Effects of free antibiotic resistance genes in the environment on intestinal microecology of mice.

The rapid spread of antibiotic resistance genes (ARGs) is a great challenge to the ecological safety and human health. The intestine of humans and animals is an important site for the increase and spread of ARGs due to the great diversity and abundance of microorganisms in the intestinal microecology. ARGs, including the intracellular (iARGs) and the extracellular (eARGs) ARGs, are usually introduced into the intestinal tract through the diet, and the iARGs are colonized and spread in the intestinal microbiota with the help of the host bacteria. However, whether the eARGs can enter the intestinal microorganisms in the absence of host bacteria is not known. Here, we show the transformation and the diffusion of the ampramycin resistance gene (Ap) carried by the free plasmid RK2 in the intestinal microbiota of mice. After two days of consecutive gavage with free RK2, the intracellular Ap gene increases from days 0-8 in the feces of mice, and has remained constant. Bacterial transformation happens in the small intestine, including proximal and distal jejuna and proximal and distal ilea, at the early stage (first two days), and the intracellular RK2 is diffused into the intestinal microbiota of mice by conjugation on days 2-8 day, which is based on the distribution of eARG and iARG and the mRNA expression levels of trbBp, trfAp, korA, korB, and trbA. The characteristics of ARGs susceptible microbiota for transformation are analyzed using 16s rRNA gene sequencing, transmission electron microscopy, and flow cytometric. The ingestion of RK2 affects the composition of intestinal microbiota especially for Proteobacteria, and the antibiotic residue promotes the increase in Escherichia coli. These findings are important to assess the risk of ARGs, especially the eARGs in the intestinal microecology.

Preparation of ß-CD-Ellagic Acid Microspheres and Their Effects on HepG2 Cell Proliferation.

OBJECTIVE: In this study, ß-cyclodextrin (ß-CD) was chosen as the coating for ellagic acid to prepare ellagic acid microspheres, and the effect of microspheres on the growth of HepG2 cells was observed. METHODS: Scanning electron microscopy, infrared spectroscopy, and release rate analysis were used to identify the formation of ellagic acid microspheres. Methyl thiazolyl tetrazolium (MTT) assay was used to detect the effect of different concentrations of ellagic acid microspheres on tumor cell proliferation at 6, 12, 24 and 36 h, and cell morphology and quantity were observed using hematoxylin-eosin (HE) staining. Single-cell gel electrophoresis was used to observe the effect of ellagic acid microspheres on the DNA damage of HepG2 cells, and the Olive tail moment and the mRNA expression of tumor suppressor protein gene p53 was measured. RESULTS: ß-CD could be used as wrapping material of ellagic acid to prepare ellagic acid microspheres. HepG2 cell proliferation could be inhibited by 0.1, 0.3 and 0.5 g/L of ellagic acid microspheres in a dose- and time-dependent manner, and the mechanism of proliferation inhibition was related to DNA damage and cell apoptosis. CONCLUSION: Preparing ellagic acid microspheres with ß-CD is feasible, and ellagic acid microspheres have potential therapeutic value (anticancer).

Preparation of ß-CD-Vitexin Microspheres and their Effects on SW480 Cell Proliferation.

OBJECTIVE: In order to overcome the insolution and low bioavailability of the vitexin in vivo, ß-cyclodextrin-vitexin (ß-CD-vitexin) microspheres were prepared, and their effects on the proliferation of SW480 cells were observed. METHODS: Scanning electron microscopy, ultraviolet spectrum, Fourier transform infrared spectroscopy, and release rate analysis identified the formation of ß-CD-vitexin microspheres. MTT assay detected the effect of ß-CD-vitexin microspheres on tumor cell proliferation at 6, 12, 24, and 48 h. Fluorescence microscopy and flow cytometry were used to observe the effect of ß-CD-vitexin microspheres on the apoptosis of SW480 cells. The mRNA expression of the p53 gene was measured by qPCR. RESULTS: ß-CD-vitexin microspheres were successfully prepared. SW480 cell proliferation was inhibited by 0.1, 0.2, and 0.4 mg/mL of ß-CD-vitexin microspheres in a dose- and time-dependent manner, and the mechanism of proliferation inhibition was related to cell apoptosis caused by the upregulated expression of p53 gene. CONCLUSION: The preparation of ß-CD-vitexin sustained release microspheres is feasible, and ß-CDvitexin microspheres have potential anti-colorectal cancer value.

