Inhibition of acetylcholinesterase by two genistein derivatives: kinetic analysis,molecular docking and molecular dynamics simulation

In this study two genistein derivatives (G1 and G2) are reported as inhibitors of acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE), and differences in the inhibition of AChE are described. Although they differ in structure by a single methyl group, the inhibitory effect of G1 (IC₅₀=264 nmol/L) on AChE was 80 times stronger than that of G2 (IC₅₀=21,210 nmol/L). Enzyme-kinetic analysis, molecular docking and molecular dynamics (MD) simulations were conducted to better understand the molecular basis for this difference. The results obtained by kinetic analysis demonstrated that G1 can interact with both the catalytic active site and peripheral anionic site of AChE. The predicted binding free energies of two complexes calculated by the molecular mechanics/generalized born surface area (MM/GBSA) method were consistent with the experimental data. The analysis of the individual energy terms suggested that a difference between the net electrostatic contributions (ΔE_ele+ΔG_GB) was responsible for the binding affinities of these two inhibitors. Additionally, analysis of the molecular mechanics and MM/GBSA free energy decomposition revealed that the difference between G1 and G2 originated from interactions with Tyr124, Glu292, Val294 and Phe338 of AChE. In conclusion, the results reveal significant differences at the molecular level in the mechanism of inhibition of AChE by these structurally related compounds.KEY WORDS: Genistein derivatives, Acetylcholinesterase (AChE), Kinetics analysis, Molecular docking, Molecular dynamics simulation, MM/GBSAAbbreviations: ACh, acetylcholine; AChEIs, acetylcholinesterase inhibitors; AChE, acetylcholinesterase; AD, Alzheimer׳s disease; BuChE, butyrylcholinesterase; BuSCh, S-butyrylthiocholine chloride; CAS, catalytic active site; DTNB, 5,5′-dithiobis-(2-nitrobenzoic acid); GAFF, generalized AMBER force field; G1, 3-(4-methoxyphenyl)-7-(2-(piperidin-1-yl)ethoxy)-4H-chromen-4-one; G2, (S)-3-(4-methoxyphenyl)-7-(2-(2-methylpiperidin-1-yl)ethoxy)-4H-chromen-4-one; iso-OMPA, tetraisopropyl pyrophosphoramide; MD, molecular dynamics; MM/GBSA, molecular mechanics/generalized born surface area; PAS, peripheral anionic site; PDB, protein data bank; PME, particle mesh Ewald; RMSD, root-mean-square deviation; S-ACh, acetylthiocholine iodide; ΔE_ele, electrostatic energy contribution; ΔE_MM, gas-phase interaction energy between receptor and ligand; ΔE_vdw, van der Waals energy contribution; SASA, solvent accessible surface area; ΔG_exp, experimental binding free energy; ΔG_GB, polar desolvation energy term; ΔG_pred, total binding free energy; ΔG_SA, nonpolar desolvation energy term; ΔS, conformational entropy contribution     

不同居群的新疆虫草发酵液体外抗氧化活性的研究

目的对不同居群的8种新疆虫草发酵液进行体外抗氧化的研究。方法采用还原能力测定法,Fenton反应法以及邻苯三酚自氧化法研究新疆虫草发酵液体外抗氧化活性。结果 DH001虫草发酵液的还原力最强,吸光度值最大,达1.216;BH001虫草发酵液和BE002虫草发酵液清除羟自由基能力最大,清除率分别达98.7%和95.4%;8种虫草发酵液清除超氧阴离子自由基活性均较低。结论新疆虫草发酵液具有一定的抗氧化活性。     

Free cholesterol levels in plasma from the maternal, placental and fetal circulations during human pregnancy

A sensitive, precise assay for free plasma cholesterol has been developed and used to measure the levels of free cholesterol in plasma samples from sites in the maternal, placental and fetal circulations during human pregnancy. These results demonstrate the removal, during passage through the placenta, of sufficient free cholesterol from the maternal circulation for use as the biosynthetic precursor of placental neutral steroid hormones. There is a suggestion from preliminary data on the arterio-venous differences in the umbilical circulation that a de novo fetal synthesis of cholesterol occurs.     

