Response signatures of intestinal microbiota and gene transcription of yellow catfish (Pelteobagrus fulvidraco) to Aeromonas hydrophila infection.

Bacterial intestinal inflammation is a common disease of yellow catfish (Pelteobagrus fulvidraco) in high-density aquaculture. Understanding the interactions between host and intestinal bacteria is helpful to intestinal inflammatory disease control. Here, we constructed a model of intestinal inflammation after Aeromonas hydrophila infection in yellow catfish, and characterized variations in gene expression and microbiome in the gut through high-throughput sequencing. Furthermore, host gene-microbiome interactions were identified. Histology observation showed disordered distribution of columnar epithelial cells and decrease of goblet cells in intestine. A total of 4741 genes showed differentially expression, mostly in comparisons between 12 hpi group with each other groups respectively, including control, 24 hpi and 48 hpi groups. These genes were enriched in immune-related pathways including the IL-17 signaling pathway, triggering strong inflammatory response at the invading stage within 12 h. Subsequently, the host strengthened energy consumption by activating carbohydrate and lipid metabolism pathways to repair the intestinal mucosal immune defense line. In addition, fish with A. hydrophila infection show decreased richness of gut microbial, reduced relative abundance of probiotics including Akkermansia, and elevated pathogenic bacteria such as Plesimonas. An integrative analysis identified A. hydrophila-related genes, such as il22 and stat3, for which expression level is closed associated with the shift of A. hydrophila-related bacteria relative abundance, such as Akkermansia and Cetobacterium. Aside from picturing the variations of intestine gene expression and mucosal microbiome of yellow catfish coping with A. hydrophila infection, our study probed the underlying host-microbe interactions in A. hydrophila infection induced intestinal inflammatory, providing new insights for disease control in aquaculture.

High temperature aggravated hypoxia-induced intestine toxicity on juvenile Chinese mitten crab (Eriocheir sinensis).

High temperature and hypoxia in water due to global warming threaten the growth and development of aquatic animals. In natural or cultured environments, stress usually does not occur independently, whereas the synergistic effect of high temperature and hypoxia on Chinese mitten crab (Eriocheir sinensis) are rarely reported. In this study, 450 juvenile crabs were equally divided into control group (24 °C ± 0.5 °C, DO 6.8 ± 0.1 mg/L), hypoxia stress group (24 °C ± 0.5 °C, DO 1 ± 0.1 mg/L) and combined stress group (30 °C ± 0.5 °C, DO 1 ± 0.1 mg/L), and the intestinal health status, microbial diversity and metabolite profiles were evaluated for 24 h treatment. The results showed that hypoxia stress induced the expression level of pro-inflammatory related genes were significantly up-regulated in intestine of juvenile E. sinensis, and intestinal peritrophic membrane factor related genes were significantly down-regulated. High temperature further amplified the effects of hypoxia on pro-inflammatory and peritrophic membrane factor-related genes. Interesting, hypoxia stress induced a significant up-regulated of intestinal antioxidant-related genes, whereas high temperature reversed this trend. In addition, single stress or/and combined stress led to changes in intestinal microbiota diversity and abundance, and intestinal metabolite profiles. Compared with hypoxia stress, the synergistic effect of high temperature and hypoxia led to an increase in the abundance of pathogenic bacteria and a decrease in the abundance of probiotic bacteria. Moreover, intestinal metabolic pathways were significantly changed, especially amino acid metabolism and glycerophospholipid metabolism. Therefore, the results indicated that hypoxia stress could induce intestinal inflammatory response and oxidative stress, and lead to abnormal changes in intestinal microbiota and metabolic profiles, whereas high temperature further aggravate the toxic effects of hypoxia on the intestine. This study preliminarily revealed the synergistic toxic effects of high temperature and hypoxia on the intestine of juvenile E. sinensis.

Sex-bias of core intestinal microbiota in different stocks of Chinese mitten crabs (Eriocheir sinensis).

