Two-Year Results of a Five-Year Personalized Integrative Obesity Coaching Program (IBO) Based upon a Systems Health Perspective and an Evolutionary Longitudinal Study Approach.

This study presents the outcomes of a 5-year personalized integrative coaching program for adults with obesity (body mass index BMI ≥ 30 kg/m2), based upon a systems health perspective, during the first 2 years. This longitudinal study, which had an evolutionary design, included all adults who enrolled in the program. Health-related quality of life (HRQoL) was measured with the Short Form-36 (SF-36), and physical outcomes included weight, waist circumference, aerobic capacity, lipid profile, and HbA1c. Subsequently, participants completed questionnaires (e.g., the Symptom Checlist-90 (SCL-90) and the Checklist Individual Strength (CIS)). Seventy-nine adults with a mean BMI of 39.5 kg/m2 (SD 5.3) were included. Forty-four participants completed 2 years in the program. Compared to baseline, there were significant improvements in the SF-36 subscales 'physical functioning' (MD 9.9 points, 95% CI: 2.1-17.5, p = 0.013) and 'general health perceptions' (MD 9.3 points, 95% CI 2.9-15.7, p = 0.006). Furthermore, significant improvements in physical outcomes and psychosocial questionnaires (e.g., weight loss (MD 3.5 kg, 95% CI: 1.2-5.7, p = 0.003), waist circumference (MD 5.1 cm, 95% CI: 2.4-7.8, p < 0.001), and CIS fatigue (MD 6.8, 95% CI: 3.1-10.5, p = 0.001) were observed. This study highlights the importance of a systems health perspective supporting the development of a personalized integrative coaching program for adults with obesity in a 'real-world' setting.

Individual Workplace Well-Being Captured into a Literature- and Stakeholders-Based Causal Loop Diagram.

This study demonstrates an innovative approach to capture the complexity of individual workplace well-being, improving our understanding of multicausal relationships and feedback loops involved. The literature shows that a high number of interacting factors are related to individual workplace well-being. However, many studies focus on subsets of factors, and causal loops are seldomly studied. The aim of the current study was, therefore, to capture individual workplace well-being in a comprehensive conceptual causal loop diagram (CLD). We followed an iterative, qualitative, and transdisciplinary systems-thinking approach including literature search, group model building sessions, retrospective in-depth interviews with employees, and group sessions with human resource professionals, managers, job coaches, and management consultants. The results were discussed with HR and well-being officers of twelve organizations for their critical reflection on the recognizability and potential of the developed CLD. The final result, a conceptual individual workplace well-being CLD, provides a comprehensive overview of multiple, measurable key factors relating to individual workplace well-being and of the way these factors may causally interact over time, either improving or deteriorating workplace well-being. In future studies, the CLD can be translated to a quantitative system dynamics model for simulating workplace well-being scenarios. Ultimately, these simulations could be used to design effective workplace well-being interventions.

Developing a Personalized Integrative Obesity-Coaching Program: A Systems Health Perspective.

Current obesity management strategies are failing to achieve sustainable and favorable long-term results. We propose a more personalized, dynamic, and systemic perspective on the interactions of key determinants and coaching advice on obesity. The aim of this study was to use a systems view on overweight, complexity science, and a transdisciplinary process to develop a five-year personalized integrative obesity-coaching and research program. Managers, medical specialists, clinical psychologists, dieticians, physical- and psychomotor therapists, and lifestyle coaches aligned their perspectives and objectives with experts in systems thinking and systems biology. A systems health model of obesity was used to identify the causal relations of variables with the most influence on obesity. The model helped to align and design a personalized integrative obesity-coaching program and to identify the key variables to monitor the progress and to adjust the personalized program, depending on the goals and needs of the participant. It was decided to use subtyping of participants by a systems biologist, based on traditional Chinese medicine symptoms, as a novel method to personalize the intervention. The collaborative transdisciplinary approach based upon a systems view on obesity was successful in developing a personalized and adaptive five-year obesity-coaching and research program.

Link-based influence maximization in networks of health promotion professionals.

