实验性超负荷血糖大鼠血脑屏障的超微结构变化

目的探讨糖尿病对血脑屏障(BBB)超微形态学及其内皮细胞ICAM-1表达的影响。方法采用44只SD大鼠尾静脉注射链脲霉素的方法,建立超负荷血糖模型。以透射电镜动态观察3d至8个月不同时程超负荷血糖条件下BBB超微结构的改变。结果超负荷血糖3d至2个月时,出现部分血脑屏障之内皮细胞、胶质细胞足突肿胀;3个月时出现明显的血脑屏障改变,整个毛细血管呈哑铃样、串珠样、动脉瘤样变形;6～8个月时改变继续加重,可见毛细血管管腔内微血栓,病变的血脑屏障周围,出现内皮细胞和神经细胞凋亡。实验中未见胶质细胞凋亡和神经细胞坏死。对照组无血脑屏障改变。结论实验提示超负荷血糖可造成血脑屏障发生明显的超微形态学异常,尤以长期作用更加明显。这可能是慢性糖尿病患者发生非梗塞性缺血缺氧性脑病的重要病理学基础。     

TCP-FA4: A derivative of tranylcypromine showing improved blood-brain permeability

A variety of approaches have been taken to improve the brain penetration of pharmaceutical agents. The amphipathic character of a compound can improve its interaction with the lipid bilayer within cell membranes, and as a result improve permeability. Fatty acid chains or lipoamino acids of various lengths were attached to tranylcypromine (TCP), in an attempt to improve the blood-brain barrier (BBB) permeability by increasing the lipophilicity as well as the amphiphatic character of the drug. TCP-FA4, one of the derivatives containing a four carbon alkyl acid chain, showed the greatest improvement in permeability. This molecule was slightly neuroprotective in a β-amyloid-induced neurodegeneration assay and may also be capable of upregulating brain derived neurotrophic factor (BDNF), as indicated by cell culture assays using human umbilical vein endothelial cells. Since decreased levels of BDNF are observed in many CNS disorders, and direct injection of BDNF is not a viable option due to its poor permeability across the BBB, small molecules capable of regulating BDNF that also cross the BBB may be an interesting treatment option.     

阿片类药物透过血脑屏障的三维构效研究

目的:建立药物透过血脑屏障的三维构效模型,为药物分子设计提供理论依据。方法与结果:利用比较分子力场分析方法建立了阿片类药物透过血脑屏障的三维定量构效模型,该模型有较高的预测能力,交叉验证系数r²_cv=0.718,相关系数r²=0.978,F₃,7=67.902,标准偏差SE=0.209。结论:根据CoMFA模型系数等势图,解释了该类药物透过血脑屏障的构效关系。     

Blood-brain barrier permeability and tPA-mediated neurotoxicity

Tissue type plasminogen activator (tPA) can induce neuronal apoptosis, disrupt the blood-brain barrier (BBB), and promote dilation of the cerebral vasculature. The timing, sequence and contributions of these and other deleterious effects of tPA and their contribution to post-ischemic brain damage after stroke, have not been fully elucidated. To dissociate the effects of tPA on BBB permeability, cerebral vasodilation and protease-dependent pathways, we developed several tPA mutants and PAI-1 derived peptides constructed by computerized homology modeling of tPA. Our data show that intravenous administration of human tPA to rats increases BBB permeability through a non-catalytic process that is associated with reversible neurotoxicity, brain damage, mortality and contributes significantly to its brief therapeutic window. Furthermore, our data show that inhibiting the effect of tPA on BBB function without affecting its catalytic activity, improves outcome and significantly extends its therapeutic window in mechanical as well as in thromboembolic models of stroke.     

Efficient LRP1-Mediated Uptake and Low Cytotoxicity of Peptide L57 In Vitro Shows Its Promise as CNS Drug Delivery Vector

Although an abundance of drug candidates exists which are aimed at the remediation of central nervous system (CNS) disorders, the utility of some are severely limited by their inability to cross the blood brain barrier. Potential drug delivery systems such as the Angiopep family of peptides have shown modest potential; however, there is a need for novel drug delivery candidates that incorporate peptidomimetics to enhance the efficiency of transcytosis, specificity, and biocompatibility. Here, we report on the first in vitro cellular uptake and cytotoxicity study of a peptidomimetic, cationic peptide, L57. It binds to cluster 4 of the low-density lipoprotein receptor-related protein 1 (LRP1) receptor which is expressed in numerous cell types, such as brain endothelial cells. We used early-passage-number brain microvascular endothelial cells and astrocytes harvested from rat pup brains that highly express LRP1, to study the uptake of L57 versus Angiopep-7 (A7). Uptake of L57 and A7 showed a concentration-dependent increase, with L57 being taken up to a greater degree than A7 at the same concentration. Additionally, peptide uptake in LRP1-deficient PEA 10 cells had greatly reduced uptake. Furthermore, L57 demonstrated excellent cell viability versus A7, showing promise as a potential drug delivery vector for CNS therapeutics.     

