Hypericum perforatum L. attenuates depression by regulating Akkermansia muciniphila, tryptophan metabolism and NFκB-NLRP2-Caspase1-IL1ß pathway.

BACKGROUND: Gut microbiota dysbiosis significantly contributes to progression of depression. Hypericum perforatum L. (HPL) is traditionally used in Europe for treating depression. However, its mechanism remains largely underexplored. PURPOSE: This study aims to investigate the pivotal gut microbiota species and microbial signaling metabolites associated with the antidepressant effects of HPL. METHODS: Fecal microbiota transplantation was used to assess whether HPL mitigates depression through alterations in gut microbiota. Microbiota and metabolic profiling of control, chronic restraint stress (CRS)-induced depression, and HPL-treated CRS mice were examined using 16S rRNA gene sequencing and metabolomics analysis. The influence of gut microbiota on HPL's antidepressant effects was assessed by metabolite and bacterial intervention experiments. RESULTS: HPL significantly alleviated depression symptoms in a manner dependent on gut microbiota and restored gut microbial composition by enriching Akkermansia muciniphila (AKK). Metabolomic analysis indicated that HPL regulated tryptophan metabolism, reducing kynurenine (KYN) levels derived from microbiota and increasing 5-hydroxytryptophan (5-HTP) levels. Notably, supplementation with KYN activated the NFκB-NLRP2-Caspase1-IL1ß pathway and increased proinflammatory IL1ß in the hippocampus of mice with depression. Interestingly, mono-colonization with AKK notably increased 5-hydroxytryptamine (5-HT) and decreased KYN levels, ameliorating depression symptoms through modulation of the NFκB-NLRP2-Caspase1-IL1ß pathway. CONCLUSIONS: The promising therapeutic role of HPL in treating depression is primarily attributed to its regulation of the NFκB-NLRP2-Caspase1-IL1ß pathway, specifically by targeting AKK and tryptophan metabolites.

A chromosome-level genome assembly and evolutionary analysis of Coregonus ussuriensis Berg.

Coregonus ussuriensis Berg, distributed widely in cold waters above 45° N latitude, is a savored freshwater whitefish that has been included in the list of endangered animals as a consequence of overfishing. Lack of genomic information seriously hampers evolutionary and genetic research on C. ussuriensis warranting the need to assemble a high-quality reference genome to promote its genetic breeding. We assembled and constructed a reference chromosome-level C. ussuriensis genome (sequence length, 2.51 Gb; contig N50 length, 4.27 Mb) using PacBio sequencing and Hi-C assembly technology, 3,109 contigs were assembled into scaffolds, resulting in a genome assembly with 40 chromosomes and a scaffold N50 length of 62.20 Mb. In addition, 43,320 protein-coding genes were annotated. The peak Ks position in the species comparison reflects the whole-genome replication event of C. ussuriensis. This chromosome-level genome provides reference data for further studies on the molecular breeding of C. ussuriensis.

Influence of increasing acclimation temperature on growth, digestion, antioxidant capacity, liver transcriptome and intestinal microflora of Ussruri whitefish Coregonus ussuriensis Berg.

For effective restoration, conservation of Ussruri whitefish Coregonus ussuriensis Berg and coping with global climate change, effects of environmental temperature on Ussruri whitefish urgently need to be explored. In current study, the effects of different acclimation temperatures on the growth, digestive physiology, antioxidant ability, liver transcriptional responses and intestinal microflora patterns of Ussruri whitefish were investigated. Ussruri whitefish (15.20 g ± 1.23 g) were reared for 42 days under different acclimation temperatures, i.e., 10, 13, 16, 19, 22 and 25 °C, respectively. Result first determined 28 °C as the semi-lethal temperature in order to design the temperature gradient test. Highest main gain rate (MGR) and specific growth rate (SGR) were observed in fish group having acclimation temperature of 19 °C. Significantly decrease (P < 0.05) in triglyceride (TG) content appeared at 19 °C as compared to the 10 °C and 13 °C temperature groups. 19 °C notablely increased protease activities of stomach and intestine and intestinal lipase and amylase activities. 19 °C group obtained the highest activities of chloramphnicol acetyltransferase (CAT) and total antioxidant capacity (T-AOC) and higher activities of superoxide dismutase (SOD). The intestinal microflora composition was most conducive to maintaining overall intestinal health when the temperature was 19 °C, compared to 10 °C and 25 °C. Ussruri whitefish exposed to 10 °C and 25 °C possessed the lower Lactobacillus abundance compared to exposure to 19 °C. Temperature down to 10 °C or up to 25 °C, respectively, triggered cold stress and heat stress, which leading to impairment in intestinal digestion, liver antioxidant capacity and intestinal microflora structure. Liver transcriptome response to 10 °C, 19 °C and 25 °C revealed that Ussruri whitefish might require the initiation of endoplasmic reticulum stress to correct protein damage from cold-temperature and high-temperature stress, and it was speculated that DNAJB11 could be regarded as a biomarker of cold stress response.Based on the quadratic regression analysis of MGR and SGR against temperature, the optimal acclamation temperature were, respectively, 18.0 °C and 18.1 °C. Our findings provide valuable theoretical insights for an in-depth understanding of temperature acclimation mechanisms and laid the foundation for conservation and development of Ussruri whitefish germplasm resources.

