Conformational preferences in a benzodiazepine series of potent nonpeptide fibrinogen receptor antagonists

Previously, we reported the direct design of highly potent nonpeptide 3-oxo-1,4-benzodiazepine fibrinogen receptor antagonists from a constrained, RGD-containing cyclic semipeptide. The critical features incorporated into the design of these nonpeptides were the exocyclic amide at the 8-position which overlaid the Arg carbonyl, the phenyl ring which maintained an extended Gly conformation, and the diazepine ring which mimicked the gamma-turn at Asp. In this paper, we investigate conformational preferences of the 8-substituted benzodiazepine analogues by examining structural modifications to both the exocyclic amide and the seven-membered diazepine ring and by studying the conformation of the benzodiazepine ring using molecular modeling, X-ray crystallography, and NMR. We found that the directionality of the amide at the 8-position had little effect on activity and the (E)-olefin analogue retained significant potency, indicating that the trans orientation of the amide, and not the carbonyl or NH groups, made the largest contribution to the observed activity. For the diazepine ring, with the exception of the closely analogous 3-oxo-2-benzazepine ring system described previously, all of the modifications led to a significant reduction in activity compared to the potent 3-oxo-1, 4-benzodiazepine parent ring system, implicating this particular type of ring system as a desirable structural feature for high potency. Energy minimizations of a number of the modified analogues revealed that none could adopt the same low-energy conformation as the one shared by the active (S)-isomer of the 3-oxo-1, 4-benzodiazepines and 3-oxo-2-benzazepines. The overall data suggest that the features contributing to the observed high potency in this series are the orientation of the 3-4 amide and the conformational constraint imposed by the seven-membered ring, both of which position the key acidic and basic groups in the proper spatial relationship.     

Molecular evidences for a role of HSV-1 in multiple sclerosis clinical acute attack

To verify the possible role of human herpesviruses as triggering or aggravating factors in relapsing-remitting multiple sclerosis (RRMS) clinical acute attack, we studied the prevalence of some herpesviruses in the peripheral blood mononuclear cells (PBMCs) collected from 22 MS patients during an MS relapse and in a stable phase and from 18 healthy controls (HC). DNA belonging to Herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2), Human cytomegalovirus (HCMV), Epstein-Barr virus (EBV) and Human Herpes virus 6 (HHV-6) has been searched by specific nested polymerase chain reaction (n-PCR). EBV and HHV6 DNA has been detected with high frequency in acute and stable MS and in healthy controls without significant differences. HCMV DNA was observed both in acute and stable MS but not in HC, and, more interestingly, HSV-1 DNA was only found in 13% of acute MS, while both stable MS and healthy controls were negative. On the basis of these results we focused on HSV-1, and to confirm them and to demonstrate that HSV-1 is actively replicating in MS patients during clinical relapse, we searched both messenger RNA (mRNA) and DNA of HSV-1 in the PBMCs of 15 acute MS patients and 15 healthy controls. We found HSV-1 mRNA and DNA in a significant number of acute MS patients but not in the control group. On the whole these data indicate that HSV-1 reactivate in the peripheral blood of MS patients during clinical acute attack and probably play a role in the triggering of MS relapses.     

