Noncompaction cardiomyopathy is caused by a novel in‐frame desmin (DES) deletion mutation within the 1A coiled‐coil rod segment leading to a severe filament assembly defect

Mutations in DES, encoding desmin protein, are associated with different kinds of skeletal and/or cardiac myopathies. However, it is unknown, whether DES mutations are associated with left ventricular hypertrabeculation (LVHT). Here, we performed a clinical examination and subsequent genetic analysis in a family, with two individuals presenting LVHT with conduction disease and skeletal myopathy. The genetic analysis revealed a novel small in‐frame deletion within the DES gene, p.Q113_L115del, affecting the α‐helical rod domain. Immunohistochemistry analysis of explanted myocardial tissue from the index patient revealed an abnormal cytoplasmic accumulation of desmin and a degraded sarcomeric structure. Cell transfection experiments with wild‐type and mutant desmin verified the cytoplasmic aggregation and accumulation of mutant desmin. Cotransfection experiments were performed to model the heterozygous state of the patients and revealed a dominant negative effect of the mutant desmin on filament assembly. DES:p.Q113_L115del is classified as a pathogenic mutation associated with dilated cardiomyopathy with prominent LVHT.     

CAPN1 mutations: Expanding the CAPN1-related phenotype: From hereditary spastic paraparesis to spastic ataxia

Aims and objectiveTo characterize the phenotype of CAPN1 (SPG76) mutations in patients diagnosed with hereditary spastic paraplegia (HSP).BackgroundThe CAPN1 gene, located on chromosome 11q13.1, is a protein-coding gene involved in neuronal plasticity, migration, microtubular regulation and cerebellar development. Several families with CAPN1 mutations have recently been reported to present with autosomal recessive (AR) HSP and/or ataxia.MethodPatients with HSP were identified through neurological and genetic clinics with detailed phenotyping. Whole exome sequencing revealed novel pathogenic CAPN1 mutations in four patients from 3 families.ResultsAffected families were of Turkish, Japanese, and Punjabi descent and all were consanguineous. Onset of spastic paraplegia in the four patients was between 20 and 37 years. Two also had mild ataxia. Three different novel, homozygous mutations in CAPN1 were found: c.2118+1G > T, c.397C > T, c.843+1G > C. The patient with the earliest onset also manifested profound muscle weakness, likely related to a second homozygous mutation in DYSF (dysferlinopathy).ConclusionsThe phenotype of AR CAPN1 mutations appears to be spastic paraplegia with or without ataxia; onset is most commonly in adulthood. Eye movement abnormalities, skeletal defects, peripheral neuropathy and amyotrophy can sometimes be seen. Occasionally, patients can present with ataxia, illustrating the genotypic and phenotypic overlap between HSP and spastic ataxia. With the advent of exome sequencing, mutations in more than one gene can be identified, which may contribute to the phenotypic variation, even within a family.     

Cocaine metabolism accelerated by a re-engineered human butyrylcholinesterase

Plasma butyrylcholinesterase (BChE) is important in the metabolism of cocaine, but natural human BChE has limited therapeutic potential for detoxication because of low catalytic efficiency with cocaine. Here we report pharmacokinetics of cocaine in rats treated with A328W/Y332A BChE, an excellent cocaine hydrolase designed with the aid of molecular modeling. Compared with wild-type BChE, this enzyme hydrolyzes cocaine with 40-fold improved k(cat) (154 min(-1) versus 4.1 min(-1)) and only slightly increased K(M) (18 microM versus 4.5 microM). In rats given this hydrolase (3 mg/kg i.v.) 10 min before cocaine challenge (6.8 mg/kg i.v.), cocaine half-life was reduced from 52 min to 18 min. Mirroring the reductions of plasma cocaine were large increases in benzoic acid, a product of BChE-mediated cocaine hydrolysis. All other pharmacokinetic parameters confirmed a large, dose-dependent acceleration of cocaine removal by the injected cocaine hydrolase. These results show that A328W/Y332A, an efficient cocaine hydrolase in vivo as well as in vitro, might promote cocaine detoxication in a clinical setting.     

