Zebrafish Embryo Model Research Articles

Live imaging reveals a conserved role of fatty acid β-oxidation in early lymphatic development in zebrafish

During embryonic development, lymphatic endothelial cells (LECs) differentiate from venous endothelial cells (VECs), a process that is tightly regulated by several genetic signals. While the aquatic zebrafish model is regularly used for studying lymphangiogenesis and offers the unique advantage of time-lapse video-imaging of lymphatic development, some aspects of lymphatic development in this model differ from those in the mouse. It therefore remained to be determined whether fatty acid β-oxidation (FAO), which we showed to regulate lymphatic formation in the mouse, also co-determines lymphatic development in this aquatic model. Here, we took advantage of the power of the zebrafish embryo model to visualize the earliest steps of lymphatic development through time-lapse video-imaging. By targeting zebrafish isoforms of carnitine palmitoyltransferase 1a (cpt1a), a rate controlling enzyme of FAO, with multiple morpholinos, we demonstrate that reducing CPT1A levels and FAO flux during zebrafish development impairs lymphangiogenic secondary sprouting, the initiation of lymphatic development in the zebrafish trunk, and the formation of the first lymphatic structures. These findings not only show evolutionary conservation of the importance of FAO for lymphatic development, but also suggest a role for FAO in co-regulating the process of VEC-to-LEC differentiation in zebrafish in vivo.

Safety evaluation of the temporary consolidant based on a zebrafish embryo model

Temporary consolidants, such as cyclododecane, menthol, coumarin, and ethyl maltol, are proved to be effective for urgent conservation in archaeological field. However, the safety of these temporary consolidants is still unknown although they are always heavily used in archaeological field. Thus reports on the toxicity of these temporary consolidants are limited. Here, a zebrafish model was used for safety evaluation of these four temporary consolidants.In the present study, four temporary consolidants, including cyclododecane, menthol, coumarin, and ethyl maltol, at different concentrations were incubated with zebrafish embryos, and their biological toxic effects were firstly evaluated.It was shown that embryo exposure to temporary consolidants resulted in an increased mortality and malformation rate, and a decreased hatching rate. The order of embryo toxicity of the four types of temporary consolidants tested was menthol > coumarin > ethyl maltol > cyclododecane. Although the embryo toxicity of cyclododecane was minimal, some studies have indicated that this temporary consolidant could be bioaccumulated. The results also suggested that the zebrafish embryos can serve as a reliable model for the evaluation of embryo toxicity of temporary consolidants, as this model could offer the possibility to perform the rapid, medium throughput, cost-effective analyses.

Bisphenol-A diminishes neutrophil recruitment in zebrafish

Abstract The prevalent use of bisphenol-A (BPA) in food containers, thermal paper, and other products provides an environmental challenge to the immune system of both humans and wildlife as BPA disrupts estrogen activity and alters signaling. We are exploring the impact of in vivo BPA exposure on innate immunity in both mice and zebrafish. Our earlier work in C57BL/6J mice showed that low dose, in vivo BPA exposure significantly decreases LPS-induced expression of CXCL-1, a chemokine that recruits neutrophils to the site of infection and the mouse homolog of human IL-8, in spleen cells. To determine if reduced chemokine secretion has a functional outcome, we expanded our studies to the zebrafish embryo model. Real time PCR analysis indicates that 4 to 6 day post fertilization (dpf) zebrafish exposed to environmentally relevant doses of BPA show reduced background expression of CXCL-8, the zebrafish homolog of mouse CXCL-1, and a reduction in the total number of neutrophils (4 dpf). BPA exposure also reduces the upregulation of CXCL-8 induced by a tail cut injury in 4 dpf zebrafish. MPO::GFP zebrafish expressing GFP under a neutrophil-specific myeloperoxidase promoter were used to track neutrophil movement for six hours following a tail cut injury. 4 dpf zebrafish exposed to BPA show a significantly reduced neutrophil recruitment to the site of injury compared to controls (p=0.004), a finding which correlates with the reduction in injury-induced CXCL-8 expression. Together our findings demonstrate that BPA exposure during early zebrafish development diminishes two key innate immune responses, thus potentially increasing susceptibility to infection.

