Biooxidation of ferrous iron by immobilized Acidithiobacillus ferrooxidans in poly(vinyl alcohol) cryogel carriers

PVA-cryogels entrapping about 10⁹ cells of Acidithiobacillus ferrooxidans per ml of gel were prepared by freezing-thawing procedure, and the biooxidation of Fe²⁺ by immobilized cells was investigated in a 0.365 l packed-bed bioreactor. Fe²⁺ oxidation fits a plug-flow reaction model well. A maximum oxidation rate of 3.1 g Fe²⁺ l^–1 h^–1 was achieved at the dilution rate of 0.4 h^–1 or higher, while no obvious precipitate was determined at this time. In addition, cell-immobilized PVA-cryogels packed in bioreactor maintained their oxidative ability for more than two months under non-sterile conditions. Nomenclature: C _A0 – Concentration of Fe²⁺ in feed stream (g l^–1) C _A – Concentration of Fe^{2 +} in outlet stream (g l^{– 1}) D – Dilution rate of the packed-bed bioreactor (h^–1) F – Volumetric flow rate of iron solution (l h^–1) F _A0 – Mass flow rate of Fe²⁺ in the feed stream (g h^–1) K – Kinetic constant (l l^–1 h^–1) r _A – Oxidation rate of Fe²⁺ (g l^–1 h^–1) V – Volume of packed-bed bioreactor (l) X _A – Conversion ratio of Fe²⁺ (%)     

啤酒酵母生产的重组水蛭素的纯化及脱色

对啤酒酵母生产的重组水蛭素变异体1(rHV1)进行多步骤的纯化。首先将分泌到培养上清液中的水蛭素进行硫酸铵沉淀和SephadexG-50凝胶过滤,再用Q-SepharoseHP阴离子交换层析分离,最后用HPLCSP-5PW阳离子交换柱脱色及HPLCC8柱反相层析。真空干燥后得到的水蛭素蛋白经SPS-PAGE、N端氨基酸序列分析、抗凝血酶活力分析鉴定,证明已获得高纯度的重组水蛭素HV1制剂,为利用基因工程方法生产重组水蛭素的规模化生产及临床应用提供了依据     

New Immunomodulating Polyhydroxypregnane Glycosides from the Roots of Cynanchum otophyllum C.K.Schneid.

Seven new polyhydroxypregnane glycosides, named cynotophyllosides P–V, together with three known analogs were isolated from the roots of Cynanchum otophyllum C.K.Schneid . Their structures were elucidated by a variety of spectroscopic techniques, as well as acid‐catalyzed hydrolysis. All isolates were tested for their immunological activities in vitro against Con A‐ and LPS‐induced proliferation of mice splenocytes. Immunoenhancing (for 1 , 9 ) and immunosuppressive (for 2 ) activities were observed. Furthermore, cynotophylloside R ( 3 ) showed immunomodulatory as it enhanced the proliferation of splenocytes in low concentration and suppressed immune cells in concentration more than 1.0 μg/ml.     

RIPK1-RIPK3 mediates myocardial fibrosis in type 2 diabetes mellitus by impairing autophagic flux of cardiac fibroblasts

Receptor-interacting protein kinase 1 (RIPK1) and 3 (RIPK3) are critical regulators of programmed necrosis or necroptosis. However, the role of the RIPK1/RIPK3 signaling pathway in myocardial fibrosis and related diabetic cardiomyopathy is still unclear. We hypothesized that RIPK1/RIPK3 activation mediated myocardial fibrosis by impairing the autophagic flux. To this end, we established in vitro and in vivo models of type 2 diabetes mellitus with high glucose fat (HGF) medium and diet respectively. HGF induced myocardial fibrosis, and impaired cardiac diastolic and systolic function by activating the RIPK1/RIPK3 pathway, which increased the expression of autophagic related proteins such as LC3-II, P62 and active-cathepsin D. Inhibition of RIPK1 or RIPK3 alleviated HGF-induced death and fibrosis of cardiac fibroblasts by restoring the impaired autophagic flux. The autophagy blocker neutralized the effects of the RIPK1 inhibitor necrostatin-1 (Nec-1) and RIPK3 inhibitor GSK872 (GSK). RIPK1/RIPK3 inhibition respectively decreased the levels of RIPK3/p-RIPK3 and RIPK1/p-RIPK1. P62 forms a complex with RIPK1-RIPK3 and promotes the binding of RIPK1 and RIPK3, silencing of RIPK1 decreased the association of RIPK1 with P62 and the binding of P62 to LC3. Furthermore, inhibition of both kinases in combination with a low dose of Nec-1 and GSK in the HGF-treated fibroblasts significantly decreased cell death and fibrosis, and restored the autophagic flux. In the diabetic rat model, Nec-1 (1.65 mg/kg) treatment for 4 months markedly alleviated myocardial fibrosis, downregulated autophagic related proteins, and improved cardiac systolic and diastolic function. In conclusion, HGF induces myocardial fibrosis and cardiac dysfunction by activating the RIPK1-RIPK3 pathway and by impairing the autophagic flux, which is obviated by the pharmacological and genetic inhibition of RIPK1/RIPK3.Subject terms: Necroptosis, Diabetes complications     