Wild-type and A328W mutant human butyrylcholinesterase tetramers expressed in Chinese hamster ovary cells have a 16-hour half-life in the circulation and protect mice from cocaine toxicity

Human butyrylcholinesterase (BChE) hydrolyzes cocaine to inactive metabolites. A mutant of human BChE, A328W, hydrolyzed cocaine 15-fold faster compared with wild-type BChE. Although the catalytic properties of human BChE secreted by Chinese hamster ovary (CHO) cells are identical to those of native BChE, a major difference became evident when the recombinant BChE was injected into rats and mice. Recombinant BChE disappeared from the circulation within minutes, whereas native BChE stayed in the blood for a week. Nondenaturing gel electrophoresis showed that the recombinant BChE consisted mainly of monomers and dimers. In contrast, native BChE is a tetramer. The problem of the short residence time was solved by finding a method to assemble the recombinant BChE into tetramers. Coexpression in CHO cells of BChE and 45 residues from the N terminus of the COLQ protein yielded 70% tetrameric BChE. The resulting purified recombinant BChE tetramers had a half-life of 16 h in the circulation of rats and mice. The 16-h half-life was achieved without modifying the carbohydrate content of recombinant BChE. The protective effect of recombinant wild-type and A328W mutant BChE against cocaine toxicity was tested by measuring locomotor activity in mice. Pretreatment with wild-type BChE or A328W tetramers at a dose of 2.8 units/g i.p. reduced cocaine-induced locomotor activity by 50 and 80%. These results indicate that recombinant human BChE could be useful for treating cocaine toxicity in humans.     

Adrenalectomy improves diabetes in A-ZIP/F-1 lipoatrophic mice by increasing both liver and muscle insulin sensitivity

The virtually fatless A-ZIP/F-1 mouse is profoundly insulin resistant, diabetic, and a good model for humans with severe generalized lipoatrophy. Like a number of other mouse models of diabetes, the A-ZIP/F-1 mouse has elevated serum corticosterone levels. Leptin infusion lowers the corticosterone levels, suggesting that leptin deficiency contributes to the hypercorticosteronemic state. To test the hypothesis that the increased glucocorticoids contribute to the diabetes and insulin resistance, we examined the effect of adrenalectomy on A-ZIP/F-1 mice. Adrenalectomy significantly decreased the blood glucose, serum insulin, and glycated hemoglobin levels. Hyperinsulinemic-euglycemic clamps were performed to characterize the changes in whole-body and tissue insulin sensitivity. The adrenalectomized A-ZIP/F-1 mice displayed a marked improvement in insulin-induced suppression of endogenous glucose production, indicating increased hepatic insulin sensitivity. Adrenalectomy also increased muscle glucose uptake and glycogen synthesis. These results suggest that the chronically increased serum corticosterone levels contribute to the diabetes of the A-ZIP/F-1 mice and that removal of the glucocorticoid excess improves the insulin sensitivity in both muscle and liver.     

Statins ameliorate endothelial barrier permeability changes in the cerebral tissue of streptozotocin-induced diabetic rats

Statins may have favorable effects on endothelial barrier function. The effect of rosuvastatin and simvastatin therapy (10 mg/kg) for 5 weeks on blood-brain barrier (BBB), blood-retinal barrier (BRB), and cardiac muscle permeability of streptozotocin-induced diabetic rats was studied. The size-selective permeability of different vascular beds to a group of fluorescein isothiocyanate dextrans of varying molecular weights was measured. The volume of distribution of 250-, 70-, and 40-kDa dextrans in the cerebral tissue of diabetic rats were significantly increased. The volume of distribution of these dextrans in cerebral tissue was normalized by both statins. Diabetes did not significantly alter the BRB, but both statins decreased the volume of distribution of 70- and 40-kDa dextrans in the retina. The volume of distribution of 40 kDa in cardiac muscle was increased in diabetes, and this change was prevented with statin treatment. Treatment with rosuvastatin and mevalonate (150 mg/kg in drinking water for 5 weeks) did not alter the volume of distribution measurements. We concluded that 1) diabetes in rats is associated with significant changes in the BBB permeability; 2) statin treatment improves the endothelial barrier function in cerebral tissue, retina, and cardiac muscle; and 3) this statin effect could not be attributed to HMGCoA reductase inhibition.     

