RNAi-mediated Knockdown of Xist Does Not Rescue the Impaired Development of Female Cloned Mouse Embryos

In mice, one of the major epigenetic errors associated with somatic cell nuclear transfer (SCNT) is ectopic expression of Xist during the preimplantation period in both sexes. We found that this aberrant Xist expression could be impeded by deletion of Xist from the putative active X chromosome in donor cells. In male clones, it was also found that prior injection of Xist-specific siRNA could significantly improve the postimplantation development of cloned embryos as a result of a significant repression of Xist at the morula stage. In this study, we examined whether the same knockdown strategy could work as well in female SCNT-derived embryos. Embryos were reconstructed with cumulus cell nuclei and injected with Xist-specific siRNA at 6–7 h after oocyte activation. RNA FISH analysis revealed that siRNA treatment successfully repressed Xist RNA at the morula stage, as shown by the significant decrease in the number of cloud-type Xist signals in the blastomere nuclei. However, blastomeres with different sizes (from “pinpoint” to “cloud”) and numbers of Xist RNA signals remained within single embryos. After implantation, the dysregulated Xist expression was normalized autonomously, as in male clones, to a state of monoallelic expression in both embryonic and extraembryonic tissues. However, at term there was no significant improvement in the survival of the siRNA-injected cloned embryos. Thus, siRNA injection was largely effective in repressing the Xist overexpression in female cloned embryos but failed to rescue them, probably because of an inability to mimic consistent monoallelic Xist expression in these embryos. This could only be achieved in female embryos by applying a gene knockout strategy rather than an siRNA approach.     

Investigating pig survival in different production phases using genomic models

Pig survival is an economically important trait with relevant social welfare implications, thus standing out as an important selection criterion for the current pig farming system. We aimed to estimate (co)variance components for survival in different production phases in a crossbred pig population as well as to investigate the benefit of including genomic information through single-step genomic best linear unbiased prediction (ssGBLUP) on the prediction accuracy of survival traits compared with results from traditional BLUP. Individual survival records on, at most, 64,894 crossbred piglets were evaluated under two multi-trait threshold models. The first model included farrowing, lactation, and combined postweaning survival, whereas the second model included nursery and finishing survival. Direct and maternal breeding values were estimated using BLUP and ssGBLUP methods. Furthermore, prediction accuracy, bias, and dispersion were accessed using the linear regression validation method. Direct heritability estimates for survival in all studied phases were low (from 0.02 to 0.08). Survival in preweaning phases (farrowing and lactation) was controlled by the dam and piglet additive genetic effects, although the maternal side was more important. Postweaning phases (nursery, finishing, and the combination of both) showed the same or higher direct heritabilities compared with preweaning phases. The genetic correlations between survival traits within preweaning and postweaning phases were favorable and strong, but correlations between preweaning and postweaning phases were moderate. The prediction accuracy of survival traits was low, although it increased by including genomic information through ssGBLUP compared with the prediction accuracy from BLUP. Direct and maternal breeding values were similarly accurate with BLUP, but direct breeding values benefited more from genomic information. Overall, a slight increase in bias was observed when genomic information was included, whereas dispersion of breeding values was greatly reduced. Combined postweaning survival presented higher direct heritability than in the preweaning phases and the highest prediction accuracy among all evaluated production phases, therefore standing out as a candidate trait for improving survival. Survival is a complex trait with low heritability; however, important genetic gains can still be obtained, especially under a genomic prediction framework.     

Cryopreservation of male and female gonial cells by vitrification in the critically endangered cyprinid honmoroko <Emphasis Type="Italic">Gnathopogon caerulescens</Emphasis>

We investigated the feasibility of cryopreservation of spermatogonia and oogonia in the critically endangered cyprinid honmoroko Gnathopogon caerulescens using slow-cooling (freezing) and rapid-cooling (vitrification) methods. Initially, we examined the testicular cell toxicities and glass-forming properties of the five cryoprotectants: ethylene glycol (EG), glycerol (GC), dimethyl sulfoxide (DMSO), propylene glycol (PG), and 1,3-butylene glycol (BG), and we determined cryoprotectant concentrations that are suitable for freezing and vitrification solutions, respectively. Subsequently, we prepared the freezing solutions of EG, GC, DMSO, PG, and BG at 3, 2, 3, 2, and 2 M and vitrification solutions at 7, 6, 5, 5, and 4 M, respectively. Following the cryopreservation of the testicular cells mainly containing early-stage spermatogenic cells (e.g., spermatogonia and primary spermatocytes), cells were cultured for 7 days and immunochemically stained against germ cell marker protein Vasa. Areas occupied by Vasa-positive cells indicated that vitrification led to better survival of germ cells than the freezing method, and the best result was obtained with 5 M PG, about 50% recovery of germ cells following vitrification. In the case of ovarian cells containing oogonia and stage I, II, and IIIa oocytes, vitrification with 5 M DMSO resulted the best survival of oogonia, with equivalent cell numbers to those cultured without vitrification. The present data suggest that male and female gonial cells of the endangered species G. caerulescens can be efficiently cryopreserved using suitable cryoprotectants for spermatogonia and oogonia, respectively.     

