Study on the intestinal permeability of lamivudine using Caco-2 cells monolayer and Single-pass intestinal perfusion

BackgroundThe aim of this work is to investigate the intestinal permeability of lamivudine and explore its absorption mechanism.MethodCaco-2 cells monolayer and single-pass intestinal perfusion (SPIP) were selected for the investigation of lamivudine under different conditions, such as different concentration, absorption time, bidirectional transportation, and transportation with efflux transporters inhibitor. The concentration of lamivudine both in Caco-2 cells monolayer samples and SPIP samples was detected by HPLC-UV. Then the permeability parameters were calculated.ResultsThe established HPLC-UV method reach the requirements for detection. There is no statistically difference between absorption parameters of lamivudine both in Caco-2 cells monolayer and SPIP (P > 0.05) under different dose groups. After transportation with efflux transporters inhibitor, the efflux rate of lamivudine in three dose groups was significantly decreased from 2.67, 2.59 and 2.59 to 1.78, 1.61, and 1.81 respectively. Lamivudine exhibits an absorption mechanism of passive diffusion.ConclusionThe absorption of lamivudine may be related to efflux transporters. In addition, lamivudine is a moderate-permeability drug in Biopharmaceutics Classification System.     

A semi-integrated method for the determination of enzyme kinetic parameters and graphical representation of the Michaelis-Menten equation

A semi-integrated method for the determination of the enzyme kinetics parameters (Km and V) and graphical representation of the Michaelis-Menten equation is proposed as a variation of determination of initial reaction rate (v) as a function of initial substrate concentration ([S]0). The method is based on the determination of the time required to exhaust half of the initial substrate concentration as a function of the initial substrate concentration. The advantages and limitations of this method are discussed.     

Comparison of three concentration methods for the recovery of canine intestinal parasites from stool samples

The aim of present study was to compare the efficiency of a commercial assay and two conventional methods for fecal concentration in detecting canine gastrointestinal parasites. Fecal samples from 254 dogs were processed by centrifugation-sedimentation (CS), centrifugation-flotation (CF) and a commercial assay for fecal concentration (TF-test®). The following parasites were detected: Ancylostoma (37.8%), Giardia (16.9%), Toxocara canis (8.7%), Trichuris vulpis (7.1%), Isospora (3.5%), and Sarcocystis (2.7%). The calculated analytical sensitivity indicated that CF was more accurate (P < 0.01) in detecting Ancylostoma, T. canis, T. vulpis and Giardia infections. However, CF showed significantly higher sensitivity only for Ancylostoma, compared to the other two methods. The kappa index value of diagnostic agreement between TF-test and CF was high for T. canis (83%) and moderate for Giardia (72%) and Ancylostoma (63%). The advantages and limitations of each method were assessed for individual diagnosis and epidemiological investigation.     

A model for the analysis of growth data from designed experiments

A model for growth data from designed experiments is presented which extends the stochastic differential equation of Sandland and McGilchrist (1979, Biometrics 35, 255-272). Residual maximum likelihood (REML) is used to estimate the parameters of the model. The model is easily extended to incomplete data and is shown to overcome some of the practical difficulties encountered with the profile model. The procedure is applied to data from experiments on pigs and sheep.     

Software sensors for the monitoring of perfusion cultures: Evaluation of the hybridoma density and the medium composition from glucose concentration measurements

New software sensors based on the Extended Kalman Filter technique have been developed for the monitoring of animal cell perfusion cultures. They use a kinetic model describing the growth, death and metabolism of hybridoma cells as a function of the medium composition. The model was initially validated on a batch culture and found to correctly predict the continuous perfusion culture kinetics, except for the production of ammonia and lactate. Using the measurement of a single component in the culture medium, in this case glucose, the Extended Kalman Filter provides an excellent evaluation of the time variation of the concentrations of living and dead cells, of glutamine and antibodies, during the whole perfusion culture for a retained cell density rising from 1 to 11×10⁶ cells.ml^–1 inside the reactor.     

