A fluorescent nanoprobe for real-time monitoring of intracellular singlet oxygen during photodynamic therapy.

Sensing of intracellular singlet oxygen (1O2) is required in order to optimize photodynamic therapy (PDT). An optical nanoprobe is reported here for the optical determination of intracellular 1O2. The probe consists of a porous particle core doped with the commercial 1O2 probe 1,3-diphenylisobenzofuran (DPBF) and a layer of poly-L-lysine. The nanoparticle probes have a particle size of ~80 nm in diameter, exhibit good biocompatibility, improved photostability and high sensitivity for 1O2 in both absorbance (peak at 420 nm) and fluorescence (with excitation/emission peaks at 405/458 nm). Nanoprobes doped with 20% of DPBF are best suited even though they suffer from concentration quenching of fluorescence. In comparison with the commercial fluorescent 1O2 probe SOSG, 20%-doped DPBF-NPs (aged) shows higher sensitivity for 1O2 generated at an early stage. The best nanoprobes were used to real-time monitor the PDT-triggered generation of 1O2 inside live cells, and the generation rate is found to depend on the supply of intracellular oxygen. Graphical abstract A fluorescent nanoprobe featured with refined selectivity and improved sensitivity towards 1O2 was prepared from the absorption-based probe DBPF and used to real-time monitoring of the generation of intracellular 1O2 produced during PDT.

Selective impairments of alerting and executive control in HIV-infected patients: evidence from attention network test.

BACKGROUND: Attention ability can be subdivided into three functionally independent networks, i.e., alerting network, orienting network, and executive network. Previous literature has documented that deficits in attention are a common consequence of HIV infection. However, the precise nature of deficits of attention in HIV-infected patients is poorly understood. Accordingly, the aim of the study was to identify whether the HIV-infected patients showed a specific attention network deficit or a general attentional impairment. METHODS: We investigated 27 HIV-infected patients and 31 normal controls with the Attention Network Test (ANT). RESULTS: The patients exhibited less efficient alerting network and executive network than controls. No significant difference was found in orienting network effect between groups. Our results also indicate a tendency for poorer efficiency on alerting attention and executive attention in patients with CD4 ≤ 200. CONCLUSIONS: Our findings suggest that HIV-infected patients exhibited selective impairments of attention network of alerting and executive control. The link between lower CD4 T cell count and poorer attention network function imply the importance of starting antiretroviral therapy earlier to avoid irreversible neurocognitive impairment.

The role of the Pax1/9 gene in the early development of amphioxus pharyngeal gill slits.

The pharynx is a major characteristic of chordates. Compared with vertebrates, amphioxus has an advantage for the study of pharynx development, as embryos lack neural crest, and the pharynx is mainly derived from endoderm cells. The Pax1/9 subfamily genes have essential roles in vertebrate pharyngeal patterning, but it is not known if the Pax1/9 gene has similar functions in amphioxus pharynx development. To answer this question, we examined the Pax1/9 gene expression pattern in amphioxus embryos at different developmental stages, and observed morphological changes following Pax1/9 knockdown. RT-qPCR analysis indicated that Pax1/9 expression was initiated during early neurula stage and rapidly peaked during mid-neurula stage. Furthermore, in situ hybridization analysis showed that Pax1/9 transcripts were localized exclusively in the most endodermal region of the developing pharynx in early neurula stage embryos; however, Pax1/9 expression was strikingly down-regulated in the region where gill slits would form from the fusion of endoderm and ectoderm in subsequent developmental stages and was maintained in the border regions between adjacent gill slits. Knockdown of Pax1/9 function using both morpholino and siRNA approaches led to embryonic defects in the first three gill slits, and fusion of the first two gill slits. Moreover, the expression levels of the pharyngeal marker genes Six1/2 and Tbx1/10 were reduced in Pax1/9 knockdown embryos. From these observations, we concluded that the Pax1/9 gene has an important role in the initial differentiation of amphioxus pharyngeal endoderm and in the formation of gill slits, most likely via modulation of Six1/2 and Tbx1/10 expression.