Bergenin Alleviates Ulcerative Colitis By Decreasing Gut Commensal Bacteroides vulgatus-Mediated Elevated Branched-Chain Amino Acids.

Ulcerative colitis is closely associated with the dysregulation of gut microbiota. There is growing evidence that natural products may improve ulcerative colitis by regulating the gut microbiota. In this research, we demonstrated that bergenin, a naturally occurring isocoumarin, significantly ameliorates colitis symptoms in dextran sulfate sodium (DSS)-induced mice. Transcriptomic analysis and Caco-2 cell assays revealed that bergenin could ameliorate ulcerative colitis by inhibiting TLR4 and regulating NF-κB and mTOR phosphorylation. 16S rRNA sequencing and metabolomics analyses revealed that bergenin could improve gut microbiota dysbiosis by decreasing branched-chain amino acid (BCAA) levels. BCAA intervention mediated the mTOR/p70S6K signaling pathway to exacerbate the symptoms of ulcerative colitis in mice. Notably, bergenin greatly decreased the symbiotic bacteria Bacteroides vulgatus (B. vulgatus), and the gavage of B. vulgatus increased BCAA concentrations and aggravated the symptoms of ulcerative colitis in mice. Our findings suggest that gut microbiota-mediated BCAA metabolism plays a vital role in the protective effect of bergenin on ulcerative colitis, providing novel insights for ulcerative colitis prevention through manipulation of the gut microbiota.

Single and joint toxic effects of waterborne exposure to copper and cadmium on Coregonus ussuriensis Berg.

Heavy metal contamination severely affects the aquatic environment and organisms. Copper (Cu) and cadmium (Cd) are two of the most common heavy metal contaminants that impair the survival, development, and reproduction of aquatic organisms. With the growth of agriculture and industry, there is a possibility of heavy metal pollution in Coregonus ussuriensis Berg's water source. However, there are no published studies on the toxicity to C. ussuriensis. Acute toxicity experiments in C. ussuriensis revealed the 96-h median lethal concentrations of copper and cadmium to be 0.492 mg·L-1 (95% confidence interval: 0.452-0.529) and 1.548 mg·L-1 (95% confidence interval: 1.434-1.657), respectively, and safe concentrations of 4.92 µg·L-1 and 15.48 µg·L-1, respectively. C. ussuriensis was then treated for 96 h with Cu (20% of 96 h LC50), Cd (20% of 96 h LC50), and a combination of Cu and Cd (20% of Cu 96 h LC50 + 20% of Cd 96 h LC50). The histological damage caused by the three different exposure modes to the liver and gills of C. ussuriensis was verified using hematoxylin and eosin staining. All three exposure modes caused different degrees of vacuolization, nuclear consolidation, and necrosis in the liver tissue of C. ussuriensis and edema, hyperplasia, laminar fusion, and epithelial elevation in the gill tissue compared with the reference group. The severity of the damage increased with increasing exposure time. Anti-oxidant activity in the gill and liver tissues were measured using enzyme activity assay kits to reflect oxidative stress induced by copper and cadmium exposure alone and in combination. The enzyme activities of superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSH) were substantially higher than those in the reference groups. However, the activities of the enzymes decreased with increasing exposure time. Malondialdehyde (MDA) activity significantly increased during exposure in relation to that in the reference group. Analysis of immune gene expression in C. ussuriensis gill and liver tissues was executed using real-time inverse transcript polymerase chain response (RT-PCR). The expression levels of the pro-inflammatory cytokines interleukin one beta (IL-1ß) and tumor necrosis factor-alpha (TNF-α) were positively correlated with exposure time and were significantly upregulated with increasing exposure time. Metallothionein (MT) gene expression levels were significantly upregulated in the short term after exposure compared to the reference group but decreased with increasing exposure time. Our results indicate that exposure to aqueous copper and cadmium solutions, either alone or in combination, causes histopathological damage, oxidative stress, and immunotoxicity in C. ussuriensis gill and liver tissue. This study investigated the toxic effects of copper and cadmium on C. ussuriensis to facilitate the monitoring of heavy metals in water sources for healthy aquaculture.

