A 3-D model of coronary vessel development

Coronary vascular disease is one of the leading causes of mortality and morbidity in the United States. Therefore, a mechanistic understanding of coronary vessel morphogenesis would aid in the innovation of new therapies targeting vascular disorders. Moreover, a functionally equivalent in vitro model system allows for the delineation of the molecular mechanisms that regulate coronary vessel development. In this study, we present a novel in vitro model system. This three-dimensional (3-D) model system consists of a tubular scaffold, which is engineered from type-I collagen and has been optimized to support the growth of embryonic cardiac tissues. In this report, proepicardial (PE) cells, the developmental precursors of coronary vessels, have been isolated from several model species and cultured on this scaffold. In this model system, the PE cells were able to recapitulate several aspects of coronary vessel morphogenesis including epicardial formation, the epicardial to mesenchymal transformation, and de novo coronary vessel development or vasculogenesis. The differentiation of PE cells was characterized using a variety of specific protein markers. The potential uses of this novel coronary developmental model are discussed.     

Modeling <Emphasis Type="Italic">Neisseria meningitidis</Emphasis> metabolism: from genome to metabolic fluxes

Background  

Neisseria meningitidis is a human pathogen that can infect diverse sites within the human host. The major diseases caused by N. meningitidis are responsible for death and disability, especially in young infants. In general, most of the recent work on N. meningitidis focuses on potential antigens and their functions, immunogenicity, and pathogenicity mechanisms. Very little work has been carried out on Neisseria primary metabolism over the past 25 years.     

Dominant Mutations in KAT6A Cause Intellectual Disability with Recognizable Syndromic Features

Through a multi-center collaboration study, we here report six individuals from five unrelated families, with mutations in KAT6A/MOZ detected by whole-exome sequencing. All five different de novo heterozygous truncating mutations were located in the C-terminal transactivation domain of KAT6A: {"type":"entrez-nucleotide","attrs":{"text":"NM_001099412.1","term_id":"150378462","term_text":"NM_001099412.1"}}NM_001099412.1: c.3116_3117 delCT, p.(Ser1039^∗); c.3830_3831insTT, p.(Arg1278Serfs^∗17); c.3879 dupA, p.(Glu1294Argfs^∗19); c.4108G>T p.(Glu1370^∗) and c.4292 dupT, p.(Leu1431Phefs^∗8). An additional subject with a 0.23 MB microdeletion including the entire KAT6A reading frame was identified with genome-wide array comparative genomic hybridization. Finally, by detailed clinical characterization we provide evidence that heterozygous mutations in KAT6A cause a distinct intellectual disability syndrome. The common phenotype includes hypotonia, intellectual disability, early feeding and oromotor difficulties, microcephaly and/or craniosynostosis, and cardiac defects in combination with subtle facial features such as bitemporal narrowing, broad nasal tip, thin upper lip, posteriorly rotated or low-set ears, and microretrognathia. The identification of human subjects complements previous work from mice and zebrafish where knockouts of Kat6a/kat6a lead to developmental defects.     

Simulation of human atherosclerotic femoral plaque tissue: the influence of plaque material model on numerical results

Background

Due to the limited number of experimental studies that mechanically characterise human atherosclerotic plaque tissue from the femoral arteries, a recent trend has emerged in current literature whereby one set of material data based on aortic plaque tissue is employed to numerically represent diseased femoral artery tissue. This study aims to generate novel vessel-appropriate material models for femoral plaque tissue and assess the influence of using material models based on experimental data generated from aortic plaque testing to represent diseased femoral arterial tissue.

Methods

Novel material models based on experimental data generated from testing of atherosclerotic femoral artery tissue are developed and a computational analysis of the revascularisation of a quarter model idealised diseased femoral artery from a 90% diameter stenosis to a 10% diameter stenosis is performed using these novel material models. The simulation is also performed using material models based on experimental data obtained from aortic plaque testing in order to examine the effect of employing vessel appropriate material models versus those currently employed in literature to represent femoral plaque tissue.

Results

Simulations that employ material models based on atherosclerotic aortic tissue exhibit much higher maximum principal stresses within the plaque than simulations that employ material models based on atherosclerotic femoral tissue. Specifically, employing a material model based on calcified aortic tissue, instead of one based on heavily calcified femoral tissue, to represent diseased femoral arterial vessels results in a 487 fold increase in maximum principal stress within the plaque at a depth of 0.8 mm from the lumen.

Conclusions

Large differences are induced on numerical results as a consequence of employing material models based on aortic plaque, in place of material models based on femoral plaque, to represent a diseased femoral vessel. Due to these large discrepancies, future studies should seek to employ vessel-appropriate material models to simulate the response of diseased femoral tissue in order to obtain the most accurate numerical results.

     

Review and classification of variability analysis techniques with clinical applications

Background

Representation of independent biophysical sources using Fourier analysis can be inefficient because the basis is sinusoidal and general. When complex fractionated atrial electrograms (CFAE) are acquired during atrial fibrillation (AF), the electrogram morphology depends on the mix of distinct nonsinusoidal generators. Identification of these generators using efficient methods of representation and comparison would be useful for targeting catheter ablation sites to prevent arrhythmia reinduction.

Method

A data-driven basis and transform is described which utilizes the ensemble average of signal segments to identify and distinguish CFAE morphologic components and frequencies. Calculation of the dominant frequency (DF) of actual CFAE, and identification of simulated independent generator frequencies and morphologies embedded in CFAE, is done using a total of 216 recordings from 10 paroxysmal and 10 persistent AF patients. The transform is tested versus Fourier analysis to detect spectral components in the presence of phase noise and interference. Correspondence is shown between ensemble basis vectors of highest power and corresponding synthetic drivers embedded in CFAE.

