Controllable mixed-charged co-assembly of dendritic lipopeptides into invisible capsid-like nanoparticles as potential drug carriers.

The feature of invisibility is vital in drug nanocarriers for prolonging blood transportation, with this generating excellent resistance to protein adsorption and clearance from the body. In this work, we report a well-designed molecular and supramolecular strategy for precisely developing mixed-charged nanoparticles with resistance to protein adsorption. We constructed anionic dendritic lipopeptides (ADLs) and cationic dendritic lipopeptides (CDLs) with eight carboxyl or amino groups as terminal groups. By regulating the molar ratio between ADLs and CDLs, amphiphilic dendritic lipopeptides were assembled into nanoparticles (NPs) with adjustable surface charge. Notably, the co-assembly of equivalent amounts of ADLs and CDLs generated neutral mixed-charged NPs as invisible capsid-like NPs (ICNPs). ICNPs were able to resist protein adsorption and serve as stealth nanocarriers for harboring guest molecules.

Transcriptional responses and mechanisms of copper nanoparticle toxicology on zebrafish embryos.

Copper nanoparticles (CuNPs) are used widely due to their attractive antimicrobial properties. However, their biosafety and kinetics on vertebrate embryogenesis are still limited. In this study, CuNPs were revealed to induce eye hypoplasia and almost no digestive gut in zebrafish embryos in a dose-dependent manner. Then, transcriptional responses of zebrafish embryos to CuNPs were investigated, and it was revealed that the genes related to wound healing and stimulus responses were up-regulated, but the genes associated with phototransduction and metabolisms were down-regulated. Differentially expressed genes (DEGs) in CuNPs-exposed and Cu2+-exposed embryos were compared further. Increased VEGF signaling and expression of fli1 were observed in CuNPs rather than Cu2+ treated embryos, but increased reactive oxygen species (ROS) and the resulting enhanced hemoglobin were observed in both CuNPs and Cu2+ treated embryos. This study for the first time revealed that CuNPs and Cu2+ both down-regulated the genes related to phototransduction and metabolisms, but up-regulated the genes associated with hemoglobin. Additionally, compared with Cu2+, CuNPs might be more effective in elevating blood vessels in embryos. Our results suggest that the biological effects of CuNPs are organogenesis-specific during fish embryogenesis, and both particles and ions might mediate their biological effects on embryogenesis.

Eaf1 and Eaf2 mediate zebrafish dorsal-ventral axis patterning via suppressing Wnt/ß-Catenin activity.

During early vertebrate embryogenesis, maternal Wnt/ß-catenin signaling is thought to locally initiate expression of dorsal-specific genes. Here, eaf1 and eaf2 were identified as important maternal and zygotic modulators of Wnt signaling to initiate and specify ventral genes. Expression of ventral ved, vent, and vox was all obviously enhanced in either maternal or zygotic eaf1/2 morphants, and in both eaf1 heterozygous and homozygous mutants, but their expression was suppressed in embryos with over-expression of eaf1/2. Additionally, eaf1/2 were revealed to suppress ventral fates in embryos via Wnt/ß-catenin1/Tcf signaling, complimentary to their roles in suppressing dorsal fates via Wnt/ß-catenin2 signaling. Moreover, eaf1/2 were also revealed to obviously suppress the expression of axin2 induced by ß-catenin2 rather than by ß-catenin1, and the dorsal expression of axin2 in embryos was obviously suppressed by ectopic expression of eaf1/2. This study uncovers a novel dorsal-ventral patterning pathway, with eaf1 and eaf2 inhibiting ventral cells via suppressing Wnt/ß-catenin1/Tcf signaling and inducing dorsal cells indirectly via suppressing ß-catenin2-induced-axin2 on the dorsal side of embryos.

Silver nanoparticles impair zebrafish skeletal and cardiac myofibrillogenesis and sarcomere formation.

Metal nanoparticles from industries contaminate the environment and affect the normal development of fish even human health. However, little is known about their biological effects on fish embryogenesis and the potential mechanisms. In this study, zebrafish embryos exposed to/injected with silver nanopaticles (AgNPs) exhibited shorter body, reduced heartbeats, and dysfunctional movements. Less, loose, and unassembled myofibrils were observed in AgNPs-treated embryos, and genes in myofibrillogenesis and sarcomere formation were found to be down-regulated in treated embryos. Down-regulated calcium (Ca2+) signaling and loci-specific DNA methylation in specific muscle genes, such as bves, shroom1, and arpc1a, occurred in AgNPs-treated embryos, which might result in the down-regulated expression of myofibrillogenesis genes and muscle dysfunctions in the treated embryos. Our results for the first time reveal that through down-regulating Ca2+ signaling and myogenic loci-specific DNA methylation in zebrafish embryos, AgNPs might induce defects of myofibril assembly and sarcomere formation via their particles mostly, which may subsequently cause heartbeat reduction and behavior dysfunctions.

