不同基质氢化可的松软膏经糖尿病模型大鼠皮肤的透过性研究

目的:研究不同基质的氢化可的松软膏经糖尿病模型大鼠皮肤的透过性。方法:分别制备氢化可的松O/W型软膏、水溶性基质软膏和油脂性基质软膏。取Wistar大鼠随机分为正常对照组和模型组,模型组大鼠ip给予链脲菌素(40 mg/kg)以复制糖尿病模型。采用Franz扩散池透皮试验和高效液相色谱法,考察氢化可的松O/W型软膏、水溶性基质软膏和油脂性基质软膏分别经两组大鼠皮肤的透过速率。结果:与正常对照组比较,氢化可的松O/W型软膏和水溶性基质软膏经模型组大鼠皮肤的透过速率明显增加,差异具有统计学意义(P<0.05);氢化可的松油脂性基质软膏经模型大鼠皮肤的透过速率差异无统计学意义(P>0.05)。结论:氢化可的松O/W型软膏、水溶性基质软膏较易透过糖尿病模型大鼠皮肤;氢化可的松油脂性基质软膏不易透过糖尿病模型大鼠皮肤。     

Permeability dominates in vivo intestinal absorption of P-gp substrate with high solubility and high permeability

Three purposes are presented in this study: (1) to study the in vivo regional dependent intestinal absorption of a P-gp substrate with high solubility and high permeability, (2) to study the gene expression difference in the various regions of the intestine, and (3) to study the contributions of P-gp or any other transporters for the absorption of a P-gp substrate. The in vivo permeability of verapamil and propranolol were determined by single-pass in situ intestinal perfusion in rat. The gene expression profiles were measured using Affymetrix GeneChip. Correlation analysis between drug in vivo permeability and expression of 3500 genes was performed with nonparametric bootstrap and ANOVA analysis. The permeability of verapamil and propranolol did not demonstrate regional dependency even though significant differences in gene expression were observed in various regions of the intestine. Verapamil permeability significantly correlates with propranolol permeability in both jejunum and ileum, but did not correlate with the permeability of other hydrophilic compounds (valacyclovir, acyclovir, and phenylalanine). Four different regions (duodenum, jejunum, ileum, and colon) showed distinct gene expression patterns with more than 70-499 genes showing at least 5-fold expression differences. Interestingly, P-gp expression is gradually increased by 6-fold from the duodenum to colon. Despite the distinct gene expression patterns in the various regions of the intestine, verapamil permeability did not correlate with any gene expression from 3500 expressed genes in the intestine. A 2-6-fold P-gp expression difference did not seem to associate verapamil permeability in the various intestinal regions in vivo. These data suggest that P-gp plays a minimal role in the in vivo intestinal absorption process of verapamil with high water solubility and high membrane permeability. The intestinal absorption of verapamil in vivo is primarily dominated by its high permeability. However, it is important to note that the findings in this paper do not undermine the importance of P-gp in oral drug bioavailability, drug disposition from the liver, drug efflux from the blood-brain barrier, and drug-drug interaction.     

聚乙二醇400对P-糖蛋白底物罗丹明123经肠黏膜的透过性研究

目的:研究聚乙二醇400对P-糖蛋白(P-gp)底物罗丹明123(R123)经肠黏膜的透过作用。方法:使用体外扩散池评价R123经空肠、回肠和结肠黏膜的经-时吸收方向和分泌方向的透过量和透过系数(Papp),并测定不同浓度聚乙二醇400对R123和荧光素钠(CF)经肠黏膜透过性的影响。R123和CF在接受室中的浓度用荧光分光光度法测定。结果:R123经肠道黏膜的透过性存在部位差,即以空肠、回肠和结肠的次序透过性依次减少,并且R123经肠道分泌方向的透过性显著地高于其吸收方向的透过性;聚乙二醇400具有增加R123经吸收方向的透过性,但实验浓度的聚乙二醇400对CF的肠道转运没有影响。结论:聚乙二醇400可通过对P-gp功能的抑制而用于改善受P-gp介导药物的吸收,有望提高此类药物的口服生物利用度。     