The differences in intestinal microbiota composition are synergistically shaped by internal and external factors of the host. The core microbiota plays a vital role in maintaining intestinal homeostasis. In this study, we conducted 16S rRNA sequencing analysis to investigate the stability of intestinal microbiota and sex-bias of six stocks of Chinese mitten crabs (105 females; and 110 males). The dominant phyla in all six stocks were Proteobacteria, Tenericutes, Bacteroidetes and Firmicutes; however, their relative abundance differed significantly. Twenty-seven core operational taxonomic units (OTUs), corresponding to 18 genera, were screened. Correlation analysis revealed that OTUs of four stocks in the Yangtze River system play important roles in maintaining the stability of intestinal microbiota. Additionally, the core intestinal microbiota was significantly sex-biased, and the top three genera in terms of relative abundance (Acinetobacter, Vibrio, and Candidatus_Hepatoplasma) were significantly dominant in female crabs. Network structure analysis also confirmed gender differences in the association pattern of intestinal microbiota. The intestinal microbiota of male crabs has a higher degree of functional enrichment. This study provided a theoretical basis for further investigating exploring the shaping effect of gender and geographical factors on the intestinal microbiota of Chinese mitten crabs.

Systematic identification and analysis of immune-related circRNAs of Pelteobagrus fulvidraco involved in Aeromonas veronii infection.

Circular RNA (circRNA) represents a type of newly discovered non-coding RNA, distinguished by its closed loop structure formed through covalent bonds. Recent studies have revealed that circRNAs have crucial influences on host anti-pathogen responses. Yellow catfish (Pelteobagrus fulvidraco), an important aquaculture fish with great economic value, is susceptible to Aeromonas veronii, a common aquatic pathogen that can cause acute death. Here, we reported the first systematic investigation of circRNAs in yellow catfish, especially those associated with A. veronii infection at different time points. A total of 1205 circRNAs were identified, which were generated from 875 parental genes. After infection, 47 circRNAs exhibited differential expression patterns (named DEcirs). The parental genes of these DEcirs were functionally engaged in immune-related processes. Accordingly, seven DEcirs (novel_circ_000226, 278, 401, 522, 736, 843, and 975) and six corresponding parental genes (ADAMTS13, HAMP1, ANG3, APOA1, FGB, and RALGPS1) associated with immunity were obtained, and their expression was confirmed by RT-qPCR. Moreover, we found that these DEcir-gene pairs likely acted through pathways, such as platelet activation, antimicrobial humoral response, and regulation of Ral protein signal transduction, to influence host immune defenses. Additionally, integrated analysis showed that, of the 7 immune-related DEcirs, three targeted 16 miRNAs, which intertwined into circRNA-miRNA networks. These findings revealed that circRNAs, by targeting genes or miRNAs are highly involved in anti-bacterial responses in yellow catfish. Our study comprehensively illustrates the roles of circRNAs in yellow catfish immune defenses. The identified DEcirs and the circRNA-miRNA network will contribute to the further investigations on the molecular mechanisms underlying yellow catfish immune responses.

Assessing the ecotoxicity of florfenicol exposure at environmental levels: A case study of histology, apoptosis and microbiota in hepatopancreas of Eriocheir sinensis.

The intensification of production practices in the aquaculture industry has led to the indiscriminate use of antibiotics to combat diseases and reduce costs, which has resulted in environmental pollution, posing serious threats to aquaculture sustainability and food safety. However, the toxic effect of florfenicol (FF) exposure on the hepatopancreas of crustaceans remains unclear. Herein, by employing Chinese mitten crab (Eriocheir sinensis) as subjects to investigate the toxic effects on histopathology, oxidative stress, apoptosis and microbiota of hepatopancreas under environment-relevant (0.5 and 5 µg/L), and extreme concentrations (50 µg/L) of FF. Our results revealed that the damage of hepatopancreas tissue structure caused by FF exposure in a dose-and time-dependent manner. Combined with the increased expression of apoptosis-related genes (Caspase 3, Caspase 8, p53, Bax and Bcl-2) at mRNA and protein levels, activation of catalase (CAT) and superoxide dismutase (SOD), and malondialdehyde (MDA) accumulation, FF exposure also induced oxidative stress, and apoptosis in hepatopancreas. Interestingly, 7 days exposure triggered more pronounced toxic effect in crabs than 14 days under environment-relevant FF concentration. Integrated biomarker response version 2 (IBRv2) index indicated that 14 days FF exposure under extreme concentration has serious toxicity effect on crabs. Furthermore, 14 days exposure to FF changed the diversity and composition of hepatopancreas microbiota leading remarkable increase of pathogenic microorganism Spirochaetes following exposure to 50 µg/L of FF. Taken together, our study explained potential mechanism of FF toxicity on hepatopancreas of crustaceans, and provided a reference for the concentration of FF to be used in culture of Chinese mitten crab.