膀胱移行细胞癌微卫星不稳定性及其机制的研究

目的　探讨膀胱移行细胞癌 (BTCC)染色体微卫星不稳定性的表现及与基因突变的关系。方法　采用聚合酶链反应 (PCR)方法检测 4 0例 BTCC患者 5个微卫星位点的改变 ,同时用同样的方法检测癌组织中BAX基因和转化生长因子 (TGF) - β 型受体基因移码突变的情况。结果　至少发生一个微卫星位点改变的阳性率为 82 % (33/ 4 0 ) ,D9S16 2、D16 S4 76、D9S5 4、FGA和干扰素 (IFN) - A1位点改变各自的阳性率分别为 5 8%(2 3/ 4 0 )、 4 2 % (17/ 4 0 )、 38% (15 / 4 0 )、 4 8% (19/ 4 0 )和 5 5 % (2 2 / 4 0 ) ,阳性检出率与良性病变差异有显著性 ,与肿瘤的分期分级无显著相关性。发生微卫星改变的 33例中 ,33% (11/ 33)和 4 2 % (14 / 33)分别可见 TGF- β 型受体基因和 BAX基因的移码突变。结论　检测染色体微卫星的改变是 BTCC早期诊断、监测复发的有效手段 ,染色体微卫星改变可能是 BTCC发生过程中多基因突变的一种表现形式     

软式膀胱尿道镜检查技巧

目的 :提高软式膀胱尿道镜检查的技术水平。方法 :报告软式膀胱尿道镜检查的成功经验及所遇问题。结果 :软式膀胱尿道镜对膀胱、尿道的观察均具有明显的优势 ,可完成活检、插管、取异物等多项操作。结论 :软式膀胱尿道镜检查痛苦小 ,可替代硬式膀胱尿道镜 ,使用广泛 ;熟练掌握软式膀胱尿道镜的操作技巧是检查成功及延长使用寿命的关键     

异丙酚对神经元缺氧损伤的保护及其作用机制

目的　研究异丙酚对离体大鼠海马神经元缺氧损伤的保护作用及其机制。方法　以原代培养的大鼠海马神经元作为研究对象 ,建立缺氧、H2 O2 和谷氨酸损伤模型 ,用MTT法测定神经元存活率。采用激光扫描共聚焦显微镜动态监测缺氧前后 ,单个海马神经元内Ca2 + 浓度和线粒体膜电位 (△Ψm)的变化。用电子自旋共振技术测定异丙酚对羟基自由基和超氧阴离子自由基的清除作用。结果　 6～ 4 8mg·L-1浓度的异丙酚可明显降低神经元缺氧损伤时的细胞死亡率 (P <0 0 1) ,作用程度均呈剂量相关 (r =0 89,P <0 0 5 ) ;2 4～ 4 8mg·L-1浓度的异丙酚可明显降低H2 O2 损伤时的细胞死亡率 (P <0 0 1) ;12～ 4 8mg·L-1浓度的异丙酚可明显降低谷氨酸损伤时的细胞死亡率 (P <0 0 1)。 3～ 4 8mg·L-1异丙酚对缺氧诱发的细胞内钙离子超载有抑制作用(P <0 0 5 ) ;12、4 8mg·L-1异丙酚能减缓缺氧引起的△Ψm降低 (P <0 0 1)。 12、4 8mg·L-1异丙酚对羟基自由基的清除率分别为 17 1%和 2 1 1% ,但对超氧阴离子自由基无清除作用。结论　异丙酚对离体培养的大鼠海马神经元缺氧性损伤具有保护作用 ,其作用机制部分与异丙酚抑制缺氧引起的 [Ca2 + ]i 异常升高、抑制线粒体膜电位的下降和清除羟基自由基有关     

丙泊酚对N-甲基-D-天冬氨酸所致PC12细胞损伤的保护作用

目的　探讨静脉麻醉药丙泊酚 (PPF)脑保护作用的可能机制。方法　乳酸脱氢酶 (LDH)法及MTT比色法判断细胞损伤程度及细胞存活率 ,Fura 2 /AM荧光标记法测定细胞内Ca2 + 浓度 ( [Ca2 + ]i)的变化 ,分光光度法测定细胞一氧化氮合酶 (NOS)活性。结果　N 甲基 D 天冬氨酸 (NMDA) 3 0 0 μmol·L-1处理4h可明显导致PC1 2细胞的损伤 ,表现为LDH释放量明显增加 ,吸光度值A570nm明显降低 ,细胞存活率降低 ,同时 [Ca2 + ]i 和NOS活性则明显增加。PPF6.2 5 ,2 5 ,1 0 0 ,40 0 μmol·L-1与NMDA同时处理PC1 2细胞则使LDH释放量显著降低 ,细胞存活率增加。PPF 1 2 .5和 1 2 5 μmol·L-1可显著降低NMDA诱导的[Ca2 + ]i 水平及NOS活性的提高。结论　PPF对NMDA所致的PC1 2细胞损伤有明显的保护作用 ,其机制可能与其抑制NMDA受体的功能 ,降低 [Ca2 + ]i,减弱Ca2 + 超载 ,并降低NMDA诱导的NOS活性增加有关。提示PPF可能是通过抑制NMDA受体 Ca2 + NOS通路的功能而产生细胞保护效应     

