Genetic diversity of VP3 of goose parvovirus isolated from Southeastern China during 2012–2013

Although vaccine and passive immunotherapy were widely used to prevent goose parvovirus (GPV) infection in goose industry, GPV still poses a big problem in Southeastern China. In this study, 23 GPV isolates were isolated from goslings suspected with GPV infection in Southeastern China during 2012–2013, and the genetic diversity of VP3 of GPV was analyzed. Phylogenetic tree revealed that these isolates could be clustered into two groups, and 11 of 23 could be further clustered into a new subgroup. Moreover, eight novel mutations and seventeen conserved amino acids were found in these 23 isolates in comparison with isolates previously deposited in GenBank. These isolates and findings not only provide insights into the etiology and molecular characteristics of GPV endemic in Southeastern China, but also enrich the GPV genetic information for better controlling the disease.     

miR-21反义寡核苷酸增强地西他滨体外抗白血病效应

目的:研究miR-21反义寡核苷酸(anti-miR-21 oligonucleotide,AMO)对地西他滨(decitabine,DCA)抗白血病效应的影响及可能机制。方法:将AMO和无义寡核苷酸(scramble oligonucleotide,SCR)通过脂质体转染导入HL-60细胞,实时荧光定量PCR(real-time PCR)验证转染效率,再分别与DCA 0.5、2.0和4.0μmol/L作用48 h。Real-time PCR分别检测人周期节律蛋白3(h Per3)mRNA表达,Annexin V/PI法检测凋亡,流式细胞术检测CD117和CD11b平均荧光强度(MFI)。结果:AMO转染组miR-21表达(0.35±0.07)低于空白组(0.71±0.07)和SCR转染组(0.66±0.05),差异有统计学意义(P0.05)。AMO转染组的HL-60细胞DCA的IC50低于空白组和SCR转染组(P0.01)。同一浓度下,AMO组的早期凋亡率、CD11b的MFI和h Per3 mRNA均高于同一浓度药物作用的空白组和SCR组,CD117 MFI均低于同一浓度药物作用的空白组和SCR组,差异均具有统计学意义(P0.01)。结论:AMO能显著促进DCA体外抗白血病效应,其机制可能与其协助激活h Per3的表达有关。     

放射性核素131I在甲亢治疗中的应用现状和进展

以Graves病为代表,探讨放射性核素131I在甲亢治疗学中的应用,归纳、总结和讨论131I在甲亢治疗中的适应症、剂量、联合治疗、疗效和转归.与传统治疗方法相比,131I治疗甲亢具有快速简便、不良反应少、治疗效果好、费用低等优点,逐渐成为甲亢治疗的主流方法,但治疗时需注意向甲减的转归.     

桂芍知母汤对类风湿关节炎VEGF的作用探讨

通过对桂芍知母汤及类风湿关节炎的全面认识,探讨该药物对类风湿关节炎血管内皮生长因子(VEGF)的作用。     

瘦素促进肺成纤维细胞向肌纤维母细胞的转分化

目的探讨瘦素对肺成纤维细胞向肌纤维母细胞转分化的影响及其作用机制。方法体外培养HFL-1人胚肺成纤维细胞,用(0~200)ng/m L重组人瘦素(r HL)干预或联合转化生长因子β1(TGF-β1)处理HFL-1细胞,Western blot法测定α平滑肌肌动蛋白(α-SMA)、蛋白激酶B(AKT)、磷酸化的AKT(p-AKT)的表达。CCK-8法测定细胞增殖,ELISA测定细胞上清液1型胶原蛋白含量。结果 (50、100、200)ng/m L的r HL处理HFL-1细胞48 h,α-SMA的蛋白表达水平均较对照组明显上调;与200 ng/m L r HL或5 ng/m L TGF-β1单独处理的细胞相比,200 ng/m L r HL和5 ng/m L TGF-β1联合处理HFL-1细胞48 h,HFL-1细胞α-SMA蛋白表达水平显著上调。r HL能够上调HFL-1细胞p-AKT的表达,LY294002可抑制r HL对HFL-1细胞α-SMA表达的诱导作用。与对照组相比,(12.5、25、50、100、200)ng/m L作用72 h,细胞存活率无显著性差异。(50~200)ng/m L r HL作用48 h,细胞上清液中1型胶原蛋白含量明显升高。结论瘦素能够促进肺成纤维细胞向肌纤维母细胞转分化,其作用机制与激活PI3K/AKT信号通路有关。     

