Snail Activity Research Articles

Efektivitas Ekstrak Daun Mangkokan (Nothopanax scutellarium Merr) dengan Pemrosesan Berbeda untuk Pengendalian Keong Mas (Pomacea canaliculata Lamarck)

Mangkokan leaves (Nothopanax scutellarium Merr) have been used to control golden snail (Pomacea canaliculata Lamarck) in several preparations. This research aimed to study the most effective preparation of the mangkokan leaf extract to control the golden snail. The research was conducted in the Insect Bioecology Laboratory and agriculture field in Batu Busuak Village, Pauh District, Padang City, from March to April 2021. This study used a Completely Randomized Design (CRD) consisting of five treatments and five replications. The treatments were several preparations of mangkokan leaves with a concentration of 5 g/l: control, fresh, boiled, sun-dried, and oven-dried. The observed variables were mortality of the golden snail, the weight of feed consumed, and the decrease in the eating activity of the golden snail. The result showed that botanical molluscicide from mangkokan leaves preparation by oven-dried and fresh were the two most effective in controlling the golden snail because they caused 100 % mortality of the golden snail, reduced the feed number consumed, with decreasing in feeding activity of 54.91% and 52.94%.

Activation of cell adhesion molecules and Snail during epithelial to mesenchymal transition prior to formation of the regenerative tail blastema in lizards.

After few days from tail amputation in lizards the stump is covered with mesenchymal cells accumulated underneath a wound epidermis and forms a regenerative blastema. During migration, some keratinocytes transit from a compact epidermis into relatively free keratinocytes in a process of "epithelial to mesenchymal transition" (EMT). EMT is also induced after damaging the regenerating epidermis by cauterization, whereas keratinocytes detach and migrate as mesenchymal-like cells among the superficial blastema cells and reconstruct a wound epidermis after about a week from the damage. In normal amputation or after cauterization, no malignant transformation is observed during the transition and migration of keratinocytes. Immunolabeling for markers of EMT confirms the histological description and shows a unique pattern of expression for l-CAM (E-cadherin), N-CAM, and SNAIL-1 and -2 (SLUG). These proteins are present in the cytoplasm and nuclei of migrating keratinocytes. It is hypothesized that the nuclear labeling for E-cadherin coupled to cytoplasmic SNAIL-labeling is somehow related to an initially regulated EMT. After the migrating keratinocytes have reached confluence over the stump, they reverse into a "mesenchymal to epithelial transition" (MET) forming the wound epidermis. The basal layers of the apical wound epidermis of the blastema show some nuclear E-cadherin labeling, while the tail regenerates. It is hypothesized that, together with other tumor suppressors proteins, the apical epidermis and mesenchyme are kept under a tight proliferative control, while in proximal regions the prevalent effect of tumor suppressors determine the differentiation of the new tail tissues.

The identification of alternative oxidase in intermediate host snails of Schistosoma and its potential role in protecting Oncomelania hupensis against niclosamide-induced stress

