Na Na Kim

Profiles of antioxidant gene expression and physiological changes by thermal and hypoosmotic stresses in black porgy (Acanthopagrus schlegeli).

We determined oxidative stress by measuring the expression and activity of 3 antioxidant enzymes [Cu/Zn-superoxide dismutase (Cu/Zn-SOD), catalase (CAT) and glutathione peroxidase (GPX)] in black porgy exposed to thermal (20 degrees C-->30 degrees C) and hypoosmotic (35 psu-->10 psu and 0 psu) stresses. The expression and activity of antioxidant enzymes were significantly higher after exposure to 30 degrees C, 10 psu, and 0psu. Furthermore, we measured H(2)O(2) and lipid peroxidation (LPO) levels. As a result, H(2)O(2) and LPO levels were significantly increased after exposure to thermal (20 degrees C-->30 degrees C) and hypoosmotic stress (35 psu-->10 psu and 0 psu) stress. These results indicate that thermal and hypoosmotic stress induces oxidative stress in black porgy. Additionally, we investigated the changes due to thermal and hypoosmotic stress by measuring plasma cortisol and ion (Na(+) and Cl(-)) levels. Plasma cortisol levels increased at 30 degrees C and at 10 psu and then decreased at 0 psu. However, plasma Na(+) and Cl(-) levels did not change after exposure to thermal stress (30 degrees C), and decreased at 10 psu and 0 psu. In conclusion, thermal and hypoosmotic environments increase oxidative stress, thereby these results may be indicators of oxidative stress in black porgy.

Effect of LED light spectra on starvation-induced oxidative stress in the cinnamon clownfish Amphiprion melanopus

The present study aimed to test starvation-induced oxidative stress in the cinnamon clownfish Amphiprion melanopus illuminated by light-emitting diodes (LEDs): red (peak at 630 nm), green (peak at 530 nm), and blue (peak at 450 nm) within a visible light. We investigated the oxidative stress induced by starvation for 12 days during illumination with 3 LED light spectra through measuring antioxidant enzyme (superoxide dismutase [SOD] and catalase [CAT]) mRNA expression and activity; CAT western blotting; and measuring lipid peroxidation [LPO]), plasma H(2)O(2), lysozyme, glucose, alanine aminotransferase (AlaAT), aspartate aminotransferase (AspAT), and melatonin levels. In green and blue lights, expression and activity of antioxidant enzyme mRNA were significantly lower than those of other light spectra, results that are in agreement with CAT protein expression level by western blot analysis. Also, in green and blue lights, plasma H(2)O(2), lysozyme, glucose, AlaAT, AspAT, and melatonin levels were significantly lower than those in other light spectra. These results indicate that green and blue LEDs inhibit oxidative stress and enhance immune function in starved cinnamon clownfish.

The effect of LED light spectra on antioxidant system by thermal stress in goldfish, Carassius auratus

We investigated the effects of the different light emitting diodes (LEDs; red, green, blue, and mixed purple) on oxidative stress by measuring the expression and activities of antioxidant enzymes, plasma H2O2, lipid peroxidation (LPO), aspartate aminotransferase (AspAT), alanine aminotransferase (AlaAT) levels, and melatonin during thermal stress (low-water temperature, 22 → 14°C; high-water temperature, 22 → 30°C) on goldfish, Carassius auratus. The expression and activity of the antioxidant enzymes and levels of H2O2, LPO, AspAT/AlaAT, and melatonin were significantly higher after the fish were exposed to low-/high-water temperature environments. Furthermore, in green and blue lights, the expression of antioxidant enzymes and mRNA of heat shock protein 70, and the activity of the antioxidant enzymes and plasma H2O2, LPO, and melatonin levels were significantly lower than those under other light spectra. The results indicated that low-/high-water temperature conditions induced oxidative stress in goldfish and green and blue LEDs inhibit oxidative stress.

Kisspeptin regulates the hypothalamus–pituitary–gonad axis gene expression during sexual maturation in the cinnamon clownfish, Amphiprion melanopus

Kisspeptins (Kiss) have been recognized as potent regulators of reproduction in teleosts, and Kiss is suggested to be a key regulator of the hypothalamus-pituitary-gonad axis (HPG). However, its regulatory role on reproduction in fish remains unclear. Therefore, to investigate the role of Kiss on fish reproduction, this study aimed to test differences in the hormones of the HPG axis, Kiss as neuropeptides, and sex steroids on the sexual maturation of paired cinnamon clownfish, Amphiprion melanopus, following treatment with Kiss. We investigated the actions of sex maturation hormones, including HPG axis hormones and sex steroid hormones, such as gonadotropin-releasing hormones, gonadotropin hormones (GTHs), GTH receptors, estrogen receptors, and vitellogenin in the pituitary, gonads, and liver following treatment with Kiss. The expression levels of HPG axis genes increased after the Kiss injection. In addition, the levels of plasma 17α-hydroxypregnenolone, estradiol-17β, and 11-ketotestosterone increased. These results support the hypothesis that Kiss play important roles in the regulation of the HPG axis and are most likely involved in gonadal development and sexual maturation in cinnamon clownfish.

