重组枯草芽孢杆菌全细胞催化合成2-O-α-D-甘油葡糖苷

2-O-α-D-甘油葡糖苷是一种在食品、化妆品、保健品及医药领域有着重大应用前景的高附加值产品,但国内仍未实现2-O-α-D-甘油葡糖苷的工业化生产,且鲜有关于2-O-α-D-甘油葡糖苷合成的相关报道。文中旨在开发一种利用食品安全级重组枯草芽孢杆菌全细胞催化合成2-O-α-D-甘油葡糖苷的方法,通过构建一株异源表达肠膜明串珠菌蔗糖磷酸化酶 (Sucrose phosphorylase,SPase) 的重组枯草芽孢杆菌Bacillus subtilis 168/pMA5-gtfA,并将其用作全细胞催化剂合成2-O-α-D-甘油葡糖苷,通过优化培养温度、时间及全细胞转化条件,提高其转化合成2-O-α-D-甘油葡糖苷的产量。结果表明,重组枯草芽孢杆菌B. subtilis 168/pMA5-gtfA在30 ℃下培养20 h,菌体裂解物酶活力最大达1.43 U/mL,并且在1 mol/L蔗糖、2.5 mol/L甘油、pH 7.0、菌体OD600为40、30 ℃下全细胞转化反应48 h,共生成2-O-α-D-甘油葡糖苷189.3 g/L,平均转化速率为15.6 mmol/(L·h),蔗糖转化率约为75.1%,是目前报道的利用重组枯草芽孢杆菌催化合成2-O-α-D-甘油葡糖苷的最高产量,这为2-O-α-D-甘油葡糖苷的工业化生产及应用奠定了理论和实验基础。     

Migration sources and pathways of the pest species Sogatella furcifera in Yunnan,China, and across the border inferred from DNA and wind analyses

The migration sources and pathways of Sogatella furcifera (Horváth) in topologically complex regions like Yunnan, China, and adjacent montane areas have long been a challenging task and a bottleneck in effective pest forecast and control. The present research reinvestigated this issue using a combination of mtDNA and long‐term historical wind field data in an attempt to provide new insights. Genetic analyses showed that the 60 populations of S. furcufera collected across Myanmar, Thailand, Laos, Vietnam, Yunnan, Guizhou, and Sichuan lack genetic structure and geographic isolation, while spatial analysis of haplotype and diversity indices discovered geographic relevance between populations. Migration rate analysis combined with high‐resolution 10‐year wind field analysis detected the following migration sources, pathways, and impacted areas which could explain the outbreak pattern in Yunnan. (a) Dominating stepwise northward migrations originated from northern Indochina, southern Yunnan, and central‐eastern Yunnan, impacting their northern areas. (b) Concurring summer–autumn southward (return) migration originated from nearly all latitude belts of Sichuan and Yunnan mainly impacting central and southern Yunnan. (c) Regular eastward and summer–autumn westward migrations across Yunnan. The northward migration reflects the temporal rhythm of gradual outbreaks from the south to the north in a year, while the return migration may explain the repeated or very severe outbreaks in the impacted areas. To form a better pest forecast and control network, attention must also be paid to the northern part of Yunnan to suppress the impact of return migration in summers and autumns.     

Erianin induces triple-negative breast cancer cells apoptosis by activating PI3K/Akt pathway

