Production of Recombinant Single-Chain Eel Luteinizing Hormone and Follicle-Stimulating Hormone Analogs in Chinese Hamster Ovary Suspension Cell Culture.

We produced rec-single chain eel luteinizing (rec-eel LH) and follicle-stimulating (rec- eel FSH) hormones displaying high biological activity in Chinese hamster ovary suspension (CHO-S) cells. We constructed several mutants, in which a linker, including an O-linked glycosylated carboxyl-terminal peptide (CTP) of an equine chorionic gonadotropin (eCG) ß-subunit, was attached between the ß- and α-subunit (LH-M and FSH-M) or in the N-terminal (C-LH and C-FSH) or C-terminal (LH-C and FSH-C) regions. The plasmids were transfected into CHO-S cells, and culture supernatants were collected. The secretion of mutants from the CHO-S cells was faster than that of eel LHß/α-wt and FSHß/α-wt proteins. The molecular weight of eel LHß/α-wt and eel FSHß/α-wt was 32-34 and 34-36 kDa, respectively, and that of LH-M and FSH-M was 40-43 and 42-45 kDa, respectively. Peptide-N-glycanase F-treatment markedly decreased the molecular weight by approximately 8-10 kDa. The EC50 value and the maximal responsiveness of the eel LH-M and eel FSH-M increased compared with the wild-type proteins. These results show that the CTP region plays a pivotal role in early secretion and signal transduction. We suggest that novel rec-eel LH and FSH proteins, exhibiting potent activity, could be produced in large quantities using a stable CHO cell system.

Stable Production of a Tethered Recombinant Eel Luteinizing Hormone Analog with High Potency in CHO DG44 Cells.

We produced a recombinant eel luteinizing hormone (rec-eel LH) analog with high potency in Chinese hamster ovary DG44 (CHO DG44) cells. The tethered eel LH mutant (LH-M), which had a linker comprising the equine chorionic gonadotropin (eLH/CG) ß-subunit carboxyl-terminal peptide (CTP) region (amino acids 115 to 149), was inserted between the ß-subunit and α-subunit of wild-type tethered eel LH (LH-wt). Monoclonal cells transfected with the tethered eel LH-wt and eel LH-M plasmids were isolated from five to nine clones of CHO DG44 cells, respectively. The secreted quantities abruptly increased on day 3, with peak levels of 5000-7500 ng/mL on day 9. The molecular weight of tethered rec-eel LH-wt was 32-36 kDa, while that of tethered rec-eel LH-M increased to approximately 38-44 kDa, indicating the detection of two bands. Treatment with the peptide N-glycanase F decreased the molecular weight by approximately 8 kDa. The oligosaccharides at the eCG ß-subunit O-linked glycosylation sites were appropriately modified post-translation. The EC50 value and maximal responsiveness of eel LH-M increased by approximately 2.90- and 1.29-fold, respectively, indicating that the mutant exhibited more potent biological activity than eel LH-wt. Phosphorylated extracellular regulated kinase (pERK1/2) activation resulted in a sharp peak 5 min after agonist treatment, with a rapid decrease thereafter. These results indicate that the new tethered rec-eel LH analog had more potent activity in cAMP response than the tethered eel LH-wt in vitro. Taken together, this new eel LH analog can be produced in large quantities using a stable CHO DG44 cell system.

Stable Production of a Recombinant Single-Chain Eel Follicle-Stimulating Hormone Analog in CHO DG44 Cells.