双歧杆菌对应激模型大鼠肠道通透性的影响

目的:研究双歧杆菌对应激模型大鼠肠道通透性的影响及其与蒙脱石散联用的协同作用。方法:取SD大鼠50只随机均分为正常对照组、模型对照组、双歧杆菌干预组(2×108cfu)、蒙脱石散干预组(0.6 g.kg-1)及其联合干预组(双歧杆菌2×108cfu+蒙脱石散0.6 g.kg-1),每日灌胃给药1次,连续给药7 d,后4组每日给药后以避水压力模型(WAS)方法建立大鼠应激模型。第8天各组分别灌胃给予探针药物甘露醇80 mg、三氯蔗糖60 mg,检测各组大鼠给药后5、24 h尿液中甘露醇、三氯蔗糖的量及其比值,以评价肠道通透性。实验结束处死大鼠,检测各组大鼠血清中促肾上腺皮质激素释放因子(CRF)和促肾上腺皮质激素(ACTH)的含量。结果:与正常对照组比较,模型对照组大鼠24 h尿液中甘露醇的量及血清中CRF、ACTH含量均明显升高(P<0.05);与模型对照组比较,双歧杆菌干预组和联合干预组ACTH含量均明显降低(P<0.05),其余指标均有降低,但没有统计学意义。结论:双歧杆菌对应激模型大鼠的肠道通透性有影响,其与蒙脱石散联用对应激所致的肠道屏障功能紊乱无协同作用。     

细胞膜糖蛋白P170抑制剂的研究进展

多药耐药性的发生是肿瘤治疗失败的原因之一，而肿瘤细胞多药耐药基因1(MDR1)编码的细胞膜糖蛋白P170表达的增加是最常见的发生机制。近年来针对细胞膜糖蛋白P170抑制剂的研究有了很大进展，本文就目前细胞膜糖蛋白P170抑制剂的研究状况进行综述。     

雷公藤甲素对糖尿病模型大鼠的经皮渗透特性研究

目的:研究雷公藤甲素经糖尿病模型大鼠皮肤的渗透特性。方法:一次性ip给予链脲佐菌素(55 mg/kg)以复制大鼠糖尿病模型,剥离鼠皮备用,同时设正常对照(正常大鼠皮)组与模型组。采用改良型Franz扩散池评价雷公藤甲素经皮渗透效果并测定大鼠皮肤各层中雷公藤甲素滞留量,采用高效液相色谱串联质谱(LC-MS/MS)法检测雷公藤甲素含量。色谱柱为Agilent ZORBAX-C18,流动相为甲醇-水(60∶40),流速为0.3 ml/min,进样量为2μl,柱温为30℃;电喷雾电离(ESI)+,电离电压为+4.0 k V,检测离子对为m/z 361.1→91.1,离子源温度为110℃,干燥气温度为350℃,流量为7 L/min,监测模式为多重反应检测(MRM)模式,裂解电压为135 V,碰撞能量为55 e V。结果:模型组大鼠皮的稳态渗透速率为12.66μg/(h·cm2),单位面积累积透过量为(165.42±8.90)μg/cm2;与正常对照组(19.22±4.32)μg/cm2比较,上述两个指标均明显增加(P<0.05)。模型组大鼠真皮层中药物含量为(34.32±0.47)μg/cm2,明显高于正常对照组大鼠真皮层的(12.48±0.35)μg/cm2。结论:糖尿病模型大鼠皮肤对雷公藤甲素的渗透性(渗透速率、单位面积累积透过量、皮肤滞留量)明显增加。     

Insulin Permeability Across an in Vitro Dynamic Model of Endothelium

Purpose. Endothelium insulin permeability was investigated using in vitro, dynamic culture of endothelial cells. Methods. Endothelial cells were cultured in a hollow fiber apparatus and continuously exposed to a flow. Transendothelial electrical resistance and permeability to [¹⁴C]sucrose and [¹⁴C]inulin were used to monitor the integrity of the endothelial monolayer. Results. Under these experimental conditions, measurements of insulin permeability, investigated at increasing hormone concentrations, suggested that the predominant transendothelial insulin fluxes were attributable to bidirectional convective transport rather than to a saturable transport mechanism, in agreement with in vivo experiment results published earlier. Analytical determinations of insulin catabolism demonstrated a low percent of insulin degradation by the endothelium, leading to production of insulin metabolites qualitatively identical to those produced by human monocytes. Conclusions. The findings of this paper indicated that (a) insulin crosses the endothelial monolayer by paracellular leak and endothelial insulin receptors have a minor (if any) role in insulin transport; (b) degradation of the hormone by BAEC is minimal; (c) the in vitro, dynamic culture of endothelial cells presented here should represent a valuable transport model system to study permeability mechanisms of insulin and many other drugs.     

