Blood–Brain Barrier Transporters: Opportunities for Therapeutic Development in Ischemic Stroke

Globally, stroke is a leading cause of death and long-term disability. Over the past decades, several efforts have attempted to discover new drugs or repurpose existing therapeutics to promote post-stroke neurological recovery. Preclinical stroke studies have reported successes in identifying novel neuroprotective agents; however, none of these compounds have advanced beyond a phase III clinical trial. One reason for these failures is the lack of consideration of blood–brain barrier (BBB) transport mechanisms that can enable these drugs to achieve efficacious concentrations in ischemic brain tissue. Despite the knowledge that drugs with neuroprotective properties (i.e., statins, memantine, metformin) are substrates for endogenous BBB transporters, preclinical stroke research has not extensively studied the role of transporters in central nervous system (CNS) drug delivery. Here, we review current knowledge on specific BBB uptake transporters (i.e., organic anion transporting polypeptides (OATPs in humans; Oatps in rodents); organic cation transporters (OCTs in humans; Octs in rodents) that can be targeted for improved neuroprotective drug delivery. Additionally, we provide state-of-the-art perspectives on how transporter pharmacology can be integrated into preclinical stroke research. Specifically, we discuss the utility of in vivo stroke models to transporter studies and considerations (i.e., species selection, co-morbid conditions) that will optimize the translational success of stroke pharmacotherapeutic experiments.     

Optimized Hemolysin Type 1 Secretion System in Escherichia coli by Directed Evolution of the Hly Enhancer Fragment and Including a Terminator Region

Type 1 secretion systems (T1SS) have a relatively simple architecture compared to other classes of secretion systems and therefore, are attractive to be optimized by protein engineering. Here, we report a KnowVolution campaign for the hemolysin (Hly) enhancer fragment, an untranslated region upstream of the hlyA gene, of the hemolysin T1SS of Escherichia coli to enhance its secretion efficiency. The best performing variant of the Hly enhancer fragment contained five nucleotide mutations at five positions (A30U, A36U, A54G, A81U, and A116U) resulted in a 2-fold increase in the secretion level of a model lipase fused to the secretion carrier HlyA1. Computational analysis suggested that altered affinity to the generated enhancer fragment towards the S1 ribosomal protein contributes to the enhanced secretion levels. Furthermore, we demonstrate that involving a native terminator region along with the generated Hly enhancer fragment increased the secretion levels of the Hly system up to 5-fold.     

Enhancing Post-Stroke Rehabilitation and Preventing Exo-Focal Dopaminergic Degeneration in Rats—A Role for Substance P

Dopaminergic signaling is a prerequisite for motor learning. Delayed degeneration of dopaminergic neurons after stroke is linked to motor learning deficits impairing motor rehabilitation. This study investigates safety and efficacy of substance P (SP) treatment on post-stroke rehabilitation, as this neuropeptide combines neuroprotective and plasticity-promoting properties. Male Sprague Dawley rats received a photothrombotic stroke within the primary motor cortex (M1) after which a previously acquired skilled reaching task was rehabilitated. Rats were treated with intraperitoneal saline (control group, n = 7) or SP-injections (250 µg/kg) 30 min before (SP-pre; n = 7) or 16 h (SP-post; n = 6) after rehabilitation training. Dopaminergic neurodegeneration, microglial activation and substance P-immunoreactivity (IR) were analyzed immunohistochemically. Systemic SP significantly facilitated motor rehabilitation. This effect was more pronounced in SP-pre compared to SP-post animals. SP prevented dopaminergic cell loss after stroke, particularly in the SP-pre condition. Despite its proinflammatory propensity, SP administration did not increase stroke volumes, post-stroke deficits or activation of microglia in the midbrain. Finally, SP administration prevented ipsilesional hypertrophy of striatal SPergic innervation, particularly in the SP-post condition. Mechanistically, SP-pre likely involved plasticity-promoting effects in the early phase of rehabilitation, whereas preservation of dopaminergic signaling may have ameliorated rehabilitative success in both SP groups during later stages of training. Our results demonstrate the facilitating effect of SP treatment on motor rehabilitation after stroke, especially if administered prior to training. SP furthermore prevented delayed dopaminergic degeneration and preserved physiological endogenous SPergic innervation.     

