fibrin and thymosin-beta(4)

Thymosin beta 4 (Tβ4) has multiple beneficial facets for myocardial injury, but its efficiency is limited by the low local concentration within the infarct. Here, we established a Tβ4 delivery system for cardiac repair based on the interaction between the abundant fibrin in the infarct zone and the fibrin-targeting moiety clot-binding peptide cysteine-arginine-glutamic acid-lysine-alanine (CREKA).. CREKA and Tβ4 were conjugated to nanoparticles (CNP-Tβ4). In vitro binding test revealed that CNP-Tβ4 had a significant binding ability to the surface of fibrin clots when compared to the control clots (NP-Tβ4). Based on the validation of fibrin expression in the early stage of ischemia injury, CNP-Tβ4 was intravenously administered to mice with acute myocardial ischemia-reperfusion injury. CNP-Tβ4 revealed a stronger fibrin-targeting ability than the NP-Tβ4 group and accumulated mainly in the infarcted area and colocalized with fibrin. Subsequently, treatment with CNP-Tβ4 resulted in a better therapeutic effect.. CRKEA modification favored Tβ4 accumulation and retention in the infarcted region, leading to augmented functional benefits. Fibrin-targeting delivery system represents a generalizable platform technology for regenerative medicine.

Topics: Animals; Drug Delivery Systems; Fibrin; Male; Mice, Inbred BALB C; Molecular Targeted Therapy; Myocardial Reperfusion Injury; Myocardium; Nanoparticles; Oligopeptides; Regenerative Medicine; Signal Transduction; Thymosin; Tissue Distribution

It was shown recently that tissue transglutaminase and presumably plasma transglutaminase, factor XIIIa, can covalently incorporate into fibrin(ogen) a physiologically active peptide, thymosin beta(4) [(Huff et al. (2002) FASEB J. 16, 691-696]. To clarify the mechanism of this incorporation, we studied the interaction of thymosin beta(4) with fibrinogen, fibrin, and their recombinant fragments, the gamma-module (gamma-chain residues 148-411), and the alphaC-domain (Aalpha-chain residues 221-610) and its truncated variants by immunoblot and ELISA. No significant noncovalent interaction between them was detected in the absence of activated factor XIII, while in its presence thymosin beta(4) was effectively incorporated into fibrin and to a lesser extent into fibrinogen. The incorporation at physiological concentrations of fibrin(ogen) and factor XIII was significant with molar incorporation ratios of thymosin beta(4) to fibrinogen and fibrin of 0.2 and 0.4, respectively. Further experiments revealed that although activated factor XIII incorporates thymosin beta(4) into the isolated gamma-module and alphaC-domain, in fibrin the latter serves as the major incorporation site. This site was further localized to the COOH-terminal portion of the alphaC-domain including residues 392-610.

Topics: Amino Acid Sequence; Factor XIIIa; Fibrin; Fibrinogen; Humans; Kinetics; Peptide Fragments; Protein Structure, Tertiary; Protein Subunits; Sequence Deletion; Thymosin

The beta-thymosins constitute a family of highly conserved and extremely water-soluble 5 kDa polypeptides. Thymosin beta4 is the most abundant member; it is expressed in most cell types and is regarded as the main intracellular G-actin sequestering peptide. There is increasing evidence for extracellular functions of thymosin beta4. For example, thymosin beta4 increases the rate of attachment and spreading of endothelial cells on matrix components and stimulates the migration of human umbilical vein endothelial cells. Here we show that thymosin beta4 can be cross-linked to proteins such as fibrin and collagen by tissue transglutaminase. Thymosin beta4 is not cross-linked to many other proteins and its cross-linking to fibrin is competed by another family member, thymosin beta10. After activation of human platelets with thrombin, thymosin beta4 is released and cross-linked to fibrin in a time- and calcium-dependent manner. We suggest that thymosin beta4 cross-linking is mediated by factor XIIIa, a transglutaminase that is coreleased from stimulated platelets. This provides a mechanism to increase the local concentration of thymosin beta4 near sites of clots and tissue damage, where it may contribute to wound healing, angiogenesis and inflammatory responses.

Topics: Actins; Binding, Competitive; Blood Platelets; Calcium; Collagen; Factor XIIIa; Fibrin; Fibrinogen; Humans; Kinetics; Models, Biological; Thymosin