Expression profiles of three types of GnRH during sex-change in the protandrous cinnamon clownfish, Amphiprion melanopus: Effects of exogenous GnRHs

Gonadotropin-releasing hormones (GnRHs) play pivotal roles in the control of reproduction and gonadal maturation in teleost fish. Fish have multiple GnRH genes that encode structurally distinct peptides. We identified salmon GnRH (sGnRH), seabream GnRH (sbGnRH), and chicken GnRH-II (cGnRH-II) by cDNA cloning in cinnamon clownfish (Amphiprion melanopus) using reverse transcription-PCR (RT-PCR) and rapid amplification of cDNA ends-PCR (RACE-PCR). Gene identity was confirmed by sequence alignment and subsequent phylogenetic analyses. We also investigated GnRH mRNA expression in the gonads by quantitative real time-PCR (Q-PCR), and measured plasma estradiol-17β (E(2)) levels in immature fish following treatment with the three molecular forms of GnRHs. The expression levels of sGnRH, sbGnRH, and cGnRH-II mRNA were higher in mature testes and ovaries, as compared to the levels in gonads at earlier stages of maturity. The levels of the three prepro-GnRH mRNA species and the plasma E(2) levels increased after injection of the three GnRH variants. These findings support the hypothesis that GnRH peptides play important roles in the regulation of the hypothalamic-pituitary-gonadal axis and are probably involved in paracrine control of gonadal development and sex change in cinnamon clownfish.

Effects of recombinant gonadotropin hormones on the expression of vitellogenin, gonadotropin subunits and gonadotropin receptors in cinnamon clownfish, Amphiprion melanopus.

Gonadotropins (GTHs) are the key regulators of reproduction in vertebrates. The present study investigated autoregulatory effects of gonadotropins, using recombinant FSH (rFSH) and LH (rLH) in cinnamon clownfish (Amphiprion melanopus). Experiments were carried out to investigate the actions of cinnamon clownfish rFSH and rLH on expression of GTH subunits, GTH receptors, and vitellogenin (Vtg) mRNA in vivo and in vitro. Plasma estradiol-17β (E(2)) level was also measured in immature fish following treatments with rFSH and rLH. The results demonstrate increasing levels of GTH subunits, GTH-receptors, Vtg mRNA levels, as well as plasma E(2) levels following injection with rFSH and rLH. The findings support the hypothesis that LH and FSH stimulate reproduction, in part, by autoregulatory mechanisms leading to upregulation of GTH receptors and GTH hormone production in cinnamon clownfish. The results provide a framework for better understanding of the mechanisms of GTH-mediated control of reproduction in cinnamon clownfish and other vertebrates.

Upregulation of estrogen receptor subtypes and vitellogenin mRNA in cinnamon clownfish Amphiprion melanopus during the sex change process: profiles on effects of 17β-estradiol.

In the present study, we investigated the expression pattern of estrogen receptors (esr) and vitellogenin (vtg) mRNA in the gonads and liver during sex change in cinnamon clownfish by using quantitative polymerase chain reaction. We divided gonadal development during the sex change from male to female into 3 stages (mature male, male at 90days after removing female, and mature female) and investigated esr and vtg mRNA expressions during the sex change. With female, the esr and vtg mRNA expressions increased. In western blot analysis, Esr1 protein was detected only in the ovaries of female cinnamon clownfish. Also, to understand the effect of 17beta-estradiol (E(2)), we investigated the esr and vtg mRNA expression patterns in the gonads and liver, and the changes in plasma E(2) level after E(2) injection. E(2) treatment increased both mRNA expression levels of esr and vtg and plasma E(2) levels. The present study describes the molecular characterization of esr subtypes and the interactions between esr and vtg after E(2) treatment in cinnamon clownfish.

Effects of gonadotropin inhibitory hormone or gonadotropin-releasing hormone on reproduction-related genes in the protandrous cinnamon clownfish, Amphiprion melanopus.