首发抑郁症患者服药前后MMSE、HAMD与ERP的对比研究

目的：对首发抑郁症患者在治疗前后进行简易智力状态量表（MMSE）、汉密尔顿抑郁量表（HAMD）及事件相关电位（ERP）检测,探讨抑郁症患者早期的认知功能。方法：对抑郁症患者组100例及正常对照组80例进行MMSE、HAMD及 ERP检查,并对最终进入研究的53例患者进行随访研究。抑郁组在抗抑郁药物治疗前和治疗12周、6个月、1年后各进行一次上述各指标的测定,并与对照组进行比较。结果：①抑郁症组治疗12周后 HAMD评分较前明显好转,差异有统计学意义（ P＜0．05）;MMSE ,总分及ERP波形较治疗前有所改善,差异有统计学意义（P＜0．05）,但MMSE仍较对照组差,P3波潜伏期较对照组延长,波形分化差,差异有统计学意义（ P＜0．05）。②抑郁症组治疗6个月时HAMD评分较前进一步好转,差异有统计学意义（P＜0．05）;MMSE评分及ERP检测较治疗前进一步改善,差异有统计学意义（P＜0．05）,但MMSE短时记忆仍较对照组差,P3波潜伏期延长,波形较分化差,差异有统计学意义（P＜0．05）。③抑郁症组治疗1年 HAMD评分较前进一步好转,差异有统计学意义（P＜0．05）;MMSE评分及ERP中N2、P3波潜伏期和波形较用药前进一步改善,差异有统计学意义（P＜0．05）。④Logistic回归分析：年龄和首发病程是影响抑郁症患者认知功能变化的独立危险因素。结论：①ERP检测能为抑郁症患者认知功能变化的诊断提供较客观的指标。②药物治疗1年后,抑郁症患者MMSE评分的注意与计算力、短时记忆好转,HAMD值降低,患者抑郁障碍改善明显,表明MMSE、HAMD及ERP检查对首发抑郁症患者认知功能、抑郁障碍研究具有重要意义。     

门电路调控血池成像技术对隐匿性冠心病灌注与舒缩功能的特征研究

目的利用门电路调控血池成像技术探讨隐匿性冠心病(LCHD)患者的心脏灌注与舒缩的功能学变化特点,旨在提高早期确诊率。方法选择164例患者,分3组。LCHD组39例患者,其中男性31例,女性8例;年龄28~68岁,平均年龄50.0岁。阴性对照(NC)组46例,其中男性33例,女性13例;年龄17~72岁,平均年龄47.3岁。阳性对照组(即陈旧心肌梗死组,OMI组)79例,其中男性73例,女性6例;年龄26~83岁,平均年龄57.4岁。分别进行心肌灌注断层显像和门电路调控心室功能闪烁显像,各组参数分析比较时,将年龄作为协变量进行协方差分析,然后进行F检验。结果除最大充盈时间(TPFR)外,3组间左心室舒缩功能参数(LVEF、PFR和PER)间差异具有显著统计学意义(P0.01);两两比较,LCHD组与NC组差异无统计学意义(P0.05);但LCHD组泵功能显著高于OMI组(P0.01)。LCHD组左心室功能参数分析的结果显示,心脏充盈、收缩变化幅度未超过20%,心脏代偿掩盖其器质性变化。结论 LCHD的功能影像学特征呈现为可逆性放射性缺损或分布减低,对心脏舒缩功能的损伤程度明显低于OMI组。门电路调控的闪烁成像技术应用于LCHD早期无创诊断,可发现心脏轻度器质性变化的特点。     

Synthesis of Novel Chain‐End‐Functionalized Polyethylenes via Thiol‐ene Click Chemistry

Chain‐end‐functionalized polyethylenes (Cef‐PEs: PE? S? SH (Cef₁), PE? S? furfuryl (Cef₂), PE? S? NH₂ (Cef₄), PE? S? NH₂?HCl (Cef₅), PE? S? COONa (Cef₆), PE? S? CHA(Cef₇), PE? S? SO₃Na (Cef₈), PE? S? OH (Cef₉) and PE? S? COOH (Cef₁₀)) are synthesized by thiol‐ene addition of vinyl‐terminated polyethylene (v‐PE) with 1,2‐ethanedithiol, furfurylmercaptan, cysteamine, cysteamine hydrochloride, sodium thioglycolate, L‐cysteine hydrochloride, sodium 2‐mercaptoethanesulfonate, mercaptoethanol, and thioglycolic acid, respectively. PE? S? t? NCO (Cef₃) is obtained by subsequent reaction of Cef₉ with anhydrous 4,4′‐MDI. Cef‐PE_1–8 are reported for the first time. The conversions of all the reactions are up to 95%–100%. All the Cef‐PEs are characterized by NMR, GPC, DSC, FT‐IR, and TGA. In the experiments, it is found that alkaline mercaptans and solvents are adverse to the thiol‐ene click reactions.

