Different diatoms induce contrasting effects on the reproductive success of the copepod Temora stylifera

Four diatoms, Thalassiosira rotula (THA), Thalassiosira weissflogii (TWEI), Skeletonema costatum (SKE) and Phaeodactylum tricornutum (PHA), and one dinoflagellate Prorocentrum minimum (PRO, control diet), were tested during the course of 14-day laboratory experiments examining the effects of these algae on the fecundity and egg viability in the copepod Temora stylifera. All algae were provided at the same mean carbon concentrations of 0.98 μg C ml⁻¹ to normalize the effects due to differences in diatom size and nutrient concentrations, at least with regards to carbon, so that variations in egg production and hatching viability were presumably due to other chemical constituents of the cells. Our results show that only PRO supported high reproductive success, with stable egg production rates and egg viability close to 100%. By contrast, all four diatoms negatively impacted egg production and hatching success, with varying degrees of suppression. The two diets SKE and PHA induced dramatic effects on reproductive success; SKE was the worst diet with naupliar production ceasing altogether within only 3 days. With PHA, fecundity never reached zero, but at the end of the experiments, initial egg production rates had dropped to 6.6% of initial rates, and viability had declined to zero within 7 days. The two Thalassiosira species mainly affected egg viability, so that after 14 days of feeding, a decrease of 90% had occurred. However, egg production rates were high and were reduced only by 15-24% towards the end of the experiments. In terms of potential recruitment rates, even though PRO induced high mean naupliar production, a slight decrease was observed during the first 5 days, likely due to the time required for T. stylifera to acclimate to PRO. After day 5, potential recruitment rate was higher, comparable to in situ values. THA and TWEI inhibited potential recruitment up to 90-100%, but these effects were not evident over a short-term period, denoting the importance of introducing the time factor when evaluating diatom effects on copepod production. With the other two diatom diets, SKE and PHA, recruitment was zero after 2 and 6 days, respectively. Experiments testing the effects of unsaturated aldehydes in THA (2-trans-4-trans-decadienal) and SKE (2-trans-4-trans-octadienal and 2-trans-4-trans-heptadienal) on the hatching viability of T. stylifera indicate that diatom-derived aldehydes may differ in terms of biological activity. Of the three molecules tested, decadienal had somewhat stronger effects on hatching success, with total blockage occurring at concentrations of 2.0 μg ml⁻¹; the concentrations to induce total blockage by octadienal and heptadienal were somewhat higher and ≥2.5 μg ml⁻¹. We suggest that the greater biological activity of SKE in terms of cell blockage may thus be due to the presence of antimitotic compounds other than these unsaturated aldehydes.     

Chemical characterization of new oxylipins from Cestrum parqui, and their effects on seed germination and early seedling growth

Isolation, chemical characterization, and phytotoxicity of five new oxylipins, together with seven already known related compounds, from Cestrum parqui L' Hérl. is reported. All the structures were elucidated on the basis of their spectral data, especially 1D-(1H- and 13C-NMR, DEPT) and 2D-NMR (COSY, TOCSY, HSQC, HMBC, and NOESY). The configurations of the stereogenic C-atoms were determined by the Mosher's method. The compounds have been assayed for their phytotoxicity on Lactuca sativa at concentrations ranging between 10(-4) and 10(-8) M. The results of the phytotoxicity tests on the germination and growth of the test species, obtained by a cluster analysis, showed interesting relationship between the chemical structures of the compounds and their biological effects.     

Effect of culture density and antioxidants on naupliar production and gene expression of the cyclopoid copepod, Paracyclopina nana

