Identification and characterization of circular RNAs in the skin of rainbow trout (Oncorhynchus mykiss) infected with infectious hematopoietic necrosis virus.

Rainbow trout (Oncorhynchus mykiss) is an economically significant freshwater-farmed fish worldwide, and the frequent outbreaks of infectious hematopoietic necrosis (IHN) in recent years have gravely compromised the healthy growth of the rainbow trout aquaculture industry. Fish skin is an essential immune barrier against the invasion of external pathogens, but it is poorly known about the role of circRNAs in rainbow trout skin. Therefore, we examined the expression profiles of circRNAs in rainbow trout skin following IHNV infection using RNA-seq. A total of 6607 circRNAs were identified, of which 34 circRNAs were differentially expressed (DE) and these DE circRNA source genes were related to immune-related pathways such as Toll-like receptor signaling pathway, NOD-like receptor signaling pathway, Cytokine-cytokine receptor interaction, ubiquitin mediated proteolysis, and ferroptosis. We used qRT-PCR, Sanger sequencing, and subcellular localization to validate the chosen DE circRNAs, confirming their localization and expression patterns in rainbow trout skin. Further, 12 DE circRNAs were selected to construct the circRNA-miRNA-mRNA regulatory network, finding one miRNA could connect one or more circRNAs and mRNAs, and some miRNAs were reported to be associated with antiviral immunity. The functional prediction findings revealed that novel_circ_002779 and novel_circ_004118 may act as sponges for miR-205-z and miR-155-y to regulate the expression of target genes TLR8 and PIK3R1, respectively, and participated in the antiviral immune responses in rainbow trout. These results shed light on the immunological mechanism of circRNAs in rainbow trout skin and offer fundamental information for further research on the innate immune system and breeding rainbow trout resistant to disease.

Dietary supplementation of Chinese herbal medicines enhances the immune response and resistance of rainbow trout (Oncorhynchus mykiss) to infectious hematopoietic necrosis virus.

Rainbow trout is a widely farmed economical cold-water fish worldwide, but the prevalence of infectious hematopoietic necrosis virus (IHNV) presents a severe risk to the aquaculture industry, resulting in high mortality and huge economic losses. In this study, the impacts of different concentrations (0, 10, 20, and 30 g/kg) of Chinese herbal medicine mixture (CHMM) on the immune response and resistance of rainbow trout to IHNV infection were evaluated. The results show that CHMM noticeably increased (P < 0.05) T-SOD, CAT, AST, ALT, ACP, and AKP activities and decreased MDA content. NF-κB, TNF-α, IFN-ß, IL-1ß, JAK1, HSP70, and HSP90 expressions were significantly upregulated (P < 0.05) in all CHMMs, while SOCS2 expression was downregulated (P < 0.05). Following infection with IHNV, feeding rainbow trout with varying amounts of CHMM resulted in noticeably increased (P < 0.05) T-SOD, ACP, and AKP activities and significantly decreased (P < 0.05) MDA content and AST and ALT activities. TNF-α, IFN-ß, IL-1ß, HSP70, and HSP90 expressions were significantly upregulated (P < 0.05) in all CHMMs, while the expressions of JAK1 and SOCS2 were downregulated. The expression level of the IHNV G protein gene at a dosage of 20 g/kg was notably lower than that of the other CHMM feeding groups. This study provides a solid scientific basis for promoting CHMM as an immunostimulant for boosting antiviral immunity in rainbow trout.

Integrative mRNA-miRNA interaction analysis associated with the immune response in the head kidney of rainbow trout (Oncorhynchus mykiss) after infectious hematopoietic necrosis virus infection.

