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Kallikrein gene-modified EPCs induce angiogenesis in rats with ischemic hindlimb and correlate with integrin αvβ3 expression.

Fu SS, Li FJ, Wang YY, You AB, Qie YL, Meng X, Li JR, Li BC, Zhang Y, Da Li Q - PLoS ONE (2013)

Bottom Line: Expressions of integrin αvβ3 and endothelial nitric oxide synthase (eNOS) were detected on the surface of EPCs.The levels of integrin αvβ3 expression were reduced by PI3K and eNOS blockade, and the inhibitor of integrin αvβ3 abrogated the migration and adhesion of hTK-transduced EPCs (P<0.05). hTK gene delivery in vivo improves the natural angiogenic response to ischemia.The ability of hTK gene-transduced EPCs can be enhanced in vitro, in which integrin αvβ3 plays a role in the process.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, Qilu Hospital, Shandong University, Jinan, Shandong Province, China.

ABSTRACT

Background: Human tissue kallikrein (hTK) plays an essential role in the physiological and pathological mechanisms of blood vessels. This study aimed to determine whether angiogenesis induced by endothelial progenitor cells (EPCs) transduced with the adenovirus-mediated hTK gene could improve blood flow in rat hindlimb ischemia in vivo and to establish a promising mechanism in vitro.

Methods: EPCs transduced with adenovirus encoding hTK-162 (i.e., Ad/hTK-transduced EPCs or Ad/GFP-transduced EPCs) were administered to Wister rats with hindlimb ischemia through therapeutic neovascularization. Muscular capillary density (MCD), blood flow (BF), and the number of myofibers were measured at days 7, 14, and 21 after treatment. Expressions of integrin αvβ3 and endothelial nitric oxide synthase (eNOS) were detected on the surface of EPCs.

Results: MCD, BF, and the number of myofibers in rats with Ad/hTK-transduced EPCs remarkably increased at day 21 after treatment compared with rats with Ad/GFP-transduced EPCs or the control group (P<0.01). Expressions of integrin αvβ3 and eNOS protein on the surface of EPCs also increased in rats with Ad/hTK-transduced EPCs. The levels of integrin αvβ3 expression were reduced by PI3K and eNOS blockade, and the inhibitor of integrin αvβ3 abrogated the migration and adhesion of hTK-transduced EPCs (P<0.05).

Conclusion: hTK gene delivery in vivo improves the natural angiogenic response to ischemia. The ability of hTK gene-transduced EPCs can be enhanced in vitro, in which integrin αvβ3 plays a role in the process.

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Related in: MedlinePlus

Surface expressions of integrinαvβ3 of EPCs.7A and 7B, surface expressions of integrinαvβ3 were analyzed using fluorescence- activated cell sorter (FACS) analysis. L-NAME was used to block the pathway of eNOS or wortmannin was used to block the pathway of PI3K. In Ad/hTK-transduced EPCs, the decreased expression levels of integrinαvβ3 are shown as compared with using saline (control). The FACS values were 18.1%, 8.29% and 7.88% for saline, L-NAME and wortmannin inhibitors, respectively. In EPCs, the FACS values were 7.51%, 5.61% and 4.87% for saline, L-NAME and wortmannin inhibitors, respectively. However expression levels of integrinαvβ3 on the surface of Ad/hTK-transduced EPCs were significant higher than on the surface of EPCs either using inhibitor of L-NAME/wortmannin or saline (P<0.05). Expression levels of integrinαvβ3 were decreased on the surface of the Ad/hTK-transduced EPCs and EPCs when compared with their controls (saline) (*P<0.05).
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pone-0073035-g007: Surface expressions of integrinαvβ3 of EPCs.7A and 7B, surface expressions of integrinαvβ3 were analyzed using fluorescence- activated cell sorter (FACS) analysis. L-NAME was used to block the pathway of eNOS or wortmannin was used to block the pathway of PI3K. In Ad/hTK-transduced EPCs, the decreased expression levels of integrinαvβ3 are shown as compared with using saline (control). The FACS values were 18.1%, 8.29% and 7.88% for saline, L-NAME and wortmannin inhibitors, respectively. In EPCs, the FACS values were 7.51%, 5.61% and 4.87% for saline, L-NAME and wortmannin inhibitors, respectively. However expression levels of integrinαvβ3 on the surface of Ad/hTK-transduced EPCs were significant higher than on the surface of EPCs either using inhibitor of L-NAME/wortmannin or saline (P<0.05). Expression levels of integrinαvβ3 were decreased on the surface of the Ad/hTK-transduced EPCs and EPCs when compared with their controls (saline) (*P<0.05).

Mentions: The migration of EPCs involved adhesion molecules. We investigated the effects on expression of adhesion molecules after NO inhibition by L-NAME and PI3K inhibition by wortmannin. Surface expressions of integrinαvβ3 were analyzed using FACS (fluorescence- activated cell sorter) analysis by flow cytometry. After blocking the pathway of eNOS by L-NAME or blocking the pathway of PI3K by wortmannin, decreased expression levels of integrin αvβ3 were observed as compared with using saline (control). However, expression levels of integrin αvβ3 on the Ad/hTK-EPCs were significant higher than those on the EPCs either using the inhibitor of L-NAME/wortmannin or saline (P<0.05) (Fig. 7A and Fig. 7B). In the pathway of eNOS inhibited by L-NAME, the FACS values of integrin αvβ3 expression for the three groups of Ad/hTK-transduced EPCs, Ad/GFP-transduced EPCs and non-transduced EPCs were 18.1%, 8.29% and 7.88%, respectively. In the pathway of PI3K inhibited by wortmannin, the FACS values of integrin αvβ3 expression for the three groups were 7.51%, 5.61% and 4.87%, respectively.


