Rhein inhibits angiogenesis and the viability of hormone-dependent and -independent cancer cells under normoxic or hypoxic conditions in vitro

Hypoxia is a hallmark of solid tumors, including breast cancer, and the extent of tumor hypoxia is associated with treatment resistance and poor prognosis. Considering the limited treatment of hypoxic tumor cells and hence a poor prognosis of breast cancer, the investigation of natural products as potential chemopreventive anti-angiogenic agents is of paramount interest. Rhein (4,5-dihydroxyanthraquinone-2-carboxylic acid), the primary anthraquinone in the roots of Cassia alata L., is a naturally occurring quinone which exhibits a variety of biologic activities including anti-cancer activity. However, the effect of rhein on endothelial or cancer cells under hypoxic conditions has never been delineated. Therefore, the aim of this study was to investigate whether rhein inhibits angiogenesis and the viability of hormone-dependent (MCF-7) or -independent (MDA-MB-435s) breast cancer cells in vitro under normoxic or hypoxic conditions. Rhein inhibited vascular endothelial growth factor (VEGF₁₆₅)-stimulated human umbilical vein endothelial cell (HUVEC) tube formation, proliferation and migration under normoxic and hypoxic conditions. In addition, rhein inhibited in vitro angiogenesis by suppressing the activation of phosphatidylinositol 3-kinase (PI3K), phosphorylated-AKT (p-AKT) and phosphorylated extracellular signal-regulated kinase (p-ERK) but showed no inhibitory effects on total AKT or ERK. Rhein dose-dependently inhibited the viability of MCF-7 and MDA-MB-435s breast cancer cells under normoxic or hypoxic conditions, and inhibited cell cycle in both cell lines. Furthermore, Western blotting demonstrated that rhein inhibited heat shock protein 90alpha (Hsp90α) activity to induce degradation of Hsp90 client proteins including nuclear factor-kappa B (NF-κB), COX-2, and HER-2. Rhein also inhibited the expression of hypoxia-inducible factor-1 alpha (HIF-1α), vascular endothelial growth factor (VEGF₁₆₅), epidermal growth factor (EGF), and the phosphorylation of inhibitor of NF-κB (I-κB) under normoxic or hypoxic conditions. Taken together, these data indicate that rhein is a promising anti-angiogenic compound for breast cancer cell viability and growth. Therefore, further studies including in vivo and pre-clinical need to be performed.     

Antioxidant protective effect of flavonoids on linoleic acid peroxidation induced by copper(II)/ascorbic acid system

Antioxidants are compounds that can delay or inhibit lipid oxidation. The peroxidation of linoleic acid (LA) in the absence and presence of Cu(II) ion–ascorbate combinations was investigated in aerated and incubated emulsions at 37 °C and pH 7. LA peroxidation induced by copper(II)–ascorbic acid system followed first order kinetics with respect to hydroperoxides concentration. The extent of copper-initiated peroxide production in a LA system assayed by ferric thiocyanate method was used to determine possible antioxidant and prooxidant activities of the added flavonoids. The effects of three different flavonoids of similar structure, i.e. quercetin (QR), morin (MR) and catechin (CT), as potential antioxidant protectors were studied in the selected peroxidation system. The inhibitive order of flavonoids in the protection of LA peroxidation was: morin > catechin ≥ quercetin, i.e. agreeing with that of formal reduction potentials versus NHE at pH 7, i.e. 0.60, 0.57 and 0.33 V for MR, CT, and QR, respectively. Morin showed antioxidant effect at all concentrations whereas catechin and quercetin showed both antioxidant and prooxidant effects depending on their concentrations. The structural requirements for antioxidant activity in flavonoids interestingly coincide with those for Cu(II)-induced prooxidant activity, because as the reducing power of a flavonoid increases, Cu(II)–Cu(I) reduction is facilitated that may end up with the production of reactive species. The findings of this study were evaluated in the light of structure–activity relationships of flavonoids, and the results are believed to be useful to better understand the actual conditions where flavonoids may act as prooxidants in the preservation of heterogeneous food samples containing traces of transition metal ions.     

