Depeng Li

Geniposide, from Gardenia jasminoides Ellis, inhibits the inflammatory response in the primary mouse macrophages and mouse models

Geniposide, a main iridoid glucoside component of gardenia fruit, has been known to exhibit antibacterial, anti-inflammatory and other important therapeutic activities. The objective of this study was to investigate the protective effects of geniposide on inflammation in lipopolysaccharide (LPS) stimulated primary mouse macrophages in vitro and LPS induced lung injury model in vivo. The expression of pro-inflammatory cytokines was determined by enzyme-linked immunosorbent assay (ELISA). Nuclear factor-kappa B (NF-κB), inhibitory kappa B (IκBα) protein, p38, extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK) and Toll-like receptor 4 (TLR4) were determined by Western blot. Further analysis was carried out in mTLR4 and mMD-2 co-transfected HEK293 cells. The results showed that geniposide markedly inhibited the LPS-induced TNF-α, IL-6 and IL-1β production both in vitro and in vivo. Geniposide blocked the phosphorylation of IκBα, p65, p38, ERK and JNK in LPS stimulated primary mouse macrophages. Furthermore, geniposide inhibited the expression of TLR4 in LPS stimulated primary mouse macrophages and inhibited the LPS-induced IL-8 production in HEK293-mTLR4/MD-2 cells. In vivo study, it was also observed that geniposide attenuated lung histopathologic changes in the mouse models. These results suggest that geniposide exerts an anti-inflammatory property by down-regulating the expression of TLR4 up-regulated by LPS. Geniposide is highly effective in inhibiting acute lung injury and may be a promising potential therapeutic reagent for acute lung injury treatment.

Emodin ameliorates lipopolysaccharide-induced mastitis in mice by inhibiting activation of NF-κB and MAPKs signal pathways

Emodin is an anthraquinone derivative from the Chinese herb Radix et Rhizoma Rhei. It has been reported that emodin possesses a number of biological properties, such as anti-inflammatory, anti-virus, anti-bacteria, anti-tumor, and immunosuppressive properties. However, the effect of emodin on mastitis is not yet known. The aim of this study was to investigate whether emodin has protective effect against lipopolysaccharide (LPS)-induced mastitis in a mouse model. The mouse model of mastitis was induced by injection of LPS through the duct of mammary gland. Emodin was administered intraperitoneally with the dose of 1, 2, and 4 mg/kg respectively 1h before and 12h after induction of LPS. Emodin significantly reduced infiltration of neutrophilic granulocyte, activation of myeloperoxidase (MPO), concentration of tumor necrosis factor-alpha (TNF-α), interleukin-1 beta (IL-1β), and interleukin-6 (IL-6), mRNA expression levels of TNF-α, IL-1β and IL-6, which were increased in LPS-induced mouse mastitis. In addition, emodin influenced nuclear factor kappa-B signal transduction pathway by inhibiting activation of nuclear transcription factor-kappaB (NF-κB) p65 and degradation inhibitor of NF-κB α (IκBα), and emodin also influenced mitogen activated protein kinases signal transduction pathway by depression activation of p38, extracellular signal-regulated kinase (ERK), and c-jun NH2-terminal kinase (JNK). In conclusion, these results indicated that emodin could exert beneficial effects on experimental mastitis induced by LPS and may represent a novel treatment strategy for mastitis.

Baicalin plays an anti-inflammatory role through reducing nuclear factor-κB and p38 phosphorylation in S. aureus-induced mastitis.

Mastitis is an inflammatory disease caused by microbial infection. Staphylococcus aureus is the major etiological microorganism responsible for both clinical and subclinical mastitis in dairy cows. A mouse model of S. aureus mastitis is available. Baicalin is isolated from Scutellaria and is known to have anti-inflammatory properties. This study was designed to evaluate the effects of baicalin in S. aureus mastitis. In the present study, the mouse model was infected with S. aureus to cause mammary gland inflammation. Baicalin treatment was administered from 6h until 24h after infection. Baicalin significantly attenuated inflammatory cell infiltration and decreased levels of TNF-α, IL-β, and IL-6. Further studies revealed that baicalin downregulated phosphorylation of NF-κB and p38 in the mammary gland with S. aureus mastitis. Our results demonstrated that baicalin reduced the expression of the proinflammatory cytokines TNF-α, IL-β, and IL-6 by inhibiting NF-κB and p38 phosphorylation and mRNA expression.

Staphylococcus aureus and Escherichia coli elicit different innate immune responses from bovine mammary epithelial cells.