Rainbow trout (Oncorhynchus mykiss) is an important cold-water fish widely cultivated in China. The frequent occurrence of viral diseases caused by infectious hematopoietic necrosis virus (IHNV) seriously restricted the healthy development of the rainbow trout farming industry. However, the immune defense mechanism induced by IHNV in rainbow trout has not been fully elucidated. In the present study, we detected mRNA and miRNA expression profiles in rainbow trout head kidney after IHNV infection using RNA-seq and identified key immune-related genes and miRNAs. The results showed that a total of 7486 genes and 277 miRNAs were differentially expressed, and numerous differentially expressed genes (DEGs) enriched in the immune-related pathways such as Toll-like receptor signaling pathway, RIG-I-like receptor signaling pathway, NOD-like receptor signaling pathway, cytokine-cytokine receptor interaction, and JAK-STAT signaling pathway were significantly up-regulated, including LGP2, MDA5, TRIM25, IRF3, IRF7, TLR3, TLR7, TLR8, MYD88, and IFN1. Integration analysis identified six miRNAs (miR-141-y, miR-200-y, miR-144-y, miR-2188-y, miR-725-y, and miR-203-y) that target at least six key immune-related genes (TRIM25, LGP2, TLR3, TLR7, IRF3, and IRF7). Further, we verified selected immune-related mRNAs and miRNAs through qRT-PCR and confirmed the reliability of the RNA-seq results. These findings improve our understanding of the immune mechanism of rainbow trout infected with IHNV and provide basic data for future breeding for disease resistance in rainbow trout.

Competing endogenous RNA (ceRNA) in a non-model animal: Non-coding RNAs respond to heat stress in rainbow trout (Oncorhynchus mykiss) through ceRNA-regulated mechanisms.

BACKGROUND: Rainbow trout (Oncorhynchus mykiss) is a typical cold-water fish. With global warming and extreme heat, high summer temperatures are the biggest threat to rainbow trout farming. Rainbow trout initiate stress defense mechanisms in response to thermal stimuli, and competing endogenous RNA (ceRNA) regulation of target genes (mRNAs) mediated by non-coding RNAs (microRNAs [miRNAs], long non-coding RNAs) may be the main strategy for responding to thermal stimuli and enhancing adaptation. RESULTS: We screened the LOC110485411-novel-m0007-5p-hsp90ab1 ceRNA relationship pairs for affect heat stress in rainbow trout and validated their targeting relationships and functions based on preliminary high-throughput sequencing analysis results. The transfection of exogenous novel-m0007-5p mimics and inhibitors into primary rainbow trout hepatocytes effectively bound and inhibited the target genes hsp90ab1 and LOC110485411 without significant effects on hepatocyte viability, proliferation, and apoptosis. The inhibitory effect of novel-m0007-5p overexpression on hsp90ab1 and LOC110485411 under heat stress was time-efficient. Similarly, small interfering RNAs (siRNAs) affected hsp90ab1 mRNA expression by silencing LOC110485411 expression time-efficiently. CONCLUSIONS: In conclusion, we found that in rainbow trout, LOC110485411 and hsp90ab1 can bind competitively to novel-m0007-5p via 'sponge adsorption' and that interference with LOC110485411 affects hsp90ab1 expression. These results provide potential for anti-stress drug screening in rainbow trout.

Comprehensive analysis of immune parameters, mRNA and miRNA profiles, and immune genes expression in the gill of rainbow trout infected with infectious hematopoietic necrosis virus (IHNV).

Rainbow trout (Oncorhynchus mykiss) is a species of cold-water fish with important economic values, widely cultivated worldwide. However, the outbreak of infectious hematopoietic necrosis virus (IHNV) caused the large-scale death of rainbow trout and seriously restricted the development of the trout farming industry. In this study, the changes of immune parameters in different periods (6-, 12-, 24-, 48-, 72-, 96-, 120-, and 144 h post-infection (hpi)), transcriptome profiles of 48 hpi (T48G) compared to control (C48G), and key immune-related genes expression patterns were measured in rainbow trout gill following IHNV challenge through biochemical methods, RNA sequencing (RNA-seq), and quantitative real-time polymerase chain reaction (qRT-PCR). The results showed that alkaline phosphatase (AKP), acid phosphatase (ACP), total superoxide dismutase (T-SOD), alanine aminotransferase (ALT), and aspartate aminotransferase (AST) activities, as well as lysozyme (LZM) and malonaldehyde (MDA) content decreased and then increased during infection, and remained at a high level after 48 hpi (P < 0.05), whereas catalase (CAT) activity showed a significant peak at 48 hpi (P < 0.05). The mRNA and miRNA analysis identified 4343 differentially expressed genes (DEGs) and 11 differentially expressed miRNAs (DEMs), and numerous immune-related DEGs involved in the Toll-like receptor signaling pathway, apoptosis, DNA replication, p53 signaling, RIG-I-like receptor signaling pathway, and NOD-like receptor signaling pathway and expression were significantly up-regulated in T48Gm group, including tlr3, tlr7, tlr8, traf3, ifih1, trim25, dhx58, ddh58, hsp90a.1, nlrc3, nlrc5, socs3, irf3, irf7, casp7, mx1, and vig2. The integrated analysis identified several important miRNAs (ola-miR-27d-3p_R+5, gmo-miR-124-3-5p, ssa-miR-301a-5p_L+2, and ssa-miR-146d-3p) that targeted key immune-related DEGs. Expression analysis showed that tlr3, tlr7, traf3, ifih1, dhx58, hap90a.1, irf3, irf7, and mx1 genes increased and then decreased during infection, and peaked at 72 hpi (P < 0.05). However, trim25 expression peaked at 96 hpi (P < 0.05). This study contributes to understanding immune response of rainbow trout against IHNV infection, and provides new insights into the immune regulation mechanisms and disease resistance breeding studies.