壳聚糖ZnSO4的络合物对Bacillus amyloliquefaciens BS20发酵液絮凝回收

目的利用壳聚糖-ZnSO_4作为絮凝剂回收B.amyloliquefaciens BS-20发酵液中产生的芽胞孢子,为益生芽胞杆菌制剂的工业化生产提供新的浓缩方法和参考。方法通过制备壳聚糖-ZnSO_4形成络合物,对B.amyloliquefaciens BS-20孢子发酵液进行絮凝处理,分别探讨絮凝的发酵液初始pH值条件、初始壳聚糖浓度以及壳聚糖-ZnSO_4的最适配比等参数对絮凝效果的影响;并对絮凝回收的芽胞活菌孢子数量进行验证。结果在发酵液初始pH值调整为4.5、壳聚糖浓度为0.5 g/L并且与ZnSO_4按照3∶1的配比进行络合后,能够有效地实现发酵液的菌体絮凝,最终回收后发酵液的浓缩率为90%,益生芽胞杆菌活菌孢子的回收率可达93%。结论该实验结果可以为在液体发酵中产生的芽胞孢子进行快速回收制备处理。     

Glycation of fibronectin inhibits VEGF‐induced angiogenesis by uncoupling VEGF receptor‐2‐c‐Src crosstalk

Glycation of extracellular matrix proteins has been demonstrated to contribute to the pathogenesis of vascular complications. However, no previous report has shown the role of glycated fibronectin (FN) in vascular endothelial growth factor (VEGF)‐induced angiogenesis. Thus, this study aimed to investigate the effects of glycated FN on VEGF signalling and to clarify the molecular mechanisms involved. FN was incubated with methylglyoxal (MGO) in vitro to synthesize glycated FN, and human umbilical vein endothelial cells (HUVECs) were seeded onto unmodified and MGO‐glycated FN. Then, VEGF‐induced angiogenesis and VEGF‐induced VEGF receptor‐2 (VEGFR‐2) signalling activation were measured. The results demonstrated that normal FN‐positive bands (260 kD) vanished and advanced glycation end products (AGEs) appeared in MGO‐glycated FN and glycated FN clearly changed to a higher molecular mass. The glycation of FN inhibited VEGF‐induced VEGF receptor‐2 (VEGFR‐2), Akt and ERK1/2 activation and VEGF‐induced cell migration, proliferation and tube formation. The glycation of FN also inhibited the recruitment of c‐Src to VEGFR‐2 by sequestering c‐Src through receptor for AGEs (RAGE) and the anti‐RAGE antibody restored VEGF‐induced VEGFR‐2, Akt and ERK1/2 phosphorylation, endothelial cell migration, proliferation and tube formation. Furthermore, the glycation of FN significantly inhibited VEGF‐induced neovascularization in the Matrigel plugs implanted into subcutaneous tissue of mice. Taken together, these data suggest that the glycation of FN may inhibit VEGF signalling and VEGF‐induced angiogenesis by uncoupling VEGFR‐2‐c‐Src interaction. This may provide a novel mechanism for the impaired angiogenesis in diabetic ischaemic diseases.     

生物发酵法制备木糖醇的研究进展 *

木糖醇由于其特殊的物理,化学性质在众多领域被广泛应用,全球需求量日益增多.目前,木糖醇的工业生产方法是由纯D-木糖在高温,高压下化学催化法制得的,该方法存在原料要求高,能耗高,条件苛刻,污染重等问题.生物发酵技术可以利用菌株发酵价格低廉的农作物废弃物来制备木糖醇,由于其原料来源广,能耗低,条件温和,环境友好等优点而备受关注,是一种潜在的具有吸引力的化学过程替代品.然而,由于发酵产物木糖醇含量低,微生物发酵法制备木糖醇的路线尚未在工业上得到实践.综述了生物法制备木糖醇过程中影响木糖醇产率的因素以及可能的菌种改造技术,并给出了生物技术生产木糖醇目前面临的挑战,进一步展望了生物法制备木糖醇的研究方向.     

