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Synthetic RGDS peptide attenuates lipopolysaccharide-induced pulmonary inflammation by inhibiting integrin signaled MAP kinase pathways.

Moon C, Han JR, Park HJ, Hah JS, Kang JL - Respir. Res. (2009)

Bottom Line: RGDS inhibited LPS-induced phosphorylation of focal adhesion kinase and MAP kinases, including ERK, JNK, and p38 MAP kinase, in lung tissue.Similarly, a blocking antibody against integrin alphav significantly inhibited LPS-induced inflammatory cell migration into the lung, protein accumulation and proinflammatory mediator production in BAL fluid, at 4 or 24 h post-LPS.These results suggest that RGDS with high specificity for alphavintegrins attenuates inflammatory cascade during LPS-induced development of acute lung injury.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Physiology, Division of Cell Biology, Ewha Medical Research Center, School of Medicine, Ewha Womans University, Seoul, Seoul, Korea. 94cmoon@hanmail.net

ABSTRACT

Background: Synthetic peptides containing the RGD sequence inhibit integrin-related functions in different cell systems. Here, we investigated the effects of synthetic Arg-Gly-Asp-Ser (RGDS) peptide on key inflammatory responses to intratracheal (i.t.) lipopolysaccharide (LPS) treatment and on the integrin signaled mitogen-activated protein (MAP) kinase pathway during the development of acute lung injury.

Methods: Saline or LPS (1.5 mg/kg) was administered i.t. with or without a single dose of RGDS (1, 2.5, or 5 mg/kg, i.p.), anti-alphav or anti-beta3 mAb (5 mg/kg, i.p.). Mice were sacrificed 4 or 24 h post-LPS.

Results: A pretreatment with RGDS inhibited LPS-induced increases in neutrophil and macrophage numbers, total protein levels and TNF-alpha and MIP-2 levels, and matrix metalloproteinase-9 activity in bronchoalveolar lavage (BAL) fluid at 4 or 24 h post-LPS treatment. RGDS inhibited LPS-induced phosphorylation of focal adhesion kinase and MAP kinases, including ERK, JNK, and p38 MAP kinase, in lung tissue. Importantly, the inhibition of the inflammatory responses and the kinase pathways were still evident when this peptide was administered 2 h after LPS treatment. Similarly, a blocking antibody against integrin alphav significantly inhibited LPS-induced inflammatory cell migration into the lung, protein accumulation and proinflammatory mediator production in BAL fluid, at 4 or 24 h post-LPS. Anti-beta3 also inhibited all LPS-induced inflammatory responses, except the accumulation of BAL protein at 24 h post-LPS.

Conclusion: These results suggest that RGDS with high specificity for alphavintegrins attenuates inflammatory cascade during LPS-induced development of acute lung injury.

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Related in: MedlinePlus

Effects of RGDS on LPS-induced TNF-α and MIP-2 productions, and on MMP-9 activity. Where indicated, mice were administered i.p. RGDS (1, 2.5 or 5 mg/kg) or RGES (5 mg/kg) once 1 h before LPS treatment and sacrificed 4 h post-LPS. Mice were also administered these peptides (5 mg/kg, i.p.) once 1 h before or 2 h after LPS and were sacrificed 24 h post-LPS (B). TNF-α and MIP-2 levels in BAL fluid samples were quantified using enzyme-linked immunosorbent assays (A, B, D). Gelatinolytic activity in BAL fluid samples (C, E). Samples were analyzed by zymography followed by scanning densitometry. The 92-kD genolytic bands corresponded to MMP-9. Densities are expressed as percentages versus saline treated controls. Values represent means ± SEM of results from 5 mice per group. *Significantly different from saline treated controls, p < 0.05; +significantly different from animals treated with LPS only, p < 0.05.
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Figure 4: Effects of RGDS on LPS-induced TNF-α and MIP-2 productions, and on MMP-9 activity. Where indicated, mice were administered i.p. RGDS (1, 2.5 or 5 mg/kg) or RGES (5 mg/kg) once 1 h before LPS treatment and sacrificed 4 h post-LPS. Mice were also administered these peptides (5 mg/kg, i.p.) once 1 h before or 2 h after LPS and were sacrificed 24 h post-LPS (B). TNF-α and MIP-2 levels in BAL fluid samples were quantified using enzyme-linked immunosorbent assays (A, B, D). Gelatinolytic activity in BAL fluid samples (C, E). Samples were analyzed by zymography followed by scanning densitometry. The 92-kD genolytic bands corresponded to MMP-9. Densities are expressed as percentages versus saline treated controls. Values represent means ± SEM of results from 5 mice per group. *Significantly different from saline treated controls, p < 0.05; +significantly different from animals treated with LPS only, p < 0.05.