Hypothalamic peptide neurohormones such as gonadotropin-releasing hormones (GnRHs) and gonadotropin-inhibitory hormone (GnIH) play pivotal roles in the control of reproduction and gonadal maturation in teleost fish. To study the effects of GnIH on fish reproduction, we investigated the influence of seabream GnRH (sbGnRH) and GnIH (both alone and in combination) on levels of reproductive genes (GnIH, GnIH-receptor [GnIH-R], melatonin receptor [MT3], sbGnRH, and gonadotropic hormones [GTHs]) during different stages of gonadal maturation in male, female, and immature cinnamon clownfish, Amphiprion melanopus. The results showed that the expression levels of GnIH, GnIH-R, and MT3 genes increased after the GnIH injection, but decreased after the sbGnRH injection. In addition, these gene expression levels gradually lowered after GnIH3 and sbGnRH combination treatment, as compared to the MT3 mRNA levels of GnIH treatment alone. However, the expression levels of the HPG (hypothalamus-pituitary-gonad) axis genes (sbGnRH and GTHs) decreased after the GnIH injection, but increased after the sbGnRH injection. In all cinnamon clownfish groups, HPG axis gene mRNA levels gradually decreased after mixed GnIH3 and sbGnRH treatment, compared to GnIH treatment alone. The present study provides novel information on the effects of GnIH and strongly supports the hypothesis that GnIH plays an important role in the negative regulation of the HPG axis in the protandrous cinnamon clownfish.

Expression of aquaporin-3 and 8 mRNAs in the parr and smolt stages of sockeye salmon, Oncorhynchus nerka: Effects of cortisol treatment and seawater acclimation

This study aimed to examine the role of 2 aquaporin (AQP) isoforms (AQP3, and -8) in sockeye salmon (Oncorhynchus nerka) in response to a hyperosmotic challenge from freshwater to seawater (SW) during the parr and smoltification (smolt) stages. AQP3 mRNA was primarily detected in the osmoregulatory organs, such as gills, while AQP8 mRNA was primarily found in the intestine. These results suggested that AQP isoforms play a role in osmoregulation in specific osmoregulatory organs. Similarly, AQP3 mRNA expression in the gills (mean values:1.06 ± 0.05 [parr] and 1.29 ± 0.07 [smolt]) was significantly higher than AQP8 mRNA levels (parr: 0.04 ± 0.003; smolt: 0.14 ± 0.004), and in the intestine, AQP8 mRNA expression (parr: 0.89 ± 0.007; smolt: 1.91 ± 0.03) was significantly higher than AQP3 mRNA levels (parr: 0.24 ± 0.006; smolt: 0.83 ± 0.005); these expression patterns were similar in vivo and in vitro. Additionally, AQP mRNA levels were lower in cortisol treated than in control groups. Therefore, these results suggest that AQPs play important roles in the water absorption mechanisms associated with multiple AQP isoforms, and that cortisol enhances the hypo-osmoregulatory capacity of fish in SW, and also controls the expression of AQPs in a hyperosmotic environment.

Effects of various LED light spectra on antioxidant and immune response in juvenile rock bream, Oplegnathus fasciatus exposed to bisphenol A.

Bisphenol A (BPA) is a monomer used in plastics and plasticizers. As an environmental toxin included in industrial wastewater, it contaminates the aquatic environment and is known to cause endocrine disruption in fish. Particular wavelengths of light-emitting diodes (LEDs) are known to affect the endocrine regulation of fish. The present study aimed to investigate the effects of green and red LED light on the antioxidant and immune systems in juvenile rock bream (Oplegnathus fasciatus) exposed to BPA. We used green and red LED exposure at two intensities (0.3 and 0.5W/m(2)) for 1, 3, and 5 days. We measured liver mRNA expression and plasma levels of antioxidant enzyme superoxide dismutase (SOD) and caspase-3. Furthermore, we measured plasma levels of hydrogen peroxide (H2O2), lipid peroxidation (LPO), melatonin, and immunoglobulin M (IgM). DNA damage and apoptotic activity were measured using comet and terminal transferase dUTP nick end labeling (TUNEL) assays, respectively. We found that SOD, H2O2, and LPO increased significantly, whereas melatonin and IgM decreased significantly, suggesting that BPA induces oxidative stress and reduces immune function. Likewise, both DNA damage and apoptotic activity increased following BPA exposure. However, we found that exposure to green LED light effectively reduced the detrimental effects induced by BPA, including decreasing DNA damage, apoptotic activity, SOD mRNA expression, and plasma levels of SOD, H2O2, and LPO. Likewise, the plasma levels of melatonin and IgM increased. Thus, our results indicate that green light conditions effectively reduces oxidative stress and promotes the immune function in juvenile rock bream.