The influence maximization problem (IMP) as classically formulated is based on the strong assumption that "chosen" nodes always adopt the new product. In this paper we propose a new influence maximization problem, referred to as the "Link-based Influence Maximization Problem" (LIM), which differs from IMP in that the decision variable of the spreader has changed from choosing an optimal seed to selecting an optimal node to influence in order to maximize the spread. Based on our proof that LIM is NP-hard with a monotonic increasing and submodular target function, we propose a greedy algorithm, GLIM, for optimizing LIM and use numerical simulation to explore the performance in terms of spread and computation time in different network types. The results indicate that the performance of LIM varies across network types. We illustrate LIM by applying it in the context of a Dutch national health promotion program for prevention of youth obesity within a network of Dutch schools. GLIM is seen to outperform the other methods in all network types at the cost of a higher computation time. These results suggests that GLIM may be utilized to increase the effectiveness of health promotion programs.

An Innovative Approach for Decision-Making on Designing Lifestyle Programs to Reduce Type 2 Diabetes on Dutch Population Level Using Dynamic Simulations.

The number of individuals suffering from type 2 diabetes is dramatically increasing worldwide, resulting in an increasing burden on society and rising healthcare costs. With increasing evidence supporting lifestyle intervention programs to reduce type 2 diabetes, and the use of scenario simulations for policy support, there is an opportunity to improve population interventions based upon cost-benefit analysis of especially complex lifestyle intervention programs through dynamic simulations. In this article, we used the System Dynamics (SD) modeling methodology aiming to develop a simulation model for policy makers and health professionals to gain a clear understanding of the patient journey of type 2 diabetes mellitus and to assess the impact of lifestyle intervention programs on total cost for society associated with prevention and lifestyle treatment of pre-diabetes and type 2 diabetes in The Netherlands. System dynamics describes underlying structure in the form of causal relationships, stocks, flows, and delays to explore behavior and simulate scenarios, in order to prescribe intervention programs. The methodology has the opportunity to estimate and simulate the consequences of unforeseen interactions in order to prescribe intervention programs based on scenarios tested through "what-if" experiments. First, the extensive knowledge of diabetes, current available data on the type 2 diabetes population, lifestyle intervention programs, and associated cost in The Netherlands were captured in one simulation model. Next, the relationships between leverage points on the growth of type 2 diabetes population were based upon available data. Subsequently, the cost and benefits of future lifestyle intervention programs on reducing diabetes were simulated, identifying the need for an integrated adaptive design of lifestyle programs while collecting the appropriate data over time. The strengths and limitations of scenario simulations of complex lifestyle intervention programs to improve the (cost)effectiveness of these programs to reduce diabetes in a more sustainable way compared to usual care are discussed.

A Proof-of-Concept System Dynamics Simulation Model of the Development of Burnout and Recovery Using Retrospective Case Data.

The phenomenon of burnout is a complex issue, which despite major efforts from researchers and organizations remains hard to prevent. The current literature highlights an increasing global prevalence of employees that are dealing with burnout. What has been largely missing is a more systemic, dynamic, and personal perspective on the interactions of the key determinants of burnout. Burnout can be seen as the outcome of a complex system involving feedback loops between individual mental models, individual behavior, and external social influences. Understanding the feedback loops involved may enable employees and organizations to intervene in burnout trajectories early and effectively. System dynamics (SD) modeling is a methodology that can describe the structure and behavior of a complex system. The current paper describes the development of an SD model of burnout. First, an expert- and literature-informed causal loop diagram (CLD) of burnout is developed. Then, a novel approach is developed to collect personal retrospective scenario data. Finally, the CLD and data are translated into a quantitative SD model. The potential of the SD model is illustrated by simulating the behavior of three realistic personas during the onset of and recovery from burnout. The process of development of an SD model of burnout is presented and the strengths and limitations of the approach are discussed.

A higher throughput and physiologically relevant two-compartmental human ex vivo intestinal tissue system for studying gastrointestinal processes.

