Co-culture of cord blood CD34(+) cells with human BM mesenchymal stromal cells enhances short-term engraftment of cord blood cells in NOD/SCID mice

BACKGROUND: The major challenge for cord blood transplantation (CBT) is higher rates of delayed and failed engraftment. In an attempt to broaden the application of CBT to more candidates, ex vivo expansion of hematopoietic stem/progenitor cells in CB is a major area of investigation. The purpose of this study was to employ human BM mesenchymal stromal cells (hBM-MSC) as the feeding-layer to expand CB cells ex vivo. METHODS: In this study, hBM-MSC were isolated and characterized by morphologic, mmunophenotypic and RT-PCR analysis. The hBM-MSC at passage 3 were employed as the feeding-layer to expand CB CD34(+) cells in vivo in the presence of thrombopoietin, flt3/flk2 ligand, stem cell factor and G-CSF. The repopulating capacity of the ex vivo-expanded CB cells was also evaluated in a NOD/SCID mice transplant experiment. RESULTS: After 1 or 2 weeks of in vitro expansion, hBM-MSC supported more increasing folds of CB in total nucleated cells, CD34(+) cells and colony-forming units (CFU) compared with CB without hBM-MSC. Furthermore, although NOD/SCID mice transplanted with CB cells expanded only in the presence of cytokines showed a higher percentage of human cell engraftment in BM than those with unexpanded CB CD34(+) cells, expanded CB cells co-cultured with hBM-MSC were revealed to enhance short-term engraftment further in recipient mice. DISCUSSION: Our study suggests that hBM-MSC enhance in vitro expansion of CB CD34(+) cells and short-term engraftment of expanded CB cells in NOD/SCID mice, which may be valuable in a clinical setting.     

EDNRB基因编码区点突变及多态性在浙江地区散发性先天性巨结肠症中的检测与分析

本实验应用聚合酶链反应-单链构象多态性(single strand confbrmation polymorphism analysis of polymerase chain reaction products,PCR-SSCP)和DNA直接测序技术,对75例浙江地区散发性先天性巨结肠病例EDNRB基因编码区的全部7个外显子,进行了点突变与单核苷酸多态性的检测与分析,探讨浙江地区先天性巨结肠患者EDNRB基因的突变特征,阐明EDNRB基因与散发性先天性巨结肠症发病之间可能存在的关系。结果有6例患者在第4外显子上检测到密码子277位点 CTG→CTA的置换,导致亮氨酸的同义突变(L277L),属于单核苷酸多态性,发生率为8％(6/75)。有 2例患者在第2外显子上检测到密码子185位点GTG→ATG的置换,导致缬氨酸到蛋氨酸的错义突变(V185M),此突变型未在国内外文献中报道过,认为是新的基因突变型,突变率2．7％(2/75)。研究结果表明,浙江地区先天性巨结肠群体可发生EDNRB基因的杂合性突变,提示EDNRB基因与先天性巨结肠症的发病存在一定程度的关联。     

MiR-455-5p ameliorates HG-induced apoptosis,oxidative stress and inflammatory via targeting SOCS3 in retinal pigment epithelial cells

Diabetic retinopathy (DR) remains the leading cause of blindness in adults with diabetes mellitus. Numerous microRNAs (miRNAs) have been identified to modulate the pathogenesis of DR. The main purpose of this study was to evaluate the potential roles of miR-455-5p in high glucose (HG)-treated retinal pigment epithelial (RPE) cells and underlying mechanisms. Our present investigation discovered that the expression of miR-455-5p was apparently downregulated in ARPE-19 cells stimulated with HG. In addition, forced expression of miR-455-5p markedly enhanced cell viability and restrained HG-induced apoptosis accompanied by decreased BCL2-associated X protein (Bax)/B-cell leukemia/lymphoma 2 (Bcl-2) ratio and expression of apoptotic marker cleaved caspase-3 during HG challenged. Subsequently, augmentation of miR-455-5p remarkably alleviated HG-triggered oxidative stress injury as reflected by decreased the production of intracellular reactive oxygen species (ROS) and malondialdehyde (MDA) content as well as NADPH oxidase 4 expression, concomitant with enhanced the activities of superoxide dismutase, catalase, and GPX stimulated with HG. Furthermore, enforced expression of miR-455-5p effectively ameliorated HG-stimulated inflammatory response as exemplified by repressing the secretion of inflammatory cytokines interleukin 1β (IL-1β), IL-6, and tumour necrosis factor-α in ARPE-19 cells challenged by HG. Most importantly, we successfully identified suppressor of cytokine signaling 3 (SOCS3) as a direct target gene of miR-455-5p, and miR-455-5p negatively regulated the expression of SOCS3. Mechanistically, restoration of SOCS3 abrogated the beneficial effects of miR-455-5p on apoptosis, accumulation of ROS, and inflammatory factors production in response to HG. Taken together, these findings demonstrated that miR-455-5p relieved HG-induced damage through repressing apoptosis, oxidant stress, and inflammatory response by targeting SOCS3. The study gives evidence that miR-455-5p may serve as a new potential therapeutic agent for DR treatment.     

