年龄增长对GRK5基因缺陷小鼠海马内肿胀轴突丛形成与积累的影响

目的研究G蛋白偶联受体（G protein-coupled receptors）激酶5（GPCR kinase-5,GRK5）基因缺陷和老化交互作用对阿尔茨海默病（Alzheimer＇s disease,AD）早期的病理改变-海马内肿胀轴突丛（swollen axonal clusters,SACs）出现和积累的影响。方法选取5、6、7、9、12～13、18—19月龄雌性GRK5基因敲除小鼠（GRK5 Knockout,GRK5KO）作为观察对象,另选取年龄匹配的雌性野生型（wild type,WT）小鼠为对照,每个年龄段GRK5KO和WT小鼠各4只。用抗人神经原纤维缠结（neurofibrillary tangles,NFTs）特异性抗体的免疫荧光染色方法观察海马内SACs的变化。结果所有小鼠随着年龄增长,海马内SACs逐渐增加;GRKSKO小鼠组海马内NFT^＋ SACs数量较WT型小鼠组显著增加（P〈0．01）;双因素方差分析显示遗传性GRK5基因缺陷和老化双因素对海马内NFT^＋SACs的影响有显著协同效应（P〈0．01）。结论在促进早期AD病理发生的过程中,GRK5缺陷和老化双因素共同加剧了雌性GRKSKO小鼠海马内SACs的形成与积累。     

Large-Scale Screens of miRNA-mRNA Interactions Unveiled That the 3′UTR of a Gene Is Targeted by Multiple miRNAs

Animal microRNA (miRNA) target prediction is still a challenge, although many prediction programs have been exploited. MiRNAs exert their function through partially binding the messenger RNAs (mRNAs; likely at 3′ untranslated regions [3′UTRs]), which makes it possible to detect the miRNA-mRNA interactions in vitro by co-transfection of miRNA and a luciferase reporter gene containing the target mRNA fragment into mammalian cells under a dual-luciferase assay system. Here, we constructed a human miRNA expression library and used a dual-luciferase assay system to perform large-scale screens of interactions between miRNAs and the 3′UTRs of seven genes, which included more than 3,000 interactions with triplicate experiments for each interaction. The screening results showed that the 3′UTR of one gene can be targeted by multiple miRNAs. Among the prediction algorithms, a Bayesian phylogenetic miRNA target identification algorithm and a support vector machine (SVM) presented a relatively better performance (27% for EIMMo and 24.7% for miRDB) against the average precision (17.3%) of the nine prediction programs used here. Additionally, we noticed that a relatively high conservation level was shown at the miRNA 3′ end targeted regions, as well as the 5′ end (seed region) binding sites.     

Mid-Neolithic Exploitation of Mollusks in the Guanzhong Basin of Northwestern China: Preliminary Results

Mollusk remains are abundant in archaeological sites in the Guanzhong Basin of Northwestern China, providing good opportunities for investigations into the use of mollusks by prehistoric humans. Here we report on freshwater gastropod and bivalve mollusks covering the time interval from about 5600 to 4500 cal. yrs BP from sites of Mid-Late Neolithic age. They are identified as Cipangopaludina chinensis and Unio douglasiae, both of which are currently food for humans. The shells are well preserved and have no signs of abrasion. They are all freshwater gastropods and bivalves found in pits without water-reworked deposits and have modern representatives which can be observed in rivers, reservoirs, and paddy fields in the studied region. Mollusk shells were frequently recovered in association with mammal bones, lithic artifacts, and pottery. These lines of evidence indicate that the mollusks are the remains of prehistoric meals. The mollusk shells were likely discarded into the pits by prehistoric humans after the flesh was eaten. However, these mollusk remains may not have been staple food since they are not found in large quantities. Mollusk shell tools and ornaments are also observed. Shell tools include shell knives, shell reaphooks and arrowheads, whereas shell ornaments are composed of pendants and loops. All the shell tools and ornaments are made of bivalve mollusks and do not occur in large numbers. The finding of these freshwater mollusk remains supports the view that the middle Holocene climate in the Guanzhong Basin may have been warm and moist, which was probably favorable to freshwater mollusks growing and developing in the region.     

Protein C‐terminal enzymatic labeling identifies novel caspase cleavages during the apoptosis of multiple myeloma cells induced by kinase inhibition

Caspase activation and proteolytic cleavages are the major events in the early stage of apoptosis. Identification of protein substrates cleaved by caspases will reveal the occurrence of the early events in the apoptotic process and may provide potential drug targets for cancer therapy. Although several N‐terminal MS‐based proteomic approaches have been developed to identify proteolytic cleavages, these methods have their inherent drawbacks. Here we apply a previously developed proteomic approach, protein C‐terminal enzymatic labeling (ProC‐TEL), to identify caspase cleavage events occurring in the early stage of the apoptosis of a myeloma cell line induced by kinase inhibition. Both previously identified and novel caspase cleavage sites are detected and the reduction of the expression level of several proteins is confirmed biochemically upon kinase inhibition although the current ProC‐TEL procedure is not fully optimized to provide peptide identifications comparable to N‐terminal labeling approaches. The identified cleaved proteins form a complex interaction network with central hubs determining morphological changes during the apoptosis. Sequence analyses show that some ProC‐TEL identified caspase cleavage events are unidentifiable when traditional N‐terminomic approaches are utilized. This work demonstrates that ProC‐TEL is a complementary approach to the N‐terminomics for the identification of proteolytic cleavage events such as caspase cleavages in signaling pathways.     