柱塞泵蜗轮箱异常磨损的原因分析及改进

分析柱塞泵运行中蜗轮箱产生异常磨损和零部件失效的原因,进行了技术改进和旧件的修复夏利用后,泵运行正常,减少了维修工作量,取得了较好的经济效益。     

简论变速压力机行程功之间的相互关系

介绍了机械压力机连续行程功与单次行程功的概念，简述了变速压力机的优点及其影响行程功的各种因素，分析了变速压力机连续行程功与连续行程次数之间、单次行程功与连续行程功之间的内在关系，并推导出它们之间关系的计算公式和计算结果。     

短行程铝挤压机中伺服电机的控制技术

介绍了前上料短行程铝挤压机中的供锭方式、供锭机械手的控制系统工艺流程、硬件构成及程序设计。开发设计的Sinamics S120伺服驱动控制系统,实现了供锭机械手的伸缩定位控制。该伺服控制系统响应快,跟随性能好,具有很高的可靠性,为短行程挤压机的研发提供了技术保障。该伺服系统已应用于实际生产,完全满足工艺要求,生产效率和经济效益均得到不同程度的提高,对后续同类产品的设计起到了借鉴作用。     

HCV核心抗原N-端片段生物素化表达质粒的构建及表达

目的构建HCV核心(C)抗原N-端1～130aa片段的生物素化表达质粒,并在大肠杆菌中进行表达。方法PCR扩增HCV C抗原N-端1～130aa片段的编码基因,拼接上生物素-蛋白连接酶底物肽序列(BSP),构建重组原核表达质粒pGEX4T-1-CN130BSP,转化大肠杆菌Rosetta,IPTG诱导表达。表达产物经SDS-PAGE后,电转至硝酸纤维素膜上,以抗HCVC抗原的单克隆抗体和辣根过氧化物酶标记的亲和素对表达产物进行分析。结果PCR扩增出约466bp的目的基因片段;质粒pGEX4T-1-CN130BSP经PCR及双酶切鉴定,与预期结果一致,测序鉴定基因无突变;SDS-PAGE显示表达产物相对分子质量约为42000,22℃诱导16h可溶性表达产物含量较高;表达的重组蛋白可被抗HCV C抗原的单克隆抗体所识别,并能特异性结合辣根过氧化物酶标记的亲和素。结论已成功构建了HCV C抗原N-端1～130aa片段生物素化表达质粒,并表达出带生物素标签的GST融合蛋白,为进一步研制HCV双抗原夹心检测试剂奠定了基础。     

化工罐区爆炸碎片多米诺效应影响概率计算模型

爆炸碎片是化工区多米诺事故的重要诱因。从爆炸碎片飞行过程的规律性及碎片产生抛射的不确定性出发,分析化工罐区爆炸碎片破坏性及事故连锁机理。将储罐爆炸瞬间视为一个多方过程,通过求解多方过程状态方程,推导了爆炸碎片初速度计算公式;通过求解碎片飞行的加速度方程并结合边界条件,得到了爆炸碎片飞行距离及飞行速度计算公式;以爆炸碎片飞行规律性分析得到的相关公式为目标函数,分析影响爆炸碎片飞行距离及飞行速度不确定性的随机变量,通过随机抽样并对目标函数不同计算结果的出现频率进行累积,得到爆炸碎片在不同飞行距离上的累积击中概率。通过分析重大危险源单元（major hazard installation unit,MHIU）之间的一次影响及多次影响,构建化工罐区多个重大危险源发生爆炸碎片多米诺影响效应的概率计算模型,揭示爆炸碎片产生、抛射、飞行及打击目标的内在机理,相关分析结果能够为化工园区多米诺事故风险分析奠定基础。