A simple method for selecting spawning-ready individuals out from laboratorial cultured amphioxus population.

Amphioxus is an emerging model organism for evolutionary developmental (Evo-Dev) studies owing to its key phylogenetic position in chordates. However, the rare supply of living embryonic materials is a major drawback for using amphioxus as a laboratorial model animal. Although the problem has been partially resolved in several recent reports, the spawning of amphioxus still remains unpredictable to some extent. In the present study, we reported an accurate method to distinguish spawning-ready and non-spawning-ready individuals of amphioxus Branchiostoma belcheri. In comparison with non-spawning-ready amphioxus, all spawning-ready individuals display following features several hours before their spawning: 1) for both males and females, the interstices between two adjacent gonads are obvious and relatively wide; and 2) the connections among eggs are loose and the crannies appear in each individual ovary of females. These morphological features were also observed in B. japonicum, indicating their conservation among different lancelet species. Based on this observable criterion, we made predictions on the spawning of about 600 ripe B. belcheri individuals and acquired an accuracy of 86.7% for females and 80.4% for males. In addition, we found that advancing or delaying onset of darkness has no detectable effect on the timing of spawning of B. belcheri. Our study makes amphioxus spawning more amenable for our experiments and will greatly facilitate its utilization as a laboratorial model animal.

Demonstration of multi-wavelength tunable fiber lasers based on a digital micromirror device processor.

Based on a digital micromirror device (DMD) processor as the multi-wavelength narrow-band tunable filter, we demonstrate a multi-port tunable fiber laser through experiments. The key property of this laser is that any lasing wavelength channel from any arbitrary output port can be switched independently over the whole C-band, which is only driven by single DMD chip flexibly. All outputs display an excellent tuning capacity and high consistency in the whole C-band with a 0.02 nm linewidth, 0.055 nm wavelength tuning step, and side-mode suppression ratio greater than 60 dB. Due to the automatic power control and polarization design, the power uniformity of output lasers is less than 0.008 dB and the wavelength fluctuation is below 0.02 nm within 2 h at room temperature.

Generating amphioxus Hedgehog knockout mutants and phenotype analysis.

The amphioxus is a promising animal model for evolutionary-developmental studies due to its key position on the animal phylogenetic tree. In the present study, we reported a genetically modified amphioxus strain on the Hedgehog (Hh) gene locus using the TALEN method. The result showed that our TALEN pair injection could bring about 34% mutations in the amphioxus Hh coding region. Further analysis on the F(0) gametic DNA revealed that the mutations had entered into gametes. So, we paired one F(0) male carrying an 8 bp deletion with a wild-type (WT) female, and carefully nursed the F(1) embryos up to adulthood. We then screened F(1) individually via analyzing their genomic DNA from a tiny tail tip, and obtained eight heterozygous mutants from the F(1) offspring. Moreover, our observation on the F(2) embryos generated by mating F(1) mutants also revealed that about 25% of early larvae developed aberrantly with head and tail curving ventrally, agenesis of the mesoblastic tissue under their anterior notochord, and no mouth opening. With the larva growth, deformities (such as twist of head and tail, mouth absent, ventrally localized endostyle and gill slits) became more severe, and eventually those malformed larvae died due to no food intake. Genetic analysis showed that all these deformed embryos were homozygous mutants and the ratio of Hh hetorozygotes vs WT agreed with Mondel's law. WT amphioxus larvae are asymmetric with the mouth on the left and gill slits on the right side. However, the homozygous mutant larvae became left-right symmetric with the gill slits on the ventral side, indicating a conserved role of Hedgehog signaling in establishing the left-right embryonic axis.

Functional analysis of the promoter region of amphioxus ß-actin gene: a useful tool for driving gene expression in vivo.

