Limits...
KLF2 Is a novel transcriptional regulator of endothelial proinflammatory activation.

SenBanerjee S, Lin Z, Atkins GB, Greif DM, Rao RM, Kumar A, Feinberg MW, Chen Z, Simon DI, Luscinskas FW, Michel TM, Gimbrone MA, García-Cardeña G, Jain MK - J. Exp. Med. (2004)

Bottom Line: Therefore, identification of the regulatory factors that mediate the effects of these stimuli on endothelial function is of considerable interest.Finally, our studies implicate recruitment by KLF2 of the transcriptional coactivator cyclic AMP response element-binding protein (CBP/p300) as a unifying mechanism for these various effects.These data implicate KLF2 as a novel regulator of endothelial activation in response to proinflammatory stimuli.

View Article: PubMed Central - PubMed

Affiliation: Cardiovascular Division, Brigham and Women's Hospital, Harvard Medical School, 75 Francis St., Boston, MA 02115, USA.

ABSTRACT
The vascular endothelium is a critical regulator of vascular function. Diverse stimuli such as proinflammatory cytokines and hemodynamic forces modulate endothelial phenotype and thereby impact on the development of vascular disease states. Therefore, identification of the regulatory factors that mediate the effects of these stimuli on endothelial function is of considerable interest. Transcriptional profiling studies identified the Kruppel-like factor (KLF)2 as being inhibited by the inflammatory cytokine interleukin-1beta and induced by laminar shear stress in cultured human umbilical vein endothelial cells. Overexpression of KLF2 in umbilical vein endothelial cells robustly induced endothelial nitric oxide synthase expression and total enzymatic activity. In addition, KLF2 overexpression potently inhibited the induction of vascular cell adhesion molecule-1 and endothelial adhesion molecule E-selectin in response to various proinflammatory cytokines. Consistent with these observations, in vitro flow assays demonstrate that T cell attachment and rolling are markedly attenuated in endothelial monolayers transduced with KLF2. Finally, our studies implicate recruitment by KLF2 of the transcriptional coactivator cyclic AMP response element-binding protein (CBP/p300) as a unifying mechanism for these various effects. These data implicate KLF2 as a novel regulator of endothelial activation in response to proinflammatory stimuli.

Show MeSH

Related in: MedlinePlus

Effect of KLF2 on cytokine-mediated induction of adhesion molecules. (A) KLF2 inhibits VCAM-1 and E-selectin but not ICAM-1 mRNA. HUVECs were infected with the adenovirus (C, Ad-GFP; K2, Ad-KLF2) at the indicated dose, stimulated with IL-1β for 4 h, and total RNA was assessed for adhesion molecule expression. In contrast to VCAM-1 and E-selectin, no effect is observed on ICAM-1 expression. Exo-KLF2 refers to exogenously expressed mouse KLF2. Endo-KLF2 refers to endogenous human KLF2. (B) KLF2 inhibits IL-1β–mediated induction of VCAM-1 and E-selectin protein levels. Experiments were performed as described in A except cells were harvested for total protein and Western blot analysis was performed. (C) KLF2 inhibits VCAM-1 and E-selectin in response to multiple cytokines. HUVECs infected with the adenovirus at the indicated dose, stimulated with cytokine for 4 h, and total RNA was assessed for adhesion molecule expression. (D) KLF2 inhibits IL-1β–mediated T cell attachment and rolling to endothelial cells. HUVECs infected with the indicated adenovirus (Ctrl, Ad-GFP), stimulated with IL-1β, and then perfused with T cells under flow conditions (0.75 dynes/cm2). *P < 0.05.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2211816&req=5