Mentions: TNF-α, MIP-2, and MMP-9 are representative proinflammatory mediators, which play major roles in neutrophil influx and lung damage. TNF-α production and MMP-9 expression require integrin signaling, as demonstrated by in vitro inflammatory cell models [21-26]. Here, we evaluated the effects of RGDS on these proinflammatory mediators in BAL fluid. As shown in Figures 4A and 4B, RGDS administered 1 h before LPS inhibited LPS-induced increases in TNF-α and MIP-2 levels in BAL fluid at 4 h post-LPS. Moreover, these inhibitions were in dose-dependent and peaked at 5 mg/kg when these inhibitions of TNF-α and MIP-2 levels were 47 and 60%, respectively. At 24 h post-LPS, LPS-induced TNF-α levels in BAL fluid were decreased by a fifth of those at 4 h post-LPS and MIP-2 levels were decreased to saline control (data not shown). Pre- or posttreatment with RGDS (5 mg/kg) significantly inhibited LPS-induced TNF-α levels by 78 and 53%, respectively (Figure 4D).


Synthetic RGDS peptide attenuates lipopolysaccharide-induced pulmonary inflammation by inhibiting integrin signaled MAP kinase pathways.

Moon C, Han JR, Park HJ, Hah JS, Kang JL - Respir. Res. (2009)

Effects of RGDS on LPS-induced TNF-α and MIP-2 productions, and on MMP-9 activity. Where indicated, mice were administered i.p. RGDS (1, 2.5 or 5 mg/kg) or RGES (5 mg/kg) once 1 h before LPS treatment and sacrificed 4 h post-LPS. Mice were also administered these peptides (5 mg/kg, i.p.) once 1 h before or 2 h after LPS and were sacrificed 24 h post-LPS (B). TNF-α and MIP-2 levels in BAL fluid samples were quantified using enzyme-linked immunosorbent assays (A, B, D). Gelatinolytic activity in BAL fluid samples (C, E). Samples were analyzed by zymography followed by scanning densitometry. The 92-kD genolytic bands corresponded to MMP-9. Densities are expressed as percentages versus saline treated controls. Values represent means ± SEM of results from 5 mice per group. *Significantly different from saline treated controls, p < 0.05; +significantly different from animals treated with LPS only, p < 0.05.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2666640&req=5