Endocytosis is enhanced in Tangier fibroblasts: possible role of ATP-binding cassette protein A1 in endosomal vesicular transport

A human genetic disorder, Tangier disease, has been linked recently to mutations in ATP-binding cassette protein A1 (ABCA1). In addition to its function in apoprotein A-I-mediated lipid removal, ABCA1 was also shown to be a phosphatidylserine (PS) translocase that facilitates PS exofacial flipping. This PS translocation is crucial for the plasma membrane to produce protrusions enabling the engulfment of apoptotic cells. In this report, we show that ABCA1 also plays a role in endocytosis. Receptor-mediated endocytosis, probed by both transferrin and low density lipoprotein, is up-regulated by more than 50% in homozygous Tangier fibroblasts in comparison with controls. Fluid-phase uptake is increased similarly. We also demonstrate that bulk membrane flow, including lipid endocytosis and exocytosis, is accelerated greatly in Tangier cells. Moreover, endocytosis is similarly enhanced in normal fibroblasts when ABCA1 function is inhibited by glyburide, whereas glyburide has no effect on endocytosis in Tangier cells. In addition, we demonstrate a decreased annexin V binding in Tangier fibroblasts as compared with controls, supporting the notion that PS transmembrane distribution is indeed defective in the presence of ABCA1 mutations. Furthermore, adding a PS analog to the exofacial leaflet of the plasma membrane normalizes endocytosis in Tangier cells. Taken together, these data demonstrate that ABCA1 plays an important role in endocytosis. We speculate that this is related to the PS translocase function of ABCA1. A loss of functional ABCA1, as in the case of Tangier cells, enhances membrane inward bending and facilitates endocytosis.     

MicroRNA-451 regulates p38 MAPK signaling by targeting of Ywhaz and suppresses the mesangial hypertrophy in early diabetic nephropathy

Diabetic nephropathy (DN) is a major diabetic complication. However, the initiating molecular events triggering DN are unknown. In this study we focused on microRNA-451 (miR-451), which is downregulated during early DN. We found that miR-451 negatively regulated the expression of Ywhaz through Ywhaz 3'UTR and that Ywhaz was required for the miR-451-mediated downregulation of p38 MAPK signalling. Moreover, over-expression of miR-451 inhibits glomerular mesangial cell proliferation in vitro and in vivo. These findings suggest that the growth-inhibitory effect of miR-451 may be explained in part by miR-451-induced suppression of Ywhaz and p38 MAPK signalling, providing evidence for the potential role of miR-451 in early DN.     

microRNA‐124 inhibits stem‐like properties and enhances radiosensitivity in nasopharyngeal carcinoma cells via direct repression of expression of JAMA

Cancer stem cells (CSCs) are a source of tumour recurrence in patients with nasopharyngeal carcinoma (NPC); however, the function of microRNA‐124 (miR‐124) in NPC CSCs has not been clearly defined. In this study, we investigated the role of miR‐124 in NPC CSCs. qRT‐PCR was performed to measure miR‐124 expression in NPC tissues and cell lines and the effects of miR‐124 on stem‐like properties and radiosensitivity of NPC cells measured. Luciferase reporter assays and rescue experiments were used to investigate the interaction of miR‐124 with the 3′UTR of junctional adhesion molecule A (JAMA). Finally, we examined the effects of miR‐124 in an animal model and clinical samples. Down‐regulation of miR‐124 was detected in cancer tissues and was inversely associated with tumour stage and lymph node metastasis. Overexpression of miR‐124 inhibited stemness properties and enhanced radiosensitivity of NPC cells in vitro and in vivo via targeting JAMA. Up‐regulation of miR‐124 was correlated with superior overall survival of patients with NPC. Our study demonstrates that miR‐124 can inhibit stem‐like properties and enhance radiosensitivity by directly targeting JAMA in NPC. These findings provide novel insights into the molecular mechanisms underlying therapy failure in NPC.