Sinomenine ameliorates rheumatoid arthritis by modulating tryptophan metabolism and activating aryl hydrocarbon receptor via gut microbiota regulation.

Gut microbiota dysbiosis is associated with the development of rheumatoid arthritis (RA). Sinomenine (SIN) is an effective immunosuppressive and anti-inflammatory drug used for treating RA, but how SIN regulates gut microbiota to alleviate RA remains underexplored. To identify the critical gut microbial species and microbial metabolites associated with the RA-protective effects of SIN, the microbiota-dependent anti-RA effects of SIN were assessed by 16S rRNA gene sequencing, antibiotic treatment, and fecal microbiota transplantation. Metabolomics analysis, transcriptional analysis, and targeted bacteria/metabolites gavage were conducted to explore how SIN regulates gut microbiota to reduce the severity of RA. SIN could restore intestinal microbial balance by mainly modulating the abundance of Lactobacillus, and significantly relieve collagen-induced arthritis (CIA) symptoms in a gut microbiota-dependent manner. SIN significantly elevated microbial tryptophan metabolites indole-3-acrylic acid (IA), indole-3-propionic acid (IPA), and indole-3-acetic acid (IAA). Tryptophan metabolites supplementation could activate aryl hydrocarbon receptor (AhR) and regulate Th17/Treg balance in CIA rats. Intriguingly, SIN relieved the arthritis symptoms involving the enrichment of two beneficial anti-CIA Lactobacillus species, L. paracasei and L. casei by mono-colonization. The promising therapeutic function of SIN was mostly attributed to the activation of AhR by explicitly targeting the Lactobacillus and microbial tryptophan metabolites. The intestinal bacterium L. paracasei and L. casei may be used to reduce the severity of CIA.

Nobiletin Ameliorates Nonalcoholic Fatty Liver Disease by Regulating Gut Microbiota and Myristoleic Acid Metabolism.

Disturbance of the gut microbiota plays a critical role in the development of nonalcoholic fatty liver disease (NAFLD). Increasing evidence supports that natural products may serve as prebiotics to regulate the gut microbiota in the treatment of NAFLD. In the present study, the effect of nobiletin, a naturally occurring polymethoxyflavone, on NAFLD was evaluated, and metabolomics, 16S rRNA gene sequencing, and transcriptomics analysis were performed to determine the underlying mechanism of nobiletin, and the key bacteria and metabolites screened were confirmed by in vivo experiment. Nobiletin treatment could significantly reduce lipid accumulation in high-fat/high-sucrose diet-fed mice. 16S rRNA analysis demonstrated that nobiletin could reverse the dysbiosis of gut microbiota in NAFLD mice and nobiletin could regulate myristoleic acid metabolism, as revealed by untargeted metabolomics analysis. Treatment with the bacteria Allobaculum stercoricanis, Lactobacillus casei, or the metabolite myristoleic acid displayed a protective effect on liver lipid accumulation under metabolic stress. These results indicated that nobiletin might target gut microbiota and myristoleic acid metabolism to ameliorate NAFLD.

New 4',5'-methylenedioxyflavone derivatives from the whole plant of sarcandra glabra.