Symbiotic bacterium modifies aphid body color

Color variation within populations of the pea aphid influences relative susceptibility to predators and parasites. We have discovered that infection with a facultative endosymbiont of the genus Rickettsiella changes the insects' body color from red to green in natural populations. Approximately 8% of pea aphids collected in Western Europe carried the Rickettsiella infection. The infection increased amounts of blue-green polycyclic quinones, whereas it had less of an effect on yellow-red carotenoid pigments. The effect of the endosymbiont on body color is expected to influence prey-predator interactions, as well as interactions with other endosymbionts.     

Fucoxanthinol,Metabolite of Fucoxanthin,Improves Obesity-Induced Inflammation in Adipocyte Cells

Fucoxanthin (Fx) is a marine carotenoid found in edible brown seaweeds. We previously reported that dietary Fx metabolite into fucoxanthinol (FxOH), attenuates the weight gain of white adipose tissue of diabetic/obese KK-A^y mice. In this study, to evaluate anti-diabetic effects of Fx, we investigated improving the effect of insulin resistance on the diabetic model of KK-A^y mice. Furthermore, preventing the effect of FxOH on low-grade chronic inflammation related to oxidative stress was evaluated on 3T3-L1 adipocyte cells and a RAW264.7 macrophage cell co-culture system. A diet containing 0.1% Fx was fed to diabetic model KK-A^y mice for three weeks, then glucose tolerance was observed. Fx diet significantly improved glucose tolerance compared with the control diet group. In in vitro studies, FxOH showed suppressed tumor necrosis factor-α (TNF-α), and monocyte chemotactic protein-1 (MCP-1) mRNA expression and protein levels in a co-culture of adipocyte and macrophage cells. These findings suggest that Fx ameliorates glucose tolerance in the diabetic model mice. Furthermore, FxOH, a metabolite of Fx, suppresses low-grade chronic inflammation in adipocyte cells.     

Macrolactone Nuiapolide,Isolated from a Hawaiian Marine Cyanobacterium,Exhibits Anti-Chemotactic Activity

A new bioactive macrolactone, nuiapolide (1) was identified from a marine cyanobacterium collected off the coast of Niihau, near Lehua Rock. The natural product exhibits anti-chemotactic activity at concentrations as low as 1.3 μM against Jurkat cells, cancerous T lymphocytes, and induces a G2/M phase cell cycle shift. Structural characterization of the natural product revealed the compound to be a 40-membered macrolactone with nine hydroxyl functional groups and a rare tert-butyl carbinol residue.     

An investigation of the relationship between duct system and A cell-rich and PP cell-rich pancreatic islets in the canine pancreas.

The pancreata of four six-month-old dogs of the same mother, two with both the pancreatic and accessory pancreatic ducts (X-type) and two with only the accessory pancreatic duct (Y-type), were examined in this study. To clarify the relationships between the type of pancreatic duct system and the composition of pancreatic endocrine cells, the pancreata were examined immunohistochemically using antiserum against four types of pancreatic hormones (glucagon, insulin, somatostatin and pancreatic polypeptide). In all areas of the X- and Y-type duct system pancreata, B cells accounted for 52-82% of the total number of islet cells, and D cells accounted for 4-15%. In the X-type ducts system, the percentages of A and PP cells in the right and left lobes of the pancreas differed greatly. It was found that A and PP cells appear in inverse proportion to each other and that there exist A cell-rich and PP cell-rich pancreatic islets. The A cell-rich pancreatic islets appeared in the left lobes along the accessory pancreatic duct, while the PP cell-rich pancreatic islets were observed in the right lobes along the pancreatic duct. The body of the pancreas contained both A cell-rich and PP cell-rich pancreatic islets. In the Y-type duct systems, A cell-rich pancreatic islets appeared in the right lobes. These findings indicate that the composition of A and PP cells in pancreatic islets is closely related to the type of duct system.     

