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Grant, Derrick S.; Rose, Wesley; Yaen, Christopher H.; Goldstein, Allan L.; Martinez, Josè; Kleinman, Hynda K.

doi:10.1023/a:1009041911493

Cited by 127 publications

(42 citation statements)

References 40 publications

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“…Since AGP is known to localize at inflammation foci, it may provide the basic molecular framework for the concentration of PAI-1. Interestingly, T␤4 is also accumulated in such areas, and particularly in wound fluid (3,23,24). It suggests that AGP may serve as a receptor for PAI-1 on cells that naturally express low concentrations of membrane PAI-1 receptors.…”

Section: Discussionmentioning

confidence: 99%

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Binding of PAI-1 to Endothelial Cells Stimulated by Thymosin β4 and Modulation of Their Fibrinolytic Potential

Boncela

Smolarczyk

Wyroba

et al. 2006

Journal of Biological Chemistry

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Our previous studies showed that thymosin ␤4 (T␤4) induced the synthesis of plasminogen activator inhibitor-1 (PAI-1) in cultured human umbilical vein endothelial cells (HUVECs) via the AP-1 dependent mechanism and its enhanced secretion. In this work we provide evidence that the released PAI-1 is accumulated on the surface of HUVECs, exclusively in its active form, in a complex with ␣1-acid glycoprotein (AGP) that is also up-regulated and released from the cells. This mechanism is supported by several lines of experiments, in which expression of both proteins was analyzed by flow cytometry and their colocalization supported by confocal microscopy. PAI-1 did not bind to quiescent cells but only to the T␤4-activated endothelial cells. In contrast, significant amounts of AGP were found to be associated with the cells overexpressing enhanced green fluorescent protein (EGFP)-␣1-acid glycoprotein (AGP) without T␤4 treatment. The AGP⅐PAI-1 complex was accumulated essentially at the basal surface of endothelial cells, and such cells showed (a) morphology characteristic for strongly adhered and spread cells and (b) significantly reduced plasmin formation. Taken together, these results provide the evidence supporting a novel mechanism by which active PAI-1 can be bound to the T␤4-activated endothelial cells, thus influencing their adhesive properties as well as their ability to generate plasmin.Thymosin ␤4 (T␤4) 2 promotes skin and corneal wound healing through its effects on cell migration, angiogenesis and possibly cell survival (1-3). However, the precise molecular mechanism through which it functions and its potential role in solid organ wound healing remains unknown. In our studies of the biological activity of T␤4 we found that this 43-amino acid peptide strongly activates endothelial cells to express and release plasminogen activator inhibitor type 1 (PAI-1). It occurs via the mechanism involving the signaling pathway leading to activation of the MAPK cascade and enhanced c-Fos/c-Jun DNA binding activity (4). Such a role of AP-1 in activation of transcriptional events by T␤4 was confirmed by independent studies (5). Recently, T␤4 was reported to stimulate migration of cardiomyocytes and endothelial cells and promote survival of cardiomyocytes by formation of a functional complex with PINCH and integrin-linked kinase, resulting in activation of the survival kinase Akt (6). These observations revealed that T␤4 affects cellular functions by activation of several signaling pathways and induces changes in the cell properties necessary for both migration and survival. Here, we show that T␤4 up-regulates expression and induces the surface accumulation of ␣1-acid glycoprotein (AGP) known to bind PAI-1 and prolong its inhibitory activity (7). AGP is expressed by activated endothelial cells and exerts its effect by interacting with components of the endothelial glycocalyx (8). In this study, we identify a previously unrecognized function of AGP, a direct and primary role in plasminogen activation. We provide evidence that t...

