scholarly journals Dynamic Imaging of Fibrin Network Formation Correlated with Other Measures of Polymerization.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 1757-1757
Author(s):  
John W. Weisel ◽  
Irina N. Chernysh
Keyword(s):  
Branch Point ◽  
Network Formation ◽  
Dynamic Imaging ◽  
Longitudinal Growth ◽  
Gel Point ◽  
Lateral Growth ◽  
Branch Points ◽  
Sequence Of Events ◽  
Fibrin Network ◽  
Length Changes

Abstract The clotting time or gel point is commonly used in a variety of clinical assays, but little has been known about what it represents structurally because of the limitations of imaging technologies. The necessity of fixation and dehydration for electron microscopy means that only final clot structures, often distorted by preparation artifacts, can be observed, and there are severe limitations of image quality for conventional light microscopy. For similar reasons, we know almost nothing about the branching and lateral aggregation of fibrin that are essential for clot or thrombus stability. Turbidity, light scattering, or clot stiffness are useful to follow the time course of polymerization but reflect only overall clot properties. We visualized in real time fibrin network formation in the hydrated state, using deconvolution microscopy, which allows optical sectioning without the bleaching that accompanies confocal microscopy. Thus, the events during polymerization could be followed quantitatively over long periods of time. Videos will demonstrate the major observations. Individual mobile fibers were observed before the gel point. After gelation, an initial fibrin network, or scaffold, was seen, which evolved over time by addition of new fibers and elongation and branching of others. Furthermore, some fibers in the network moved chaotically for some time. A detailed, quantitative morphological analysis of network formation was carried out by superposition of images from different time points colorized to distinguish changes. We quantified network formation by the number of branch points, and longitudinal and lateral growth of fibers as a function of time. The distributions of fibers that reached a maximum of longitudinal growth and branch point formation both had maxima at the gel point but, surprisingly, some longitudinal growth continued and new branch points appeared after the gel point, requiring modification of existing models of fibrin polymerization. The cumulative percentage of fibers reaching their final length and the number of branch points attained maximum values at the time corresponding to that at which the turbidity reached approximately 60% of its maximum. Lateral growth reached a plateau at the same time as turbidity. Measurements of clot mechanical properties revealed that the clots achieved maximum stiffness and minimum plasticity well after branch point, as well as length changes and lateral growth of fibers, were completed. These results provide new information on the time sequence of events during fibrin network formation, which is important to understand both clotting and thrombosis and to allow modulation of clot properties.

scholarly journals Dynamic imaging of fibrin network formation correlated with other measures of polymerization

Blood ◽  
2008 ◽  
Vol 111 (10) ◽  
pp. 4854-4861 ◽  
Author(s):  
Irina N. Chernysh ◽  
John W. Weisel
Keyword(s):  
Network Formation ◽  
Structural Parameters ◽  
Dynamic Imaging ◽  
Gel Point ◽  
Branch Points ◽  
Sequence Of Events ◽  
New Information ◽  
Maximum Stiffness ◽  
Fibrin Network ◽  
Final Length

Abstract Using deconvolution microscopy, we visualized in real time fibrin network formation in the hydrated state. Individual mobile fibers were observed before the gel point determined by eye. After gelation, an initial fibrin network was seen, which evolved over time by addition of new fibers and elongation and branching of others. Furthermore, some fibers in the network moved for a time. We quantified network formation by number of branch points, and longitudinal and lateral growth of fibers. Eighty percent of branch points were formed, and 70% of all fibers reached their maximum length at the gel point. In contrast, at the gel point, fiber diameter, measured as fluorescence intensity, was less than 25% and turbidity was less than 15% of the maximum values of the fully formed clot. The cumulative percentage of fibers reaching their final length and the number of branch points attained maximum values at 60% of maximum turbidity. Lateral fiber growth reached a plateau at the same time as turbidity. Measurements of clot mechanical properties revealed that the clots achieved maximum stiffness and minimum plasticity after the structural parameters reached their maxima. These results provide new information on the relative time sequence of events during fibrin network formation.

