Pro‐cognitive activity in rats of 3‐furan‐2‐yl‐N‐p‐tolyl‐acrylamide,a positive allosteric modulator of the α7 nicotinic acetylcholine receptor

Background and Purpose

α7 nicotinic acetylcholine receptors (α7 nAChRs) may represent useful targets for cognitive improvement. The aim of this study is to compare the pro‐cognitive activity of selective α7‐nAChR ligands, including the partial agonists, DMXBA and A‐582941, as well as the positive allosteric modulator, 3‐furan‐2‐yl‐N‐p‐tolyl‐acrylamide (PAM‐2).

Experimental Approach

The attentional set‐shifting task (ASST) and the novel object recognition task (NORT) in rats, were used to evaluate the pro‐cognitive activity of each ligand [i.e., PAM‐2 (0.5, 1.0, and 2.0 mg·kg⁻¹), DMXBA and A‐582941 (0.3 and 1.0 mg·kg⁻¹)], in the absence and presence of methyllycaconitine (MLA), a selective competitive antagonist. To determine potential drug interactions, an inactive dose of PAM‐2 (0.5 mg·kg⁻¹) was co‐injected with inactive doses of either agonist ‐ DMXBA: 0.1 (NORT); 0.3 mg·kg⁻¹ (ASST) or A‐582941: 0.1 mg·kg⁻¹.

Key Results

PAM‐2, DMXBA, and A‐582941 improved cognition in a MLA‐dependent manner, indicating that the observed activities are mediated by α7 nAChRs. Interestingly, the co‐injection of inactive doses of PAM‐2 and DMXBA or A‐582941 also improved cognition, suggesting drug interactions. Moreover, PAM‐2 reversed the scopolamine‐induced NORT deficit. The electrophysiological results also support the view that PAM‐2 potentiates the α7 nAChR currents elicited by a fixed concentration (3 μM) of DMXBA with apparent EC₅₀ = 34 ± 3 μM and E_max = 225 ± 5 %.

Conclusions and Implications

Our results support the view that α7 nAChRs are involved in cognition processes and that PAM‐2 is a novel promising candidate for the treatment of cognitive disorders.

Abbreviations

α7 nAChR

nicotinic acetylcholine receptor with α7 subunit

AD

Alzheimer''s disease

apparent EC₅₀

enhancement potency

ASST

attentional set‐shifting task

CD

compound discrimination

DI

discrimination index

E

exploration time

ED

extra‐dimensional

E_max

ligand efficacy

ID

intra‐dimensional

ITI

inter‐trial interval

MLA

methyllycaconitine

NORT

novel object recognition task

n_H

Hill coefficient

PAM

positive allosteric modulator

PAM‐2

3‐furan‐2‐yl‐N‐p‐tolyl‐acrylamide

Rev

reversal of discrimination

SD

simple discrimination

T1

familiarisation trial

T2

retention trial

     

显微手术联合分泌BMP4的自体MSCs靶向BTSC治疗复发脑胶质瘤

目的探讨显微外科手术切除联合分泌BMP4的自体MSCs靶向脑肿瘤干细胞(brain tumor stem cells,BTSC)治疗复发脑胶质瘤的临床疗效和安全性。方法对10例复发脑胶质瘤患者行开颅显微外科手术全切除,术中瘤腔内安置Ommaya囊,术后2 W、4 W、6 W、8 W经由Ommaya囊行瘤腔内给予分泌BMP4的自体MSCs靶向BTSC治疗。随访6～18个月,观察治疗效果。结果 10例患者均获随访,3个月无复发,无死亡;6个月复发3例,死亡1例,生存期明显延长。全部患者均无明显的不良反应,生存质量得到明显改善。结论显微外科手术切除联合分泌BMP4的自体MSCs靶向BTSC治疗复发脑胶质瘤安全显效,值得推广。     

