A New Benzo[6,7]oxepino[3,2-b] Pyridine Derivative Induces Apoptosis in Canine Mammary Cancer Cell Lines.

CMC is the most frequently diagnosed cancer and one of the leading causes of death in non-spayed female dogs. Exploring novel therapeutic agents is necessary to increase the survival rate of dogs with CMC. MPOBA is a BZOP derivative that has a significant anticancer effect in a human cell line. The main goal of this study was to investigate the anticancer properties of MPOBA against two CMC cell lines (REM134 and CMGT071020) using a 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay, a wound healing assay, a transwell migration assay, an Annexin V-FITC apoptosis assay with a flow cytometry analysis, a mRNA expression analysis using quantitative real-time PCR (qRT-PCR), and an immunohistochemistry (IHC). According to the accumulated studies, MPOBA caused significant concentration- and time-dependent reductions in cell proliferation and cell migration and induced apoptosis in both CMC cell lines. In gene expression analysis, nine canine genes, including TP53, BCL-2, BAX, epidermal growth factor receptor (EGFR), snail transcription factor (SNAIL), snail-related zinc-finger transcription factor (SLUG), TWIST, E-cadherin, and N-cadherin, were investigated. The mRNA expression results revealed that MPOBA induced upregulation of TP53 and overexpression of the pro-apoptotic gene BAX, together with an inhibition of BCL-2. Moreover, MPOBA also suppressed the mRNA expression levels of SNAIL, EGFR, and N-cadherin and induced upregulation of E-cadherin, crucial genes related to the epithelial-to-mesenchymal transition (EMT). However, there was no significant difference in the IHC results of the expression patterns of vimentin (VT) and cytokeratin (CK) between MPOBA-treated and control CMC cells. In conclusion, the results of the present study suggested that MPOBA exhibited significant anticancer activity by inducing apoptosis in both CMCs via upregulation of TP53 and BAX and downregulation of BCL-2 relative mRNA expression. MPOBA may prove to be a potential candidate drug to be further investigated as a therapeutic agent for CMC.

A New Benzaldehyde Derivative Exhibits Antiaflatoxigenic Activity against Aspergillus flavus.

Aflatoxin B1 (AFB1) is the most potent naturally occurring carcinogen for humans and animals produced by the common fungus Aspergillus flavus (A. flavus). Aflatoxin (AF) contamination in commodities is a global concern related to the safety of food and feed, and it also impacts the agricultural economy. In this study, we investigated the AFB1-inhibiting activity of a new benzaldehyde derivative, 2-[(2-methylpyridin-3-yl)oxy]benzaldehyde (MPOBA), on A. flavus. It was found that MPOBA inhibited the production of AFB1 by A. flavus, with an IC50 value of 0.55 mM. Moreover, the inhibition of conidiation was also observed at the same concentration. The addition of MPOBA resulted in decreased transcript levels of the aflR gene, which encodes a key regulatory protein for the biosynthesis of AF, and also decreased transcript levels of the global regulator genes veA and laeA. These results suggested that MPOBA has an effect on the regulatory mechanism of the development and differentiation of conidia, leading to the inhibition of AFB1 production. In addition, the cytotoxicity study showed that MPOBA had a very low cytotoxic effect on the Madin-Darby canine kidney (MDCK) cell line. Therefore, MPOBA may be a potential compound for developing practically effective agents to control AF contamination.

Anti-cancer potentials of Gynura procumbens leaves extract against two canine mammary cancer cell lines.