Modulation of P-gp expression by lapatinib

Chemotherapy drug resistance is a major obstacle in the treatment of cancer. It can result from an increase in levels of cellular drug efflux pumps, such as P-glycoprotein (P-gp). Lapatinib, a growth factor receptor tyrosine kinase inhibitor, is currently in clinical trials for treatment of breast cancer. We examined the impact of co-incubation of chemotherapy drugs in combination with lapatinib in P-gp over-expressing drug resistant cells. Unexpectedly, lapatinib treatment, at clinically relevant concentrations, increased levels of the P-gp drug transporter in a dose- and time-responsive manner. Conversely, exposure to the epidermal growth factor (EGF), an endogenous growth factor receptor ligand, resulted in a decrease in P-gp expression. Despite the lapatinib-induced alteration in P-gp expression, use of accumulation, efflux and toxicity assays demonstrated that the induced alteration in P-gp expression by lapatinib had little direct impact on drug resistance.     

Effect of Cyclodextrin Complexation on the Liposome Permeability of a Model Hydrophobic Weak Acid

Purpose  This study examines the effect of a chemically modified β-cyclodextrin on the liposome bilayer permeability of a liposomally entrapped model hydrophobic weak acid, DB-67 (7-t-butyldimethylsilyl-10-hydroxycamptothecin). Materials and Methods  Permeability studies were conducted in liposomes prepared by hydration–extrusion in the presence or absence of entrapped hydroxypropyl-β-cyclodextrin (HPβCD). A gradient HPLC method with evaporative light scattering detection was developed for analysis of HPβCD. DB-67 was analyzed by HPLC with fluorescence detection. Results  HPβCD entrapped in the aqueous compartment of liposomes was found to be membrane impermeable. Gel phase liposomes were stable in the presence of HPβCD. HPβCD complexation did not significantly alter the apparent permeability of DB67 lactone, due to its high membrane binding. However, lactone ring-opening and ionization significantly decreased the apparent permeability and improved the liposomal retention of DB-67, an effect that was amplified in the presence of 50 mM HPβCD. Conclusions  In liposomes, cyclodextrin complexation competes with liposomal membrane binding which may temper the potential benefit of complexation in prolonging hydrophobic drug retention. Cyclodextrin complexation combined with drug ionization may nevertheless significantly enhance the retention of ionizable hydrophobic drugs in liposomes as complexation may compete more favorably with membrane binding when the drug is ionized.     

Concentration-Dependent Effects of Polyethylene Glycol 400 on Gastrointestinal Transit and Drug Absorption

Purpose. The aim of the study was to investigate the effect of different concentrations of polyethylene glycol 400 (PEG 400) on liquid transit through, and ranitidine absorption from, the gastrointestinal tract. Methods. Six healthy male volunteers received, on four separate occasions, 150 mL water containing 150 mg ranitidine and either 0 (control), 1, 2.5, or 5 g PEG 400. The solutions were radiolabeled with technetium-99m to allow their gastrointestinal transit to be followed using a gamma camera. Urine samples were collected over a 24-h period to assess the amount of ranitidine excreted and hence absorbed. Results. No significant differences in gastric emptying were noted between the four solutions. In contrast, the presence of 1, 2.5, and 5 g PEG 400 reduced the mean small intestinal transit times of the solutions by 9, 20, and 23%, respectively, against the control. In terms of drug absorption, the mean cumulative amount of ranitidine excreted was reduced by 38% in the presence of both 2.5 and 5 g PEG 400, although it was significantly increased by 41% in the presence of 1 g PEG 400. Conclusions. The results show that low concentrations of PEG 400 enhance the absorption of ranitidine possibly via modulation of intestinal permeability, while high concentrations have a detrimental effect on ranitidine absorption presumably via a reduction in the small intestinal transit time.     

