Molecular cloning and expression analysis of adiponectin and its receptors (AdipoR1 and AdipoR2) in the hypothalamus of the Huoyan goose during different stages of the egg-laying cycle

Multiple amino acid sequence alignment of the Huoyan goose adiponectin protein with other vertebrate species. The colour black denotes 100 % conserved sequences, and the colour grey indicates non-conservative sequences. Gaps (−) were introduced to maximize the alignment. Sequences for the alignment were obtained from GenBank (accession numbers are in brackets): Anas platyrhynchos (ADA68839.1); Ovis aries (AHV91023.1); Canis lupus familiaris (BAD15362.1); Felis catus (BAF52934.1); Gallus (AAX40986.1); Homo sapiens (NP_004788.1); Meleagris gallopavo (XP_010714799.1); Mus musculus (NP_033735.3); and Sus scrofa (ABQ95350.1). (TIFF 3483 kb

Licochalcone A Protects the Blood–Milk Barrier Integrity and Relieves the Inflammatory Response in LPS-Induced Mastitis

Background/Aims: Mastitis is an acute clinical inflammatory response. The occurrence and development of mastitis seriously disturb women's physical and mental health. Licochalcone A, a phenolic compound in Glycyrrhiza uralensis, has anti-inflammatory properties. Here, we examined the effect of licochalcone A on blood-milk barrier and inflammatory response in LPS-induced mice mastitis.Methods:In vivo, we firstly established mice models of mastitis by canal injection of LPS to mammary gland, and then detected the effect of licochalcone A on pathological indexes, inflammatory responses and blood-milk barrier in this model. In vivo, Mouse mammary epithelial cells (mMECs) were treated with licochalcone A prior to the incubation of LPS, and then the inflammatory responses, tight junction which is the basic structure of blood-milk barrier were analyzed. Last, we elucidated the anti-inflammatory mechanism by examining the activation of mitogen-activated protein kinase (MAPK) and AKT/NF-κB signaling pathways in vivo and in vitro.Result: The in vivo results showed that licochalcone A significantly decreased the histopathological impairment and the inflammatory responses, and improved integrity of blood-milk barrier. The in vitro results demonstrated that licochalcone A inhibited LPS-induced inflammatory responses and increase the protein levels of ZO-1, occludin, and claudin3 in mMECs. The in vivo and in vitro mechanistic study found that the anti-inflammatory effect of licochalcone A in LPS-induced mice mastitis was mediated by MAPK and AKT/NF-κB signaling pathways.Conclusions and Implications: Our experiments collectively indicate that licochalcone A protected against LPS-induced mice mastitis via improving the blood–milk barrier integrity and inhibits the inflammatory response by MAPK and AKT/NF-κB signaling pathways

GLP-2 Suppresses LPS-Induced Inflammation in Macrophages by Inhibiting ERK Phosphorylation and NF-κB Activation

Background/Aims: GLP-2 has been shown to exert anti-inflammatory effects, but the underlying molecular mechanisms remained undefined. As macrophages are important in the development and maintenance of inflammation, we investigated whether exogenous GLP-2 modulates the expression of pro-inflammatory proteins in LPS stimulated murine peritoneal macrophages. Methods: Macrophages were pretreated with various concentrations of GLP-2 for 1 h and then stimulated with LPS. The effects on pro-inflammatory enzymes (iNOS and COX-2), and pro-inflammatory cytokines (TNF-a, IL-1ß and IL-6) were analysed by Western blotting, ELISA and qRT-PCR. We also examined whether NF-κB or MAPK signaling was involved in the effects of GLP-2. Results: In macrophages, GLP-2 blunted the effect of LPS on protein and mRNA expression levels of iNOS, COX-2, TNF-a, IL-1ß and IL-6. Pre-incubation of macrophages with GLP-2 also blunted LPS-induced IκB-a degradation, IκB-a phosphorylation and NF-κB translocation. In the presence of GLP-2, the effect of LPS treatment on ERK phosphorylation was also profoundly blunted. GLP-2 did, however, not significantly modify the effects of LPS on p38 and JNK activities. Conclusions: These findings demonstrate that in LPS primed macrophages, GLP-2 reduced pro-inflammatory enzymes and cytokine production via mechanisms involving the suppression of NF-κB activity and ERK phosphorylation