BACKGROUND: The anti-cancer effects of Gynura procumbens leaves extract (GPE) have been reported in various human cancers. However, the anti-cancer effects and molecular mechanisms of this extract on canine mammary cancer (CMC) have not yet been elucidated. OBJECTIVES: The main goal of this study was to investigate the anti-cancer properties of GPE against two CMC cell lines (CHMp-13a and CHMp-5b). METHODS: The GP leaves were extracted with 80% ethanol. Anti-cancer potentials of GPE on CHMp-13a and CHMp-5b cancer cell lines using dimethyl-2-thiazolyl-2,5-diphenyl-2H-tetrazolium bromide (MTT), wound healing, transwell migration, and caspase 3/7 activity assays were evaluated. The mRNA expression levels of two oncogenes: epidermal growth factor receptor (EGFR) and twist family bHLH transcription factor 1 (TWIST) and one tumour suppressor gene: phosphatase and tensin homolog (PTEN) in these cell lines were determined by quantitative real-time PCR (qRT-PCR). In addition, The EGFR and PTEN protein levels as well as protein kinase B (AKT) and extracellular signal-regulated kinase 1/2 (ERK1/2) phosphorylation levels expression were also evaluated by western blot analysis. RESULTS: The results showed that GPE caused a significant concentration- and time-dependent reduction in cell proliferation of both CHMp-13a and CHMp-5b cells, detected by MTT assays. This extract also significantly suppressed cancer cell migration in both cell lines, tested by wound healing and transwell migration assays. Additionally, the increase in caspase 3/7 activity observed in both CMC cell treated with GPE suggests that GPE induced caspase 3/7 dependent apoptosis. Moreover, GPE significantly decreased EGFR mRNA and protein expression levels compared to control in both cell lines in a dose-dependent manner. CONCLUSION: These findings emphasized that GPE has an in vitro anti-cancer activity against CMC by inhibiting EGFR signalling pathway. Thus, GPE may serve as an alternative therapy in CMC with high EGFR expression.

Evaluation of Aflatoxin Concentrations and Occurrence of Potentially Toxigenic Fungi in Imported Chia Seeds Consumed in Thailand.

ABSTRACT: This study was conducted to investigate possible contamination by aflatoxins (AFs) and aflatoxigenic fungi in imported chia seeds consumed in Thailand. A survey was performed on 100 samples of imported chia seeds collected from supermarkets and health food stores in Bangkok from May 2017 to February 2018. Ten mold species belonging to Aspergillus and Penicillium were isolated, and Aspergillus flavus was the most prevalent aflatoxigenic fungi. Chia seed samples were cleaned with an immunoaffinity column and analyzed for AFs by high-performance liquid chromatography with fluorescence detection using precolumn derivatization. AFs were detected in 40% of total samples at concentrations of 0.4 to 10.99 ng/g. Among the positive samples, three were contaminated with total AFs at concentrations higher than the European Union regulatory limit (4 ng/g). The most commonly found AF found in chia seeds was AFB1.

Pharmacokinetics of ceftriaxone in Green sea turtles (Chelonia mydas) following intravenous and intramuscular administration at two dosages.

Green sea turtles are widely distributed in tropical and subtropical waters. Adult green sea turtles face many threats, primarily from humans, including injuries from boat propellers, being caught in fishing nets, pollution, poaching, and infectious diseases. To the best of our knowledge, limited pharmacokinetic information to establish suitable therapeutic plans is available for green sea turtles. Therefore, the present study aimed to describe the pharmacokinetic characteristics of ceftriaxone (CEF) in green sea turtles, Chelonia mydas, following single intravenous and intramuscular administrations at two dosages of 10 and 25 mg/kg body weight (b.w.). Blood samples were collected at assigned times up to 96 hr. The plasma concentrations of CEF were measured by liquid chromatography tandem mass spectrometry. The concentrations of CEF in the plasma were quantified up to 24 and 48 hr after i.v. and i.m. administrations at dosages of 10 and 25 mg/kg b.w., respectively. The Cmax values of CEF were 15.43 ± 3.71 µg/ml and 43.48 ± 4.29 µg/ml at dosages of 10 and 25 mg/kg, respectively. The AUClast values increased in a dose-dependent fashion. The half-life values were 2.89 ± 0.41 hr and 5.96 ± 0.26 hr at dosages of 10 and 25 mg/kg b.w, respectively. The absolute i.m. bioavailability was 67% and 108%, and the binding percentage of CEF to plasma protein was ranged from 20% to 29% with an average of 24.6%. Based on the pharmacokinetic data, susceptibility break-point and PK-PD index (T > MIC, 0.2 µg/ml), i.m. administration of CEF at a dosage of 10 mg/kg b.w. might be appropriate for initiating treatment of susceptible bacterial infections in green sea turtles.