Two new natural products named 5,7-dihydroxy-3,3',6,8-tetramethoxy-4',5'-methylenedioxyflavone (1) and 3,3',5,7-tetramethoxy-4',5'-methylenedioxyflavone (2), along with thirteen known compounds, ß-sitosterol (3), desmethoxyyangonin (4), hexadecane (5), 3,9-bis(2,4-di-tert-butylphenoxy)-2,4,8,10-tetraoxa-3,9-diphosphaspiro [5.5] undecane 3,9-dioxide (6), 2'6'-dihydroxy-4'-methoxydihydrochalcone (7), cardamonin (8), 3,3',5,6,7,8-hexamethoxy-4',5'-methylenedioxyflavone (9), isofraxidin (10), aniba dimer A (11), 3,3',4',5,5',8-hexamethoxy-6,7-methylenedioxyflavone (12), quercetin (13), quercitrin (14) and isoquercitrin (15) were isolated from Sarcandra glabra (Thunb.) Nakai by various chromatographic methods. Compounds 1, 2, 4, 6, 11, and 12 were isolated from S. glabra for the first time. Their chemical structures were identified through the analysis of NMR and HR-MS spectra. The anti-inflammatory and cytotoxic activities of compounds 1-15 were evaluated in cell assays. The results indicated that compounds 1, 7, 8, 10, 14, and 15 significantly inhibited the NO production in LPS-induced RAW 264.7 murine macrophage cells. Moreover, compounds 1, 3, 4, 7, 8, 9, 10 and 12 exhibited a cytotoxic effect on the human HepG2 cell line.

Physiological responses to heat stress in the liver of rainbow trout (Oncorhynchus mykiss) revealed by UPLC-QTOF-MS metabolomics and biochemical assays.

Rainbow trout (Oncorhynchus mykiss) is one of the world's most widely farmed cold-water fish. However, the rise in water temperature caused by global warming has seriously restricted the development of rainbow trout aquaculture. In this study, we investigated the physiological responses in the liver of rainbow trout exposed to 20 â„ƒ and 24 â„ƒ and returning to the initial temperature (14 â„ƒ) by combining biochemical analyses and UPLC-QTOF-MS metabolomics. The results of the biochemical analysis showed that serum aminotransferase, lysozyme, total bilirubin, alkaline phosphatase and liver superoxide dismutase, glutathione peroxidase, and malondialdehyde in rainbow trout under heat stress changed significantly. Even after the temperature recovery, some of the above indicators were still affected. Compared to the control group, 115, 130, and 121 differentially expressed metabolites were identified in the 20 â„ƒ, 24 â„ƒ, and recovery groups, respectively. Further pathway enrichment of these metabolites revealed that heat stress mainly affected the linoleic acid metabolism, α-linolenic acid metabolism, glycerophospholipid metabolism, and sphingolipid metabolism in the liver of rainbow trout, and continuously affected these metabolic pathways during the recovery period. Notably, the enrichment of glutathione metabolic pathways was consistent with the changes in glutathione peroxidase in the biochemical results. The results above suggest that heat stress can induce immune responses and oxidative stress inside the rainbow trout. After temperature recovery, some of the hepatic functions of fish return to normal gradually. The biochemical analysis and UPLC-QTOF-MS metabolomics tools provide insight into the physiological regulation of rainbow trout in response to heat stress.

miR-301b-5p and its target gene nfatc2ip regulate inflammatory responses in the liver of rainbow trout (Oncorhynchus mykiss) under high temperature stress.

