Common carp Peptidoglycan Recognition Protein 2 (CcPGRP2) plays a role in innate immunity for defense against bacterial infections. Fish & shellfish immunology PGRP is a family of pattern recognition molecules of the innate immune system. PGRPs are conserved from insects to mammals and have diverse functions in antimicrobial defense. Here we cloned a common carp PGRP ortholog, CcPGRP2 containing a conserved C-terminal PGRP domain. We tested the expression levels of CcPGRP2 in the liver, spleen, kidney, foregut, midgut, and hindgut of the highest level in the liver. The expression of CcPGRP2 upregulated in common carp infected with Aeromonas hydrophila (A. hydrophila) or Staphylococcus aureus (S. aureus). Recombinant CcPGRP2 protein expressed in Escherichia coli (E. coli) system and the purified CcPGRP2 could maintain the integrity of intestinal mucosa of common carp infected with A. hydrophila. In addition, CcPGRP2 could agglutinate or bind both gram-positive and gram-negative bacteria in a Zn-dependent manner. CcPGRP2 has a stronger agglutination and bacterial binding ability in gram-positive bacteria than in gram-negative bacteria. It is perhaps because CcPGRP2 could bind peptidoglycan (PGN) with a higher degree to lipopolysaccharide (LPS). And CcPGRP2 shows antimicrobial activities in the presence of Zn. Our results of CcPGRP2 provided new insight into the function of PGRP in the innate immunity of the common carp. 10.1016/j.fsi.2023.108564

Luteolin reduces inflammation in Staphylococcus aureus-induced mastitis by inhibiting NF-kB activation and MMPs expression. Guo Ying-Fang,Xu Nian-Nian,Sun Weijing,Zhao Yifan,Li Cheng-Ye,Guo Meng-Yao Oncotarget Mastitis is a serious and prevalent disease caused by infection by pathogens such as Staphylococcus aureus. We evaluated the anti-inflammatory effects and mechanism of luteolin, a natural flavonoid with a wide range of pharmacological activities, in a mouse model of S. aureus mastitis. We also treated cultured mouse mammary epithelial cells (mMECs) with S. aureus and luteolin. Histopathological changes were examined by H&E staining and the levels of inflammatory cytokine proteins were analyzed using ELISAs. We determined mRNA levels with qPCR and the level of NF-κB and matrix metalloproteinase (MMP) proteins by Western blotting. The observed histopathological changes showed that luteolin protected mammary glands with S. aureus infection from tissue destruction and inflammatory cell infiltration. Luteolin inhibited the expression of TNF-α, IL-1β, and IL-6, all of which were increased with S. aureus infection of mammary tissues and mMECs. S. aureus-induced TLR2 and TLR4 was suppressed by luteolin, as were levels of IκBα and NF-κB p65 phosphorylation and expression of MMP-2 and MMP-9. Levels of tissue inhibitor of metalloproteinases (TIMP)-1 and TIMP-2 were enhanced. These findings suggest luteolin is a potentially effective new treatment to reduce tissue damage and inflammation from S. aureus-induced mastitis. 10.18632/oncotarget.16092

Establishment and characterization of Yellow River carp (Cyprinus carpio haematopterus) muscle cell line and its application to fish virology and immunology. Fish & shellfish immunology The Yellow River carp (Cyprinus carpio haematopterus) is a vital economically farmed fish of the Cyprinidae family. With the development of intensive aquaculture, carp production has increased dramatically, leading to the frequent occurrence of various diseases. Cell lines are considered the most cost-effective resource for in vitro studies and are widely used for physiological and pathological studies because of accessibility and convenience. This research established a novel immortal cell line CCM (Yellow River carp muscle cells) derived from the carp muscle. CCM has been passed over 71 generations for 1 year. The morphology of CCM and the adhesion and extension processes were captured by light and electron microscopy. CCM were passaged every 3 days with 20% FBS DMEM/F12 at 1:3. The optimum conditions for CCM growth were 28 °C and 20% FBS concentration. DNA sequencing of 16S rRNA and COI showed that CCM was derived from carp. CCM positively reacts to anti-PAX7 and anti-MyoD antibodies of carp. Analysis of chromosomes revealed that the chromosomal pattern number of CCM was 100. Transfection experiment demonstrated that CCM might be utilized to express foreign genes. Furthermore, cytotoxicity testing showed that CCM was susceptible to Aeromonas hydrophila, Aeromonas salmonicida, Aeromonas veronii, and Staphylococcus Aureus. The organophosphate pesticides (chlorpyrifos and glyphosate) or heavy metals (Hg, Cd, and Cu) exhibited dose-dependent cytotoxicity against CCM. After LPS treatment, the MyD88-IRAKs-NFκB pathway stimulates inflammatory-related factor il1β, il8, il10, and nfκb expression. LPS did not seem to cause oxidative stress in CCM, and the expression of cat and sod was not affected. Poly (I:C) through TLR3-TRIF-MyD88-TRAF6-NFκB and TRIF-TRAF3-TBK1-IRF3 activated the transcription of related factors, increased expression of anti-viral protein, but no changes in apoptosis-related genes. To our knowledge, this is the first muscle cell line in Yellow River carp and the first study on the immune response signal pathways of Yellow River carp based on the muscle cell line. CCM cell line provides a more rapid and efficient experimental material for fish immunology research, and this study preliminarily elucidated its immune response strategy to LPS and poly (I:C). 10.1016/j.fsi.2023.108859