白术对抗体形成细胞的作用研究

目的：观察白术对小鼠抗体形成细胞的作用。方法：用100％浓度白术浸出液定期给小鼠灌胃，然后检测小鼠抗体产生能力．特异玫瑰花环结形成细胞检测(SRFC)、血清IgG检测。结果：用药组以上三项免疫学指标均增高，结论：白术有明显促进抗体形成细胞的作用。     

高效液相色谱法测定淡豆豉药材中异黄酮的含量

目的:建立高效液相色谱法测定淡豆豉中异黄酮含量。方法:采用HPLC法,色谱柱为YWG C18柱(250 mm×4.5 mm,10μm),以甲醇-水-乙酸(10:10:1)为流动相,流速为0.8 mL·min-1,检测波长260 nm,测定淡豆豉中染料木素(genistein)、大豆黄素(daidzein)含量。结果:染料木素及大豆黄素的线性范围分别为2.00-40.00μg·mL-1和2.64-52.70μg·mL-1,相关系数分别为0.9994和0.9993,不同产地的淡豆豉中异黄酮含量差异较大。结论:本法准确、简便、快速,对于控制和评价淡豆豉的品质有重要意义。     

循环式扩张治疗食管化学灼伤后狭窄20例

目的　探讨食管化学灼伤后狭窄的简单、有效治疗方法 .方法　胃造瘘术后 ,食管内置入丝线 ,从鼻腔和胃造瘘口拉出连结 .用自制的食管扩张器 ,在丝线引导下进行扩张 .结果　食管上段狭窄 4例 ,中段 4例 ,下段 2例 ,全程不规则线状狭窄 1 0例 .经 3～ 8m o间断扩张治疗 ,全部顺利通过 1 8mm扩张器 ,进普食顺利 .经 2～ 5 a随访 ,效果良好 .结论　食管化学灼伤后狭窄行循环式扩张治疗是一种简单、有效、安全、经济的好方法 ,值得推广应用     

双调蛋白诱导乳腺癌细胞表达尿激酶型纤溶酶原活化物

目的:在人乳腺癌模型上研究双调蛋白与尿激酶型纤溶酶原活化物表达之间的关系.方法:乳腺癌NS2T2A1细胞经双调蛋白反义cDNA质粒转染后经潮霉素B筛选获得表达双调蛋白反义RNA的AR-AS1及AR-AS3两个细胞克隆,转染空载体获得NS2T2A1 V对照细胞,接种至裸鼠皮下形成肿瘤.测定细胞及肿瘤uPA表达水平,并研究uPA与细胞侵袭性之间的关系.结果:AR-AS1及AR-AS3细胞体外及体内uPA表达均被抑制.外源性双调蛋白可刺激对照细胞uPA的表达,并部分恢复AR-AS1及AR-AS3细胞uPA的表达水平.双调蛋白反义cDNA质粒转染及抗uPA抗体均导致乳腺癌细胞体外侵袭性的降低.结论:在乳腺癌模型上,uPA表达与肿瘤细胞的侵袭密切相关.双调蛋白反义RNA表达可有效地抑制uPA的表达,进而抑制肿瘤细胞的侵袭性.