BackgroundSnail intermediate hosts are mandatory for the transmission of schistosomiasis, which has to date infected more than 200 million people worldwide. Our previous studies showed that niclosamide treatment caused the inhibition of aerobic respiration and oxidative phosphorylation, and the disruption of energy supply, in one of the intermediate hosts of schistosomiasis, Oncomelania hupensis, which eventually led to the death of the snails. Meanwhile, the terminal oxidase in the mitochondrial respiratory chain, alternative oxidase (AOX), was significantly up-regulated, which was thought to counterbalance the oxidative stress and maintain metabolic homeostasis in the snails. The aims of the present study are to identify the AOXs in several species of snails and investigate the potential activation of O. hupensis AOX (OhAOX) under niclosamide-induced stress, leading to enhanced survival of the snail when exposed to this molluscicide.MethodsThe complete complementary DNA was amplified from the AOXs of O. hupensis and three species of Biomphalaria; the sequence characteristics were analysed and the phylogenetics investigated. The dynamic expression and localisation of the AOX gene and protein in O. hupensis under niclosamide-induced stress were examined. In addition, the expression pattern of genes in the mitochondrial respiratory complex was determined and the production of reactive oxygen species (ROS) calculated. Finally, the molluscicidal effect of niclosamide was compared between snails with and without inhibition of AOX activity.ResultsAOXs containing the invertebrate AOX-specific motif NP-[YF]-XPG-[KQE] were identified from four species of snail, which phylogenetically clustered together into Gastropoda AOXs and further into Mollusca AOXs. After niclosamide treatment, the levels of OhAOX messenger RNA (mRNA) and OhAOX protein in the whole snail were 14.8 and 2.6 times those in untreated snails, respectively, but varied widely among tissues. Meanwhile, the level of cytochrome C reductase mRNA showed a significant decrease in the whole snail, and ROS production showed a significant decrease in the liver plus gonad (liver-gonad) of the snails. At 24 h post-treatment, the mortality of snails treated with 0.06–0.1 mg/L niclosamide and AOX inhibitor was 56.31–76.12% higher than that of snails treated with 0.1 mg/L niclosamide alone.ConclusionsAOX was found in all the snail intermediate hosts of Schistosoma examined here. AOX was significantly activated in O. hupensis under niclosamide-induced stress, which led to a reduction in oxidative stress in the snail. The inhibition of AOX activity in snails can dramatically enhance the molluscicidal effect of niclosamide. A potential target for the development of an environmentally safe snail control method, which acts by inhibiting the activity of AOX, was identified in this study.Graphical abstract

Akt/mTOR Activation in Lung Cancer Tumorigenic Regulators and Their Potential Value as Biomarkers

The high incidence and modest therapeutic outcomes of lung cancer have prompted the identification of cell molecular targets/biomarkers within the complex networks of interactions involved in cell malignancy. Most of the EMT-related regulatory mediators underline patients’ biologic variations, therapeutic refractory events, and tumor cell heterogeneity. Patient stratification based on the understanding of the relevant pathways, such as the PI3K/Akt axis crucial in EMT initiation, could favorably alter disease management. Significant clinical advantage could be expected when overexpressed Akt tyrosine kinase (Akt2) is addressed as a malignant biomarker to guide clinical management decisions, improving prognosis in lung cancer patients. Moreover, one should not miss the opportunity of using it as a druggable target aiming at the inhibition of the downstream complexity that underlies cell proliferation and survival, expression of stemness markers and drug resistance. The value of mTOR, as a downstream target of Akt, and the further activation of EMT transcription factors Twist, Snail and Zeb1 are revisited in this review. An in-depth state-of-the-art assessment provides evidence of its role in the mechanistic inhibition of epithelial markers, such as E-cadherin and miR-200, while inducing the expression of the mesenchymal ones, such as vimentin, N-cadherin, and miR-21. Lastly, evidence suggesting another transcription factor, FOXM1, as the link between the PI3K/Akt and Wnt/β-catenin pathways, prompting cell metabolism through the regulation of p70S6K, is analyzed. A more realistic approach is advised to address unmet clinical needs and support decision making at a clinical level. Taking into consideration several complex intracellular interactions might further improve patient stratification and result in better outcomes.

JNK-mediated Ser27 phosphorylation and stabilization of SIRT1 promote growth and progression of colon cancer through deacetylation-dependent activation of Snail.

Sirtuin 1 (SIRT1), an NAD+‐dependent histone/protein deacetylase, has multifaceted functions in various biological events such as inflammation, aging, and energy metabolism. The role of SIRT1 in carcinogenesis, however, is still under debate. Recent studies have indicated that aberrant overexpression of SIRT1 is correlated with metastasis and poor prognosis in several types of malignancy, including colorectal cancer. In the present study, we found that both SIRT1 and SIRT1 phosphorylated on serine 27 were coordinately upregulated in colon cancer patients’ tissues and human colon cancer cell lines. This prompted us to investigate a role of phospho‐SIRT1 in the context of colon cancer progression. A phosphorylation‐defective mutant form of SIRT1, in which serine 27 was substituted by alanine (SIRT1‐S27A), exhibited lower protein stability compared to that of wild‐type SIRT1. Notably, human colon cancer (HCT‐116) cells harboring the SIRT1‐S27A mutation showed decreased cell proliferation and reduced capability to form xenograft tumor in athymic nude mice, which was accompanied by diminished transcriptional activity of Snail. HCT‐116 cells carrying SIRT1‐S27A were less capable of deacetylating the Snail protein, with a concomitant decrease in the levels of interleukin (IL)‐6 and IL‐8 mRNA transcripts. Taken together, these observations suggest that SIRT1 stabilized through phosphorylation on serine 27 exerts oncogenic effects at least partly through deacetylation‐dependent activation of Snail and subsequent transcription of IL‐6 and IL‐8 in human colon cancer cells.

