchange in all except the 8Fr design. Highest presence of shear stresses >10Pa were also found in the 8Fr CVL while highest PLI in the 10F model. CFD analysis on the anatomical model showed higher levels of wall shear stress (Figure 1b) and an increased disturbance of the flow inside the vein after catheter placement.
CFD results were in accordance with the clinical data which showed a higher recurrence rate of thrombosis for the 8Fr and 10Fr CVLs (Figure 1c). Also, microCT of the explanted lines confirmed presence of thrombosis at the level of the catheter tips.
Conclusions
This study provides a novel characterization of fluid- dynamics of paediatric CVLs. These findings may explain the poor outcomes recorded in clinical practice and guide the optimisation of CVL design in the future.
Seminar 1: Celebrating our 60th anniversary abstracts
Invited 15-minute presentations:
Authors:
Tanya Hutter,1,2 Liz Norgett,1,2 Fiona Karet,1,2 Tom Collings2
Affiliations:
1University of Cambridge, 2Kalium Health Ltd
Abstract:
Kidney patients and others can be endangered by very high blood potassium levels (hyperkalaemia). Patients with certain rare diseases, and particularly those on blood-pressure lowering medications, can have the opposite problem of severely low potassium (hypokalaemia). Currently, there are no practical means of measuring potassium levels outside of specialist hospital or laboratory facilities, which means delays and inconvenience to both patients and their healthcare providers. There is no way to immediately identify potentially life-threatening potassium levels and consequently no opportunity to rapidly treat patients at risk.
Providing a rapid, low-cost, easy to use test for use at home, at the GP or at a community-based dialysis unit would improve health management and safety in a home or healthcare setting, and rapidly identify potentially life-threatening situations.
We are developing a small electrochemical sensor which, in response to the presence of potassium, creates a detectable signal. We have integrated this sensor onto a disposable test strip that requires only a small fingerprick drop of blood. This talk will present our technology development journey from the initial idea to founding a startup company for technology commercialization.
Authors:
Wen Y. Ding1, Sarah Hunter1, Abigail Lay1, Bryony Hayes1, Valeryia Kuzmuk1, Matthew Beesley2, Ruth Rollason1, Jenny Hurcombe1, Fern Barrington1, William Cathery1, Catrin Masson1, Carl J May1, Timothy Roberts1, Geraldine Mollet3, Colin Chu4, Jenny McIntosh5, Richard J. Coward1, Corinne Antignac3, Amit Nathwani5, Gavin I. Welsh1, Moin A. Saleem1
Affiliations:
1Bristol Renal, Bristol Medical School, University of Bristol, Bristol, UK, 2Department of Histopathology, Cheltenham General Hospital, Cheltenham, UK, 3INSERM U983, Imagine Institute, Paris, France, 4Academic Unit of Ophthalmology, Bristol Medical School, University of Bristol, Bristol, UK, 5Research Department of Haematology, University College London, London, UK.
Abstract:
Kidney gene therapy has proven challenging. Adeno-Associated Virus (AAV) is used successfully for gene therapy targeting other organs, with particular success demonstrated in monogenic diseases like Leber’s Congenital Amaurosis, Haemophilia B and Spinal Muscular Atrophy. We aimed to progress kidney gene therapy by targeting a monogenic disease of the kidney podocyte. The most common causes of childhood genetic nephrotic syndrome are mutations in the podocyte gene NPHS2 which encodes podocin. We used AAV gene therapy to rescue this genetic defect in human and mouse genetic models. In vitro transduction identified AAV-LK03 serotype as a highly efficient transducer of human podocytes. AAV-LK03 transduction of podocin mutant human podocytes showed functional rescue in vitro, while AAV 2/9 gene transfer in the inducible podocin knock-out and knock-in mouse resulted in successful amelioration of kidney disease. This is the first successful AAV mediated gene rescue in monogenic renal disease and establishes the podocyte as a tractable target for gene therapy approaches.