抑制P-糖蛋白的羟基喜树碱纳米混悬剂的制剂学评价

采用微沉淀-高压匀质法制备具有高过饱和溶出水平、高渗透性和物理稳定的口服羟基喜树碱纳米混悬剂(hydroxycamptothecin nanosuspensions,HCPT-Nano),并对其粒径及分布、zeta电位、粒子形态、物理存在状态、贮存稳定性及过饱和溶出水平等制剂学性质进行了评价。结果表明,HCPT-Nano的粒径小于300 nm且分布均匀,粒子多呈棒状或块状,药物以结晶形式存在,能长期维持较高的过饱和溶出水平。因此,结合P糖蛋白(P-glycoprotein,P-gp)的抑制剂环孢素A(cyclosporin,CsA)的加入,为提高HCPT的口服生物利用度提供了可能性。     

Concentration-Dependent Plasma Protein Binding of Flurbiprofen in the Rat: An in Vivo Microdialysis Study

Pharmaceutical Research - Purpose. The in vivo plasma protein binding and pharmacokinetics of flurbiprofen were studied in awake, unrestrained rats using intravenous microdialysis sampling....     

Nasal delivery of P-gp substrates to the brain through the nose-brain pathway

The objective of this study was to evaluate in rats the potential utility of the nasal route to enhance central nervous system (CNS) delivery of drugs recognized by P-glycoprotein (P-gp). Well-known P-gp substrates verapamil and talinolol were perfused nasally or infused intravenously, and when plasma concentrations following intravenous infusion and nasal perfusion showed similar profiles. The concentration of verapamil in the brain after nasal perfusion was twice that after intravenous infusion. Although talinolol in the brain and the cerebrospinal fluid after i.v. infusion were below the detection limit, it was detected after nasal perfusion. When rats were treated with cyclosporin A, brain concentrations of verapamil after both administration modes were increased significantly, while those of talinolol were not significantly changed. Since the permeability of talinolol is low, talinolol in the brain which was transported directly from the nasal cavity has little chance of transport by P-gp localized in the apical membrane of cerebral microvessel endothelial cells. The potential for drug delivery utilizing the nose-CNS route was confirmed for P-gp substrates. The advantage of nasal delivery over i.v. delivery of talinolol to the brain was more significant than that of verapamil, suggesting that nasal administration is more useful strategy for the brain delivery of low-permeability P-gp substrates than the use of P-gp inhibitors.     

P糖蛋白在大肠癌中的表达及意义

目的 探讨P糖蛋白（P-gp）在大肠癌组织中的表达及意义。方法 应用免疫组织化学方法 检测48例大肠癌组织中P-gP的表达，并分析与临床病理特征的关系。结果 P-gP表达与淋巴结转移有关，与性别、年龄、原发肿瘤部位、肿瘤大小、分化程度、局部浸润深度、远处转移、Duke’s分期无关。结论临床检测大肠癌P-gP表达对大肠癌化疗方案的选择具有重要的意义。     

P-糖蛋白与中枢抑制药物相互作用的体外研究

目的:基于"上调MDR(multiple drug resistance)基因、诱导P-糖蛋白(P-gp,P-glycoprotein)外向转运解除中枢抑制药物中毒"的机制研究,对临床中毒多发的中枢抑制药物建立P-gp底物谱,明确适合该解救治疗方案的药物(毒物)。方法:采用三磷酸腺苷酶(adenosine triphosphate,ATP)酶活性测试法确定待测中枢抑制药物是否为P-gp的底物。结果:舒必利、齐拉西酮、佐匹克隆、氯米帕明、西酞普兰及吗氯贝胺均为P-gp底物。舒必利、齐拉西酮与西酞普兰为非浓度依赖的ATP酶抑制剂,佐匹克隆、氯米帕明与吗氯贝胺浓度依赖的ATP酶双向调节子。结论:该研究结果将为新的中枢药物中毒解救方案的临床应用提供参考和依据。     

Chemogenomic Analysis Identifies Geldanamycins as Substrates and Inhibitors of ABCB1

