Differences in Glycerolipid Response of Chlamydomonas reinhardtii Starchless Mutant to High Light and Nitrogen Deprivation Stress Under Three Carbon Supply Regimes.

Carbon source serves as a crucial factor for microalgal lipid biosynthesis. The supplied exogenous inorganic or organic carbon affects lipid accumulation in microalgae under stress conditions. However, the impacts of different carbon availability on glycerolipid metabolism, triacylglycerol (TAG) metabolism in particular, still remain elusive in microalgae. Chlamydomonas starchless mutant BAFJ5 has emerged as a model system to study TAG metabolism, due to its property of hyper-accumulating TAG. In this study, the glycerolipidomic response of the starchless BAFJ5 to high light and nitrogen-deprived (HL-N) stress was deciphered in detail to distinguish glycerolipid metabolism under three carbon supply regimes. The results revealed that the autotrophically and mixotrophically grown BAFJ5 cells aerated with air containing 2% CO2 presented similar changes in growth, photosynthetic activity, biochemical components, and glycerolipid metabolism under HL-N conditions. But the mixotrophically grown BAFJ5 aerated with air containing 0.04% CO2 exhibited more superior accumulation in TAG, which was esterified with a significantly higher proportion of C18:1n9 and prominently the lower proportions of polyunsaturated fatty acids. In addition, these cells increased the relative levels of C18:2n6 in the membrane lipids, i.e., monogalactosyldiacylglycerol (MGDG) and digalactosyldiacylglycerol (DGDG), in priority, and decreased that of C18:3n3 and C18:4n3 in the betaine lipid, N,N,N-trimethylhomoserine diacylglycerol (DGTS), subsequently, to adapt to the HL-N stress conditions, compared to the cells under the other two conditions. Thus, it was suggested that C. reinhardtii starchless mutant appeared to present distinct metabolism for TAG biosynthesis involving membrane lipid remodeling under distinct carbon supply regimes. This study provides insights into how the different carbon supply regimes affect lipid metabolism in Chlamydomonas starchless cells, which will benefit the optimized production of storage lipids in microalgae.

Achieving Partial Nitrification-Anammox Process Dependent on Microalgal-Bacterial Consortia in a Photosequencing Batch Reactor.

Partial nitrification coupled with anammox (PN/A) process is an energy-efficient approach for nitrogen removal from low C/N wastewater. In this study, PN/A was achieved with optimal oxygen supply from a green microalga, Chlorella sorokiniana. The PN process was first initiated within 35 days, and the following algae-intensified PN then reached the steady state within the next 32 days. The dissolved oxygen (DO) concentration was gradually maintained at 0.6 mg L-1 via adjusting the photoperiod to 6-h light/18-h dark cycles, when the accumulation ratio of NO2 --N and the removal ratio of NH4 +-N were both more than 90%. The nitrogen removal capability of anammox was acclimated via elevating the individual effluent NH4 +-N and NO2 --N levels from 100 to 200, to 300 mg L-1. After acclimation, the removal rates of NH4 +-N and total nitrogen (TN) reached more than 70 and 80%, respectively, and almost all the NO2 --N was removed. Then, the algae-intensified PN/A, algammox biofilm system, was successfully started up. When the NH4 +-N level increased from 100 to 300 mg L-1, the TN removal varied between 78 and 82%. In the photosequencing bioreactor, C. sorokiniana, ammonia-oxidizing bacteria (AOB), and anammox coexisted with an illumination of 200 µmol m-2 s-1 and a 6-h light/18-h dark cycles. The DO levels ranged between 0.4 and 0.5 mg L-1. In addition, the microbial community analysis by Illumina MiSeq sequencing showed that the dominant functional bacteria in the algae-intensified PN/A reactors included Nitrosomonas (AOB) and Candidatus Brocadia (anammox), while Nitrospira and Nitrobacter (nitrite oxidizing bacteria), together with Denitratisoma (denitrifier) were largely inhibited. Further studies are required to optimize the microalgal-bacterial consortia system to achieve superior nitrogen removal rates under controllable conditions.

Inhibitory effect of luteolin on the proliferation of human breast cancer cell lines induced by epidermal growth factor.