Background: Triple-negative breast cancer (TNBC) is a refractory subtype of breast cancer, 25–30% of which have dysregulation in the PI3K/AKT pathway. The present study investigated the anticancer effect of erianin on TNBC cell line and its underlying mechanism.Methods: After treatment with erianin, MTT assay was employed to determine the MDA-MB-231 and EFM-192A cell proliferation, the nucleus morphological changes were observed by DAPI staining. The cell cycle and apoptotic proportion were detected by flow cytometry. Western blot was performed to determine the cell cycle and apoptosis-related protein expression and PI3K pathways. Finally, the antiproliferative activity of erianin was further confirmed by adding or not adding PI3K agonists SC79.Results: Erianin inhibited the proliferation of MDA-MB-231 and EFM-192A cells in a dose-dependent manner, the IC₅₀ were 70.96 and 78.58 nM, respectively. Erianin could cause cell cycle arrest at the G₂/M phase, and the expressions of p21 and p27 were up-regulated, while the expressions of CDK1 and Cyclin B1 were down-regulated. Erianin also induced apoptosis via the mitochondrial pathway, with the up-regulation of the expression of Cyto C, PARP, Bax, active form of Caspase-3, and Caspase-9. Furthermore, p-PI3K and p-Akt expression were down-regulated by erianin. After co-incubation with SC79, the cell inhibition rate of erianin was decreased, which further confirmed that the attenuated PI3K/Akt pathway was relevant to the pro-apoptotic effect of erianin.Conclusions: Erianin can inhibit the proliferation of TNBC cells and induce cell cycle arrest and apoptosis, which may ascribe to the abolish the activation of the PI3K/Akt pathway.     

Efficient Editing of an Adenoviral Vector Genome with CRISPR/Cas9

Immunotherapy based on genetic modification of T cells has played an important role in the treatment of tumors and viral infections. Moreover, adenoviral vectors engineered with improved safety due to their inability to integrate into the host genome have been key in the clinical application of T cell therapy. However, the commonly used adenoviral vector Ad5 exhibits low efficiency of infection of human T cells and the details of the intracellular trafficking pathway of adenoviral vectors in human primary T cells remains unclear. Resolution of these issues will depend on successful modification of the adenoviral vector. To this end, here we describe the successful establishment of a simple and efficient method for editing adenoviral vectors in vitro using the CRISPR-Cas9 gene editing system to target the adenoviral fiber gene. Electronic supplementary materialThe online version of this article (10.1007/s12088-020-00905-3) contains supplementary material, which is available to authorized users.     

Sirt3 Protects Against Ischemic Stroke Injury by Regulating HIF-1α/VEGF Signaling and Blood–Brain Barrier Integrity

Sirtuin 3 (Sirt3) is a member of the Sirtuin family proteins and known to regulate multiple physiological processes such as metabolism and aging. As stroke is an aging-related disease, in this work, we attempt to examine the role and potential mechanism of Sirt3 in regulating ischemic stroke by using a permanent middle cerebral artery occlusion (pMCAO) model in wild type (WT) and Sirt3 knockout (KO) mice, coupled with oxygen glucose deprivation (OGD) experiments in cultured primary astrocytes. Sirt3 deficiency aggravated neuronal cell apoptosis and neurological deficits after brain ischemia. In addition, Sirt3 KO mice showed more severe blood–brain barrier (BBB) disruption and inflammatory responses compared with WT group in the acute phase. Furthermore, specific overexpression of Sirt3 in astrocytes by injecting glial fibrillary acidic protein (GFAP)::Sirt3 virus in ischemic region showed protective effect against stroke-induced damage. Mechanistically, Sirt3 could regulate vascular endothelial growth factor (VEGF) expression by inhibiting hypoxia inducible factor-1α (HIF-1α) signaling after ischemia (OGD). Our results have shown that Sirt3 plays a protective role in ischemic stroke via regulating HIF-1α/VEGF signaling in astrocytes, and reversal of the Sirt3 expression at the acute phase could be a worthy direction for stroke therapy.

     

Astragaloside IV alleviates atherosclerosis through targeting circ_0000231/miR-135a-5p/CLIC4 axis in AS cell model in vitro

Molecular and Cellular Biochemistry - Non-coding RNAs (ncRNAs) have shown to act as crucial mediators in atherosclerosis (AS) development. The purpose of our study was to explore the role of...     