Demographic and spatial structure at the stage of expansion in thepopulations of some alien land snails in Belgorod city (CentralRussian Upland)

Studying of the demographic characteristics of populations of alien species at the stage of expansion makes it possible to assess their invasive potential. Field studies of alien terrestrial gastropods Xeropicta derbentina (Gastropoda,Stylommatophora, Hygromiidae), Brephulopsis cylindrica (Gastropoda, Stylommatophora, Enidae) and Harmozica ravergiensis (Gastropoda, Stylommatophora, Hygromiidae) have been carried out in Belgorod city, the southern part of the Central Russian Upland (Russia). All studied species are native to the Black Sea region and the Caucasus; in the southern part of the Central Russian Upland, they have been registered over the past ten years. The population density, demographic structure, and spatial structure have been analyzed for three years of observations (2017, 2019, 2020). During this period, the changes in the population density have been noted for all studied species. The most pronounced changes have been observed in X. derbentina, whose population density has significantly decreased, and the spatial structure has changed from clustered type to random distribution. In addition, the age structure of X. derbentina population had a pronounced change at the beginning of the snail activity season of 2020. The expansion of the other two invaders, B. cylindrica and H. ravergiensis, is proved as more successful. At the same time, H. ravergiensis is distributed throughout the city. This species has stable population density and random type of spatial population structure on most sites. B. cylindrica has the highest population density: in 2020, it has reached 406 ind./m2 on average, with the maximum of 1215 ind./m2.Meantime, the species keeps the clustered spatial structure, which is explained by both high population density and adaptation to the arid conditions of its natural range.

INHIBITORY ACTIVITY OF SNAIL (Achatina fulica (Lam.) Bowdich) MUCUS ON GROWTH OF MAMMARY CANCER IN RAT INDUCED WITH DMBA (7,12-DIMETHYLBENZ(α)ANTHRACENE)

Back to nature as a medication concept has been accepted widely because it has fewer side effects than modern medicines. Researches on natural products as anticancer agent therapies are in progress. This present research was conducted to determine that the inhibitory activity of snail (Achatina fulica (Lam.) Bowdich) mucus inhibits the growth rate of mammary cancer in SpragueDawley rats. Five groups of female rats, with four individuals each, were induced with 7,12-dimethylbenz(α)anthracene (DMBA) for five weeks. Snail mucus was applied every seven days to the treated rats with three different dosages (15, 20, and 25 mg/kg BW). Observations were done on diameters and growth rate of the mammary cancer lump developed at the end of week 8, 10, 12, 14, and 16. Histopathological examination was carried out at the end of the 16th week. Inhibitory activity results showed a row of average diameter of the cancerous lumps on rats with the following details. The results obtained from the application of snail mucus at dosage of 15, 20, and 25 mg/kg BW were at 0.40, 0.60, and 0.09 cm respectively along with average growth rate of cancerous lumps at 1.50, 0.75, and 0.25. The histopathology results of the snail mucus treatment at dosage of 15, 20, and 25 mg/kg BW showed normal tissue depiction and similarly normal histopathological form indicated by the presence of their sub-clinic components. The results showed that a snail mucus dose of 25 mg/kg BW was able to perform inhibitory activity on growing mammary cancer in rats induced by DMBA.

Trophic Transfer and Toxicity of (Mixtures of) Ag and TiO2 Nanoparticles in the Lettuce-Terrestrial Snail Food Chain.

