Development of polymorphic microsatellite markers for distylous-homostylous Primula secundiflora (Primulaceae) using HiSeq sequencing.

Primula secundiflora is an insect-pollinated, perennial herb belonging to the section Proliferae (Primulaceae) that exhibits considerable variation in its mating system, with predominantly outcrossing populations comprising long-styled and short-styled floral morphs and selfing populations comprising only homostyles. To facilitate future investigations of the population genetics and mating patterns of this species, we developed 25 microsatellite markers from P. secundiflora using next-generation sequencing and measured polymorphism and genetic diversity in a sample of 30 individuals from three natural populations. The markers displayed high polymorphism, with the number of observed alleles per locus ranging from three to 16 (mean = 8.36). The observed and expected heterozygosities ranged from 0.100 to 1.000 and 0.145 to 0.843, respectively. Twenty-one of the loci were also successfully amplified in P. denticulata. These microsatellite markers should provide powerful tools for investigating patterns of population genetic diversity and the evolutionary relationships between distyly and homostyly in P. secundiflora.

Parallel evolution of morphological and genomic selfing syndromes accompany the breakdown of heterostyly.

Evolutionary transitions from outcrossing to selfing in flowering plants have convergent morphological and genomic signatures and can involve parallel evolution within related lineages. Adaptive evolution of morphological traits is often assumed to evolve faster than nonadaptive features of the genomic selfing syndrome. We investigated phenotypic and genomic changes associated with transitions from distyly to homostyly in the Primula oreodoxa complex. We determined whether the transition to selfing occurred more than once and investigated stages in the evolution of morphological and genomic selfing syndromes using 22 floral traits and both nuclear and plastid genomic data from 25 populations. Two independent transitions were detected representing an earlier and a more recently derived selfing lineage. The older lineage exhibited classic features of the morphological and genomic selfing syndrome. Although features of both selfing syndromes were less developed in the younger selfing lineage, they exhibited parallel development with the older selfing lineage. This finding contrasts with the prediction that some genomic changes should lag behind adaptive changes to morphological traits. Our findings highlight the value of comparative studies on the timing and extent of transitions from outcrossing to selfing between related lineages for investigating the tempo of morphological and molecular evolution.

Environmental cue difference and training duration modulate spatial learning and cue preference in detour task.

In this study, we investigated how different environmental cue and the proficiency of body motion influenced detour learning behaviour and cue preference in cue conflict situations. Domestic chicks were trained to detour around an obstacle and follow a fixed route to rejoin with their partners. When the environmental cue was red versus blue vertical stripes, the chicks learned the detour task quicker, and as the number of training trials after route acquisition increased, they switched their preference from the environmental cue to a body-motion cue in the cue conflict test. On the other hand, when the environmental cue was vertical versus horizontal blue stripes, the chicks learned the detour task slower and showed a dependence on the body-motion cue regardless of the number of training trials performed after route acquisition. When the environmental cue was removed, most chicks could still successfully detour according to the specific route on which they had been trained. Furthermore, a significant difference in detour latency was found between chicks using the environmental cue and chicks using the body-motion cue, suggesting separate neuronal circuits responsible for processing the two types of information. Our results demonstrated that young domestic chicks could use both environmental cue and body-motion cues to memorize the route during the detour learning task; however, the detour route preference could be dynamically modulated by difference of the environmental cue and the number of training trials they received.

The complete chloroplast genome of a distylous-homostylous species, Primula homogama (Primulaceae).

Primula homogama F. H. Chen & C. M. Hu (Primulaceae) is endemic to the Emei Mountain of China. In this study, we characterized the complete chloroplast genome of P. homogama based on next-generation sequencing (NGS). The complete chloroplast genome of P. homogama was 154,677 bp in size with a typical quadripartite structure, containing a large single-copy (LSC) region of 85,299 bp and a small single-copy (SSC) region of 17,816 bp. These two regions were separated by a pair of inverted repeat regions (IRs), each of 25,781 bp. A total of 130 functional genes were encoded, consisted of 86 protein-coding genes (PCG), 36 tRNA genes, and eight ribosomal RNA (rRNA) genes.

Phylogenomic analysis reveals multiple evolutionary origins of selfing from outcrossing in a lineage of heterostylous plants.

