Expression pattern and cellular localization of pepsinogen in early development and induced by different diets in the spotted knifejaw (Oplegnathus punctatus).

To solve the high mortality rate of early-stage larval feed conversion during aquaculture in Oplegnathus punctatus, the investigation of the structural and functional characteristics of the gastric tissue was conducted. Histological results showed that the gastric gland rudiment appeared at 17 dph. The basic structure of the stomach was fully developed between 26 and 35 dph. Two pepsinogen genes, named OpPGA1 and OpPGA2, were identified in the spotted knifejaw genome. qPCR results of developmental period showed that the two genes were low in expression during early development (5 and 15 dph). At 20 dph, the two genes started to show trace expression, and at 30 dph the mRNA expression levels of OpPGA1 and OpPGA2 reached the highest levels. Results of pepsin activity detection during the development period showed lower activity was detected 22 dph, followed by a peak at 30 dph. Under different feeding inductions, OpPGA1 showed the highest expression in the basic diet group and hard-shell group, while the expression level in the phytophagous group remained consistently low. The mRNA expression level of OpPGA2 in the phytophagous group was significantly higher than in other groups. Enzyme activity determination under different feeding inductions showed slightly higher enzyme activity in the basic diet group and crustacean group. The results of in situ hybridization showed that the mRNA of both OpPGA1 and OpPGA2 genes was both expressed in gastric gland cells. These information can contribute to the development of practical feeding methods in terms of digestive physiology for the development of larvae.

Drynaria Naringin alleviated mechanical stress deficiency-caused bone loss deterioration via Rspo1/Lgr4-mediated Wnt/ß-catenin signalling pathway.

Osteoporosis is a metabolic condition distinguished by the degradation of bone microstructure and mechanical characteristics. Traditional Chinese medicine (TCM) has been employed in China for the treatment of various illnesses. Naringin, an ingredient found in Drynariae TCM, is known to have a significant impact on bone metabolism. For this research, we studied the precise potential effect of Drynaria Naringin on protecting against bone loss caused by stress deficiency. In this study, a tail-suspension (TS) test was performed to establish a mouse model with hind leg bone loss. Some mice received subcutaneous injections of Drynaria Naringin for 30 d. Trabecular bone microarchitecture was evaluated using micro-computed tomography analysis and bone histological analysis. Bone formation and resorption markers were quantified in blood samples from mice or in the supernatant of MC3T3-E1 cells by ELISA analysis, Western blotting, and PCR. Immunofluorescence was utilized to visualize the location of ß-catenin. Additionally, siRNA was employed to knockdown-specific genes in the cells. Our findings highlight the efficacy of Drynaria Naringin in protecting against the deterioration of bone loss and promoting bone formation and Rspo1 expression in a mouse model following the TS test. Specifically, in vitro experiments also indicated that Drynaria Naringin may promote osteogenesis through the Wnt/ß-catenin signalling pathway. Moreover, our results suggest that Drynaria Naringin upregulates the expression of Rspo1/Lgr4, leading to the promotion of osteogenesis via the Wnt/ß-catenin signalling pathway. Therefore, Drynaria Naringin holds potential as a therapeutic medication for osteoporosis. Drynaria Naringin alleviates bone loss deterioration caused by mechanical stress deficiency through the Rspo1/Lgr4-mediated Wnt/ß-catenin signalling pathway.

Tea polyphenols, astaxanthin, and melittin can significantly enhance the immune response of juvenile spotted knifejaw (Oplegnathus punctatus).