Autoradiographic mapping of dopamine-D2/D3 receptor stimulated [35S]GTPgammaS binding in the human brain

Agonist stimulated [35S]guanosine 5'-gamma-thiotriphosphate ([35S]GTPgammaS) binding autoradiography was established for the examination of dopamine-D2/D2 receptors in human brain sections. The distribution of G proteins activated by dopamine-D2/D3 receptors was studied in whole hemisphere cryosections. Dopamine stimulated [35S]GTPgammaS binding in brain regions with high densities of dopamine D2-like receptors, i.e. putamen (23 +/- 2%, mean +/- SEM,% stimulation over basal binding), caudate (20 +/- 0%) and substantia nigra (22 +/- 2%), but also in regions with lower receptor densities such as amygdala (17 +/- 8%), hippocampus (16 +/- 6%), anterior cingulate (13 +/- 3%), and thalamus (12 +/- 2%). Dopamine stimulated [35S]GTPgammaS binding to significantly higher levels in the dorsal than in the ventral part of the striatum. Dopamine caused low or very low stimulation in all cortical areas. Raclopride, a selective D2/D3 receptor antagonist, potently inhibited dopamine stimulated [35S]GTPgammaS binding, whereas R(+)-7-chloro-8-hydroxy-3-methyl-1-phenyl-2,3,4,5-tetrahydro-1H-3-benzazepine hydrochloride (SCH23390), a selective D1 antagonist, did not block the [35S]GTPgammaS binding response stimulated by dopamine. Hence, the stimulatory effect of dopamine was primarily mediated by D2/D3 receptors. Quinpirole stimulated [35S]GTPgammaS binding in the same regions as dopamine. The maximal level of stimulation induced by dopamine and quinpirole was not significantly different. The present study demonstrates that agonist stimulated [35S]GTPgammaS binding autoradiography could be a suitable technique for the examination of dopamine-D2/D3 receptors in the human brain. This functional assay could provide useful new information about dopamine receptor/G protein coupling in the postmortem human brain, and reveal possible disease related alterations of the interaction between D2/D3 receptors and G proteins.     

HLA—B27表达阴性的早中期强直性脊柱炎患者调查

目的：对4年多在我院门诊和病房确诊的早中期强直性脊柱炎（AS）患者进行HLA—B27检测的结果做了对比分析,以了解本地HLA—B27表达阴性与表达阳性的确诊早中期AS患者之间有何差异。方法：采用流式细胞术检测确诊的298例早中期AS患者的HLA-27抗原。研究样本来源于2006年9月～2011年2月在我院临床诊断为早中期AS的患者。结果：临床确诊的早中期AS患者中,HLA—B27表达阴性患者,男女比例约为3．3：1;HIJA—B27表达阳性患者．男女比例约为2．9：1,HLA—B27阴性患者与同性别阳性患者平均年龄比较,差异无统计学意义（P〉0．05）。临床确诊的早中期AS患者中,HIJA—B27表达阴性患者各年龄段的阴性率比较,差异无统计学意义（P〉0．05）,HLA—B27表达阳性患者各年龄段的阳性率比较,差异无统计学意义（P〉O．05）。结论：本地HLA—B27表达阴性与表达阳性的确诊早中期AS患者之间,男女比例、平均年龄、HLA—B27阳性率和阴性率无显著差异。在临床诊断时,对HLA—B27表达阴性的疑似患者也要给予同样的重视,以免漏诊和误诊。     

La « folie à deux » dans l'œuvre de Clérambault

In this text, the author presents G. Gatian de Clérambault's interventions concerning « folie à deux ». These are viewed from a historical perspective from a draft of a nosography of mental diseases. The evolution of his different contributions shows de Clérambault's concern with the concept of mental automatism in his nosography, which, as it is based on the clinical identification of a supposed psychoses-generating mechanism, is similar to a structural approach. To that extent, de Clérambault's work brought a new vision to psychotic disorders in small groups and deserves recognition.     

p27蛋白和细胞周期素D1在喉鳞状细胞癌中的表达及意义

目的 :探讨喉癌及癌旁组织中抑癌基因p2 7及癌基因细胞周期素D1的表达状况 ,以及p2 7与细胞周期素D1的表达与喉癌临床特征之间的关系。方法 :用免疫组织化学S P法对 38例喉癌组织 ,癌旁组织进行p2 7及细胞周期素D1检测。结果 :p2 7及细胞周期素D1表达在喉癌和癌旁组织之间各有统计学差异。p2 7及细胞周期素D1表达与颈淋巴结转移和患者预后有相关性 (P <0 .0 5 ) ;且大多数受检喉癌组织中 ,p2 7与细胞周期素D1呈反方向表达。结论 :检测p2 7与细胞周期素D1有助于判断喉癌预后。     

Massive hemangiomas and lymphangiomas causing dystocia

The concentration of alkaline phosphatase in cervical mucus was serially determined during a menstrual cycle in five normal ovulatory women and correlated with the time of ovulation as monitored by the basal body temperature and radioimmunoassay of serum lutenizing hormone (LH), progesterone, and estradiol. The activity of alkaline phosphatase decreased significantly at midcycle just prior to the LH surge and began to rise after ovulation. Self-detection of cervical mucus alkaline phosphatase may provide a practical method of ovulation prediction.     