Results

The ensemble basis is orthogonal, and efficient for representation of CFAE components as compared with Fourier analysis (p ≤ 0.002). When three synthetic drivers with additive phase noise and interference were decomposed, the top three peaks in the ensemble power spectrum corresponded to the driver frequencies more closely as compared with top Fourier power spectrum peaks (p ≤ 0.005). The synthesized drivers with phase noise and interference were extractable from their corresponding ensemble basis with a mean error of less than 10%.

Conclusions

The new transform is able to efficiently identify CFAE features using DF calculation and by discerning morphologic differences. Unlike the Fourier transform method, it does not distort CFAE signals prior to analysis, and is relatively robust to jitter in periodic events. Thus the ensemble method can provide a useful alternative for quantitative characterization of CFAE during clinical study.     

Importance of temporal flow gradients and integrin alphaIIbbeta3 mechanotransduction for shear activation of platelets

Disturbances of blood flow play an important role in promoting platelet activation and arterial thrombus formation in stenosed, injured, atherosclerotic arteries. To date, glycoprotein Ib (GPIb) has been considered the primary platelet mechanosensory receptor, responding to increased shear with enhanced adhesive and signaling function. We demonstrate here that von Willebrand factor-GPIb interaction is inefficient at inducing platelet activation even when platelets are exposed to very high wall shear stresses (60 dyn/cm(2)). Rapid platelet activation under flow was only observed under experimental conditions in which transiently adherent platelets were exposed to sudden accelerations in blood flow. Platelet responsiveness to temporal shear gradients was integrin alpha(IIb)beta(3)-dependent and occurred only on a von Willebrand factor substrate, as platelets forming integrin alpha(IIb)beta(3) adhesive contacts with immobilized fibrinogen were unresponsive to sudden increases in shear. The calcium response induced by temporal shear gradients was distinct from previously identified integrin alpha(IIb)beta(3) calcium responses in terms of its transient nature, its requirement for platelet co-stimulation by the P2Y(1) purinergic ADP receptor, and its dependence on the influx of extracellular calcium. Our studies demonstrate a key role for temporal shear gradients in promoting platelet activation. Moreover, they define for the first time the involvement of P2Y receptors in integrin mechanotransduction.     

Lipoyl synthase requires two equivalents of S-adenosyl-L-methionine to synthesize one equivalent of lipoic acid

Lipoyl synthase (LipA) catalyzes the formation of the lipoyl cofactor, which is employed by several multienzyme complexes for the oxidative decarboxylation of various alpha-keto acids, as well as the cleavage of glycine into CO(2) and NH(3), with concomitant transfer of its alpha-carbon to tetrahydrofolate, generating N(5),N(10)-methylenetetrahydrofolate. In each case, the lipoyl cofactor is tethered covalently in an amide linkage to a conserved lysine residue located on a designated lipoyl-bearing subunit of the complex. Genetic and biochemical studies suggest that lipoyl synthase is a member of a newly established class of metalloenzymes that use S-adenosyl-l-methionine (AdoMet) as a source of a 5'-deoxyadenosyl radical (5'-dA(*)), which is an obligate intermediate in each reaction. These enzymes contain iron-sulfur clusters, which provide an electron during the cleavage of AdoMet, forming l-methionine in addition to the primary radical. Recently, one substrate for lipoyl synthase has been shown to be the octanoylated derivative of the lipoyl-bearing subunit (E(2)) of the pyruvate dehydrogenase complex [Zhao, S., Miller, J. R., Jian, Y., Marletta, M. A., and Cronan, J. E., Jr. (2003) Chem. Biol. 10, 1293-1302]. Herein, we show that the octanoylated derivative of the lipoyl-bearing subunit of the glycine cleavage system (H-protein) is also a substrate for LipA, providing further evidence that the cofactor is synthesized on its target protein. Moreover, we show that the 5'-dA(*) acts directly on the octanoyl substrate, as evidenced by deuterium transfer from [octanoyl-d(15)]H-protein to 5'-deoxyadenosine. Last, our data indicate that 2 equiv of AdoMet are cleaved irreversibly in forming 1 equiv of [lipoyl]H-protein and are consistent with a model in which two LipA proteins are required to synthesize one lipoyl group.     

Homogeneity of the 16S rDNA sequence among geographically disparate isolates of Taylorella equigenitalis

Background

At present, six accessible sequences of 16S rDNA from Taylorella equigenitalis (T. equigenitalis) are available, whose sequence differences occur at a few nucleotide positions. Thus it is important to determine these sequences from additional strains in other countries, if possible, in order to clarify any anomalies regarding 16S rDNA sequence heterogeneity. Here, we clone and sequence the approximate full-length 16S rDNA from additional strains of T. equigenitalis isolated in Japan, Australia and France and compare these sequences to the existing published sequences.

Results

Clarification of any anomalies regarding 16S rDNA sequence heterogeneity of T. equigenitalis was carried out. When cloning, sequencing and comparison of the approximate full-length 16S rDNA from 17 strains of T. equigenitalis isolated in Japan, Australia and France, nucleotide sequence differences were demonstrated at the six loci in the 1,469 nucleotide sequence. Moreover, 12 polymorphic sites occurred among 23 sequences of the 16S rDNA, including the six reference sequences.

Conclusion

High sequence similarity (99.5% or more) was observed throughout, except from nucleotide positions 138 to 501 where substitutions and deletions were noted.