The increasing application of biosolids and agrochemicals containing silver nanoparticles (AgNPs) and titanium dioxide nanoparticles (TiO2NPs) results in their inevitable accumulation in soil, with unknown implications along terrestrial food chains. Here, the trophic transfer of single NPs and a mixture of AgNPs and TiO2NPs from lettuce to snails and their associated impacts on snails were investigated. Both AgNPs and TiO2NPs were transferred from lettuce to snails with trophic transfer factors (defined as the ratio of the Ag/Ti concentration in snail tissues to the Ag/Ti concentration in lettuce leaves) of 0.2–1.1 for Ag and 3.8–47 for Ti. Moreover, the majority of Ag captured by snails in the AgNP-containing treatments was excreted via feces, whereas more than 70% of Ti was distributed in the digestive gland of snails in the TiO2NP-containing treatments. Additionally, AgNP-containing treatments significantly inhibited the activity of snails, while TiO2NP-containing treatments significantly reduced feces excretion of snails. Furthermore, the concurrent application of AgNPs and TiO2NPs did not affect the biomagnification and distribution patterns of Ag and Ti in snails, whereas their co-existence exhibited more severe inhibition of the growth and activity of snails than in the case of applying AgNPs or TiO2NPs alone. This highlights the possibility of nanoparticle transfer to organisms of higher trophic levels via food chains and the associated risks to ecosystem health.

Omeprazole suppresses aggressive cancer growth and metastasis in mice through promoting Snail degradation.

Omeprazole is a proton pump inhibitor that has recently been reported to exhibit anticancer activity against several types of cancer. However, the anticancer mechanisms of omeprazole remain elusive. Snail is an oncogenic zinc finger transcription factor; aberrant activation of Snail is associated with the occurrence and progression of cancer. In this study, we investigated whether Snail acted as a direct anticancer target of omeprazole. We showed that omeprazole displayed a high binding-affinity to recombinant Snail protein (Kd = 0.076 mM), suggesting that omeprazole directly and physically binds to the Snail protein. We further revealed that omeprazole disrupted CREB-binding protein (CBP)/p300-mediated Snail acetylation and then promoted Snail degradation through the ubiquitin-proteasome pathway in HCT116 cells. Omeprazole treatment markedly suppressed Snail-driven epithelial-mesenchymal transition (EMT) in aggressive HCT116, SUM159, and 4T1 cancer cells in vitro and reduced EMT-associated tumor invasion and metastasis in cancer cell xenograft models. Omeprazole also inhibited tumor growth by limiting Snail-dependent cell cycle progression. Overall, this study, for the first time, identifies Snail as a target of omeprazole and reveals a novel mechanism underlying the therapeutic effects of omeprazole against cancer. This study strongly suggests that omeprazole may be an excellent auxiliary drug for treating patients with malignant tumors.

SNHG8 Promotes the Progression of Epstein-Barr Virus-Associated Gastric Cancer via Sponging miR-512-5p and Targeting TRIM28.

SNHG8, a family member of small nucleolar RNA host genes (SNHG), has been reported to act as an oncogene in gastric carcinoma (GC). However, its biological function in Epstein–Barr virus (EBV)-associated gastric cancer (EBVaGC) remains unclear. This study investigated the role of SNHG8 in EBVaGC. Sixty-one cases of EBVaGC, 20 cases of non-EBV-infected gastric cancer (EBVnGC), and relative cell lines were studied for the expression of SNHG8 and BHRF1 (BCL2 homolog reading frame 1) encoded by EBV with Western blot and qRT-PCR assays. The relationship between the expression levels of SNHG8 and the clinical outcome in 61 EBVaGC cases was analyzed. Effects of overexpression or knockdown of BHRF1, SNHG8, or TRIM28 on cell proliferation, migration, invasion, and cell cycle and the related molecules were determined by several assays, including cell proliferation, colony assay, wound healing assay, transwell invasion assay, cell circle with flow cytometry, qRT-PCR, and Western blot for expression levels. The interactions among SNHG8, miR-512-5p, and TRIM28 were determined with Luciferase reporter assay, RNA immunoprecipitation (RIP), pull-down assays, and Western blot assay. The in vivo activity of SNHG8 was assessed with SNHG8 knockdown tumor xenografts in zebrafish. Results demonstrated that the following. (1) BHRF1 and SNHG8 were overexpressed in EBV-encoded RNA 1-positive EBVaGC tissues and cell lines. BHRF1 upregulated the expressions of SNHG8 and TRIM28 in AGS. (2) SNHG8 overexpression had a significant correlation with tumor size and vascular tumor thrombus. Patients with high SNHG8 expression had poorer overall survival (OS) compared to those with low SNHG8 expression. (3) SNHG8 overexpression promoted EBVaGC cell proliferation, migration, and invasion in vitro and in vivo, cell cycle arrested at the G2/M phase via the activation of BCL-2, CCND1, PCNA, PARP1, CDH1, CDH2 VIM, and Snail. (4) Results of dual-luciferase reporter assay, RNA immunoprecipitation, and pull-down assays indicated that SNHG8 sponged miR-512-5p, which targeted on TRIM28 and promoted cancer malignant behaviors of EBVaGC cells. Our data suggest that BHRF1 triggered the expression of SNHG8, which sponged miR-512-5p and upregulated TRIM28 and a set of effectors (such as BCL-2, CCND1, CDH1, CDH2 Snail, and VIM) to promote EBVaGC tumorigenesis and invasion. SNHG8 could be an independent prognostic factor for EBVaGC and sever as target for EBVaGC therapy.