A majority of the preclinical intestinal screening models do not properly reflect the complex physiology of the human intestinal tract, resulting in low translational value to the clinical situation. The often used cell lines such as Caco-2 or HT-29 are not well suited to investigate the different processes that predict oral bioavailability in real life, or processes involved in general gut health aspects. Therefore, highly realistic models resembling the human in vivo situation are needed; application of ex vivo intestinal tissue is an interesting and feasible alternative. After previously using porcine intestinal tissue as a predictive model for human intestinal absorption, we now have successfully applied human intestinal tissue into a newly developed InTESTine™ two-compartmental disposable device suitable for standard 6- or 24-well plate format. With this set-up we demonstrated (regional differences in) drug absorption, by using a subset of compounds with known varying Fa (fraction absorbed) values. A rank-order relationship of R2 = 0.85 could be established between the Fa and Papp of these commercially available drugs. Additionally, comparison between the InTESTine system and the established Ussing chamber technology showed a correlation of R2 = 0.94 (10 drugs) with respect to Papp values, indicating good comparison of both models. Besides absorption, intestinal wall metabolism of testosterone (CYP3A4) was determined by showing a linear formation (R2 = 0.99; up to 165 min) of the main metabolites androstenedione and 6Beta-hydroxytestosterone, indicating no loss of metabolic capacity of the intestinal tissue within the system. Enteroendocrine responses were assessed of the satiety hormones GLP-1 and PYY after stimulation with rebaudioside A and casein, resulting in significantly increased secretion to the luminal side as well as to the basolateral side. Incubation with the probiotic strain LGG showed to enhance the viability of the tissue by showing to decrease the LDH secretion compared to blank intestinal tissue. In conclusion, we show that human ex vivo intestinal tissue mounted in the higher throughput InTESTine 6- 24-transwell plate system is easy to handle and a suitable system to study diverse functional GI processes.

The effect of chitosan on the bioaccessibility and intestinal permeability of acyclovir.

Chitosan is object of pharmaceutical research as a candidate permeability enhancer. However, chitosan was recently shown to reduce the oral bioavailability of acyclovir in humans. The effect of chitosan on two processes determining the oral bioavailability of acyclovir, bioaccessibility and intestinal absorption, was now investigated. Acyclovir's bioaccessibility was studied using the dynamic TNO gastro-Intestinal Model (TIM-1). Four epithelial models were used for permeability experiments: a Caco-2 cell model in absence and presence of mucus and both rat and porcine excised intestinal segments. Study concentrations of acyclovir (0.8 g/l) and chitosan (1.6 g/l and 4 g/l) were in line with those used in the aforementioned human study. No effect of chitosan was measured on the bioaccessibility of acyclovir in the TIM-1 system. The results obtained with the Caco-2 models were not in line with the in vivo data. The tissue segment models (rat and porcine intestine) showed a negative trend of acyclovir's permeation in presence of chitosan. The Ussing type chamber showed to be the most biopredictive, as it did point to an overall statistically significantly reduced absorption of acyclovir. This model thus seems most appropriate for pharmaceutical development purposes, in particular when interactions between excipients and drugs are to become addressed.

Proteomics of human liver membrane transporters: a focus on fetuses and newborn infants.

