Transepithelial transport of putrescine across monolayers of the human intestinal epithelial cell line, Caco-2

AIM To study the transepithelial transport characteristics of the polyamine putrescine in human intestinal Caco-2 cell monolayers to elucidate the mechanisms of the putrescine intestinal absorption.METHODS The transepithelial transport and the cellular accumulation of putrescine was measured using Caco 2 cell monolayers grown on permeable filters.RESULTS Transepithelial transport of putrescine in physiological concentrations (>0.5 mM)from the apical to basolateral side was linear. Intracellular accumulation of putrescine was higher in confluent than in fully differentiated Caco-2 cells, but still negligible (less than 0.5%) of the overall transport across the monolayers in apical-to-basolateral direction. EGF enhanced putrescine accumulation in Caco-2 cells by four-fold, as well as putrescine conversion to spermidine and spermine by enhancing the activity of Sadenosylmethionine decarboxylase. However,EGF did not have any significant influence on putrescine flux across the Caco-2 cell monolayers. Excretion of putrescine from Caco-2cells into the basolateral medium did not exceed 50 picomoles, while putrescine passive flux from the apical to the basolateral chamber,contributed hundreds of micromoles polyamines to the basolateral chamber.CONCLUSION Transepithelial transport of putrescine across Caco-2 cell monolayers occurs in passive diffusion, and is not influenced when epithelial cells are stimulated to proliferate by a potent mitogen such as EGF.

Human small intestine is capable of restoring barrier function after short ischemic periods

AIM To assess intestinal barrier function during human intestinal ischemia and reperfusion(IR).METHODS In a human experimental model,6 cm of jejunum was selectively exposed to 30 min of ischemia(I) followed by 30 and 120 min of reperfusion(R). A sham procedure was also performed. Blood and tissue was sampled at all-time points. Functional barrier function was assessed using dual-sugar absorption tests with lactulose(L) and rhamnose(R). Plasma concentrations of citrulline,an amino acid described as marker for enterocyte function were measured as marker of metabolic enterocytes restoration. Damage to the epithelial lining was assessed by immunohistochemistry for tight junctions( TJs),by plasma marker for enterocytes damage(I-FABP) and analyzed by electron microscopy(EM) using lanthanum nitrate as an electrondense marker.RESULTS Plasma L/R ratio's were significantly increased after 30 min of ischemia(30 I) followed by 30 min of reperfusion(30 R) compared to control(0.75 ± 0.10 vs 0.20 ± 0.09,P < 0.05). At 120 min of reperfusion(120 R),ratio's normalized(0.17 ± 0.06) and were not significantly different from control. Plasma levels of I-FABP correlated with plasma L/R ratios measured at the same time points(correlation: 0.467,P < 0.01). TJs staining shows distortion of staining at 30 I. An intact lining of TJs was again observed at 30 I120 R. Electron microscopy analysis revealed disrupted TJs after 30 I with paracellular leakage of lanthanum nitrate,which restored after 30 I120 R. Furthermore,citrulline concentrations closely paralleled the histological perturbations during intestinal IR.CONCLUSION This study directly correlates histological data with intestinal permeability tests,revealing that the human gut has the ability of to withstand short episodes of ischemia,with morphological and functional recovery of the intestinal barrier within 120 min of reperfusion.

基于人工智能的药物人体肠道吸收性质预测

人体肠道吸收性(human intestinal absorption,HIA)是衡量药物口服生物利用度的重要标志之一。利用人工智能方法在药物发现早期对药物的HIA进行预测评估,能够加速药物发现过程并且降低成本。本研究分别使用分子模拟软件MOE(molecular operating environment)的2D、3D描述符和ECFP4(extended connectivity fingerprints)对分子进行表征,针对2061条HIA数据建立支持向量机(SVM)、随机森林(RF)、深度神经网络(DNN)等8种模型。结果表明,基于2D、3D描述符和ECFP4指纹的组合描述符构建的SVM模型在各项评价指标上进行综合评价后是最优的,最优模型的受试者工作特征曲线下面积(AUC)、马修斯系数和Kappa系数分别为0.94,0.75及0.74。综上,本研究建立一个鲁棒性高、泛化能力强的预测HIA性质的机器学习模型,该模型可以用于为药物药代动力学性质研究提供指导及早期的分子筛选。

从药物的三维分子结构预测人体小肠吸收(英文)