Targeting oncogenic Ras by the Clostridium perfringens toxin TpeL.

Clostridium perfringens toxin TpeL belongs to the family of large clostridial glycosylating toxins. The toxin causes N-acetylglucosaminylation of Ras proteins at threonine35 thereby inactivating the small GTPases. Here, we show that all main types of oncogenic Ras proteins (H-Ras, K-Ras and N-Ras) are modified by the toxin in vitro and in vivo. Toxin-catalyzed modification of Ras was accompanied by inhibition of the MAP kinase pathway. Importantly, TpeL inhibited the paradoxical activation of the MAP kinase pathway induced by the BRAF inhibitor Vemurafenib in the human melanoma cell line SBCL2. The toxin also blocked Ras signaling in a zebrafish embryo model expressing oncogenic H-RasG12V, resulting in a reduction of melanocyte number. By using the binding and translocation component of anthrax toxin (protective antigen), the glucosyltransferase domain of TpeL was effectively introduced into target cells that were not sensitive to native TpeL toxin. To reach a higher specificity towards cancer cells, a chimeric TpeL toxin was engineered that possessed the knob region of adenovirus serotype 35 fiber, which interacts with CD46 of target cells frequently overexpressed in cancer cells. The chimeric TpeL fusion toxin efficiently inhibited Ras and MAP kinases in human pancreatic cancer Capan-2 cells, which were insensitive to the wild-type toxin. The data reveal that TpeL and TpeL-related immunotoxins provide a new toolset as Ras-inactivating agents.

Antitubercular Bis-Substituted Cyclam Derivatives: Structure-Activity Relationships and in Vivo Studies.

We recently reported the discovery of nontoxic cyclam-derived compounds that are active against drug-resistant Mycobacterium tuberculosis. In this paper we report exploration of the structure-activity relationship for this class of compounds, identifying several simpler compounds with comparable activity. The most promising compound identified, possessing significantly improved water solubility, displayed high levels of bacterial clearance in an in vivo zebrafish embryo model, suggesting this compound series has promise for in vivo treatment of tuberculosis.

The extract of Polygoni Cuspidati Rhizoma et Radix suppresses the vascular endothelial growth factor-induced angiogenesis

BackgroundPolygoni Cuspidati Rhizoma et Radix (PCRR; the root and rhizome of Polygonum cuspidatum Sieb. et Zucc) is a traditional Chinese medicine for the treatment of inflammation, hyperlipemia, favus, jaundice and scald. Hypothesis/PurposeThe extract of PCRR inhibits vascular endothelial growth factor (VEGF)-induced angiogenesis. The hypothesis is supported by analysis of PCRR extract and investigation of pharmacological role and signaling mechanism of PCRR extract in regulating angiogenic responses. Study designThe PCRR ethanolic extract was examined for its inhibitory effects on angiogenesis based on VEGF-treated human umbilical vein endothelial cells and in zebrafish model MethodsThe effects and signaling mechanism of a standardized ethanolic extract of PCRR were tested on cell proliferation, migration and tube formation in VEGF-treated human umbilical vein endothelial cells, and which was further validated in zebrafish embryo model. ResultsThe treatment of PCRR extract in cultured endothelial cells inhibited VEGF-induced cell proliferation, cell migration and tube formation in a dose-dependent manner and also suppressed the formation of sub-intestinal vessels in zebrafish embryos. Moreover, the applied PCRR extract suppressed VEGF-induced phosphorylations of VEGF receptor 2 (VEGFR2) and JNK. Thus, the site of effect triggered by PCRR was proposed to be mediated by VEGFR2. To further support this notion, the phosphorylations of Erk, Akt and eNOS, induced by VEGF, were markedly reduced under the challenge of PCRR extract: the reductions were subsequently further decreased in the present of inhibitors of Erk, Akt and eNOS. In parallel, the formation of ROS induced by VEGF in cultured endothelial cells was markedly reduced in the present of PCRR extract. ConclusionCollectively, our studies demonstrated the pharmacological role and signaling mechanism of PCRR in regulation of angiogenic responses, which supported further evaluation and development of PCRR as a potential therapeutic agent for the treatment and prevention of diseases related with angiogenesis.