Rainbow trout (Oncorhynchus mykiss) is a typical cold-water aquaculture fish and a high-end aquatic product. When water temperature exceeds its optimal range of 12-18 °C, the immune system of rainbow trout becomes weakened and unbalanced. High temperature in summer and global warming severely impact rainbow trout industry. The focus of this study was to explore the mechanisms regulating the immune response of rainbow trout under high temperature stress and identify molecular elements that account for resistance to high temperature. In this study, individual fish were screened in a high temperature stress experiment and divided into resistant (R) and sensitive (S) groups. The hepatic transcriptome sequencing and analysis of mRNAs and microRNAs of the R, S, and control groups showed that the number of the differentially expressed genes (DEGs) in the S group (9259) was higher than that in the R group (5313). Furthermore, the 1233 genes differentially expressed between S and R groups were mainly enriched in immune-related pathways, including cytokine-cytokine receptor interaction, TNF signaling and IL-17 signaling. Among these DEGs were miR-301b-5p and its target gene that encodes nuclear factor of activated T cells two interacting protein (nfatc2ip). The dual-luciferase reporter system and immunofluorescence experiments verified the relationship between miR-301b-5p and nfatc2ip. We also showed that expression levels of miR-301b-5p and nfatc2ip significantly negatively correlated in the liver of rainbow trout under high temperature stress. By performing functional experiments, we showed that activation of miR-301b-5p expression or inhibition of nfatc2ip expression stimulated the phosphorylation of p65, p38, and JNK in the classical nuclear factor kappa-B and mitogen-activated protein kinase pathways under high temperature stress. These manipulations initially promoted the secretion of the pro-inflammatory factor IL-1ß and then increased the levels of IL-6, IL-12, and TNF-α. In addition, activation of miR-301b-5p expression or inhibition of nfatc2ip expression stimulated the repair of the hepatic ultrastructural damage caused by high temperature stress by activating the inflammatory response in rainbow trout liver.

Baicalin attenuate diet-induced metabolic syndrome by improving abnormal metabolism and gut microbiota.

In this work, we examined whether baicalin (BC), a bioactive flavonoid in Scutellaria baicalensis Georgi, can reduce high-fat diet (HFD)-induced metabolic syndrome (MetS) in mice. The UPLC-QTOF/MS was used for metabolome profiles analysis, and an analysis of bacterial 16S rDNA in feces was used to examine the effects of BC on gut microbiota composition. Our results showed that BC (400 mg/kg) could reduce the body weight gain, decrease hepatic fat accumulation and abnormal blood lipids, and increase insulin sensitivity after 8 weeks of treatment. BC could reverse the alteration of 7 metabolites induced by HFD and the metabolic pathways responsive to BC intervention including citrate cycle, alanine, aspartate and glutamate metabolism, glycerophospholipid metabolism, and aminoacyl-tRNA biosynthesis. 16S rDNA analysis demonstrated that BC altered the composition and function of gut microbiota in MetS mice. Notably, we found that the change in succinic acid was negatively associated with the changes in Bacteroides and Sutterella, and positively associated with the change in Mucispirillum. Moreover, we confirmed that succinic acid displayed a metabolic protective effect on MetS mice. The antibiotic treatment verified that BC exerts metabolic protection through gut microbiota. Our findings suggested BC may be a potential therapeutic drug to ameliorate diet induced MetS and gut microbiome may be a novel mechanistic target of BC for treatment of MetS.

Screening and Validation of p38 MAPK Involved in Ovarian Development of Brachymystax lenok.

Brachymystax lenok (lenok) is a rare cold-water fish native to China that is of high meat quality. Its wild population has declined sharply in recent years, and therefore, exploring the molecular mechanisms underlying the development and reproduction of lenoks for the purposes of artificial breeding and genetic improvement is necessary. The lenok comparative transcriptome was analyzed by combining single molecule, real-time, and next generation sequencing (NGS) technology. Differentially expressed genes (DEGs) were identified in five tissues (head kidney, spleen, liver, muscle, and gonad) between immature [300 days post-hatching (dph)] and mature [three years post-hatching (ph)] lenoks. In total, 234,124 and 229,008 full-length non-chimeric reads were obtained from the immature and mature sequencing data, respectively. After NGS correction, 61,405 and 59,372 non-redundant transcripts were obtained for the expression level and pathway enrichment analyses, respectively. Compared with the mature group, 719 genes with significantly increased expression and 1,727 genes with significantly decreased expression in all five tissues were found in the immature group. Furthermore, DEGs and pathways involved in the endocrine system and gonadal development were identified, and p38 mitogen-activated protein kinases (MAPKs) were identified as potentially regulating gonadal development in lenok. Inhibiting the activity of p38 MAPKs resulted in abnormal levels of gonadotropin-releasing hormone, follicle-stimulating hormone, and estradiol, and affected follicular development. The full-length transcriptome data obtained in this study may provide a valuable reference for the study of gene function, gene expression, and evolutionary relationships in B. lenok and may illustrate the basic regulatory mechanism of ovarian development in teleosts.