Evaluation of the effect of transgenic Bt cotton on snails Bradybaena (Acusta) ravida and Bradybaena similaris (Ferussac)

The impact of transgenic crops on non-target organisms is a key aspect of environmental safety assessment to transgenic crops. In the present study, we fed two snail species, Bradybaena (Acusta) ravida (B. ravida) and Bradybaena similaris (Ferussac)(B. similaris), with the leaves of transgenic Bt cotton Zhong 30 (Z30) and control cotton, its parent line zhong 16 (Z16), to assess the environmental safety of Bt cotton to common non-target organisms in the field. Survival, body weight, shell diameter, helix number, reproduction rate, superoxide dismutase (SOD) activity and Bt protein concentration in snails were monitored in 15 days and 180 days experiments. We also monitored the population dynamics of B. ravida and B. similaris in Z30 and Z16 cotton fields for two successive years. Compared to the snails fed on the control cotton Z16, there was no significant difference in survival, growth, reproduction, and SOD activity on Bt cotton Z30. Bt protein concentrations were significantly between different treatments, and Bt protein residues were only detected in the feces of the Z30 treatment. According to the field data, the number of B. ravida and B. similaris fluctuated considerably across seasons over the entire cotton-growing season; however, there were no significant differences between the Bt and control cotton fields at similar time. As the results showed, in our experiments, Bt cotton Z30 had no adverse effects on the two snail species, both in the laboratory and in the fields.

Abstract 2881: Snail-mediated regulation of MMP1 expression in the context of ovarian cancer invasiveness

Abstract Ovarian cancer ranks fifth among the causes of cancer death in US women. Despite initial positive response to cytoreductive surgery and chemotherapy treatments with platinum-based therapies, over 80% of patients with high-grade serous ovarian cancer (HGSOC), the most common type of ovarian cancer, exhibit relapse. Most recurrent cases of HGSOC are resistant to chemotherapy, and cells are highly invasive. Therefore, further studies are required to understand the mechanisms of invasiveness and chemoresistance in relapse patients. In HGSOC, invasiveness can result when epithelial cancer cells undergo epithelial-mesenchymal transition (EMT). The transcription factor SNAI1 (Snail) initiates the EMT process and increased Snail expression can result in enhancement of metastatic potential, acquisition of stem cell-like characteristics, and resistance to chemotherapeutic drugs. However, the mechanism of activation of invasiveness in ovarian cancer by Snail has not been defined. In OVCAR8, a HGSOC cell line, comparison of shSnail knockdown to control through RNA-sequencing revealed that Snail expression is directly correlated to matrix metalloprotease 1 (MMP1) expression. MMP1 is an extracellular matrix remodeler known to digest interstitial collagens. High MMP1 expression has been linked to increased cancer invasion, metastasis, angiogenesis, and apoptosis. Furthermore, in breast cancer, MMP1 has been reported to be a downstream target of Slug, another major EMT transcription factor. Some evidence for Snail activation of MMPs in liver cancer has been presented, but a direct relationship has not been established. Therefore, we hypothesize that overexpression of Snail in ovarian cancer cells results in MMP1 upregulation, leading to increased invasion. RT-qPCR was performed to analyze relative mRNA expression levels of Snail and MMP1 in eight cell lines. We found that Snail and MMP1 levels were higher in the more mesenchymal-like cell lines, some of which are known to be more invasive. Additionally, Snail overexpression led to a significant corresponding increase in MMP1 mRNA expression and resistance to cisplatin. Chromatin immunoprecipitation (ChIP) experiments demonstrate Snail binding to the MMP1 promoter. Thus, our preliminary findings have shown strong evidence of a Snail-mediated regulation of MMP1 expression in the context of HGSOC invasiveness. Citation Format: Tise Suzuki, Casey Curow, Hanmin Wang, Asis Martinez, Nozomi Hojo, Juli Unternaehrer. Snail-mediated regulation of MMP1 expression in the context of ovarian cancer invasiveness [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2881.