This study aimed to produce single-chain recombinant Anguillid eel follicle-stimulating hormone (rec-eel FSH) analogs with high activity in Cricetulus griseus ovary DG44 (CHO DG44) cells. We recently reported that an O-linked glycosylated carboxyl-terminal peptide (CTP) of the equine chorionic gonadotropin (eCG) ß-subunit contributes to high activity and time-dependent secretion in mammalian cells. We constructed a mutant (FSH-M), in which a linker including the eCG ß-subunit CTP region (amino acids 115-149) was inserted between the ß-subunit and α-subunit of wild-type single-chain eel FSH (FSH-wt). Plasmids containing eel FSH-wt and eel FSH-M were transfected into CHO DG44 cells, and single cells expressing each protein were isolated from 10 and 7 clones. Secretion increased gradually during the cultivation period and peaked at 4000-5000 ng/mL on day 9. The molecular weight of eel FSH-wt was 34-40 kDa, whereas that of eel FSH-M increased substantially, with two bands at 39-46 kDa. Treatment with PNGase F to remove the N glycosylation sites decreased the molecular weight remarkably to approximately 8 kDa. The EC50 value and maximal responsiveness of eel FSH-M were approximately 1.23- and 1.06-fold higher than those of eel FSH-wt, indicating that the mutant showed slightly higher biological activity. Phosphorylated extracellular-regulated kinase (pERK1/2) activation exhibited a sharp peak at 5 min, followed by a rapid decline. These findings indicate that the new rec-eel FSH molecule with the eCG ß-subunit CTP linker shows potent activity and could be produced in massive quantities using the stable CHO DG44 cell system.

The Religious Dimensions of Epidemic Disease: Cholera, the Ghost Rite, and Missionary Medicine in Nineteenth-Century Korea.

One of the most catastrophic pandemics in human history was the repeated spread of cholera in the nineteenth century. In spite of its historical significance, few scholars have studied cholera's influence in East Asia. This paper illustrates how cholera was considered, conceptualized, and treated by Korean people prior to contact with North American medical missionaries in 1885. In particular, the article compares the government-ordered public health measures during the Joseon dynasty, focusing on the "ghost rite" performed during outbreaks of epidemic disease with the work of medical missionaries in the late nineteenth century. This study finds that even after the introduction of Western biomedicine, the Korean people persisted with a religious-based etiology of cholera and other infectious diseases until the twentieth century.

Contesting a Medical Gaze: 'The 1888 Baby Riot' in Seoul - Social, Cultural, and Religious Collision.

This paper focuses on the encounter and collision of two different cultural systems in the influx of biomedicine and Protestant Christianity during the 1888 'baby riot' in Seoul, Korea. This research scrutinizes the relationship between religion and medicine in modern Korea to illustrate why Korean people contested against the nexus of Protestant Christianity and biomedicine that had been introduced from the West. For them, biomedicine was not simply a way of treating sickness or disease, but a way in which their bodies were examined and manipulated by placing a new gaze on the human body. These disputes developed due to the significant differences between biomedicine and the traditional perspective that was based on the values of Confucian teachings. The Korean people desired to protect the belief that the human body was a locus of virtue and should be preserved without modification, which was considered as a pivotal part of their social identity. Therefore, people who lacked social and political power tried to express their opposition to the medicine and religion from the West by spreading rumors of cannibalism. Through the spread of the rumors, the people interpreted and manifested that Western medicine was cruel, ignorant, immoral and dehumanizing. This paper states that Koreans were not merely passive recipients of biomedicine and Protestant Christianity, but also appropriated the new medical and religious discourses to maintain their dignity and cultural authority against the current of colonization.

Characterization of Latilactobacillus curvatus MS2 isolated from Korean traditional fermented seafood and cholesterol reduction effect as synbiotics with isomalto-oligosaccharide in BALB/c mice.

This study investigated the properties of Latilactobacillus curvatus MS2 isolated from Korean traditional fermented seafood as probiotics and the effect of reducing cholesterol as a synbiotic with isomalto-oligosaccharide (IMO) in BALB/c mice. The isolated strain showed high resistance to acids and bile acids and exhibited a high DPPH scavenging capacity of 72.27 ± 0.38 %. In the intestinal adhesion test using HT-29 cells, the adhesion rate of MS2 was 17.10 ± 1.78 %, which was higher than the adhesion rate of the other investigated probiotics. MS2 showed good antimicrobial activity against food-borne pathogens, especially Staphylococcus aureus, S. epidermidis, Escherichia coli, and Vibrio vulnificus. This strain had high availability for IMO among the prebiotics of fructo-oligosaccharide, inulin and IMO. Oral administration of MS2 and IMO to BALB/c mice for 5 weeks resulted in a significant reduction in blood cholesterol levels by regulating liver lipid metabolism. These results suggest that the combination of MS2 and IMO has potential for application in functional foods.