Developmental Effects and Estrogenicity of Bisphenol A Alternatives in a Zebrafish Embryo Model.

In order to understand the negative effects of bisphenol A (BPA) alternatives comprehensively, zebrafish embryos were used to assess the lethality, developmental effects, and estrogenic activity of bisphenol analogues. The in silico estrogenic activities of bisphenol analogues were assayed by binding simulation. According to our results, the lethality of bisphenol analogues decreased in order of bisphenol AF (BPAF) > BPA > bisphenol F (BPF) > bisphenol S (BPS). BPAF and BPF induced significant effects on zebrafish embryos, including decreased heart rate, hatching inhibition, and teratogenic effects. The binding potentials of bisphenol analogues toward zebrafish ERs (zfERS) decreased in the following order: BPAF > BPA > BPF > BPS. Among the three subtypes of zfERs, zfERβ2 showed the highest binding activity toward the bisphenols, followed by zfERα and zfERβ1. In vivo estrogenic activity tests showed that BPAF, BPA, and BPF significantly enhanced the protein levels of ERα along with the mRNA levels of esr1, esr2a, esr2b, and vtg1 in zebrafish embryos. Esr2b showed the strongest response to BPAF and BPA exposure among the three esrs. In contrast, BPS did not significantly regulate ER protein level or ER transcription. In conclusion, BPAF showed the highest lethality, developmental effects, and estrogenic activity (both in silico and in vivo) followed by BPA and BPF. BPS showed the weakest toxicity and estrogenic activity. zfERβ2 might act as the main target among the three ER subtypes of zebrafish after exposure to BPAF and BPA.

The MRPS18-2 protein levels correlate with prostate tumor progression and it induces CXCR4-dependent migration of cancer cells

We have earlier found abnormal expression of the mitochondrial ribosomal protein S18-2 (MRPS18-2, S18-2) in endometrial cancer, compared to the expression in hyperplasia and in normal endometrium. Here we report that expression of S18-2 was increased with disease progression in clinical specimens of prostate cancer (PCa). The level of induction of epithelial to mesenchymal cell transition (EMT) correlated with the expression level of S18-2 in PCa cell lines. Moreover, cells acquired increased ability of migration upon S18-2 overexpression, as was evaluated in zebrafish embryo model and in trans-well assay. We found that this is due to increased CXCR4 cell surface expression. Neutralizing CXCR4 protein or abrogating S18-2 expression in cells significantly reduced their migratory ability directed toward CXCL12. The mRNA expression of TWIST2, encoding one of transcription factors that induce EMT upon CXCR4 increase, positively correlated with the S18-2 protein level. Together, these data suggest that the S18-2 protein induces EMT through the TWIST2/E-cadherin signalling and, consequently, CXCR4-mediated migration of PCa cells.

Carbazole alkaloids with antiangiogenic activities from Clausena sanki

Two new carbazole alkaloids 1 and 2, and eleven known congeners 3–13 were isolated and identified from Clausena sanki for the first time. Their structures were elucidated on the basis of extensive UV, IR, MS, NMR spectroscopic data and comparison with literatures. The compounds 1–13 were evaluated by MTT assay to determine whether they decreased VEGF-mediated cell proliferation in HUVECs with Axitinib as positive control. Among them, compounds 1, 2, 6, 8, and 13 (μM) exhibited moderate antiangiogenic activities, which inhibited VEGF-induced HUVEC proliferation in vitro with IC50 values of 12.1 (C.I. 8.2–15.2), 58.1 (C.I. 56.3–63.4), 13.7 (C.I. 9.2–15.4), 16.0 (C.I. 9.5–16.4), and 63.2 (C.I. 57.8–65.7) μM, respectively. Moreover, the antiangiogenic activities of compounds 1–13 were evidenced in vivo in the zebrafish embryo model. As a result, compounds 1, 2, 6, 8, and 13 showed effectively suppress angiogenesis. These research results may guide the search for new natural products with antiangiogenic attributes.