Escherichia coli and Staphylococcus aureus are the most important pathogenic bacteria causing bovine clinical mastitis and subclinical mastitis, respectively. However, little is known about the molecular mechanisms underlying the different host response patterns caused by these bacteria. The aim of this study was to characterize the different innate immune responses of bovine mammary epithelium cells (MECs) to heat-inactivated E. coli and S. aureus. Gene expression of Toll-like receptor 2 (TLR2) and TLR4 was compared. The activation of nuclear factor kappa B (NF-κB) and the kinetics and levels of cytokine production were analyzed. The results show that the mRNA for TLR2 and TLR4 was up-regulated when the bovine MECs were stimulated with heat-inactivated E. coli, while only TLR2 mRNA was up-regulated when the bovine MECs were stimulated with heat-inactivated S. aureus. The expression of tumor necrosis factor-α (TNF-α), interleukin (IL)-1β, IL-6 and IL-8 increased more rapidly and higher when the bovine MECs were stimulated with heat-inactivated E. coli than when they were stimulated with heat-inactivated S. aureus. E. coli strongly activated NF-κB in the bovine MECs, while S. aureus failed to activate NF-κB. Heat-inactivated S. aureus could induce NF-κB activation when bovine MECs cultured in medium without fetal calf serum. These results were confirmed using TLR2- and TLR4/MD2-transfected HEK293 cells and suggested that differential TLR recognition and the lack of NF-κB activation account for the impaired immune response elicited by heat-inactivated S. aureus.

Lipopolysaccharide increases Toll-like receptor 4 and downstream Toll-like receptor signaling molecules expression in bovine endometrial epithelial cells

The endometrium is easily contaminated with bacteria and the endometrial epithelial cells (EECs) play an important role in defence against invading pathogens which recognized pathogen-associated molecular patterns (PAMPs) via pattern recognition receptors (PRRs). Toll-like receptor 4 (TLR4) can recognize lipopolysaccharide (LPS) from Gram-negative bacteria and initiates innate immune responses. In this study, we stimulated bovine EECs with LPS from Escherichia coli (E. coli). The expression of TLR4 was detected by quantitative real-time polymerase chain reaction (qRT-PCR) and western blot. The expression of downstream TLR4 signaling molecules was detected by qRT-PCR. The results showed that TLR4 and downstream adaptor molecules, transcription factors and cytokines were up-regulated when bovine EECs were stimulated with LPS. Furthermore, the expression of TOLLIP and β-defensin 5 were up-regulated when cells were stimulated with LPS. The results demonstrated that both MyD88 dependent and independent pathways in TLR4 were activated by LPS in bovine EECs. Bovine EECs have the immune repertoires required in defending against E. coli and play an important role in innate immune defence of the bovine endometrium.

Astragalin suppresses inflammatory responses via down-regulation of NF-κB signaling pathway in lipopolysaccharide-induced mastitis in a murine model

Mastitis is a prevalent and economic disease around the world and defined as infection and inflammation of the mammary gland. Astragalin, a bioactive component isolated from persimmon or Rosa agrestis, has been reported to have anti-inflammatory properties. To investigate the potential therapeutic effect of astragalin in mastitis, a murine model of mastitis was induced by administration of LPS in mammary gland. Astragalin was applied 1h before and 12h after LPS treatment. The results showed that astragalin attenuated the infiltration of inflammatory cells, the activity of myeloperoxidase (MPO) and the expression of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and interleukin-1β (IL-1β) in a dose-dependent manner. Additionally, Western blotting results showed that astragalin efficiently blunt decreased nuclear factor-kappaB (NF-κB) activation by inhibiting the degradation and phosphorylation of IκBα and the nuclear translocation of p65. These results suggested that astragalin exerts anti-inflammatory properties in LPS-mediated mastitis, possibly through inhibiting inhibition of the NF-κB signaling pathway, which mediates the expression of pro-inflammatory cytokines. Astragalin may be a potential therapeutic agent against mastitis.

RP105 involved in activation of mouse macrophages via TLR2 and TLR4 signaling

RP105 is a member of the toll-like receptor family of proteins that transmits an activation signal in B cells, playing a role in regulation of B cell growth and death; in macrophages and dendritic cells, RP105 is a specific inhibitor of TLR4 signaling. RP105 is uniquely important for regulating TLR4-dependent signaling. It also proved that RP105 is closely related to TLR2 in macrophage activation by Mycobacterium tuberculosis lipoproteins. The aim of our study is to investigate the role of RP105 in mouse macrophages activation of TLR4 and TLR2 signaling by lipopolysaccharides (LPS) and Pam3CysSerLys4 (Pam3CSK4) alone or in combination, and the interaction between TLR2 and TLR4 signaling through RP105. Our results indicate that besides exhibiting negative regulation of TNF-α and IL12-p40 secretion in macrophage activated by LPS, RP105 is also involved in macrophages activation by Pam3CSK4 through TLR2 signaling and exhibited regulation to IL-10 and RANTES production by mouse peritoneal macrophage activated by Pam3CSK4. In macrophages activation by LPS and Pam3CSK4 in combination, TLR2 signaling can overcome RP105-mediated regulation of TLR4 signaling. Thus, our data demonstrate that not only TLR4 signaling, but also RP105 appears to be an essential accessory for immune responses through TLR2 signaling. The function of TLR2 and TLR4 in response to TLR ligands could be associated with each other by RP105. These results can help us understanding the unique role of RP105 in macrophages response to TLR ligands.

