UPLC(超高效液相色谱)/MS/MS联用技术测定全血中的西罗莫司

目的 建立全血中西罗莫司的UPLC（超高效液相色谱）／MS／MS测定方法,方法 全血样品中加入他克莫司（FK506）作内标，用叔丁基甲醚进行提取。以乙腈与含千分之五甲酸的水溶液为流动相（40：50），色谱柱为AcquityUPLCTMBEHC。（50mm×2．1mm，1．7μm），流速0．5ml／min。三重四极杆质谱采用正离子模式，离子采集方式为多反应监测模式（MRM），离子源温度105℃，离子源电离电压为3300V，雾化气流速500L／h。采集离子（母离子／子离子）西罗莫司为931．2／864．1，FK506为822．0／577．0。结果 西罗莫司在1～240腿／L浓度范围内呈良好的线性（n=0．9995）。日内、日间精密度均在15％以内，提取回收率大于75．0％，方法回收率为95．0％～98．5％，最低检测限为0．2μg／L。结论 本方法灵敏、准确，适合临床西罗莫司的全血分析。     

MK801慢性处理引起皮层神经元3H-GABA释放的改变

Excitatory amino acids are involved in acute and chronic neurodegenerativediseases.Little is known about the potential consequences of chronic blockade of NMDA receptors (onesubtype of excitatory amino acid receptors).Receptor function measured as ³H-GABA release inculture media after pretreatment with MK801 was studied in rat cortical neurons in primary cultures.Cultured neurons were exposed to 1μmol·L^-1 MK801 for 4 days since the 14th day.Glutamate( 1 mmol·L^-1 )evoked ³H-GABA release was shown to be significantly increased(control0.2174‰±1.40‰; MK801 treatment 0.763%±0.192%).KCl 40 mmol·L^-1 stimulation showedno such effect.This result suggests that the NMDA receptor function of releasing neurotransmitterschanged after chronic treatment with noncompetitive antagonists.     

灵芝多糖锗的抗肿瘤及免疫增强作用研究

目的 :研究灵芝多糖锗对实验性肉瘤和腹腔巨噬细胞活性的影响。方法 :利用接种肉瘤S180 腹水型诱导的小鼠肿瘤模型 ,检测对S180 肉瘤的抑制率 ;利用酸性磷酸酶测定法 ,检测对荷瘤小鼠腹腔巨噬细胞活性的影响。结果 :灵芝多糖锗可抑制小鼠S180 肉瘤的生长 ,能增强荷瘤小鼠腹腔巨噬细胞的活性。结论 :灵芝多糖锗具有抗肿瘤及免疫增强活性。     

短暂性脑缺血发作短期预后与脑动脉狭窄特征的相关性研究

目的 研究短暂性脑缺血发作（TIA）患者首发症状后90 d预后与脑供血动脉的狭窄特征包括狭窄程度、分布部位及斑块性质的关系.方法 对87例首发TIA住院患者进行回顾性分析,观察颅脑动脉病变情况,对其进行90 d随访.根据患者预后情况分为稳定组、频发组、脑梗死组,收集数据进行统计分析.结果 87例患者失访8例,完成随访的79例TIA患者中稳定组34例（43.0%）,频发组37例（46.8%）,脑梗死组8例（10.2%）.3组患者血管病变特点：狭窄血管共135条,稳定组血管狭窄构成比：轻度77.4%（24/31）,中度16.1%（5/31）,重度3.2%（1/31）,闭塞3.2%（1/31）;频发组血管狭窄构成比;轻度73.9%（48/65）,中度10.8%（7/65）,重度4.6%（3/65）,闭塞10.8%（7/65）;脑梗死组血管狭窄构成比：轻度56.4%（22/39）,中度10.3%（4/39）,重度15.4%（6/39）,闭塞17.9%（7/39）.中轻度血管狭窄构成比以稳定组最高,其次为频发组,脑梗死组最低.重度和闭塞血管构成比脑梗死组最高,其次为频发组,稳定组最低.相关分析结果显示TIA预后与血管狭窄程度呈正相关（r=0.199,P=0.021）.发现斑块117个,稳定组斑块构成比：钙化斑51.4%（18/35）,混合斑34.3%（12/35）,软斑14.3%（5/35）;频发组斑块构成比：钙化斑23.5%（12/51）,混合斑47.1（24/51）,软斑29.4%（15/51）;脑梗死组斑块构成比：钙化斑12.9%（4/31）,混合斑48.4%（15/31）,软斑38.7%（12/31）.中稳定组钙化斑构成比最高,其次为频发组,脑梗死组最低.混合斑和软斑脑梗死组所占比例最高,其次为频发组,稳定组最低.相关分析显示TIA与血管斑块性质呈正相关（r=0.341,P=0.001）.稳定组前循环血管狭窄以颈总动脉38.9%（7/18）最多见,后循环以椎动脉颅内段30.8%（4/13）多见;频发组和脑梗死组前循环狭窄以大脑中动脉32.5%（13/40）、36.8%（7/19）最多见,后循环以椎动脉颅外段36.0%（9/25）、35.0%（7/20）多见,但差异无统计学意义（P＞0.05）.结论 TIA预后与脑供血动脉的狭窄程度及斑块性质有关,狭窄程度越重、斑块性质越不稳定,预后越差.     