SOS1 promotes epithelial-mesenchymal transition of Epithelial Ovarian Cancer(EOC) cells through AKT independent NF-κB signaling pathway

We aimed to explore the role and mechanism of SOS1 (Son of sevenless homolog 1) in malignant behaviors of epithelial ovarian cancer (EOC) cells Hey with high metastatic potential. Firstly, compared with Hey-WT (wild type) and Hey-NT (none targeted) cells, Hey-SOS1i cells showed decreased polarities, disorders in cytoskeleton arrangement. Numbers of transwell migrated, invaded, intravasation cells and extravasated cells were decreased significantly. Hey-NT cells and Hey-SOS1i cells were employed to establish a peritoneal dissemination model in nude mice. Hey-SOS1i cells formed less implantation metastatic foci in the abdominal cavity than Hey-NT cells, especially on the intestine and diaphragm in the 5th week after the tumor cells were injected intraperitoneally. SOS1 knockdown in Hey cells resulted in increased E-cadherin and decreased Vimentin, N-cadherin, MMP2, and MMP9, together with reduced Snail and activation of NF-κB pathway. Together, these results suggest SOS1 might induce EMT through activating AKT independent NF-κB pathway and the transcriptive activity of Snail, and subsequently regulate the cytoskeleton reprogramming and cell motility of Hey, one of EOC cells with high metastatic potential. This may provide some new targets for the treatment of ovarian cancer with high metastatic potential.

LINC01348 suppresses hepatocellular carcinoma metastasis through inhibition of SF3B3-mediated EZH2 pre-mRNA splicing.

Long non-coding RNAs (lncRNA) play crucial roles in hepatocellular carcinoma (HCC) progression. However, the specific functions of lncRNAs in alternative splicing (AS) and the metastatic cascade in liver cancer remain largely unclear. In this study, we identified a novel lncRNA, LINC01348, which was significantly downregulated in HCC and correlated with survival functions in HCC patients. Ectopic expression of LINC01348 induced marked inhibition of cell growth, and metastasis in vitro and in vivo. Conversely, these phenotypes were reversed upon knockdown of LINC01348. Mechanistically, LINC01348 complexed with splicing factor 3b subunit 3 (SF3B3) acted as a modulator of EZH2 pre-mRNA AS, and induced alterations in JNK/c-Jun activity and expression of Snail. Notably, C-terminal truncated HBx (Ct-HBx) negatively regulated LINC01348 through c-Jun signaling. Our data collectively highlight those novel regulatory associations involving LINC01348/SF3B3/EZH2/JNK/c-Jun/Snail are an important determinant of metastasis in HCC cells and support the potential utility of targeting LINC01348 as a therapeutic strategy for HCC.

Cinnamon oil as a co-chemotherapy agent through inhibition of cell migration and MMP-9 expression on 4T1 cells.