Authors:
William G. Herrington, David Kerr
Affiliations:
University of Oxford
Abstract:
UK primary care data 2000-2011 showed the prevalence of overweight/obese (a body mass index [BMI] ≥25kg/m2) peaked in men at ages 50–54y at ~75%, and peaked at ages 65–69y in women at ~67%. Conventional observational analyses showed that, compared to “ideal” BMI, being overweight (BMI ≥25<30) increased risk of stage 4-5 chronic kidney disease (CKD) by about one-third, and being obese (BMI ≥30<35) and severely obese (BMI ≥35) approximately doubled and tripled risk, respectively. These associations were similar among those with and without diabetes mellitus (DM) or hypertension. Assuming causality, an estimated 39% (95%CI 36–42%) of CKD stage 4-5 in women, and 26% (22–30%) in men aged 40–79y resulted from being overweight/obese. Whether these associations are causal, the contribution of central versus general adiposity, and the underlying mechanisms are uncertain.
To address these uncertainties, we used data from 281,228 UK Biobank participants and estimated the relevance of waist-to-hip ratio and BMI to CKD prevalence, comparing genetic and conventional approaches. Genetic analyses used Mendelian randomization (MR) and data from 394 waist-to-hip ratio and 773 BMI-associated loci. Models assessed the role of known mediators (DM/BP) by adjusting for measured values (conventional analyses) or genetic associations of the selected loci (multivariable MR).
Evidence of CKD was found in 18,034 (6.4%) UK Biobank participants. Each 0.06 higher measured waist-to-hip ratio and each 5kg/m2 increase in BMI were associated with 69% (odds ratio, 1.69, 95%CI 1.64-1.74) and 58% (1.58; 1.55-1.62) higher odds of CKD, respectively. In analogous MR analyses, each 0.06–genetically-predicted higher waist-to-hip ratio was associated with a 29% (1.29; 1.20-1.38) increased odds of CKD, and each 5kg/m2 genetically-predicted higher BMI was associated with a 49% (1.49; 1.39-1.59) increased odds. After adjusting for DM and measured BP, chi-squared values for associations for waist-to-hip ratio and BMI fell by 56%. In contrast, mediator adjustment using multivariable MR found 83% and 69% reductions in chi-squared values for genetically-predicted waist-to-hip ratio and BMI models, respectively.
In conclusion, genetic analyses suggest that conventional associations between excess adiposity with CKD are largely causal, but underestimate mediating roles of diabetes, BP, and their correlates. Genetic approaches suggest these mediators explain most of adiposity-CKD–associated risk.
Short, selected presentations:
Authors:
Elizabeth Ralston 1, Amanda Clery 2, Yanzhong Wang 1, Kate Wiles 1, Hannah Blakey 3, Graham Lipkin 4, Matt Hall 5, Liz Lightstone 6, Lucy Chappell 1, Philip Webster 7, Sue Carr 8, Kate Bramham 1
Affiliations:
1King’s College London, 2University College London, 3Queen Elizabeth Hospital Birmingham, 4University Hospitals Birmingham NHS Foundation Trust, 5Nottingham University Hospitals NHS Trust, 6Imperial College London, 7Imperial College Healthcare NHS Trust, 8Univeristy of Leicester
Abstract:
Pregnancy can accelerate the decline in kidney function in women with Chronic Kidney Disease (CKD). These pregnancies are also at greater risk of adverse pregnancy outcomes. Our aim is to develop a prediction tool that estimates the degree of pregnancy associated progression of renal disease and adverse pregnancy outcomes in women with CKD.
