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Exogenous pathogen and plant 15-lipoxygenase initiate endogenous lipoxin A4 biosynthesis.

Bannenberg GL, Aliberti J, Hong S, Sher A, Serhan C - J. Exp. Med. (2004)

Bottom Line: Hence, we incubated STAg itself with arachidonic acid and found using LC-UV-MS-MS-based lipidomics that STAg produced both 15-HETE and 5,15-diHETE, indicating that T. gondii carries 15-lipoxygenase activity.Local administration of a plant (soybean) lipoxygenase itself reduced neutrophilic infiltration in murine peritonitis, demonstrating that 15-lipoxygenase possesses antiinflammatory properties.Together, these results indicate that 15-lipoxygenase expressed by a pathogen as well as exogenously administered 15-lipoxygenase can interact with host biosynthetic circuits for endogenous "stop signals" that divert the host immune response and limit acute inflammation.

View Article: PubMed Central - PubMed

Affiliation: Center for Experimental Therapeutics and Reperfusion Injury, Department of Anesthesiology, Perioperative and Pain Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA.

ABSTRACT
Lipoxin A4 (LXA4) is a potent endogenous lipoxygenase-derived eicosanoid with antiinflammatory and proresolving properties. Supraphysiological levels of LXA4 are generated during infection by Toxoplasma gondii, which in turn reduces interleukin (IL) 12 production by dendritic cells, thus dampening Th1-type cell-mediated immune responses and host immunopathology. In the present work, we sought evidence for the structural basis of T. gondii's ability to activate LXA4 biosynthesis. Proteomic analysis of T. gondii extract (soluble tachyzoite antigen [STAg]), which preserves the immunosuppressive and antiinflammatory activity of the parasite, yielded several peptide matches to known plant lipoxygenases. Hence, we incubated STAg itself with arachidonic acid and found using LC-UV-MS-MS-based lipidomics that STAg produced both 15-HETE and 5,15-diHETE, indicating that T. gondii carries 15-lipoxygenase activity. In addition, T. gondii tachyzoites (the rapidly multiplying and invasive stage of the parasite) generated LXA4 when provided with arachidonic acid. Local administration of a plant (soybean) lipoxygenase itself reduced neutrophilic infiltration in murine peritonitis, demonstrating that 15-lipoxygenase possesses antiinflammatory properties. Administration of plant 15-lipoxygenase generated endogenous LXA4 and mimicked the suppression of IL-12 production by splenic dendritic cells observed after T. gondii infection or STAg administration. Together, these results indicate that 15-lipoxygenase expressed by a pathogen as well as exogenously administered 15-lipoxygenase can interact with host biosynthetic circuits for endogenous "stop signals" that divert the host immune response and limit acute inflammation.

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Local administration of plant 15-lipoxygenase diminishes zymosan A–stimulated leukocyte infiltration, a hallmark of acute inflammation. Peritonitis was induced by intraperitoneal injection of 1 mg zymosan A. Soybean lipoxygenase (10 or 100 × 103 U) or vehicle alone was administered 5 min before zymosan A (1 mg/ml saline i.p.), and after 2 h, peritoneal exudates were collected. (A) Inhibition of peritoneal exudate cell numbers (mean ± SEM; n = 5–7; *, significantly different from vehicle; P < 0.05). (inset) In-gel lipoxygenase activity stain with substrate arachidonic acid after native isoelectric focusing of soybean 15-lipoxygenase (LO; 5 × 103 U). (B) Peritoneal exudate LXA4 levels expressed as mean values ± SEM (n = 7–8).
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fig4: Local administration of plant 15-lipoxygenase diminishes zymosan A–stimulated leukocyte infiltration, a hallmark of acute inflammation. Peritonitis was induced by intraperitoneal injection of 1 mg zymosan A. Soybean lipoxygenase (10 or 100 × 103 U) or vehicle alone was administered 5 min before zymosan A (1 mg/ml saline i.p.), and after 2 h, peritoneal exudates were collected. (A) Inhibition of peritoneal exudate cell numbers (mean ± SEM; n = 5–7; *, significantly different from vehicle; P < 0.05). (inset) In-gel lipoxygenase activity stain with substrate arachidonic acid after native isoelectric focusing of soybean 15-lipoxygenase (LO; 5 × 103 U). (B) Peritoneal exudate LXA4 levels expressed as mean values ± SEM (n = 7–8).

Mentions: To determine whether exogenous 15-lipoxygenase itself carries antiinflammatory properties, we administered neat soybean lipoxygenase into murine peritoneum before inducing leukocytic inflammation with zymosan A. By native isoelectric focusing and in-gel lipoxygenase staining, the soybean lipoxygenase preparation used in this analysis appeared to consist of a single active isoenzyme with an apparent pI of ∼6.4 (Fig. 4 A, inset). By SDS-PAGE, followed by LC-MS-MS and peptide mapping, this lipoxygenase preparation was shown to be mainly composed of soybean lipoxygenase type-1 (unpublished data). Unexpectedly, as shown in Fig. 4 A, local administration of soybean 15-lipoxygenase type-1 gave a marked reduction in inflammation. Peritoneal exudate cell viability was >95% by trypan blue staining. Administration of soybean lipoxygenase gave a dose-dependent increase in exudate LXA4 levels, indicating that the enzyme remained active in vivo (Fig. 4 B). Together, these results indicate that administration of plant 15-lipoxygenase promotes local endogenous LXA4 biosynthesis, and attenuates zymosan A–stimulated murine peritonitis.