Morphine dependence in human neuroblastoma SH-SY5Y cells is associated with adaptive changes in both the quantity and functional interaction of PGE1 receptors and stimulatory G proteins

Chronic exposure of all-trans-retinoic acid-differentiated SH-SY5Y cells to morphine (10 μM; 2 days) results in sensitization of adenylate cyclase as characterized by a significant increase in both PGE1 receptor-mediated as well as receptor-independent (NaF, 10 mM; forskolin, 100 μM) stimulation of effector activity. To investigate the underlying biochemical alterations, chronic opioid regulation of each of the components comprising the stimulatory PGE, receptor system was examined. On receptor level, chronic morphine treatment was found to reduce PGE₁ receptor number (B_max) by approximately 40%, whereas their affinity slightly increased. Binding experiments performed in the presence of GTPγS (100 μM) further indicate that the decrease in PGE1 receptor density is associated with a loss of functionally G protein-coupled receptors. On post-receptor level, chronic morphine treatment substantially increased the abundance and functional activity of stimulatory G proteins, as assessed by cholera toxin-catalyzed ADP-ribosylation of G_sα and S49 cyc⁻ reconstitution assays. No changes were found on the level of adenylate cyclase. Evaluation of the functional interaction between PGE₁ receptors and G_s in situ by application of a C-terminal anti-G_sα antibody revealed a more intense coupling efficiency between these two entities, since a significant higher amount of antibody (2.3-fold) was required in morphine dependent cell membranes to half-maximally attenuate PGE₁ receptor-stimulated adenylate cyclase activity. In addition, limitation of the amount of functionally available G_sα within the PGE₁ receptor/adenylate cyclase signal transduction cascade abolished the generation of a supersensitive adenylate cyclase response during the state of naloxone (100 μM)-precipitated withdrawal. These data demonstrate that in human neuroblastoma SH-SY5Y cells chronic morphine-induced sensitization of adenylate cyclase is associated with distinct quantitative and qualitative adaptations within the stimulatory adenylate cyclase-coupled PGE₁ receptor system. Thus, alterations in the functional activity of stimulatory receptor systems are suggested to contribute to the cellular mechanisms underlying opioid dependence.     

IgG4相关性疾病泌尿系统损害的临床特点分析

目的 总结和分析IgG4相关性疾病(IgG4-RD)泌尿系统损害的临床特点.方法 收集24例IgG4相关性疾病泌尿系统损害患者的临床资料,回顾性分析患者肾脏受累、治疗前后Scr、Hb、炎性反应指标和IgG4亚类水平的变化情况.结果 2010年8月至2014年5月经北京协和医学院肾内科确诊的IgG4相关性疾病泌尿系统损害病例24例,平均受累器官数目(4.38±1.55)个.治疗前血清IgG4[M(范围)]为12 750(1 460～ 59 400) mg/L,均伴炎性反应指标明显升高.20例患者有检测尿蛋白量,其中19例尿蛋白量升高,6例伴肾病综合征.21例患者Scr升高,均值为(410.48±352.17) μmol/L,其中3例需接受透析治疗.14例患者伴贫血,8例伴单侧或双侧肾脏体积缩小.21例Scr升高的患者中,11例患者肾功能不全因肾后性梗阻所致,3例病理证实为肾小管间质肾炎.与治疗前相比,24例患者糖皮质激素联合或不联合免疫抑制剂治疗1周后Scr明显下降(P＜0.05),治疗1个月后Scr下降更为明显(P＜0.01),3例透析患者均脱离透析.血沉于治疗1个月后亦明显下降(P＜0.01).IgG4亚类水平在治疗1个月时无显著下降(P＞0.05),治疗2个月时明显下降(P＜0.01).结论 IgG4相关疾病合并肾功能不全并不少见,对中至大量糖皮质激素治疗反应良好,治疗后Scr下降早于血沉和IgG4亚类.