LncRNA pol-lnc78 as a ceRNA regulates antibacterial responses via suppression of pol-miR-n199-3p-mediated SARM down-regulation in Paralichthys olivaceus.

Long non-coding RNAs (lncRNAs) function as key modulators in mammalian immunity, particularly due to their involvement in lncRNA-mediated competitive endogenous RNA (ceRNA) crosstalk. Despite their recognized significance in mammals, research on lncRNAs in lower vertebrates remains limited. In the present study, we characterized the first immune-related lncRNA (pol-lnc78) in the teleost Japanese flounder ( Paralichthys olivaceus). Results indicated that pol-lnc78 acted as a ceRNA for pol-miR-n199-3p to target the sterile alpha and armadillo motif-containing protein (SARM), the fifth discovered member of the Toll/interleukin 1 (IL-1) receptor (TIR) adaptor family. This ceRNA network regulated the antibacterial responses of flounder via the Toll-like receptor (TLR) signaling pathway. Specifically, SARM acted as a negative regulator and exacerbated bacterial infection by inhibiting the expression of inflammatory cytokines IL-1ß and tumor necrosis factor-α (TNF-α). Pol-miR-n199-3p reduced SARM expression by specifically interacting with the 3' untranslated region (UTR), thereby promoting SARM-dependent inflammatory cytokine expression and protecting the host against bacterial dissemination. Furthermore, pol-lnc78 sponged pol-miR-n199-3p to ameliorate the inhibition of SARM expression. During infection, the negative regulators pol-lnc78 and SARM were significantly down-regulated, while pol-miR-n199-3p was significantly up-regulated, thus favoring host antibacterial defense. These findings provide novel insights into the mechanisms underlying fish immunity and open new horizons to better understand ceRNA crosstalk in lower vertebrates.

Integrated analysis of transcriptome, translatome and proteome reveals insights into yellow catfish (Pelteobagrus fulvidraco) brain in response to hypoxia.

Brain plays a central role in adapting to environmental changes and is highly sensitive to the oxygen level. Although previous studies investigated the molecular response of brain exposure to acute hypoxia in fish, the lack of studies at the translational level hinders further understanding of the regulatory mechanism response to hypoxia from multi-omics levels. Yellow catfish (Pelteobagrus fulvidraco) is an important freshwater aquaculture species; however, hypoxia severely restricts the sustainable development of its breeding industry. In the present study, the transcriptome, translatome, and proteome were integrated to study the global landscapes of yellow catfish brain response to hypoxia. The evidently increased amount of cerebral cortical cells with oedema and pyknotic nuclei has been observed in hypoxia group of yellow catfish. A total of 2750 genes were significantly changed at the translational level. Comparative transcriptional and translational analysis suggested the HIF-1 signaling pathway, autophagy and glycolysis/gluconeogenesis were up-regulated after hypoxia exposure. KEGG enrichment of translational efficiency (TE) differential genes suggested that the lysosome and autophagy were highly enriched. Our result showed that yellow catfish tends to inhibit the TE of genes by increasing the translation of uORFs to adapt to hypoxia. Correlation analysis showed that transcriptome and translatome exhibit higher correlation. In summary, this study demonstrated that hypoxia dysregulated the cerebral function of yellow catfish at the transcriptome, translatome, and proteome, which provides a better understanding of hypoxia adaptation in teleost.

Acute cold stress leads to zebrafish ovarian dysfunction by regulating miRNA and mRNA.