Dietary taurine stimulates the hepatic biosynthesis of both bile acids and cholesterol in the marine teleost, tiger puffer (Takifugu rubripes).

Taurine (TAU) plays important roles in the metabolism of bile acids, cholesterol and lipids. However, little relevant information has been available in fish where TAU has been identified as a conditionally essential nutrient. The present study aimed to investigate the effects of dietary TAU on the metabolism of bile acids, cholesterol and lipids in tiger puffer, which is both an important aquaculture species and a good research model, having a unique lipid storage pattern. An 8-week feeding trial was conducted in a flow-through seawater system. Three experimental diets differed only in TAU level, that is, 1·7, 8·2 and 14·0 mg/kg. TAU supplementation increased the total bile acid content in liver but decreased the content in serum. TAU supplementation also increased the contents of total cholesterol and HDL-cholesterol in both liver and serum. The hepatic bile acid profile mainly includes taurocholic acid (94·48 %), taurochenodeoxycholic acid (4·17 %) and taurodeoxycholic acid (1·35 %), and the contents of all these conjugated bile acids were increased by dietary TAU. The hepatic lipidomics analysis showed that TAU tended to decrease the abundance of individual phospholipids and increase those of some individual TAG and ceramides. The hepatic mRNA expression study showed that TAU stimulated the biosynthesis of both bile acids and cholesterol, possibly via regulation of farnesoid X receptor and HDL metabolism. TAU also stimulated the hepatic expression of lipogenic genes. In conclusion, dietary TAU stimulated the hepatic biosynthesis of both bile acids and cholesterol and tended to regulate lipid metabolism in multiple ways.

Daily variation of D2 dopamine receptor transcription in the brain of the Japanese eel Anguilla japonica and its regulation with dopamine and melatonin.

Dopamine plays a crucial role in controlling reproduction in eels, and its action is mediated through D2-type dopamine receptors. D2A and D2B receptors in the Japanese eel Anguilla japonica were cloned and characterized in the present study. Attention (daily expression patterns in the brain and endogenous regulation) was paid to D2B receptor because it is considered to play a crucial role in eel reproduction. The cDNAs of D2A and D2B receptors had open reading frames comprising 456 and 454 amino acid residues, respectively, which were phylogenetically clustered with those of other teleost species. Both receptors were highly expressed in the brain. D2B receptor transcript levels exhibited high day/low night variation in the midbrain and pituitary, suggesting that its transcription in these tissues is regulated in a daily manner, possibly under influence of melatonin. Intraperitoneal injection of dopamine downregulated D2B receptor transcription significantly in the midbrain and moderately in the pituitary within 1 h, but upregulated its transcription in the forebrain. Co-injection of dopamine with its antagonist (domperidone) reversed the effect of dopamine in the pituitary and forebrain, but not in the midbrain, suggesting that the effect of dopamine on D2B receptor transcription differs among brain regions. The same treatment with melatonin resulted in decreased D2B receptor transcription in the midbrain. These findings indicate that dopamine and melatonin have key roles in the daily variation in D2B receptor transcription in the brain of Japanese eel, and that they are related to a daily base secretion of hormones in the hypothalamic-pituitary-gonadal axis in this species.

Contribution of chitooligosaccharides to biofilm formation, antibiotics resistance and disinfectants tolerance of Listeria monocytogenes.

Listeria monocytogenes is a food-borne pathogen present in various environmental reservoirs. It exhibits resistance and tolerance to antibiotics and sanitizing agents used in several food processing industries. It has been reported that L. monocytogenes chitinase can catalyze hydrolysis of chitin polymeric carbohydrate present in the environment and act as a virulence factor that support its survival in mammalian host cells. By taking advantage of chitinase, L. monocytogenes has both saprophytic and pathogenic lifestyles in the soil and the living host, respectively. The objective of the present study was to determine the involvement of chitin degradation products such as chitooligosaccharides (COS) in biofilm formation of L. monocytogenes. Results showed that different concentrations of COS with various molecular weight enhanced biofilm formation of L. monocytogenes. Such enhancement in biofilm formation contributed to the development of antibiotics resistance and disinfectants tolerance of cells present in the biofilm. The present article also described diverse roles of chitin, chitinase, and degradation of chitin and chitin-like substrates in saprophytic and pathogenic lifestyles of L. monocytogenes. This study offers a new direction for further exploration of the mechanisms of pathogenesis caused by L. monocytogenes.