Effect of neurotransmitters on the system that transports Tyr-MIF-1 and the enkephalins across the blood-brain barrier: a dominant role for serotonin

Neurotransmitters and neuropeptides interact in several ways. We studied a new type of interaction: the effect of neurotransmitters on the saturable system that transports Tyr-MIF-1 and the enkephalins out of the central nervous system (CNS). The neurotransmitters were introduced into the lateral ventricle of the brain with radioiodinated peptide, using an established method previously shown to accurately quantify the amount of peptide being transported from the CNS to the blood. Serotonin inhibited transport, histamine stimulated transport, and dopamine, acetylcholine, epinephrine, GABA, kainic acid, cAMP and cGMP were without effect. Cyproheptadine, a serotonin antagonist, stimulated transport. Of several psychotropic agents tested, only tranylcypromine had a statistically significant effect and stimulated transport. Of the serotonin receptor specific agents tested, those with 5HT1 activity most consistently affected transport. We conclude that serotonin, and perhaps histamine, are important modulators of the system that transports Tyr-MIF-1 and the enkephalins out of the CNS.     

Challenges for blood–brain barrier (BBB) screening

Whilst blood–brain barrier permeability is an important determinant in achieving efficacious central nervous system drug concentrations, it should not be viewed or measured in isolation. Recent studies have highlighted the need for an integrated approach where optimal central nervous system penetration is achieved through the correct balance of permeability, a low potential for active efflux, and the appropriate physicochemical properties that allow for drug partitioning and distribution into brain tissue. Integrating data from permeability studies performed incorporating an assessment of active efflux by P-glycoprotein in combination with drug-free fraction measurements in blood and brain has furthered the understanding of the impact of the blood–brain barrier on central nervous system uptake and the underlying physicochemical properties that contribute to central nervous system drug disposition. This approach moves away from screening and ranking compounds in assays designed to measure or predict central nervous system penetration in the somewhat arbitrary units of brain–blood (or plasma) ratios.     

Investigation on the effect of nanoparticle size on the blood–brain tumour barrier permeability by in situ perfusion via internal carotid artery in mice

The blood–brain barrier (BBB) is a limiting factor in nanoparticle drug delivery to the brain, and various attempts have been made to overcome it for efficient drug delivery. Nowadays, it was considered as further issue for brain–drug delivery that the nanoparticle delivered to brain through the BBB reach cancer cells in tumour tissue. In this study, we investigated the effect of nanoparticle size on blood–brain tumour barrier (BBTB) permeation of fluorescence-labelled gold nanoparticles (AuNPs) in a mouse model of orthotopic glioblastoma multiforme (GBM), established by intracranial implantation of luciferase-expressing human glioblastoma U87MG cells. AuNPs sized 10, 50, and 100?nm were perfused into the GBM mice via internal carotid artery (ICA) for 5?min. Immediately after perfusion, the brains were fixed and prepared for LSCM observation. The AuNPs distribution in the normal and tumorous brain tissues was analysed qualitatively and quantitatively. Higher distribution of AuNPs was observed in the tumorous tissue than in the normal tissue. Furthermore, the smallest nanoparticle, 10?nm AuNPs, was widely distributed in the brain tumour tissue, whereas the 50 and 100?nm AuNPs were located near the blood vessels. Therefore, nanoparticle size affected the permeation of nanoparticles from the blood into brain tumour tissue.     

Carbidopa attenuation of L-DOPA emesis in dogs: evidence for a cerebral site of action outside the blood-brain barrier

L-Dopa and dopamine administered intracerebroventricularly (i.c.vent.) to dogs were approximately 50 times more potent as emetic agents than when administered i.v. The fact that dopamine, which penetrates the CNS only poorly, was more potent than L-dopa by both routes of administration, suggested a central extra-blood-brain barrier site of action for L-dopa-induced emesis. Systemically administered D,L-carbidopa, an inhibitor of aromatic amino acid decarboxylase, did not affect the emetic action of systemically administered dopamine but attenuated the emetic response to L-dopa administered both i.v. or i.c.vent. Since D,L-carbidopa does not penetrate whole brain to a significant extent it was concluded that carbidopa attenuates L-dopa-induced emesis by inhibiting the decarboxylation of L-dopa to dopamine at a CNS site lying outside the blood-brain barrier.     