Insulin resistance in the liver-specific IGF-1 gene-deleted mouse is abrogated by deletion of the acid-labile subunit of the IGF-binding protein-3 complex: relative roles of growth hormone and IGF-1 in insulin resistance

Liver IGF-1 deficient (LID) mice demonstrate a 75% reduction in circulating IGF-1 levels and a corresponding fourfold increase in growth hormone (GH) levels. At 16 weeks of age, LID mice demonstrate, using the hyperinsulinemic-euglycemic clamp, insulin insensitivity in muscle, liver, and fat tissues. In contrast, mice with a gene deletion of the acid-labile subunit (ALSKO) demonstrate a 65% reduction in circulating IGF-1 levels, with normal GH levels and no signs of insulin resistance. To further clarify the relative roles of increased GH and decreased IGF-1 levels in the development of insulin resistance, we crossed the two mouse lines and created a double knockout mouse (LID+ALSKO). LID+ALSKO mice demonstrate a further reduction in circulating IGF-1 levels (85%) and a concomitant 10-fold increase in GH levels. Insulin tolerance tests showed an improvement in insulin responsiveness in the LID+ALSKO mice compared with controls; LID mice were very insulin insensitive. Surprisingly, insulin sensitivity, while improved in white adipose tissue and in muscle, was unchanged in the liver. The lack of improvement in liver insulin sensitivity may reflect the absence of IGF-1 receptors or increased triglyceride levels in the liver. The present study suggests that whereas GH plays a major role in inducing insulin resistance, IGF-1 may have a direct modulatory role.     

Large-scale comparative genomics meta-analysis of Campylobacter jejuni isolates reveals low level of genome plasticity

We have used comparative genomic hybridization (CGH) on a full-genome Campylobacter jejuni microarray to examine genome-wide gene conservation patterns among 51 strains isolated from food and clinical sources. These data have been integrated with data from three previous C. jejuni CGH studies to perform a meta-analysis that included 97 strains from the four separate data sets. Although many genes were found to be divergent across multiple strains (n = 350), many genes (n = 249) were uniquely variable in single strains. Thus, the strains in each data set comprise strains with a unique genetic diversity not found in the strains in the other data sets. Despite the large increase in the collective number of variable C. jejuni genes (n = 599) found in the meta-analysis data set, nearly half of these (n = 276) mapped to previously defined variable loci, and it therefore appears that large regions of the C. jejuni genome are genetically stable. A detailed analysis of the microarray data revealed that divergent genes could be differentiated on the basis of the amplitudes of their differential microarray signals. Of 599 variable genes, 122 could be classified as highly divergent on the basis of CGH data. Nearly all highly divergent genes (117 of 122) had divergent neighbors and showed high levels of intraspecies variability. The approach outlined here has enabled us to distinguish global trends of gene conservation in C. jejuni and has enabled us to define this group of genes as a robust set of variable markers that can become the cornerstone of a new generation of genotyping methods that use genome-wide C. jejuni gene variability data.     

Presenilin 1 Glu318Gly polymorphism: interpret with caution

BACKGROUND: The significance of the presenilin 1 (PSEN1) Glu318Gly polymorphism has been described as either a causal mutation with reduced penetrance or a benign polymorphism. When this polymorphism is found in a symptomatic person with a family history of dementia, counseling on recurrence risk becomes very problematic. OBJECTIVE: To demonstrate that the PSEN1 Glu318Gly polymorphism should be interpreted cautiously. DESIGN: Case histories of 2 patients with presenile dementia and family histories of dementia are described. The PSEN1 gene was sequenced in the patients and in 11 family members of patient 1. RESULTS: Two patients with presenile dementia and personality change were found to carry the PSEN1 Glu318Gly polymorphism. The presence of the polymorphism was confirmed in several family members of patient 1 but was absent in 1 symptomatic relative. CONCLUSIONS: The Glu318Gly polymorphism may be associated with risk for neurodegenerative disease; however, in the cases described here, it did not appear to be a risk factor. Until there is consensus on whether it is associated with disease, families should be informed that the clinical significance of the polymorphism is uncertain.