Temperature is a critical factor that regulates the reproduction processes in teleost. However, the gonadal response mechanism to cold stress in fish remains largely unknown. In the present study, female zebrafish were exposed to different extents of low temperatures at 18 °C and 10 °C for 48 h. The ovarian histology was remarkably damaged after cold stress exposure. Integrated analysis of miRNA-mRNA was used to investigate the ovarian response to acute cold stress. A large number of mRNAs and miRNAs were altered by cold stress, which are involved in extensive biological processes. It is indicated that the signal transduction of MAPK and Calcium signaling pathway is highly engaged in zebrafish ovary to adapt to cold stress. The immune system was dysregulated by cold stress while the ovarian autophagy was activated. Remarkably increased gene number related to reproductive functions was identified in the cold stress at 10 °C compared to the control. The cold stress-induced dysregulated reproductive genes include star, hsd3b1, hsd17b1, inha, insl3, amh, nanos1 and foxl2. Combined with the dysregulated insulin, IGF and progesterone signaling, it is suggested that cold stress affects ovarian function in multiple aspects, including oocyte meiosis, folliculogenesis, final maturation and ovarian maintenance. On the other hand, the ovarian miRNA-mRNA regulatory network response to cold stress was also constructed. Overall, our result revealed the ovarian response to cold stress in zebrafish and provided insight into the fish adaptation mechanism to acute temperature change.

Hypoxia stress induces complicated miRNA responses in the gill of Chinese mitten crab (Eriocheir sinensis).

Hypoxia caused by global climate change and human activities has become a growing concern eliciting serious damages to aquatic animals. microRNAs (miRNAs) as non-coding regulatory RNAs exert vital effects on hypoxia responses. Chinese mitten crab (Eriocheir sinensis) with the habitat on the sediment surface or the pond bottom is susceptible to oxygen deficiency. However, whether miRNAs are involved in the response of the crabs to hypoxia stress remains enigmas. In this study, we conducted the whole transcriptome-based miRNA-mRNA integrated analysis of Chinese mitten crab gill under hypoxic condition for 3 h and 24 h We found that the acute hypoxia induces complex miRNA responses with the extensive influences on their target genes that engaged in various bio-processes, especially those associated with immunity, metabolism and endocrine. The impact of hypoxia on crab miRNAs is severer, as the exposure lasts longer. In response to the dissolved oxygen fluctuation, the HIF-1 signaling is activated by miRNAs to cope with the hypoxia stress through strategies including balancing inflammatory and autophagy involved in immunity, changing metabolism to reducing energy consumption, and enhancing oxygen-carrying and delivering capacities. The miRNAs and their corresponding target genes engaged in hypoxia response were intertwined into an intricate network. Moreover, the top hub molecular, miR-998-y and miR-275-z, discovered from the network might serve as biomarkers for hypoxia response in crabs. Our study provides the first systemic miRNA profile of Chinese mitten crab induced by hypoxia stress, and the identified miRNAs and the interactive network add new insights into the mechanism of hypoxia response in crabs.

Effects of hypoxia and reoxygenation on apoptosis, oxidative stress, immune response and gut microbiota of Chinese mitten crab, Eriocheir sinensis.

Hypoxia causes irreversible damage to aquatic animals. However, few reports have explored the effect of hypoxia stress and reoxygenation on intestinal homeostatic imbalance and consequent hepatopancreas-intestine axis health in crustacean. Herein, 180 Chinese mitten crabs (Eriocheir sinensis) were equally divided into control (DO 7.0 ± 0.2 mg/L) and treatment groups. The treatment group was exposed with continuous hypoxic stress (DO 3.0 ± 0.1 mg/L) for 96 h and then reoxygenated (DO 6.9 ± 0.1 mg/L) for 96 h. The effects on intestines and hepatopancreas of Chinese mitten crab were investigated, and the role of gut microbiota in hypoxia induced damages was explored. Hypoxia impaired intestinal tissue structure, and decreased swelling and the number of goblet cells, which are features that did not significantly improve after reoxygenation. With prolonged hypoxic stress, the activities of antioxidant enzymes (LDH, SOD and CAT) and MDA content in intestine were significantly elevated. Moreover, the level of oxidative stress increased, which led to upregulated apoptosis rate and expression of apoptosis-related genes (Caspase 3, Caspase 8 and BAX). In addition, the expression of immune related genes (MyD88, ALF1, Relish and Crustin) in hepatopancreas and intestine was both significantly induced under hypoxia, which activated the immune defense mechanism of Chinese mitten crab to adapt to the hypoxic environment. Furthermore, diversity and relative abundance of gut microbiota decreased noticeably during hypoxic stress; the number of beneficial bacteria downregulated. Finally, KEGG pathway analysis revealed that nine pathways were significantly enriched in intestinal microorganisms, including autoimmune disease and environmental adaptation. Collectively, these results suggested that hypoxia negatively affected E. sinensis health by triggering oxidative stress, altering the composition of the gut microbiota and inhibiting the immune response.