The frequent occurrence of diseases seriously hampers the sustainable development of the spotted knifejaw (Oplegnathus punctatus) breeding industry. Our previous genome-wide scan and cross-species comparative genomic analysis revealed that the immune gene family (Toll-like receptors, TLR) members of O. punctatus underwent a significant contraction event (tlr1, tlr2, tlr14, tlr5, and tlr23). To address immune genetic contraction may result in reduced immunity, we investigated whether adding different doses (0, 200, 400, 600, and 800 mg/kg) of immune enhancers (tea polyphenols, astaxanthin, and melittin) to the bait after 30 days of continuous feeding could stimulate the immune response of O. punctatus. We found that the expression of tlr1, tlr14, tlr23 genes in immune organs (spleen and head kidney) was stimulated when tea polyphenols were added at 600 mg/kg. The tlr2 (400 mg/kg), tlr14 (200 mg/kg), tlr5 (200 mg/kg), and tlr23 (200 mg/kg) genes expression of intestine were elevated in the tea polyphenol group. When the addition of astaxanthin is 600 mg/kg, it can effectively stimulate the expression of tlr14 gene in immune organs (liver, spleen and head kidney). In the astaxanthin group, the expression of the genes tlr1 (400 mg/kg), tlr14 (600 mg/kg), tlr5 (400 mg/kg) and tlr23 (400 mg/kg) reached their highest expression in the intestine. Besides, the addition of 400 mg/kg of melittin can effectively induce the expression of tlr genes in the liver, spleen and head kidney, except the tlr5 gene. The tlr-related genes expression in the intestine was not significantly elevated in the melittin group. We hypothesize that the immune enhancers could enhance the immunity of O. punctatus by increasing the expression of tlr genes, and thereby leading to increased resistance to diseases. Meanwhile, our findings further demonstrated that significant increases in weight gain rate (WGR), visceral index (VSI), and feed conversion rate (FCR) were observed at 400 mg/kg, 200 mg/kg and 200 mg/kg of tea polyphenols, astaxanthin and melittin in the diet, respectively. Overall, our study provided valuable insights for future immunity enhancement and viral infection prevention in O. punctatus, as well as offered guidance for the healthy development of the O. punctatus breeding industry.

Portable ultrasonic humidifier exacerbates indoor bioaerosol risks by raising bacterial concentrations and fueling pathogenic genera.

Portable ultrasonic humidifiers are frequently used in heating rooms to ease air dryness. However, it has also posed serious health concerns such as "humidifier fever" because the bioaerosol concentration and community in the humidified space may alter quickly before the occupants could even notice. We compared the microbial proliferation rates in the humidifiers' reservoirs filled with three commonly used water types and investigated the impacts of the ultrasonic humidifiers on the temporal concentration, size distribution, and community variations of indoor bacterial and fungal aerosols during two-week humidification. The concentration of indoor bacterial aerosols increased exponentially, concentrating in the respiratory size ranges (≤1.1 µm), and was proportional to the humidification level, which soon exceeded 1000 CFU/m3 in one week (at RH = 70%), while the fungal concentration always remained low (≤177 CFU/m3 ). The indoor bioaerosol community, significantly associated with the humidifier water, was substantially distorted after humidification and dominated by the pathogenic Pseudomonas spp. (40.50%), Brevundimonas spp. (3.02%), Acinetobacter spp. (0.98%) and Legionella spp. (0.69%). Our results show that ultrasonic humidification contaminates indoor air by raising bacterial concentrations and fueling the pathogenic genera. To minimize the exposure risks, occupants should avoid long-term and excessive humidification (RH ≥ 70%) and clean the ultrasonic humidifier weekly.

MiR-495 regulates cell proliferation and apoptosis in H2O2 stimulated rat spinal cord neurons through targeting signal transducer and activator of transcription 3 (STAT3).