Molecular characterization and antibacterial immunity functional analysis of the antimicrobial peptide hepcidin from Coregonus ussuriensis berg.

Antimicrobial peptides are immune system molecules existing in different organisms including mollusks, crustaceans and vertebrates. Hepcidins are a group of cysteine rich antimicrobial peptides, which plays an important role in fish response to a variety of pathogens. In this study, we cloned and identified Hepcidin from the Coregonus ussuriensis Berg, and its functions in vivo and in vitro was investigated. Our results showed that, CuHepc contains a 267 bp coding sequence (CDS) region that encodes 88 putative amino acids with a molecular weight of 9.77 kD. Hepcidin transcripts were most abundant in the liver of healthy C. ussuriensis Berg. The synthesized Hepcidin peptide exhibited a wide range of antibacterial activity against Gram-positive and Gram-negative bacteria in vitro, and the results of in vivo bacterial attack assays showed that the CuHepc gene was differentially up-regulated in the six tissues investigated after infection with Aeromonas hydrophila. To analyze the changes in protein levels in C. ussuriensis, we generated Hepc polyclonal antibodies in rabbits and verified that the protein expression was increased after bacterial infection with Western blot assay. MIC assay results showed a geometric mean value of 5.513 µM for CuHepc peptide. In the in vivo experiment, immune-related genes IL-10, NF-κB, TLR3 were up-regulated post-infection CuHepc peptide in liver and intestine. Finally, CuHepc peptide reduced the tissues microbial load compared to infection with Aeromonas hydrophila. The above results indicate that Hepc plays a role in the immune response of C. ussuriensis to exogenous disturbances, indicate that CuHepc might act a candidate for modulation of the innate immune system in C. ussuriensis.

Comprehensive analysis of miRNA-mRNA/lncRNA during gonadal development of triploid female rainbow trout (Oncorhynchus mykiss).

Chromosomal ploidy manipulation is one of the means to create excellent germplasm. Triploid fish could provide an ideal sterile model for searching of a underlying mechanism of abnormality in meiosis. The complete understanding of the coding and noncoding RNAs regulating sterility caused by meiosis abnormality is still not well understood. By high-throughput sequencing, we compared the expression profiles of gonadal mRNA, long non-coding RNA (lncRNA), and microRNA (miRNA) at three different developmental stages between the diploid (XX) and triploid (XXX) female rainbow trout. These stages were gonads before differentiation (65 days post fertilisation, dpf), at the beginning of morphological differences (180 dpf) and showing clear difference between diploids and triploids (600 dpf), respectively. A majority of differentially expressed (DE) RNAs were identified, and 22 DE mRNAs related to oocyte meiosis and homologous recombination were characterized. The predicted miRNA-mRNA/lncRNA networks of 3 developmental stages were constructed based on the target pairs of DE lncRNA-miRNA and DE mRNA-miRNA. According to the networks, meiosis-related gene of ccne1 was targeted by dre-miR-15a-5p_R + 1, and 6 targeted DE lncRNAs were identified. Also, qRT-PCR was performed to validate the credibility of the network. Overall, this study explored the potential interplay between coding and noncoding RNAs during the gonadal development of polyploid fish. The mRNA, lncRNA and miRNA screened in this study may be helpful to identify the functional elements regulating fertility of rainbow trout, which may provide reference for character improvement in aquaculture.

Flavonoids from sea buckthorn: A review on phytochemistry, pharmacokinetics and role in metabolic diseases.

Sea buckthorn (Hippophae rhamnoides L., SBT) is being used as a folk medicine for their diverse medicinal properties. Flavonoids are generally considered as the main bioactive and characteristic ingredients in SBT. This review was conducted using a comprehensive literature search on the chemical components, quality control, pharmacokinetics of flavonoids from SBT (FSBT). Particularly, we highlighted the therapeutic potential in metabolic diseases and clinical applications of FSBT. More than 95 flavonoids have been identified from SBT. Although the oral bioavailability of FSBT was relatively low, FSBT displays significant effect on the regulation of metabolism to ameliorate metabolic disorders and their complications. There is a heightened need to explore the bioactive compounds in SBT and mechanism(s) of action of FSBT in order to fully understand the pathways of their activities. PRACTICAL APPLICATIONS: For years, due to the increasing emergence of metabolic syndrome and diverse functions of FSBT in regulating metabolism, they can be efficiently utilized for human health and have an urgent need to become a hotspot for research. This review will broaden the understanding of FSBT, providing some directions for further development and expanding the therapeutic applications of FSBT.

