Mutation Screening of the HGD Gene Identifies a Novel Alkaptonuria Mutation with Significant Founder Effect and High Prevalence

Alkaptonuria (AKU) is an autosomal recessive disorder; caused by the mutations in the homogentisate 1, 2‐dioxygenase (HGD) gene located on Chromosome 3q13.33. AKU is a rare disorder with an incidence of 1: 250,000 to 1: 1,000,000, but Slovakia and the Dominican Republic have a relatively higher incidence of 1: 19,000. Our study focused on studying the frequency of AKU and identification of HGD gene mutations in nomads. HGD gene sequencing was used to identify the mutations in alkaptonurics. For the past four years, from subjects suspected to be clinically affected, we found 16 positive cases among a randomly selected cohort of 41 Indian nomads (Narikuravar) settled in the specific area of Tamil Nadu, India. HGD gene mutation analysis showed that 11 of these patients carry the same homozygous splicing mutation c.87 + 1G > A; in five cases, this mutation was found to be heterozygous, while the second AKU‐causing mutation was not identified in these patients. This result indicates that the founder effect and high degree of consanguineous marriages have contributed to AKU among nomads. Eleven positive samples were homozygous for a novel mutation c.87 + 1G > A, that abolishes an intron 2 donor splice site and most likely causes skipping of exon 2. The prevalence of AKU observed earlier seems to be highly increased in people of nomadic origin.     

Correlation of creatinine‐ and specific gravity‐normalized free and glucuronidated urine cannabinoid concentrations following smoked,vaporized, and oral cannabis in frequent and occasional cannabis users

Variability in urine dilution complicates urine cannabinoid test interpretation. Normalizing urine cannabinoid concentrations to specific gravity (SG) or creatinine was proposed to account for donors' hydration states. In this study, all urine voids were individually collected from eight frequent and eight occasional cannabis users for up to 85 hours after each received on separate occasions 50.6 mg Δ9‐tetrahydrocannabinol (THC) by smoking, vaporization, and oral ingestion in a randomized, within‐subject, double‐blind, double‐dummy, placebo‐controlled protocol. Each urine void was analyzed for 11 cannabinoids and phase I and II metabolites by liquid chromatography?tandem mass spectrometry (LC–MS/MS), SG, and creatinine. Normalized urine concentrations were log₁₀ transformed to create normal distributions, and Pearson correlation coefficients determined the degree of association between the two normalization methods. Repeated‐measures linear regression determined if the degree of association differed by frequent or occasional cannabis use, or route of administration after adjusting for gender and time since dosing. Of 1880 urine samples examined, only 11‐nor‐9‐carboxy‐THC (THCCOOH), THCCOOH‐glucuronide, THC‐glucuronide, and 11‐nor‐9‐carboxy‐Δ9‐tetrahydrocannabivarin (THCVCOOH) were greater than the method's limits of quantification (LOQs). Associations between SG‐ and creatinine‐normalized concentrations exceeded 0.90. Repeated‐measures regression analysis found small but statistically significant differences in the degree of association between normalization methods for THCCOOH and THCCOOH‐glucuronide in frequent vs occasional smokers, and in THCVCOOH and THC‐glucuronide by route of administration. For the first time, SG‐ and creatinine‐normalized urine cannabinoid concentrations were evaluated in frequent and occasional cannabis users and following oral, smoked, and inhaled cannabis. Both normalization methods reduced variability, improving the interpretation of urine cannabinoid concentrations and methods were strongly correlated.     

Cardiac autonomic neuropathy predicts renal function decline in patients with type 2 diabetes: a cohort study

Aims/hypothesis

The aim of this work was to assess the impact of cardiac autonomic neuropathy (CAN) on the development and progression of chronic kidney disease (CKD) in patients with type 2 diabetes.

Methods

We conducted a cohort study in adults with type 2 diabetes. Patients with end-stage renal disease were excluded. CKD was defined as the presence of albuminuria (albumin/creatinine ratio GFR >?3.4 mg/mmol) or an estimated (eGFR) <?60 ml min^?1 1.73 m^?2. CKD progression was based on repeated eGFR measurements and/or the development of albuminuria. CAN was assessed using heart rate variability.

Results

Two hundred and four patients were included in the analysis. At baseline, the prevalence of CKD and CAN was 40% and 42%, respectively. Patients with CAN had lower eGFR and higher prevalence of albuminuria and CKD. Spectral analysis variables were independently associated with eGFR, albuminuria and CKD at baseline. After a follow-up of 2.5 years, eGFR declined to a greater extent in patients with CAN than in those without CAN (?9.0?±?17.8% vs ?3.3?±?10.3%, p?=?0.009). After adjustment for baseline eGFR and baseline differences, CAN remained an independent predictor of eGFR decline over the follow-up period (β?=??3.5, p?=?0.03). Spectral analysis variables were also independent predictors of eGFR decline.