Amphioxus is a promising new animal model for developmental biology. To develop molecular tools for this model, we characterized the promoter region of a cytoplasmic ß-actin gene (Bb-actin-6-2) from the Chinese amphioxus Branchiostoma belcheri. In situ hybridization and real time-quantitative PCR analyses showed that this gene is expressed in many tissues throughout embryonic development. Cloning of cDNA revealed two isoforms with distinct transcription start sites. Isoform #1 exhibits a similar exon/intron and regulatory element organization to that of vertebrate ß-actin, whereas isoform #2 lacks the first exon of isoform #1 and recruits its first intron as a promoter. The activities of upstream promoter regions in the two isoforms were examined using the lacZ reporter system in amphioxus embryos. The proximal promoter of isoform #1 drove reporter gene expression broadly in 58.6 % of injected embryos. That of isoform #2 exhibited much higher activity (91.5 %) than that of isoform #1 or the human EF-1-α gene (38.2 %). We determined the minimal promoter regions of the two isoforms via functional analysis. These two regions, alone or inserted a random DNA fragment upstream, had no detectable activity, but when an upstream enhancer was inserted, the promoters directed reporter gene expression in 61.0 and 93.8 %, respectively, of injected embryos in a tissue-specific manner. Our study not only provides insight into the regulatory mechanism underlying amphioxus Bb-actin-6-2 gene expression, but also identifies two sets of efficient proximal and minimal promoters. These promoters could be used to construct gene expression vectors for transgenic studies using amphioxus as a model.

Complete mitochondrial genome of Epigonichthys cultellus (Cephalochordata: Branchiostomatidae).

The systematic position of the amphioxus species with dextral gonads distributed in the southeast Pacific and Indian Oceans has remained to be clarified due to the adoption of different names by different researchers. Mitochondrial (mt) DNA is generally considered to be a powerful molecular marker in taxonomic studies. For a reliable systematics of the amphioxi collected from the South China Sea, we sequenced the complete mtDNA from a single specimen and compared it with those of the other eight amphioxus species. The present mtDNA genome contains 13 protein-coding genes, 22 tRNA genes, and two rRNA genes, with the same gene order as those in Branchiostoma and Epigonichthys, which, however, is different from that in Asymmetron. Based on our morphological data (including measurements of some characters) and the features of the mt genome, together with the distribution records of the dextral-gonad amphioxi, we conclude that the present mtDNA sequence most likely represents that of E. cultellus.

[A survey of the quality of life among 740 men after surgical sterilization].

OBJECTIVE: To evaluate the life quality of men after surgical sterilization. METHODS: We investigated the incidence of psychosomatic reactions, their influential factors and the quality of life among 740 men after surgical sterilization. We also evaluated the effectiveness of mental intervention for those with psychosomatic reactions. RESULTS: Totally, 59 (8%) of the subjects were not satisfied with their life status, including 42 (5.67%) suffering from psychosomatic reactions, and their psychosomatic reactions were related to their education background, economic status and understanding of sterilization. After 2 - 24 months of mental intervention, 34 (80.95%) of the 42 cases were cured, 6 (14.29%) relieved, and 2 (4.76%) unimproved. CONCLUSION: Mental intervention is effective for psychosomatic reactions in men after surgical sterilization.

An efficient microinjection method for unfertilized eggs of Asian amphioxus Branchiostoma belcheri.