Fructose inhibits pear pollen germination on agar medium without loss of viability

Pollen of Japanese pear (Pyrus pyrifolia Nakai) germinated well on agar medium containing 10% sucrose or glucose, but not on agar containing fructose. The inhibitory effect of fructose was dose-dependent. Sucrose enhanced pollen tube growth much more effectively than glucose. Addition of 5% fructose to 5% or 10% sucrose or glucose media suppressed germination completely. Ungerminated pollen, however, showed similar respiration rate and stainability against acetocarmine dye as germinated pollen. When pollen was transferred onto fructose medium after culturing it on glucose or sucrose medium for 1–2 h, germination was completely impeded. Reversely, pollen transferred to sucrose or glucose medium from fructose medium germinated at almost the same ratio as pollen on sucrose or glucose medium without transfer. Thus, pollen inhibition by fructose is reversible. Compared with uncultured pollen, cultured pollen contained less than half amount of total sugars, even if failed in germination on fructose medium. Germinated pollen on sucrose and glucose media contained sucrose and glucose, but ungerminated pollen on fructose medium contained only trace levels of these sugars, suggests that pollen on fructose medium predominantly uses sucrose and glucose as respiration substrates and cannot maintain the constant levels of these sugars. However, as pollen germination occurred on agar medium without any sugar, fructose may impede a physiological factor that triggers germination, and once the trigger is impeded, many physiological pathways including sugar biosynthesis may be blocked.     

梨花柱S糖蛋白对离体花粉萌发及花粉管生长的影响

应用CM和SP琼脂糖凝胶 (Sepharose)离子交换层析法从‘新水’梨花柱可溶性蛋白质中分离纯化出具有RNase活性的S糖蛋白 ,将其加入花粉培养基 3～ 12h ,新水花粉 (自交不亲和 )的萌发率及花粉管生长受到强烈抑制 ,当浓度为 1.2 5μg·μL- 1时 ,分别为对照的 6 5%和 38% ,低浓度 (0 .4 μg·μL- 1)抑制作用不明显 ;与新水杂交不亲和的‘幸水’花粉也出现同样的结果 ;而与其杂交亲和的‘长十郎’花粉萌发率不受影响 ,仅花粉管生长受到轻微的抑制 ,其长度为对照的 79%。不含S糖蛋白的花柱可溶性蛋白质对自花及异花的花粉萌发及花粉管生长均无影响。     

Effect of distance from early flowering pollinizers ‘Maypole’ and ‘Dolgo’ on ‘Fiji’ fruit set

An apple orchard consisting of a single cultivar under the condition of natural pollination must have suitable pollinizers such as Crab apples to ensure stable fruit production. We selected ‘Maypole’ and ‘Dolgo’ as pollinizers for the cultivar ‘Fuji’, and investigated the rate of fruit and seeds in ‘Fuji’ fruits produced by pollen of the pollinizers. We developed a method for tracing pollen flow based on the leaf color of progeny and S-RNase allele of ‘Maypole’, and on Simple Sequence Repeat (SSR) analyses of ‘Maypole’ and ‘Dolgo’. These were powerful tools for determining the distance insects (mainly Osmia cornifrons) carry pollen from the pollinizers to ‘Fuji’. Although the fruit set of ‘Fuji’ apples was not reduced with increasing distance between ‘Fuji’ apple trees and pollinizers (probably due to pollen flow from other commercial cultivars planted outside the area), the rate of ‘Fuji’ apple fruit produced by the pollen of the pollinizers decreased with increasing distance. The rate of fruit produced by the pollinizers was 84% and 77% when ‘Fuji’ was 2.5 m from ‘Maypole’ and ‘Dolgo’, respectively, and 71% and 64% when ‘Fuji’ was 5 m from ‘Maypole’ and ‘Dolgo’, respectively, but was reduced to 47% and 39% when ‘Fuji’ was 10 m from ‘Maypole’ and ‘Dolgo’, respectively. However, the spacing could cause reduced fruit size, and require extra fruit thinning for producing large fruits. It could also cause reduction of the yield for increasing the planting area of pollinizers. We recommend that pollinizers should be planted not more than 10 m from ‘Fuji’.