Conclusions/interpretation

CAN was independently associated with CKD, albuminuria and eGFR in patients with type 2 diabetes. In addition, CAN was an independent predictor of the decline in eGFR over the follow-up period. CAN could be used to identify patients with type 2 diabetes who are at increased risk of rapid decline in eGFR, so that preventative therapies might be intensified.     

Direct evidence for a magnetic f-electron–mediated pairing mechanism of heavy-fermion superconductivity in CeCoIn5

To identify the microscopic mechanism of heavy-fermion Cooper pairing is an unresolved challenge in quantum matter studies; it may also relate closely to finding the pairing mechanism of high-temperature superconductivity. Magnetically mediated Cooper pairing has long been the conjectured basis of heavy-fermion superconductivity but no direct verification of this hypothesis was achievable. Here, we use a novel approach based on precision measurements of the heavy-fermion band structure using quasiparticle interference imaging to reveal quantitatively the momentum space (k-space) structure of the f-electron magnetic interactions of CeCoIn₅. Then, by solving the superconducting gap equations on the two heavy-fermion bands Ekα,β with these magnetic interactions as mediators of the Cooper pairing, we derive a series of quantitative predictions about the superconductive state. The agreement found between these diverse predictions and the measured characteristics of superconducting CeCoIn₅ then provides direct evidence that the heavy-fermion Cooper pairing is indeed mediated by f-electron magnetism.Superconductivity of heavy fermions is of abiding interest, both in its own right (1–7) and because it could exemplify the unconventional Cooper pairing mechanism of high-temperature superconductors (8–11). Heavy-fermion compounds are intermetallics containing magnetic ions in the 4f- or 5f-electronic state within each unit cell. At high temperatures, each f-electron is localized at a magnetic ion (Fig. 1A). At low temperatures, interactions between f-electron spins (red arrows Fig. 1A) lead to the formation of a narrow but the subtly curved f-electron band εkf near the chemical potential (red curve, Fig. 1B), and Kondo screening hybridizes this band with the conventional c-electron band εkc of the metal (black curve, Fig. 1B). As a result, two new heavy-fermion bands Ekα,β (Fig. 1C) appear within a few millielectron volts of the Fermi energy. Their electronic structure is controlled by the hybridization matrix element s_k for interconversion of conduction c-electrons to f-electrons and vice-versa, such thatEkα,β=εkc+εkf2±(εkc−εkf2)2+sk2.[1]The momentum structure of the narrow bands of hybridized electronic states (Eq. 1 and Fig. 1C, blue curves at left) near the Fermi surface then directly reflects the form of magnetic interactions encoded within the parent f-electron band εkf. It is these interactions that are conjectured to drive the Cooper pairing (1–5) and thus the opening up of a superconducting energy gap (Fig. 1C, yellow curves at right).Open in a separate windowFig. 1.Effects of f-electron magnetism in a heavy-fermion material. (A) The magnetic subsystem of CeCoIn₅ consists of almost localized magnetic f-electrons (red arrows) with a weak hopping matrix element yielding a very narrow band with strong magnetic interactions between the f-electron spins. (B) The heavy f-electron band is shown schematically in red and the light c-electron band in black. (C) On the left, schematic of the result of hybridizing the c- and f-electrons in B into new composite electronic states referred to as heavy fermions (blue). On the right, the opening of a superconducting energy gap is schematically shown by back-bending bands near the chemical potential. The microscopic interactions driving Cooper pairing of these states, and thus of heavy-fermion superconductivity, have not been identified unambiguously for any heavy-fermion compound.     

Uterine development and fertility are dependent on gene dosage of the nuclear receptor coregulator REA

Although the effectiveness of nuclear hormone-receptor complexes is known to depend on coregulator partner proteins, relatively little is known about the roles of coregulators in uterine development and early stages of pregnancy and implantation. Because conventional genetic deletion of the coregulator, repressor of estrogen receptor activity (REA), was embryonic lethal, we here study REA conditional knockout mice generated by cre-loxP recombination, in which REA function was abrogated only in progesterone receptor-expressing tissues, to define the roles of REA in postembryonic stages and in a tissue-specific manner. We find that REA has gene dose-dependent activity impacting uterine development and fertility. Conditional homozygous mutant (REA(d/d)) mice developed to adulthood and showed normal ovarian function, but females were infertile with severely compromised uterine development and function characterized by cell cycle arrest, apoptosis, and altered adenogenesis (endometrial gland morphogenesis), resulting in failure of implantation and decidualization. By contrast, mice heterozygous for REA (REA(f/d)) had a very different phenotype, with estradiol treatment resulting in hyperstimulated, large uteri showing increased proliferation of luminal epithelial cells, and enhanced fluid imbibition associated with altered regulation of aquaporins. These REA(f/d) female mice showed a subfertility phenotype with reduced numbers and sizes of litters. These findings highlight that uterine development and regulation of estrogen receptor activities show a bimodal dependence on the gene dosage of REA. Optimal uterine development and functional activities require the normal gene dosage of REA, with partial or complete deletion resulting in hyperresponsiveness or underresponsiveness to hormone and subfertility or infertility, respectively.     