Gene ssa-miR-301a-3p improves rainbow trout (Oncorhynchus mykiss) resistance to heat stress by targeting hsp90b2.

Rainbow trout (Oncorhynchus mykiss) is a cold-water fish that is commonly harmed by high temperatures. MicroRNAs (miRNAs) are being investigated intensively because they act as essential metabolic regulators and have a role in the heat stress response. Although there have been numerous studies on rainbow trout heat stress, research on miRNA implicated in rainbow trout heat stress is quite restricted. Rainbow trout were sampled at 18 and 24 °C, respectively, to examine the mechanism of miRNA under heat stress, and we identified a heat stress-induced miRNA, ssa-miR-301a-3p, for further investigation based on our bioinformatics analysis of rainbow trout small RNA sequencing data. Bioinformatics research suggested that hsp90b2 is a probable target gene for ssa-miR-301a-3p. QRT-PCR was used to confirm the expression levels of ssa-miR-301a-3p and hsp90b2. Meanwhile, the dual-luciferase reporter assay was employed to validate the ssa-miR-301a-3p-hsp90b2 targeted connection. The results indicated that at 24 °C, the relative expression of ssa-miR-301a-3p was considerably lower than at 18 °C. On the other hand, hsp90b2 expression, followed the opposite pattern. The binding of ssa-miR-301a-3p to the 3'-UTR of hsp90b2 resulted in a substantial decrease in luciferase activity. The findings showed that ssa-miR-301a-3p was implicated in heat stress, and our findings provide fresh insights into the processes of miRNA in response to heat stress in rainbow trout.

Integrative mRNA-miRNA interaction analysis reveals the molecular mechanism of skin color variation between wild-type and yellow mutant rainbow trout (Oncorhynchus mykiss).

Rainbow trout (Oncorhynchus mykiss) is an important economic fish in China. Skin color affects the economic value of trout. However, the molecular mechanism of the skin color variation between wild-type (WR) and yellow mutant rainbow trout (YR) is unclear. We sequenced mRNAs and miRNAs of dorsal skin to identify key color variation-associated mRNAs and miRNAs between WR and YR. Overall, 2060 out of 3625 differentially expressed genes were upregulated in YR, and 196 out of 275 differentially expressed miRNAs were downregulated in WR. We identified three key YR-upregulated genes related to the formation of xanthophores (GCH1, SLC2A11, and SOX10). Interestingly, several genes related to melanogenesis (TYR, TYRP1, TYRP2, MC1R, MITF, PMEL, SLC45A2, and OCA2) were downregulated in WR. Integrated analysis identified five miRNAs that target at least two skin color-related genes (miR-495-y, miR-543-y, miR-665-z, miR-433-y, and miR-382-x). Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses of target genes identified noncoding RNA metabolic process as the most significantly enriched GO term, and several metabolic pathways associated with skin color were enriched significantly, such as tyrosine metabolism, histidine metabolism, and vitamin B6 metabolism. Quantitative real-time PCR of selected mRNAs and miRNAs validated the reliability of the integrated analysis. This study provides in-depth insights into the molecular mechanism of skin color variation between WR and YR, which will accelerate the genetic selection and breeding of rainbow trout with consumer-favored traits.

miR-434 and miR-242 have a potential role in heat stress response in rainbow trout (Oncorhynchus mykiss).