BACKGROUND: Hepatic membrane transporters are involved in the transport of many endogenous and exogenous compounds, including drugs. We aimed to study the relation of age with absolute transporter protein expression in a cohort of 62 mainly fetus and newborn samples. METHODS: Protein expressions of BCRP, BSEP, GLUT1, MCT1, MDR1, MRP1, MRP2, MRP3, NTCP, OCT1, OATP1B1, OATP1B3, OATP2B1 and ATP1A1 were quantified with LC-MS/MS in isolated crude membrane fractions of snap-frozen post-mortem fetal and pediatric, and surgical adult liver samples. mRNA expression was quantified using RNA sequencing, and genetic variants with TaqMan assays. We explored relationships between protein expression and age (gestational age [GA], postnatal age [PNA], and postmenstrual age); between protein and mRNA expression; and between protein expression and genotype. RESULTS: We analyzed 36 fetal (median GA 23.4 weeks [range 15.3-41.3]), 12 premature newborn (GA 30.2 weeks [24.9-36.7], PNA 1.0 weeks [0.14-11.4]), 10 term newborn (GA 40.0 weeks [39.7-41.3], PNA 3.9 weeks [0.3-18.1]), 4 pediatric (PNA 4.1 years [1.1-7.4]) and 8 adult liver samples. A relationship with age was found for BCRP, BSEP, GLUT1, MDR1, MRP1, MRP2, MRP3, NTCP, OATP1B1 and OCT1, with the strongest relationship for postmenstrual age. For most transporters mRNA and protein expression were not correlated. No genotype-protein expression relationship was detected. DISCUSSION AND CONCLUSION: Various developmental patterns of protein expression of hepatic transporters emerged in fetuses and newborns up to four months of age. Postmenstrual age was the most robust factor predicting transporter expression in this cohort. Our data fill an important gap in current pediatric transporter ontogeny knowledge.

Grip on health: A complex systems approach to transform health care.

This article addresses the urgent need for a transition in health care to deal with the increasing prevalence of chronic diseases and associated rapid rise of health care costs. Chronic diseases evolve and are predominantly related to lifestyle and environment. A shift is needed from a reductionist repair mode of thinking, toward a more integrated biopsychosocial way of thinking about health. The aim of this article is to discuss the opportunities that complexity science offer for transforming health care toward optimal treatment and prevention of chronic lifestyle diseases. Health and health care is discussed from a complexity science perspective. The benefits of concepts developed in the field of complexity science for stimulating transitions in health care are explored. Complexity science supports the elucidation of the essence of health processes. It provides a unique perspective on health with a focus on the relationships within networks of dynamically interacting factors and the emergence of health out of the organization of those relationships. Novel types of complexity science-based intervention strategies are being developed. The first application in practice is the integrated obesity treatment program currently piloted in the Netherlands, focusing on health awareness and healing relationships. Complexity science offers various theories and methods to capture the path toward unhealthy and healthy states, facilitating the development of a dynamic integrated biopsychosocial perspective on health. This perspective offers unique insights into health processes for patients and citizens. In addition, dynamic models driven by personal data provide simulations of health processes and the ability to detect transitions between health states. Such models are essential for aligning and reconnecting the many institutions and disciplines involved in the health care sector and evolve toward an integrated health care ecosystem.

Variability in Mass Spectrometry-based Quantification of Clinically Relevant Drug Transporters and Drug Metabolizing Enzymes.

Many different methods are used for mass-spectrometry-based protein quantification in pharmacokinetics and systems pharmacology. It has not been established to what extent the results from these various methods are comparable. Here, we compared six different mass spectrometry-based proteomics methods by measuring the expression of clinically relevant drug transporters and metabolizing enzymes in human liver. Mean protein concentrations were in general quantified to similar levels by methods using whole tissue lysates. Methods using subcellular membrane fractionation gave incomplete enrichment of the proteins. When the enriched proteins were adjusted to levels in whole tissue lysates, they were on average 4-fold lower than those quantified directly in whole tissue lysates. The differences in protein levels were propagated into differences in predictions of hepatic clearance. In conclusion, caution is needed when comparing and applying quantitative proteomics data obtained with different methods, especially since membrane fractionation is common practice for protein quantification used in drug clearance predictions.

Regional Expression Levels of Drug Transporters and Metabolizing Enzymes along the Pig and Human Intestinal Tract and Comparison with Caco-2 Cells.