SHH/BMP4信号在肛门直肠畸形肠神经系统发育中的作用研究

目的研究不同类型先天性肛门直肠畸形(ARM)胎鼠直肠末端肠神经系统(ENS)发育程度,探讨SHH/BMP4在其发育过程中的作用。方法利用全反式维甲酸(ATRA)诱导大鼠产生肛门直肠畸形胚胎,孕20d剖宫取胎,应用免疫组织化学方法检测对照组和高、低位ARM组直肠末端神经元特异性烯醇化酶(NSE)及骨形态发生蛋白4(BMP4)的表达;反转录-聚合酶链反应(RT-PCR)方法检测各组直肠末端SHH/BMP4mRNA的表达差异。结果对照组肠壁肌间及黏膜下神经丛可见NSE和BMP4抗体染色阳性细胞,ARM高位组与ARM低位组、对照组比较,阳性细胞的平均光密度(MOD)值明显降低,差异均有统计学意义(P<0.05);ARM低位组和对照组相比,阳性细胞的MOD值略降低,差异有统计学意义(P<0.05)。在ARM直肠末端SHH和BMP4基因表达呈正相关(P<0.01,r=0.884),对照组与ARM组相比较,BMP4和SHHmRNA的表达水平明显高于ARM组,差异均有统计学意义(P<0.05),不同ARM组之间比较,高位ARM组的表达水平显著减弱低于低位ARM组,差异也有统计学意义(P<0.05)。结论①不同类型ARM胎鼠直肠末端ENS发育程度存在差异,不仅与闭锁的位置密切相关,还可能与BMP4基因的表达水平有关。②过量的ATRA可能抑制了SHH/BMP4信号表达,干扰了直肠ENS的正常发育。     

左乙拉西坦对致痫大鼠海马组织中BMP4表达的影响

目的：建立氯化锂－匹罗卡品幼年大鼠癫痫模型,观察左乙拉西坦对癫痫海马组织中骨形成蛋白（BMP4）表达的影响,以探讨左乙拉西坦的抗癫痫作用机制。方法：将116只21d龄Wistar雄性大鼠随机分为4组,其中空白对照组（NS组）24只、左乙拉西坦对照组（LEV对照组）24只、氯化锂一匹罗卡品模型组（PILO组）34只、氧化锂一匹罗卡品模型＋左乙拉西坦治疗组（LEV治疗组）34只。PILO组和LEV治疗组大鼠首先给予腹腔注射氯化锂（125mg／kg）进行诱导,16h后腹腔注射硫酸阿托品（1mg／kg）,30min后再以匹罗卡品（60mg／kg）腹腔注射,如果没有癫痫发作,可在半小时后,再次腹腔注射匹罗卡品15rag／kg,直至充分点燃建立癫痫模型。其中,LEV治疗组,每日给予左乙拉西坦200mg／kg,连续3周进行灌胃治疗,并分别于1周,2周,3周,处死各组大鼠,并通RT--PCR的方法分别检测及观察海马组织中BMP4表达的变化。结果：PILO组与NS对照组相比,BMP4的表达增加,第1周达高峰（P〈0．01）,第2周、第3周BMP4表达逐渐下降,但较NS对照组仍高表达（P〈0．05）,LEV治疗组与PILO对照组相比,BMP4表达逐渐降低（P〈0．05）,LEV对照组与NS对照组无统计学意义（P〉0．05）。结论：癫痫发作后BMP4表达增加,提示BMP4与癫痫形成密切相关,左乙拉西坦可能通过抑制海马组织中BMP4表达而发挥抗癫痫发作。     

Intravesical delivery of 5‐aminolevulinic acid from water‐in‐oil nano/submicron‐emulsion systems