Figure 4: Effects of RGDS on LPS-induced TNF-α and MIP-2 productions, and on MMP-9 activity. Where indicated, mice were administered i.p. RGDS (1, 2.5 or 5 mg/kg) or RGES (5 mg/kg) once 1 h before LPS treatment and sacrificed 4 h post-LPS. Mice were also administered these peptides (5 mg/kg, i.p.) once 1 h before or 2 h after LPS and were sacrificed 24 h post-LPS (B). TNF-α and MIP-2 levels in BAL fluid samples were quantified using enzyme-linked immunosorbent assays (A, B, D). Gelatinolytic activity in BAL fluid samples (C, E). Samples were analyzed by zymography followed by scanning densitometry. The 92-kD genolytic bands corresponded to MMP-9. Densities are expressed as percentages versus saline treated controls. Values represent means ± SEM of results from 5 mice per group. *Significantly different from saline treated controls, p < 0.05; +significantly different from animals treated with LPS only, p < 0.05.
Mentions: TNF-α, MIP-2, and MMP-9 are representative proinflammatory mediators, which play major roles in neutrophil influx and lung damage. TNF-α production and MMP-9 expression require integrin signaling, as demonstrated by in vitro inflammatory cell models [21-26]. Here, we evaluated the effects of RGDS on these proinflammatory mediators in BAL fluid. As shown in Figures 4A and 4B, RGDS administered 1 h before LPS inhibited LPS-induced increases in TNF-α and MIP-2 levels in BAL fluid at 4 h post-LPS. Moreover, these inhibitions were in dose-dependent and peaked at 5 mg/kg when these inhibitions of TNF-α and MIP-2 levels were 47 and 60%, respectively. At 24 h post-LPS, LPS-induced TNF-α levels in BAL fluid were decreased by a fifth of those at 4 h post-LPS and MIP-2 levels were decreased to saline control (data not shown). Pre- or posttreatment with RGDS (5 mg/kg) significantly inhibited LPS-induced TNF-α levels by 78 and 53%, respectively (Figure 4D).

Bottom Line: RGDS inhibited LPS-induced phosphorylation of focal adhesion kinase and MAP kinases, including ERK, JNK, and p38 MAP kinase, in lung tissue.Similarly, a blocking antibody against integrin alphav significantly inhibited LPS-induced inflammatory cell migration into the lung, protein accumulation and proinflammatory mediator production in BAL fluid, at 4 or 24 h post-LPS.These results suggest that RGDS with high specificity for alphavintegrins attenuates inflammatory cascade during LPS-induced development of acute lung injury.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Physiology, Division of Cell Biology, Ewha Medical Research Center, School of Medicine, Ewha Womans University, Seoul, Seoul, Korea. 94cmoon@hanmail.net

ABSTRACT

Background: Synthetic peptides containing the RGD sequence inhibit integrin-related functions in different cell systems. Here, we investigated the effects of synthetic Arg-Gly-Asp-Ser (RGDS) peptide on key inflammatory responses to intratracheal (i.t.) lipopolysaccharide (LPS) treatment and on the integrin signaled mitogen-activated protein (MAP) kinase pathway during the development of acute lung injury.

Methods: Saline or LPS (1.5 mg/kg) was administered i.t. with or without a single dose of RGDS (1, 2.5, or 5 mg/kg, i.p.), anti-alphav or anti-beta3 mAb (5 mg/kg, i.p.). Mice were sacrificed 4 or 24 h post-LPS.

Results: A pretreatment with RGDS inhibited LPS-induced increases in neutrophil and macrophage numbers, total protein levels and TNF-alpha and MIP-2 levels, and matrix metalloproteinase-9 activity in bronchoalveolar lavage (BAL) fluid at 4 or 24 h post-LPS treatment. RGDS inhibited LPS-induced phosphorylation of focal adhesion kinase and MAP kinases, including ERK, JNK, and p38 MAP kinase, in lung tissue. Importantly, the inhibition of the inflammatory responses and the kinase pathways were still evident when this peptide was administered 2 h after LPS treatment. Similarly, a blocking antibody against integrin alphav significantly inhibited LPS-induced inflammatory cell migration into the lung, protein accumulation and proinflammatory mediator production in BAL fluid, at 4 or 24 h post-LPS. Anti-beta3 also inhibited all LPS-induced inflammatory responses, except the accumulation of BAL protein at 24 h post-LPS.

Conclusion: These results suggest that RGDS with high specificity for alphavintegrins attenuates inflammatory cascade during LPS-induced development of acute lung injury.

Show MeSH
Related in: MedlinePlus