Protective Effects of 28-O-Caffeoyl Betulin (B-CA) on the Cerebral Cortex of Ischemic Rats Revealed by a NMR-Based Metabolomics Analysis

28-O-caffeoyl betulin (B-CA) has been demonstrated to reduce the cerebral infarct volume caused by transient middle cerebral artery occlusion (MCAO) injury. B-CA is a novel derivative of naturally occurring caffeoyl triterpene with little information associated with its pharmacological target(s). To date no data is available regarding the effect of B-CA on brain metabolism. In the present study, a ¹H-NMR-based metabolomics approach was applied to investigate the therapeutic effects of B-CA on brain metabolism following MCAO in rats. Global metabolic profiles of the cortex in acute period (9 h after focal ischemia onset) after MCAO were compared between the groups (sham; MCAO?+?vehicle; MCAO?+?B-CA). MCAO induced several changes in the ipsilateral cortex of ischemic rats, which consequently led to the neuronal damage featured with the downregulation of NAA, including energy metabolism dysfunctions, oxidative stress, and neurotransmitter metabolism. Treatment with B-CA showed statistically significant rescue effects on the ischemic cortex of MCAO rats. Specifically, treatment with B-CA ameliorated the energy metabolism dysfunctions (back-regulating the levels of succinate, lactate, BCAAs, and carnitine), oxidative stress (upregulating the level of glutathione), and neurotransmitter metabolism disturbances (back-regulating the levels of γ-aminobutyric acid and acetylcholine) associated with the progression of ischemic stroke. With the administration of B-CA, the levels of three phospholipid related metabolites (O-phosphocholine, O-phosphoethanolamine, sn-glycero-3-phosphocholine) and NAA improved significantly. Overall, our findings suggest that treatment with B-CA may provide neuroprotection by augmenting the metabolic changes observed in the cortex following MCAO in rats.

     

Plant Growth-Promoting Effect of the Chitosanolytic Phosphate-Solubilizing Bacterium Burkholderia gladioli MEL01 After Fermentation with Chitosan and Fertilization with Rock Phosphate

Phosphate-solubilizing bacteria (PSB) are important plant growth-promoting rhizobacteria that can increase soil fertility through the solubilization of insoluble inorganic phosphate and organophosphorus. In this study, a PSB, Burkholderia gladioli MEL01, was isolated and identified from rice–wheat rotation rhizosphere soil. MEL01 had an excellent phosphate-solubilizing capacity (reaching 107.69 mg/L) toward insoluble inorganic phosphate rock phosphate. HPLC analysis revealed that the mechanism of phosphate solubilization of MEL01 was probably due to secreted oxalic acid and gluconic acid transformation of phosphate from insoluble to soluble. MEL01 also exhibited 4030 U/L specific chitosanase activity when cultured with chitosan fermentation medium. Interestingly, the chitosan hydrolysis product chitooligosaccharide could significantly enhance the MEL01 phosphate-solubilizing capacity. Pot experiments showed that MEL01 chitosan medium fermentation liquor (MCMFL) could promote improvement of soil available phosphorus and pakchoi growth when supplemented with phosphate rock phosphate as the phosphate fertilizer. In addition, pot experiments demonstrated that MCMFL could also promote the growth of wheat, which could decrease the amount of compound fertilizer used. Microbial diversity analysis showed that the genera Pseudomonas, Burkholderia, Mycoplana, and Cellvibrio were enriched, which might participate in synergetic phosphate solubilization. Therefore, after fermentation with chitosan and fertilization with rock phosphates, MEL01 has potential as a phosphate biofertilizer in ecological agricultural production.

     

成长记录袋评价法提升细胞工程课堂教学质量

首次将成长记录袋评价法引入到高等院校生物类专业课程《细胞工程》的课堂教学改革中,构建了较为完备的课堂评价体系,将成长记录袋课堂评价体系的建立分为4个阶段,即准备阶段、演练阶段、实施阶段和作品展示阶段。并从实施成长记录袋评价法的可行性与必要性、评价体系的构建、执行过程中应注意的要点及事项分别展开论述。结果表明：通过该评价法的施行,不仅活跃了课堂气氛,增强了学生学习的主动性和自主性,还提高了学生分析、解决与细胞工程技术相关的专业问题的能力,执行效果良好。本课程课堂教学改革的实施,可为高校同类性质的其他专业课程提供借鉴与参考。