C-Phycocyanin exerts anti-cancer effects via the MAPK signaling pathway in MDA-MB-231 cells

Abstract Background Triple-negative breast cancer is a biological subtype of breast cancer, which is unresponsive to conventional chemotherapies and has a poor prognosis. C-Phycocyanin (C-PC), a marine natural purified from Spirulina platensis, has been investigated that has anti-cancer function. The mitogen activated protein kinase (MAPK) pathway plays a crucial role in the development and progression of cancer. Therefore, we would like to study the anti-cancer effects of C-phycocyanin in the treatment of triple-negative breast cancer, and explore the role of MAPK pathway in the anti-tumor effects of C-phycocyanin. Methods Cell proliferation, cell cycle, cell apoptosis and cell migration were explored in breast cancer MDA-MB-231 cell lines. AKT, MAPK and membrane death receptor signaling were evaluated in MDA-MB-231 cell lines. Results Our study indicated that C-phycocyanin inhibited cell proliferation and reduced colony formation ability of MDA-MB-231 cells. Furthermore, C-phycocyanin induced cell cycle G0/G1 arrest by decreasing protein expression levels of Cyclin D1 and CDK-2 and increasing protein expression levels of p21 and p27. In addition, C-phycocyanin induced cell apoptotic by activating cell membrane surface death receptor pathway. Besides, C-phycocyanin down-regulated the protein expression levels of cyclooxygenase-2, and further inhibited MDA-MB-231 cells migration. We also found cell death induced by C-phycocyanin was carried through the MAPK signaling pathways. C-Phycocyanin was able to induce MDA-MB-231 cell apoptosis by activating p38 MAPK and JNK signaling pathways while inhibiting ERK pathway. Conclusions C-Phycocyanin exerted anti-cancer activity via the MAPK signaling pathway in MDA-MB-231 cells

Clinical and Pathological Analysis of 4910 Patients Who Received Renal Biopsies at a Single Center in Northeast China

Purpose. To identify the epidemiology and pathological types of kidney diseases and their changes during the past decade, in a population from Northeast China. Methods. We retrospectively analysed clinical and renal pathological data from 4910 patients who received renal biopsies in the Second Hospital of Jilin University from 2008 to 2017. Results. Males received more renal biopsies than females (p < 0.001). The average age (p < 0.001) and percentage of elderly patients (p < 0.001) increased over time. The pathological types were primary glomerulonephritis (PGN, 73.2%), secondary glomerulonephritis (SGN, 23.7%), tubular-interstitial nephropathy (TIN, 2.8%), and hereditary nephropathy (HN, 0.3%). The most common forms of PGN were membranous nephropathy (MN, 37.2%) and IgA nephropathy (IgAN, 29.9%). Over time, the prevalence of IgAN decreased, but the prevalence of MN increased. MN was more common in middle-aged and elderly patients, but IgAN was most common in young adults. Analysis of SGN data indicated that lupus nephritis (LN, 34.0%), Henoch-Schönlein purpura glomerulonephritis (HSPN, 17.9%), and diabetic nephropathy (DN, 11.7%) were the most common forms. Over time, the prevalence of DN (p = 0.003), hypertension-associated renal damage (p = 0.005), and systemic vasculitis-associated nephritis (SVARD, p < 0.001) increased, but the prevalence of HSPN (p < 0.001) and hepatitis B virus-associated glomerulonephritis (HBV-GN, p = 0.001) decreased. Nephrotic syndrome was the main clinical manifestation of PGN. Conclusion. From 2008 to 2017, renal biopsies were increasingly performed in the elderly. There were notable changes in the epidemiology and pathological types of kidney disease among renal biopsy patients at our centre

Development and screening of EMS mutants with altered seed oil content or fatty acid composition in Brassica napus

Brassica napus is an important oilseed crop in the world and the mechanism of seed oil biosynthesis in B. napus remains unclear. In order to study the mechanism of oil biosynthesis and generate germplasms for breeding, an EMS mutant population with ~100,000 M2 lines was generated using Zhongshuang 11 as the parent line. The EMS-induced genome-wide mutations in M2-M4 plants were assessed. The average number of mutations including single nucleotide polymorphisms and insertion/deletion in M2-M4 was 21,177, 28,675 and 17,915, respectively. The effects of the mutations on gene function were predicted in M2-M4 mutants, respectively. We screened the seeds from 98,113 M2 lines and 9,415 seed oil content and fatty acid mutants were identified. We further confirmed 686 mutants with altered seed oil content and fatty acid in advanced generation (M4 seeds). Five representative M4 mutants with increased oleic acid were re-sequenced and the potential causal variations in FAD2 and ROD1 genes were identified. This study generated and screened a large scale of B. napus EMS mutant population and the identified mutants could provide useful genetic resources for the study of oil biosynthesis and genetic improvement of seed oil content and fatty acid composition of B. napus in the future.Accepted versionThis research was supported by The National Key Research and Development Program of China (2016YFD0101000) and the National Natural Science Foundation of China (31871658)