     

Layered hydrogels accelerate iPSC-derived neuronal maturation and reveal migration defects caused by MeCP2 dysfunction

Probing a wide range of cellular phenotypes in neurodevelopmental disorders using patient-derived neural progenitor cells (NPCs) can be facilitated by 3D assays, as 2D systems cannot entirely recapitulate the arrangement of cells in the brain. Here, we developed a previously unidentified 3D migration and differentiation assay in layered hydrogels to examine how these processes are affected in neurodevelopmental disorders, such as Rett syndrome. Our soft 3D system mimics the brain environment and accelerates maturation of neurons from human induced pluripotent stem cell (iPSC)-derived NPCs, yielding electrophysiologically active neurons within just 3 wk. Using this platform, we revealed a genotype-specific effect of methyl-CpG-binding protein-2 (MeCP2) dysfunction on iPSC-derived neuronal migration and maturation (reduced neurite outgrowth and fewer synapses) in 3D layered hydrogels. Thus, this 3D system expands the range of neural phenotypes that can be studied in vitro to include those influenced by physical and mechanical stimuli or requiring specific arrangements of multiple cell types.Neuronal migration and maturation is a key step in brain development. Defects in this process have been implicated in many disorders, including autism (1) and schizophrenia (2). Thoroughly understanding how neural progenitor cell (NPC) migration is affected in neurodevelopmental disorders requires a means of dissecting the process using cells with genetic alterations matching those in patients. Existing in vitro assays of migration generally involve measurement of cell movement across a scratch or gap or through a membrane toward a chemoattractant in 2D culture systems. Although widely used, such assays may not accurately reveal in vivo differences, as neuronal migration is tightly regulated by physical and chemical cues in the extracellular matrix (ECM) that NPCs encounter as they migrate.In vitro 3D culture systems offer a solution to these limitations (3–7). Compared with 2D culture, a 3D arrangement allows neuronal cells to interact with many more cells (4); this similarity to the in vivo setting has been shown to lengthen viability, enhance survival, and allow formation of longer neurites and more dense networks in primary neurons in uniform matrices or aggregate culture (8, 9). Indeed, 3D culture systems have been used to study nerve regeneration, neuronal and glial development (10–12), and amyloid-β and tau pathology (13). Thus, measuring neuronal migration through a soft 3D matrix would continue this trend toward using 3D systems to study neuronal development and pathology.We sought to develop a 3D assay to examine potential migration and neuronal maturation defects in Rett syndrome (RTT), a genetic neurodevelopmental disorder that affects 1 in 10,000 children in the United States and is caused by mutations in the X-linked methyl-CpG-binding protein-2 (MECP2) gene (14). Studies using induced pluripotent stem cells (iPSCs) from RTT patients in traditional 2D adherent culture have revealed reduced neurite outgrowth and synapse number, as well as altered calcium transients and spontaneous postsynaptic currents (1). However, 2D migration assays seemed unlikely to reveal inherent defects in this developmental process, which could be affected because MeCP2 regulates multiple developmental related genes (15). Migration of RTT iPSC-derived NPCs has not previously been studied.Using a previously unidentified 3D tissue culture system that allows creation of layered architectures, we studied differences in migration of MeCP2-mutant iPSC-derived versus control iPSC-derived NPCs. This approach revealed a defect in migration of MeCP2-mutant iPSC-derived NPCs induced by either astrocytes or neurons. Further, this 3D system accelerated maturation of neurons from human iPSC-derived NPCs, yielding electrophysiologically active neurons within just 3 wk. With mature neurons derived from RTT patients and controls, we further confirmed defective neurite outgrowth and synaptogenesis in MeCP2-mutant neurons. Thus, this 3D system enables study of morphological features accessible in 2D system as well as previously unexamined phenotypes.     