Lower-Molecular-Weight Chitosan-Treated Polyethyleneimine: a Practical Strategy For Gene Delivery to Mesenchymal Stem Cells

Background/Aims: Genetic modification of mesenchymal stem cells (MSCs) is an essential requirement for their use as a delivery vehicle. To achieve higher transfection efficiency and better reproducibility than previously synthesized chitosan (100 kDa)-polyethylenimine (PEI; 1200 Da), we synthesized a low molecular weight PEI (1200 Da)-grafted chitosan (50 kDa) (CP). Methods: Safety of CP/DNA or PEI (25 kDa)/DNA was evaluated by an MTT assay using A549 cells or MSCs and a zebrafish embryo model. Effects of CP/DNA on the characteristics of MSCs were evaluated using flow cytometry. Additionally, a pGL3 plasmid was used to investigate the transfection efficiency of PEI (25 kDa), chitosan (100 kDa)-PEI (1200 Da), and CP with different N/P mass ratios on A549 cells and MSCs. Furthermore, CP/pGL3 was used to investigate the effect of serum on transfection, and intracellular transport was assessed by observing the intracellular location of DNA using laser scanning confocal microscopy. In addition, the effect of endocytosis on transfection efficiency was evaluated using A549 cells pre-treated with different inhibitors. Investigations related to analysis of transfection efficiency were all performed using the BCA protein assay to standardize the data. Furthermore, TGF-β1-and CXCR4-expressing plasmids were applied to evaluate the gene transfer efficiency of CP, including its effects on the osteogenic differentiation and migratory ability of MSCs. Results: The safety evaluation demonstrated that CP/DNA had significantly lower toxicity than PEI (25 kDa)/DNA. Additionally, DNA entered MSCs transfected by CP without changing their properties, while the examination of intracellular transport demonstrated that CP/pGL3 was internalized rapidly into MSCs. Furthermore, studies of the internalization mechanism showed that CP/pGL3 complexes entered the cells through caveolae-mediated endocytosis, thereby suggesting that the CP coating helped DNA enter A549 cells without the requirement for receptors. Compared to PEI (25 kDa), the interference of serum on transfection was reduced significantly with the use of CP in both A549 cells and MSCs. To evaluate the effects of gene delivery using the constructed CP complex and the possibility of obtaining gene-engineered MSCs, TGF-β1- and CXCR4-expressing plasmids were successfully delivered into MSCs, confirming their ability to induce osteogenesis and change the migratory ability of MSCs, respectively. Conclusion: These results demonstrated that CP could be used to deliver genes into MSCs and could potentially be used in gene therapy based on MSCs.

MiR-378 and MiR-1827 Regulate Tumor Invasion, Migration and Angiogenesis in Human Lung Adenocarcinoma by Targeting RBX1 and CRKL, Respectively.

MicroRNAs (miRNAs) have been extensively studied over the decades and have been proposed as potential molecular targets for cancer treatment. Studies have shown that miR-378 participates in numerous biological processes in various cancers; whereas miR-1827 has only been reported in pediatric glioma. The mechanism of how miRNAs modulate lung cancer metastasis remains unclear. Our previous study demonstrated that miR-378 is up-regulated while miR-1827 is down-regulated in high invasive lung cancer sub-cell lines, and their biological functions have been described. Here, we report that miR-378 and miR-1827 modulate lung cancer cell invasion and migration via epithelial-mesenchymal transition (EMT). We also demonstrated that cells treated with miR-378 inhibitors or miR-1827 mimics had reduced number of metastases and ectopic vessels in the zebrafish embryo model. We then showed that miR-378 promoted invasion and miR-1827 suppressed migration by targeting RBX1 and CRKL, respectively. Restored protein expression in miRNA-overexpressed/ miRNA-suppressed cells attenuated the inhibitory/ inducing effect of the miRNA on lung cancer cells. Collectively, our findings highlight that miR-378 and miR-1827 could serve as novel therapeutic targets in lung cancer.

Ecotoxicological assessment of Ti3C2Tx(MXene) using a zebrafish embryo model

We studied the biocompatibility of Ti3C2TxMXene sheets by analyzing their potential toxicityin vivoand their potential risks on marine biota and ecosystems using a zebrafish embryo model.