“Tau Protein Targeting Via Radioiodinated Azure A For Brain Theranostics: Radiolabeling,Molecular Docking,in vitro And in vivo Biological Evaluation”

Azure-A is one of the phenothiazines (PTZs) derivatives which for decades have been used as antipsychotic drugs due to good lipophilic characteristics which enable them to pass through the blood brain barrier (BBB), besides the important property of enabeling investigation of the pathological forms of aggregated tau protein found in the neurons of the central nervous system. Radioiodination of Azure-A was carried out via an electrophilic substitution reaction using chloramine-T as oxidizing agent. The influence of various reaction parameters and conditions on radioiodination efficiency was investigated, and a high radiochemical yield of 92.07 ± 0.9 % was obtained. An in vitro cytotoxicity study of iodinated Azure-A on three cell lines (HCT-116, human colon carcinoma cell line; Hep-G2, liver carcinoma cell line and HFB-4, normal human melanocytes) was carried out, and the data revealed that ioiodinated Azure A has no to very low toxic effect. The in vivo biodistribution study of ¹³¹I-Azure A showed a high brain uptake of 6.15 ± 0.09 % injected dose/g tissue organ at 30 minutes post-injection, and its retention in brain remained high up to 2 hours, whereas the clearance from the body appeared to proceed via the renal system. The experimental data were confirmed by the molecular docking studies to predict the effect of radioiodination on the binding affinity of the parent molecule (Azure A) to tau paired helical filaments (PHFs). Both ligands showed better binding to S2 and S3 pockets of (PHFs). Consequently, radioiodinated Azure A seems to be a good candidate as an imaging agent for taupathies such as Alzheimer's disease, chronic traumatic encephalopathy, and corticobasal degeneration. Furthermore, it could be a very potent theranostics agent for brain tumors.     

Effect of a Chemical Delivery System for Dexamethasone (Dex-CDS) on Peritumoral Edema in an Experimental Brain Tumor Model

Pharmaceutical Research -     

Solid lipid nanoparticles containing tamoxifen characterization and in vitro antitumoral activity

Solid lipid nanoparticles (SLNs) containing tamoxifen, a nonsteroidal antiestrogen used in breast cancer therapy, were prepared by microemulsion and precipitation techniques. Tamoxifen loaded SLNs seem to have dimensional properties useful for parenteral administration, and in vitro plasmatic drug release studies demonstrated that these systems are able to give a prolonged release of the drug in the intact form. Preliminary study of antiproliferative activity in vitro, carried out on MCF-7 cell line (human breast cancer cells), demonstrated that SLNs, containing tamoxifen showed an antitumoral activity comparable to free drug. The results of characterization studies and of in vitro antiproliferative activity strongly support the potential application of tamoxifen-loaded SLNs as a carrier system at prolonged release useful for intravenous administration in breast cancer therapy.     

双配体修饰聚合物泡囊的制备及细胞摄取

目的构建一种主动靶向的新型纳米药物载体——聚合物泡囊(polymer vesicles,PVs),并考察其细胞摄取。方法以马来酰亚胺-聚乙二醇-聚乳酸-羟基乙酸共聚物(MAL-PEG-PLGA)为载体材料,通过自组装制备PVs,用转铁蛋白(Tf)与Tet-1对PVs进行修饰,构建纳米药物载体(Tf/Tet-1-PVs)。以香豆素-6作为荧光探针包载于药物载体,考察脑微血管内皮细胞(BCEC)及神经细胞(Neuro-2a)对载体系统的摄取。结果 PVs粒径约80nm,形态圆整,电镜观察具有明显膜层结构。BCEC细胞和Neuro-2a细胞对Tf/Tet-1-PVs的摄取均显著优于空白对照组和单配体修饰对照组。结论 PVs经双配体Tf及Tet-1修饰后可促进脑微血管内皮细胞和神经细胞的摄取。     

Effect of Chronic Hypertension on the Blood–Brain Barrier Permeability of Libenzapril

Very little information is available on the permeability of theblood–brain barrier (BBB) to small polar drugs inchronic hypertension. The blood and cerebrospinal fluid (CSF)pharmacokinetics of liben-zapril (LZP), a potentangiotensin converting enzyme inhibitor, were investigated inhypertensive (SH) and normotensive (SD) rats.Following intravenous bolus administration of this hydrophilic drug, theterminal rate constant for elimination (),steady-state volume of distribution ( ), and systemic clearance (CL) were similar in these two animalgroups. Other pharmacokinetic parameters (C_p°,, k _l2, and k ₂₁)were significantly (P < 0.05) greater in thehypertensive group, except for the volume of the central compartment(V_c) and ratio of V_c to , which were smaller in SH rats. The ratio ofarea under the concentration–time curve (AUC) in CSF toblood was about twofold higher in SH rats compared to normotensive rats,showing increased BBB permeability in hypertensive rats. An acute brainuptake study was also performed in SH, SD, and WK rats by intracarotidadministration of ¹⁴C-LZP along with³H₂O as a reference marker. Both LZP and watertransport was found to be significantly higher (about two-to five-fold) in six of the seven different brain regions inSH rats as compared to the normotensive (SD and WK) controls.Because of this simultaneous increase in concentrations of both the drugand the reference marker, BUI values were not affected. Regional brainconcentrations in SH rats were also linearly correlated with the meanarterial pressure (MAP) values, providing further evidence ofthe systemic pressure related increase in BBB permeability.