Man5GlcNAc2哺乳动物甘露糖型糖蛋白的毕赤酵母表达系统构建

蛋白的糖基化对蛋白的活性、高级结构及功能都有重要的影响。酵母表达的糖蛋白不同于哺乳动物表达的杂合型或复杂型糖蛋白,而是高甘露糖型或过度甘露糖化糖蛋白。在前期成功敲除毕赤酵母α-1,6-甘露糖转移酶(Och1p)基因、阻断毕赤酵母过度糖基化,获得毕赤酵母过度糖基化缺陷菌株GJK01 (ura3、och1) 的基础上,通过表达不同物种来源的α-1,2-甘露糖苷酶I (MDSI) 的活性区与酵母自身定位信号的融合蛋白,并通过DSA-FACE (基于DNA测序仪的荧光辅助糖电泳) 分析筛选报告蛋白HSA/GM-CSF (人血清白蛋白与粒细胞-巨噬细胞集落刺激因子融合蛋白) 的糖基结构,发现当编码酿酒酵母α-1,2-甘露糖苷酶 (MnsI) 基因的内质网定位信号与带有完整C-端催化区的拟南芥MDSI基因融合表达时,毕赤酵母工程菌株能够合成Man5GlcNAc2哺乳动物甘露糖型糖蛋白。这为在酵母体内合成类似于哺乳动物杂合型或复杂型糖基化修饰的糖蛋白奠定了基础。     

CO2浓度升高和施氮条件下小麦根际呼吸对土壤呼吸的贡献

依托FACE技术平台,采用稳定13C同位素技术,通过将小麦(C3作物)种植于长期单作玉米(C4作物)的土壤上,研究了大气CO2浓度升高和不同氮肥水平对土壤排放CO2的δ13C值及根际呼吸的影响.结果表明:种植小麦后土壤排放CO2的δ13C值随作物生长逐渐降低,CO2浓度升高200 μmol·mol-1显著降低了孕穗、抽穗期(施氮量为250 kg·hm-2,HN)与拔节、孕穗期(施氮量为150 kg·hm-2,LN)土壤排放CO2的δ13C值,显著提高了孕穗、抽穗期的根际呼吸比例.拔节至成熟期,根际呼吸占土壤呼吸的比例在高CO2浓度下为24％～48％ (HN)和21％ ～48％ (LN),在正常CO2浓度下为20％ ～36％ (HN)和19％～32％(LN).不同CO2浓度下土壤排放CO2的δ13C值和根际呼吸对氮肥增加的响应不同,CO2浓度与氮肥用量在拔节期对根际呼吸的交互效应显著.     

Altered connectivity pattern of hubs in default-mode network with Alzheimer's disease: an Granger causality modeling approach

Background

Evidences from normal subjects suggest that the default-mode network (DMN) has posterior cingulate cortex (PCC), medial prefrontal cortex (MPFC) and inferior parietal cortex (IPC) as its hubs; meanwhile, these DMN nodes are often found to be abnormally recruited in Alzheimer''s disease (AD) patients. The issues on how these hubs interact to each other, with the rest nodes of the DMN and the altered pattern of hubs with respect to AD, are still on going discussion for eventual final clarification.