Exposure to copper nanoparticles or copper sulfate dysregulated the hypothalamic-pituitary-gonadal axis, gonadal histology, and metabolites in Pelteobagrus fulvidraco.

This study evaluated the effects of chronic exposure to copper nanoparticles (Cu-NPs) and waterborne copper (CuSO4) on the reproductive system of yellow catfish (Pelteobagrus fulvidraco). Juvenile yellow catfish were exposed to 100 and 200 µg Cu/L Cu-NPs and 100 µg Cu/L CuSO4 for 42 days. The results showed clear reproductive defects in both female and male yellow catfish in the 200 µg Cu/L Cu-NPs and 100 µg Cu/L CuSO4 groups. Exposure to Cu-NPs or CuSO4 inhibited folliculogenesis and vitellogenesis in the ovaries, and spermatogenesis in the testes, accompanied by elevation of the apoptotic signal. Ultrastructural observations also revealed damaged organelles of gonadal cells in both testes and ovaries. Most of the hypothalamic-pituitary-gonadal (HPG) axis genes examined and serum sex steroid hormones tended to be downregulated after Cu exposure. Metabolomic analysis suggested that gonadal estradiol level is sensitive to Cu-NPs or CuSO4. The heat map of gonadal metabolomics suggested a similar effect of 200 µg Cu/L Cu-NPs and 100 µg Cu/L CuSO4 in both the ovaries and testes. Additionally, metabolomics data showed that the reproductive toxicity due to Cu-NPs and CuSO4 may occur via different metabolic pathways. Cu-NPs tend to dysregulate the metabolic pathways of sphingolipid and linoleic acid metabolism in the ovary and the biosynthesis of amino acids and pantothenate and CoA in the testis. Overall, these findings revealed the toxicological effects of Cu-NPs and CuSO4 on the HPG axis and gonadal metabolism in yellow catfish.

Diagnosis, differential diagnosis, and treatment for sudden sensorineural hearing loss: Current otolaryngology practices in China.

Introduction: Although sudden sensorineural hearing loss (SSNHL) has been attempted to be understood for 70 years, diagnosis and treatment strategies still have strong heterogeneity worldwide, which are reflected in the guidelines issued by countries and the clinical practice of otolaryngologists. Methods: Questionnaires were sent to registered otolaryngologists nationwide via an online questionnaire system. We investigated the current views and clinical practices of otolaryngologists in mainland China about the diagnosis, examination, and treatment strategies of SSNHL. Results: Most otolaryngologists supported diagnostic classification via audiograms. Regional economic situation and hospital grade affected application strategies for differential diagnosis. Regarding corticosteroid therapy, 54.9% of respondents opted to discontinue the drug 5 days after systemic administration. Both intratympanic therapy and post-auricular injections were selected by more than half of the respondents as initial and salvage treatments. Discussion: Chinese otolaryngologists exhibit heterogeneity in clinical practices for SSNHL, including distinct approaches to combination therapy and local application of steroids. This study pointed out Chinese doctors' similarities, differences, and unique strategies in diagnosing and treating SSNHL and analyzed the possible reasons to help the world understand the current otolaryngology practices in China.

Multi-omics analysis revealed the brain dysfunction induced by energy metabolism in Pelteobagrus vachelli under hypoxia stress.