Three UK obstetric prospective cohort studies were combined to create a model development dataset. Women with an estimated glomerular filtration rate (eGFR CKD-EPI without ethnicity adjustment) <90 mls/min/1.73m2 within 24 months preconception were included. The primary outcome was a binary indicator variable for whether there is a 25% or greater reduction in eGFR between 6 weeks and 12 months after delivery. The secondary outcome was a composite indicator variable: premature birth (<34 weeks’ gestation) and/or small for gestational age (SGA). For both outcomes, each predictor was tested in a univariable logistic regression model to estimate crude associations with the outcome, Odds Ratios (OR) with confidence intervals are reported. Significant and clinically relevant predictors were included in each multivariable model using backwards elimination. Model calibration and discrimination were assessed and area under the curve (AUROC) values were calculated with 95% confidence intervals. Internal validation in the same dataset was assessed using k-fold cross validation.
Pre-pregnancy eGFR (OR 1.0, 95% CI 0.9-1.0), diabetic nephropathy (OR 6.5, 95% CI 1.2-48.0) and clinically relevant proteinuria (OR 3.2, 95% CI 1.6-6.6) were independently associated with an ≥25% decline in eGFR postpartum. Together in the multivariable model demonstrated strong performance with an AUROC of 0.731 (95% CI 0.643-0.818). Pre-pregnancy eGFR (OR 1.0, 95% CI 1.0-1.0), diabetic nephropathy (OR 5.3, 95% CI 1.8-16.3) and chronic hypertension (OR 3.7, 95% CI 1.9–7.5) were independently associated with having a premature birth and/or SGA. Together in the model, they demonstrated strong performance with an AUROC of 0.721 (95% CI 0.655-0.787).
We have developed a prediction tool with high performance for estimation of a 25% or greater decline in eGFR postpartum and estimation of preterm birth and/or SGA. This could have important clinical value to both women and healthcare professionals. External validation is needed prior to introduction into routine care.
Authors:
Catherine E Lovegrove1,2, Akira Wiberg3, Thomas Littlejohns4, Naomi Allen4,
Benjamin W Turney 1, Anubha Mahajan5, Fadil Hannan6, Rajesh Thakker2, Mark I McCarthy5, Dominic Furniss3, Sarah A Howles1,2
Affiliations:
1 University of Oxford Nuffield Department of Surgical Sciences , 2 Academic Endocrine Unit, Radcliffe Department of Medicine, University of Oxford 3 University of Oxford Nuffield Department of Orthopaedics, Rheumatology and Musculoskeletal Sciences, 4 University of Oxford Nuffield Department of Population Health, 5 Wellcome Centre for Human Genetics, University of Oxford, 6 University of Oxford Nuffield Department of Women’s and Reproductive Health
Abstract:
Kidney stone disease (KSD) affects up to 18% of men and 11% of women in their lifetime and ≈50% of individuals will suffer from recurrent disease. The pathogenesis of KSD is poorly understood but has been linked to features of metabolic syndrome (MetS). We sought to increase understanding of these associations using conventional and genetic epidemiological approaches.
Observational data from 492,380 UK Biobank participants demonstrated that high waist hip ratio (WHR, ≥0.9 in men, ≥0.85 in women) confers a >40% increased risk of stone formation in patients with BMI ≥25 and <30kg/m2 compared to individuals with a normal WHR in this BMI range. Using a linear regression model we observed that an increased WHR was associated with increased adjusted serum calcium and decreased 25-OH vitamin D concentrations.
Using Mendelian randomisation (MR), we assessed causal associations between MetS phenotypes (WHR adjusted for BMI (WHRadjBMI), hypertension, hyperlipidaemia, hypercholesterolaemia, type 2 diabetes (T2DM),fasting insulin, and fasting glucose), and both KSD and metabolites of calcium homeostasis. One standard deviation (SD) increase in WHRadjBMI was found to be associated with a 45% increased risk of KSD. Gender-stratified analysis did not confer strong evidence of a gender-associated effect of WHRadjBMI on risk of KSD. Furthermore, type 2 diabetes and increased serum HDL concentrations conferred an increased risk of KSD (8% and 18% increased risk per 1 SD increase, respectively); the risk conferred by high HDL on KSD demonstrated horizontal pleiotropy. Exploring associations of WHR, T2DM, and HDL with biochemical phenotypes we found that elevated HDL concentrations were causally associated with increased adjusted serum calcium (OR 1.05 (95% CI 1.01-1.09), P=0.01), and that a 1SD increase in adjusted serum calcium conferred a 69% increased risk of KSD. No other phenotypes associated with MetS demonstrated causality with KSD.