Evolutionary transitions from outcrossing to selfing often occur in heterostylous plants. Selfing homostyles originate within distylous populations and frequently evolve to become reproductively isolated species. We investigated this process in 10 species of Primula section Obconicolisteri using phylogenomic approaches and inferred how often homostyly originated from distyly and its consequences for population genetic diversity and floral trait evolution. We estimated phylogenetic relationships and reconstructed character evolution using the whole plastome comprised of 76 protein-coding genes. To investigate mating patterns and genetic diversity we screened 15 microsatellite loci in 40 populations. We compared floral traits among distylous and homostylous populations to determine how phenotypically differentiated homostyles were from their distylous ancestors. Section Obconicolisteri was monophyletic and we estimated multiple independent transitions from distyly to homostyly. High selfing rates characterised homostylous populations and this was associated with reduced genetic diversity. Flower size and pollen production were reduced in homostylous populations, but pollen size was significantly larger in some homostyles than in distylous morphs. Repeated transitions to selfing in section Obconicolisteri are likely to have been fostered by the complex montane environments that species occupy. Unsatisfactory pollinator service is likely to have promoted reproductive assurance in homostyles leading to subsequent population divergence through isolation.

The complete chloroplast genome of an endangered endemic herb species in China, Primula filchnerae (Primulaceae).

Primula filchnerae (Primulaceae) is an endangered endemic herb species in China. In this study, we characterized the complete chloroplast genome of P. filchnerae based on next generation sequencing (NGS). The chloroplast genome of P. filchnerae was 151,547 bp in size, containing a large single-copy (LSC) region of 82, 662 bp and a small single-copy (SSC) region of 17,749 bp. These two regions were separated by a pair of inverted repeat regions (IRs), each of 25,568 bp. A total of 130 functional genes were encoded, consisted of 86 protein-coding genes, 36 tRNA genes, and eight rRNA genes.

Current Perspective in the Discovery of Anti-aging Agents from Natural Products.

Aging is a process characterized by accumulating degenerative damages, resulting in the death of an organism ultimately. The main goal of aging research is to develop therapies that delay age-related diseases in human. Since signaling pathways in aging of Caenorhabditis elegans (C. elegans), fruit flies and mice are evolutionarily conserved, compounds extending lifespan of them by intervening pathways of aging may be useful in treating age-related diseases in human. Natural products have special resource advantage and with few side effect. Recently, many compounds or extracts from natural products slowing aging and extending lifespan have been reported. Here we summarized these compounds or extracts and their mechanisms in increasing longevity of C. elegans or other species, and the prospect in developing anti-aging medicine from natural products.

Otophylloside B Protects Against Aß Toxicity in Caenorhabditis elegans Models of Alzheimer's Disease.

Alzheimer's disease (AD) is a major public health concern worldwide and the few drugs currently available only treat the symptoms. Hence, there is a strong need to find more effective anti-AD agents. Cynanchum otophyllum is a traditional Chinese medicine for treating epilepsy, and otophylloside B (Ot B), isolated from C. otophyllum, is the essential active component. Having previously identified anti-aging effects of Ot B, we evaluated Ot B for AD prevention in C. elegans models of AD and found that Ot B extended lifespan, increased heat stress-resistance, delayed body paralysis, and increased the chemotaxis response. Collectively, these results indicated that Ot B protects against Aß toxicity. Further mechanistic studies revealed that Ot B decreased Aß deposition by decreasing the expression of Aß at the mRNA level. Genetic analyses showed that Ot B mediated its effects by increasing the activity of heat shock transcription factor (HSF) by upregulating the expression of hsf-1 and its target genes, hsp-12.6, hsp-16.2 and hsp-70. Ot B also increased the expression of sod-3 by partially activating DAF-16, while SKN-1 was not essential in Ot B-mediated protection against Aß toxicity.

Differentiation of bel-7402 human hepatocarcinoma cells induced by aqueous extracts of fresh gecko (AG) and its anti-tumor activity in vivo.