Hypoxia in water environment has become increasingly frequent and serious due to global warming and environmental pollution. Revealing the molecular mechanism of fish hypoxia adaptation will help to develop markers of environmental pollution caused by hypoxia. Here, we used a multi-omics method to identify the hypoxia-associated mRNA, miRNA, protein, and metabolite involved in various biological processes in Pelteobagrus vachelli brain. The results showed that hypoxia stress caused brain dysfunction by inhibiting energy metabolism. Specifically, the biological processes involved in energy synthesis and energy consumption are inhibited in P. vachelli brain under hypoxia, such as oxidative phosphorylation, carbohydrate metabolism and protein metabolism. Brain dysfunction is mainly manifested as blood-brain barrier injury accompanied by neurodegenerative diseases and autoimmune diseases. In addition, compared with previous studies, we found that P. vachelli has tissue specificity in response to hypoxia stress and the muscle suffers more damage than the brain. This is the first report to the integrated analysis of the transcriptome, miRNAome, proteome, and metabolome in fish brain. Our findings could provide insights into the molecular mechanisms of hypoxia, and the approach could also be applied to other fish species. DATA AVAILABILITY: The raw data of transcriptome has been uploaded to NCBI database (ID: SUB7714154 and SUB7765255). The raw data of proteome has been uploaded to ProteomeXchange database (PXD020425). The raw data of metabolome has been uploaded to Metabolight (ID: MTBLS1888).

Blood redistribution preferentially protects vital organs under hypoxic stress in Pelteobagrus vachelli.

Blood redistribution occurs in mammals under hypoxia but has not been reported in fish. This study investigated the tissue damage, hypoxia-inducible factor (HIF) activation level, and blood flow changes in the brain, liver, and muscle of Pelteobagrus vachelli during the hypoxia process for normoxia-hypoxia-asphyxia. The results showed that P. vachelli has tissue specificity in response to hypoxic stress. Cerebral blood flow increased with less damage than in the liver and muscle, suggesting that P. vachelli may also have a blood redistribution mechanism in response to hypoxia. It is worth noting that severe hypoxia can lead to a sudden increase in the degree of brain tissue damage. In addition, higher dissolved oxygen levels activate HIF and may have contributed to the reduced damage observed in the brain. This study provides basic data for investigating hypoxic stress in fish.

Integrated Analysis of Transcriptome and Metabolome Reveals Distinct Responses of Pelteobagrus fulvidraco against Aeromonas veronii Infection at Invaded and Recovering Stage.

Yellow catfish (Pelteobagrus fulvidraco) is an important aquaculture fish susceptible to Aeromonas veronii infection, which causes acute death resulting in huge economic losses. Understanding the molecular processes of host immune defense is indispensable to disease control. Here, we conducted the integrated and comparative analyses of the transcriptome and metabolome of yellow catfish in response to A. veronii infection at the invaded stage and recovering stage. The crosstalk between A. veronii-induced genes and metabolites uncovered the key biomarkers and pathways that strongest contribute to different response strategies used by yellow catfish at corresponding defense stages. We found that at the A. veronii invading stage, the immune defense was strengthened by synthesizing lipids with energy consumption to repair the skin defense line and accumulate lipid droplets promoting intracellular defense line; triggering an inflammatory response by elevating cytokine IL-6, IL-10 and IL-1ß following PAMP-elicited mitochondrial signaling, which was enhanced by ROS produced by impaired mitochondria; and activating apoptosis by up-regulating caspase 3, 7 and 8 and Prostaglandin F1α, meanwhile down-regulating FoxO3 and BCL6. Apoptosis was further potentiated via oxidative stress caused by mitochondrial dysfunction and exceeding inflammatory response. Additionally, cell cycle arrest was observed. At the fish recovering stage, survival strategies including sugar catabolism with D-mannose decreasing; energy generation through the TCA cycle and Oxidative phosphorylation pathways; antioxidant protection by enhancing Glutathione (oxidized), Anserine, and α-ketoglutarate; cell proliferation by inducing Cyclin G2 and CDKN1B; and autophagy initiated by FoxO3, ATG8 and ATP6V1A were highlighted. This study provides a comprehensive picture of yellow catfish coping with A. veronii infection, which adds new insights for deciphering molecular mechanisms underlying fish immunity and developing stage-specific disease control techniques in aquaculture.