The present work reports on the development of water‐in‐oil (w/o) emulsions for the intravesical administration of 5‐aminolevulinic acid (ALA). The physicochemical properties of droplet size, zeta potential, and viscosity of the emulsions are characterized and the ability of the emulsions to release ALA following in vitro application is tested. The delivery systems are administered intravesically for 1 and 3 h in rats to examine the drug accumulation in bladder tissue. The mean size and zeta potential of the emulsions are 50–200 nm and ?3 to ?14 mV, respectively. The loading of ALA into the emulsions resulted in a slower and sustained release. The release extent was found to be inversely related to the droplet size of the emulsions. The emulsions did not increase the drug permeation into tissues during short exposure duration (1 h). When the dwell time was extended to 3 h, the systems showed a 2.7‐fold increase in the ALA concentration in the bladder wall. Images of confocal laser scanning microscopy demonstrated a higher and deeper fluorescence signal, with emulsion administration, as compared to the aqueous control. Intravesical emulsion delivery provides a significant advantage for drugs targeting bladder tissues. © 2009 Wiley‐Liss, Inc. and the American Pharmacists Association J Pharm Sci 99: 2375–2385, 2010     

Texture Optimization of Water‐in‐Oil Emulsions

The aim of this research is to demonstrate the effect of variations in certain parameters of the oily phase (OP) in water‐in‐oil (W/O) emulsions on rheological and texture properties of finished products. The formulated emulsions were selected according to an optimal experimental procedure. The applied variations were nature of the OP, its volume fraction, the hydrophilic‐lipophilic balance (HLB) value, and the surfactant proportion. Results are presented for the followed tests carried out on the emulsions: texture analysis, rheology, and particle size analysis. The oils used in the study were sweet almond oil, liquid paraffin, maize oil, cyclomethicone, dimethicone, and wheat germ oil. The resulting data demonstrate a notable influence of the volume fraction oil on hardness, viscosity, adhesiveness, and cohesiveness of W/O emulsions. Emulsion hardness and viscosity increased as the OP percentage increased; this effect being even more pronounced for the vegetable oils. In contrast, emulsion adhesiveness and cohesiveness decreased as the volume fraction oil increased. The HLB value of the surfactant mixture of the emulsion also influenced hardness, adhesiveness, and elasticity, increasing or decreasing as HLB value did.     

Differences in perimenstrual symptoms between cocaine‐abusing and non‐cocaine‐abusing women

Cocaine abuse among women has become a major health problem in the United States, yet there is little information in the literature concerning the effects of this form of substance abuse on a woman's reproductive system. This study of 65 women in residential treatment for cocaine abuse and 65 non‐cocaine‐abusing women was undertaken to determine if there are differences in frequency or severity of perimenstrual symptoms between these two groups of women. Data were collected by questionnaire and included demographics, a substance use history, and the Menstrual Distress Questionnaire developed by Moos. Findings suggested that there are statistically significant differences in frequency and severity of perimenstrual symptoms between the two groups of women.     

BMP调控成骨细胞生物活性的实验研究

目的 探讨骨形态发生蛋白（bone morphogenetic protein,BMP）对培养的正常人成骨细胞主要生物学行为的影响。方法分别以0．02、0．1、1μg/m3种浓度的BMP作用于成骨细胞,通过MTY法了解BMP的效应。以0．1μg/ml浓度的BMP作用于成骨细胞,测定BMP对成骨细胞碱性磷酸酶（alkaline phospbatase,ALP）活性及骨钙素（bone gla protein,BGP）合成等主要生物学特性的影响。结果BMP作用于成骨细胞,能促进ALP活性及BGP的合成。结论BMP能促进成骨细胞活性,并且其促进成骨增殖的效能具有浓度依赖性。     