Mangiferin inhibits mastitis induced by LPS via suppressing NF-ĸB and NLRP3 signaling pathways

&NA; During the past era, small molecules derived from various plants have attracted extensive attention for their versatile medicinal benefits. Among these, one organic molecule called mangiferin from certain plant species including mangoes and honey bush tea is widely used in treating inflammation. In this study, a LPS‐induced mastitis model in mouse is established to investigate the anti‐inflammatory effects and mechanism of mangiferin. The result shows that mangiferin significantly alleviates LPS‐induced histopathology, meanwhile, also decreases LPS‐induced MPO activity. Furthermore, mangiferin treatment remarkably impeded the expression of pro‐inflammatory cytokines TNF‐&agr;, IL‐1&bgr;, and IL‐6. In addition, mangiferin was found to inhibit LPS‐induced NF‐&kgr;B and NLRP3 inflammasome activation. In conclusion, these results suggested that LPS‐induced mastitis can be abated by mangiferin through inhibiting NF‐&kgr;B and NLRP3 signaling pathways. HighlightsMangiferin significantly inhibited mammary gland histopathologic changes and MPO activity.Mangiferin significantly inhibited LPS‐induced NF‐&kgr;B activation.Mangiferin significantly inhibited LPS‐induced NLRP3 activation.

Involvement of RP105 and toll-like receptors in the activation of mouse peritoneal macrophages by Staphylococcus aureus.

Staphylococcus aureus is the aetiological agent of many hospital‐ and community‐acquired infections. Toll‐like receptor 2 (TLR2) has been shown to play a crucial role in the host defence against S. aureus infection. The aim of this study is to investigate the roles of the heterogeneous TLR family proteins TLR2, TLR4 and RP105 during S. aureus infection. Peritoneal macrophages from mice were exposed to S. aureus. Their production of inflammatory cytokines and chemokines, their expression of cell‐surface markers and interactions between TLR2, TLR4 and RP105 were assessed in the presence or absence of inhibitory antibodies against TLR2, TLR4/MD‐2 and RP105/MD‐1 complexes. Our results demonstrate that not only TLR2 but also TLR4 and RP105 are involved in the response of macrophages to S. aureus, that TLR2, TLR4 and RP105 physically interact with each other during S. aureus infection, and that TLR2, TLR4 and RP105 both cooperate and play unique roles in the production of inflammatory cytokines (TNF‐α, IL‐12p40 and IL‐10) and chemokine (RANTES) by macrophages after S. aureus infection. This study characterizes the important roles that TLR2, TLR4 and RP105 play in host resistance against S. aureus infection.

Melatonin inhibits endoplasmic reticulum stress-associated TXNIP/NLRP3 inflammasome activation in lipopolysaccharide-induced endometritis in mice

ABSTRACT Endometritis, an inflammatory response of the uterus tissue, is characterized by the production of inflammatory cytokines and migration of neutrophil (PMN) into the uterus tissue. Melatonin has been demonstrated to have anti‐inflammatory and antioxidant effects. The purpose of this study was to investigate the protective effects of melatonin on lipopolysaccharide (LPS)‐induced endometritis in mice. An endometritis model was induced by LPS and melatonin was given 1 h before LPS treatment. The results showed that melatonin inhibited LPS‐induced pathologic changes, Myeloperoxidase (MPO) activity, and levels of interleukin‐1 beta (IL‐1&bgr;). Melatonin also inhibited LPS‐induced thioredoxin‐interacting protein (TXNIP)/NOD‐like receptor family, pyrin domain containing 3 (NLRP3) inflammasome and nuclear factor kappa B (NF‐&kgr;B) activation, reactive oxygen species (ROS) production, and endoplasmic reticulum (ER) stress. Furthermore, melatonin was found to increase AMPK activity. In conclusion, our results demonstrated that melatonin inhibited ER stress‐associated TXNIP/NLRP3 inflammasome activation with a regulation of adenosine monophosphate activated protein kinase (AMPK) in LPS‐induced endometritis. Melatonin may serve as a promising nutritional supplement for the treatment of endometritis. Graphical abstract Figure. No Caption available. HighlightsMelatonin protects LPS‐induced endometritis through anti‐inflammation and anti‐oxidant pathways.Melatonin inhibits TXNIP/NLRP3 inflammasome activation induced by LPS.Melatonin increases AMPK activity induced by LPS.