M1 macrophage-derived exosomes and their key molecule lncRNA HOTTIP suppress head and neck squamous cell carcinoma progression by upregulating the TLR5/NF-κB pathway.

Exosomes serve as a crucial mode of communication between tumor-associated macrophages (TAMs) and cancer cells. This study attempted to explore the function of M1-derived exosomes and clarify their specific mechanism in head and neck squamous cell carcinoma (HNSCC). Moreover, the functional roles of M1-derived exosomes and their key molecule long noncoding RNA (lncRNA) HOXA transcript at the distal tip (HOTTIP) in HNSCC were investigated by conducting a series of in vitro and in vivo experiments. The dual-luciferase test was utilized to clarify the binding capacities between HOTTIP/mRNA and miRNAs. Accordingly, HOTTIP was found to be upregulated in M1-derived exosomes. Meanwhile, the in vitro experiments indicated that M1 exosomes suppressed proliferation, migration and invasion but induced apoptosis of cancer cells. This function was noted to be enhanced by HOTTIP-overexpressed M1 exosomes but was weakened by HOTTIP-knockdown ones, indicating that HOTTIP serves as a key molecule in M1 exosomes. Therefore, the function of HOTTIP in cancer cells was explored, for which overexpression of HOTTIP was found to inhibit proliferation, migration and invasion but induced apoptosis of cancer cells in vitro. A mechanism study further showed that M1 exosomes and HOTTIP activated the TLR5/NF-κB signaling pathway by competitively sponging miR-19a-3p and miR-19b-3p. Furthermore, cancer cells expressing HOTTIP were noted to induce the polarization of both local M1 and M2 macrophages; however, M1 exosomes were observed to reprogram local TAMs into M1 macrophages. More importantly, both cancer cells expressing HOTTIP and M1 exosomes reeducated circulating monocytes to express the M1 phenotype. The corresponding data demonstrated that the M1 exosomal lncRNA HOTTIP suppresses HNSCC progression by upregulating the TLR5/NF-κB signaling pathway through competitively sponging miR-19a-3p and miR-19b-3p. In particular, M1 exosomes and HOTTIP induce the polarization of M1 in circulating monocytes, thus providing novel insight into HNSCC immunotherapy.

Tissue distribution and seasonal accumulation of carotenoids in Yesso scallop (Mizuhopecten yessoensis) with orange adductor muscle.

Carotenoids are colored compounds with important physiological functions. The Haida golden scallop, which has an orange adductor muscle, is a carotenoid-enriched variety of scallop Mizuhopecten yessoensis, an important aquaculture shellfish. In this study, we investigated the tissue distribution of the carotenoids, pectenolone and pectenoxanthin, in both Haida golden scallop and normal Yesso scallop. Both carotenoids were detected in all the sampled tissues of the two scallops, except in the adductor muscle of normal scallop. There were significantly more carotenoids in Haida golden scallop than in normal scallop, in the tissues of the mantle, female gonad, kidney, and adductor muscle. Increased carotenoid concentrations were detected in Haida golden scallop adductor muscle during the spring spawning season, indicating the effects of reproduction on muscle carotenoids accumulation. This study was the first systematic investigation of carotenoid distribution in Yesso scallop tissues and will benefit future research on carotenoid accumulation and function in scallops.

Identification and characterization of immune-related lncRNAs and lncRNA-miRNA-mRNA networks of Paralichthys olivaceus involved in Vibrio anguillarum infection.

