Molecular characterization of avian-like H1N1 swine influenza a viruses isolated in Eastern China, 2011

Currently, three predominant subtypes of influenza virus are prevalent in pig populations worldwide: H1N1, H3N2, and H1N2. European avian-like H1N1 viruses, which were initially detected in European pig populations in 1979, have been circulating in pigs in eastern China since 2007. In this study, six influenza A viruses were isolated from 60 swine lung samples collected from January to April 2011 in eastern China. Based on whole genome sequencing, molecular characteristics of two isolates were determined. Phylogenetic analysis showed the eight genes of the two isolates were closely related to those of the avian-like H1N1 viruses circulating in pig populations, especially similar to those found in China. Four potential glycosylation sites were observed at positions 13, 26, 198, 277 in the HA1 proteins of the two isolates. Due to the presence of a stop codon at codon 12, the isolates contained truncated PB1-F2 proteins. In this study, the isolates contained 591Q, 627E and 701N in the polymerase subunit PB2, which had been shown to be determinants of virulence and host adaptation. The isolates also had a D rather than E at position 92 of the NS1, a marker of mammalian adaptation. Both isolates contained the GPKV motif at the PDZ ligand domain of the 3′ end of the NS1, a characteristic marker of the European avian-like swine viruses since about 1999, which is distinct from those of avian, human and classical swine viruses. The M2 proteins of the isolates have the mutation (S31N), a characteristic marker of the European avian-like swine viruses since about 1987, which may confer resistance to amantadine and rimantadine antivirals. Our findings further emphasize the importance of surveillance on the genetic diversity of influenza A viruses in pigs, and raise more concerns about the occurrence of cross-species transmission events.     

Aspergillus niger DLFCC-90 rhamnoside hydrolase, a new type of flavonoid glycoside hydrolase

A novel rutin-α-L-rhamnosidase hydrolyzing α-L-rhamnoside of rutin, naringin, and hesperidin was purified and characterized from Aspergillus niger DLFCC-90, and the gene encoding this enzyme, which is highly homologous to the α-amylase gene, was cloned and expressed in Pichia pastoris GS115. The novel enzyme was classified in glycoside-hydrolase (GH) family 13.     

鳜鱼Mx蛋白全长cDNA的克隆和序列分析

Mx蛋白是一类由I型干扰素诱导表达的抗病毒蛋白.本研究以感染了鳜传染性脾肾坏死病毒(Infectious spleen and kidney necrosis virus,ISKNV)的鳜鱼为材料,提取肝脏总RNA,通过逆转录-聚合酶链式反应(RT-PCR)扩增出Mx蛋白基因的核心片段序列,再应用3'和5'快速扩增cDNA末端(RACE)方法PCR扩增Mx蛋白cDNA末端,最终获得鳜鱼Mx蛋白cDNA序列(GenBank登陆号AY392097).序列分析表明鳜鱼Mx蛋白cDNA含有2391bp,其中编码区长1881bp,编码627个氨基酸残基,推测蛋白质分子量大小为7.15kDa.鳜鱼Mx蛋白具有脊椎动物Mx蛋白共有的结构特征一个三联体GTP结合区域(GXXXSGKS/T、DXXG、T/NKXD);一个发动蛋白家族的典型结构特征序列(LPRGS/KGIVTR);以及C端高度保守的Leu拉链结构域.鳜鱼Mx蛋白全基因的获得为下一步研究鱼类Mx蛋白的抗病毒活性、作用机制,以及干扰素的检测奠定了基础.     

热处理肿瘤细胞抗原负载树突细胞抗小鼠结肠癌效应

研究了热处理肿瘤细胞抗原负载骨髓来源的树突细胞（dendritic cell,DC）对结肠癌小鼠的治疗作用.将小鼠结肠癌细胞CT26热处理后超声破膜,以其细胞裂解液负载BALB/c小鼠骨髓来源的DC,观察DC诱导肿瘤特异性细胞毒性T淋巴细胞（cytotoxic T lymphocyte,CTL）杀伤活性;并将DC接种于荷瘤小鼠皮下,观察其对肿瘤生长的抑制作用及对荷瘤小鼠生存期的影响.发现致敏DC诱导的CTL对CT26肿瘤细胞具有显著的杀伤作用.用致敏DC免疫小鼠后,对小鼠肿瘤的生长具有显著的抑制作用,并能显著延长荷瘤小鼠的生存时间.热处理肿瘤细胞抗原负载的树突细胞对结肠癌小鼠具有显著的治疗效果.     