The Ability of Different Ketohexoses to Alter Apo-A-I Structure and Function In Vitro and to Induce Hepatosteatosis, Oxidative Stress, and Impaired Plasma Lipid Profile in Hyperlipidemic Zebrafish.

In the current study, we have tested the nonenzymatic glycation activities of ketohexoses, such as tagatose and psicose. Although tagatose-treated apoA-I (t-A-I) and psicose-treated apoA-I (p-A-I) exerted more inhibitory activity you cupric ion-mediated low-density lipoprotein (LDL) oxidation and oxidized LDL (oxLDL) phagocytosis into macrophage than fructose-treated apoA-I (f-A-I). In the lipid-free state, t-A-I and f-A-I showed more multimerized band without crosslinking. Since t-A-I lost its phospholipid binding ability, the rHDL formation was not as successful as f-A-I. However, injecting t-A-I showed more antioxidant activities in zebrafish embryo under the presence of oxLDL. Three weeks of consumption of fructose (50% of wt in Tetrabit/4% cholesterol) showed a 14% elevation of serum triacylglycerol (TG), while tagatose-administered group showed 30% reduction in serum TG compared to high cholesterol control. Fructose-fed group showed the biggest area of Oil Red O staining with the intensity as strong as the HCD control. However, tagatose-consumed group showed much lesser Oil Red O-stained area with the reduction of lipid accumulation. In conclusion, although tagatose treatment caused modification of apoA-I, the functional loss was not as much severe as the fructose treatment in macrophage cell model, zebrafish embryo, and hypercholesterolemic zebrafish model.

Francisella noatunensis subspecies noatunensis clpB deletion mutant impairs development of francisellosis in a zebrafish model

BackgroundFrancisella noatunensis ssp. noatunensis (F.n.n.) is the causative agent of francisellosis in Atlantic cod and constitutes one of the main challenges for future aquaculture on this species. A facultative intracellular bacterium like F.n.n. exert an immunologic challenge against which live attenuated vaccines in general are most effective. Thus, we constructed a deletion in the F.n.n. clpB gene as ΔclpB mutants are among the most promising vaccine candidates in human pathogenic Francisella. PurposeCharacterization of F.n.n. ΔclpB using primary Atlantic cod head kidney leukocytes, the zebrafish embryo and adult zebrafish model with focus on potential attenuation, relevant immune responses and immunogenic potential. Main resultsInterleukin 1 beta transcription in Atlantic cod leukocytes was significantly elevated from 24 to 96 h post infection with F.n.n. ΔclpB compared to F.n.n. wild-type (wt). Growth attenuation of the deletion mutant in zebrafish embryos was observed by fluorescence microscopy and confirmed by genome quantification by qPCR. In the immunization experiment, adult zebrafish were immunized with 7 × 106 CFU of F.n.n. ΔclpB before challenge four weeks later with 6 × 108 CFU of F.n.n. wt. One day after challenge, immunized zebrafish responded with significantly lower interleukin 8 levels compared to the non-immunized control. Immunized fish were protected against the acute mortality observed in non-immunized zebrafish after challenge and bacterial genomes quantified by qPCR were reduced to a minimum 28 days post challenge, indicating protective immunity stimulated by F.n.n. ΔclpB. ConclusionDeletion mutation of clpB in F.n.n. causes in vitro and in vivo attenuation and elicits a protective immune response in adult zebrafish against a lethal dose of F.n.n. wt. Taken together, the results presented increases the knowledge on protective immune responses against F.n.n.

β-Cyclodextrin Attenuates Perfluorooctanoic Acid Toxicity in the Zebrafish Embryo Model.