Although attempts have been made to use mass cultures of marine copepods as live foods in marine aquaculture, some limitations such as low density culture still exist. The brackish water cyclopoid copepod, Paracyclopina nana has the potential for mass culturing as live food. In this study, we not only investigated the effect of culture density on the naupliar production and specific gene expressions of P. nana, but also the effect of several antioxidants under the conditions of a high density culture. The naupliar production of the copepod decreased with increasing culture density. The expression of glutathione reductase (GR), selenium-dependent glutathione peroxidase (SeGPx), glutathione S-transferase kappa (GST kappa), heat shock protein 40 (Hsp40), and Hsp70 genes of P. nana increased in the high density treatment but vitellogenin genes (Vg1 and Vg2) showed downregulation. In the condition with 20 inds./mL, vitamin C had a significant decrease but sodium selenite induced the naupliar production of P. nana greatly. The expressions of GR, SeGPx, Hsp70, and Vg genes increased with the vitamin C treatment. Sodium selenite caused a decrease of SeGPx and Hsp40 but GST kappa increased in the treatment with 20 inds./mL. These results suggest that sodium selenite is a positive antioxidant which can increase the culture efficiency of the copepod.     

Long-term effects of dietary isoflavones on uterine gene expression profiles

Isoflavones (ISOs) are bioactive food ingredients of the traditional East Asian diet and currently discussed as alternatives to classical hormone replacement therapies and for reducing the prevalence of hormone-dependent cancers. Although there are many studies on ISOs, not much is known about their long-term effects.Therefore, we performed an animal experiment analyzing the effects of three different diets: a phytoestrogen-free diet, a diet supplemented with genistein (700 μg/g diet) and an ISO-high diet (232 μg daidzein and 240 μg genistein/g) at two distinct time points, juvenile (21 days) and adult (97 days). Exposure started prior to mating of the parents and throughout the life of the offspring.We observed a stronger increase of uterine wet weights in juvenile offspring with genistein exposure (1018 ± 350 mg/kg BW) than with ISO-high diet (497 ± 133 mg/kg BW). Whereas the expression of proliferation related genes (PCNA; Ki67; IGF-1; IGF-1R), analyzed by real-time-qPCR and Western blot, were significantly down-regulated in juvenile animals exposed to genistein. Additionally, genistein exposure led to estrogenic responses, observed upon increase of complement C3 and decrease of estrogen receptors gene expressions, while the exposure to ISO-high diet did not show these effects.In conclusion, both the time point on which phytoestrogen exposure starts together with the composition of the ingested phytoestrogen containing diet are of great importance for the biological response of the offspring.     

Separation of chitooligosaccharides and the potent effects on gene expression of cell surface receptor CR3

Chitooligosaccharides were prepared through hydrolysis of colloidal chitosan by enzyme source from Aspergillus fumigatus BSF114. Chitosan pentamer (COS5) and chitosan hexamer (COS6) were isolated and purified from COS by the ultra-filtration, nano-filtration, ethanol precipitation and the CM-Sephadex C-25 column. COS5 consisted of (GlcN)₄ (59.84%) and (GLcN)₅ (40.16%). COS6, however, mainly consisted of (GLcN)₆ (93.11%) and (GLcN)₅ (6.89%). Effects of COS5 and COS6 in vivo and in vitro on gene expression of cell surface CR3 receptor were investigated by relatively quantitative RT-PCR and ELISA. The results showed that the expression of CR3 mRNA could be promoted by both COS5 and COS6. The promotion effect caused by COS6 was greater than that of COS5.     

Endometrial effects of selective estrogen receptor modulators (SERMs) on estradiol-responsive gene expression are gene and cell-specific

Three selective estrogen receptor modulator (SERM) drugs which included 4-OH-tamoxifen (Tam), EM-800 (EM) and GW 5638 (GW) were investigated to determine their ability to inhibit estradiol-responsive gene expression in sheep endometrium. The uteri of ovariectomized ewes (10 ewes per SERM group) were infused with 10⁻⁷ M SERMs for 24 h prior to hysterectomy. Five ewes from each group received 50 μg 17β-estradiol (E2) and the remaining five ewes received vehicle 18 h prior to hysterectomy. Northern blot analyses and in situ hybridization demonstrated that E2 treatment increased estrogen receptor (ER), progesterone receptor (PR), glyceraldehyde 3-phosphate dehydrogenase (GAPDH), and cyclophilin (CYC) mRNA levels in most endometrial cells examined. Tam and GW exhibited characteristics similar to E2 by increasing ER gene expression, but they antagonized the E2-induced increases in PR and CYC mRNA levels. EM acted as an E2-agonist of GAPDH gene expression, but antagonized the E2 up-regulation of ER, PR and CYC gene expression in most endometrial cells. Immunohistochemistry determined that EM decreased ER protein levels in the glandular epithelium, and the SERMs investigated antagonized increases in PR protein levels in endometrium. In conclusion, GW and EM exhibit fewer agonist effects than Tam on endometrial gene expression. EM demonstrated the greatest antagonism of E2-enhanced levels of ER, PR and CYC, likely due to the inhibition of ER gene expression at both mRNA and protein levels.     