BMP4基因rs17563位点多态性与中国华东地区部分人群非综合征性唇腭裂的关系

目的探讨骨形态发生蛋白4(BMP4)基因rs17563位点多态性与中国华东地区部分人群非综合征性唇腭裂(NSCL/P)的关系。方法收集65个唇裂核心家庭血样,分为患儿(患儿组)和患儿父母(父母组)组,另设正常儿童65例为对照组。对BMP4外显子4的rs17563位点突变碱基进行测序。比较患儿与正常儿童中的分布差异,比较患儿组与父母组的基因型和等位基因频率,并对核心家庭的数据进行单倍型相对风险分析(HRR),对含杂合子父母的核心家庭进行传递不平衡检验(TDT)。结果患儿组与对照组的基因型分布差异有统计学意义(P<0.05)。患儿组与父母组各位点基因型和等位基因分布差异无统计学意义(P>0.05),HRR结果和TDT结果无统计学意义(P>0.05)。结论 BMP4基因rs17563位点多态性与中国华东部分人群NSCL/P存在相关性。     

A comparison of medical symptoms reported by cocaine‐, opiate‐, and alcohol‐dependent patients

Substance abuse is frequently associated with adverse medical consequences. The differences in medical symptoms reported by 101 alcohol‐, 113 cocaine‐, and 107 opiate‐dependent individuals receiving outpatient treatment were studied using a 134‐item questionnaire (MILCOM). Data analysis revealed interesting and unexpected findings, with cocaine patients reporting the fewest total symptoms among the three groups. Moreover, cocaine patients reported significantly fewer CNS and musculo‐skeletal symptoms compared to both alcohol and opiate patients and significantly fewer GI and urinary symptoms than the alcohol but not the opiate patients. In addition, there were sex‐ and race‐related differences in the pattern of symptoms reported. Women reported significantly more CVS, mood, nose/throat, CNS, skin, and GI symptoms than men. Similarly, Caucasians reported significantly more mood, CNS, nose/throat, head/neck, musculoskeletal, and GI symptoms than African‐Americans. The study highlights the influence of drug of choice, gender, and race on medical needs of substance‐abusing persons.     

PLA作为BMP载体修复骨缺损实验研究

为探讨聚乳酸（PLA）作为骨形态发生蛋白（BMP）载体的可行性,及观察其成骨效果。于兔尺骨12mm骨缺损实验模型中,实验组植入以PLA为载体的BMP10mg、对照组植入以牛松质骨基质为载体的BMP10mg、空白对照组不作任何植入,分别于植入后第8、12、16、20周X线观察骨缺损修复情况,观察第12、16、20周缺损部组织学变化并图像分析各组骨小梁相对面积。结果显示术后第16周实验组和对照组骨缺损基本修复,两组成骨没有明显差异,而空白组缺损内主要形成纤维组织。研究结果表明PLA是可以用作BMP载体用于骨缺损修复的,它具有和异种松质骨基…     

骨形成蛋白（BMP—5）与IA型BMP受体在口腔鳞癌中的表达及意义

目的：认识和分析BMP及其受体BMPR-IA与口腔鳞癌发生、发展的可能关系。方法：用免疫组织化学方法分析BMP-5、BMP受体BMPR-IA在口号颊部粘膜正常上皮、慢性炎症和癌变中的表达，标本为18例正常上皮（normal buccal mucosa,NB)，24例慢性炎症（nonspecofic chronic inflammation.NCI)，58例鳞癌（squamous cell carcinoma,SCC)。结果：BMP-5、BMPR-IA在口腔粘膜的正常与慢性炎症上皮中有弱表达，NB与NCI明显差别；在鳞癌上皮中除2例BMP-5表达为阴性外，其余均有阳性或强阳性表达，明显高于NB、NCI两组；鳞癌中高分化、中分化、低分化三组间的表达无明显差别。结论：BMP-5、BMPR-IA可能参与对口腔鳞癌发生、发展的调控。     

High‐throughput screening assays for SARS‐CoV‐2 drug development: Current status and future directions

In response to the ongoing coronavirus disease 2019 (COVID-19) pandemic, a panel of assays has been developed and applied to screen collections of approved and investigational drugs for anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) activity in a quantitative high-throughput screening (qHTS) format. In this review, we applied data-driven approaches to evaluate the ability of each assay to identify potential anti-SARS-CoV-2 leads. Multitarget assays were found to show advantages in terms of accuracy and efficiency over single-target assays, whereas target-specific assays were more suitable for investigating compound mechanisms of action. Moreover, strict filtering with counter screens might be more detrimental than beneficial in identifying true positives. Thus, developing novel HTS assays acting simultaneously against multiple targets in the SARS-CoV-2 life cycle will benefit anti-COVID-19 drug discovery.     