Perfluorooctanoic acid (PFOA) has been linked to negative health outcomes including cancer, thyroid disease, infertility, and developmental delays. β-Cyclodextrin (β-CD), a cyclic sugar, has been previously shown to form strong host–guest complexes with PFOA, and is proposed as a means of environmental remediation with respect to this widespread contaminant. In the present study, β-CD was directly examined with regards to possible attenuation of the toxicity of PFOA specifically employing the zebrafish (Danio rerio) embryo model. Zebrafish embryos were exposed to various concentrations of PFOA without β-CD, and with equimolar (1:1) and excess (2:1) molar ratios of β-CD to PFOA, and assessed for lethality and developmental toxicity through seven days post-fertilization (dpf). Rapid onset of lethality with limited morphological abnormalities was observed at relatively low concentrations of PFOA (LC50 ≈ 50 ppm), along with effects on morphometric and neurobehavioral parameters in surviving embryos. A highly significant difference (p < 0.0001) was observed between the 2:1 treatment, and both 1:1 and PFOA only treatments, with respect to lethal concentration and apparent neurobehavioral effects, suggesting an effectively reduced toxicity of the fully complexed PFOA. In contrast, however, neither β-CD treatment reduced developmental toxicity with respect to the morphometric endpoint (i.e., interocular distance). Whereas LC50 of PFOA alone did not change over 7 dpf, the 1:1 and 2:1 values decreased slightly over time, suggesting either delayed or alternative toxic effects on later developmental stages at presumptively lowered levels. This study, therefore, indicates β-CD may be an effective agent to reduce toxicity of and mitigate environmental health concerns associated with PFOA, but that further study is required to elucidate the mechanism of complexation as it relates to the attenuation of toxicity.

In vitro and in vivo anti-inflammatory activities of high molecular weight sulfated polysaccharide; containing fucose separated from Sargassum horneri: Short communication

Recent studies on crude and pure compounds from Sargassum horneri have shown promising bioactive properties. However, anti-inflammatory potentials of fucose-rich sulfated polysaccharides from S. horneri have not yet been discovered. In the present study, we evaluated the in vitro and in vivo anti-inflammatory activities of four polysaccharide fractions separated from membrane filters according to their molecular weights (<5kDa (f1), 5–10kDa (f2), 10–30kDa (f3), and >30kDa (f4)). According to the results, F4 fraction inhibited the lipopolysaccharides (LPS)-induced nitric oxide (NO) (IC50=87.12μg/mL) and prostaglandin E2 production as well as pro-inflammatory cytokine production in RAW 264.7 cells through down-regulating nuclear factor-κB signaling cascade. According to the results, f4 has a potential to down-regulate LPS-induced toxicity, cell death and NO production levels in LPS-induced in vivo zebrafish embryo model. These results suggest that f4 fraction has the potential to develop functional materials or drugs to treat inflammatory diseases.

Zebrafish In-Vivo Screening for Compounds Amplifying Hematopoietic Stem and Progenitor Cells: - Preclinical Validation in Human CD34+ Stem and Progenitor Cells

The identification of small molecules that either increase the number and/or enhance the activity of human hematopoietic stem and progenitor cells (hHSPCs) during ex vivo expansion remains challenging. We used an unbiased in vivo chemical screen in a transgenic (c-myb:EGFP) zebrafish embryo model and identified histone deacetylase inhibitors (HDACIs), particularly valproic acid (VPA), as significant enhancers of the number of phenotypic HSPCs, both in vivo and during ex vivo expansion. The long-term functionality of these expanded hHSPCs was verified in a xenotransplantation model with NSG mice. Interestingly, VPA increased CD34+ cell adhesion to primary mesenchymal stromal cells and reduced their in vitro chemokine-mediated migration capacity. In line with this, VPA-treated human CD34+ cells showed reduced homing and early engraftment in a xenograft transplant model, but retained their long-term engraftment potential in vivo, and maintained their differentiation ability both in vitro and in vivo. In summary, our data demonstrate that certain HDACIs lead to a net expansion of hHSPCs with retained long-term engraftment potential and could be further explored as candidate compounds to amplify ex-vivo engineered peripheral blood stem cells.

P62/SQSTM1 interacts with vimentin to enhance breast cancer metastasis.