A Three-Dimensional Equivalent Stiffness Model of Composite Laminates with Wrinkle Defects

The stiffness of composite laminates is easily affected by wrinkle defects. In this paper, a new effective analytical model was proposed to predict the three-dimensional equivalent elastic properties of multidirectional composite laminates with wrinkle defects. Firstly, a geometric model was established according to the microscopic characteristics of wrinkle defects. Then, based on the classical laminate theory and homogenization method, the constitutive equation and flexibility matrix of the wrinkle region were established. Finally, the equivalent stiffness parameters of unidirectional and multidirectional laminates were derived, and the effects of different wrinkle parameters and ply-stacking sequences on the stiffness of unidirectional and multidirectional laminates were studied by using the analytical model. The results show that the mechanical properties of the lamina and laminates are affected by the out-of-plane angle and in-plane angle of the wrinkle defects. The accuracy of the analytical model has been verified by the numerical model and other theoretical models, and it has the characteristics of few parameters and a high efficiency. The analytical model can be used to predict the stiffness of composite structures with wrinkle defects simply, effectively, and quantitatively. It can also be used as a tool to provide the mechanical response information of laminates with wrinkle defects.     

晶状体前囊膜下上皮细胞增殖性病变的临床病理分析

目的：观察晶状体前囊膜下上皮细胞增殖的组织病理学变化,并探讨其发生原因。方法：取不同病因致手术摘除的伴晶状体前囊膜下上皮细胞增殖的眼球标本64例,其中眼外伤36例,绝对期青光眼10例,角巩膜葡萄肿6例,视网膜母细胞瘤5例,眼内炎4例,白内障手术后2例,糖尿病性白内障1例,在光镜下进行组织病理学观察研究。结果：增殖的晶状体前囊膜下上皮细胞形态不规则,排列不整齐、向后囊膜下延伸;囊膜破裂或部分缺如 前囊膜下纤维结缔组织化生,纤维组织形成;晶状体纤维崩解、液化和钙化,结论：眼内多种病状态,如眼外伤、眼内炎、青光眼,以及糖尿病等全身疾患均可命名晶状体前囊膜下上皮细胞增殖,导致晶状体发生病变,眼内炎和眼外伤后晶状体发生增殖的程度最重。     

Chemical constituents from Amentotaxus yunnanensis and Torreyayunnanensis

In a chemical study of taxonomically related Taxaceae plants of Yunnan Province, China, seven compounds, including a new amentoflavone biflavonoid, 2,3-dihydro-7,7' '-dimethoxyamentoflavone (1), were isolated from Amentotaxus yunnanensis, and 12 isolates were obtained from Torreya yunnanensis. From the latter plant, a new abietane diterpene, torreyayunnin (7), is reported for the first time. The known isolates from A. yunnanensis have been identified as sequoiaflavone (3), sotetsuflavone (4), 7,7' '-dimethoxyamentoflavone (5), lutein, beta-sitosterol, and sequoyitol. Amentoflavone (2), sotetsuflavone (4), sciadopitysin (6), 12-hydroxydehydroabietinol, meridinol, balanophonin, (+)-pinoresinol monomethyl ether, (+)-pinoresinol monomethyl ether glucoside, erythro-1-(4-hydroxy-3-methoxyphenyl)-2-[4-[2-formyl-(E)-vinyl]-2- methoxyphenoxy]propane-1,3-diol, threo-1-(4-hydroxy-3-methoxyphenyl)-2- [4-[2-formyl-(E)-vinyl]-2-methoxyphenoxy] propane-1,3-diol, and (E)-2-butenedioic acid were identified as known isolates from T. yunnanensis. The presence of the amentoflavone biflavonoids (1, 3-5) in A. yunnanensis supports its placement in the Taxaceae. The occurrence of the biflavonoid sotetsuflavone (4) in both A. yunnanensis and T. yunnanensis suggests that these two genera are closely related. The identification and structural elucidation of these isolates were based on spectral data analysis including 1D and 2D NMR.     