Kallikrein gene-modified EPCs induce angiogenesis in rats with ischemic hindlimb and correlate with integrin αvβ3 expression.

Fu SS, Li FJ, Wang YY, You AB, Qie YL, Meng X, Li JR, Li BC, Zhang Y, Da Li Q - PLoS ONE (2013)

Surface expressions of integrinαvβ3 of EPCs.7A and 7B, surface expressions of integrinαvβ3 were analyzed using fluorescence- activated cell sorter (FACS) analysis. L-NAME was used to block the pathway of eNOS or wortmannin was used to block the pathway of PI3K. In Ad/hTK-transduced EPCs, the decreased expression levels of integrinαvβ3 are shown as compared with using saline (control). The FACS values were 18.1%, 8.29% and 7.88% for saline, L-NAME and wortmannin inhibitors, respectively. In EPCs, the FACS values were 7.51%, 5.61% and 4.87% for saline, L-NAME and wortmannin inhibitors, respectively. However expression levels of integrinαvβ3 on the surface of Ad/hTK-transduced EPCs were significant higher than on the surface of EPCs either using inhibitor of L-NAME/wortmannin or saline (P<0.05). Expression levels of integrinαvβ3 were decreased on the surface of the Ad/hTK-transduced EPCs and EPCs when compared with their controls (saline) (*P<0.05).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3760867&req=5

pone-0073035-g007: Surface expressions of integrinαvβ3 of EPCs.7A and 7B, surface expressions of integrinαvβ3 were analyzed using fluorescence- activated cell sorter (FACS) analysis. L-NAME was used to block the pathway of eNOS or wortmannin was used to block the pathway of PI3K. In Ad/hTK-transduced EPCs, the decreased expression levels of integrinαvβ3 are shown as compared with using saline (control). The FACS values were 18.1%, 8.29% and 7.88% for saline, L-NAME and wortmannin inhibitors, respectively. In EPCs, the FACS values were 7.51%, 5.61% and 4.87% for saline, L-NAME and wortmannin inhibitors, respectively. However expression levels of integrinαvβ3 on the surface of Ad/hTK-transduced EPCs were significant higher than on the surface of EPCs either using inhibitor of L-NAME/wortmannin or saline (P<0.05). Expression levels of integrinαvβ3 were decreased on the surface of the Ad/hTK-transduced EPCs and EPCs when compared with their controls (saline) (*P<0.05).
Mentions: The migration of EPCs involved adhesion molecules. We investigated the effects on expression of adhesion molecules after NO inhibition by L-NAME and PI3K inhibition by wortmannin. Surface expressions of integrinαvβ3 were analyzed using FACS (fluorescence- activated cell sorter) analysis by flow cytometry. After blocking the pathway of eNOS by L-NAME or blocking the pathway of PI3K by wortmannin, decreased expression levels of integrin αvβ3 were observed as compared with using saline (control). However, expression levels of integrin αvβ3 on the Ad/hTK-EPCs were significant higher than those on the EPCs either using the inhibitor of L-NAME/wortmannin or saline (P<0.05) (Fig. 7A and Fig. 7B). In the pathway of eNOS inhibited by L-NAME, the FACS values of integrin αvβ3 expression for the three groups of Ad/hTK-transduced EPCs, Ad/GFP-transduced EPCs and non-transduced EPCs were 18.1%, 8.29% and 7.88%, respectively. In the pathway of PI3K inhibited by wortmannin, the FACS values of integrin αvβ3 expression for the three groups were 7.51%, 5.61% and 4.87%, respectively.

Bottom Line: Expressions of integrin αvβ3 and endothelial nitric oxide synthase (eNOS) were detected on the surface of EPCs.The levels of integrin αvβ3 expression were reduced by PI3K and eNOS blockade, and the inhibitor of integrin αvβ3 abrogated the migration and adhesion of hTK-transduced EPCs (P<0.05). hTK gene delivery in vivo improves the natural angiogenic response to ischemia.The ability of hTK gene-transduced EPCs can be enhanced in vitro, in which integrin αvβ3 plays a role in the process.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, Qilu Hospital, Shandong University, Jinan, Shandong Province, China.

ABSTRACT

Background: Human tissue kallikrein (hTK) plays an essential role in the physiological and pathological mechanisms of blood vessels. This study aimed to determine whether angiogenesis induced by endothelial progenitor cells (EPCs) transduced with the adenovirus-mediated hTK gene could improve blood flow in rat hindlimb ischemia in vivo and to establish a promising mechanism in vitro.

Methods: EPCs transduced with adenovirus encoding hTK-162 (i.e., Ad/hTK-transduced EPCs or Ad/GFP-transduced EPCs) were administered to Wister rats with hindlimb ischemia through therapeutic neovascularization. Muscular capillary density (MCD), blood flow (BF), and the number of myofibers were measured at days 7, 14, and 21 after treatment. Expressions of integrin αvβ3 and endothelial nitric oxide synthase (eNOS) were detected on the surface of EPCs.

Results: MCD, BF, and the number of myofibers in rats with Ad/hTK-transduced EPCs remarkably increased at day 21 after treatment compared with rats with Ad/GFP-transduced EPCs or the control group (P<0.01). Expressions of integrin αvβ3 and eNOS protein on the surface of EPCs also increased in rats with Ad/hTK-transduced EPCs. The levels of integrin αvβ3 expression were reduced by PI3K and eNOS blockade, and the inhibitor of integrin αvβ3 abrogated the migration and adhesion of hTK-transduced EPCs (P<0.05).

Conclusion: hTK gene delivery in vivo improves the natural angiogenic response to ischemia. The ability of hTK gene-transduced EPCs can be enhanced in vitro, in which integrin αvβ3 plays a role in the process.

Show MeSH
Related in: MedlinePlus