BACKGROUND: MicroRNA-495 (miR-495) is a post-translational modulator that performs several functions, and it is involved in several disease states. On the other hand, the physiological functions of miR-495 in H2O2 stimulated mouse spinal cord neuronal dysfunction have not yet been fully understood. METHODS: In this study, we speculated that miR-495 may regulate the expression of STAT3 in the processes of neuronal proliferation and apoptosis following spinal cord injury (SCI). Cell viability was assessed with methyl thiazolyl tetrazolium (MTT) assay. Caspase-3 activity was assayed with ELISA. Cellular apoptotic changes were measured with TUNEL assay. Intracellular ROS production was determined by measuring uptake of dichlorodihydrofluorescein diacetate (DCFH-DA; PCR was used to assay the mRNA expression of STAT3 gene bearing predicted targeting positions for miR-495, while qRT-PCR was used to measure miR-495 mRNA. RESULTS: The results demonstrated that treatment of SCNs with H2O2 led to a significant decrease in cell survival, while it enhanced apoptosis. The H2O2 treatment induced cell membrane dysfunction, and increased ROS levels and DNA damage. Interestingly, the expression of miR-495 was markedly suppressed when SCNs were exposed to H2O2. However, miR-495 overexpression reversed H2O2-induced cytotoxicity and apoptosis in SCNs. Moreover, H2O2 exposure elevated protein and mRNA concentrations of STAT3 in SCNs. Bioinformatics analysis showed likely binding domains of miR-495 in the 3'-untranslated regions of STAT3 in SCNs. MiR-495 loss-of-function and gain-of-function significantly up-regulated and down-regulated both STAT3 mRNA and protein expressions, respectively, in SCNs. CONCLUSIONS: miR-495 overexpression inhibited H2O2-induced SCN dysfunction. This mechanism was mediated through the down-regulation of STAT3 expression.

Identification and analysis of spinal cord injury subtypes using weighted gene co-expression network analysis.

BACKGROUND: Spinal cord injury (SCI) has an immediate and devastating impact on the control over various movements and sensations. However, no effective therapies for SCI currently exist. METHODS: To identify and analyze SCI subtypes, we obtained the expression profile data of the 1,057 genes (889 intersection genes) in GSE45550 using weighted gene co-expression network analysis (WGCNA), and 14 co-expression gene modules were identified. Next, we filtered out the network degree top 10 (degree >80) genes, considered the final key SCI genes. A multifactor regulatory network (105 interaction pairs), consisting of messenger RNAs (mRNAs), long non-coding RNAs (lncRNAs), and transcription factors (TFs) was constructed. This network was involved in the co-expression of key genes. We selected the top 10 regulatory factors (degree >4) as core regulators in the multifactor regulatory network. RESULTS: The results of functional enrichment analysis of the target gene expressing the core regulatory factor [1,059] showed that these target genes were enriched in pathways for human cytomegalovirus infection, chronic myeloid leukemia, and pancreatic cancer. Further, we used the key genes in the co-expression network to categorize the SCI samples in GSE45550. The expression levels of the top 6 genes (CCNB2, CCNB1, CKS2, COL5A1, KIF20A, and RACGAP1) may act as potential marker genes for different SCI subtypes. On the basis of these different subtypes, 8 SCI core gene CDK1-associated drugs were also found to provide potential therapeutic options for SCI. CONCLUSIONS: These results may provide a novel therapeutic strategy for the treatment of SCI.

Analysis of the relationship between electromagnetic radiation characteristics and urban functions in highly populated urban areas.

The electromagnetic environment (EME) in cities is becoming increasingly complex, and the resulting potential health hazards have attracted widespread attention. Large-scale field observations and monitoring of electromagnetic fields were performed in Xiamen Island over the past six years. The results show that the integrated electric field intensity in Xiamen Island ranged from 0.32 V/m to 1.70 V/m, while the integrated magnetic flux density ranged from 0.11 µT to 0.50 µT; where more electric power facilities and electronic equipment are present in the island, the electric and magnetic field strengths are higher; the radiation power of 2nd Generation mobile communication (2G) is higher than that of 3rd Generation mobile communication (3G) and 4th Generation mobile communication (4G), the coverage of the 3G signal was more uniform than the others and the 4G communication signal's coverage is still developing. The relationship between the EME characteristics and urban functions has been analyzed in this study. Results showed that electric field intensity had no correlation with urban functional areas, magnetic flux density had a positive correlation with residential area (q = 0.29); 2G and 4G radiation power are positively related to the educational (Edu) function area (960 MHz q = 0.22, 1.8 GHz q = 0.47, 2.61 GHz q = 0.28); there was a positive relationship between 2G (1.8 GHz) radiation power and residential area (q = 0.2). We concluded that there is a strong link between the Xiamen Island's EME and the distribution of electromagnetic radiation (EMR) sources, the denser and wider distributed EMR sources lead to a more complicated urban EME.