Amphioxus is a promising model animal for evolutionary and developmental studies. However, as an emerging model organism, amphioxus lacks most molecular techniques applied in other well-developed model animals. Microinjection is a powerful technique for gene manipulation, and thus it undoubtedly is one of useful approaches in the studies of gene function and embryonic development. Although the method has been exploited in Florida and European amphioxus, it still remains to be optimized and introduced into other amphioxus species. In order to introduce the technique into our lab, we followed and optimized the previous description and successfully performed microinjection on unfertilized eggs of Asian amphioxus Branchiostoma belcheri. We made six solutions for practice: 200 mM KCl, 100 ng/µl actin-LacZ or 100 ng/µl actin-RFP vectors, LacZ or RFP vectors without promoter and RFP capped mRNA. More than 99.2 % of eggs injected with KCl were able to be fertilized, 94.3 % of them could hatch normally and 55.9 % survived until 2-day larvae, all of which were nearly equivalent to those obtained from normally fertilized eggs. Embryos injected with two plasmid constructs also showed very high fertilizing and hatching ratios, but normally developing ratios were slightly lower than that of KCl injection. Of those injected embryos, 91.8 % expressed exogenous gene LacZ and 80.5 % exhibited foreign RFP expression, which were driven by a promoter from amphioxus ß-actin gene. The data indicated a successful modified microinjection method for the unfertilized eggs of Asian amphioxus, and those modifications improved the feasibility and efficiency of microinjection on amphioxus.

[Research progress of conserved non-coding elements in metazoan].

In addition to protein coding sequence, the organism genome contains a significant amount of regulatory DNA. Comparative genomics reveals that the organism genomes of vertebrates, tunicate, cephalochordate, flies, and nematodes contain cis-regulatory elements with highly conserved non-coding elements (CNEs). CNEs that cluster around trans-dev genes are part of core gene regulatory networks (GRNs), and usually, they can act as transcriptional enhancers. In this review, we described the identification of CNEs and summarized their key properties across the metazoans, and then discussed the evolution of CNEs after large-scale genome duplication events and the role of CNEs in the evolution of ani-mal body plan.

Diffraction of digital micromirror device gratings and its effect on properties of tunable fiber lasers.

A digital micromirror device (DMD) is a kind of widely used spatial light modulator. We apply DMD as wavelength selector in tunable fiber lasers. Based on the two-dimensional diffraction theory, the diffraction of DMD and its effect on properties of fiber laser parameters are analyzed in detail. The theoretical results show that the diffraction efficiency is strongly dependent upon the angle of incident light and the pixel spacing of DMD. Compared with the other models of DMDs, the 0.55 in. DMD grating is an approximate blazed state in our configuration, which makes most of the diffracted radiation concentrated into one order. It is therefore a better choice to improve the stability and reliability of tunable fiber laser systems.

[Wnt signaling pathway and the Evo-Devo of deuterostome axis].

A series of signal transduction pathways have been found to regulate the polarity establishment and formation of animal primary body axis. Among them, Wnt signaling pathway is extremely conserved and several key components in the pathway have been identified in the demosponge lineage. This implies that it is one of the earliest pathways involved in the ancestral metazoan axis development and might play an important role in specification and development of posterior and ventral fate of animal axis. Recently, with the establishment of functional experiments in vitro, the body plan formation has been found to be affected, in varying degrees, by many genes in the Wnt signaling pathway, such as members of wnt gene family, maternal gene beta-catenin and some transcription factor encoding genes. In this review, we analyzed the evolutionary origin of the wnt gene family involved in development of metazoan body plans, and then made a brief review on the roles of canonical Wnt/beta-catenin signaling in the polarity establishment and formation of primary body axis in diverse deuterostomes including sea urchin, amphioxus, zebrafish, frog, and mouse.

Status and causes of soil compaction at apple orchards in the Weibei Dry Highland, Northwest China.

To find out the recessive factors of soil degradation threatening the healthy development of Weibei apple orchards, we examined soil compaction status and its inducing factors. This study could provide a theoretical basis for apple orchard scientific management. We evalua-ted the changes of soil bulk density and compaction with the depth of soil layer in 0-60 cm in four apple orchards with various planting period, including <10 years (4-6 years), 10-20 years (14-16 years) and ＞20 years (24-26 years). The location and degradation degree of soil compaction in orchard were investigated. Through analyzing the number of soil aggregates and the stability, soil clay and organic matter contents, we tried to find the reasons for the internal compacting of soil in Weibei orchards. The results showed that soil bulk density and compactness in the 0-60 cm soil layer significantly increased with increasing planting years and soil depth. With the 20 cm soil layer as a boundary, soil of Weibei orchards in different planting years showed obvious variation characteristics of loose in upper and compact in lower. The above indicators in soil layer above 20 cm basically met the normal requirements of apple trees, whereas soil layer below 20 cm exceeded the threshold for healthy growth of apple trees. The main reasons for soil compaction below the subsurface layer were poor soil aggregation, the lack of soil organic matter, less human disturbance during fruit planting, and the movement of scattered clay particles to the lower layer. With increasing years of fruit planting, soil compaction became more severe.