MicroRNAs (miRNAs) are being extensively studied as they function as key metabolic regulators which play a role in the heat stress response. However, the role of miRNAs in heat stress remains uncertain and many new miRNAs have not yet been discovered. In a previous study, we performed high-throughput sequencing of differentially expressed miRNAs identified on exposing rainbow trout (Oncorhynchus mykiss) to heat stress (18 vs. 24°C), which led to the identification of two novel miRNAs, temporarily named novel miR-434 and -242. The differential expression level of these miRNAs was extremely significant (P < 0.01); we analysed target gene mRNA transcripts by bioinformatics software (miRanda). We found novel miR-434 and -242 were predicted to regulate the transcripts of heat shock 70-kDa protein 4-like (HSPA4L) and calreticulin (CRT), respectively, by bioinformatics software. Here our core objective was to validate if HSPA4L and CRT are indeed the target genes of novel miR-434 and -242, respectively, and for this purpose we used the dual-luciferase reporter assay system. Target gene sequences were synthesized and cloned into a dual-luciferase vector. To better understand the function of the target genes, we combined the previous sequencing results of Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis. We found that novel miR-434 regulated HSPA4L expression by binding to a putative binding site in the 3'-UTR of HSPA4L, and luciferase activity inhibition was observed. In contrast, novel miR-242 was not involved in regulating CRT expression. To conclude, we believe our results should serve as a foundation for future studies aiming to comprehensively understand the mechanisms used by rainbow trout to cope with heat stress.

Integrated analysis of the responses of a circRNA-miRNA-mRNA ceRNA network to heat stress in rainbow trout (Oncorhynchus mykiss) liver.

BACKGROUND: With the intensification of global warming, rainbow trout (Oncorhynchus mykiss) suffer from varying degrees of thermal stimulation, leads to mass mortality, which severely restrict the development of aquaculture. Understanding the molecular regulatory mechanisms of rainbow trout under heat stress is useful to develop approaches to relieve symptoms. RESULTS: Changes in nonspecific immune parameters revealed that a strong stress response was caused in rainbow trout at 24 °C, so we performed multiple transcriptomic analyses of rainbow trout liver under heat stress (HS, 24 °C) and control conditions (CG, 18 °C). A total of 324 DEcircRNAs, 105 DEmiRNAs, and 1885 DEmRNAs were identified. A ceRNA regulatory network was constructed and a total of 301 circRNA-miRNA and 51 miRNA-mRNA negative correlation pairs were screened, and three regulatory correlation pairs were predicted: novel_circ_003889 - novel-m0674-3p - hsp90ab1, novel_circ_002325 - miR-18-y - HSPA13 and novel_circ_002446 - novel-m0556-3p - hsp70. Some target genes involved in metabolic processes, biological regulation or response to stimulus were highly induced at high temperatures. Several important pathways involved in heat stress were characterized, such as protein processing in the ER, the estrogen signaling pathway, and the HIF-1 signaling pathway. CONCLUSIONS: These results extend our understanding of the molecular mechanisms of the heat stress response and provide novel insight for the development of strategies that relieve heat stress.

Proteome analysis of rainbow trout (Oncorhynchus mykiss) liver responses to chronic heat stress using DIA/SWATH.

Aquaculture of rainbow trout (Oncorhynchus mykiss) is severely hampered by high temperatures in summer, and understanding the regulatory mechanisms controlling responses to chronic heat stress may assist the development of measures to relieve heat stress. In the present study, biochemical parameters revealed a strong stress response in rainbow trout at 24 °C, including activation of stress defence and immune systems. Liver proteome analysis under heat stress (24 °C) and control (18 °C) conditions using DIA/SWATH identified precursors (90,827), peptides (67,028), proteins (6770) and protein groups (5124), among which 460 differentially abundant proteins (DAPs; q-value < 0.05, fold change >1.5), 201 and 259 were up- and down-regulated, respectively. Many were related to heat shock proteins (HSPs), metabolism and immunity. Gene Ontology (GO) analysis showed that some DAPs induced at high temperature were involved in regulating cell homeostasis, metabolism, adaptive stress and stimulation. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis identified metabolic pathways, protein processing in endoplasmic reticulum, PPAR signalling, and complement and coagulation cascades. Protein-protein interaction (PPI) network analysis indicated that HSP90b1 and C3 may cooperative to affect cell membrane integrity under heat stress. Our findings assist the development of strategies to relieve heat stress in rainbow trout.