Coupling of oxidative stress responses to tricarboxylic acid cycle and prostaglandin E2 alterations in Caenorhabditis elegans under extremely low-frequency electromagnetic field.

Purpose: With all-pervasive presence of extremely low-frequency electromagnetic field (ELF-EMF) in modern life, ELF-EMF has been regarded as an essential factor which may induce changes in many organisms. The objective of the present study was to investigate the physiological responses of Caenorhabditis elegans (C. elegans) to 50 Hz, 3 mT ELF-EMF exposure. Materials and methods: Worms were exposed to ELF-EMF from the egg stage until reaching the fourth larva (L4) stage. After exposure, expressions of the tricarboxylic acid (TCA) cycle enzymes were examined by qRT-PCR and western blot analysis. Two lipid metabolites were detected by GC-MS. Reactive oxygen species (ROS) level was detected by dichlorofluorescein staining and worm antioxidant system was investigated by enzymatic activity analysis, including detection of the superoxide dismutase and catalase (CAT) activity and the total antioxidant capacity (T-AOC). Results: The TCA cycle enzyme, fumarase was found with decreased expression under ELF-EMF exposure. And arachidonic acid (ArA) and prostaglandin E2(PGE2) showed elevated concentrations, with increased expression of prostaglandin E2 synthase (PGES-2) in ELF-EMF exposed worms. Significant elevation of ROS level was identified accompanied with the significant depression of T-AOC in response to ELF-EMF. Conclusions: Our results suggested that exposure to 50 Hz, 3 mT ELF-EMF in C. elegans can elicit disruptions of the TCA cycle metabolism and PGE2 formation, coupling ELF-EMF-induced oxidative stress responses. Our study probably will attract increasing attentions to the controllable application of ELF-EMF associated with health and disease.

Electromagnetic Radiation Disturbed the Photosynthesis of Microcystis aeruginosa at the Proteomics Level.

Photosynthesis of Microcystis aeruginosa under Electromagnetic Radiation (1.8 GHz, 40 V/m) was studied by using the proteomics. A total of 30 differentially expressed proteins, including 15 up-regulated and 15 down-regulated proteins, were obtained in this study. The differentially expressed proteins were significantly enriched in the photosynthesis pathway, in which the protein expression levels of photosystems II cytochrome b559 α subunit, cytochrome C550, PsbY, and F-type ATP synthase (a, b) decreased. Our results indicated that electromagnetic radiation altered the photosynthesis-related protein expression levels, and aimed at the function of photosynthetic pigments, photosystems II potential activity, photosynthetic electron transport process, and photosynthetic phosphorylation process of M. aeruginosa. Based on the above evidence, that photoreaction system may be deduced as a target of electromagnetic radiation on the photosynthesis in cyanobacteria; the photoreaction system of cyanobacteria is a hypothetical "shared target effector" that responds to light and electromagnetic radiation; moreover, electromagnetic radiation does not act on the functional proteins themselves but their expression processes.

Generation and propagation of yeast prion [URE3] are elevated under electromagnetic field.

In this study, we studied the effect of 2.0 GHz radio frequency electromagnetic field (RF-EMF) and 50 Hz extremely low frequency electromagnetic field (ELF-EMF) exposure on prion generation and propagation using two budding yeast strains, NT64C and SB34, as model organisms. Under exposure to RF-EMF or ELF-EMF, the de novo generation and propagation of yeast prions [URE3] were elevated in both strains. The elevation increased over time, and the effects of ELF-EMF occurred in a dose-dependent manner. The transcription and expression levels of the molecular chaperones Hsp104, Hsp70-Ssa1/2, and Hsp40-Ydj1 were not statistically significantly changed after exposure. Furthermore, the levels of ROS, as well as the activities of superoxide dismutase (SOD) and catalase (CAT), were significantly elevated after short-term, but not long-term exposure. This work demonstrated for the first time that EMF exposure could elevate the de novo generation and propagation of yeast prions and supports the hypothesis that ROS may play a role in the effects of EMF on protein misfolding. The effects of EMF on protein folding and ROS levels may mediate the broad effects of EMF on cell function.