fig3: Effect of KLF2 on cytokine-mediated induction of adhesion molecules. (A) KLF2 inhibits VCAM-1 and E-selectin but not ICAM-1 mRNA. HUVECs were infected with the adenovirus (C, Ad-GFP; K2, Ad-KLF2) at the indicated dose, stimulated with IL-1β for 4 h, and total RNA was assessed for adhesion molecule expression. In contrast to VCAM-1 and E-selectin, no effect is observed on ICAM-1 expression. Exo-KLF2 refers to exogenously expressed mouse KLF2. Endo-KLF2 refers to endogenous human KLF2. (B) KLF2 inhibits IL-1β–mediated induction of VCAM-1 and E-selectin protein levels. Experiments were performed as described in A except cells were harvested for total protein and Western blot analysis was performed. (C) KLF2 inhibits VCAM-1 and E-selectin in response to multiple cytokines. HUVECs infected with the adenovirus at the indicated dose, stimulated with cytokine for 4 h, and total RNA was assessed for adhesion molecule expression. (D) KLF2 inhibits IL-1β–mediated T cell attachment and rolling to endothelial cells. HUVECs infected with the indicated adenovirus (Ctrl, Ad-GFP), stimulated with IL-1β, and then perfused with T cells under flow conditions (0.75 dynes/cm2). *P < 0.05.

Mentions: As shown in Fig. 1 A, KLF2 mRNA expression is inhibited by IL-1β. Inflammatory cytokines are known to induce several effects on endothelial cells, such as the expression of key adhesion molecules like VCAM-1, E-selectin, and ICAM-1 (41, 5, 14). To determine the effect of KLF2 on endothelial activation in response to inflammatory cytokines, HUVECs were infected with Ad-GFP (control; C) or Ad-KLF2 (K2) for 24 h, stimulated with IL-1β for an additional 4 h, and assessed for adhesion molecule mRNA and protein abundance. As shown in Fig. 3 A, treatment of HUVECs with IL-1β strongly induced VCAM-1, E-selectin, and ICAM-1 in both uninfected and control virus-infected cells. In contrast, adenoviral overexpression of KLF2 (10 MOI) strongly inhibited the induction of VCAM-1 and E-selectin but not ICAM-1 mRNA (Fig. 3 A). This effect was dose dependent, since no significant effect was observed at a dose of 1 MOI. Consistent with these observations, KLF2 inhibited VCAM-1 and E-selectin protein expression but did not affect ICAM-1 (Fig. 3 B). We also found that the ability of KLF2 to inhibit adhesion molecule expression was seen with other proinflammatory agents such as LPS and TNFα (Fig. 3 C).


KLF2 Is a novel transcriptional regulator of endothelial proinflammatory activation.

SenBanerjee S, Lin Z, Atkins GB, Greif DM, Rao RM, Kumar A, Feinberg MW, Chen Z, Simon DI, Luscinskas FW, Michel TM, Gimbrone MA, García-Cardeña G, Jain MK - J. Exp. Med. (2004)

Effect of KLF2 on cytokine-mediated induction of adhesion molecules. (A) KLF2 inhibits VCAM-1 and E-selectin but not ICAM-1 mRNA. HUVECs were infected with the adenovirus (C, Ad-GFP; K2, Ad-KLF2) at the indicated dose, stimulated with IL-1β for 4 h, and total RNA was assessed for adhesion molecule expression. In contrast to VCAM-1 and E-selectin, no effect is observed on ICAM-1 expression. Exo-KLF2 refers to exogenously expressed mouse KLF2. Endo-KLF2 refers to endogenous human KLF2. (B) KLF2 inhibits IL-1β–mediated induction of VCAM-1 and E-selectin protein levels. Experiments were performed as described in A except cells were harvested for total protein and Western blot analysis was performed. (C) KLF2 inhibits VCAM-1 and E-selectin in response to multiple cytokines. HUVECs infected with the adenovirus at the indicated dose, stimulated with cytokine for 4 h, and total RNA was assessed for adhesion molecule expression. (D) KLF2 inhibits IL-1β–mediated T cell attachment and rolling to endothelial cells. HUVECs infected with the indicated adenovirus (Ctrl, Ad-GFP), stimulated with IL-1β, and then perfused with T cells under flow conditions (0.75 dynes/cm2). *P < 0.05.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2211816&req=5