Myometrial effects of selective estrogen receptor modulators on estradiol-responsive gene expression are gene and cell-specific

We examined in vivo effects of selective estrogen receptor modulators (SERMs) 4-OH-tamoxifen (Tam), GW 5638 (GW) and EM-800 (EM) on myometrial gene expression. The uteri of ovariectomized ewes were infused with 10⁻⁷ M of one SERM via indwelling catheters for 24 h preceding hysterectomy. Half of the ewes in each SERM group received an intramuscular injection of 50 μg 17β-estradiol (E2) 18 h prior to hysterectomy. Northern blot analysis and in situ hybridization demonstrated that E2 increased estrogen receptor (ER), progesterone receptor (PR) and cyclophilin (CYC) gene expression in the cells of both inner layer of myometrium (IM) and outer layer of myometrium (OM) as well as glyceraldehyde 3-phosphate dehydrogenase (GAPDH) gene expression in OM. Tam also increased ER mRNA levels in OM. EM appeared to increase ER gene expression, but antagonized E2’s up-regulation of PR and CYC gene expression in both IM and OM. Tam and GW also antagonized E2 up-regulation of PR gene expression in OM but not IM. No SERM affected GAPDH gene expression with or without E2. Immunohistochemistry indicated that E2 increased nuclear ER and PR protein levels in both IM and OM. EM was unique in up-regulating ER protein levels, opposite to its effects in endometrial cells. All SERMs tested antagonized this increase in PR immunostaining preferentially in OM compared to the IM layer. These results illustrate gene and cell layer-specific effects of SERMs in sheep myometrium.     

Description of a fugu CXC chemokine and two CXC receptor genes, and characterization of the effects of different stimulators on their expression

The primary structures of a CXC chemokine (CXCL8) and two CXC receptors (CXCR) have been characterized in fugu, Takifugu rubripes. Unlike mammalian and avian species, CXCL8 of teleosts including fugu lacks the ELR motif that appears to be important in ligand/receptor interactions on neutrophils. Genomic organization shows that fugu CXCL8 gene consists of four exons and three introns. As in other vertebrates, two CXCR genes isolated from fugu encode proteins CXCR1 and CXCR2 that possess characteristic seven transmembrane domains. Each receptor consists of two exons separated by an intron. Synteny analysis indicates that these two CXCRs were derived from whole genome duplication in teleosts, differing from mammalian CXCR1 and CXCR2. All of these genes are primarily expressed in the lymphoid tissues. Immune stimulation with PHA showed that the expression of both CXCL8 and CXCRs in PBL are upregulated even after only a short time period, but downregulated by LPS stimulation, implying that these genes are involved in the regulation of the immune response in fugu.     

Effects of feed restriction during pregnancy on maternal reproductive outcome,foetal hepatic IGF gene expression and offspring performance in the rabbit