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Section: Discussionmentioning

confidence: 99%

“…Thymosin ␤4 (T␤4) 2 promotes skin and corneal wound healing through its effects on cell migration, angiogenesis and possibly cell survival (1)(2)(3). However, the precise molecular mechanism through which it functions and its potential role in solid organ wound healing remains unknown.…”

mentioning

confidence: 99%

Binding of PAI-1 to Endothelial Cells Stimulated by Thymosin β4 and Modulation of Their Fibrinolytic Potential

Boncela

Smolarczyk

Wyroba

et al. 2006

Journal of Biological Chemistry

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“…Although it is a typical intracellular polypeptide, it plays numerous roles, both intracellularly and extracellularly. Numerous observations indicate that T␤4 is involved in adhesion and spreading of fibroblasts (1,2), differentiation of endothelial cells (3,4), directional migration of endothelial cells and keratinocytes (5)(6)(7)(8), angiogenesis (3-6, 9, 10), wound healing (6,7,11), hair follicle growth (8), and apoptosis (12,13), and has been described to possess anti-inflammatory properties (11,14). In addition, elevated T␤4 expression has been observed in various malignant cell lines and tumors (15)(16)(17) and its levels seem to be associated with increased tumorigenicity and metastatic potential (10, 13, 18 -20).…”

mentioning

confidence: 99%

“…Thus, T␤4 externally added to endothelial cells: (a) induces expression and release of plasminogen activator inhibitor type 1 (PAI-1) by the mechanism involving activation of the mitogen-activated protein kinase cascade leading to enhanced c-Fos/c-Jun binding to the AP-1-like element present in the PAI-1 promoter (25,26); (b) binds to PINCH and integrinlinked kinase, resulting in activation of the survival kinase Akt, and thus promotes myocardial and endothelial cell migration in the embryonic heart (27) and (c) promotes skin and corneal wound healing through its effects on cell migration, angiogenesis, and possibly cell survival (4,6,11).…”

mentioning

confidence: 99%

Ku80 as a Novel Receptor for Thymosin β4 That Mediates Its Intracellular Activity Different from G-actin Sequestering

Bednarek

Boncela

Smolarczyk

et al. 2008

Journal of Biological Chemistry

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Our data demonstrate that increased intracellular expression of thymosin ␤4 (T␤4) is necessary and sufficient to induce plasminogen activator inhibitor type 1 (PAI-1) gene expression in endothelial cells. To describe the mechanism of this effect, we produced T␤4 mutants with impaired functional motifs and tested their intracellular location and activity. Cytoplasmic distributions of T␤4 (AcSDKPT/4A) , T␤4 (KLKKTET/7A) , and T␤4 (K16A) mutants fused with green fluorescent protein did not differ significantly from those of wild-type T␤4. Overexpression of T␤4, T␤4 (AcSDKPT/4A) , and T␤4 (K16A) affected intracellular formation of actin filaments. As expected, T␤4 (K16A) uptake by nuclei was impaired. On the other hand, overexpression of T␤4 (KLKKTET/7A) resulted in developing the actin filament network typical of adhering cells, indicating that the mutant lacked the actin binding site. The mechanism by which intracellular T␤4 induced the PAI-1 gene did not depend upon the N-terminal tetrapeptide AcSDKP and depended only partially on its ability to bind G-actin or enter the nucleus. Both T␤4 and T␤4 (AcSDKPT/4A) induced the PAI-1 gene to the same extent, whereas mutants T␤4 (KLKKTET/7A) and T␤4 (K16A) retained about 60% of the original activity. By proteomic analysis, the Ku80 subunit of ATPdependent DNA helicase II was found to be associated with T␤4. Ku80 and T␤4 consistently co-immunoprecipitated in a complex from endothelial cells. Co-transfection of endothelial cells with the Ku80 deletion mutants and T␤4 showed that the C-terminal arm domain of Ku80 is directly involved in this interaction. Furthermore, down-regulation of Ku80 by specific short interference RNA resulted in dramatic reduction in PAI-1 expression at the level of both mRNA and protein synthesis. These data suggest that Ku80 functions as a novel receptor for T␤4 and mediates its intracellular activity.2 is the most abundant member of the highly conserved family of acidic polypeptides called ␤-thymosins. Although it is a typical intracellular polypeptide, it plays numerous roles, both intracellularly and extracellularly. Numerous observations indicate that T␤4 is involved in adhesion and spreading of fibroblasts (1, 2), differentiation of endothelial cells (3, 4), directional migration of endothelial cells and keratinocytes (5-8), angiogenesis (3-6, 9, 10), wound healing (6, 7, 11), hair follicle growth (8), and apoptosis (12, 13), and has been described to possess anti-inflammatory properties (11,14). In addition, elevated T␤4 expression has been observed in various malignant cell lines and tumors (15-17) and its levels seem to be associated with increased tumorigenicity and metastatic potential (10, 13, 18 -20). The increased expression of T␤4 correlates with the invasive capability of the cells, the degree of morphologic transformation, and disintegration of actin filaments (21). Recent studies have also correlated increased T␤4 expression with potentiated cell growth (13, 22) but this observation seems not to be universal (2, 10).T␤4 is conside...