Platelet Microtubules in Clot Structure Formation and Contractile Force Generation: Investigation of a Controversy

1986 ◽  
Vol 56 (01) ◽  
pp. 023-027 ◽  
Author(s):  
C J Jen ◽  
L V McIntire
Keyword(s):  
Network Formation ◽  
Contractile Force ◽  
Second Phase ◽  
Clot Retraction ◽  
Forming Process ◽  
Microtubule Organization ◽  
Physiological Processes ◽  
Fibrin Network ◽  
Two Parameters

SummaryWhether platelet microtubules are involved in clot retraction/ contraction has been controversial. To address this question we have simultaneously measured two clotting parameters, clot structural rigidity and isometric contractile force, using a rheological technique. For recalcified PRP clots these two parameters began rising together at about 15 min after CaCl2 addition. In the concentration range affecting microtubule organization in platelets, colchicine, vinca alkaloids and taxol demonstrated insignificant effects on both clotting parameters of a recalcified PRP clot. For PRP clots induced by adding small amounts of exogenous thrombin, the kinetic curves of clot rigidity were biphasic and without a lag time. The first phase corresponded to a platelet-independent network forming process, while the second phase corresponded to a platelet-dependent process. These PRP clots began generating contractile force at the onset of the second phase. For both rigidity and force parameters, only the second phase of clotting kinetics was retarded by microtubule affecting reagents. When PRP samples were clotted by adding a mixture of CaCl2 and thrombin, the second phase clotting was accelerated and became superimposed on the first phase. The inhibitory effects of micro tubule affecting reagents became less pronounced. Thrombin clotting of a two-component system (washed platelets/ purified fibrinogen) was also biphasic, with the second phase being microtubule-dependent. In conclusion, platelet microtubules are important in PRP clotted with low concentrations of thrombin, during which fibrin network formation precedes platelet-fibrin interactions. On the other hand they are unimportant if a PRP clot is induced by recalcification, during which the fibrin network is constructed in the presence of platelet-fibrin interactions. The latter is likely to be more analogous to physiological processes in vivo.

scholarly journals Assessment of branch point prediction tools to predict physiological branch points and their alteration by variants

2019 ◽  
Author(s):  
Raphael Leman ◽  
Hélène Tubeuf ◽  
Sabine Raad ◽  
Isabelle Tournier ◽  
Céline Derambure ◽  
...  
Keyword(s):  
Branch Point ◽  
Mrna Splicing ◽  
Large Set ◽  
Branch Points ◽  
Splice Sites ◽  
Prediction Tools ◽  
Mature Mrna ◽  
Bioinformatics Tools ◽  
The Impact

Abstract Background: Branch points (BPs) map within short motifs upstream of acceptor splice sites (3’ss) and are essential for splicing of pre-mature mRNA. Several BP-dedicated bioinformatics tools, including HSF, SVM-BPfinder, BPP, Branchpointer, LaBranchoR and RNABPS were developed during the last decade. Here, we evaluated their capability to detect the position of BPs, and also to predict the impact on splicing of variants occurring upstream of 3’ss. Results: We used a large set of constitutive and alternative human 3’ss collected from Ensembl (n = 264,787 3’ss) and from in-house RNAseq experiments (n = 51,986 3’ss). We also gathered an unprecedented collection of functional splicing data for 120 variants (62 unpublished) occurring in BP areas of disease-causing genes. Branchpointer showed the best performance to detect the relevant BPs upstream of constitutive and alternative 3’ss (99.48 % and 65.84 % accuracies, respectively). For variants occurring in a BP area, BPP emerged as having the best performance to predict effects on mRNA splicing, with an accuracy of 89.17 %. Conclusions: Our investigations revealed that Branchpointer was optimal to detect BPs upstream of 3’ss, and that BPP was most relevant to predict splicing alteration due to variants in the BP area. Keywords: Branch Point, Prediction, RNA, Benchmark, HSF, SVM-BPfinder, BPP, Branchpointer, LaBranchoR, RNABPS, Variants