Antibiofilm and antivirulence properties of chitosan-polypyrrole nanocomposites to Pseudomonas aeruginosa.

Pseudomonas aeruginosa is an opportunistic human pathogen which exhibits its property of pathogenesis due to several factors, including the formation of biofilm and production of several virulence factors. Development of resistance properties against antibiotics leads to the discovery of certain alternative strategies to combat its pathogenesis. In the present study, a highly stable, biocompatible and water soluble nanocomposites (NCs) are synthesized from chitosan (CS) and the polypyrrole (PPy). The resultant chitosan-polypyrrole nanocomposites (CS-PPy NCs) inhibit the establishment of biofilm and also eradicate the preformed matured biofilm formed by P. aeruginosa. CS-PPy NCs inhibit the hemolytic and protease activities of P. aeruginosa. The NCs significantly reduce the production of many virulence factors such as pyocyanin, pyroverdine and rhamnolipid. CS-PPy NCs also suppress the bacterial motility such as swimming and swarming. The present study showed that highly stable CS-PPy NCs act as a potent antibiofilm and antivirulence drug for the treatment of P. aeruginosa infection.

Site-specific roles of N-linked oligosaccharides in recombinant eel follicle-stimulating hormone for secretion and signal transduction.

Eel follicle-stimulating hormone (eelFSH) is composed of a common α-subunit and a hormone specific ß-subunit, both of which contain two N-linked carbohydrate residues. We characterized the biologically active single chains by fusing the α-subunit to the carboxyl terminal region of the eelFSH ß-subunit. Expression vectors were constructed and the biological activity of the recombinant hormones (rec-hormones) was characterized using Chinese hamster ovary (CHO) K1 cells expressing the eelFSH receptor gene. Mutagenesis of the individual and double glycosylated sites was performed to determine the functions of the oligosaccharide chains on signal transduction. The absence of the Asn22 (eelFSHßΔ22/α) and Asn5.22 (eelFSHßΔ5.22/α) N-linked oligosaccharide chain in the eelFSH ß-subunit completely reduced the secretion level in the medium and cell lysate of CHO-K1 cells. The expression levels of eelFSHß/α wild-type in CHO suspension (CHO-S) cells was approximately 4-fold higher in CHO-k1 cells. The molecular weight of rec-eelFSHß/α wild-type by western blotting analysis was found to be 34 kDa. Mutants (ß/αΔ56, ß/αΔ79, and ßΔ5/α) lacking single oligosaccharide sites showed molecular weights that were reduced by approximately 10%. The digestion of N-linked oligosaccharides using PNGaseF treatment showed that the molecular weights of all mutants were reduced to 27-kDa. The oligosaccharide chains in rec-eelFSHß/α wild-type were modified to a molecular weight of approximately 7-10 kDa in CHO-K1 and CHO-S cells. Oligosaccharide site deletions at positions Asn56 and Asn79 on the α-subunit and Asn5 on the ß-subunit were found to play an essential role in cAMP signal transduction through the eelFSH receptor. The EC50 values of Asn56 and Asn5 resulted in a significant decrease in potency to 64% and 53% of the wild type, respectively. Specifically, the removal of the carbohydrates at Asn79 of the α-subunit (ß/αΔ79) was drastically reduced to 53.8% of the wild-type levels in maximum response. These results have allowed for the identification of the site-specific roles of carbohydrate residues in eel FSH. Our data suggest that N-linked oligosaccharide chains play a pivotal role in biological activity through the eelFSH receptor as suggested in similar studies of other mammalian FSH hormones.

Identification and molecular characterization of peroxiredoxin 6 from Japanese eel (Anguilla japonica) revealing its potent antioxidant properties and putative immune relevancy.