Structure of Arabidopsis CESA3 catalytic domain with its substrate UDP-glucose provides insight into the mechanism of cellulose synthesis

Cellulose is synthesized by cellulose synthases (CESAs) from the glycosyltransferase GT-2 family. In plants, the CESAs form a six-lobed rosette-shaped CESA complex (CSC). Here we report crystal structures of the catalytic domain of Arabidopsis thaliana CESA3 (AtCESA3^CatD) in both apo and uridine diphosphate (UDP)-glucose (UDP-Glc)–bound forms. AtCESA3^CatD has an overall GT-A fold core domain sandwiched between a plant-conserved region (P-CR) and a class-specific region (C-SR). By superimposing the structure of AtCESA3^CatD onto the bacterial cellulose synthase BcsA, we found that the coordination of the UDP-Glc differs, indicating different substrate coordination during cellulose synthesis in plants and bacteria. Moreover, structural analyses revealed that AtCESA3^CatD can form a homodimer mainly via interactions between specific beta strands. We confirmed the importance of specific amino acids on these strands for homodimerization through yeast and in planta assays using point-mutated full-length AtCESA3. Our work provides molecular insights into how the substrate UDP-Glc is coordinated in the CESAs and how the CESAs might dimerize to eventually assemble into CSCs in plants.

Cellulose, a linear homopolymer of d-glucopyranose linked by β-1,4-glycosidic bonds, is the major structural component of the cell walls of plants, oomycetes, and algae and constitute the most abundant biopolymer on Earth (1). Cellulose is synthesized by cellulose synthases (CESAs) that belongs to the glycosyltransferase GT-2 superfamily (1, 2). In land plants, cellulose is produced at the plasma membrane by six-lobed rosette-shaped CESA complexes (CSCs) where each CESA is thought to synthesize one cellulose chain (3). The precise number of CESAs per CSC is unresolved but estimated to range between 18 and 36 (4–6).Plants contain multiple cesa genes, with 10 found in the Arabidopsis genome (7). Of these, CESA1, CESA3, CESA6, and the CESA6-like CESAs (i.e., CESA2, CESA5, and CESA9) are involved in primary cell wall formation, whereas CESA4, CESA7, and CESA8 participate in secondary cell wall formation (8–12). These two types of CSCs form heterotrimeric complexes with a ratio of 1:1:1 (13, 14). The Arabidopsis CESAs share an overall sequence identity of ∼60% and have seven transmembrane helices (15). In plants, the catalytic domain (CatD) of the CESAs is located between the second and third transmembrane helices and contains a canonical D, D, D, QxxRW motif (1). While there are similarities between the plant CatD and its counterpart in bacterial cellulose synthases, the CatD is flanked by two plant-specific domains, the so-called plant-conserved region (P-CR) and class-specific region (C-SR) (16). These domains are proposed to have important functions in cellulose synthesis and CESA oligomerization (17).The oligomerization of plant CESAs is thought to be important for the final CSC assembly, and multiple oligomeric states of CESAs, including homodimers, have been reported (18, 19). For example, immunoprecipitation assays using CESA7 fused to a dual His/STRP-tag demonstrated that CESA4, CESA7, and CESA8 could form independent homodimers, and it was hypothesized that the CESA homodimerization may contribute to early stages of CSC assembly. These homodimers might then be converted into CSC heterotrimeric configurations (19). This feature poses a marked difference from the bacterial cellulose synthase complex. However, how CESA homodimers are formed and how they function in cellulose synthesis are unknown.To comprehend the mechanisms behind plant cellulose synthesis, it is essential to acquire structural information about plant CESAs. Indeed, the BcsA–BcsB complex structure from Rhodobacter greatly aided our understanding of the cellulose synthesis in bacteria (20). Nevertheless, there are many differences between bacterial and plant CESAs and the corresponding protein complexes. Extensive efforts have been undertaken to acquire plant CESA structural information, including homology modeling and small-angle X-ray scattering analyses (5, 6, 16, 21, 22). While these efforts have been important to form new hypotheses, they did not reveal significant insights into substrate coordination, cellulose chain extrusion, and complex assembly. Recently, a homotrimeric CESA8 structure from Populus tremula × tremuloides was resolved by cryogenic electron microscopy (cryo-EM), which offered significant new molecular understanding of cellulose microfibril biosynthesis and CESA coordination within the CSC (15). Here we report the crystal structures of Arabidopsis CESA3 CatD (AtCESA3^CatD) in apo and uridine diphosphate (UDP)-glucose (UDP-Glc) bound forms and outline how the CatD might contribute to CESA homodimerization and substrate coordination.