Identification and characterization of long noncoding RNAs provide insight into the regulation of gene expression in response to heat stress in rainbow trout (Oncorhynchus mykiss).

Rainbow trout are typical cold-water fish species. However, with the intensification of global warming, high temperatures have severely restricted the development of aquaculture during the summer. Understanding the molecular regulatory mechanisms of rainbow trout responses to heat stress will be beneficial for alleviating heat stress-related damage. In this study, we performed RNA-seq of liver tissues from rainbow trout under heat stress (24 °C) and control conditions (18 °C) to identify lncRNAs and target genes by strand-specific library. Changes in nonspecific immune parameters revealed that a strong stress response occurred in rainbow trout at 24 °C. More than 658 million filtered reads and 5916 lncRNAs were identified from six libraries. A total of 927 novel lncRNAs were identified, and 428 differentially expressed lncRNAs were screened with stringent thresholds. The RNA-seq results were verified by RT-qPCR. In addition, a regulatory network of lncRNA-mRNA functional interactions was constructed, and the potential antisense, cis and trans targets of lncRNAs were predicted. GO and KEGG enrichment analyses showed that many target genes involved in maintenance of homeostasis or adaptation to stress and stimuli were highly induced under heat stress. Several regulatory pathways were also found to be involved in heat stress, including the thyroid hormone signaling pathway, the PI3K-Akt signaling pathway, and the estrogen signaling pathway, among others. These results broaden our understanding of lncRNAs associated with heat stress and provide new insights into the lncRNA mediated regulation of the rainbow trout heat stress response.

Genome-Wide Identification of hsp90 Gene in Rainbow Trout (Oncorhynchus mykiss) and Their Regulated Expression in Response to Heat Stress.

In this study, we analyzed the gene structure, chemical characterizations, chromosome locations, evolutionary relationship, and expression profile of hsp90 genes with online database. In addition, the expression levels of hsp90s were also investigated under heat stress by quantitative real-time (qRT)-PCR. A total of eight hsp90 genes were identified from the rainbow trout genome. They were all distributed on chromosomes 2, 4, 8, and 13. The molecular weight ranged from 78.93 to 91.39 kDa, and the isoelectric point ranged from 4.84 to 4.96. The eight hsp90 genes were clustered into six subfamilies (A, B, C, D, E, and F). Genetic structure and conserved domain analysis revealed that all eight hsp90 genes had only one exon, and motif 1-motif 10 was shared by most genes. According to RNA-seq analysis of rainbow trout liver and head kidney, a total of seven out of eight genes were significantly upregulated under heat stress, and qRT-PCR was carried out on these seven genes; the expression levels of these genes were significantly upregulated under heat stress. The significantly regulated expressions of hsp90 genes under heat stress indicated that hsp90 genes are involved in heat stress response in rainbow trout. This study provides a theoretical basis for further study on the role of hsp90 in the heat stress tolerance of rainbow trout.

High-throughput sequencing reveals microRNAs in response to heat stress in the head kidney of rainbow trout (Oncorhynchus mykiss).

Recently, the research of animal microRNAs (miRNAs) has attracted wide attention for its regulatory effect in the development process and the response to abiotic stresses. Rainbow trout is a commercially and cold water fish species, and usually encounters heat stress, which affects its growth and leads to a huge economic loss. But there were few investigations about the roles of miRNAs in heat stress in rainbow trout. In this study, miRNAs of rainbow trout which were involved in heat stress were identified by high-throughput sequencing of six small RNA libraries from head kidney tissues under control (18 °C) and heat-treated (24 °C) conditions. A total of 392 conserved miRNAs and 989 novel miRNAs were identified, of which 78 miRNAs were expressed in different response to heat stress. Ten of these miRNAs were further validated by quantitative real-time PCR. In addition to, including 393 negative correlation miRNA-target gene pairs, several important regulatory pathways were involved in heat stress of the potential target genes, including protein processing in endoplasmic reticulum, NOD-like receptor signaling pathway, and phagosome. Our data significantly advance understanding of heat stress regulatory mechanism of miRNA in the head kidney of rainbow trout, which provide a useful resource for the cultivation of rainbow trout.