Characterization and expression of AmphiBMP3 /3b gene in amphioxus Branchiostoma japonicum.

Bone morphogenetic proteins (BMPs) are responsible for regulating embryo development and tissue homeostasis beyond osteogenesis. However, the precise biological roles of BMP3 and BMP3b remain obscure to a certain extent. In the present study, we cloned an orthologous gene (AmphiBMP3/3b) from amphioxus (Branchiostoma japonicum) and found its exon/intron organization is highly conserved. Further, in situ hybridization revealed that the gene was strongly expressed in the dorsal neural plate of the embryos. The gene also appeared in Hatschek's left diverticulum, neural tube, preoral ciliated pit and gill slit of larvae, and adult tissues including ovary, neural tube and notochordal sheath. Additionally, real-time quantitative polymerase chain reaction (RTqPCR) analysis revealed that the expression displayed two peaks at gastrula and juvenile stages. These results indicated that AmphiBMP3/3b, a sole orthologue of vertebrate BMP3 and BMP3b, might antagonize ventralizing BMP2 orthologous signaling in embryonic development, play a role in the evolutionary precursors of adenohypophysis, as well as act in female ovary physiology in adult.

[Construction of genome BAC library for single Branchiostoma belcheri individual].

As one of the closest living invertebrate relatives of vertebrates, amphioxus (subphylum Cephalochordata) occupies a key position in animal evolution and is becoming the best available proxy and model animal for studying the last common ancestor of all chordates, especially vertebrates. As long-term continuous culturing of amphioxus in laboratory became reliable, for pushing this animal to be a more successful model system, whole-genome sequencing of one or more species derived from this branch will be another urgent issue needed to address. In the present study, we described the construction and characterization of a bacterial artificial chromosome (BAC) library, using a single individual of Chinese amphioxus (Branchiostoma belcheri). High quality genomic DNA extracted from the spermary was partially digested with EcoRand EcoRmethylase. Desirable DNA fragments were isolated by pulsed field gel electrophoresis (PFGE), ligated to linearized and phosphorylased carrier pCC1BAC, and then transformed to Escherichia coli EPI300. The constructed library consists of 44 706 clones with the average insert fragment size around 80 kb as estimated by PFGE. The representation of the library is about 9 equivalents to the amhioxus genome. These results indicate that the BAC library will be useful for functional genomic studies and facilitate the whole-genome sequencing of Chinese amphioxus.

Phylogeography of the rice frog, Fejervarya multistriata (Anura: Ranidae), from China based on mtDNA D-loop sequences.

The rice frog, Fejervarya multistriata, is an amphibian widely distributed in China. In this study, we sampled the species across its distributional area in China and sequenced the mtDNA D-loop to investigate the genetic diversity and geographical pattern of the frog population. The results revealed 38 haplotypes in the population, with K2P values varying from 0.19% to 4.22%. Both a phylogenetic analysis and a nested clade analysis (NCA) detected two geographically isolated lineages respectively distributed around the Yangtze drainage (Yangtze lineage) and the south of China (southern lineage). NCA inferred a contiguous range expansion within the Yangtze lineage and allopatric fragmentation within the southern lineage, which might be partly due to the limited samples from this lineage. Accordingly, Fu's Fs test also indicated a population expansion after glacial movement. Therefore, we assumed that the species history responding to glacial events shaped the present population pattern of F. multistriata on the Chinese mainland.