Inhibitory Effects of Thai Essential Oils on Potentially Aflatoxigenic Aspergillus parasiticus and Aspergillus flavus.

　The antiaflatoxigenic and antifungal activities of essential oils (EOs) of finger root (Boesenbergia rotunda (L.) Mansf.), pine (Pinus pinaster), rosewood (Aniba rosaedora), Siam benzoin (Styrax tonkinensis), Thai moringa (Moringa oleifera), and ylang ylang (Cananga odorata) were tested for Aspergillus parasiticus and Aspergillus flavus in potato dextrose broth. Aflatoxin B1 (AFB1) was extracted from culture using a QuEChERS-based extraction procedure and analyzed with high performance liquid chromatography (HPLC) coupled to a fluorescence detector. EO of pine showed the greatest inhibition of growth and AFB1 production of A. parasiticus, followed by EOs of rosewood, finger root, Siam benzoin, and ylang ylang. EO of finger root gave the best inhibitory effects on A. flavus, followed by EOs of rosewood, pine, ylang ylang, and Siam benzoin. EO of Thai moringa did not show any significant inhibition of aflatoxigenic fungi. The antiaflatoxigenic activities of EOs correlated with their antifungal activities in the dosedependent manner. Comparison of the application of the five selected EOs in peanut pods by direct and vapor exposure indicated that the AFB1 production inhibitory effects of the five EOs by direct exposure were faster and more effective than by vapor exposure. EO of finger root showed the best inhibition of AFB1 production of A. flavus in peanut pods by direct exposure, followed by EOs of pine, rosewood, ylang ylang, and Siam benzoin.

Health risk associated with the consumption of duck egg containing endosulfan residues.

Eight duck farms and a local market in Suphanburi province, Thailand adjacent to paddy fields were selected for this study. The concentrations of endosulfan isomers (α- and ß-endosulfan) and endosulfan sulfate in environmental matrices (water, soil, feed) and duck eggs were determined. Human health risk via the contaminated egg consumption was also evaluated. Analysis of environmental matrices found both endosulfan isomers (α- and ß-endosulfan) and endosulfan sulfate in most samples. Endosulfan sulfate was predominantly found in all matrices followed by ß- and α-endosulfan, respectively. The total endosulfan concentrations were in the following order: feed > soil > water. However, the levels of endosulfan detected were lower than the regulatory maximum residue limit of endosulfan, except in water (>0.200 ng mL(-1)). Endosulfan sulfate in duck egg samples was also predominantly detected in both yolk and albumin. The average total endosulfan residues (∑endosulfan) in yolk (6.73 ng g(-1)) were higher than in albumin (4.78 ng g(-1)). According to principle component analysis, we found that paddy soil surrounding the duck farms is the suspected source of endosulfan contamination in husbandry water which subsequently contaminates duck eggs. The estimated daily intakes (EDIs) of these endosulfan-contaminated eggs were well below the acceptable daily intake (ADI) for endosulfan (6 µg kg(-1) day(-1)). However, the consumption of this contaminated duck eggs should be of concerns in regard to chronic exposure. Therefore, the better environmental managements to reduce endosulfan residues can play a crucial role for decreasing human health risk.

Search for aflatoxin and trichothecene production inhibitors and analysis of their modes of action.

Mycotoxin contamination of crops is a serious problem throughout the world because of its impact on human and animal health as well as economy. Inhibitors of mycotoxin production are useful not only for developing effective methods to prevent mycotoxin contamination, but also for investigating the molecular mechanisms of secondary metabolite production by fungi. We have been searching for mycotoxin production inhibitors among natural products and investigating their modes of action. In this article, we review aflatoxin and trichothecene production inhibitors, including our works on blasticidin S, methyl syringate, cyclo(L-Ala-L-Pro), respiration inhibitors, and precocene II.

The complete mitogenome of the freshwater fairy shrimp Streptocephalus sirindhornae (Crustacea: Anostraca: Streptocephalidae).