The long-term and high-dose use of doxorubicin as chemotherapy for triple-negative breast cancer (TNBC) patients induces epithelial-to-mesenchymal transition (EMT) and stimulates cancer metastasis. Cinnamaldehyde is a major compound of cinnamon oil (CO) suppressing Snail and NFκB activity that are involved in cell migration. This study aims to explore the activity of CO as a co-chemotherapeutic agent on 4T1 breast cancer cells. The CO was obtained by water and steam distillation and was characterized phytochemically by gas chromatography-mass spectrometry (GC-MS). Cytotoxic activity of single CO or in combination with doxorubicin was observed by MTT assay. Cell migration and MMP-9 expression were measured by scratch wound healing and gelatin zymography assays. The intracellular reactive oxygen species (ROS) levels were observed by 2',7'-dichlorofluorescin diacetate (DCFDA) staining flowcytometry. The phytochemical analysis with GC-MS showed that CO contains 14 compounds with cinnamaldehyde as the major compound. CO exhibited cytotoxicity on 4T1 cells with the IC50 value of 25μg/mL and its combination with doxorubicin decreased cell viability and inhibited cell migration compared to a single use. Furthermore, the combination of CO and doxorubicin inhibited MMP-9 expression and elevated intracellular ROS levels compared to control. CO has the potential to be developed as a co-chemotherapy agent through inhibition of cell migration, and intracellular ROS levels elevation.

No effects of waterproof marking on the behaviour and growth of Physa acuta Draparnaud, 1805 (Gastropoda: Hygrophila: Physidae) in the laboratory

Physa acuta Draparnaud, 1805 is one of the most common freshwater gastropod species, with worldwide distribution. It is an effective periphyton grazer and a potential keystone species in shallow-water systems, where it can boost macrophyte well-being and thus help maintain high water clarity even in nutrient-rich habitats. P. acuta also has been extensively studied in ecotoxicological and behavioural investigations. Such investigations may require observations on individual snails. A method to distinguish individual snails in small-scale experiments is marking their shells with paint dots. However, such marking must not influence snail behaviour (nutritional, reproductive, respiratory, etc.) or growth to avoid confounding effects. Earlier investigations point to no or very limited effects of marking on aquatic and terrestrial snail survival, behaviour, and growth. We tested whether marking could affect the behaviour (as snail activity) and growth of P. acuta using a waterproof, oil-based, non-toxic, fine-point car-body paint marker. Snails were divided into a “marked” and an “unmarked” (control) group of ten snails each in an eight-day experiment. The marking had no effect on the snail activity or growth. The snails survived the experiment and produced egg clutches well beyond the eight-day period. The marking persisted without fading during and beyond the experimental period. Our results support earlier findings that the use of oil-based, non-toxic markers can assist in carrying out reliable observations on individual snails, including the small-bodied P. acuta. Combinations of two dots of different colours allow simultaneous observations on a high number of replicate individuals.

A Novel Relative High-Density Lipoprotein Index to Predict the Structural Changes in High-Density Lipoprotein and Its Ability to Inhibit Endothelial–Mesenchymal Transition

Therapeutic elevation of high-density lipoprotein (HDL) is thought to minimize atherogenesis in subjects with dyslipidemia. However, this is not the case in clinical practice. The function of HDL is not determined by its concentration in the plasma but by its specific structural components. We previously identified an index for the prediction of HDL functionality, relative HDL (rHDL) index, and preliminarily explored that dysfunctional HDL (rHDL index value > 2) failed to rescue the damage to endothelial progenitor cells (EPCs). To confirm the effectiveness of the rHDL index for predicting HDL functions, here we evaluated the effects of HDL from patients with different rHDL index values on the endothelial–mesenchymal transition (EndoMT) of EPCs. We also analyzed the lipid species in HDL with different rHDL index values and investigated the structural differences that affect HDL functions. The results indicate that HDL from healthy adults and subjects with an rHDL index value < 2 protected transforming growth factor (TGF)-β1-stimulated EndoMT by modulating Smad2/3 and Snail activation. HDL from subjects with an rHDL index value > 2 failed to restore the functionality of TGF-β1-treated EPCs. Lipidomic analysis demonstrated that HDL with different rHDL index values may differ in the composition of triglycerides, phosphatidylcholine, and phosphatidylinositol. In conclusion, we confirmed the applicability of the rHDL index value to predict HDL function and found structural differences that may affect the function of HDL, which warrants further in-depth studies.