Principal Findings

To address these issues, we investigated the causal influences between any pair of nodes within the DMN using Granger causality analysis and graph-theoretic methods on resting-state fMRI data of 12 young subjects, 16 old normal controls and 15 AD patients respectively. We found that: (1) PCC/MPFC/IPC, especially the PCC, showed the widest and distinctive causal effects on the DMN dynamics in young group; (2) the pattern of DMN hubs was abnormal in AD patients compared to old control: MPFC and IPC had obvious causal interaction disruption with other nodes; the PCC showed outstanding performance for it was the only region having causal relation with all other nodes significantly; (3) the altered relation between hubs and other DMN nodes held potential as a noninvasive biomarker of AD.

Conclusions

Our study, to the best of our knowledge, is the first to support the hub configuration of the DMN from the perspective of causal relationship, and reveal abnormal pattern of the DMN hubs in AD. Findings from young subjects provide additional evidence for the role of PCC/MPFC/IPC acting as hubs in the DMN. Compared to old control, MPFC and IPC lost their roles as hubs owing to the obvious causal interaction disruption, and PCC was preserved as the only hub showing significant causal relations with all other nodes.     

不吸水链霉菌梧州新亚种胆固醇氧化酶基因的克隆与序列分析

克隆不吸水链霉菌梧州新亚种胆固醇氧化酶基因.以不吸水链霉菌梧州新亚种基因组DNA作为模板,根据已知的胆固醇氧化酶基因的保守序列设计简并引物进行PCR扩增,PCR产物与载体连接、转化,构建重组质粒.获得了不吸水链霉菌梧州新亚种胆固醇氧化酶基因重组质粒,序列测定并分析.     

Effect of UV-B radiation on the growth and antioxidant enzymes of Antarctic sea ice microalgae <Emphasis Type="Italic">Chlamydomonas</Emphasis> sp. ICE-L

The effect of ultraviolet-B (UV-B) radiation on Antarctic phytoplankton has become an attractive ecological issue as a result of annual springtime ozone depletion. The effects of UV-B radiation on the growth and antioxidant enzymes were investigated using Antarctic sea ice microalgae Chlamydomonas sp. ICE-L as the material in this study. The results demonstrated that UV-B radiation could notably inhibit the growth, especially at high UV-B radiation intensity (70 μW cm⁻²). Malondialdehyde and O₂ ^·− content in ICE-L increased rapidly in early days (1–3 days) exposed to UV-B radiation enhancement, then decreased rapidly. In the stress of UV-B radiation enhancement, the superoxide dismutase, peroxidase and Catalase activities of 1–4 days in ICE-L were obviously higher than those in the control, and their activities became higher at high UV-B radiation intensity (70 μW cm⁻²). These enzymes activity of 7 days would kept stable at low UV-B radiation intensity (35 μW cm⁻²), but kept high level at high UV-B radiation intensity (70 μW cm⁻²). However, the ascorbate peroxidase activity in ICE-L kept stable under the stress of UV-B radiation enhancement. The above experimental results indicated that the antioxidant enzyme system played an important role in the adaptation of Antarctic ice microalgae under the UV-B radiation change of Antarctic ecosystems.     

荻幼苗对复合盐碱胁迫的生理响应

该研究以荻(Miscanthus sacchariflorus)幼苗为材料,模拟中国东北大庆盐碱地的低、中、高浓度盐碱土壤环境,考察不同盐碱处理浓度及其不同处理时间(0~25d)对荻幼苗各项生理指标的影响,以界定荻幼苗的复合盐碱耐受范围,为荻类能源植物的耐盐碱性筛选和在园林中的应用提供理论依据。结果表明:(1)随着复合盐碱胁迫时间的延长和胁迫程度加剧,荻幼苗叶绿素含量不断降低,根活力在低、中浓度处理下随时间不断升高,而在高浓度处理时先降低后上升再降低;同时,幼苗相对电导率和MDA含量则不断升高。(2)随着复合盐碱胁迫时间延长,荻幼苗SOD、POD、CAT、APX、GR活性和ASA、GSH含量一般在低浓度处理时缓慢升高,而高浓度处理时则呈先升高后降低的趋势。研究发现,荻幼苗能够通过调节自身保护酶系统活性和抗氧化物积累,清除体内过多活性氧和自由基,保护细胞结构,有效抵御外界低、中浓度的复合盐碱胁迫,但其对高浓度的复合盐碱胁迫基本没有抵抗能力。