Enhanced aerobic granular sludge with micro-electric field for sulfamethoxazole degradation: Efficiency, mechanism, and microbial community.

In this study, an aerobic granular sludge electrochemical system (AGES) was established by applying the micro-electric field to an aerobic granular sludge (AGS) reactor for the degradation of sulfamethoxazole (SMZ). Under the stimulation of the micro-electric field, the granulation of sludge was improved and the degradation rate of SMZ was enhanced. The features of granular sludge were characterized by scanning electron microscopy and X-ray diffraction. The optimal degradation rate of SMZ (88%) was obtained at the voltage of 3 V and the effective electrode area of 800 mm2. The results of kinetics analyses revealed that the degradation of SMZ by AGES can be fitted with the second-order kinetic equation, showing a degradation rate constant (k) of 0.001 L mol-1·min-1. The degradation products of SMZ in the AGES system were detected by LC-MS and their possible degradation routes were elucidated. The micro-electric field in the AGES system played a selective role in microbes' enrichment and growth, changing the diversity of the microbial community. Pseudomonas, Tolumonas, and Acidovorax were the dominant bacteria in the AGES system, which is accountable for the abatement of SMZ and nutrients. This work provides a green means for improving AGS and paves the way for applying the AGS process to real-world wastewater treatment.

The MKK3-MPK7 cascade phosphorylates ERF4 and promotes its rapid degradation to release seed dormancy in Arabidopsis.

Seeds establish dormancy to delay germination until the arrival of a favorable growing season. In this study, we identify a fate switch comprised of the MKK3-MPK7 kinase cascade and the ethylene response factor ERF4 that is responsible for the seed state transition from dormancy to germination. We show that dormancy-breaking factors activate the MKK3-MPK7 module, which affects the expression of some α-EXPANSIN (EXPA) genes to control seed dormancy. Furthermore, we identify a direct downstream substrate of this module, ERF4, which suppresses the expression of these EXPAs by directly binding to the GCC boxes in their exon regions. The activated MKK3-MPK7 module phosphorylates ERF4, leading to its rapid degradation and thereby releasing its inhibitory effect on the expression of these EXPAs. Collectively, our work identifies a signaling chain consisting of protein phosphorylation, degradation, and gene transcription , by which the germination promoters within the embryo sense and are activated by germination signals from ambient conditions.

Glutathione S-transferase P1 suppresses iNOS protein stability in RAW264.7 macrophage-like cells after LPS stimulation.

Glutathione S-transferase P1 (GSTP1) is a ubiquitous expressed protein which plays an important role in the detoxification and xenobiotics metabolism. Previous studies showed that GSTP1 was upregulated by the LPS stimulation in RAW264.7 macrophage-like cells and GSTP1 overexpression downregulated lipopolysaccharide (LPS) induced inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) expression. Here we show that GSTP1 physically associates with the oxygenase domain of iNOS by the G-site domain and decreases the protein level of iNOS dimer. Both overexpression and RNA interference (RNAi) experiments indicate that GSTP1 downregulates iNOS protein level and increases S-nitrosylation and ubiquitination of iNOS. The Y7F mutant type of GSTP1 physically associates with iNOS, but shows no effect on iNOS protein content, iNOS S-nitrosylation, and changes in iNOS from dimer to monomer, suggesting the importance of enzyme activity of GSTP1 in regulating iNOS S-nitrosylation and stability. GSTM1, another member of GSTs shows no significant effect on regulation of iNOS. In conclusion, our study reveals the novel role of GSTP1 in regulation of iNOS by affecting S-nitrosylation, dimerization, and stability, which provides a new insight for analyzing the regulation of iNOS and the anti-inflammatory effects of GSTP1.

[Pilot-scale study on treatment of municipal sewage by moving-bed biofilm reactor with the hydrophobically modified polyurethane cubes as biofilm carriers].