In this study, we amplified, sequenced and analyzed the complete mitogenome of the freshwater fairy shrimp Streptocephalus sirindhornae (Crustacea: Anostraca: Streptocephalidae). The full-length of the S. sirindhornae mitogenome is a circular molecule of 16,887 bp in size with an A + T content of 64.5%. It has the largest putative control region (2794 bp) with the lowest A + T content (62.6%) for all determined branchiopod mitogenomes. The genome consisted of 37 genes that are involved in the respiration chain as well as the mitochondrial translation system. The S. sirindhornae mitogenome exhibits an identical gene arrangement as the Artemia pattern, which shows translocation and inversion of two transfer-RNA genes compared to the pancrustacean ancestral pattern. This is by far the first determined mitogenome of a freshwater fairy shrimp. The results of our study will provide significant data for reconstructing the consensus Branchiopoda tree of life.

Using Nassarius stolatus as a potential heavy metal biomonitor.

Nassarius stolatus was evaluated as the potential heavy metal (Cd, Fe, Mn, Ni and Pb) accumulator in Don Hoi Lot sandbar, Samut Songkhram province, Thailand. This selected species belongs to the Gastropoda, which is widely distributed in the coastal areas from the upper Gulf through the southern part of Thailand. From our findings, the heavy metal accumulations in N. stolatus tissues were Fe > Pb > Mn > Ni > Cd. The retrieved bioaccumulation factor (BAF) indicated that N. Stolatus has high potential to be a biomonitor for the contaminations of Fe and Mn in water and Cd, Ni and Pb in sediment.

Prevention of aflatoxin contamination by a soil bacterium of Stenotrophomonas sp. that produces aflatoxin production inhibitors.

A soil bacterium, designated strain no. 27, was found to produce aflatoxin-production inhibitors. The strain was identified as a species of the genus Stenotrophomonas, and was found to be closely related to Stenotrophomonas rhizophila. Two diketopiperazines, cyclo(L-Ala-L-Pro) and cyclo(L-Val-L-Pro), were isolated from the bacterial culture filtrate as main active components. These compounds inhibited aflatoxin production of Aspergillus parasiticus and Aspergillus flavus in liquid medium at concentrations of several hundred µM without affecting fungal growth. Both inhibitors inhibited production of norsorolinic acid, a biosynthetic intermediate involved in an early step of the aflatoxin biosynthetic pathway, and reduced the mRNA level of aflR, which is a gene encoding a key regulatory protein necessary for the expression of aflatoxin-biosynthetic enzymes. These results indicated that the inhibitors targets are present in early regulatory steps leading to AflR expression. Co-culture of strain no. 27 with aflatoxigenic fungi in liquid medium effectively suppressed aflatoxin production of the fungus without affecting fungal growth. Furthermore, application of the bacterial cells to peanuts in laboratory experiments and at a farmer's warehouse in Thailand by dipping peanuts in the bacterial cell suspension strongly inhibited aflatoxin accumulation. The inhibitory effect was dependent on bacterial cell numbers. These results indicated that strain no. 27 may be a practically effective biocontrol agent for aflatoxin control.

Isolation of methyl syringate as a specific aflatoxin production inhibitor from the essential oil of Betula alba and aflatoxin production inhibitory activities of its related compounds.

Methyl syringate was isolated from the essential oil of Betula alba as an aflatoxin production inhibitor. It inhibited aflatoxin production of Aspergillus parasiticus and Aspergillus flavus with IC(50) values of 0.9 and 0.8 mM, respectively, without significantly inhibiting fungal growth. Methyl syringate reduced mRNA levels of genes (aflR, pksA, and omtB) [corrected] encoding proteins required for aflatoxin biosynthesis. Methyl gallate, methyl 3,4,5-trimethoxybenzoate, and methyl 3-O-methylgallate inhibited both aflatoxin production and fungal growth of A. parasiticus and A. flavus. However, their acids and syringic acid did not inhibit aflatoxin production and growth of A. parasiticus significantly, although gallic acid inhibited aflatoxin production of A. flavus with selectivity. The 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity of methyl syringate was much weaker than that of gallic acid. These results showed that methyl syringate has a unique inhibitory activity toward aflatoxin production with a different mode of action from that of gallic acid.