The aim of the present study was to investigate the mechanism of the inhibitory effect of luteolin on the proliferation of breast cancer cells induced by epidermal growth factor (EGF) in vitro. MTT assay was used to detect the inhibitory effect of luteolin on the proliferation of MCF-7 and MDA-MB-231 cells as well as the effect on the proliferation of MCF-7 cells induced by EGF. Western blotting was used to detect the effects of luteolin on the expression of epidermal growth factor receptor (EGFR), phosphatidylinositol 3-kinase (PI3K)/Akt, mitogen-activated protein kinase (MAPK)/extracellular-signal-regulated kinases (Erk) 1/2 and signal transducers and activators of transcription-3 (STAT3) in MCF-7 cells induced by EGF. The results showed that luteolin could significantly inhibit the proliferation of MCF-7 and MDA-MB-231 cells, and the inhibitory effect on MCF-7 cells was more prominent. Moreover, luteolin could inhibit the proliferation of MCF-7 cells induced by EGF. Western blotting results showed that luteolin and AG1478 (an inhibitor of EGFR signaling) could inhibit the expression of p-EGFR and p-STAT3 in MCF-7 cells induced by EGF. Luteolin, LY294002 (an inhibitor of Akt signaling) and PD98059 (an inhibitor of Erk1/2 signaling) could inhibit the expression of p-Akt and p-Erk1/2 respectively in MCF-7 cells induced by EGF. Our data suggest that luteolin may inhibit EGF-induced activities of EGFR signaling pathway in human breast cancer cell lines, and PI3K/Akt, MAPK/Erk1/2, STAT3 signal pathways may be the major pathways that mediate the inhibitory effect of luteolin on EGFR signaling. Overall, our results may provide a theoretical foundation for the development of luteolin as anti-tumor drug.

Emodin inhibits breast cancer cell proliferation through the ERα-MAPK/Akt-cyclin D1/Bcl-2 signaling pathway.

BACKGROUND: The aim of the present study was to investigate the involvement of emodin on the growth of human breast cancer MCF-7 and MDA-MB-231 cells and the estrogen (E2) signal pathway in vitro. MATERIALS AND METHODS: MTT assays were used to detect the effects of emodin on E2 induced proliferation of MCF-7 and MDA-MB-231 cells. Flow cytometry (FCM) was applied to determine the effect of emodin on E2-induced apoptosis of MCF-7 cells. Western blotting allowed detection of the effects of emodin on the expression of estrogen receptor α, cyclin D1 and B-cell lymphoma-2 (Bcl-2), mitogen-activated protein kinases (MAPK) and phosphatidylinostiol 3-kinases (PI3K). Luciferase assays were emplyed to assess transcriptional activity of ERα. RESULTS: Emodin could inhibit E2-induced MCF-7 cell proliferation and anti-apoptosis effects, and arrest the cell cycle in G0/G1 phase, further blocking the effect of E2 on expression and transcriptional activity of ERα. Moreover, Emodin influenced the ER α genomic pathway via downregulation of cyclin D1 and Bcl-2 protein expression, and influenced the non-genomic pathway via decreased PI3K/Akt protein expression. CONCLUSIONS: These findings indicate that emodin exerts inhibitory effects on MCF-7 cell proliferation via inhibiting both non-genomic and genomic pathways.

[Inhibitory effect of luteolin on the angiogenesis of chick chorioallantoic membrane and invasion of breast cancer cells via downregulation of AEG-1 and MMP-2].

The purpose of the present study was to investigate the effect of luteolin on the angiogenesis and invasion of breast cancer cells. MTT assay was used to examine breast cancer proliferation. The chick chorioallantoic membrane model was used to assess the angiogenesis effect. Wound healing assay was used to assess cell invasion ability. Western blot was used to analyze Bcl-2, AEG-1 and MMP-2 expression levels. The results showed luteolin inhibited MCF-7 cells proliferation in a dose- and time-dependent manner, and the expression of Bcl-2 protein was decreased. Luteolin had a strong anti-angiogenesis of chick chorioallantoic membrane. After treatment of MCF-7 cells with luteolin at 60 µmol/L for 48 h, migration rate was reduced by 71.07% compared with control (P < 0.01). After treatment of MCF-7 cells with luteolin at 60 µmol/L for 48 h, the expression of AEG-1 and MMP-2 was reduced by 82.34% (P < 0.05) and 85.70% (P < 0.05) respectively, compared with control. In conclusion, the results suggest that luteolin can inhibit the proliferation of breast cancer cells, and suppress the expression of Bcl-2. Furthermore, luteolin has strong anti-angiogenesis of chick chorioallantoic membrane and anti-invasive activity on breast cancer cells, and down-regulates the expression of AEG-1 and MMP-2.

[Relationship between the inhibitory effect of fraxetin on breast cancer and estrogen signaling pathway].