Evaluation of reference genes for quantitative real-time PCR analysis of messenger RNAs and microRNAs in rainbow trout Oncorhynchus mykiss under heat stress.

We assessed the expression stability of several messenger (m)RNAs and micro (mi)RNAs from liver and head kidney of rainbow trout Oncorhynchus mykiss using high-throughput RNA sequencing (RNA-seq) and miRNA-seq data. Additionally, four commonly used reference genes and one small non-coding RNA (u6) were also selected to identify ideal reference mRNAs and miRNAs for quantitative real-time (qrt)-PCR analysis of heat stress responses. GeNorm, NormFinder, BestKeeper and comparative ΔCt were employed for analysis of qrt-PCR data to systematically assess the expression stability of candidate mRNAs and miRNAs and stability was ranked using geometric means. ß-actin and ef1-α were the most stably expressed reference mRNAs in liver and head kidney, respectively and ssa-mir-26a-5p and ssa-mir-462b-5p were the most stably expressed miRNAs in these tissues. This is the first identification of appropriate reference mRNAs and miRNAs for qrt-PCR analysis of O. mykiss under heat stress.

Label-free quantification of protein expression in the rainbow trout (Oncorhynchus mykiss) in response to short-term exposure to heat stress.

Rainbow trout (Oncorhynchus mykiss) are a cold-water salmonid species that is highly susceptible to heat stress. Summer temperature stress is a common issue in trout aquaculture. To better understand the molecular mechanisms of the heat-stress response in the trout, we used label-free quantitative proteome techniques to identify differentially expressed proteins in the livers of rainbow trout exposed to heat stress. We identified 3362 proteins and 152 differentially expressed proteins (p < 0.05; fold-change >2). Of these, 37 were uniquely expressed in the heat-stress group and 35 were uniquely expressed in the control group. In addition, 42 proteins were significantly upregulated (fold-change >2) and 38 proteins were significantly downregulated (fold-change >2). GO (Gene Ontology) analysis indicated that these differentially expressed proteins were primarily expressed in the nucleus, extracellular matrix, and cytoplasm, and were associated with a variety of functions, including protein binding/bridging and enzyme facilitation. KEGG (Kyoto Encyclopedia of Genes and Genomes) analysis of the differentially expressed proteins showed that, during high temperature stress, many biological processes were extensively altered, particularly the estrogen signaling pathway, the complement and coagulation cascades, and the platelet activation pathway. Our study focused on the identification of a systematic approach for the characterization of regulatory networks. Our results provide a framework for further studies of the heat-stress response in fish.

Identification and characterization of microRNAs in the liver of rainbow trout in response to heat stress by high-throughput sequencing.

MicroRNAs (miRNAs) play key roles in the stress response in animals. The rainbow trout is a widely cultivated cold-water fish species that usually encounters heat stress, which affects its growth and may cause diseases. However, there is limited information on the miRNAs involved in heat stress in this species. To identify the heat-responsive miRNAs at the genome level in rainbow trout, we conducted high-throughput sequencing of 6 small RNA libraries from the liver tissues of rainbow trout specimens under control and heat-treated conditions. A total of 363 conserved miRNAs and 638 novel miRNAs were identified, among which 39 (21 conserved and 18 novel) miRNAs were differentially expressed in response to heat stress. Eight important miRNAs were further validated using quantitative real-time PCR. GO enrichment and KEGG pathway analyses of the potential target genes of the DE miRNAs, revealed DNA replication and protein folding and unfolding were affected by heat stress. Several key pathways involved in the heat stress response were characterized, such as protein processing in the endoplasmic reticulum, NOD-like receptor signaling pathway, and progesterone-mediated oocyte maturation. In addition, 20 important functional interaction pairs among the 65 negatively correlated miRNA-mRNA pairs were identified using integrated miRNA-mRNA analysis and miRNA prediction algorithms, which including potential important miRNAs and target genes such as miR-301a, miR-301d, let-7c, novel_1181 and HSP90BA, HSP90BB, HSP40, PDIA3. This study showed that the identified miRNAs probably play important roles in the heat stress response and lay a foundation for further studies on the thermal adaptation mechanisms in rainbow trout.