在口服药物的发展过程中,人体小肠吸收的预测是候选药物设计、优化和选择的一个主要目标.VolSurf/GRID计算方法作为一个新的工具被用来预测被测化合物的人体小肠吸收,以及测定人体小肠吸收所必需的重要的分子特征.被测化合物包括112个结构不同的类似药物的化合物.使用偏最小二乘判别分析方法在实验数据和人体小肠吸收的理论分子特征之间建立相关性.建立的两个模型之间具有较高的一致性.小肠吸收实验数据与分子特征之间好的相关性(r2=0.82,q2=0.67)表明,从化合物的三维分子结构能够预测人体小肠吸收.有利于人体小肠吸收的药物分子特征包括,分子量中心与亲水区重心的不平衡性,较大的疏水区域以及分子内较少的氢键给体.

延胡索甲素和乙素的人源肠内菌丛生物转化及在人源Caco-2细胞单层模型的吸收转运

目的:研究延胡索甲素(d-corydaline,CDL)和延胡索乙素(tetrahydropalmatine,THP)的人源肠内菌丛生物转化,在人源结肠腺癌细胞系Caco-2细胞单层模型的吸收转运。方法:应用人源肠内菌丛在厌氧、37℃条件下分别与CDL和THP共温孵培养;利用人源结肠腺癌细胞系Caco-2细胞单层模型测试CDL和THP从绒毛面(AP端)到基底面(BL端)、BL端到AP端双向的转运过程。计算转运参数和表观渗透系数(Papp),并与吸收良好的阳性对照药普萘洛尔和吸收不良的阿替洛尔进行比较。应用高效液相色谱法对CDL和THP进行分析。结果:CDL和THP在人源肠内菌丛温孵体系没有发生生物转化。它们与普萘洛尔在Caco-2细胞单层模型双向转运的Papp皆在1×10-5cm.s-1数量级;在0～180 min,转运效率与转运时间呈正相关。结论:CDL和THP在人源肠内菌丛温孵体系中稳定;在Caco-2细胞单层模型的转运机制可能为被动扩散。

红参水提物中人参三萜在人源肠Caco-2细胞单层模型上的吸收转运研究

目的研究红参水提物中人参三萜在人小肠上皮细胞的吸收特性。方法利用人源结肠腺癌细胞系Caco-2细胞单层模型测试红参水提物中人参三萜从顶端(apical,AP)侧到底端(basolateral,BL)侧、BL侧到AP侧2个方向的转运过程。应用色谱-质谱技术对人参三萜进行定量分析,计算转运参数和表观渗透系数(Papp)。结果在Caco-2细胞单层模型中,发现大多数人参三萜皂苷吸收程度中等或不良,但是,唯有2个苷元20(S)-原人参三醇(protopanaxatriol,PPT)和20(R)-PPT吸收程度良好(Papp值接近1×10-5 cm/s);此外,还发现人参三萜皂苷20(S)-G-Rh1、20(R)-G-Rh1、G-Rk3、G-Rh4、PG-RT5这5个单糖苷的Papp值皆在1×10-6 cm/s数量级,提示它们吸收程度中等。所考察的42个人参三萜其中包括8对R、S差向异构体,结果显示R/S异构体的吸收情况并没有显著性差异。结论随着人参三萜皂苷苷元上糖取代基数目的增多,其Papp值越来越小,提示多糖基的人参三萜皂苷在体内很难被吸收,可能是前药。

短肠综合征的康复治疗(英文)

目的 探讨短肠康复治疗对改善短肠病人营养状况、促进肠功能代偿的疗效。方法 短肠康复治疗包括营养支持、应用谷氨酰胺和生长激素 ,以及膳食纤维。共 2 7例短肠患者接受了 2 9次康复治疗 ,患者平均年龄 38 5± 19 3岁。剩余小肠长度范围 15 - 80cm ,平均 4 6 8± 2 3 4cm ,14例有回盲瓣。从肠切除至接受康复治疗的平均时间为 86± 10 5天。结果 治疗后病人的营养状况明显改善 ,肠道吸收功能有所增强。随访超过 2年者 8例 ,4例 (5 0 % )完全脱离肠外营养 ,随访时间超过 1年者 13例 ,有 10例 (76 9% )完全脱离肠外营养。结论 短肠康复治疗能够有效地改善短肠病人的营养状况、并能促进肠功能代偿 ,治疗效果与残留小肠长度、治疗开始的时间和病人年龄有关 ,及早进行康复治疗能够促进肠功能代偿 ,减少病人对肠外营养的依赖。