Estrogen signaling pathways play an important role in the regulation of the physiological function of breast cancer cell proliferation and apoptosis. The article used MTT assay, flow cytometer analysis and Western blot to detect the inhibition of fraxetin on MCF-7 cell cycle distribution and apoptosis, ERα, cyclin D1 and Bcl-2 expression levels, MAPK and PI3K signaling pathway to investigate the mechanism of anti-breast cancer of fraxetin. The results showed fraxetin inhibited E2ß-stimulated MCF-7 cell proliferation in a dose- and time-dependent manner, reversed E2ß-induced anti-apoptosis and promoted G0/G1 phase arrest. After treatment with fraxetin, the expression of ERα in MCF-7 cell was decreased, and estrogen genomic signaling pathway was inhibited by down-regulating the expression of cyclin D1 and Bcl-2 proteins. After MCF-7 cells were treated with fraxetin, the expressions of MAPK/Erk1/2 protein were reduced, which affected estrogen non-genomic signaling pathway. The results suggest fraxetin plays a part in anti-breast cancer function through E2ß-mediated genomic and non-genomic signaling pathways.

Luteolin inhibits proliferation induced by IGF-1 pathway dependent ERα in human breast cancer MCF-7 cells.

The growth of many breast tumors is stimulated by IGF-1, which activates signal transduction pathways inducing cell proliferation. ERα is important in this process. The aim of the study was to investigate relationships in vitro among inhibitory effects of luteolin on the growth of MCF-7 cells, IGF-1 pathway and ERα. Our results showed that luteolin could effectively block IGF-1-stimulated MCF-7 cell proliferation in a dose- and time- dependent manner and block cell cycle progression and induce apoptosis evidenced by the flow cytometric detection of sub-G1DNA content. Luteolin markedly decreased IGF-1-dependent IGF-1R and Akt phosphorylation without affecting Erk1/2 phosphorylation. Further experiments pointed out that ERα was directly involved in IGF-1 induced cell growth inhibitory effects of luteolin, which significantly decreased ERα expression. Knockdown of ERα in MCF-7 cells by an ERα-specific siRNA decreased the IGF-1 induced cell growth inhibitory effects of luteolin. ERα is thus a possible target of luteolin. These findings indicate that the inhibitory effect of luteolin on the growth of MCF-7 cells is via inhibiting IGF-1 mediated PI3K-Akt pathway dependent on ERα expression.

[Wogonin inhibits IGF-1-stimulated cell growth and estrogen receptor α expression in breast adenocarcinoma cell and angiogenesis of chick chorioallantoic membrane].

The aim of the present study was to investigate the involvements of insulin-like growth factor-1 (IGF-1) and estrogen receptor α (ERα) in the inhibitory effect of wogonin on the breast adenocarcinoma growth. Moreover, the effect of wogonin on the angiogenesis of chick chorioallantoic membrane (CAM) was also investigated. MCF-7 cells (human breast adenocarcinoma cell line) were subjected to several drugs, including IGF-1, wogonin and ER inhibitor ICI182780, alone or in combination. MTT assay was used to detect breast cancer proliferation. Western blot was used to analyze ERα and p-Akt expression levels. CAM models prepared from 6-day chicken eggs were employed to evaluate angiogenesis inhibition. The results showed wogonin and ICI182780 both exhibited a potent ability to blunt IGF-1-stimulated MCF-7 cell growth. Either of wogonin and ICI182780 significantly inhibited ERα and p-Akt expressions in IGF-1-treated cells. The inhibitory effect of wogonin showed no difference from that of ICI182780 on IGF-1-stimulated expressions of ERα and p-Akt. Meanwhile, wogonin at different concentrations showed significant inhibitory effect on CAM angiogenesis. These results suggest the inhibitory effect of wogonin on breast adenocarcinoma growth via inhibiting IGF-1-mediated PI3K-Akt pathway and regulating ERα expression. Furthermore, wogonin has a strong anti-angiogenic effect on CAM model.

[Antibacterial activity and mechanism of baicalein].

Baicalein (BAI) is an effective bactericide. The antibacterial activity and mechanism experiments were carried out by determining conductivity and content of macromolecules of membrane penetrability, the oxidative respiratory metabolism and protein synthesis changes and the inhibition of DNA topoisomerase activities. Electrical conductivity and the number of large molecules of BAI increased 2.48% and 1.8%, respectively, than that of the control. However, the membrane integrity did not destroyed by BAI directly. With BAI treatment, inhibition rates of activities for SDH and MDH were 56.2% and 57.4%, respectively, demonstrating that BAI could inhibit cell respiratory. After treated with BAI for 20 h, the total soluble content of proteins decreased by 42.83%. Moreover, the activities of DNA topoisomerase I and II were inhibited completely by 0.2 mmol x L(-1) BAI. These results indicated that BAI had obvious antibacterial activity on Staphylococcus aureus. The mechanism is that it could affect bacterial membrane penetrability, inhibit protein synthesis and influence SDH, MDH and DNA topoisomerase I and II activities to exert its antibacterial functions.