胃肠道间质瘤的临床病理分析

目的:观察胃肠道间质瘤临床病理及生物学行为。方法:45例胃肠道间质瘤行常规检查及生物学分型,同时免疫组化染色CD117、CD34、SMA和S-100。结果:45例GIST中,年龄15-79岁,平均53岁。发生于胃的25例,小肠14例,结肠5例,食道1例。良性10例,交界性15例,恶性20例。细胞梭形或上皮样形或者两者混合存在,免疫组化CD117、CD34、SMA和S-100阳性率分别是91.1%、80%、35.6%和28.9%,阳性表达率与肿瘤危险性无关。结论:GIST主要见于老年人,好发于胃和小肠,CD117和CD34联合应用提高其诊断率,免疫表型与危险程度无关。     

ADAMTS4 and its proteolytic fragments differentially affect melanoma growth and angiogenesis in mice

The metalloproteinase ADAMTS4 (ADAMTS, a disintegrin‐like and metalloproteinase with thrombospondin motif)/aggrecanase‐1 is highly expressed in cartilage and has been implicated in human arthritis. Although abundantly expressed in many types of cancer, its role in cancer remains unknown. In this work, we demonstrate for the first time that full‐length ADAMTS4 and its catalytically more active N‐terminal 53 kDa autocatalytic fragment both promote B16 melanoma growth and angiogenesis in mice. In contrast, overexpression of its catalytically inactive E362A mutant or truncated fragments containing only the C‐terminal ancillary domains suppresses melanoma growth and angiogenesis under similar conditions. Structure–function mapping revealed that the single thrombospondin‐type 1 repeat domain is essential and sufficient for the antitumorigenic activity displayed by the catalytically inactive ADAMTS4 isoforms. Suppression of tumor growth and angiogenesis in mice is accompanied by a significant increase in tumor cell apoptosis, whereas tumor cell proliferation is not affected. Importantly, we identified and demonstrated the presence of novel proteolytic fragments of ADAMTS4 containing essentially only the C‐terminal ancillary domains in cultured cells, and also in human cancer tissues, coexisting with full‐length and catalytically active N‐terminal fragments. The contrasting functions toward tumor growth in mice by the wild‐type proteinase and its catalytically inactive mutant correlate with their contrasting influences on angiogenesis signaling pathway molecules in B16 melanoma in mice. Our results suggest a complex role for ADAMTS4 in cancer with the functional balance of protumorigenic and antitumorigenic isoforms likely to act as an important parameter in determining the net influence of this metalloproteinase on tumor growth in vivo.     

人NGAL克隆表达及抗体的制备与鉴定

目的：高效表达人中性粒细胞明胶酶相关脂质运载蛋白（neutrophil gelatinase-associated lipocalin,NGAL）重组蛋白并制备其多克隆及单克隆抗体。方法：人工合成经密码子优化的NGAL基因,构建原核表达重组质粒pW28-NGAL,IPTG诱导表达后分离纯化并分析在溶液中的聚集状态;获得的rhNGAL抗原免疫兔子制备多克隆抗体,免疫小鼠筛选高效价的特异性单克隆抗体并鉴定抗体亚型;Protein G亲合柱纯化,等温滴定量热法及非竞争ELISA法检测抗体的亲和力,SDS-PAGE分析纯度;最后运用Western blot、免疫荧光和免疫组化对抗体进行鉴定。结果：高效获得高纯度NGAL重组蛋白,主要以单体形式存在;成功制备兔多抗,同时筛选获得6株高效价单克隆抗体;其中,25号单抗为IgG1亚型,余者均属IgG2a亚型,且19、35号单抗的亲和常数分别为3.06×109、2.14×109。SDS-PAGE分析表明纯度均大于90%。进一步的鉴定结果表明制备的抗体皆具有良好免疫反应性及特异性,且单抗的特异性明显高于多抗。结论：本研究利用密码子优化技术高效表达制备了NGAL重组蛋白,获得了其高效价多克隆抗体和高特异性单克隆抗体,为NGAL的病理生理作用等功能研究提供了良好的实验材料,为NGAL临床免疫检测诊断试剂的国产化奠定了有力的基础。