Exposure of ELF-EMF and RF-EMF Increase the Rate of Glucose Transport and TCA Cycle in Budding Yeast.

In this study, we investigated the transcriptional response to 50 Hz extremely low frequency electromagnetic field (ELF-EMF) and 2.0 GHz radio frequency electromagnetic field (RF-EMF) exposure by Illumina sequencing technology using budding yeast as the model organism. The transcription levels of 28 genes were upregulated and those of four genes were downregulated under ELF-EMF exposure, while the transcription levels of 29 genes were upregulated and those of 24 genes were downregulated under RF-EMF exposure. After validation by reverse transcription quantitative polymerase chain reaction (RT-qPCR), a concordant direction of change both in differential gene expression (DGE) and RT-qPCR was demonstrated for nine genes under ELF-EMF exposure and for 10 genes under RF-EMF exposure. The RT-qPCR results revealed that ELF-EMF and RF-EMF exposure can upregulate the expression of genes involved in glucose transportation and the tricarboxylic acid (TCA) cycle, but not the glycolysis pathway. Energy metabolism is closely related with the cell response to environmental stress including EMF exposure. Our findings may throw light on the mechanism underlying the biological effects of EMF.

Coupling Mechanism of Electromagnetic Field and Thermal Stress on Drosophila melanogaster.

Temperature is an important factor in research on the biological effects of extremely low-frequency electromagnetic field (ELF-EMF), but interactions between ELF-EMF and temperature remain unknown. The effects of ELF-EMF (50 Hz, 3 mT) on the lifespan, locomotion, heat shock response (HSR), and oxidative stress (OS) of Canton-Special (CS) and mutant w1118 flies were investigated at 25°C and 35°C (thermal stress). Results showed that thermal stress accelerated the death rates of CS and w1118 flies, shortened their lifespan, and influenced their locomotion rhythm and activity. The upregulated expression levels of heat shock protein (HSP) 22, HSP26, and HSP70 indicated that HSR was enhanced. Thermal stress-induced OS response increased malondialdehyde content, enhanced superoxide dismutase activity, and decreased reactive oxygen species level. The effects of thermal stress on the death rates, lifespan, locomotion, and HSP gene expression of flies, especially w1118 line, were also enhanced by ELF-EMF. In conclusion, thermal stress weakened the physiological function and promoted the HSR and OS of flies. ELF-EMF aggravated damages and enhanced thermal stress-induced HSP and OS response. Therefore, thermal stress and ELF-EMF elicited a synergistic effect.

Sperm of the giant grouper: cryopreservation, physiological and morphological analysis and application in hybridizations with red-spotted grouper.

In order to develop excellent germplasm resources for giant grouper (Epinephelus lanceolatus), cryopreservation of giant grouper sperm was examined in the present study. Firstly, 13 kinds of sperm dilution (ELS1-3, EM1-2, TS-2, MPRS, ELRS0-6) were prepared with physiological salt, sucrose, glucose and fetal bovine serum. The physiological parameters of ELRS3 (ratio of fast motion, ratio of slow motion, time of fast motion, time of slow motion, lifespan and motility) and ELS3 (sperm ratio of slow motion, time of slow motion and motility) were significantly higher than those of the other dilutions (P < 0.05). Secondly, after adding 15% DMSO and 10% FBS to ELRS3 and ELS3, most physiological parameters of frozen sperm were also significantly higher than the other gradients (P < 0.05), and sperm motility was as high as 63.68 ± 4.16% to74.75 ± 12.71% (fresh sperm motility, 80.70 ± 1.37% to 80.71 ± 1.49%). Mixed with the above dilutions, a final volume of 105 ml semen was cryopreserved. Finally, the sperm of giant grouper cryopreserved with cryoprotectants (ELRS3 + 15% DMSO + 10% FBS) was used for electron-microscopic observation and crossbreeding with red-spotted groupers (Epinephelus akaara). The electron-microscopic observation revealed that part of the frozen-thawed sperm was cryodamaged, e.g., flagellum fracturing and mitochondria falling out, while the ultrastructure of sperm membrane, mitochondria and flagellum remained intact. Also, the fertilization and hatchability rates of giant grouper frozen sperm and red-spotted grouper eggs were as high as 94.56% and 75.56%, respectively. Thus, a technique for cryopreservation of giant grouper sperm was successfully developed and applied to crossbreeding with red-spotted grouper eggs.

