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Morphometric characterization of fibrinogen's ?C regions and their role in fibrin self-assembly and molecular organization.

ABSTRACT: The flexible C-terminal parts of fibrinogen's A? chains named the ?C regions have been shown to play a role in fibrin self-assembly, although many aspects of their structure and functions remain unknown. To examine the involvement of the ?C regions in the early stages of fibrin formation, we used high-resolution atomic force microscopy to image fibrinogen and oligomeric fibrin. Plasma-purified full-length human fibrinogen or des-?C fibrinogen lacking most of the ?C regions, untreated or treated with thrombin, was imaged. Up to 80% of the potentially existing ?C regions were visualized and quantified; they were highly heterogeneous in their length and configurations. Conversion of fibrinogen to fibrin was accompanied by an increase in the incidence and length of the ?C regions as well as transitions from more compact conformations, such as a globule on a string, to extended and more flexible offshoots. Concurrent dynamic turbidimetry, confocal microscopy, and scanning electron microscopy revealed that trimming of the ?C regions slowed down fibrin formation, which correlated with longer protofibrils, thinner fibers, and a denser network. No structural distinctions, except for the incidence of the ?C regions, were revealed in the laterally aggregated protofibrils made of the full-length or des-?C fibrinogens, suggesting a pure kinetic effect of the ?C regions on the fibrin architecture. This work provides a structural molecular basis for the promoting role of the ?C regions in the early stages of fibrin self-assembly and reveals this stage of fibrin formation as a potential therapeutic target to modulate the structure and mechanical properties of blood clots.

SUBMITTER: Protopopova AD

PROVIDER: S-EPMC6501582 | biostudies-literature | 2017 Sep

REPOSITORIES: biostudies-literature

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Morphometric characterization of fibrinogen's αC regions and their role in fibrin self-assembly and molecular organization.

Protopopova Anna D AD Litvinov Rustem I RI Galanakis Dennis K DK Nagaswami Chandrasekaran C Barinov Nikolay A NA Mukhitov Alexander R AR Klinov Dmitry V DV Weisel John W JW

Nanoscale 20170901 36

The flexible C-terminal parts of fibrinogen's Aα chains named the αC regions have been shown to play a role in fibrin self-assembly, although many aspects of their structure and functions remain unknown. To examine the involvement of the αC regions in the early stages of fibrin formation, we used high-resolution atomic force microscopy to image fibrinogen and oligomeric fibrin. Plasma-purified full-length human fibrinogen or des-αC fibrinogen lacking most of the αC regions, untreated or treated ...[more]

PMID: 28884176

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Project description:Introduction: New vessel formation requires a continuous and tightly regulated interplay between endothelial cells with cells of the perivascular microenvironment supported by mechanic-physical and chemical cues from the extracellular matrix. Aim: Here we investigated the potential of small fragments of synovial tissue to form de novo vascular structures in the context of inflammation within three dimensional (3D) fibrin-based matrices in vitro, and assessed the contribution of mesenchymal stromal cell (MSC)-immune cell cross-talk to neovascularization considering paracrine signals in a fibrin-based co-culture model. Material and Methods: Synovial tissue fragments from patients with rheumatoid arthritis (RA) and inflammatory osteoarthritis (OA) were cultivated within 3D fibrin matrices for up to 4 weeks. Cellular and structural re-arrangement of the initially acellular matrix were documented by phase contrast microscopy and characterized by confocal laser-scanning microscopy of topographically intact 3D cultures and by immunohistochemistry. MSC-peripheral blood mononuclear cell (PBMC) co-cultures in the 3D fibrin system specifically addressed the influence of perivascular cell interactions to neo-vessel formation in a pro-inflammatory microenvironment. Cytokine levels in the supernatants of cultured explant tissues and co-cultures were evaluated by the Bio-Plex cytokine assay and ELISA. Results: Vascular outgrowth from the embedded tissue into the fibrin matrix was preceded by leukocyte egress from the tissue fragments. Neo-vessels originating from both the embedded sample and from clusters locally formed by emigrated mononuclear cells were consistently associated with CD45+ leukocytes. MSC and PBMC in co-culture formed vasculogenic clusters. Clusters and cells with endothelial phenotype emerging from them, were surrounded by a collagen IV scaffold. No vascular structures were observed in control 3D monocultures of PBMC or MSC. Paracrine signals released by cultured OA tissue fragments corresponded with elevated levels of granulocyte-colony stimulating factor, vascular endothelial growth factor and interleukin-6 secreted by MSC-PBMC co-cultures. Conclusion: Our results show that synovial tissue fragments with immune cell infiltrates have the potential to form new vessels in initially avascular 3D fibrin-based matrices. Cross-talk and cluster formation of MSC with immune cells within the 3D fibrin environment through self-organization and secretion of pro-angiogenic paracrine factors can support neo-vessel growth.

Dataset Information

Morphometric characterization of fibrinogen's ?C regions and their role in fibrin self-assembly and molecular organization.

Publications

Morphometric characterization of fibrinogen's αC regions and their role in fibrin self-assembly and molecular organization.

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