Abstract 329: The Role of TIE1 in Flow Mediated Lymphatic Vessel Remodeling and Valvulogenesis

2016 ◽  
Vol 36 (suppl_1) ◽  
Author(s):  
Cristina Harmelink ◽  
Bin Zhou ◽  
Xianghu Qu ◽  
H. Scott Baldwin
Keyword(s):  
Network Formation ◽  
Lymphatic Vessels ◽  
Mechanical Stimulus ◽  
Turbulent Shear ◽  
Branch Points ◽  
Turbulent Shear Stress ◽  
Valve Development ◽  
Lymphatic Vasculature ◽  
Cellular Machinery ◽  
Molecular Landscape

Recently, it has been shown that the mechanical stimulus of turbulent shear stress caused by onset of lymph flow is required for lymphatic remodeling, maturation, and lymphatic valve (LV) development. Homeostasis of the adult lymphatic vasculature also relies on flow-mediated signal transduction. However, the cellular machinery responsible for transducing mechanosensory signals required for lymphatic network formation and maintenance is unknown. Our laboratory has previously shown that TIE1 is at least partially responsible for mechanotransduction of turbulent flow required for initiation and maintenance of atherosclerotic plaque formation at the branch points of systemic vasculature in the adult animal. Moreover, TIE1 is expressed throughout lymphatic vasculature during mouse embryogenesis into adulthood, with enrichment in LVs. To circumvent the embryonic lethality caused by global Tie1 disruption, we conditionally deleted Tie1 using Nfatc1Cre. Nfatc1Cre drives recombination in lymphatic endothelial cells, with strong expression in the LVs. Nfatc1Cre:Tie1fl/fl mutants survive to birth but accumulate chyle in the peritoneal and pleural cavities by postnatal day 2. The lymphatic vessels in the mutants are dilated and tortuous, and do not undergo normal hierarchical remodeling. The constrictions that normally indicate intraluminal valve development are lacking in the mutant lymphatic vessels. Underlying these defects in the Nfatc1Cre:Tie1fl/fl mutants is loss of the normal molecular landscape associated with lymphatic patterning and valvulogenesis. Therefore, we hypothesize that Tie1 orchestrates the mechanotransduction necessary for intraluminal LV development and postnatal maintenance.

On the threshold branch-point of many-body channels

1963 ◽  
Vol 276 (1367) ◽  
pp. 492-501
Keyword(s):  
Branch Point ◽  
Scattering Amplitude ◽  
Cube Root ◽  
Branch Points ◽  
Many Body ◽  
Realistic Potential ◽  
Break Up ◽  
Analytic Behaviour ◽  
The Masses ◽  
Special Case

The analytic behaviour of the elastic and break-up scattering amplitudes in a soluble one-dimensional model has been examined by Nussenzveig. It was found that the break-up threshold gave rise to rather curious cube-root branch-points in the scattering amplitude. In this paper, we shall examine the scattering amplitude in a soluble model which reduces Nussenzveig’s model to the special case where the incident and ionized particles have equal masses. It will be shown that the threshold branch-point is a function of the ratio of the masses of the two particles and that the cube-root occurs only when the masses are equal. In general, there are an infinite number of Riemann sheets associated with the threshold branch-point. An examination into the physical origin of such a threshold behaviour will also be made to determine if a more complicated branch-point than a simple square-root may exist for a more realistic potential model.

scholarly journals ZFAT is essential for endothelial cell assembly and the branch point formation of capillary-like structures in an angiogenesis model