Additional file 1: Figure S1. of Molecular cloning and expression analysis of adiponectin and its receptors (AdipoR1 and AdipoR2) in the hypothalamus of the Huoyan goose during different stages of the egg-laying cycle

Nucleotide and deduced amino acid sequences of adiponectin. The nucleotide (black) and deduced amino acid (blue) sequences are shown and numbered on the left. The nucleotide sequence is numbered from the 5’ end. The first methionine (M) is the first deduced amino acid. The C1Q domain (amino acids 105–241) is shaded. The start codons (ATG) and the stop codons (TAA) are marked in bold red. (TIFF 315 kb

Additional file 2: Figure S2. of Molecular cloning and expression analysis of adiponectin and its receptors (AdipoR1 and AdipoR2) in the hypothalamus of the Huoyan goose during different stages of the egg-laying cycle

Nucleotide and deduced amino acid sequences of AdipoR1. The nucleotide (black) and deduced amino acid (blue) sequences are shown and numbered on the left. The nucleotide sequence is numbered from the 5’ end. The first methionine (M) is the first deduced amino acid. The start codons (ATG) and the stop codons (TAA) are marked in bold red. (TIFF 727 kb

Additional file 9: Figure S9. of Molecular cloning and expression analysis of adiponectin and its receptors (AdipoR1 and AdipoR2) in the hypothalamus of the Huoyan goose during different stages of the egg-laying cycle

The phylogenetic tree of the AdipoR2 protein was constructed using the neighbour-joining method with MEGA4. The amino acid sequences of adiponectin for these species were downloaded from the NCBI protein database. Their corresponding accession numbers are the same as those given in Table 2. The number at the branches denotes the bootstrap majority consensus values on 1000 replicates; the branch lengths represent the relative genetic distances among these species. (TIFF 7477 kb

Additional file 5: Figure S5. of Molecular cloning and expression analysis of adiponectin and its receptors (AdipoR1 and AdipoR2) in the hypothalamus of the Huoyan goose during different stages of the egg-laying cycle

Multiple amino acid sequence alignment of the Huoyan goose AdipoR1 protein with other vertebrate species. The colour black denotes 100 % conserved sequences, and the colour grey indicates non-conservative sequences. Gaps (−) were introduced to maximize the alignment. Sequences for the alignment were obtained from GenBank (accession numbers are in brackets): Anas platyrhynchos (ABI49513.2); Ovis aries (AHV91022.1); Canis lupus familiaris (XP_005622230.1); Felis catus (NP_001128153.1); Gallus (NP_001026198.1); Homo sapiens (NP_001277558.1); Taeniopygia guttata (XP_002198547.1); Mus musculus (NP_082596.2); and Sus scrofa (NP_001007194.1). (TIFF 4230 kb

Additional file 6: Figure S6. of Molecular cloning and expression analysis of adiponectin and its receptors (AdipoR1 and AdipoR2) in the hypothalamus of the Huoyan goose during different stages of the egg-laying cycle

Multiple amino acid sequence alignment of the Huoyan goose AdipoR2 protein with other vertebrate species. The colour black denotes 100 % conserved sequences, and the colour grey indicates non-conservative sequences. Gaps (−) were introduced to maximize the alignment. Sequences for the alignment were obtained from GenBank (accession numbers are in brackets): Anas platyrhynchos (ABC75392.1); Bos taurus (NP_001035589.1); Canis lupus familiaris (XP_005637426); Felis catus (XP_011282054.1); Gallus (NP_001007855.1); Homo sapiens (NP_078827.2); Meleagris gallopavo (XP_003202489.1); Mus musculus (NP_932102.2); and Sus scrofa (NP_001007193.1). (TIFF 5171 kb