YKL-40 is elevated in patients with peripheral arterial disease and diabetes or pre-diabetes

ObjectiveYKL-40 is secreted by macrophages in atherosclerotic lesions and involved in plaque rupture. YKL-40 is elevated in coronary artery disease, and predicts cardiovascular mortality. Experimental in vivo and in vitro data suggest a role of YKL-40 in tissue remodeling. A disease modulating potency of YKL-40 was not investigated in peripheral arterial disease (PAD).MethodsWe measured YKL-40 in 460 subjects: 316 PAD: 71 normal glucose metabolism (PAD-NGM), 90 pre-diabetes (PAD-PREDM) and 155 diabetes (PAD-DM); 20 diabetes with atherosclerosis but without PAD (AS-DM); 85 diabetes without macro-vascular complications (DM) and 39 healthy controls (CO).ResultsYKL-40 is higher in PAD vs. CO (median [25–75 percentile]: 103 [69–159] vs. 43 [30–80] ng/ml; p < 0.001). In addition, YKL-40 is elevated in DM (p < 0.001), PAD-NGM (p = 0.001), PAD-PREDM (p < 0.001), PAD-DM (p < 0.001) and AS-DM (p = 0.002) compared to CO. Among PAD, YKL-40 is increased in PAD-PREDM (p = 0.001) and PAD-DM (p = 0.01) vs. PAD-NGM. By multivariate regression YKL-40 is significantly associated with age (beta = 0.272), triglycerides (beta = 0.216), aspartate-amino-transferase (beta = 0.177) and c-reactive-protein (beta = 0.178). Underpinning its role YKL-40 was found to be associated with micro-/macroalbuminuria (p = 0.014/p = 008) – a strong remodeling inducer. In addition, YKL-40 was elevated in existence of mediasclerosis (p = 0.008), a remodeling process.ConclusionWe are first to show that YKL-40 is higher in subjects with peripheral arterial disease. YKL-40 was higher in PAD patients with pre-/diabetes. In addition, YKL-40 was associated with the “severity” of generalized atherosclerosis estimated by affected vascular beds. All our findings point towards a role of YKL-40 in the progression/prognosis of patients with PAD and concomitant diabetes.     

Role of dimethylarginine dimethylaminohydrolase activity in regulation of tissue and plasma concentrations of asymmetric dimethylarginine in an animal model of prolonged critical illness

High plasma concentrations of asymmetric dimethylarginine (ADMA), an endogenous nitric oxide synthase inhibitor, are associated with adverse outcome in critically ill patients. Asymmetric dimethylarginine is released within cells during proteolysis of methylated proteins and is either degraded by dimethylarginine dimethylaminohydrolase (DDAH) or exported to the circulation via cationic amino acid transporters. We aimed to establish the role of DDAH activity in the regulation of tissue and plasma concentrations of ADMA. In 33 critically ill rabbits, we measured DDAH activity in kidney, liver, heart, and skeletal muscle and related these values to concentrations of ADMA in these tissues and in the circulation. Both DDAH activity and ADMA concentration were highest in kidney and lowest in skeletal muscle, with intermediate values for liver and heart. Whereas ADMA content was significantly correlated between tissues (r = 0.40-0.78), DDAH activity was not. Significant inverse associations between DDAH activity and ADMA content were only observed in heart and liver. Plasma ADMA was significantly associated with ADMA in the liver (r = 0.41), but not in the other tissues. In a multivariable regression model, DDAH activities in muscle, kidney, and liver, but not in heart, were negatively associated with plasma ADMA concentration, together explaining approximately 50% of its variation. In critical illness, plasma ADMA poorly reflects intracellular ADMA. Furthermore, tissue DDAH activity is a stronger predictor of plasma ADMA than of intracellular ADMA, indicating that, compared with DDAH activity, generation of ADMA and cationic amino acid transporter-mediated exchange may be more important regulators of intracellular ADMA.