J Vet Sci.  2008 Sep;9(3):267-272. 10.4142/jvs.2008.9.3.267.

The inhibitory effect of quercitrin gallate on iNOS expression induced by lipopolysaccharide in Balb/c mice

Affiliations
  • 1College of Veterinary Medicine, Chungbuk National University, Cheongju 361-763, Korea. bwahn@cbu.ac.kr
  • 2College of Pharmacy, Chungbuk National University, Cheongju 361-763, Korea.

Abstract

Quercetin 3-O-beta-(2''-galloyl)-rhamnopyranoside (QGR) is a naturally occurring quercitrin gallate, which is a polyphenolic compound that was originally isolated from Persicaria lapathifolia (Polygonaceae). QGR has been shown to have an inhibitory effect on nitric oxide (NO) production in lipopolysaccharide (LPS)-stimulated macrophage RAW 264.7 cells. Therefore, this study was conducted to investigate the inhibitory effect of QGR on nitric oxide production and inducible nitric oxide synthases (iNOS) expression in LPS-stimulated Balb/c mice. To accomplish this, 10 mg/kg of QGR was administered via gavage once a day for 3 days. iNOS was then induced by intraperitoneal injection of LPS. Six hours after the LPS treatment the animals were sacrificed under ether anethesia. The serum levels of NO were then measured to determine if QGR exerted an inhibitory effect on NO production in vivo. LPS induced an approximately 6 fold increase in the expression of NO. However, oral administration of QGR reduced the LPS induced increase in NO by half. Furthermore, RT-PCR and western blot analysis revealed that the increased levels of iNOS expression that occurred in response to treatment with LPS were significantly attenuated in response to QGR pretreatment. Histologically, LPS induced the infiltration of polymorphonuclear neutrophils in portal veins and sinusoids and caused the formation of a large number of necrotic cells; however, pretreatment with QGR attenuated these LPS induced effects. Taken together, these results indicate that QGR inhibits iNOS expression in vivo as well as in vitro and has antiinflammatory potentials.

Keyword

Balb/c mice; iNOS; lipopolysaccharide; quercitrin gallate

MeSH Terms

Animals
DNA Primers
Gene Expression Regulation, Enzymologic/drug effects
Lipopolysaccharides/*pharmacology
Liver/drug effects/enzymology
Mice
Mice, Inbred BALB C
Nitric Oxide/blood
Nitric Oxide Synthase Type II/drug effects/*genetics
Quercetin/*analogs & derivatives/pharmacology
RNA, Messenger/genetics
Reverse Transcriptase Polymerase Chain Reaction

Figure

  • Fig. 1 Chemical structure of Quercetin 3-O-β-(2"-galloyl) rhamnopyranoside.

  • Fig. 2 Effect of Quercetin 3-O-β-(2"-galloyl) rhamnopyranoside (QGR) on nitric oxide (NO) production in serum. Lipopolysaccharide (LPS) induced an approximately 6 fold increase in NO when compared to controls. Pretreatment of QGR attenuated approximately 50% of the LPS induced increase in NO. *Significantly different from control (p < 0.05).

  • Fig. 3 RT-PCR analysis of inducible nitric oxide synthases (iNOS) mRNA in liver samples. A shows representative bands from each group. B shows the normalized densitometric ratios of iNOS to GAPDH. Pretreatment with Quercetin 3-O-β-(2"-galloyl) rhamnopyranoside (QGR) significantly inhibited the iNOS mRNA expression that was induced by lipopolysaccharide (LPS). *Significantly different from control (p < 0.05), ‡Significantly different from LPS treatment (p < 0.05).

  • Fig. 4 Western blot analysis of inducible nitric oxide synthases (iNOS) protein in liver samples. A shows representative bands from each group. B shows normalized densitometric ratios of iNOS to β-actin. Pretreatment with Quercetin 3-O-β-(2"-galloyl) rhamnopyranoside (QGR) inhibited the iNOS protein expression that was induced by LPS. *Significantly different from control (p < 0.05).

  • Fig. 5 Effect of Quercetin 3-O-β-(2"-galloyl) rhamnopyranoside (QGR) on polymorphonuclear neutrophil (PMN) infiltration in the liver. A, B: control; C, D: lipopolysaccharide (LPS) treatment; E, F: QGR + LPS treatment. A, C, E: ×200; B, D, F: ×400. Arrows indicate the infiltration of PMN.

  • Fig. 6 The number of polymorphonuclear neutrophils (PMN) in liver samples. The number of PMN in 10 randomly selected high-power fields. Lipopolysaccharide (LPS) induced an obvious increase in the infiltration of PMN, but this increase was attenuated by pretreatment with Quercetin 3-O-β-(2"-galloyl) rhamnopyranoside (QGR). *Significantly different from control (p < 0.05), ‡Significantly different from LPS treatment (p < 0.05).


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