In situ silica coating-directed synthesis of orthorhombic methylammonium lead bromide perovskite quantum dots with high stability.

Luminescent perovskite quantum dots (QDs) had attracted great attention by virtue of the merits of color-tunable and narrow-band emissions. However, sofar reported perovskite QDs suffered from instability more or less. In this work, a type of silica-coated orthorhombic CH3NH3PbBr3 QDs (SiO2-QDs) with greatly improved stability was reported. The SiO2-QDs were one-pot synthesized using a reprecipitation-encapsulation method assisted with an amine functional silane, which not conly controlled the crystallization of QDs, but also encapsulated QDs with a silica layer simultaneously. More interestingly, the in situ encapsulation of silica shell induced the presence of orthorhombic perovskite that was thought to be unstable at room temperature. This is the first report of orthorhombic CH3NH3PbBr3 QDs, as far as we are concerned. The orthorhombic SiO2-QDs exhibited narrow-band green luminescence with a quantum yield of 78%, and a high production yield of ∼70wt%. Moreover, stability of SiO2-QDs was considerably improved due to silica-coating. White-LEDs were also successfully fabricated with the green SiO2-QDs and a red commercial phosphors using a noncontact configuration. These results demonstrated that the orthorhombic SiO2-QDs held great promise for high-performance display or lighting technology.

Ultrastable Luminescent Organic-Inorganic Perovskite Quantum Dots via Surface Engineering: Coordination of Methylammonium Bromide and Covalent Silica Encapsulation.

Encapsulation of luminescent perovskite quantum dots (QDs) into a solid matrix has been approved to be an efficient way to improve their stability. In this work, we reported a green encapsulation method to produce ultrastable CH3NH3PbBr3 QDs incorporated into the SiO2 matrix. Specifically, fresh-prepared CH3NH3PbBr3 QDs were covalently embedded into silica by an aqueous sol-gel method assisted with CH3NH3Br, which not only effectively inhibited the water-driven degradation of QDs through surface coordination, but also strongly stabilized the QDs in solid powder via concentration gradient. As far as we know, this silica encapsulation of perovskite QDs in aqueous environments is reported for the first time. Luminescent properties of perovskite QDs during the course of gelation as well as in resulting composite powder were investigated using steady-state and time-resolved spectroscopies, and a 2 wt % QD-doped sample treated with 11.5 mM of CH3NH3Br was demonstrated to be the optimal phosphor. The green-emissive phosphor had a PLQY of 60.3% and a full width at half maxima of ∼25 nm, exhibiting ultrahigh stability tested by cycle heating (120 °C), continuous heating (80 °C, 60 h), and light irradiation (450 nm light, 350 h). The phosphor was readily blended with polymers and applied as a color-converting layer on blue light-emitting diodes.

Origin and evolution of vertebrate ABCA genes: a story from amphioxus.

Previous studies showed that the vertebrate ABCA subfamily, one subgroup of the ATP-binding-cassette superfamily, has evolved rapidly in terms of gene duplication and loss. To further uncover the evolutionary history of the ABCA subfamily, we characterized ABCA members of two amphioxus species (Branchiostoma floridae and B. belcheri), the closest living invertebrate relative to vertebrates. Phylogenetic analysis indicated that these two species have the same set of ABCA genes (both containing six members). Five of these genes have clear orthologs in vertebrate, including one cephalochordate-specific duplication and one vertebrate-specific duplication. In addition, it is found that human orthologs of amphioxus ABCA1/4/7 and its neighboring genes mainly localize on chromosome 1, 9, 19 and 5. Considering that most of analyzed amphioxus genes have clear orthologs in zebrafish, we conclude these four human paralogous regions might derive from a common ancestral region by genome duplication occurred prior to teleost/tetrapod split. Therefore, the present results provide new evidence for 2R hypothesis.