The influence of 5% and 10% dietary apple pectin on parameters of fermentation in faeces and caecal digesta of weaning pigs

In order to determine the effect of pectin on fermentation parameters in the faeces and caecal digesta of weaned pigs 18 castrated male crossbred pigs with an average body weight of 8?kg were fitted with T-cannulas at the caecum. The animals were randomly distributed into three groups and fed with diets supplemented with 0, 5 and 10% pectin. Faeces were collected over a period of 3 days. Thereafter the diets were withdrawn for 24?h followed by ad libitum feeding to enhance the feed intake. Caecal chyme was collected 0, 8 and 24?h postprandial. In the faeces the addition of 5% pectin to the diet lowered the content of dry matter and lactic acid. The pH and the digestibility of pectins, the concentration of total SCFA, acetate, propionate, butyrate, bicarbonate and chloride increased. Dietary pectin of 10% increased the content of total SCFA and acetate further. When the diets were withdrawn and fed ad libitum 24?h later, a decline of the pH and an increased concentration of lactate in the caecal chyme could be observed in all groups up to 8?h after feeding. With an interval of 8 to 24?h after feeding, a further decline in pH and a rise of lactate only occurred when the diet was not supplemented with pectin. It was concluded that pectin might be beneficial for the development of fermentative processes in the large intestine.     

The Logic,Experimental Steps,and Potential of Heterologous Natural Product Biosynthesis Featuring the Complex Antibiotic Erythromycin A Produced Through E. coli

The heterologous production of complex natural products is an approach designed to address current limitations and future possibilities. It is particularly useful for those compounds which possess therapeutic value but cannot be sufficiently produced or would benefit from an improved form of production. The experimental procedures involved can be subdivided into three components: 1) genetic transfer; 2) heterologous reconstitution; and 3) product analysis. Each experimental component is under continual optimization to meet the challenges and anticipate the opportunities associated with this emerging approach.Heterologous biosynthesis begins with the identification of a genetic sequence responsible for a valuable natural product. Transferring this sequence to a heterologous host is complicated by the biosynthetic pathway complexity responsible for product formation. The antibiotic erythromycin A is a good example. Twenty genes (totaling >50 kb) are required for eventual biosynthesis. In addition, three of these genes encode megasynthases, multi-domain enzymes each ~300 kDa in size. This genetic material must be designed and transferred to E. coli for reconstituted biosynthesis. The use of PCR isolation, operon construction, multi-cystronic plasmids, and electro-transformation will be described in transferring the erythromycin A genetic cluster to E. coli.Once transferred, the E. coli cell must support eventual biosynthesis. This process is also challenging given the substantial differences between E. coli and most original hosts responsible for complex natural product formation. The cell must provide necessary substrates to support biosynthesis and coordinately express the transferred genetic cluster to produce active enzymes. In the case of erythromycin A, the E. coli cell had to be engineered to provide the two precursors (propionyl-CoA and (2S)-methylmalonyl-CoA) required for biosynthesis. In addition, gene sequence modifications, plasmid copy number, chaperonin co-expression, post-translational enzymatic modification, and process temperature were also required to allow final erythromycin A formation.Finally, successful production must be assessed. For the erythromycin A case, we will present two methods. The first is liquid chromatography-mass spectrometry (LC-MS) to confirm and quantify production. The bioactivity of erythromycin A will also be confirmed through use of a bioassay in which the antibiotic activity is tested against Bacillus subtilis. The assessment assays establish erythromycin A biosynthesis from E. coli and set the stage for future engineering efforts to improve or diversify production and for the production of new complex natural compounds using this approach.     

试论肠菌与中草药联合和发酵防治疾病

【摘 要】 本文阐述了肠道原始固有菌群（肠菌）与天然药用植物（中草药）联合应用和用肠菌发酵中草药的可行性及应用前景。