In conclusion, we have demonstrated that central adiposity and T2DM are causally associated with KSD whilst elevated HDL demonstrates a pleiotropic relationship with KSD that may be mediated via alterations in calcium homeostasis. These findings motivate weight management and glycaemic control in KSD prevention and provide a basis for further studies to define causal metabolic pathways.
Authors:
William J Mason1,2, Daniyal J Jafree2,3, Gideon Pomeranz2, Maria Kolatsi-Joannou2, Antje K Grotz2, Sabrina Pacheco2, Dale A Moulding2, Anja Wolf4, Christian Kupatt4, Claire Peppiatt-Wildman5, Eugenia Papakrivopoulou2,6, Paul R Riley7, David A Long2, Elisavet Vasilopoulou1,2,8
Affiliations:
1Medway School of Pharmacy, Division of Natural Sciences, University of Kent, Chatham, Kent, UK, 2Developmental Biology and Cancer Programme, UCL Great Ormond Street Institute of Child Health, London, UK, 3UCL MB/PhD Programme, Faculty of Medical Science, University College London, London, UK, 4Medizinische Klinik und Poliklinik I, TUM Munich, Germany 5Division of Natural Sciences, University of Kent, Chatham, Kent, UK, 6Department of Internal Medicine and Nephrology, Clinique Saint Jean, Brussels, Belgium, 7Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford, UK, 8Comparative Biomedical Sciences, The Royal Veterinary College, London, UK
Abstract
Glomerular disease is associated with the injury of podocytes, epithelial cells that wrap around the glomerular capillaries. Podocytes have a unique shape maintained by a complex cytoskeleton, which becomes disrupted in glomerular disease resulting in defective filtration and albuminuria. Targeting the podocyte cytoskeleton is therefore a promising new avenue to slow the progression of glomerular disease. Thymosin β4 (TB4), is a peptide that regulates actin polymerisation. Lack of endogenous TB4 exacerbates glomerular injury and disrupts the organisation of the podocyte actin cytoskeleton, however, the effect of treatment with exogenous TB4 on glomerular injury is unknown. We hypothesised that exogenous TB4 would alleviate the phenotypes of Adriamycin nephropathy, a rodent model of focal segmental glomerulosclerosis. Through interrogating single-cell RNA-sequencing data of isolated glomeruli we demonstrate that Adriamycin injury results in reduced levels of podocyte TB4 compared with vehicle treated mice (P<0.05). Systemic administration of an adeno-associated virus vector encoding TB4 (AAV-Tmsb4x) successfully transduced liver cells (10-fold upregulation, P<0.001) and resulted in a 2-fold increase in circulating TB4 levels (P<0.01) compared with mice administered the control virus (AAV-LacZ). TB4 gene therapy prevented Adriamycin-induced podocyte loss (P<0.001) and albuminuria (P<0.01). Whilst there was no difference in the amount of F-actin in podocytes, Adriamycin injury was associated with disorganisation of the actin cytoskeleton in vitro (P<0.001), which was ameliorated by exogenous TB4 (P<0.001). Furthermore, Adriamycin administration in mice was associated with increased prevalence of podocyte vesicles (P<0.001), a mechanism by which albumin may leak into the urine, which was also prevented by TB4 (P<0.01). Collectively, we propose that systemic TB4 gene therapy prevents podocyte injury and maintains glomerular filtration via modulation of the podocyte cytoskeleton thus presenting a novel treatment strategy for glomerular disease.