Purpose A prerequisite for geldanamycin (GA, NSC122750) to targeting heat shock protein 90 and inhibiting tumor growth is sufficient intracellular drug accumulation. We hypothesized that membrane transporters on tumor cells determine at least in part the response to GA analogues. Materials and Methods To facilitate a systematic study of chemosensitivity across a group of GA analogues with similar chemical structures, we correlated mRNA expression profiles of most known transporters with growth inhibitory potencies of compounds in 60 tumor cell lines (NCI-60). We subsequently validated the gene-drug correlations using cytotoxicity and transport assays. Results Geldanamycin analogues displayed a range of negative correlations coefficients with ABCB1 (MDR1, or P-glycoprotein) expression. Suppressing ABCB1 in multidrug resistant cells (NCI/ADR-RES and K562/DOX) and ABCB1-transfected cells (BC19) increased sensitivity to GA analogues, as expected for substrates. Moreover, ABCB1-mediated efflux of daunorubicin in K562/DOX cells could be blocked markedly by GA analogues in a dose-dependent fashion. The IC₅₀ values (half-maximum inhibition of daunorubicin efflux) were 5.5, 7.3 and 12 μM for macbecin II (NSC330500), 17-AAG (NSC330507) and GA, respectively. Conclusions These observations demonstrate that GA analogues are substrates as well as inhibitors of ABCB1, suggesting that drug interactions between GA analogues and other agents that are ABCB1 substrates may occur via ABCB1 in normal or tumor cells. Electronic supplementary material The online version of this article (doi: ) contains supplementary material, which is available to authorized users.     

Antiinflammatory Drug-Induced Small Intestinal Permeability: The Rat Is a Suitable Model

Excretion of orally administrated ⁵¹Cr-EDTA as a marker of small intestinal permeability (a proposed prerequisite for human enteropathy) is increased by corticosteroids and non-steroidal anti-inflammatory drugs (NSAIDs). We have investigated the suitability of the rat as an animal model of small intestinal permeability using orally administered ⁵¹Cr-EDTA. We dosed Sprague-Dawley rats with NSAIDs and corticosterone followed by ⁵¹Cr-EDTA under conditions reported for humans and measured urinary excretion of the marker. In control rats, the urinary excretion of ⁵¹Cr-EDTA exhibited a skewed-to-the-left frequency distribution curve with a median of 2.13% of the dose. No sex-related differences were noticed in the baseline permeability. In male rats, single therapeutically equivalent doses of indomethacin, flurbiprofen, ibuprofen, naproxen, diclofenac, sulindac, nambumetone, and corticosterone, increased the intestinal permeability by different extents with indomethacin eliciting the maximum effect, and the last four drugs showing minimal potencies. Therapeutically relevant doses of aspirin did not have any significant effect. The increase in permeability was dependent upon the NSAIDs dose. Administration of glucose/citrate, misoprostol and sulfasalazine significantly reduced the effect of indomethacin. Misoprostol antagonized the effect of naproxen but H₂-antagonists and sucralfate did not. All the above observations made in the rat were similar to those previously reported for humans. Thus the rat is a suitable model for studies of small intestinal permeability.     

Analysis of Unstirred Water Layer in In Vitro Permeability Experiments

In vitro permeability experiments are used widely in drug discovery and other areas of pharmaceutical research. Much effort has been expended in developing novel epithelial models but generally much less attention has been paid to the hydrodynamic barrier in the actual experiments. The restricted liquid flow in the vicinity of solid surfaces leads to a zone where the diffusional movement of molecules exceeds the convection. This leads to formation of a concentration gradient between the bulk solution and the surface. The formed unstirred water layer (UWL) reduces the apparent permeability (P_app) of compounds that rapidly pass through the actual epithelial layer. This lowers the resolution of P_app versus fraction-absorbed assay, complicates the structure-permeability analysis and skews apparent kinetic parameters of transporter substrates. This review describes the techniques that can be used to determine the UWL thickness in permeability experiments and apparatuses described in the literature to control the in vitro hydrodynamics. © 2009 Wiley-Liss, Inc. and the American Pharmacists Association J Pharm Sci 98:4469–4479, 2009