ETHNOPHARMACOLOGICAL RELEVANCE: Gecko, a kind of reptile, has been widely used as a traditional Chinese medicine to treat various diseases including cancer in China for thousands of years. The aim of this study was to investigate the anti-tumor effect of AG (aqueous extracts of fresh gecko) on human hepatocellular carcinoma cell Bel-7402 in vitro and mouse H22 hepatocellular in vivo. Further to underlie the molecular mechanism of AG inducing the differentiation of Bel-7402 cells. MATERIALS AND METHODS: AG was obtained by water extracting method and qualitatively analyzed through High Performance Liquid Chromatography. The total protein concentration of AG was measured by BCA (bicinchoninic acid disodium) assay. The anti-tumor activities in vivo were analyzed through H22 (mouse hepatocellular carcinoma cell line H22) tumor xenografts mice. The cytotoxic activity of AG on Bel-7402 cells was evaluated by MTT assays. AFP (alpha fetoprotein) was detected by radioimmunoassay. ALB (albumin), ALP (alkaline phosphatase) and γ-GT (γ-glutamyl transpeptidase) were detected by biochemical methods with commercial kits. While morphological changes were observed through an inverted microscope. Moreover, the expression level of the proteins involved in MAPK (mitogen-activated protein kinase) signal pathway which was closely related to cellular differentiation was assessed by Western blot. RESULTS: AG showed obviously anti-tumor activity in vivo and anti-proliferative activity on Bel-7402 cells in vitro both dose-dependently. The number of clones of Bel-7402 cells treated with AG reduced and the cells were displaying differentiation state such as relatively bigger size and dispersed growth. The biochemical function markers of the cells were significantly changed after being treated with AG. The data showed that AFP secretion of the cells decreased 42.5%, ALB secretion increased 58.9%, the activity of ALP and γ-GT markedly decreased 67.0% and 48.5% separately when the concentration of AG was 10µl/ml, and those effects were all in a dose-dependent manner. The major original and phosphorylated signal proteins (ERK1/2 (extracellular sigal-regualted kinase 1/2), P38 (p38 MAPK) and JNK1/2 (c-Jun N-terminal kinase 1/2)) involved in MAPK signal pathway were measured and the results showed that AG activated the ERK1/2 of Bel-7402 cells. CONCLUSIONS: AG has anti-tumor activity in vivo and inhibits Bel-7402 cell proliferation in vitro through inducing cell differentiation, and the mechanism involves the activation of ERK1/2.

[Establishment of tree shrew chronic morphine dependent model].

The clinical use of morphine to reduce pain is limited because of its drug tolerance, dependence and addiction. In the present study, the tree shrews (Tupaia belangeri chinensis) developed morphine tolerance and chronic morphine dependence by morphine injections with increasing doses (5, 10, 15, 20 mg/kg body weight for 7 days). Meanwhile, the naloxone (1.25 mg/kg body weight)-induced conditioned place aversion (CPA) and the withdrawal symptom were also found. The tree shrew model of chronic morphine dependence can be used to investigate the withdrawal symptoms and to select potential withdrawal symptoms reducing drugs in the future.

[Dynamics of hippocampal sensory gating during the chronic morphine administration, withdrawal and re-exposure to morphine in rats].

Drug addiction is considered as a chronic, recurrent brain disease characterized by relapse. Repeated exposure to certain drugs, such as morphine, can produce deleterious sequelae, such as drug dependence, tolerance and compulsive drug seeking. In the present study, we investigated the dependence and psychological craving for morphine in rats using the conditioned place preference (CPP) paradigm. On the other hand, to study the effect of morphine on hippocampal sensory gating (N40), double click auditory-evoked potential was recorded during the chronic morphine administration, withdrawal and re-exposure to morphine in rats. The rats in morphine group received a course of morphine (10 mg/kg, i.p.) injection for 12 d, followed by 12 d of withdrawal, 1 d of re-exposure to morphine (2.5 mg/kg, i.p.) and 2 d of the second withdrawal. The rats in the control group were treated in the same way except that saline was applied instead of morphine. CPP test demonstrated that the method of drug administration in the present study induced dependence and psychological craving for morphine in rats. The results in the double click auditory-evoked potential experiment showed that during the chronic morphine administration, hippocampal N40 gating was damaged. In the initial 2 d of the first withdrawal hippocampal N40 gating in morphine group was reduced compared with that in the control group and it was significantly greater on the 3rd day, and then recovered gradually to the normal level from day 4 to day 12. After re-exposure to morphine, hippocampal N40 gating in morphine group was significantly reduced compared with that in the control group, and it remained at a lower level during the following 2 d, suggesting that hippocampal N40 gating in rats was more sensitive to morphine during re-exposure. Our results suggest that long-term repeated morphine administration and re-exposure to morphine disrupt hippocampal N40 gating, and that the effect of morphine addiction on the brain is possibly long-term.

[Effects of morphine on the development of chick embryos].

AIM: To investigate the effect of morphine on fetal movement, heart rate, hatch weight, hatch days and hatch rate. METHODS: Morphine was injected into airspace of eggs and fetal movement, heart rate, hatch weight, hatch days and hatch rates were recorded. RESULTS: Hatch days were shorter, hatch rates were lower and some chicks became motor disorder for morphine. Chicks with morphine exposure 20 mg/kg from E 12 to E 16 had highest hatch rate and lowest disable rate. Morphine reduced fetal movement, increased heart rate (P < 0.05). CONCLUSION: The development of chick embryo is impaired by morphine exposure and the magnitude of these effects depends on the drug dose and the length of time that the developing organism is exposed to morphine.