Media replenishment: A tool for the analysis of high-cell density perfusion systems

A simple method to analyze media consumption and by-product build-up in high-cell-density perfusion cultures has been developed. This technique makes use of media replenishment strategy which can be used to decouple the two phenomena. Also, the replenished media method can be used to identify limitation by complex and undefined media components such as bovine serum.     

Genetic algorithms applied to multi-class prediction for the analysis of gene expression data

MOTIVATION: An important challenge in the use of large-scale gene expression data for biological classification occurs when the expression dataset being analyzed involves multiple classes. Key issues that need to be addressed under such circumstances are the efficient selection of good predictive gene groups from datasets that are inherently 'noisy', and the development of new methodologies that can enhance the successful classification of these complex datasets. METHODS: We have applied genetic algorithms (GAs) to the problem of multi-class prediction. A GA-based gene selection scheme is described that automatically determines the members of a predictive gene group, as well as the optimal group size, that maximizes classification success using a maximum likelihood (MLHD) classification method. RESULTS: The GA/MLHD-based approach achieves higher classification accuracies than other published predictive methods on the same multi-class test dataset. It also permits substantial feature reduction in classifier genesets without compromising predictive accuracy. We propose that GA-based algorithms may represent a powerful new tool in the analysis and exploration of complex multi-class gene expression data. AVAILABILITY: Supplementary information, data sets and source codes are available at http://www.omniarray.com/bioinformatics/GA.     

Validation of the Boussinesq equation for use in traction field determination

Cell traction force plays an important role in many biological processes. Several traction force microscopy methods have been developed to determine cell traction forces based on the Boussinesq solution. This approach, however, is rooted in a half-space assumption. The purpose of this study was to determine the error induced in the half-space assumption using a finite element method (FEM). It demonstrates that displacement error between the FEM and the Boussinesq equation can be used to measure the accuracy of the Boussinesq equation, although singularity exists in the loading point. For one concentrated force, significant difference between the FEM and the Boussinesq equation occurs in the whole field; this difference decreases with an increase in the plate thickness. However, in the case of the balanced forces, the offset of the balanced forces decreases the errors in the middle area. Overall, this study demonstrates that increasing the thickness of the polyacrylamide gel is important for reducing the error of the Boussinesq equation when determining the displacement field of the gel under loads.     

Segmented linear regression models applied to the analysis of data from a cross-sectional growth experiment

Several different methods of analysis are applied to data consisting of weight measurements, taken at specified post-treatment times, of harvested thyroids from rats given one of four treatments. Previous studies of this type of data indicated that the growth is initially rapid, and that a second phase of less rapid growth is followed by a final phase in which little additional growth occurs. The data are further characterized by increasing variance through time. The primary purpose of the analysis is to study the effect of the treatments at the end of the study period. One-way analysis of variance tests among groups are performed on each day, but the results are not particularly helpful. However, results from two-way analyses of variance (over subsets of days and groups) are consistent with the three phase model and accordingly indicate significant group differences during each. Finally, maximum likelihood methods are used to fit a three part segmented linear regression model.     

Gas chromatographic method for detection of urinary sucralose: application to the assessment of intestinal permeability

We developed a capillary column gas chromatography (CCGC) method for the measurement of urinary sucralose (S) and three other sugar probes including, sucrose, lactulose (L) and mannitol (M) for use in in vivo studies of intestinal permeability. We compared the capillary method with a packed column gas chromatography (PCGC) method. We also investigated a possible role for sucralose as a probe for the measurement of whole gut permeability. Sample preparation was rapid and simple. The above four sugars were detected precisely, without interference. We measured intestinal permeability using 5- and 24-h urine collections in 14 healthy volunteers. The metabolism of sugars was evaluated by incubating the intestinal bacteria with an iso-osmolar mixture of mannitol, lactulose and sucralose at 37 degrees C for 19 h. Sugar concentrations and the pH of the mixture were monitored. The use of the CCGC method improved the detection of sucralose as compared to PCGC. The average coefficient of variation decreased from 15% to 4%. It also increased the sensitivity of detection by 200-2000-fold. The GC assay was linear between sucralose concentrations of 0.2 and 40 g/l (r=1.000). Intestinal bacteria metabolized lactulose and acidified the media but did not metabolize sucralose or mannitol. The new method for the measurement of urinary sucralose permits the simultaneous quantitation of sucrose, mannitol and lactulose, and is rapid, simple, sensitive, accurate and reproducible. Because neither S nor M is metabolized by intestinal bacteria, and because only a tiny fraction of either sugar is absorbed, this pair of sugar probes appears to be available for absorption throughout the GI tract. Thus, the 24-h urinary concentrations of S and M, or the urinary S/M ratio following an oral dose of a sugar mixture, might be good markers for whole gut permeability.     