Intestinal transporter proteins and metabolizing enzymes play a crucial role in the oral absorption of a wide variety of drugs. The aim of the current study was to characterize better available intestinal in vitro models by comparing expression levels of these proteins and enzymes between porcine intestine, human intestine, and Caco-2 cells. We therefore determined the absolute protein expression of 19 drug transporters and the mRNA expression of 12 metabolic enzymes along the pig intestinal tract (duodenum, jejunum, ileum; N = 4), in human intestine (jejunum; N = 9), and Caco-2 cells. Expression of the included transporters and enzymes was in general well comparable between porcine and human intestinal tissue, although breast cancer resistance protein, monocarboxylate transporter 5, multidrug resistance protein (MRP) 1, MRP1, MRP3 (∼2-fold), and organic anion-transporting polypeptide (OATP) 4A1 (∼6-fold) was higher expressed in pig compared with human jejunum. Alternatively, expression level of relevant transporter proteins (glucose transporter 1, OATP4A1, MRP2, MRP1, and OATP2B1) was significantly higher (3- to 130-fold) in Caco-2 cells compared with human jejunum. Moreover, all examined CYPs showed at least a fivefold lower gene expression in Caco-2 cells compared with human jejunum, with the smallest differences for CYP1A1 and CYP3A5 and the largest difference for CYP3A4 (871-fold higher expression in human jejunum compared with Caco-2 cells). In conclusion, a comprehensive overview is provided of the expression levels of clinically relevant transporter proteins and metabolic enzymes in porcine and human intestinal tissue and Caco-2 cells, which may assist in deciding upon the most suitable model to further improve our understanding of processes that determine intestinal absorption of compounds.

Nutrient-induced glucagon like peptide-1 release is modulated by serotonin.

Glucagon like peptide-1 (GLP-1) and serotonin are both involved in food intake regulation. GLP-1 release is stimulated upon nutrient interaction with G-protein coupled receptors by enteroendocrine cells (EEC), whereas serotonin is released from enterochromaffin cells (ECC). The central hypothesis for the current study was that nutrient-induced GLP-1 release from EECs is modulated by serotonin through a process involving serotonin receptor interaction. This was studied by assessing the effects of serotonin reuptake inhibition by fluoxetine on nutrient-induced GLP-1, PYY and CCK release from isolated pig intestinal segments. Next, serotonin-induced GLP-1 release was studied in enteroendocrine STC-1 cells, where effects of serotonin receptor inhibition were studied using specific and non-specific antagonists. Casein (1% w/v), safflower oil (3.35% w/v), sucrose (50mM) and rebaudioside A (12.5mM) stimulated GLP-1 release from intestinal segments, whereas casein only stimulated PYY and CCK release. Combining nutrients with fluoxetine further increased nutrient-induced GLP-1, PYY and CCK release. Serotonin release from intestinal tissue segments was stimulated by casein and safflower oil while sucrose and rebaudioside A had no effect. The combination with fluoxetine (0.155µM) further enhanced casein and safflower oil induced-serotonin release. Exposure of ileal tissue segments to serotonin (30µM) stimulated GLP-1 release whereas it did not induce PYY and CCK release. Serotonin (30 and 100µM) also stimulated GLP-1 release from STC-1 cells, which was inhibited by the non-specific 5HT receptor antagonist asenapine (1 and 10µM). These data suggest that nutrient-induced GLP-1 release is modulated by serotonin through a receptor mediated process.

Proteomic Analysis of the Developmental Trajectory of Human Hepatic Membrane Transporter Proteins in the First Three Months of Life.

Human hepatic membrane-embedded transporter proteins are involved in trafficking endogenous and exogenous substrates. Even though impact of transporters on pharmacokinetics is recognized, little is known on maturation of transporter protein expression levels, especially during early life. We aimed to study the protein expression of 10 transporters in liver tissue from fetuses, infants, and adults. Transporter protein expression levels [ATP-binding cassette transporter (ABC)B1, ABCG2, ABCC2, ABCC3, bile salt efflux pump, glucose transporter 1, monocarboxylate transporter 1, organic anion transporter polypeptide (OATP)1B1, OATP2B1, and organic cation/carnitine transporter 2) were quantified using ultraperformance liquid chromatography tandem mass spectrometry in snap-frozen postmortem fetal, infant, and adult liver samples. Protein expression was quantified in isolated crude membrane fractions. The possible association between postnatal and postmenstrual age versus protein expression was studied. We studied 25 liver samples, as follows: 10 fetal [median gestational age 23.2 wk (range 16.4-37.9)], 12 infantile [gestational age at birth 35.1 wk (27.1-41.0), postnatal age 1 wk (0-11.4)], and 3 adult. The relationship of protein expression with age was explored by comparing age groups. Correlating age within the fetal/infant age group suggested four specific protein expression patterns, as follows: stable, low to high, high to low, and low-high-low. The impact of growth and development on human membrane transporter protein expression is transporter-dependent. The suggested age-related differences in transporter protein expression may aid our understanding of normal growth and development, and also may impact the disposition of substrate drugs in neonates and young infants.