Urolithin A Inhibits Epithelial-Mesenchymal Transition in Lung Cancer Cells via P53-Mdm2-Snail Pathway.

PurposeThe epithelial-to-mesenchymal transition (EMT) is a fundamental process in tumor progression that endows cancer cells with migratory and invasive potential. Snail, a zinc finger transcriptional repressor, plays an important role in the induction of EMT by directly repressing the key epithelial marker E-cadherin. Here, we assessed the effect of urolithin A, a major metabolite from pomegranate ellagitannins, on Snail expression and EMT process.MethodsThe role of Snail in urolithin A-induced EMT inhibition in lung cancer cells was explored by wound healing assay and cell invasion assay. The qRT-PCR and CHX assay were performed to investigate how urolithin A regulates Snail expression. Immunoprecipitation assays were established to determine the effects of urolithin A in mdm2-Snail interaction. In addition, the expression of p53 was manipulated to explore its effect on the expression of mdm2 and Snail.ResultsThe urolithin A dose-dependently upregulated epithelial marker and decreased mesenchymal markers in lung cancer cells. In addition, exposure to urolithin A decreased cell migratory and invasive capacity. We have further demonstrated that urolithin A inhibits lung cancer cell EMT by decreasing Snail protein expression and activity. Mechanistically, urolithin A disrupts the interaction of p53 and mdm2 which leads Snail ubiquitination and degradation.ConclusionWe conclude that urolithin A could inhibit EMT process by controlling mainly Snail expression. These results highlighted the role of pomegranate in regulation of EMT program in lung cancer.

Invader to invader facilitation: ice plant mats prompt increased densities and grazing impacts of white garden snails in a Pampean coastal dune system

Non-native species can facilitate other non-native species via their effects on the physical environment. Here, I evaluate whether the ice plant, Carpobrotus edulis, prompts increased densities and grazing impacts of the white garden snail, Theba pisana, in coastal dunes of the Argentinean Pampas, where both species are non-native. Snail densities were higher in ice plant patches than in native graminoid patches over four seasonal samplings. Snail activity and grazing impacts on two co-occurring forbs (the native Hydrocotyle bonariensis and the non-native Calystegia soldanella) were also higher within ice plant patches. Experimental ice plant transplant into graminoid-dominated areas led to increased snail densities and grazing impacts, which demonstrates cause-effect relationships linking ice plant cover with increasing snail densities and grazing damage to forbs. The findings here suggest that ice plant mats provide suitable microclimatic conditions for T. pisana, and thereby facilitate greater snail numbers in ice plant patches leading to increased grazing rates on other plants. On that basis, it can be expected that the cumulative effect of both invaders on co-occurring plants would be larger than their individual effects.

To Eat or Not to Eat: the Importance of Starvation on Behavioral Bioassays

Behavioral endpoints are important parameters to assess the effects of toxicants and other stressors on natural ecosystems. The relevance of these parameters has caused a rise in their use in aquatic ecotoxicology. However, abiotic and biotic parameters may interact causing changes in the behavioral responses. Among those parameters, starvation of animals is a factor that is usually applied in ecotoxicological short-term bioassays. This could alter animal behavior, along with the toxicant. Therefore, the study of the effects of starvation on baseline behaviors of invertebrates is a relevant issue. This study assessed the behavior of the aquatic snail Potamopyrgus antipodarum under a combination of four treatments: (1) animals normally fed (control treatment), (2) starved animals, (3) animals normally fed and exposed to a high conductivity, and (4) starved animals exposed to a high conductivity. The behavior activity of snails was monitored for 14 days. Results show that animals of the second treatment (starved animals) increased their activity. On the contrary, animals of the third and fourth treatments reduced their activity. Animals from the control treatment showed an activity in between starved animals and animals exposed to high conductivity (both starved and normally fed). These results show that starvation increases the snail activity, but under another environmental stressor (i.e., high conductivity), this trend was reversed. The influence of starvation on behavior should be taken into account in the development of behavioral bioassays.