A new proposal for a principal component-based test for high-dimensional data applied to the analysis of PhyloChip data

A modification of the principal component test is presented. It uses a weighted combination of the sums of squares for different principal components and is thus more powerful in high-dimensional settings with small sample sizes. Under usual normality assumptions, a rotation test is proposed which enables an exact conditional parametric test. The procedure is demonstrated with microarray data for the bacterial composition in the rhizosphere of different potato cultivars. In simulation studies, the power of the proposed statistic is compared with the competing multivariate parametric tests.     

Mathematical modeling of elution curves for a protein mixture in ion exchange chromatography applied to high protein concentration

Protein elution curves in ion exchange chromatography (IEC) were simulated with a rate model. Three pure proteins and their mixture were used (α‐lactalbumin, BSA, and conalbumin) under different operational conditions. The anionic matrix Q‐Sepharose FF was used packed in a 1 mL column. A high protein concentration (37.5 mg/mL of total protein injected into the column) was used in order to extend the utility of the model. Mass transfer parameters were calculated using empiric correlations, where the axial dispersion was negligible (Pe > 300) and the mass transfer was controlled by the intraparticle diffusion (Bi > 10). The model assumes a modulator–eluite relationship were the equilibrium constant of the Langmuir isotherm was a function of salt concentration. Adsorption kinetic parameters were estimated from experimental data. The parameters for pure proteins were determined, and elution curves for changes in flow rate, ionic strength gradient, concentration, and sample size were predicted by the model. Then the kinetic parameters of the mixture were determined under the same operational conditions and some of the parameters had to be modified to take into account effects such as protein–protein interactions, competition, and displacement. Experimental elution curves obtained for changes in operational conditions such as flow rate and ionic strength gradient were simulated by the rate model for the protein mixture with a relative error in retention time of visible peaks <5%. IEC operational conditions and the peak fraction collection can be selected using a cost function of the production process which considers yield, purity, concentration, and process time that are obtained from simulations. Operational conditions that gave the minimum cost were selected. Simulations allows to diminish experimental time and cost. Biotechnol. Bioeng. 2009; 104: 572–581 © 2009 Wiley Periodicals, Inc.     

微生态制剂治疗肝硬化肠功能紊乱患者的临床观察

目的探讨微生态制剂治疗肝硬化肠功能紊乱患者的疗效。方法选择80例肝硬化肠功能紊乱患者,随机分成治疗组和对照组。治疗组:常规保肝治疗加金双歧(4粒/次,2次/d);对照组:常规保肝治疗。疗程均为4周。结果治疗组和对照组相比,腹胀、腹泻、腹部不适症状明显改善,血氨水平降低,血浆内毒素水平下降,2组相比差异有显著性(P<0.05)。结论微生态制剂对于改善肝硬化患者临床症状有肯定的价值,并可降低血氨及血浆内毒素水平,有利于肝功能的改善。     

Regression analysis of multivariate interval-censored failure time data with application to tumorigenicity experiments

This paper discusses multivariate interval-censored failure time data that occur when there exist several correlated survival times of interest and only interval-censored data are available for each survival time. Such data occur in many fields. One is tumorigenicity experiments, which usually concern different types of tumors, tumors occurring in different locations of animals, or together. For regression analysis of such data, we develop a marginal inference approach using the additive hazards model and apply it to a set of bivariate interval-censored data arising from a tumorigenicity experiment. Simulation studies are conducted for the evaluation of the presented approach and suggest that the approach performs well for practical situations.     