The carrier is the key influencers in moving bed biofilm reactor( MBBR), in this paper, a pilot scale apparatus was set up for treating municipal wastewater using modified cubic polyurethane carriers. For MBBR, the capacity of 3-3.5 t x d(-1), hydraulic residence time of 7-8 h, under the condition of continuous feed water (COD:140-280 mg x L(-1), NH4+ -N:30-50 mg x L(-1), TN: 45-65 mg x L(-1), TP:2.5-4.0 mg x L(-1)), the speed of biofilm formation and removal effects of COD, nitrogen and phosphorus were studied. After 140 days, the results showed that the formation of biofilm on the carrier was very quickly under 24-28 degrees C, and obtained stable treatment effect about 6 days. The COD, NH4+ -N, TN, TP average removal rates were 70%, 97%, 70%, 39%, respectively. As the temperature gradually decreased to about 12 degrees C, a high NH4+ -N removal rate (97%) could still be maintained, which indicating that the modified carrier can be achieved a high nitrification rate at low temperature.

Effects of packing rates of cubic-shaped polyurethane foam carriers on the microbial community and the removal of organics and nitrogen in moving bed biofilm reactors.

The effects of packing rates (20%, 30%, and 40%) of polyurethane foam (PUF) to the removal of organics and nitrogen were investigated by continuously feeding artificial sewage in three aerobic moving bed biofilm reactors. The results indicated that the packing rate of the PUF carriers had little influence on the COD removal efficiency (81% on average). However, ammonium removal was affected by the packing rates, which was presumably due to the different relative abundances of nitrifying bacteria. A high ammonium removal efficiency of 96.3% at a hydraulic retention time of 5h was achieved in 40% packing rate reactor, compared with 37.4% in 20% packing rate. Microprofiles of dissolved oxygen and nitrate revealed that dense biofilm limits the DO transfer distance and nitrate diffusion. Pyrosequencing analysis of the biofilm showed that Proteobacteria, Bacteroidetes and Verrucomicrobia were the three most abundant phyla, but the proportions of the microbial community varied with the packing rate of the PUF carriers.

Inhibition of cadmium-induced apoptosis by glutathione S-transferase P1 via mitogen-activated protein kinases and mitochondrial pathways.

Cadmium is a well-known toxic metal for the kidney. Glutathione S-transferase P1 (GSTP1) plays an important role in the detoxification and xenobiotics metabolism. Here, we investigated whether GSTP1 affected Cd(2+)-induced apoptotic cell death in human embryonic kidney cell line (HEK) 293 cells. We showed that in HEK293 cells, silencing of GSTP1 expression through RNA interference reinforced the loss in cell viability induced by Cd(2+). Overexpression of GSTP1 inhibited loss of mitochondrial membrane potential, prevented cytochrome c release from mitochondria and caspase-3 activation, inhibited mitogen-activated protein kinases (MAPKs) including ERK, JNK and p38, and suppressed apoptosis induced by Cd(2+). The oligonucleosomal DNA fragmentation assay also demonstrated that overexpression of GSTP1 by adenovirus infection prevented Cd(2+)-induced apoptosis in primary renal tubule cells. Our data suggest that GSTP1 was an endogenous inhibitor of Cd(2+)-induced apoptosis.

CHIP facilitates ubiquitination of inducible nitric oxide synthase and promotes its proteasomal degradation.

Inducible nitric oxide synthase (iNOS) is responsible for nitric oxide (NO) synthesis from l-arginine in response to inflammatory mediators. It is reported that iNOS is degraded mainly by the ubiquitin-proteasome pathway in RAW264.7 cells and human embryonic kidney (HEK) 293 cells. In this study, we showed that iNOS was ubiquitinated and degraded dependent on CHIP (COOH terminus of heat shock protein 70-interacting protein), a chaperone-dependent ubiquitin ligase. The results from overexpression and RNAi experiments demonstrated that CHIP decreased the protein level of iNOS, shortened the half-life of iNOS and attenuated the production of NO. Furthermore, CHIP promoted ubiquitination and proteasomal degradation of iNOS by associating with iNOS. These results suggest that CHIP plays an important role in regulation iNOS activity.

Chlorogenic acid inhibits lipopolysaccharide-induced cyclooxygenase-2 expression in RAW264.7 cells through suppressing NF-kappaB and JNK/AP-1 activation.