The signalling adaptor p62 is frequently overexpressed in numerous cancer types. Here, we found that p62 expression was elevated in metastatic breast cancer and its overexpression correlated with reduced metastasis- and relapse-free survival times. Analysis of p62 expression in breast cancer cell lines demonstrated that high p62 expression was associated with the invasive phenotypes of breast cancer. Indeed, silencing p62 expression attenuated the invasive phenotypes of highly metastatic cells, whereas overexpressing p62 promoted the invasion of non-metastatic cells in in vitro microfluidic model. Moreover, MDA-MB-231 cells with p62 depletion which were grown in a three-dimensional culture system exhibited a loss of invasive protrusions. Consistently, genetic ablation of p62 suppressed breast cancer metastasis in both zebrafish embryo and immunodeficient mouse models, as well as decreased tumourigenicity in vivo. To explore the molecular mechanism by which p62 promotes breast cancer invasion, we performed a co-immunoprecipitation-mass spectrometry analysis and revealed that p62 interacted with vimentin, which mediated the function of p62 in promoting breast cancer invasion. Vimentin protein expression was downregulated upon p62 suppression and upregulated with p62 overexpression in breast cancer cells. Linear regression analysis of clinical breast cancer specimens showed a positive correlation between p62 and vimentin protein expression. Together, our findings provide strong evidence that p62 functions as a tumour metastasis promoter by binding vimentin and promoting its expression. This finding might help to develop novel molecular therapeutic strategies for breast cancer metastasis treatment.

Identification of sterols from the soft coral Dendronephthya gigantea and their anti-inflammatory potential

Exploration of anti-inflammatory phytochemicals has received tremendous attention worldwide owing to the rapid increase in inflammatory diseases. Current study reveals the identification of eight 3β-hydroxy-Δ5-steroidal congeners from a nonpolar column fraction of the ethanol solubles from the soft coral Dendronephthya gigantea collected from Jeju Island South Korea, using GC–MS/MS analysis. The sterol-rich fraction (DGEH21) showed a significant anti-inflammatory activity as exhibited by the inhibition of NO production (IC50 4.33±0.50μg/mL) and PGE2 production in LPS-stimulated RAW 264.7 macrophages. It also suppressed the expression levels of proinflammatory cytokines, TNF-α, IL-1β, and IL-6 in a dose-dependent manner. Furthermore, DGEH21 effectively downregulated the expression levels of iNOS, and COX-2 and reduced NO and ROS production as well as cell death in LPS-stimulated in-vivo zebrafish embryo model. However, DGEH21 at relatively high concentrations indicated cytotoxicity in both RAW cells and zebrafish embryos with RAW cell viability being nearly 80% after treatment with 25μg/mL DGEH21. This study highlights the synergistic anti-inflammatory activity of several steroids found in D. gigantea. Their actions may be useful in the development of anti-inflammatory cosmeceuticals, pharmaceutical agents, and other consumer products.

The novel autophagy inhibitor elaiophylin exerts antitumor activity against multiple myeloma with mutant TP53 in part through endoplasmic reticulum stress-induced apoptosis

ABSTRACTElaiophylin is a natural compound and a novel and potent inhibitor of late stage autophagy with outstanding antitumor activity in human ovarian cancer cells. However, the possible biologic effects and functional linkage between elaiophylin and multiple myeloma (MM) have not been explored. This study aimed to assess the effect of elaiophylin on MM cells with mutant TP53 and the possible molecular mechanism. The results suggested that elaiophylin exerted anti-myeloma activity by inducing apoptosis and proliferation arrest. As expected, elaiophylin blocked autophagy flux in MM cells. Subsequently, persistent activation of endoplasmic reticulum (ER) stress was induced. Moreover, the apoptotic effect was to some extent attenuated by the ER stress inhibitor tauroursodeoxycholic acid (TUDCA). Further studies indicated that elaiophylin effectively suppressed MM cell growth without obvious side effects in zebrafish embryo and mouse xenograft models. Taken together, our data are the first to demonstrate that exposure of human MM cells with mutant TP53 to elaiophylin blocked autophagy flux and thus induced cell death, which partially involved ER stress-associated apoptosis. Targeted disruption of the cellular protein handling system by elaiophylin is therefore a promising therapeutic strategy for overcoming incurable MM, even when TP53 mutations are present.