Peroxiredoxins (Prdx) are thiol specific antioxidant enzymes that play a pivotal role in cellular oxidative stress by reducing toxic peroxide compounds into nontoxic products. In this study, we identified and characterized a peroxiredoxin 6 counterpart from Japanese eel (Anguilla japonica) (AjPrdx6) at molecular, transcriptional and protein level. The identified full-length coding sequence of AjPrdx6 (669 bp) coded for a polypeptide of 223 aa residues (24.9 kDa). Deduced protein of AjPrdx6 showed analogy to characteristic structural features of 1-cysteine peroxiredoxin sub-family. According to the topology of the generated phylogenetic reconstruction AjPrdx6 showed closest evolutionary relationship with Salmo salar. As detected by Quantitative real time PCR (qPCR), AjPrdx6 mRNA was constitutively expressed in all the tissues examined. Upon the immune challenges with Edwardsiella tarda, lipopolysaccharides and polyinosinic:polycytidylic acid, expression of AjPrdx6 mRNA transcripts were significantly induced. The general functional properties of Prdx6 were confirmed using purified recombinant AjPrdx6 protein by deciphering its potent protective effects on cultured vero cells (kidney epithelial cell from an African green monkey) against H2O2-induced oxidative stress and protection against oxidative DNA damage elicited by mixed function oxidative (MFO) system. Altogether, our findings suggest that AjPrdx6 is a potent antioxidant protein in Japanese eels and its putative immune relevancy in pathogen stress mounted by live-bacteria or pathogen associated molecular patterns (PAMPs).

Introducing Open Dialogue as part of the WHO QualityRights Project in South Korea: experiences and opinions from an introductory workshop and 1-year pilot practice.

This study explores the subjective experiences of participants in a 5-day Open Dialogue (OD) workshop and a 1-year pilot practice, conducted as part of the WHO QualityRights Project in South Korea. Twenty-four participants, selected through purposive sampling, completed surveys immediately after the workshop and 1 year later. Data were analyzed through both statistical and thematic approaches. A statistically significant decrease in the availability of "Flexibility and Mobility" was observed across all participants (p = 0.044) and a significant reduction in the availability of "Tolerance of Uncertainty" (p = 0.04) was noted among participants who engaged in network meetings over the course of 1 year. Qualitative analysis revealed that participants initially felt ambivalent toward OD due to systemic, cultural, and professional challenges. However, through experiential learning, their ambivalence shifted to hope, fostering solidarity and a more positive outlook for future OD practice. Participants recognized that implementing OD supported human rights, while addressing personal, organizational, and policy challenges. The findings provide important insights for developing OD training and implementation guidelines in South Korea. Recommendations include focusing on experiential learning and selecting mixed-group trainees from catchment area institutions, emphasizing the support of client rights, and considering individual, organizational, and systemic levels for successful implementation. This study represents a new case of OD dissemination through a top-down national research and development project and its integration into the WHO QualityRights service package, suggesting complementary potential between OD and global human rights-based mental health initiatives.

Morphological and Allometric Changes in Anguilla japonica Larvae.

The freshwater eel Anguilla japonica is rapidly decreasing in number and has not yet been successfully mass produced. This may be at least partially attributable to the unique and long early life history of the eel. Therefore, we investigated its ontogeny of morphometry and growth pattern in larval stages to provide baseline information for understanding the early life history and improving seed rearing technology. This study was conducted for 200 days after hatching (DAH) and analyzed morphometry and allometry for eel larvae. The following cultured eel larval stages were identified: the yolk sac larvae stage (0-6 DAH, 3.23-6.85 mm total length (TL)), the pre-leptocephalus stage (7-30 DAH, 6.85-15.31 mm TL), and the leptocephalus stage (50-200 DAH, 15.31-60.06 mm TL). Cultured and wild eel larvae could be divided into characteristic larval stages at similar sizes. However, compared to wild eels, cultured eels had a slower growth rate and fewer preanal myomeres. Meanwhile, cultured eel larvae rarely had a mixed feeding period as the absorption of endogenous reserves was completed by 7 DAH. The lower jaw of eel larvae was significantly longer than the upper jaw from 50 DAH. In the pre-leptocephalus and leptocephalus stages, eel larvae showed continuous positive allometric growth at trunk height and tail muscle height with change to the willow leaf-like form. These growth characteristics may be the result of adaptation to the migration over long distances and to a diel vertical migration. The inflection point in the body parts growth patterns showed only before 30 DAH, and mass mortality appeared at this period. Therefore, to improve the growth and survival rates of cultured eel seed, it is necessary to focus on improving the feeding and rearing protocol until 30 DAH.