Predicting carrier-mediated hepatic disposition of rosuvastatin in man by scaling from individual transfected cell-lines in vitro using absolute transporter protein quantification and PBPK modeling.

In contrast to primary hepatocytes, estimating carrier-mediated hepatic disposition by using a panel of single transfected cell-lines provides direct information on the contribution of the individual transporters to the net disposition. The most direct way to correct for differences in transporter abundance between cell-lines and tissue is by using absolute protein quantification. In the present study, the performance of this strategy to predict human hepatic uptake transport was investigated and compared with traditional scaling from primary human hepatocytes. Rosuvastatin was used as a model compound. The uptake activity was measured in HEK293 cell-lines stably overexpressing OATP1B1(∗)1a, OATP1B3 or OATP2B1, the major transporters involved in human hepatic uptake of rosuvastatin, or expressing OATP1B1(∗)15, associated with reduced hepatic uptake of rosuvastatin. The abundance of these transporter proteins in the outer membranes of HEK293-cells, in human primary hepatocytes and in human liver tissue was determined by LC-MS/MS. The measured activity, corrected for protein abundance and scaled to the whole liver, gave a very accurate prediction of the hepatic intrinsic clearance observed in vivo. Embedded in a PBPK model describing the hepatic disposition and enterohepatic circulation, the collective in vitro data resulted in a good explanation of the observed oral and intravenous pharmacokinetic profiles of rosuvastatin. The model allowed simulation of the effect of polymorphic variants of OATP1B1 on rosuvastatin pharmacokinetics. These results encourage a larger scale validation. This approach may facilitate prediction of drug-drug interactions, scaling of transporter processes across subpopulations (children, diseased patients), and may be extended to tissues for which primary cells may be more difficult to obtain.

Steviol glycoside rebaudioside A induces glucagon-like peptide-1 and peptide YY release in a porcine ex vivo intestinal model.

Glucagon-like peptide-1 (GLP-1) and peptide YY (PYY) are hormones important for satiation and are involved in the process called "ileal brake". The aim of this study was to investigate the GLP-1- and PYY-stimulating efficacy of rebaudioside A, casein, and sucrose. This was studied using tissue segments collected from various regions of the pig small intestine. GLP-1 release was strongest from the distal ileum. There, control release was 0.06 ± 0.01 (GLP-1) and 0.07 ± 0.01 (PYY) pmol/cm(2) of tissue. Rebaudioside A (2.5, 12.5, and 25 mM) stimulated GLP-1 release (0.14 ± 0.02, 0.16 ± 0.02, and 0.13 ± 0.02 pmol/cm(2) of tissue, p < 0.001) and PYY release (0.19 ± 0.02, 0.42 ± 0.06, and 0.27 ± 0.03 pmol/cm(2) of tissue, p < 0.001). Sucrose stimulated GLP-1 release (0.08 ± 0.01 pmol/cm(2) of tissue, p < 0.05) only at 10 mM. Casein (0.5%, 1%, and 2.5%, w/v) stimulated GLP-1 release (0.15 ± 0.03, 0.13 ± 0.02, and 0.14 ± 0.01 pmol/cm(2) of tissue, p < 0.001) and PYY release (0.13 ± 0.02, 0.20 ± 0.03, and 0.27 ± 0.03 pmol/cm(2) of tissue, p < 0.01). These findings may help in developing dietary approaches for weight management.

A new approach to predict human intestinal absorption using porcine intestinal tissue and biorelevant matrices.

