Recent clinical trials have led to the worldwide suspension of aprotinin, the most commonly used antifibrinolytic agent in fibrin-based tissue engineering. For future clinical applications of fibrin-based scaffolds, a suitable, alternative fibrinolysis inhibitor must be identified. The present study aimed to evaluate tranexamic acid (trans-4-aminomethyl-cyclohexane-1-carboxylic acid [t-AMCA]) as an alternative fibrinolysis inhibitor to aprotinin for cardiovascular tissue engineering applications. The effects of various concentrations of t-AMCA (30-160 microg/mL) and aprotinin on fibrin gel-lysis were spectrophotometrically quantified in vitro. Cytotoxic effects of t-AMCA and aprotinin on carotid artery-derived cells, in addition to their influence on fibrin gel mechanical strength, were examined. Further, the influence of t-AMCA versus aprotinin on three-dimensional fibrin-based constructs was analyzed using light microscopy, scanning electron microscopy, and transmission electron microscopy. The results demonstrated that neither t-AMCA (30-160 microg/mL) nor aprotinin elicited cytotoxic effects on cultured cells. Although aprotinin showed reduced fibrinolysis in the presence of plasmin compared to t-AMCA, no significant difference was obtained under standard culture conditions. Additionally, t-AMCA had no negative influence on the mechanical stability of fibrin gels, which also demonstrated excellent cell morphology, tissue development, and ultrastructure. The results from the present study demonstrate that t-AMCA may be a suitable alternative to aprotinin for controlling the in vitro degradation rate of fibrin-based tissue-engineered constructs.

Development of dendritic cells (DCs) commences in the bone marrow, from where pre-DCs migrate to peripheral organs to differentiate into mature DCs in situ. However, the factors that regulate organ-specific differentiation to give rise to tissue-specific DC subsets remain unclear. Here we show that the Ras-PI3Kγ-Akt-mTOR signaling axis acted downstream of FLT3L signaling and was required for development of lung CD103(+) DCs and, to a smaller extent, for lung CD11b(+) DCs, but not related DC populations in other non-lymphoid organs. Furthermore, we show that in lymphoid organs such as the spleen, DCs depended on a similar signaling network to respond to FLT3 ligand with overlapping and partially redundant roles for kinases PI3Kγ and PI3Kδ. Thus we identified PI3Kγ as an essential organ-specific regulator of lung DC development and discovered a signaling network regulating tissue-specific DC development mediated by FLT3.

The results of the present study demonstrated that ethanol precipitation is a simple and effective method for isolation of fibrinogen and a suitable alternative to cryoprecipitation. This technique allows minimization of the necessary blood volume for fibrinogen isolation, particularly important for pediatric applications, and also has no negative influence on microstructure, mechanical properties, cell proliferation, or tissue development.

Fibrin gel has proven a valuable scaffold for tissue engineering. Complex geometries can be produced by injection molding; it offers effective cell seeding and can be produced autologous. In order to evaluate its suitability for respiratory tissue engineering, we examined proliferation, functionality, and differentiation of respiratory epithelial cells on fibrin gel in comparison to culture on collagen-coated, microporous membranes. Respiratory epithelial cells formed a confluent layer by day 4, and proliferation showed no significant difference with respect to surface. Measurement of the transepithelial electrical resistance reflected the development of a confluent epithelial cell layer and the subsequent initiation of adequate ion-transfer processes. Appearance of ciliae could be detected at similar time points, and ciliary beating could be observed for cells on both surfaces. Histology and immunohistochemistry of cells grown on fibrin gel revealed the onset of adequate differentiation. As no significant differences in respiratory epithelial cells' proliferation, function, and differentiation could be observed between cells grown on fibrin gel compared to cells on a collagen-coated, microporous surface, we concluded that fibrin gel might prove a suitable scaffold for respiratory tissue engineering and merits further investigation to overcome the limitations associated with scaffolds currently in use.