TRPC6 Single Nucleotide Polymorphisms and Progression of Idiopathic Membranous Nephropathy

Background

Activating mutations in the Transient Receptor Potential channel C6 (TRPC6) cause autosomal dominant focal segmental glomerular sclerosis (FSGS). TRPC6 expression is upregulated in renal biopsies of patients with idiopathic membranous glomerulopathy (iMN) and animal models thereof. In iMN, disease progression is characterized by glomerulosclerosis. In addition, a context-dependent TRPC6 overexpression was recently suggested in complement-mediated podocyte injury in e.g. iMN. Hence, we hypothesized that genetic variants in TRPC6 might affect susceptibility to development or progression of iMN.

Methods & Results

Genomic DNA was isolated from blood samples of 101 iMN patients and 292 controls. By direct sequencing of the entire TRPC6 gene, 13 single nucleotide polymorphisms (SNPs) were identified in the iMN cohort, two of which were causing an amino acid substitution (rs3802829; Pro15Ser and rs36111323, Ala404Val). No statistically significant differences in genotypes or allele frequencies between patients and controls were observed. Clinical outcome in patients was determined (remission n = 26, renal failure n = 46, persistent proteinuria n = 29, follow-up median 80 months {range 51–166}). The 13 identified SNPs showed no association with remission or renal failure. There were no differences in genotypes or allele frequencies between patients in remission and progressors.

Conclusions

Our data suggest that TRPC6 polymorphisms do not affect susceptibility to iMN, or clinical outcome in iMN.     

“First experience with JenaValve?: a single-centre cohort”

Aims

Since the introduction of transcatheter aortic valve implantation (TAVI), newer generation and novel devices such as the retrievable JenaValve™ have been developed. We evaluated the procedural and 6-month results of our first experience with implantation of the JenaValve™.

Methods and results

From June 2012 to December 2013, 24 consecutive patients (mean age 80 ± 7 years, 42 % male) underwent an elective transapical TAVI with the JenaValve™. Device success was 88 %. The mortality rate was 4 % at 30 days and 31 % at 6 months. TAVI reduced the mean transvalvular gradient (44.2 ± 11.1 mmHg vs. 12.3 ± 4.3 mmHg, p < 0.001) and increased the mean aortic valve area (0.8 3 ± 0.23 to 1.70 ± 0.44 cm²). A mild paravalvular leakage (PVL) occurred in 4 patients (18 %) and a moderate PVL in 1 patient (4 %). Mean New York Heart Association Functional Class improved from 2.9 ± 0.5 to 2.0 ± 0.8 at 30 days.

Conclusion

TAVI using the JenaValve™ prosthesis seems adequate and safe in this first experience cohort.     

Feasibility of multiplane microtransoesophageal echocardiographic guidance in structural heart disease transcatheter interventions in adults

Introduction

Structural heart interventions are guided by transoesophageal or intracardiac echocardiography (TEE/ICE). MicroTEE, developed for paediatric purposes, is smaller and therefore less invasive and traumatic, avoiding the need for general anaesthesia. We aimed to show feasibility of procedural guidance by comparing image quality of microTEE with standard TEE and ICE during adult transcatheter interventions, and assess the accuracy in obtaining left atrial appendage (LAA) measurements between the microTEE probe and standard TEE.

Methods and results

We prospectively included 49 patients (20 women, 64 ± 18 years). Intraprocedural images were obtained by using the microTEE probe and standard (2D and 3D) TEE (LAA closure, MitraClip implantation) or ICE (interatrial communication closure, transseptal puncture for left atrial ablation). Two echocardiographers independently assessed image quality from 1 (excellent) to 4 (poor) and performed LAA measurements. Use of microTEE was not related to significant discomfort. Image quality obtained with the microTEE probe was lower than with standard TEE (2 [1–2] vs. 1 [1–2]; p = 0.04) and comparable with ICE images (2 [1–2] vs. 2 [1–2], p = 0.13). MicroTEE showed a wider field of view than ICE. LAA measurements on images obtained by microTEE were strongly associated with standard TEE.

Conclusions

MicroTEE seems feasible for guidance during transcatheter heart interventions in adults. MicroTEE imaging offers a wider field of view than ICE, and its accuracy is comparable with TEE. In transcatheter interventions performed under conscious sedation, microTEE might be a viable and advantageous alternative to standard TEE or ICE.

     

Synthesis of a novel potent cyclic peptide MC4-ligand by ring-closing metathesis

The synthesis of a novel potent cyclic peptide MC4-ligand by ring-closing metathesis (RCM) is described. Based on the Ac-Nle-Gly-Lys-D-Phe-Arg-Trp-Gly-NH2-MC4 ligand, Ac-Nle-Alg-Lys-D-Phe-Arg-Trp-Alg-NH2 was designed and synthesized followed by cyclization using RCM. Both compounds are high affinity and selective MC4-R-agonists. The cyclic RCM-peptide was more potent in a rat-grooming assay.     