Chlorogenic acid (CGA) is a naturally occurring phenolic acid in human diet. Data obtained from in vivo and in vitro experiments demonstrate that CGA mostly presents anti-oxidant and anti-carcinogenic activities. Here we show that CGA also inhibits lipopolysaccharide (LPS)-induced inflammatory response[AU1] in RAW 264.7 cells. Our results indicated that CGA significantly decreased LPS-induced up-regulation of cyclooxygenase (COX-2) at protein and mRNA levels in RAW 264.7 cells and as a result it inhibited prostaglandin E(2) (PGE(2)) release from LPS-treated RAW 264.7 cells. In the further experiments, LPS-induced activation of nuclear factor-kappaB (NF-kappaB) and c-Jun N-terminal kinase (JNK)-c-Jun-activator protein (AP-1) pathway were suppressed significantly by CGA. In addition, CGA did not affect phosphorylation of extracellular signal-regulated kinase 1/2 (ERK1/2) and p38. In conclusion, CGA suppresses LPS-induced COX-2 expression via attenuating the activation of NF-kappaB and JNK/AP-1 signaling pathways suggesting that CGA, the polyphenol compound in our food, could exert anti-inflammatory effects through inhibiting PGE(2) production.

Gamma-linolenic acid induces apoptosis and lipid peroxidation in human chronic myelogenous leukemia K562 cells.

Various polyunsaturated fatty acids, especially gamma-linolenic acid (GLA), inhibit the growth of a variety of tumor cells. Some evidence indicates that polyunsaturated fatty acid can kill cells by apoptosis. In the current study, we tested the apoptotic effect of GLA on human chronic myelogenous leukemia K562 cells. GLA induced K562 cell death in a dose-dependent manner. Typical apoptotic nuclei were shown by staining of K562 cells with DNA-binding fluorochrome Hoechst 33342, characterized by chromatin condensation and nuclear fragmentation. Flow cytometric analysis also demonstrated that GLA caused dose-dependent apoptosis of K562 cells. The apoptosis could be inhibited by a pancaspase inhibitor (z-VAD-fmk), suggesting the involvement of caspases. Further, release of cytochrome c, activation of caspase-3 and cleavage of PARP were found in GLA-induced apoptosis. GLA treatment could also elevate lipid peroxidation in K562 cells, and antioxidant alpha-tocopherol could reverse the cytotoxicity of GLA. The saturated fatty acid SA, which did not exhibit significant increase in lipid peroxidation, also did not induce cytotoxicity. Intracellular GSH was also determined, and there was no marked change of GSH levels in cells after incubation with GLA compared with the control. These results demonstrate that GLA could induce apoptosis in K562 cells. Apoptosis is mediated by release of cytochrome c, activation of caspase-3. Lipid peroxidation may play a role in GLA cytotoxicity.

Gamma-linolenic acid modulates the response of multidrug-resistant K562 leukemic cells to anticancer drugs.

Gamma-linolenic acid (GLA) is known to have selective tumoricidal action. It is also reported that GLA may increase the cytotoxic activity of cancer chemotherapeutic agents in some cancer cells. The present study examined whether GLA pretreatment could modulate the response of multidrug-resistant K562/ADM leukemic cells to anticancer drugs. The cell viability assay results showed that GLA at 10 microg/ml enhanced cell growth inhibition induced by the MDR-type drugs doxorubicin, etoposide and vincristine, but could not enhance or even attenuated cell growth inhibition induced by the non-MDR-type drug cisplatin, mitomycin and fluorouracil in K562/ADM cells. Further flow cytometry results showed that GLA decreased the expression of P-glycoprotein, enhanced the accumulation of doxorubicin and rhodamine 123 in K562/ADM cells and inhibited the efflux of rhodamine 123 from K562/ADM cells, lowered the efflux rate. These results suggest that GLA could modulate the response to anti-cancer agents in P-gp overexpressing multidrug-resistant cells, and the mechanism may be by decreasing the P-gp expression and inhibiting P-gp function.

[Effects of gamma linolenic acid on atherosclerosis induced by cholesterol-rich diet in rats].