BMP载体的运载系统及其分类的研究进展

骨形成蛋白(bone morphogenetic protein,BMP),作为转化生长因子β(TGF-β)超家族的成员,是一类多功能细胞因子。BMP能诱导未分化间充质细胞向骨和软骨发生不可逆转化,在细胞增殖、凋亡、分化和形态遗传方面具有重要作用。但是,目前BMP在临床应用上存在困难,主要因为单纯种植BMP易在体内扩散稀释,难以在新骨形成过程中充分发挥其诱导成骨作用,因此需要合适的载体在发挥支架作用的同时使BMP得到缓慢释放。本文就构成天然、无机、合成和复合材料载体的条件、载体的特性进行综述,进一步拓展BMP的临床应用范围,并对BMP载体的应用前景进行简要分析。     

BMP信号通路与牙齿发育关系的研究进展

哺乳动物牙齿发育是一个由多种信号分子进行调控的上皮—间充质相互作用的过程。其中骨形态发生蛋白(BMP)是口腔上皮中最早表达的信号分子之一,它在上皮—间充质相互诱导中起关键的调节作用。BMP信号通路是细胞内外多种因子构成的复杂精细的调控网络,这一网络中各个因子的相互作用对维持牙齿正常发育起重要作用。本文综述了BMP信号通路与牙齿发育的研究进展。     

Antisense‐mediated reduction of proprotein convertase subtilisin/kexin type 9 (PCSK9): a first‐in‐human randomized,placebo‐controlled trial

AimsLDL‐receptor expression is inhibited by the protease proprotein convertase subtilisin/kexin type 9 (PCSK9), which is considered a pharmacological target to reduce LDL‐C concentrations in hypercholesterolaemic patients. We performed a first‐in‐human trial with SPC5001, a locked nucleic acid antisense inhibitor of PCSK9.MethodsIn this randomized, placebo‐controlled trial, 24 healthy volunteers received three weekly subcutaneous administrations of SPC5001 (0.5, 1.5 or 5 mg kg^–1) or placebo (SPC5001 : placebo ratio 6 : 2). End points were safety/tolerability, pharmacokinetics and efficacy of SPC5001.ResultsSPC5001 plasma exposure (AUC(0,24 h)) increased more than dose‐proportionally. At 5 mg kg^–1, SPC5001 decreased target protein PCSK9 (day 15 to day 35: −49% vs. placebo, P < 0.0001), resulting in a reduction in LDL‐C concentrations (maximal estimated difference at day 28 compared with placebo −0.72 mmol l^–1, 95% confidence interval − 1.24, −0.16 mmol l^–1; P < 0.01). SPC5001 treatment (5 mg kg^–1) also decreased ApoB (P = 0.04) and increased ApoA1 (P = 0.05). SPC5001 administration dose‐dependently induced mild to moderate injection site reactions in 44% of the subjects, and transient increases in serum creatinine of ≥20 μmol l^–1 (15%) over baseline with signs of renal tubular toxicity in four out of six subjects at the highest dose level. One subject developed biopsy‐proven acute tubular necrosis.ConclusionsSPC5001 treatment dose‐dependently inhibited PCSK9 and decreased LDL‐C concentrations, demonstrating human proof‐of‐pharmacology. However, SPC5001 caused mild to moderate injection site reactions and renal tubular toxicity, and clinical development of SPC5001 was terminated. Our findings underline the need for better understanding of the molecular mechanisms behind the side effects of compounds such as SPC5001, and for sensitive and relevant renal toxicity monitoring in future oligonucleotide studies.     