Distribution and model prediction of antibiotic resistance genes in Weishan Lake based on the indication of Chironomidae larvae.

The widespread contamination of antibiotic resistance genes (ARGs) in freshwater environment are becoming a serious challenge to human health and ecological safety. Rapid and efficient monitoring of ARGs pollution is of great significance to ARGs control. Water, bottom mud, and fish have all been used to indicate ARG contamination in aquatic environments. However, it is unclear whether macrobenthic invertebrates in the food chain of aquatic environments can be indicators of ARG contamination. In this study, we demonstrated that ARGs including tetA gene, sul2 gene, and km gene were distributed in Chironomidae larvae in Weishan Lake. The ARG distribution was related to animal species, body parts, sampling sites, time, urban environment, animal farming, south-to-north water diversion, food chain, antibiotics, and water storage. Mathematical model predictions of ARG contamination in Weishan Lake were constructed based on the structural equation model (SEM) and the distribution of ARG sul2 in Chironomidae larvae. Influencing factors such as water storage, metal elements, antibiotic, and temperature were found to be closely related to the prediction of ARG contamination. This study provided a new indicator for ARG contamination in freshwater environments and a method to predict ARGs contamination.

Protective effect of hawthorn vitexin on the ethanol-injured DNA of BRL-3A hepatocytes.

ABSTRACT: Vitexin is a natural active ingredient in hawthorn leaves, which has a wide range of anti-tumor effects. This study was conducted to assess the protective effect of hawthorn vitexin on the ethanol-injured DNA of hepatocytes in vitro and to explore its mechanism. The effect of different concentrations of hawthorn vitexin on ethanol-injured hepatocytes was detected via the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide method to study the protective effect of hawthorn vitexin on ethanol-injured DNA damage in hepatocytes. Single-cell gel electrophoresis was used to observe the effect of hawthorn vitexin on ethanol-induced DNA damage in hepatocytes, and the Olive tail moment was measured. Cell physiological and biochemical indexes, such as superoxide dismutase activity, malonaldehyde content, and glutathione peroxidase activity, were detected with kits. The mRNA expression of the superoxide dismutase gene was measured via real-time quantitative polymerase chain reaction. It was showed that 0.2, 0.4, and 0.8 mg mL-1 hawthorn vitexin could significantly repair hepatocyte growth and ethanol-induced DNA damage. This effect was closely related to the improvement in superoxide dismutase, malonaldehyde, and glutathione peroxidase. Hawthorn vitexin could be used to repair ethanol-injured hepatocytes through antioxidation effects, and showed potential for the treatment of liver injury.

Preparation of ß-CD-Quercetin Complex and its Effects on Ethanol- Damaged BRL-3A Hepatocytes.

OBJECTIVE: To prepare the sustained-release complex, quercetin was incorporated with ß- cyclodextrin (ß-CD) and the effect of ß-CD-quercetin complex on the growth of ethanol-injuried hepatocytes was studied. METHODS: By using scanning electron microscopy, infrared spectroscopy, and release rate analysis, ß- CD-quercetin complex was identified. The effect of different concentrations of ß-CD-quercetin complex on the growth of ethanol-damaged hepatocytes at different time was observed by using MTT assay, and the cell quantity and morphology were observed by using hematoxylin-eosin staining. By using single-cell gel electrophoresis, the prevention of ß-CD-quercetin complex from the DNA damage of ethanol-damaged BRL-3A cells was studied, and Olive tail moment was calculated. RESULTS: ß-CD-quercetin complex as the sustained-release complex was successfully prepared. The ethanol induced damage of BRL-3A cells could be prevented by 20, 40 and 80 mg/L of quercetin complex, and the protection mechanism of hepatocyte was related to the antioxidation of DNA. CONCLUSION: Quercetin sustained-release complex could be prepared with ß-CD, and it might be used to treat alcoholic liver disease.

Preparation of Chitosan/Alginate-ellagic Acid Sustained-release Microspheres and their Inhibition of Preadipocyte Adipogenic Differentiation.