Improved production of citric acid by Yarrowia lipolytica using oleic acid as the oxygen‐vector and co‐substrate

Yarrowia lipolytica is able to secrete large amounts of citric acid (CA), which is greatly affected by the dissolved oxygen concentration (DOC) in the fermentation medium. In this study, oleic acid was selected as oxygen‐vector to improve DOC during CA fermentation. When 2% (v/v) of oleic acid was added to the culture broth, higher DOC (>42.1%) was determined throughout the CA synthesis phase. The yield of CA reached a maximum of 32.1 g/L (25.4% higher than the control) and the biomass was 8.8 g/L. The substrate uptake rate, products formation rate and key enzyme activities were also determined, and the results indicated that CA synthesis was strengthened with oleic acid addition. Furthermore, it was detected that oleic acid could be assimilated by the cells, which means that oleic acid could be served both as oxygen‐vector and co‐substrate for CA synthesis by Y. lipolytica. In a bioreactor with working volume of 3 L, the highest concentration of CA reached to 36. 4 g/L in the presence of 2% (v/v) oleic acid after 192 h of fermentation. These results confirmed that oleic acid could be applied in the large‐scale production of CA by Y. lipolytica.     

The protonated 2-halogenated imidazolium cation as the noncovalent interaction donor: the σ-hole and π-hole interactions

The σ-hole and π-hole of the protonated 2-halogenated imidazolium cation (XC₃H₄N₂ ⁺; X = F, Cl, Br, I) were investigated and analyzed. The monomers of (CH₃)₃SiY(Y=F, Cl, Br, I), considered as the Lewis base, were combined with the σ-hole and π-hole of XC₃H₄N₂ ⁺ to form the σ-hole and π-hole interactions in the bimolecular complexes (CH₃)₃SiY?·?·?·?XC₃H₄N₂ ⁺ and (CH₃)₃SiY?·?·?·?C₃(X)H₄N₂ ⁺(X/Y=F, Cl, Br, I), respectively. For both the σ-hole and π-hole interactions, the equilibrium geometries of complexes show regular changes according to the sequence of heavy sequence of the noncovalent interaction acceptors and donors. The electrostatic energy is the main contribution in the formation of both kinds of interactions, it has linear relations with the V _S,max values of σ-hole and the V′ _S,max values of π-hole. Both the σ-hole and π-hole interactions belong to the closed-shell and noncovalent interactions. The π-hole interactions are stronger than the σ-hole interactions. For the π-hole interactions, the contribution percents of the dispersion energies are somewhat greater than those of the σ-hole interactions, while it is contrary for the polarization energy.

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Graphical Abstract The protonated 2-halogenated imidazolium cation as the noncovalent interaction donor: the σ-hole and π-hole interactions?

     

Placenta Growth Factor in Eyes with Neovascular Glaucoma Is Decreased after Intravitreal Ranibizumab Injection

Purpose

To evaluate changes in the concentrations of placental growth factor (PlGF) and vascular endothelial growth factor-A (VEGF-A) in aqueous humor of patients with neovascular glaucoma (NVG) before and after an intravitreal injection of ranibizumab (IVR) and to determine the underlying correlation between the levels.

Methods

The prospective interventional comparative study involved 20 eyes of 20 patients with surgery-required advanced NVG and 20 control subjects from January 2013 to November 2013. The NVG eyes received the IVR treatment before glaucoma surgery. Aqueous humor was collected at the time of the IVR injection (pre- IVR) and at the time of antiglaucomatous surgery (post-IVR). Aqueous humor was also collected at the time of cataract surgery in normal control. Aqueous humor and plasma VEGF-A and PlGF levels were measured with an enzyme-linked immunosorbent assay methods, respectively.

Results

The mean aqueous humor PlGF and VEGF-A concentrations in the pre-IVR eyes were significantly higher than in those of the control subjects (p<0.001), whereas the plasma levels showed no significant difference. There was a statistically significant correlation between the aqueous humor PlGF and the VEGF-A concentration (r = 0.612, p = 0.003). The mean aqueous humor PlGF in the post-IVR eyes dramatically decreased from 1078.36 ± 755.83 to 177.64 ± 151.73 pg/mL (p<0.001). The VEGF-A level showed a similar trend from 3697.64 ± 2104.47 pg/mL to 183.54 ± 130.35 pg/mL (p<0.001).

Conclusions

Aqueous humor concentrations of VEGF-A and PlGF were significantly elevated in the eyes with NVG, and there was a positive correlation between the levels. After an IVR treatment, VEGF-A and PlGF were significantly decreased in NVG eyes.     

冬眠动物骨骼肌生理适应及机制的研究进展

非冬眠动物的骨骼肌在废用条件下会发生明显的萎缩。冬眠动物在历经数月的冬眠期骨骼肌废用后,仍能保持较完整的形态结构与良好的收缩功能,成为天然的抗废用性肌萎缩动物模型。探明冬眠动物骨骼肌对废用的生理适应机制,是生理生态学领域的重要课题之一。本文从形态结构、肌纤维类型和收缩功能等方面综述了冬眠动物对冬眠期骨骼肌废用状态的生理适应,并从蛋白质代谢、生长与分化的调控、代谢类型的调控、氧化应激以及线粒体结构与氧化能力等方面分析了冬眠期骨骼肌生理适应的可能机制。