Interaction between phenylpropane metabolism and oil accumulation in the developing seed of Brassica napus revealed by high temporal-resolution transcriptomes

Abstract Background Brassica napus is an important oilseed crop providing high-quality vegetable oils for human consumption and non-food applications. However, the regulation between embryo and seed coat for the synthesis of oil and phenylpropanoid compounds remains largely unclear. Results Here, we analyzed the transcriptomes in developing seeds at 2-day intervals from 14 days after flowering (DAF) to 64 DAF. The 26 high-resolution time-course transcriptomes are clearly clustered into five distinct groups from stage I to stage V. A total of 2217 genes including 136 transcription factors, are specifically expressed in the seed and show high temporal specificity by being expressed only at certain stages of seed development. Furthermore, we analyzed the co-expression networks during seed development, which mainly included master regulatory transcription factors, lipid, and phenylpropane metabolism genes. The results show that the phenylpropane pathway is prominent during seed development, and the key enzymes in the phenylpropane metabolic pathway, including TT5, BAN, and the transporter TT19, were directly or indirectly related to many key enzymes and transcription factors involved in oil accumulation. We identified candidate genes that may regulate seed oil content based on the co-expression network analysis combined with correlation analysis of the gene expression with seed oil content and seed coat content. Conclusions Overall, these results reveal the transcriptional regulation between lipid and phenylpropane accumulation during B. napus seed development. The established co-expression networks and predicted key factors provide important resources for future studies to reveal the genetic control of oil accumulation in B. napus seeds

Na4P2O7-Modified Biochar Derived from Sewage Sludge: Effective Cu(II)-Adsorption Removal from Aqueous Solution

With the rapid development of industrialization, the amount of copper-containing wastewater is increasing, thereby posing a threat to the aquatic ecological environment and human health. Sludge biochar has received extensive concern in recent years due to its advantages of low cost and sustainability for the treatment of heavy-metal-containing wastewater. However, the heavy-metal-adsorption capacity of sludge biochar is limited. This study prepared a sodium pyrophosphate- (Na4P2O7-) modified municipal sludge-based biochar (SP-SBC) and evaluated its adsorption performance for Cu(II). Results showed that SP-SBC had higher yield, ash content, pH, Na and P content, and surface roughness than original sewage sludge biochar (SBC). The Cu(II)-adsorption capacity of SP-SBC was 4.55 times than that of SBC at room temperature. For Cu(II) adsorption by SP-SBC, the kinetics and isotherms conformed to the pseudo-second-order model and the Langmuir–Freundlich model, respectively. The maximum adsorption capacity of SP-SBC was 38.49 mg·g−1 at 35°C. Cu(II) adsorption by SP-SBC primarily involved ion exchange, electrostatic attraction, and precipitation. The desired adsorption performance for Cu(II) in the fixed-bed column experiment indicated that SP-SBC can be reused and had good application potential to treat copper-containing wastewater. Overall, this study provided a desirable sorbent (SP-SBC) for Cu(II) removal, as well as a new simple chemical-modification method for SBC to enhance Cu(II)-adsorption capacity

A Regulatory Feedback Loop between HIF-1α and PIM2 in HepG2 Cells

<div><p>To survive under hypoxic conditions, cancer cells remodel glucose metabolism to support tumor progression. HIF transcription factor is essential for cellular response to hypoxia. The underlying mechanism how HIF is constitutively activated in cancer cells remains elusive. In the present study, we characterized a regulatory feedback loop between HIF-1α and PIM2 in HepG2 cells. Serine/threonine kinase proto-oncogene PIM2 level was induced upon hypoxia in a HIF-1α-mediated manner in cancer cells. HIF-1α induced PIM2 expression via binding to the hypoxia-responsive elements (HREs) of the PIM2 promoter. In turn, PIM2 interacted with HIF-1α, especially a transactivation domain of HIF-1α. PIM2 as a co-factor but not an upstream kinase of HIF-1α, enhanced HIF-1α effect in response to hypoxia. The positive feedback loop between PIM2 and HIF-1α was correlated with glucose metabolism as well as cell survival in HepG2 cells. Such a regulatory mode may be important for the adaptive responses of cancer cells in antagonizing hypoxia during cancer progression.</p></div

Schematic diagram of the proposed hypoxia-induced PIM2 signaling pathway.