2010 ◽  
Vol 15 (4) ◽  
Author(s):  
Yasuhiro Yoshida ◽  
Toshiyuki Tsunoda ◽  
Yasuo Takashima ◽  
Takahiro Fujimoto ◽  
Keiko Doi ◽  
...  
Keyword(s):  
Endothelial Cell ◽  
Branch Point ◽  
Molecular Mechanisms ◽  
Network Formation ◽  
Umbilical Vein ◽  
Susceptibility Gene ◽  
Cell Assembly ◽  
Autoimmune Thyroid ◽  
Umbilical Vein Endothelial Cells ◽  
Angiogenesis Model

AbstractZFAT, originally identified as a susceptibility gene for autoimmune thyroid disease, encodes a transcriptional regulator with one AT-hook and 18 C2H2-type zinc-finger domains. It is highly conserved among species. Here, we demonstrate that ZFAT is clearly expressed in human umbilical vein endothelial cells (HUVECs). Furthermore, we show that endothelial cell assembly and the branch point formation of capillary-like structures in HUVECs is impaired by the reduction of ZFAT expression through the use of ZFAT-miRNAs, whereas differences in cell proliferation or apoptotic features were not observed after the reduction in ZFAT expression. These results suggest that ZFAT may have critical roles in the capillary-like network formation that is involved in vascular remodeling. Elucidating the ZFAT-mediated transcriptional network will lead to a better understanding of the molecular mechanisms of angiogenesis.

scholarly journals An inventory of early branch points in microbial phosphonate biosynthesis

2021 ◽  
Author(s):  
Siwei Li ◽  
Geoff P Horsman
Keyword(s):  
Branch Point ◽  
Sequence Similarity ◽  
Bond Formation ◽  
Branch Points ◽  
Bacterial Genomes ◽  
Naturally Occurring ◽  
Carbon Carbon Bond ◽  
The Common ◽  
Almost All ◽  
Biosynthetic Machinery

Microbial phosphonate biosynthetic machinery has been identified in ~5% of bacterial genomes and encodes natural products like fosfomycin as well as cell surface decorations. Almost all biological phosphonates originate from the rearrangement of phosphoenolpyruvate (PEP) to phosphonopyruvate (PnPy) catalyzed by PEP mutase (Ppm), and PnPy is often converted to phosphonoacetaldehyde (PnAA) by PnAA decarboxylase (Ppd). Seven enzymes are known or likely to act on either PnPy or PnAA as early branch points en route to diverse biosynthetic outcomes, and these enzymes may be broadly classified into three reaction types: hydride transfer, aminotransfer, and carbon-carbon bond formation. However, the relative abundance of these branch points in microbial phosphonate biosynthesis is unknown. Also unknown is the proportion of ppm-containing gene neighborhoods encoding new branch point enzymes and potentially novel phosphonates. In this study we computationally sorted 882 ppm-containing gene neighborhoods based on these seven branch point enzymes. Unsurprisingly, the majority (56%) of these pathways encode for production of the common naturally occurring compound 2-aminoethylphosphonate (AEP) or a hydroxylated derivative. The next most abundant genetically encoded intermediates were phosphonoacetate (PnAc, 16%), phosphonoalanine (PnAla, 10%), and 2-hydroxyethylphosphonate (HEP, 5%). Significantly, about 9% of the gene neighborhoods could not be assigned to any of the seven branch points and may encode novel phosphonates. Sequence similarity network analysis revealed families of unusual gene neighborhoods including possible production of phosphonoacrylate and phosphonofructose, the apparent biosynthetic use of the C-P lyase operon, and a virus-encoded phosphonate. Overall, these results highlight the utility of branch point inventories to identify novel gene neighborhoods and guide future phosphonate discovery efforts.

scholarly journals Unitarity and Regge Branch Points

1974 ◽  
Vol 27 (6) ◽  
pp. 745
Author(s):  
GC Joshi ◽  
Antonio Pagnamenta
Keyword(s):  
Branch Point ◽  
Branch Points ◽  
End Point

We show how t-channel unitarity imposes constraints on the discontinuity associated with the leading Regge branch point. If the discontinuity across the leading Regge cut does not vanish at the end point it leads to the 'Gribov paradox'.

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