The Energy Metabolism in Caenorhabditis elegans under The Extremely Low-Frequency Electromagnetic Field Exposure.

A literal mountain of documentation generated in the past five decades showing unmistakable health hazards associated with extremely low-frequency electromagnetic fields (ELF-EMFs) exposure. However, the relation between energy mechanism and ELF-EMF exposure is poorly understood. In this study, Caenorhabditis elegans was exposed to 50 Hz ELF-EMF at intensities of 0.5, 1, 2, and 3 mT, respectively. Their metabolite variations were analyzed by GC-TOF/MS-based metabolomics. Although minimal metabolic variations and no regular pattern were observed, the contents of energy metabolism-related metabolites such as pyruvic acid, fumaric acid, and L-malic acid were elevated in all the treatments. The expressions of nineteen related genes that encode glycolytic enzymes were analyzed by using quantitative real-time PCR. Only genes encoding GAPDH were significantly upregulated (P < 0.01), and this result was further confirmed by western blot analysis. The enzyme activity of GAPDH was increased (P < 0.01), whereas the total intracellular ATP level was decreased. While no significant difference in lifespan, hatching rate and reproduction, worms exposed to ELF-EMF exhibited less food consumption compared with that of the control (P < 0.01). In conclusion, C. elegans exposed to ELF-EMF have enhanced energy metabolism and restricted dietary, which might contribute to the resistance against exogenous ELF-EMF stress.

Gene expression and reproductive abilities of male Drosophila melanogaster subjected to ELF-EMF exposure.

Extremely low frequency electromagnetic field (ELF-EMF) exposure is attracting increased attention as a possible disease-inducing factor. The in vivo effects of short-term and long-term ELF-EMF exposure on male Drosophila melanogaster were studied using transcriptomic analysis for preliminary screening and QRT-PCR for further verification. Transcriptomic analysis indicated that 439 genes were up-regulated and 874 genes were down-regulated following short-term exposures and that 514 genes were up-regulated and 1206 genes were down-regulated following long-term exposures (expression >2- or <0.5-fold, respectively). In addition, there are 238 up-regulated genes and 598 down-regulated genes in the intersection of short-term and long-term exposure (expression >2- or <0.5-fold). The DEGs (differentially expressed genes) in D. melanogaster following short-term exposures were involved in metabolic processes, cytoskeletal organization, mitotic spindle organization, cell death, protein modification and proteolysis. Long-term exposure let to changes in expression of genes involved in metabolic processes, response to stress, mitotic spindle organization, aging, cell death and cellular respiration. In the intersection of short-term and long-term exposure, a series of DEGs were related to apoptosis, aging, immunological stress and reproduction. To check the ELF-EMF effects on reproduction, some experiments on male reproduction ability were performed. Their results indicated that short-term ELF-EMF exposure may decrease the reproductive ability of males, but long-term exposures had no effect on reproductive ability. Down-regulation of ark gene in the exposed males suggests that the decrease in reproductive capacity may be induced by the effects of ELF-EMF exposure on spermatogenesis through the caspase pathway. QRT-PCR analysis confirmed that jra, ark and decay genes were down regulated in males exposed for 1 Generation (1G) and 72 h, which suggests that apoptosis may be inhibited in vivo. ELF-EMF exposure may have accelerated cell senescence, as suggested by the down-regulation of both cat and jra genes and the up-regulation of hsp22 gene. Up-regulation of totA and hsp22 genes during exposure suggests that exposed flies might induce an in vivo immune response to counter the adverse effects encountered during ELF-EMF exposure. Down-regulation of cat genes suggests that the partial oxidative protection system might be restrained, especially during short-term exposures. This study demonstrates the bioeffects of ELF-EMF exposure and provides evidence for understanding the in vivo mechanisms of ELF-EMF exposure on male D. melanogaster.