<p>Schematic diagram of the proposed hypoxia-induced PIM2 signaling pathway.</p

PIM2 augments HIF-1α–mediated transcription under hypoxia.

<p>(A) HEK293T cells were transfected Flag-tagged PIM2 with HA-tagged HIF-1α (575-826aa) for 24 h and cultured under hypoxia for a further 24 h. Co-IP assays were performed with anti-HA antibody, followed by immunoblot assays. (B) GST pull-down assays were performed with GST, GST-tagged HIF-1α (575-826aa) and His-tagged PIM2 proteins which were purified from <i>Ecoli</i> expressing system. (C and D) HepG2 cells were transfected empty vector or Flag-tagged PIM2 (C); scramble siRNA or PIM2 siRNA (D) for 24 h and cultured under normoxia or hypoxia for a further 24 h. Protein Levels of PIM2 were determined in immunoblot assays using the indicated antibodies. (E and F) HepG2 cells were transfected empty vector or Flag-tagged PIM2 (E); scramble siRNA or PIM2 siRNA (F) with p2.1 luciferase reporter plasmid for 24 h and cultured under normoxia or hypoxia for a further 24 h before luciferase activity was measured. Transfection efficiency was normalized against Renilla luciferase expression. (G and H) HepG2 cells were transfected with empty vector or Flag-tagged PIM2 (G); scramble siRNA or PIM2 siRNA (H) for 24 h and cultured under normoxia or hypoxia for a further 24 h. mRNA levels of <i>Glut1</i>, <i>ENO1</i>, <i>VEGF</i> and <i>LDHA</i> were determined by real-time PCR assays. All data represent the means ± SEM of three independent experiments, *<i>p</i><0.05, **<i>p</i><0.01.</p

PIM2 is a HIF-1α target gene.

<p>(A) Schematic representation of the hypoxic response element (HRE) binding sites in the PIM2 promoter region. Sequences for putative binding sites and their positions were indicated. (B) HEK293T cells were transfected with HRE-luciferase reporter plasmids (P1, P2 or P3) under normoxia or hypoxia. After 48 h, luciferase activity was measured. Transfection efficiency was normalized against Renilla luciferase expression. (C) HEK293T cells were transfected with HRE-luciferase reporter plasmids (P4 or P5) under normoxia or hypoxia. After 48 h, luciferase activity was measured. Transfection efficiency was normalized against Renilla luciferase expression. (D) Schematic diagrams of the regulating sequences with the putative HREs of PIM2. Mut: CG was changed into AA. HEK293T cells were transfected with the HRE (+355mu), HRE (+458mu), HRE (+635mu) or HRE (+789mu)-luciferase reporter plasmids under normoxia or hypoxia. After 48h, luciferase activity was measured. Transfection efficiency was normalized against Renilla luciferase expression. (E and F) Anti-IgG, anti-HIF-1α (E) or anti-HIF-2α (F) antibodies were used in the chromatin immunoprecipitation (ChIP) assays using HEK293T cells which were treated with hypoxia for 12 h. All data represent the means ± SEM of three independent experiments, *<i>p</i><0.05, **<i>p</i><0.01.</p

PIM2 augments glycolysis and cell survival under hypoxia.

<p>(A and B) HepG2 cells were transfected with empty vector or Flag-tagged PIM2 for 24 h and cultured under normoxia or hypoxia for a further 12–16 h. Levels of glucose (A) or lactate (B) in mediums were determined. (C and D) HepG2 cells were transfected with scramble siRNA or PIM2 siRNA for 24 h and cultured under normoxia or hypoxia for a further 12–16 h. Levels of glucose (C) or lactate (D) in mediums were determined. (E). HepG2 cells were transfected with scramble siRNA or PIM2 siRNA. After 24 h, the cells were re-plated and cultured under normoxia or hypoxia for a further 72 h. Cell numbers were counted every 24 h for the analysis of cell proliferation. (F) HepG2 cells were transfected with scramble siRNA or PIM2 siRNA. After 24 h, the cells were re-plated and cultured under normoxia or hypoxia for a further 24 h. The apoptosis changes were determined using the flow cytometry BD FaCSAria3 apparatus. All data represent the means ± SEM of three independent experiments, *<i>p</i><0.05, **<i>p</i><0.01.</p