Primiparous female rabbits have high nutritional requirements and, while it is recommended that they are subjected to an extensive reproductive rhythm, this could lead to overweight, affecting reproductive outcomes. We hypothesised that restricting food intake during the less energetic period of gestation could improve reproductive outcome without impairing offspring viability. This study compares two groups of primiparous rabbit does in an extensive reproductive programme, one in which feed was restricted from Day 0 to Day 21 of gestation (R021), and another in which does were fed ad libitum (control) throughout pregnancy. The mother and offspring variables compared were (1) mother reproductive outcomes at the time points pre-implantation (Day 3 postartificial insemination [AI]), preterm (Day 28 post-AI) and birth; and (2) the prenatal offspring characteristic IGF system gene expression in foetal liver, liver fibrosis and foetus sex ratio, and postnatal factor viability and growth at birth, and survival and growth until weaning. Feed restriction did not affect the conception rate, embryo survival, or the number of morulae and blastocysts recovered at Day 3 post-AI. Preterm placenta size and efficiency were similar in the two groups. However, both implantation rate (P < 0.001) and the number of foetuses (P = 0.05) were higher in the R021 mothers than controls, while there was no difference in foetal viability. Foetal size and weight, the weights of most organs, organ weight/BW ratios and sex ratio were unaffected by feed restriction; these variables were only affected by uterine position (P < 0.05). Conversely, in the R021 does, foetal liver IGBP1 and IGF2 gene expression were dysregulated despite no liver fibrosis and a normal liver structure. No effects of restricted feed intake were produced on maternal fertility, prolificacy, or offspring birth weight, but control females weaned more kits. Litter weight and mortality rate during the lactation period were also unaffected. In conclusion, pre-implantation events and foetal development were unaffected by feed restriction. While some genes of the foetal hepatic IGF system were dysregulated during pregnancy, liver morphology appeared normal, and the growth of foetuses and kits until weaning was unmodified. This strategy of feed restriction in extensive reproductive rhythms seems to have no significant adverse effects on dam reproductive outcome or offspring growth and viability until weaning.     

Isomer-specific effects of conjugated linoleic acid on gene expression in RAW 264.7

Conjugated linoleic acid (CLA) is a mixture of dietary fatty acids that has various beneficial effects including decreasing cancer, atherosclerosis, diabetes and inflammation in animal models. Some controversy exists on the specific isomers of CLA that are responsible for the benefits observed. This study was conducted to examine how different CLA isomers regulate gene expression in RAW 264.7. A mouse macrophage cell line, RAW 264.7, was treated with five different CLA isomers (9E,11E-, 9Z,11E-, 9Z,11Z-, 10E,12Z- and 11Z,13E-CLA). Gene expression microarrays were performed, and several significantly regulated genes of interest were verified by a real-time polymerase chain reaction (PCR). Examination of the biological functions of various significantly regulated genes by the five CLA isomers showed distinct properties. Isomers 9E,11E-, 9Z,11Z-, 10E,12Z- and 11Z,13E-CLA decreased production of proinflammatory cytokines such as interleukin (IL)-1α, IL-1β and IL-6. Many of CLA's effects are believed to be mediated by the fatty acid receptors such as the peroxisome proliferator-activated receptors (PPAR) and retinoid-X-receptors (RXR). Using PPAR and RXR specific antagonists and coactivator recruitment assays, it was evident that multiple mechanisms were responsible for gene regulation by CLA isomers. Coactivator recruitment by CLA isomers showed their distinct properties as selective receptor modulators for PPARγ and RXRα. These studies demonstrate distinct isomer differences in gene expression by CLA and will have important ramifications for determining the potential therapeutic benefit of these dietary fatty acids in prevention of inflammation-related diseases.     

mRNA levels of SREBP-1c do not coincide with the changes in adipose lipogenic gene expression

Physiological differences in lipid metabolism exist according to adipose sites. To delineate at which step such gene regulation could occur, mRNA levels of various proteins involved in the overall lipogenic process were determined in subcutaneous (SC) and retroperitoneal (RP) adipose tissues. Fatty acid synthase, malic enzyme, ATP citrate lyase, insulin-sensitive glucose transporter, and glucose-6-phosphate dehydrogenase mRNA levels were coordinately reduced (by up to 50-fold) during fasting in RP and in SC relative to fed rats, and restored or overexpressed (by up to 5- to 6-fold) during refeeding. The response was most often delayed and lower in SC compared to RP. This could contribute to site-specific differences. Interestingly, SREBP-1c mRNA levels were markedly decreased by fasting in SC but remained unchanged in RP. Refeeding tended to restore levels close to fed group values. We conclude that mRNA levels of SREBP-1c do not coincide with the expected changes in adipose lipogenic gene expression of fasted/refed rats.