NADPH oxidase-mediated generation of reactive oxygen species: A new mechanism for X-ray-induced HeLa cell death

Oxidative damage is an important mechanism in X-ray-induced cell death. Radiolysis of water molecules is a source of reactive oxygen species (ROS) that contribute to X-ray-induced cell death. In this study, we showed by ROS detection and a cell survival assay that NADPH oxidase has a very important role in X-ray-induced cell death. Under X-ray irradiation, the upregulation of the expression of NADPH oxidase membrane subunit gp91^phox was dose-dependent. Meanwhile, the cytoplasmic subunit p47^phox was translocated to the cell membrane and localized with p22^phox and gp91^phox to form reactive NADPH oxidase. Our data suggest, for the first time, that NADPH oxidase-mediated generation of ROS is an important contributor to X-ray-induced cell death. This suggests a new target for combined gene transfer and radiotherapy.     

Using the lymph fistula rat model to study the potentiation of GIP secretion by the ingestion of fat and glucose

Glucose-dependent insulinotropic polypeptide (GIP) is an important incretin produced in the K cells of the intestine and secreted into the circulating blood following ingestion of carbohydrate- and fat-containing meals. GIP contributes to the regulation of postprandial insulin secretion and is essential for normal glucose tolerance. We have established a method of assaying GIP in response to nutrients using the intestinal lymph fistula model. Administration of Ensure, a mixed-nutrient liquid meal, stimulated a significant increase in intestinal lymphatic GIP levels that were approximately threefold those of portal plasma. Following the meal, lymph GIP peaked at 60 min (P < 0.001) and remained elevated for 4 h. Intraduodenal infusions of isocaloric and isovolumetric lipid emulsions or glucose polymer induced lymph GIP concentrations that were four and seven times the basal levels, respectively. The combination of glucose plus lipid caused an even greater increase of lymph GIP than either nutrient alone. In summary, these findings demonstrated that intestinal lymph contains high concentrations of GIP that respond to both enteral carbohydrate and fat absorption. The change in lymphatic GIP concentration is greater than the change observed in the portal blood. These studies allow the detection of GIP levels at which they exert their local physiological actions. The combination of glucose and lipid has a potentiating effect in the stimulation of GIP secretion. We conclude from these studies that the lymph fistula rat is a novel approach to study in vivo GIP secretion in response to nutrient feeding in conscious rats.     

Transmitted/Founder Simian Immunodeficiency Virus Envelope Sequences in Vesicular Stomatitis and Semliki Forest Virus Vector Immunized Rhesus Macaques

Identification of transmitted/founder simian immunodeficiency virus (SIV) envelope sequences responsible for infection may prove critical for understanding HIV/SIV mucosal transmission. We used single genome amplification and phylogenetic analyses to characterize transmitted/founder SIVs both in the inoculum and in immunized-infected rhesus monkeys. Single genome amplification of the SIVsmE660 inoculum revealed a maximum diversity of 1.4%. We also noted that the consensus sequence of the challenge stock differed from the vaccine construct in 10 amino acids including 3 changes in the V4 loop. Viral env was prepared from rhesus plasma in 3 groups of 6 immunized with vesicular stomatitis virus (VSV) vectors and boosted with Semliki forest virus (SFV) replicons expressing (a) SIVsmE660 gag-env (b) SIVsmE660 gag-env plus rhesus GM-CSF and (c) control influenza hemagglutinin protein. Macaques were immunized twice with VSV-vectors and once with SFV vector and challenged intrarectally with 4000 TCID₅₀. Single genome amplification characterized the infections of 2 unprotected animals in the gag-env immunized group, both of which had reduced acute plasma viral loads that ended as transient infections indicating partial immune control. Four of 6 rhesus were infected in the gag-env + GM-CSF group which demonstrated that GM-CSF abrogated protection. All 6 animals from the control group were infected having high plasma viral loads. We obtained 246 full-length envelope sequences from SIVsmE660 infected macaques at the peak of infection and determined the number of transmitted/founder variants per animal. Our analysis found that 2 of 2 gag-env vaccinated but infected macaques exhibited single but distinct virus envelope lineages whereas rhesus vaccinated with gag-env-GM-CSF or HA control exhibited both single and multiple env lineages. Because there were only 2 infected animals in the gag-env vaccinated rhesus compared to 10 infected rhesus in the other 2 groups, the significance of finding single env variants in the gag-env vaccinated group could not be established.