A reliable prediction of the oral bioavailability in humans is crucial and of high interest for pharmaceutical and food industry. The predictive value of currently used in silico methods, in vitro cell lines, ex vivo intestinal tissue and/or in vivo animal studies for human intestinal absorption, however, is often insufficient, especially when food-drug interactions are evaluated. Ideally, for this purpose healthy human intestinal tissue is used, but due to its limited availability there is a need for alternatives. The aim of this study was to evaluate the applicability of healthy porcine intestinal tissue mounted in a newly developed InTESTine™ system to predict human intestinal absorption of compounds with different chemical characteristics, and within biorelevant matrices. To that end, first, a representative set of compounds was chosen of which the apparent permeability (Papp) data in both Caco-2 cells and human intestinal tissue mounted in the Ussing chamber system, and absolute human oral bioavailability were reported. Thereafter, Papp values of the subset were determined in both porcine jejunal tissue and our own Caco-2 cells. In addition, the feasibility of this new approach to study regional differences (duodenum, jejunum, and ileum) in permeability of compounds and to study the effects of luminal factors on permeability was also investigated. For the latter, a comparison was made between the compatibility of porcine intestinal tissue, Caco-2 cells, and Caco-2 cells co-cultured with the mucin producing HT29-MTX cells with biorelevant samples as collected from an in vitro dynamic gastrointestinal model (TIM). The results demonstrated that for the paracellularly transported compounds atenolol, cimetidine, mannitol and ranitidine porcine Papp values are within 3-fold difference of human Papp values, whereas the Caco-2 Papp values are beyond 3-fold difference. Overall, the porcine intestinal tissue Papp values are more comparable to human Papp values (9 out of 12 are within 3-fold difference), compared to Caco-2 Papp values (4 out of 12 are within 3-fold difference). In addition, for the selected hydrophilic compounds a significant increase in the permeability was observed from duodenum to ileum. Finally, this study indicated that porcine jejunal tissue segments can be used with undiluted luminal samples to predict human intestinal permeability and the effect of biorelevant matrices on this. In conclusion, viable porcine intestinal tissue mounted in the InTESTine™ system can be applied as a reliable tool for the assessment of intestinal permeability in the absence and presence of biorelevant samples. This would enable an accessible opportunity for a reliable prediction of human intestinal absorption, and the effect of luminal compounds such as digested foods, early in drug development.

Interaction of digitalis-like compounds with liver uptake transporters NTCP, OATP1B1, and OATP1B3.

Digitalis-like compounds (DLCs) such as digoxin, digitoxin, and ouabain, also known as cardiac glycosides, are among the oldest pharmacological treatments for heart failure. The compounds have a narrow therapeutic window, while at the same time, DLC pharmacokinetics is prone to drug-drug interactions at the transport level. Hepatic transporters organic anion transporting polypeptide (OATP) 1B1, OATP1B3, and Na(+)-dependent taurocholate co-transporting polypeptide (NTCP) influence the disposition of a variety of drugs by mediating their uptake from blood into hepatocytes. The interaction of digoxin, digitoxin, and ouabain with hepatic uptake transporters has been studied before. However, here, we systematically investigated a much wider range of structurally related DLCs for their capability to inhibit or to be transported by these transporters in order to better understand the relation between the activity and chemical structure of this compound type. We studied the uptake and inhibitory potency of a series of 14 structurally related DLCs in Chinese hamster ovary cells expressing NTCP (CHO-NTCP) and human embryonic kidney cells expressing OATP1B1 and OATP1B3 (HEK-OATP1B1 and HEK-OATP1B3). The inhibitory effect of the DLCs was measured against taurocholic acid (TCA) uptake in CHO-NTCP cells and against uptake of ß-estradiol 17-ß-d-glucuronide (E217ßG) in HEK-OATP1B1 and HEK-OATP1B3 cells. Proscillaridin A was the most effective inhibitor of NTCP-mediated TCA transport (IC50 = 22 µM), whereas digitoxin and digitoxigenin were the most potent inhibitors of OATP1B1 and OAPTP1B3, with IC50 values of 14.2 and 36 µM, respectively. Additionally, we found that the sugar moiety and hydroxyl groups of the DLCs play different roles in their interaction with NTCP, OATP1B1, and OATP1B3. The sugar moiety decreases the inhibition of NTCP and OATP1B3 transport activity, whereas it enhances the inhibitory potency against OATP1B1. Moreover, the hydroxyl group at position 12 reinforces the inhibition of NTCP but decreases the inhibition of OATP1B1 and OATP1B3. To investigate whether DLCs can be translocated, we quantified their uptake in transporter-expressing cells by LC-MS. We demonstrated that convallatoxin, ouabain, dihydroouabain, and ouabagenin are substrates of OATP1B3. No transport was observed for the other compounds in any of the studied transporters. In summary, this work provides a step toward an improved understanding of the interaction of DLCs with three major hepatic uptake transporters. Ultimately, this can be of use in the development of DLCs that are less prone to transporter-mediated drug-drug interactions.