GDC-0575, a CHK1 Inhibitor, Impairs the Development of Colitis and Colitis-Associated Cancer by Inhibiting CCR2+ Macrophage Infiltration in Mice.

BACKGROUND: Checkpoint kinase 1 (CHK1) plays an important role in DNA damage response and cell cycle progression. Thus, targeting CHK1 is an efficient strategy for cancer therapy. PURPOSE: The present study aimed to investigate the potential therapeutic effects of GDC-0575, a CHK1-specific inhibitor, in colitis-associated cancer (CAC) and colitis. METHODS: We established a DSS-induced acute colitis model and an azoxymethane/dextran sodium sulfate (DSS)-induced CAC model using mice and tested the effect of GDC-0575 on them. Flow cytometry and immunofluorescence were employed to investigate the infiltration of immune cells, and inflammatory cytokine expression in the colon of mice with CAC or colitis was investigated using ELISA and qPCR. We also investigated the correlation between CHK1 and CCL2/CCR2 in human colorectal cancer (CRC) tissues. RESULTS: Administration of GDC-0575 significantly inhibited CHK1 expression in the colon and dramatically impaired the development of CAC and colitis in mice. Moreover, the inhibition of CHK1 expression resulted in efficient inhibition of infiltration by iNOS-positive macrophages, but had no significant effect on CD4 T cells, CD8 T cells, and myeloid-derived suppressor cells (MDSCs). Significant downregulation of TNF-α, IL-6, and IL-1ß and dramatic upregulation of IL-10 were observed in the colons of both mice with CAC and colitis treated with GDC-0575. CCL2 expression was also downregulated by GDC-0575 in both mice with CAC and colitis; this was followed by the inhibition of CCR2+ macrophage infiltration in the colon. Furthermore, we report a positive correlation between CHK1 expression and CCL2/CCR2 expression in the malignant tissues of patients with CRC. CONCLUSION: Taken together, we infer that GDC-0575 impairs the development of CAC and colitis by regulating cytokine expression and inhibiting CCR2+ macrophage infiltration in mice colon.

The complete mitochondrial genome of Neolissochilus benasi (Cypriniformes: Cyprinidae).

In this study, the complete mitochondrial genome of Neolissochilus benasi has been determined by polymerase chain reaction method for the first time. The overall base composition of N. benasi mitogenome is 31.8% for A, 27.4% for C, 15.9% for G and 25.0% for T. The percentage of G + C content is 41.3%. The mitogenome is a circular DNA molecule of 16 583 bp in length with a D-loop region and contains 22 transfer RNA (tRNA) genes, two ribosomal RNA (rRNA) genes and 13 protein-coding genes. The mitochondrial genome sequencing for N. benasi in this study provides important molecular data for further evolutionary analysis for Cyprinoidea.

Network pharmacology approach to elucidate possible action mechanisms of Sinomenii Caulis for treating osteoporosis.

ETHNOPHARMACOLOGICAL RELEVANCE: Sinomenii Caulis (SC) is a well-konwn traditional Chinese medicine used for treatment of rheumatoid arthritis (RA), dermatophytosis and paralysis. Patients with RA are usually secondary to osteoporosis, but the potential protective effect of SC on osteoporosis (OP) is seldom reported and its possible action mechanism is little known. AIM: The purpose of this study was to demonstrate the anti-osteoporosis effects of SC extract and alkaloids in prednisolone (Pre)-induced OP of zebrafish, and then to explore the potential mechanism of SC on system level by network pharmacology. METHODS: Firstly, zebrafish OP model was established to investigate the anti-osteoporosis effect of SC. Secondly, the targets of SC and OP from multiple databases were collected, and Compound-Target-Pathway network based on protein-protein interaction (PPI) was constructed. Moreover, gene enrichment and annotation were performed via the DAVID server. Finally, the reliability of the network pharmacology prediction results in Pre-induced OP of zebrafish was verified by qRT-PCR. RESULTS: The results indicated that SC extract and alkaloids have remarkable ability to promote bone formation of cranial bones and reduce TRAP contents in Pre-induced OP of zebrafish. 32 OP-related ingredients in SC and 77 OP-related targets were screened from multiple databases, and 15 OP-related pathways were enriched by the KEGG database. Further experimental validation indicated that SC extract and alkaloids could regulate the expression of MAPK14, CASP3, CXCL8, IL-1ß, IL6, PTGS2, TNF-α, ESR1, and MMP9 for treatment of OP. CONCLUSION: In summary, we conducted an integrative analysis to provide convincing evidence that SC may partially alleviate OP by inhibiting pro-inflammatory cytokines and regulating of RANK/RANKL/OPG system.