Effect of Bacillus sp. Supplementation Diet on Survival Rate and Microbiota Composition in Artificially Produced Eel Larvae (Anguilla japonica).

This study was performed to investigate the effect of microbial supplementation diet on the survival rate and microbiota composition of artificially produced eel larvae. Microorganisms supplemented in the diet were isolated from wild glass eel intestines and identified as Bacillus sp. through 16S rRNA sequencing analysis. In vitro tests confirmed that the strain had no hemolytic activity and virulence genes. Microbial supplemental feeding significantly increased the survival rate of artificially produced eel larvae for 30 days post-hatchling compared with that of the control group. It also caused changes in the α-diversity, ß-diversity, and relative abundance of the bacterial communities. Analysis via phylogenetic investigation of communities by reconstruction of unobserved states predicted that these microbial community changes would significantly increase the carbohydrate metabolism, membrane transport, and cellular community pathway of the microbial supplementation group. Therefore, microbial supplementation feeding for eel aquaculture could increase the viability of artificially produced eel larvae and alter the microbial composition to induce metabolic changes.

DNA barcoding and morphological identification of spiny lobsters in South Korean waters: a new record of Panulirus longipes and Panulirus homarus homarus.

To date, 19 species of spiny lobsters from the genus Panulirus have been discovered, of which only P. japonicus, P. penicilatus, P. stimpsoni, and P. versicolor have been documented in South Korean waters. In this study, we aimed to identify and update the current list of spiny lobster species that inhabit South Korean waters based on the morphological features and the phylogenetic profile of cytochrome oxidase I (COI) of mitochondrial DNA (mtDNA). Spiny lobsters were collected from the southern and eastern coasts of Jeju Island, South Korea. Phylogenetic analyses were performed using neighbor-joining (NJ), maximum likelihood (ML), and Bayesian inference (BI) methods. The ML tree was used to determine the spiny lobster lineages, thereby clustering the 17 specimens collected in this study into clades A, B, C, and D, which were reciprocally monophyletic with P. japonicus, P. homarus homarus, P. longipes, and P. stimpsoni, respectively. These clades were also supported by morphological examinations. Interestingly, morphological variations, including the connected pleural and transverse groove at the third abdominal somite, were observed in four specimens that were genetically confirmed as P. japonicus. This finding is novel within the P. japonicus taxonomical reports. Additionally, this study updates the documentation of spiny lobsters inhabiting South Korean waters as P. longipes and P. homarus homarus were recorded for the first time in this region.

Comparison between wild and hatchery populations of Korean pen shell (Atrina pectinata) using microsatellite DNA markers.

Pen shell (Atrina pectinata) is a popular food source with a high commercial value in a number of Asian Pacific areas. The natural A. pectinata population has been declining continuously over the past several decades. Microsatellite DNA markers are a useful DNA-based tool for monitoring the genetic variation of pen shell populations. In this study, 20 polymorphic microsatellite (MS) DNA markers were identified from a partial genomic pen shell DNA library enriched in CA repeats, and used to compare allelic variation between wild and hatchery pen shell populations in Korea. A total of 438 alleles were detected at the 20 MS loci in the two populations. All loci were easily amplified and demonstrated allelic variability, with the number of alleles ranging from 5 to 35 in the wild population and from 5 to 22 in the farmed population. The average observed and expected heterozygosities were 0.69 and 0.82, respectively, in the hatchery samples and 0.69 and 0.83, respectively, in the wild samples. Statistical analysis of fixation index (F(ST)) and analysis of molecular variance (AMOVA) showed minor, but significant, genetic differences between the wild and hatchery populations (F(ST) = 0.0106, CI(95%) = 0.003-0.017). These microsatellite loci may be valuable for future aquaculture and population genetic studies for developing conservation and management plans. Further studies with additional pen shell samples are needed to conclusively determine the genetic diversity between the wild and hatchery populations.

Testis development in the Japanese eel is affected by photic signals through melatonin secretion.

