Sequencing of LRP2 Reveals Multiple Rare Variants Associated with Urinary Trefoil Factor-3

Novel biomarkers are being investigated to identify patients with kidney disease. We measured a panel of 13 urinary biomarkers in participants from the Offspring Cohort of the Framingham Heart Study. Using an Affymetrix chip with imputation to 2.5 M single-nucleotide polymorphisms (SNPs), we conducted a GWAS of these biomarkers (n=2640) followed by exonic sequencing and genotyping. Functional studies in zebrafish were used to investigate histologic correlation with renal function. Across all 13 biomarkers, there were 97 significant SNPs at three loci. Lead SNPs at each locus were rs6555820 (P=6.7×10⁻⁴⁹; minor allele frequency [MAF]=0.49) in HAVCR1 (associated with kidney injury molecule-1), rs7565788 (P=2.15×10⁻¹⁶; MAF=0.22) in LRP2 (associated with trefoil factor 3 [TFF3]), and rs11048230 (P=4.77×10⁻⁸; MAF=0.10) in an intergenic region near RASSF8 (associated with vascular endothelial growth factor). Validation in the CKDGen Consortium (n=67,093) showed that only rs7565788 at LRP2, which encodes megalin, was associated with eGFR (P=0.003). Sequencing of exons 16–72 of LRP2 in 200 unrelated individuals at extremes of urinary TFF3 levels identified 197 variants (152 rare; MAF<0.05), 31 of which (27 rare) were nonsynonymous. In aggregate testing, rare variants were associated with urinary TFF3 levels (P=0.003), and the lead GWAS signal was not explained by these variants. Knockdown of LRP2 in zebrafish did not alter the renal phenotype in static or kidney injury models. In conclusion, this study revealed common variants associated with urinary levels of TFF3, kidney injury molecule-1, and vascular endothelial growth factor and identified a cluster of rare variants independently associated with TFF3.Serum creatinine, as used in most GFR estimating equations, is the primary biomarker of CKD.¹ Creatinine has significant limitations as a biomarker. It is insensitive to early declines in kidney function, and there are important extrarenal factors that influence its concentration, thus increasing the potential for misclassification when it is used to diagnose renal disease.² As a result, novel biomarkers are being investigated that could allow earlier and more accurate diagnosis of CKD.³ It is likely, however, that these biomarkers also have non-GFR determinants, including genetic factors. For example, levels of cystatin C, a novel GFR biomarker, are known to be influenced by both nongenetic factors, such as adiposity and the metabolic syndrome,⁴ and the presence of specific genetic variants in the cystatin C-gene cluster.⁵Genome-wide association studies (GWAS) allow for the evaluation of genetic associations of biomarkers in an unbiased manner. These studies could illuminate previously unrecognized pathways for CKD pathogenesis, thus identifying new potential molecular targets for therapeutic intervention. Multiple variants have been identified in association with kidney function through GWAS,⁶ whereas a variant in CUBN encoding cubilin, a proximal tubular transport protein, has been associated with albuminuria.⁷ Recently, a GWAS identified a locus at PTGDS in association with another kidney biomarker, β-trace protein.⁸To gain additional insight into biologic pathways of renal function and kidney injury, we measured a panel of 13 urinary biomarkers in participants from the Framingham Heart Study (FHS) and performed a GWAS of their levels. Here, we report the primary results from these studies as well as follow-up work to identify whether rare variants in LRP2 are associated with levels of trefoil factor 3 (TFF3).     

Replication Kinetics of B.1.351 and B.1.1.7 SARS-CoV-2 Variants of Concern Including Assessment of a B.1.1.7 Mutant Carrying a Defective ORF7a Gene

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the etiological agent of COVID-19, is a readily transmissible and potentially deadly pathogen which is currently re-defining human susceptibility to pandemic viruses in the modern world. The recent emergence of several genetically distinct descendants known as variants of concern (VOCs) is further challenging public health disease management, due to increased rates of virus transmission and potential constraints on vaccine effectiveness. We report the isolation of SARS-CoV-2 VOCs imported into Australia belonging to the B.1.351 lineage, first described in the Republic of South Africa (RSA), and the B.1.1.7 lineage originally reported in the United Kingdom, and directly compare the replication kinetics of these two VOCs in Vero E6 cells. In this analysis, we also investigated a B.1.1.7 VOC (QLD1516/2021) carrying a 7-nucleotide deletion in the open reading frame 7a (ORF7a) gene, likely truncating and rendering the ORF7a protein of this virus defective. We demonstrate that the replication of the B.1.351 VOC (QLD1520/2020) in Vero E6 cells can be detected earlier than the B.1.1.7 VOCs (QLD1516/2021 and QLD1517/2021), before peaking at 48 h post infection (p.i.), with significantly higher levels of virus progeny. Whilst replication of the ORF7a defective isolate QLD1516/2021 was delayed longer than the other viruses, slightly more viral progeny was produced by the mutant compared to the unmutated isolate QLD1517/2021 at 72 h p.i. Collectively, these findings contribute to our understanding of SARS-CoV-2 replication and evolutionary dynamics, which have important implications in the development of future vaccination, antiviral therapies, and epidemiological control strategies for COVID-19.     