OBJECTIVE: In this study, chitosan/alginate-ellagic acid sustained-release microspheres were prepared, and the effect of sustained-release microspheres on preadipocyte adipogenic differentiation was analyzed. METHODS: Chitosan/alginate-ellagic acid microspheres were prepared and identified by scanning electron microscopy (SEM) and infrared spectroscopy (IR). The drug release rates were measured at pH 6.8, 7.0, 7.2, 7.4 to determine sustained release of ellagic acid from microspheres. The effects of 0.1, 1, 10 mg/L chitosan/alginate-ellagic acid microsphere on 3T3-F442A preadipocyte proliferation were determined by Methyl thiazolyl tetrazolium assay (MTT), and cell morphology was checked by hematoxylin/ eosin staining (HE staining). The effect of chitosan/alginate-ellagic acid microspheres on preadipocyte adipogenic differentiation was also determined by Oil red O staining, and lipogenesis was measured by isopropanol extraction. The molecular mechanism was investigated by detecting the mRNA expression of CCAAT/enhancer binding protein alpha (C/EBPα) and peroxisome proliferatorsactivated receptor gamma (PPARγ). RESULTS: Chitosan/alginate-ellagic acid sustained-release microspheres were successfully prepared, and the inhibition of proliferation and adipogenic differentiation of preadipocytes was found to be dosedependent. The mechanism of differentiation inhibition was found to be closely related to the expression of transcription factor C/EBPα and PPARγ. CONCLUSION: Chitosan/alginate can be used as a good material to prepare ellagic acid sustained-release microspheres, and these microspheres can be used for treating the obesity.

Self-defense of Escherichia coli against damages caused by nanoalumina.

Although studies showed effects of nanoalumina (nano-Al2O3) on Escherichia coli, no study completely provides understanding on how bacterial cells respond to damages, especially on how they initiate self-defense. In this study, we showed three types of responses of E. coli to damages caused by nano-Al2O3. Live, dead, and injured, bacteria showed improved survival rates reaching 104%, 116%, and 104% after exposure to 0.1, 1, and 10mmol/L of nano-Al2O3 respectively. Survival rates improved from 100% to 114%, corresponding to an exposure time of 0-9h, and from 100% to 127%, corresponding to 0-1000µg/L Al3+. Improvements were noted in survival rates of E. coli K12 MG1655, HB101, DH5α, and K12 MG1655 △lexA treated by nano-Al2O3 in Luria-Bertani (LB) exposure system or K12 MG1655 in LB, normal saline(NS) and H2O exposure system. Bacterial cells transformed from long rods to ellipsoidal or nearly spherical as form of self-preservation mechanism; this phenomenon may be related to changes in membrane potential induced by free Al3+ released from nano-Al2O3 particles. Molecular mechanism of this response involved inhibited gene expression of sythesis and metabolism of carbohydrates, lipids and proteins. Findings presented in this study may improve understanding of potential danger of nanomaterials and control their spread to environmen.

Polycyclic aromatic hydrocarbons (PAHs) in soils and vegetation near an e-waste recycling site in South China: concentration, distribution, source, and risk assessment.

This study determined the concentrations of PAHs generated from e-waste recycling activities and their potential impacts on soil, vegetation, and human health. The total PAH concentrations in soils and plants ranged from 127 to 10,600 and 199 to 2420 ng/g, respectively. Samples from an e-waste burning site had higher PAH concentrations than samples from adjacent locations. The PAHs in plants varied with plant species and tissue, and Lactuca sativa L. contained the highest PAHs of all the vegetable species. Various land use types showed different PAH concentrations in soils, with vegetable fields showing higher concentrations than paddy fields. Low molecular weight PAHs, such as phenanthrene, were the predominant congeners in soils, whereas high molecular weight PAHs, such as fluoranthene, pyrene, and benzo[a]anthracene, were enriched in plants relative to soils. Dissimilar PAH profiles in soil and the corresponding vegetation indicated that the uptake of PAHs by plants was selective. A source analysis showed that the contamination by PAHs originated primarily from the open burning of e-waste. The total daily intakes of PAHs and carcinogenic PAHs through vegetables at the e-waste dismantling site were estimated to be 279 and 108 ng/kg/d, respectively, indicating that the consumption of vegetables grown near e-waste recycling sites is risky and should be completely avoided.