Transcriptional factors Eaf1/2 inhibit endoderm and mesoderm formation via suppressing TGF-ß signaling.

Eaf family genes act in multiple cellular responses such as tumor suppression and embryonic development. In our previous work, Eaf1/2 was found to modulate convergence and extension (C&E) movements and pattern the embryonic anterior-posterior axis during zebrafish embryogenesis. Here, we found that loss-of-function of eaf1/2 caused expanded mesoderm and endoderm in zebrafish embryos and led to the recovery of endoderm specification in TGF-ß factor-mzoeptz257 mutants, while gain-of-function of eaf1/2 induced reduced mesoderm and endoderm. Analyses of gene expression profiles in Eaf deleted or over-expressed mammalian cells indicated that the roles of Eaf1 and Eaf2 in inhibiting TGF-ß signals were conserved from fish to mammals. By taking advantages of TGF-ß reporters, eaf1/2-fused engrailed proteins, and P53M214K mutant, we revealed that Eaf1 and Eaf2 might suppress TGF-ß transduction by synergistically inhibiting none-P53 and P53-required TGF-ß signaling. Furthermore, Eaf1/2 might co-localize and interact with TGF-ß transcriptional factors in the transcriptional complex as repressors to target and suppress TGF-ß signaling activity. Our study unveiled a previously unrecognized link of Eaf1/2 genes with TGF-ß and P53 in vertebrates and demonstrated a conservation of TGF-ß suppression activity for Eaf1/2 family genes from fish to mammals, which might shed some light on the molecular mechanistic basis of Eaf1 and Eaf2 in tumor suppression.

Mechanisms of silver_nanoparticles induced hypopigmentation in embryonic zebrafish.

Silver_nanoparticles (AgNPs) have been reported to inhibit specification of erythroid cells and to induce spinal cord deformities and cardiac arrhythmia in vertebrates, but have not been implicated in development of neural crest (NC) and pigment cells in an in vivo model yet. In current study, down-regulated expressions of NC genes pax7 and foxd3, melanophore genes mitfa and dct, and xanthophore gene gch2 in AgNPs-exposed embryos were revealed by microarray, qRT-PCR and whole-mount in situ hybridization (WISH). Then, the down-regulated expressions of melanophore genes mitfa and dct but not xanthophore gene gch2 in AgNPs-exposed embryos were found to be recovered by melanogenesis agonists palmitic acid and dibutyryl cyclic AMP (dbcAMP). Finally, Ag+ chelating and AgNPs coating compound l-cysteine was found to neutralize AgNPs-induced hypopigmentation in AgNPs-exposed embryos, and to recover the down-regulated expressions of both dct and gch2 to nearly normal level in embryos, suggesting that AgNPs-releasing Ag+ might mediate their biological effects on zebrafish pigmentation mostly. This study was firstly to unveil that AgNPs might specifically act up-stream of mitfa and pax7 genes to suppress specification and differentiation of melanophore and xanthophore lineages respectively by their releasing Ag+ during vertebrate embryogenesis.

Silver_ nanoparticles inhibited erythrogenesis during zebrafish embryogenesis.

Silver_ nanoparticles (AgNPs), for their attractive antimicrobial properties, have become one of the most commercial nanomaterials used recently. AgNPs are reported to be toxic to blood cells of aquatic organisms and humans, however, few studies related to toxic effects of AgNPs in hematopoiesis using an in vivo model were available. Firstly, microarrays were applied to reveal transcriptional responses of zebrafish embryos to AgNPs at 24h post-fertilization (hpf)in this study, and hemoglobin genes were found to be down-regulated by AgNPs and to be enriched in the top 10 categories by Gene Ontology (GO) analysis. The reduced expressions of hemoglobin were further demonstrated by qRT-PCR detection, whole-mount in situ hybridization, and O-dianisidine staining at transcriptional and translational level. Next, the commitment of mesoderm, specification of hematopoietic progenitor cells and differentiation of erythroids were detected at different developmental stages in AgNPs-exposed embryos, and erythrogenesis were found to be inhibited by AgNPs in developmental-stage-specific and cell-specific manners. Finally, it was pointed out that AgNPs affected erythrogenesis mostly by their particles other than their releasing ions.