Transcriptomic responses to heat stress in rainbow trout Oncorhynchus mykiss head kidney.

Rainbow trout (Oncorhynchus mykiss) are widely cultured throughout the word for commercial aquaculture. However, as a cold-water species, rainbow trout are highly susceptible to heat stress, which may cause pathological signs or diseases by alleviating the immune roles and then lead to mass mortality. Understanding the molecular mechanisms that occur in the rainbow trout in response to heat stress will be useful to decrease heat stress-related morbidity and mortality in trout aquaculture. In the present study, we conducted transcriptome analysis of head kidney tissue in rainbow trout under heat-stress (24 °C) and control (18 °C) conditions, to identify heat stress-induced genes and pathways. More than 281 million clean reads were generated from six head kidney libraries. Using an adjusted P-value of P < 0.05 as the threshold, a total of 443 differentially expressed genes (DEGs) were identified, including members of the HSP90, HSP70, HSP60, and HSP40 family and several cofactors or cochaperones. The RNA-seq results were confirmed by RT-qPCR. Gene ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analysis of DEGs were performed. Many genes involved in maintaining homeostasis or adapting to stress and stimuli were highly induced in response to high temperature. The most significantly enriched pathway was "Protein processing in endoplasmic reticulum (ER)", a quality control system that ensures correct protein folding or degradation of misfolded polypeptides by ER-associated degradation. Other signaling pathways involved in regulation of immune system and post-transcriptional regulation of spliceosome were also critical for thermal adaptation. These findings improve our understanding of the molecular mechanisms of heat stress responses and are useful to develop strategies for the improvement of rainbow trout survival rate during summer high-temperature period.

Ocular toxicity of reduced graphene oxide or graphene oxide exposure in mouse eyes.

With the wide application and mass production of nanoparticle products, environmental nanopollutants will become increasingly common. The eye is an important organ responsible for vision in most living organisms, and it is directly exposed to the atmosphere. Direct contact between the eye and nanoparticles in the environment can potentially lead to ocular damage. However, publications focusing on the eye-damaging potential of nanoparticles are scarce. Therefore, to evaluate the impact of nanoparticles on the eyes, we investigated the ocular toxicity of reduced graphene oxide (RGO) and graphene oxide (GO) using morphological and molecular biological methods in vivo and in vitro in the present work. The findings show that short-term repeated GO exposure can cause obvious intraocular inflammation, an incrassated corneal stromal layer, cell apoptosis in the cornea, iris neovascularization and significant cytotoxicity of rat corneal epithelial cells (rCECs), while RGO causes no significant ocular toxicity in mice.

Long non-coding RNA SNHG1 predicts a poor prognosis and promotes colon cancer tumorigenesis.

Colon cancer is the main cause of cancer mortality worldwide. Its poor prognosis is mainly ascribed to high recurrence rates. Identifying novel prognostic biomarkers and therapeutic key points for management is crucial and important. Long non-coding RNAs (lncRNAs) are a class of RNAs, which have various roles in carcinogenicity and molecular mechanisms. The lncRNA small nucleolar RNA host gene 1 (SNHG1) contributes to the promotion of tumor development, however, the connections between SNHG1 and colon cancer are still unclear. The aim of the present study was to investigate the clinical significance, the biological functions, and the potential mechanism of SNHG1 in colon cancer. In the present study, we referred to the Oncomine database and used RT-qPCR to determine that SNHG1 expression was significantly higher both in colon cancer tissues and cancerous cell lines than in normal samples. Cell functional experiments were performed after knockdown of SNHG1, including Cell Counting Kit-8 assay, colony formation assay, Transwell® assay, and flow cytometric analyses of cell apoptosis, which suggested that SNHG1 stimulated colon cancer cell proliferation, promoted cell invasion and migration, and inhibited apoptosis. Immunohistochemical staining and western blotting experiments revealed that in colon cancer cells with SNHG1 knockdown, ß-catenin, c-Myc and cyclin D1 protein levels were decreased, while E-cadherin was increased, which suggested that SNHG1 promoted colon cancer cell proliferation, migration and invasion through the Wnt/ß-catenin signaling pathway. Our results indicated that SNHG1 and its interrelated components may be future therapeutic targets of carcinoma of the colon.