BACKGROUND: Long non-coding RNAs (lncRNAs) structurally resemble mRNAs and exert crucial effects on host immune defense against pathogen infection. Japanese flounder (Paralichthys olivaceus) is an economically important marine fish susceptible to Vibrio anguillarum infection. To date, study on lncRNAs in flounder is scarce. RESULTS: Here, we reported the first systematic identification and characterization of flounder lncRNAs induced by V. anguillarum infection at different time points. A total of 2,368 lncRNAs were identified, 414 of which were differentially expressed lncRNAs (DElncRNAs) that responded significantly to V. anguillarum infection. For these DElncRNAs, 3,990 target genes (named DETGs) and 42 target miRNAs (named DETmiRs) were identified based on integrated analyses of lncRNA-mRNA and lncRNA-miRNA expressions, respectively. The DETGs were enriched in a cohort of functional pathways associated with immunity. In addition to modulating mRNAs, 36 DElncRNAs were also found to act as competitive endogenous RNAs (ceRNAs) that regulate 37 DETGs through 16 DETmiRs. The DETmiRs, DElncRNAs, and DETGs formed ceRNA regulatory networks consisting of 114 interacting DElncRNAs-DETmiRs-DETGs trinities spanning 10 immune pathways. CONCLUSIONS: This study provides a comprehensive picture of lncRNAs involved in V. anguillarum infection. The identified lncRNAs and ceRNA networks add new insights into the anti-bacterial immunity of flounder.

Megalocytivirus Induces Complicated Fish Immune Response at Multiple RNA Levels Involving mRNA, miRNA, and circRNA.

Megalocytivirus is an important viral pathogen to many farmed fishes, including Japanese flounder (Paralichthys olivaceus). In this study, we examined megalocytivirus-induced RNA responses in the spleen of flounder by high-throughput sequencing and integrative analysis of various RNA-seq data. A total of 1327 microRNAs (miRNAs), including 368 novel miRNAs, were identified, among which, 171 (named DEmiRs) exhibited significantly differential expressions during viral infection in a time-dependent manner. For these DEmiRs, 805 differentially expressed target mRNAs (DETmRs) were predicted, whose expressions not only significantly changed after megalocytivirus infection but were also negatively correlated with their paired DEmiRs. Integrative analysis of immune-related DETmRs and their target DEmiRs identified 12 hub DEmiRs, which, together with their corresponding DETmRs, formed an interaction network containing 84 pairs of DEmiR and DETmR. In addition to DETmRs, 19 DEmiRs were also found to regulate six key immune genes (mRNAs) differentially expressed during megalocytivirus infection, and together they formed a network consisting of 21 interactive miRNA-messenger RNA (mRNA) pairs. Further analysis identified 9434 circular RNAs (circRNAs), 169 of which (named DEcircRs) showed time-specific and significantly altered expressions during megalocytivirus infection. Integrated analysis of the DETmR-DEmiR and DEcircR-DEmiR interactions led to the identification of a group of competing endogenous RNAs (ceRNAs) constituted by interacting triplets of circRNA, miRNA, and mRNA involved in antiviral immunity. Together these results indicate that complicated regulatory networks of different types of non-coding RNAs and coding RNAs are involved in megalocytivirus infection.

Systematic Identification and Analysis of Circular RNAs of Japanese Flounder (Paralichthys olivaceus) in Response to Vibrio anguillarum Infection.

Circular RNA (circRNA) is a new class of non-coding RNA that is structured into a closed loop without polyadenylation. Recent studies showed that circRNAs are involved in the host immune response to pathogen infection. Japanese flounder (Paralichthys olivaceus), an important economical marine fish cultured in north Asia, is affected by Vibrio anguillarum, a pathogenic bacterium that can infect a large number of fish. In this study, we systematically explored the circRNAs in the spleen of V. anguillarum-infected flounder at different infection time points. A total of 6581 circRNAs were identified, 148 of which showed differential expression patterns after V. anguillarum infection and were named DEcirs. Most of the DEcirs were strongly time-specific. The parental genes of the DEcirs were identified and functionally classified into diverse pathways, including immune-related pathways. Among the immune-related DEcirs, seven were predicted to sponge 18 targeted miRNAs that were differentially expressed during V. anguillarum infection (named DETmiRs). Further analysis showed that the DEcirs and their corresponding DETmiRs intertwined into complicated immune related networks. These results indicate that in flounder, circRNAs are regulated by V. anguillarum and form interactive networks with mRNAs and miRNAs that likely play important roles in the immune defense against pathogen infection.