Improved Real-World Glycemic Control With Continuous Glucose Monitoring System Predictive Alerts

Background:Most standalone real-time continuous glucose monitoring (RT-CGM) systems provide predictive low and high sensor glucose (SG) threshold alerts. The durations and risk of low and high SG excursions following Guardian™ Connect CGM system predictive threshold alerts were evaluated.Methods:Continuous glucose monitoring system data uploaded between January 2, 2017 and May 22, 2018 by 3133 individuals using multiple daily injections (MDIs) or continuous subcutaneous insulin infusion (CSII) therapy were deidentified and retrospectively analyzed. Glucose excursions were defined as SG values that went beyond a preset low or high SG threshold for ≥15 minutes. For a control group, thresholds were based on the median of the low SG threshold limit (70 mg/dL) and the high SG threshold limit (210 mg/dL) preset by all system users. During periods when alerts were not enabled, timestamps were identified when a predictive alert would have been triggered. The time before low horizon was 17.5 minutes and the time before high horizon was 15 minutes, of all users who enabled alerts. Excursions occurring after a low SG or high SG predictive alert were segmented into prevented, ≤20, 20-60, and >60 minutes.Results:Excursions were prevented after 59% and 39% of low and high SG predictive alerts, respectively. The risk of a low or high excursion occurring was 1.9 (P < 0.001, 95% CI, 1.88-1.93) and 3.3 (P < 0.001, 95% CI, 3.20-3.30) times greater, respectively, when alerts were not enabled.Conclusions:The predictive alerts of the RT-CGM system under study can help individuals living with diabetes prevent some real-world low and high SG excursions. This can be especially important for those unable to reach or maintain glycemic control with basic RT-CGM or CSII therapy.     

A commonly carried genetic variant in the delta opioid receptor gene,OPRD1, is associated with smaller regional brain volumes: Replication in elderly and young populations

Delta opioid receptors are implicated in a variety of psychiatric and neurological disorders. These receptors play a key role in the reinforcing properties of drugs of abuse, and polymorphisms in OPRD1 (the gene encoding delta opioid receptors) are associated with drug addiction. Delta opioid receptors are also involved in protecting neurons against hypoxic and ischemic stress. Here, we first examined a large sample of 738 elderly participants with neuroimaging and genetic data from the Alzheimer's Disease Neuroimaging Initiative. We hypothesized that common variants in OPRD1 would be associated with differences in brain structure, particularly in regions relevant to addictive and neurodegenerative disorders. One very common variant (rs678849) predicted differences in regional brain volumes. We replicated the association of this single‐nucleotide polymorphism with regional tissue volumes in a large sample of young participants in the Queensland Twin Imaging study. Although the same allele was associated with reduced volumes in both cohorts, the brain regions affected differed between the two samples. In healthy elderly, exploratory analyses suggested that the genotype associated with reduced brain volumes in both cohorts may also predict cerebrospinal fluid levels of neurodegenerative biomarkers, but this requires confirmation. If opiate receptor genetic variants are related to individual differences in brain structure, genotyping of these variants may be helpful when designing clinical trials targeting delta opioid receptors to treat neurological disorders. Hum Brain Mapp 35:1226–1236, 2014. © 2013 Wiley Periodicals, Inc.     

Predictors of CNS injury as measured by proton magnetic resonance spectroscopy in the setting of chronic HIV infection and CART

The reasons for persistent brain dysfunction in chronically HIV-infected persons on stable combined antiretroviral therapies (CART) remain unclear. Host and viral factors along with their interactions were examined in 260 HIV-infected subjects who underwent magnetic resonance spectroscopy (MRS). Metabolite concentrations (NAA/Cr, Cho/Cr, MI/Cr, and Glx/Cr) were measured in the basal ganglia, the frontal white matter, and gray matter, and the best predictive models were selected using a bootstrap-enhanced Akaike information criterion (AIC). Depending on the metabolite and brain region, age, race, HIV RNA concentration, ADC stage, duration of HIV infection, nadir CD4, and/or their interactions were predictive of metabolite concentrations, particularly the basal ganglia NAA/Cr and the mid-frontal NAA/Cr and Glx/Cr, whereas current CD4 and the CPE index rarely or did not predict these changes. These results show for the first time that host and viral factors related to both current and past HIV status contribute to persisting cerebral metabolite abnormalities and provide a framework for further understanding neurological injury in the setting of chronic and stable disease.