fig3: Effect of KLF2 on cytokine-mediated induction of adhesion molecules. (A) KLF2 inhibits VCAM-1 and E-selectin but not ICAM-1 mRNA. HUVECs were infected with the adenovirus (C, Ad-GFP; K2, Ad-KLF2) at the indicated dose, stimulated with IL-1β for 4 h, and total RNA was assessed for adhesion molecule expression. In contrast to VCAM-1 and E-selectin, no effect is observed on ICAM-1 expression. Exo-KLF2 refers to exogenously expressed mouse KLF2. Endo-KLF2 refers to endogenous human KLF2. (B) KLF2 inhibits IL-1β–mediated induction of VCAM-1 and E-selectin protein levels. Experiments were performed as described in A except cells were harvested for total protein and Western blot analysis was performed. (C) KLF2 inhibits VCAM-1 and E-selectin in response to multiple cytokines. HUVECs infected with the adenovirus at the indicated dose, stimulated with cytokine for 4 h, and total RNA was assessed for adhesion molecule expression. (D) KLF2 inhibits IL-1β–mediated T cell attachment and rolling to endothelial cells. HUVECs infected with the indicated adenovirus (Ctrl, Ad-GFP), stimulated with IL-1β, and then perfused with T cells under flow conditions (0.75 dynes/cm2). *P < 0.05.
Mentions: As shown in Fig. 1 A, KLF2 mRNA expression is inhibited by IL-1β. Inflammatory cytokines are known to induce several effects on endothelial cells, such as the expression of key adhesion molecules like VCAM-1, E-selectin, and ICAM-1 (41, 5, 14). To determine the effect of KLF2 on endothelial activation in response to inflammatory cytokines, HUVECs were infected with Ad-GFP (control; C) or Ad-KLF2 (K2) for 24 h, stimulated with IL-1β for an additional 4 h, and assessed for adhesion molecule mRNA and protein abundance. As shown in Fig. 3 A, treatment of HUVECs with IL-1β strongly induced VCAM-1, E-selectin, and ICAM-1 in both uninfected and control virus-infected cells. In contrast, adenoviral overexpression of KLF2 (10 MOI) strongly inhibited the induction of VCAM-1 and E-selectin but not ICAM-1 mRNA (Fig. 3 A). This effect was dose dependent, since no significant effect was observed at a dose of 1 MOI. Consistent with these observations, KLF2 inhibited VCAM-1 and E-selectin protein expression but did not affect ICAM-1 (Fig. 3 B). We also found that the ability of KLF2 to inhibit adhesion molecule expression was seen with other proinflammatory agents such as LPS and TNFα (Fig. 3 C).

Bottom Line: Therefore, identification of the regulatory factors that mediate the effects of these stimuli on endothelial function is of considerable interest.Finally, our studies implicate recruitment by KLF2 of the transcriptional coactivator cyclic AMP response element-binding protein (CBP/p300) as a unifying mechanism for these various effects.These data implicate KLF2 as a novel regulator of endothelial activation in response to proinflammatory stimuli.

View Article: PubMed Central - PubMed

Affiliation: Cardiovascular Division, Brigham and Women's Hospital, Harvard Medical School, 75 Francis St., Boston, MA 02115, USA.

ABSTRACT
The vascular endothelium is a critical regulator of vascular function. Diverse stimuli such as proinflammatory cytokines and hemodynamic forces modulate endothelial phenotype and thereby impact on the development of vascular disease states. Therefore, identification of the regulatory factors that mediate the effects of these stimuli on endothelial function is of considerable interest. Transcriptional profiling studies identified the Kruppel-like factor (KLF)2 as being inhibited by the inflammatory cytokine interleukin-1beta and induced by laminar shear stress in cultured human umbilical vein endothelial cells. Overexpression of KLF2 in umbilical vein endothelial cells robustly induced endothelial nitric oxide synthase expression and total enzymatic activity. In addition, KLF2 overexpression potently inhibited the induction of vascular cell adhesion molecule-1 and endothelial adhesion molecule E-selectin in response to various proinflammatory cytokines. Consistent with these observations, in vitro flow assays demonstrate that T cell attachment and rolling are markedly attenuated in endothelial monolayers transduced with KLF2. Finally, our studies implicate recruitment by KLF2 of the transcriptional coactivator cyclic AMP response element-binding protein (CBP/p300) as a unifying mechanism for these various effects. These data implicate KLF2 as a novel regulator of endothelial activation in response to proinflammatory stimuli.

Show MeSH
Related in: MedlinePlus