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“…It enhances endothelial cell migration and adhesion [13][14][15]. Thus, Tβ4 has been shown to promote tissue regeneration [14][15][16].…”

Section: Discussionmentioning

confidence: 99%

Thymosin β4 promotes angiogenesis and increases muscle progenitor cell density in ischemic skeletal muscle in a mouse model of hind limb ischemia

Zhou¹,

Martinez²,

Su³

et al. 2012

OJRM

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Aim: To determine the therapeutic effect of thymosin β4 (Tβ4) for treatment of ischemic limb disease in a mouse model. Methods: A mouse model of hindlimb ischemia was created by permanent ligation of femoral arteries and internal iliac artery. Tβ4 was dissolved in sterile saline and intramuscularly injected into the centre and periphery of ligation area in the treatment group (n = 10) starting from the surgery day until 4 weeks after surgery, while control animals received saline injection only (n = 9). All animals were sacrificed at 6 weeks after surgery and used for immunohistochemistry studies. Results: Tβ4 stimulated angiogenesis was evidenced by increased vascular density based on CD31 immunostaining, which was signifycantly increased in Tβ4 group (562.5 ± 78.4/mm 2 ) as compared with control group (371.1 ± 125.7/mm 2 ; p < 0.05). The arteriole density based on CD31 and SMA dual immunostaining was similar between the Tβ4 (27.2 ± 16.9/mm 2 ) and control (35.3 ± 6/mm 2 ; p > 0.05) groups. Tβ4 increased Pax3/7 + skeletal muscle progenitor cell density. Pax3/7 + cell density of Tβ4 group (13.7% ± 2%) was significantly higher than that of the control group (4.3% ± 1.6%, p < 0.05). However, the numbers of central nuclei fiber and central nuclei per fiber were insignificantly increased in Tβ4 group as compared to control group. The numbers of central nuclei fiber were 8.9 ± 2.1 and 9.5 ± 1.6, and the central nuclei per fiber were 0.25 ± 0.07 and 0.48 ± 0.09 for control and Tβ4 groups, respectively. Conclusions: This preliminary study suggests that localized delivery of Tβ4 increased angiogenesis and skeletal muscle progenitor cell density in ischemic skeletal muscle, but failed to promote skeletal muscle regeneration.

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Cited by 127 publications

References 40 publications

Binding of PAI-1 to Endothelial Cells Stimulated by Thymosin β4 and Modulation of Their Fibrinolytic Potential

Binding of PAI-1 to Endothelial Cells Stimulated by Thymosin β4 and Modulation of Their Fibrinolytic Potential

Ku80 as a Novel Receptor for Thymosin β4 That Mediates Its Intracellular Activity Different from G-actin Sequestering

Thymosin β4 promotes angiogenesis and increases muscle progenitor cell density in ischemic skeletal muscle in a mouse model of hind limb ischemia

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