Exogenous pathogen and plant 15-lipoxygenase initiate endogenous lipoxin A4 biosynthesis.

Bannenberg GL, Aliberti J, Hong S, Sher A, Serhan C - J. Exp. Med. (2004)

Local administration of plant 15-lipoxygenase diminishes zymosan A–stimulated leukocyte infiltration, a hallmark of acute inflammation. Peritonitis was induced by intraperitoneal injection of 1 mg zymosan A. Soybean lipoxygenase (10 or 100 × 103 U) or vehicle alone was administered 5 min before zymosan A (1 mg/ml saline i.p.), and after 2 h, peritoneal exudates were collected. (A) Inhibition of peritoneal exudate cell numbers (mean ± SEM; n = 5–7; *, significantly different from vehicle; P < 0.05). (inset) In-gel lipoxygenase activity stain with substrate arachidonic acid after native isoelectric focusing of soybean 15-lipoxygenase (LO; 5 × 103 U). (B) Peritoneal exudate LXA4 levels expressed as mean values ± SEM (n = 7–8).
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Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2211821&req=5

fig4: Local administration of plant 15-lipoxygenase diminishes zymosan A–stimulated leukocyte infiltration, a hallmark of acute inflammation. Peritonitis was induced by intraperitoneal injection of 1 mg zymosan A. Soybean lipoxygenase (10 or 100 × 103 U) or vehicle alone was administered 5 min before zymosan A (1 mg/ml saline i.p.), and after 2 h, peritoneal exudates were collected. (A) Inhibition of peritoneal exudate cell numbers (mean ± SEM; n = 5–7; *, significantly different from vehicle; P < 0.05). (inset) In-gel lipoxygenase activity stain with substrate arachidonic acid after native isoelectric focusing of soybean 15-lipoxygenase (LO; 5 × 103 U). (B) Peritoneal exudate LXA4 levels expressed as mean values ± SEM (n = 7–8).
Mentions: To determine whether exogenous 15-lipoxygenase itself carries antiinflammatory properties, we administered neat soybean lipoxygenase into murine peritoneum before inducing leukocytic inflammation with zymosan A. By native isoelectric focusing and in-gel lipoxygenase staining, the soybean lipoxygenase preparation used in this analysis appeared to consist of a single active isoenzyme with an apparent pI of ∼6.4 (Fig. 4 A, inset). By SDS-PAGE, followed by LC-MS-MS and peptide mapping, this lipoxygenase preparation was shown to be mainly composed of soybean lipoxygenase type-1 (unpublished data). Unexpectedly, as shown in Fig. 4 A, local administration of soybean 15-lipoxygenase type-1 gave a marked reduction in inflammation. Peritoneal exudate cell viability was >95% by trypan blue staining. Administration of soybean lipoxygenase gave a dose-dependent increase in exudate LXA4 levels, indicating that the enzyme remained active in vivo (Fig. 4 B). Together, these results indicate that administration of plant 15-lipoxygenase promotes local endogenous LXA4 biosynthesis, and attenuates zymosan A–stimulated murine peritonitis.

Bottom Line: Hence, we incubated STAg itself with arachidonic acid and found using LC-UV-MS-MS-based lipidomics that STAg produced both 15-HETE and 5,15-diHETE, indicating that T. gondii carries 15-lipoxygenase activity.Local administration of a plant (soybean) lipoxygenase itself reduced neutrophilic infiltration in murine peritonitis, demonstrating that 15-lipoxygenase possesses antiinflammatory properties.Together, these results indicate that 15-lipoxygenase expressed by a pathogen as well as exogenously administered 15-lipoxygenase can interact with host biosynthetic circuits for endogenous "stop signals" that divert the host immune response and limit acute inflammation.

View Article: PubMed Central - PubMed

Affiliation: Center for Experimental Therapeutics and Reperfusion Injury, Department of Anesthesiology, Perioperative and Pain Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA.

ABSTRACT
Lipoxin A4 (LXA4) is a potent endogenous lipoxygenase-derived eicosanoid with antiinflammatory and proresolving properties. Supraphysiological levels of LXA4 are generated during infection by Toxoplasma gondii, which in turn reduces interleukin (IL) 12 production by dendritic cells, thus dampening Th1-type cell-mediated immune responses and host immunopathology. In the present work, we sought evidence for the structural basis of T. gondii's ability to activate LXA4 biosynthesis. Proteomic analysis of T. gondii extract (soluble tachyzoite antigen [STAg]), which preserves the immunosuppressive and antiinflammatory activity of the parasite, yielded several peptide matches to known plant lipoxygenases. Hence, we incubated STAg itself with arachidonic acid and found using LC-UV-MS-MS-based lipidomics that STAg produced both 15-HETE and 5,15-diHETE, indicating that T. gondii carries 15-lipoxygenase activity. In addition, T. gondii tachyzoites (the rapidly multiplying and invasive stage of the parasite) generated LXA4 when provided with arachidonic acid. Local administration of a plant (soybean) lipoxygenase itself reduced neutrophilic infiltration in murine peritonitis, demonstrating that 15-lipoxygenase possesses antiinflammatory properties. Administration of plant 15-lipoxygenase generated endogenous LXA4 and mimicked the suppression of IL-12 production by splenic dendritic cells observed after T. gondii infection or STAg administration. Together, these results indicate that 15-lipoxygenase expressed by a pathogen as well as exogenously administered 15-lipoxygenase can interact with host biosynthetic circuits for endogenous "stop signals" that divert the host immune response and limit acute inflammation.

Show MeSH
Related in: MedlinePlus