OBJECTIVE: To study the effect of gamma linolenic acid (GLA) on atherogenesis in rats. METHOD: Sixty healthy male rats were randomly divided into 6 groups: normal contro 1, fed by normal feed; atherogenesis mode 1, fed by high lipid diet; positive control group 0.9 mg x kg(-1) x d(-1) of lovastatin and group IV 250 mg x kg(-1) x d(-1) duoxikang; high dose of 375 mg x kg(-1) x d(-1) GLA; low dose of 187.5 mg x kg(-1) x d(-1) GLA. After the model group received atherogenic diet for six weeks, serum triglyceride (TG), total cholesterol (TC), low density lipoprotein cholesterol (LDL-C) and high density lipoprotein cholesterol (HDL-C) were detected by enzyme method to confirm the formation of atherogenic. After fed for another five weeks, morphologic atherosclerosis of aorta in rats was observed by HE staining methods. The blood samples were collected and serum TC, TG, HDL-C, LDL-C, T-AOC, HL, LPL, NO, NOS, MDA and GSH were determined. RESULT: GLA attenuated the formation of atherosclerotic plaques, inhibited the level of serum TC, TG, MDA, OX-LDL, NO, NOS, HL, LPL and LDL-C and increased the level of T-AOC. CONCLUSION: GLA might significantly attenuate the development of atherosclerosis in rats fed with high lipid diet through improving the antioxidation capacity of the body.

Induction of apoptosis in K562/ADM cells by gamma-linolenic acid involves lipid peroxidation and activation of caspase-3.

Numerous studies have revealed that gamma-linolenic acid (GLA) possesses effective tumoricidal properties while not inducing damage to normal cells or creating harmful systemic side effects. It can exert anti-tumor efficacy against a variety of cancers including leukemia. However, little is known about the effects of GLA on leukemia resistant to chemotherapy, emerging as a serious clinical problem. The present study tested GLA-induced apoptosis in K562/ADM multidrug-resistant (MDR) leukemic cells and investigated its possible mechanisms. Using cell viability, fluorescent staining of nuclei, flow cytometric Annexin V/PI double staining and lactate dehydrogenase (LDH) release, we found that GLA could inhibit cell growth and induce apoptosis and secondary necrosis. The results showed that incubation with GLA concentrations of 10-60 microg/ml caused a dose- and time-dependent decrease of K562/ADM cell viability, and the IC50 value was 50.5 microg/ml at 24 h and 31.5 microg/ml at 48 h. Flow cytometry using Annexin V/PI double staining assessed apoptosis, necrosis and viability. Typical apoptotic nuclei were shown by staining of K562/ADM cells with DNA-binding fluorochrome Hoechst 33342, characterized by chromatin condensation and nuclear fragmentation. On the other hand, after treated K562/ADM cells with 20 microg/ml GLA for 48 h and with 40 microg/ml GLA for 12 h, the LDH release significantly increased, indicated losses of plasma membrane integrity and presence of necrosis. Further, the inhibition of GLA-induced apoptosis by a pan-caspase inhibitor (z-VAD-fmk) suggested the involvement of caspases. The increase of caspase-3 activity with GLA concentration confirmed its role in the process. The results also showed that the malondialdehyde (MDA) content was also significantly elevated, and antioxidant BHT could block GLA cytotoxity, indicating the cytotoxity induced by GLA may be due to lipid peroxidation.

Induction of apoptosis in pheochromocytoma (PC12) cells exposed to eicosapentaenoic acid in vitro.

The effect of different concentrations of eicosapentaenoic acid (EPA), on rat pheochromocytoma PC12 cells were evaluated using cell viability, lactate dehydrogenase (LDH) activity, flow cytometric DNA analysis and electronic microscopy. A time- and dose-dependent decrease in the cell viability was observed in the cultures of PC12 cells, supplemented with EPA. The incubation with 200 microM EPA for 48 and 72 hr induced a decrease in the cell viability by 53.40 and 53.43%, respectively. Treatment of PC12 cells with EPA induced apoptosis in a concentration-dependent manner, as evidenced by flow cytometry analysis. The highest percentage of apoptotic cells accumulated to 30.32%, following treatment with 200 microM EPA. The LDH levels increased significantly on treatment with 100 and 200 microM EPA, by 144.4 and 197.3%, respectively, compared with the untreated control. In addition, the cell morphology change was also observed by electron microscopy. The results suggest that EPA mediates its effect on the PC12 cells, at least in part, via the induction by apoptosis.