微生态制剂对慢性肝衰竭患者肠道菌群的影响

目的探讨微生态调节剂双歧杆菌三联活菌对慢性肝衰竭患者肠道菌群的影响。方法根据治疗方法不同将46例慢性肝衰竭患者随机分为治疗组和对照组各23例,两组均给予慢性肝衰竭常规治疗,治疗组在上述基础上给予双歧杆菌三联活菌胶囊,210mg／粒,每次2粒,2次／d,疗程2周。观察治疗前后两组患者的肠道菌群菌落计数变化及血清总胆红素（TBiL）、凝血酶原时间（PT）变化。结果两组患者在实验前肠道菌群、TBiL、PT比较差异均无统计学意义（P〉0．05）。治疗2周后,对照组患者治疗前后肠道菌群无明显变化（P〉0．05）,TBiL、PT有所下降但不显著,治疗组患者的肠球菌、双歧杆菌、乳杆菌以及酵母样真菌的菌落数均较治疗前明显增加,TBiL、PT较前明显好转（P〈0．05）。结论口服双歧杆菌三联活菌制剂可以促进慢性肝衰竭患者肠道正常菌群的恢复及改善肝功能。     

Optimization of medium with perfusion microbioreactors for high density CHO cell cultures at very low renewal rate aided by design of experiments

A novel approach of design of experiment (DoE) is developed for the optimization of key substrates of the culture medium, amino acids, and sugars, by utilizing perfusion microbioreactors with 2 mL working volume, operated in high cell density continuous mode, to explore the design space. A mixture DoE based on a simplex-centroid is proposed to test multiple medium blends in parallel perfusion runs, where the amino acids concentrations are selected based on the culture behavior in presence of different amino acid mixtures, and using targeted specific consumption rates. An optimized medium is identified with models predicting the culture parameters and product quality attributes (G0 and G1 level N-glycans) as a function of the medium composition. It is then validated in runs performed in perfusion microbioreactor in comparison with stirred-tank bioreactors equipped with alternating tangential flow filtration (ATF) or with tangential flow filtration (TFF) for cell separation, showing overall a similar process performance and N-glycosylation profile of the produced antibody. These results demonstrate that the present development strategy generates a perfusion medium with optimized performance for stable Chinese hamster ovary (CHO) cell cultures operated with very high cell densities of 60 × 10⁶ and 120 × 10⁶ cells/mL and a low cell-specific perfusion rate of 17 pL/cell/day, which is among the lowest reported and is in line with the framework recently published by the industry.     

Comparative analysis of iterative reconstruction algorithms with resolution recovery and new solid state cameras dedicated to myocardial perfusion imaging

New technologies are available in myocardial perfusion imaging. They include new software that recovers image resolution and limits image noise, multifocal collimators and dedicated cardiac cameras in which solid-state detectors are used and all available detectors are constrained to imaging just the cardiac field of view.These innovations resulted in shortened study times or reduced administered activity to patients, while preserving image quality.Many single center and some multicenter studies have been published during the introduction of these innovations in the clinical practice. Most of these studies were lead in the framework of “agreement studies” between different methods of clinical measurement. They aimed to demonstrate that these new software/hardware solutions allow the acquisition of images with reduced acquisition time or administered activity with comparable results (as for image quality, image interpretation, perfusion defect quantification, left ventricular volumes and ejection fraction) to the standard-time or standard-dose SPECT acquired with a conventional gamma camera and reconstructed with the traditional FBP method, considered as the gold standard.The purpose of this review is to provide the reader with a comprehensive understanding of the pro and cons of the different approaches summarizing the achievements reached so far and the issues that need further investigations.     

A computer program for the statistical analysis of disease prevalence data from survival/sacrifice experiments

This paper presents a computer program for analyzing disease prevalence data from animal survival experiments in which there may also be some serial sacrifice. The method has been described in Biometrics 35 (1979) 221-234. The user is interrogated about the details of particular models he wishes to fit. Then a generalized EM algorithm is used to compute maximum likelihood estimates of various quantities of interest concerning the effects of treatment, time and presence of other diseases on the prevalences and lethalities of specific diseases of interest.