Novel Starch‐Based PVA Thermoplastic Capsules for Hydrophilic Lipid‐Based Formulations

For decades, gelatin has been used in the rotary die process as a shell‐forming material of soft capsules because of its unique physicochemical properties. However, with respect to the encapsulation of comparatively hydrophilic lipid‐based formulations, gelatin has one considerable drawback: Immediately after production, the capsule shell contains a large amount of water (up to 35%). There is the potential for water to migrate from the capsule shell into the formulation, which will lead to a decrease in drug solubility and, in turn, the potential for drug crystallization. The present study introduces a novel capsule material that was obtained from extrusion. The starch‐based polyvinyl alcohol thermoplastic capsules (S‐PVA‐C) mainly comprised a blend of starch and PVA. Gelatin and the novel material were used to encapsulate a hydrophilic lipid‐based system of fenofibrate. Considerable water migration was observed from the soft gelatin shell to the hydrophilic formulation during drying and drug crystallization resulted in soft gelatin capsules. In contrast, S‐PVA‐C displayed no substantial water exchange or drug crystallization upon storage. The thermoplastic capsule material further exhibited more surface roughness and higher resistance to mechanical deformation compared with gelatin. In conclusion, S‐PVA‐C provided a robust drug product following encapsulation of a rather hydrophilic lipid‐based formulation.     

Role of endothelial TRPV4 channels in vascular actions of the endocannabinoid, 2‐arachidonoylglycerol

Background and Purpose

Metabolites of the endocannabinoid, 2‐arachidonoylglycerol (2‐AG) have been postulated to act as endogenous activators of TRPV4, a Ca²⁺‐permeable cation channel that plays a critical role in endothelium‐dependent relaxation. However, it is unclear if TRPV4 contributes to the vascular actions of 2‐AG.

Experimental Approach

Isometric tension recording of rat small mesenteric arteries and aortae were used to assess the effect of 2‐AG and the synthetic TRPV4 activator, GSK1016790A (GSK) on vascular reactivity. Changes in intracellular Ca²⁺ concentration and single‐channel currents were measured in TRPV4‐expressing human coronary endothelial cells.

Key Results

In mesenteric arteries, endothelium‐dependent relaxation to both 2‐AG and GSK was attenuated by structurally distinct TRPV4 antagonists, HC067047, RN1734 and ruthenium red. The responses were inhibited by K_Ca inhibitors (apamin + charybdotoxin) and a gap junction inhibitor (18α‐glycyrrhetinic acid). In contrast to GSK, 2‐AG elicited considerable relaxation independently of the endothelium or TRPV4. Inhibition of 2‐AG metabolism via monoacylglycerol lipase and COX (by MAFP and indomethacin) caused potentiation, while cytochrome P450 and lipoxygenase inhibitors had no effect on 2‐AG relaxation. In coronary endothelial cells, 2‐AG (with and without MAFP) induced HC067047‐sensitive increases in intracellular Ca²⁺ concentration. 2‐AG also increased TRPV4 channel opening in inside‐out patches. However, in aortae, GSK induced a relaxation sensitive to HC067047 and ruthenium red, whereas 2‐AG induced contractions.

Conclusions and Implications

These data suggest that 2‐AG can directly activate endothelial TRPV4, which partly contributes to the relaxant response to 2‐AG. However, the functional role of TRPV4 is highly dependent on the vascular region.

Abbreviations

4α‐PDD

4α‐phorbol‐12,13‐didecanoate

GSK

GSK1016790A

JTE907

N‐(1,3‐benzodioxol‐5‐ylmethyl)‐1,2‐dihydro‐7‐methoxy‐2‐oxo‐8‐(pentyloxy)‐3‐quinolinecarboxamide

SKF525A

α‐phenyl‐α‐propylbenzeneacetic acid 2‐(diethylamino)ethyl ester N,N‐diethylaminoethyl 2,2‐diphenylvalerate