Characterization of hydrogenase and reductive dehalogenase activities of Dehalococcoides ethenogenes strain 195

Dehalococcoides ethenogenes strain 195 reductively dechlorinates tetrachloroethene (PCE) and trichloroethene (TCE) to vinyl chloride and ethene using H2 as an electron donor. PCE- and TCE-reductive dehalogenase (RD) activities were mainly membrane associated, whereas only about 20% of the hydrogenase activity was membrane associated. Experiments with methyl viologen (MV) were consistent with a periplasmic location for the RDs or a component feeding electrons to them. The protonophore uncoupler tetrachlorosalicylanilide did not inhibit reductive dechlorination in cells incubated with H2 and PCE and partially restored activity in cells incubated with the ATPase inhibitor N,N'-dicyclohexylcarbodiimide. Benzyl viologen or diquat (Eo' approximately -360 mV) supported reductive dechlorination of PCE or TCE at rates comparable to MV (-450 mV) in cell extracts.     

Myasthenia Gravis,Autoantibodies, and HL-A Antigens

The serum of 100 patients with myasthenia gravis and 441 of their first-degree relatives was studied for the presence of autoantibodies against several antigens. Antibodies to skeletal muscle were present in 22% of the patients and in 2% of the relatives. Both these frequencies were significantly higher than those in matched control subjects. Also, antinuclear antibodies were present more often both in the patients and in the relatives. Typing for HL-A antigens had shown a positive correlation between HL-A 8 and myasthenia gravis which was significantly higher in women than in men. Antibodies to skeletal muscle and thymomas were found to be much rarer in HL-A 8-positive patients than in HL-A 8-negative patients; HL-A 8-positive patients acquired the disease at an earlier age.HL-A 2-positive patients more often had thymomas and antibodies to skeletal muscle than HL-A 2-negative patients; HL-A 2-positive patients acquired myasthenia gravis at a later age.The fact that the clinical aspects of the HL-A 8-negative and HL-A 2-positive patients were different from those of the HL-A 8-positive and HL-A 2-negative patients justifies the hypothesis that there are two forms of myasthenia gravis.     

Familial benign hypercalcaemia (FBH; McK. No. 14598, 1983): Linkage studies in a large Dutch family

Summary By screening 27 hypercalcaemic and 21 normocalcaemic subjects in a large Dutch pedigree with familial benign hypercalcaemia (FBH; McK. No. 14598) (McKusick 1983) for more than 35 genetic markers, it was found that linkage of FBH can be excluded at about 25 centimorgans (cM) from GM, 20 cM from ABO, 15 cM from MNS and HLA, 10cM from JK and PI, and 5cM each from ACP1, AK1, ADA, GPT1, and PGP.     

ABAP: antibody-based assay for peptidylarginine deiminase activity

Members of the family of peptidylarginine deiminases (PADs, EC 3.5.3.15) catalyze the posttranslational modification of peptidylarginine into peptidylcitrulline. Citrulline-containing epitopes have been shown to be major and specific targets of autoantibodies produced by rheumatoid arthritis patients. Recently, the citrullination of histone proteins by PAD enzyme was reported to influence gene expression levels. These findings greatly increase the interest in the PAD enzymes and their activities. A few procedures to monitor PAD activity in biological samples have been described previously. However, these assays either have low sensitivity or are rather laborious. Here we describe a reliable and reproducible method for the determination of PAD activity in both purified and crude samples. The method is based on the quantification of PAD-dependent citrullination of peptides, immobilized in microtiter plates, using antibodies that are exclusively reactive with the reaction product(s). Our results demonstrate that this antibody-based assay for PAD activity, called ABAP, is very sensitive and can be applied to monitor PAD activity in biological samples.     

Common requirements for melanocortin-4 receptor selectivity of structurally unrelated melanocortin agonist and endogenous antagonist, Agouti protein

The activity of melanocortin receptors (MCR) is regulated by melanocortin peptide agonists and by the endogenous antagonists, Agouti protein and AgRP (Agouti-related protein). To understand how the selectivity for these structurally unrelated agonists and antagonist is achieved, chimeric and mutants MC3R and MC4R were expressed in cell lines and pharmacologically analyzed. A region containing the third extracellular loop, EC3, of MC4R was essential for selective Agouti protein antagonism. In addition, this part of MC4R, when introduced in MC3R, conferred Agouti protein antagonism. Further mutational analysis of this region of MC4R demonstrated that Tyr(268) was required for the selective interaction with Agouti protein, because a profound loss of the ability of Agouti protein to inhibit (125)I-labeled [Nle(4),d-Phe(7)]alpha-melanocyte-stimulating hormone (MSH) binding was observed by the single mutation of Tyr(268) to Ile. This same residue conferred selectivity for the MC4R selective agonist, [d-Tyr(4)]MT-II, whereas it inhibited interaction with the MC3R-selective agonist, [Nle(4)]Lys-gamma(2)-MSH. Conversely, mutation of Ile(265) in MC3 (the corresponding residue of Tyr(268)) to Tyr displayed a gain of affinity for [d-Tyr(4)]MT-II, but not for Agouti protein, and a loss of affinity for [Nle(4)]Lys-gamma(2)-MSH as compared with wild-type MC3R. This single amino acid mutation thus confers the selectivity of MC3R toward a pharmacological profile like that observed for MC4R agonists but not for the antagonist, Agouti protein. Thus, selectivity for structurally unrelated ligands with opposite activities is achieved in a similar manner for MC4R but not for MC3R.