Identification and expression of the hepcidin gene from brown trout (Salmo trutta) and functional analysis of its synthetic peptide.

Hepcidin, a hepatic antimicrobial peptide, is a key player of the nonspecific immune system. The structure of hepcidin gene from brown trout (Bthepc) has been characterized at the molecular level. The 1158-bp mRNA generates a coding sequence (CDS) of 267 bp, which encodes an 88-amino acid protein. Molecular evolution analysis classified Bthepc to the family Salmonidae. Amino acid sequence homologies between Bthepc and hepcidin in other species such as Oncorhynchus mykiss, Salmo salar, and Hucho taimen were found to be 93.18%, 96.59%, and 92.05% respectively. The mature peptide and the signal peptide of Bthepc are made of 25 and 24 amino acids, respectively. Similar to the other species, eight conserved cysteines in the mature peptide of Bthepc are held together by four disulphide bonds. Expression profiling of Bthepc indicated its highest expression in the liver. Further, iron levels or inflammation did not induce the age-dependent expression of Bthepc. Bthepc mRNA expression analysis in six immune tissues (liver, gill, spleen, skin, head kidney and intestine) indicated different levels of increase when challenged with Aeromonas salmonicida and Aeromonas hydrophila. The antimicrobial activity of synthetic Bthepc to typical pathogens was verified in vitro. In addition, Bthepc showed moderate haemolytic activity to mammalian erythrocytes. The antimicrobial activity of Bthepc was attributed to the disruption of the bacterial outer membrane integrity, which was evident from our scanning electron microscopy results. In summary, hepcidin gene of brown trout was characterized, and its antimicrobial activity was verified on different levels.

Characterization, expression, and functional analysis of the hepcidin gene from Brachymystax lenok.

Hepcidin, a cysteine-rich antimicrobial peptide, is an important effector molecule in the innate immune system. Recently, Brachymystax lenok has become to be a valuable cold-water fish in China, particularly as the wild resources are rapidly declining. In this study, the hepcidin gene of Brachymystax lenok (Blhepc) has been cloned. The 870-bp mRNA contains a coding sequence (CDS) of 267 bp that encodes 88 amino acid residues. Amino acid sequence identities of Blhepc with hepcidin in Oncorhynchus mykiss, Salmo salar, and Hucho taimen were found to be 93.18%, 89.77% and 93.18%, respectively. Phylogenetic analysis indicated that Blhepc was clustered in the family Salmonidae. The putative signal peptide and the mature peptide contained 24 and 25 amino acid residues, respectively. The RXXR motif for recruitment of propeptide convertase was identified upstream of the mature peptide of Blhepc by sequence analysis. The N-terminal amino acid residues of the mature Blhepc peptide were Q-SH-L, a structure involved in regulating iron metabolism. Eight conserved cysteine residues in the mature peptide were held together by four disulfide bonds. Expression profiling of Blhepc indicated its highest level in the liver; its expression was stronger in males than in similar-aged females. Moreover, its expression in the liver increased significantly with age. Expression of Blhepc in six immune tissues showed increase in various degrees when challenged with Aeromonas salmonicida and Aeromonas hydrophila. A synthetic Blhepc mature peptide was validated to have significant antimicrobial activity against gram-negative and gram-positive bacteria and fungi in vitro. These results show that Blhepc may be an important component in the innate immunity of Brachymystax lenok, which could provide antimicrobial activities against invading pathogens.