OBJECTIVE: According to reported spawning characteristics of Japanese eel, Anguilla japonica, which exhibit spawning and migration patterns that are synchronized with lunar cycles and photoperiod, we hypothesized that a close association exists between specific photic signals (daylight, daylength, and moonlight) and endocrinological regulation. Given the photic control in melatonin secretion, this hypothesis was tested by investigating whether melatonin signals act as mediators relaying photic signals during testis development in the eel. METHODS: We examined changes in melatonin-secretion patterns using time-resolved fluorescence immunoassays in sexually immature and mature male Japanese eels under the condition of a new moon (NM) and a full moon (FM). RESULTS: The eye and plasma melatonin levels exhibited a nocturnal pattern under a 12-h light: dark cycle (12L12D) or under constant darkness (DD), but not with constant light (LL). Eye melatonin levels were similar under the 12L12D and short-day (9L15D) conditions. In the long-day condition (15L9D), secreted plasma melatonin levels were stable, whereas short-day melatonin secretion began when darkness commenced. Sexual maturation began at 8 weeks following intraperitoneal injection of human chorionic gonadotropin (hCG), and NM exposure led to significantly higher eye and plasma melatonin levels compared with those detected under FM exposure.

Involvement of Estrogen and Its Receptors in Morphological Changes in the Eyes of the Japanese Eel, Anguilla japonica, in the Process of Artificially-Induced Maturation.

During the long migration from river habitats to the spawning ground, the Japanese eel undergoes sexual maturation. This spawning migration occurs concurrently with morphological changes, such as increases in eye size; however, the mechanisms by which sex steroids and their receptors influence these changes in peripheral tissues remain unclear. The aim of this study was to investigate changes in the eyes of female Japanese eels during sexual maturation, and our research focused on estrogen receptor (ER)α and ERß transcripts. During ovarian development, the gonadosomatic index increased and yolk-laden oocytes developed rapidly. These changes occurred in conjunction with a steady increase in plasma levels of estradiol-17ß (E2). Concomitant increases in transcript levels of ERα and ERß in eye, brain, pituitary, and ovary were also observed. Fluorescence in-situ hybridization analyses revealed that ERα and ERß transcripts were present in the choriocapillary layer and photoreceptor layer of the eyes, and the analysis also revealed that their signals in these layers became stronger in mature females compared to those observed in immature females, suggesting that under the influence of gonadotropins, morphological changes in the eyes are regulated by E2 through the activation of its receptors. In conclusion, E2 plays a crucial role in physiological adaptations that occur in peripheral tissues during the spawning migration.

Effects of Lactococcus lactis subsp. lactis I2 with ß-Glucooligosaccharides on Growth, Innate Immunity and Streptococcosis Resistance in Olive Flounder (Paralichthys olivaceus).

To identify and quantify the effects of a combination of dietary 1 × 108 CFU/g Lactococcus lactis subsp. lactis I2 (LI2) and 0.1% ß-glucooligosaccharides (BGO) on the growth and immunity of olive flounder (Paralichthys olivaceus), a feeding experiment was conducted. Flounder (14 ± 0.5 g) were divided into two groups and fed control and synbiotic feeds for 8 weeks. Investigations were carried out on growth and feed utilization, innate immunity, serum biochemical parameters, intestinal lactic acid bacterial (LAB) viability, microvillus length, and changes in the expression levels of genes encoding pro-inflammatory cytokines (tumor necrosis factor [TNF]-α, interleukin [IL]-1ß, and IL-6). Results demonstrated the synbiotic diet had significantly better (p < 0.05) responses in terms of weight gain and specific growth rate, three innate immune parameters (respiratory burst, serum lysozyme, and superoxide dismutase), intestinal LAB viability, and the relative TNF-α expression level (p < 0.05). Moreover, after challenge with Streptococcus iniae (1 × 108 CFU/ml), the synbiotically fed group exhibited significantly higher (p < 0.05) protection against streptococcosis, validating the observed changes in immune parameters and induction of the cytokine-encoding gene. Therefore, according to the results of the present study, synbiotic feed (LI2 + BGO) increased growth, modulated innate immune parameters and protected olive flounder against streptococcosis.