Protein transport across the small intestine in food allergy.

In view of the imminent deficiency of protein sources for human consumption in the near future, new protein sources need to be identified. However, safety issues such as the risk of allergenicity are often a bottleneck, due to the absence of predictive, validated and accepted methods for risk assessment. The current strategy to assess the allergenic potential of proteins focuses mainly on homology, stability and cross-reactivity, although other factors such as intestinal transport might be of added value too. In this review, we present an overview of the knowledge of protein transport across the intestinal wall and the methods currently being used to measure this. A literature study reveals that protein transport in sensitised persons occurs para-cellularly with the involvement of mast cells, and trans-cellularly via enterocytes, while in non-sensitised persons micro-fold cells and enterocytes are considered most important. However, there is a lack of comparable systematic studies on transport of allergenic proteins. Knowledge of the multiple protein transport pathways and which model system can be useful to study these processes may be of added value in the risk assessment of food allergenicity.

Interaction of immunosuppressive drugs with human organic anion transporter (OAT) 1 and OAT3, and multidrug resistance-associated protein (MRP) 2 and MRP4.

Renal proximal tubule transporters can play a key role in excretion, pharmacokinetic interactions, and toxicity of immunosuppressant drugs. Basolateral organic anion transporters (OATs) and apical multidrug resistance-associated proteins (MRPs) contribute to the active tubular uptake and urinary efflux of these drugs, respectively. We studied the interaction of 12 immunosuppressants with OAT1- and OAT3-mediated [(3)H]-methotrexate (MTX) uptake in cells, and adenosine triphosphate-dependent [(3)H]-MTX transport in membrane vesicles isolated from human embryonic kidney 293 cells overexpressing human MRP2 and MRP4. Our results show that at a clinically relevant concentration of 10 µM, mycophenolic acid inhibited both OAT1- and OAT3-mediated [(3)H]-MTX uptake. Cytarabine, vinblastine, vincristine, hydrocortisone, and mitoxantrone inhibited only OAT1, whereas tacrolimus, azathioprine, dexamethasone, cyclosporine, and 6-mercaptopurine had no effect on both transporters. Cyclophosphamide stimulated OAT1, but did not affect OAT3. With regard to the apical efflux transporters, mycophenolic acid, cyclophosphamide, hydrocortisone, and tacrolimus inhibited MRP2 and MRP4, whereas mitoxantrone and dexamethasone stimulated [(3)H]-MTX transport by both transporters. Cyclosporine, vincristine, and vinblastine inhibited MRP2 only, whereas 6-mercaptopurine inhibited MRP4 transport activity only. Cytarabine and azathioprine had no effect on either transporter. In conclusion, we charted comprehensively the differences in inhibitory action of various immunosuppressive agents against the 4 key renal anion transporters, and we provide evidence that immunosuppressant drugs can modulate OAT1-, OAT3-, MRP2-, and MRP4-mediated transport of MTX to different extents. The data provide a better understanding of renal mechanisms underlying drug-drug interactions and nephrotoxicity concerning combination regimens with these compounds in the clinic.