Elsevier

Fitoterapia

Volume 82, Issue 7, October 2011, Pages 976-982
Fitoterapia

Berberine hydrochloride attenuates cyclooxygenase-2 expression in rat small intestinal mucosa during acute endotoxemia

https://doi.org/10.1016/j.fitote.2011.05.013Get rights and content

Abstract

The effect of berberine hydrochloride (BBR) on inducible cyclooxygenase-2 (COX-2) in small intestinal mucosa and related mechanisms was investigated in a rat model of acute endotoxemia. The results showed that lipopolysaccharide (LPS) increased COX-2 expression, whereas SB202190 and BBR curtailed it. LPS increased phosphorylation of mucosal p38 MAPK and ATF2 as well as production of ATF2, whereas BBR attenuated these effects. LPS upregulated mucosal peroxisome proliferator-activated receptor gamma (PPARγ), but BBR reduced this receptor. GW9662 aggravated LPS-induced and reversed BBR-attenuated COX-2 expression. The findings showed that BBR ameliorated COX-2 overexpression partially via modulation of p38 and PPARγ pathways during acute endotoxemia.

Introduction

Cyclooxygenase-2, a key enzyme for prostaglandins (PGs) biosynthesis, is expressed at low or undetectable levels in healthy intestinal tract [1]. However, COX-2 can be induced by LPS and a plethora of proinflammatory cytokines such as TNFα and IL-1 during intestinal and systemic infection [2]. Abnormally high levels of COX-2 and PGs are detrimental to intestinal mucosal barrier leading to IBD and necrotizing enterocolitis (NEC) [2], [3], and so forth.

p38 is a member of the MAPK family that plays a crucial role in transducing extracellular signals into the nucleus. Evidence shows that p38 is involved in enterocyte apoptosis [4] and gut inflmmation [3], [4]. p38 activates downstream ATFs and C/EBPs to mediate COX-2 gene transcription [5]. In enterocyte, p38 is required and sufficient for COX-2 induction ex vivo [2], [3]. Additionally, p38 also participates in inducing COX-2 expression in vascular endothelial cells [6] and smooth muscle cells [7], and so on.

Other than p38, PPARγ in most cases exerts anti-inflammatory properties via competition with molecular targets such as NF-κB, AP-1, STAT-1 for co-factor such as CBP and p300 [8] or inhibition of IκBα degradation [9]. In gut PPARγ is mainly expressed in enterocyte, macrophage [10], [11] and vascular endothelium [12]. PPARγ is implicated in maintaining gut mucosal integrity and ameliorating gut inflammation including NEC [13], IBD [14], [15], gut ischemia reperfusion injury [16], [17] and postoperative ileus [18].

Berberine hydrochloride (Fig. 1), a natural extract from Rhizoma coptidis, has been used to treat gastrointestinal infections for years [19]. Oral berberine improves morphological damage and colonic shortening, inhibits MPO activity and lipid peroxidation as well as IL-8, iNOS, IL-1β, IL-6 and TNFα production, augments SOD, GSH-Px and IL-10 in experimental colitis [20], [21], [22]. In vitro, berberine represses COX-2 in oral cancer cells and enterocytes [23], [24], [25]. Further studies reveals that berberine negatively interferes with molecular targets such as AP-1 [25], TLR-4 [26], NF-kB [27] and p38 [28], [29]. Recently, berberine has been reported to repress TNFα, MCP-1, CRP and IL-6 in 3T3-L1 adipocyte and macrophage via activation of PPARγ pathway [30], [31].

The aim of this study was to investigate the effect of BBR on mucosal COX-2 expression and related mechanisms. The results demonstrated that BBR's attenuating LPS-induced COX-2 is linked with suppression of p38 pathway and activation of PPARγ pathway.

Section snippets

Experimental animal

The Nanjing University Animal Care and Use Committee approved this protocol. Male Sprague–Dawley rats (200–250 g) were used in this study. The animals were purchased from Laboratory Animal Center of Jinling Hospital and housed at an ambient temperature of 22–23 °C, maintained in a 12 h/12 h light–dark cycle, and were fed water and standard rat chow ad libitum.

Pre-treatment and post-treatment with BBR

BBR was obtained from Sigma-Aldrich (St Louis, USA). This botanic alkaloid was dissolved in DMSO solution and diluted in distilled water

Endotoxin upregulates COX-2 immunoreactivity

Positive immunostaining for mucosal COX-2 was seen in both control and LPS group rats (Fig. 2). Without LPS stimulation, COX-2 immunostaining was scatteredly distributed within the mucosal layer, mainly in crypt cells (CCs), villus cells (VCs) and vascular endothelial cells (VECs). On the contrary, LPS challenge significantly upregulated mucosal COX-2 immunoreactivity, and the positive cells were largely confined to the VCs, followed by the CCs and VECs.

BBR inhibits LPS-induced COX-2 overexpression

Besides immunostaining, mucosal COX-2

Discussion

Suppression of COX-2 ovexpression contributes to maintain gut mucosal integrity. In this study, we observed that endotoxin significantly upregulated COX-2 immunoreactivity in rat distal ileal mucosa. Positive cells were largely confined to enterocytes, especially villus epithelium, which was consistent with previous studies [3]. Other cell types such as vascular endothelial cell also produced COX-2. Semi-quantitative analysis revealed that LPS significantly raised mucosal COX-2 mRNA and

Acknowledgement

This work was supported by the National Basic Research Program (973 Program) in China (nos. 2007CB513005 and 2009CB522405), the Key Project of National Natural Science Foundation in China (30830098), the National Natural Science Foundation in China (30672061).

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