A Human Stem Cell-Derived Brain-Liver Chip for Assessing Blood-Brain-Barrier Permeation of Pharmaceutical Drugs.

Significant advancements in the field of preclinical in vitro blood-brain barrier (BBB) models have been achieved in recent years, by developing monolayer-based culture systems towards complex multi-cellular assays. The coupling of those models with other relevant organoid systems to integrate the investigation of blood-brain barrier permeation in the larger picture of drug distribution and metabolization is still missing. Here, we report for the first time the combination of a human induced pluripotent stem cell (hiPSC)-derived blood-brain barrier model with a cortical brain and a liver spheroid model from the same donor in a closed microfluidic system (MPS). The two model compounds atenolol and propranolol were used to measure permeation at the blood-brain barrier and to assess metabolization. Both substances showed an in vivo-like permeation behavior and were metabolized in vitro. Therefore, the novel multi-organ system enabled not only the measurement of parent compound concentrations but also of metabolite distribution at the blood-brain barrier.

An Individual Patient's "Body" on Chips-How Organismoid Theory Can Translate Into Your Personal Precision Therapy Approach.

The first concepts for reproducing human systemic organismal biology in vitro were developed over 12 years ago. Such concepts, then called human- or body-on-a-chip, claimed that microphysiological systems would become the relevant technology platform emulating the physiology and morphology of human organisms at the smallest biologically acceptable scale in vitro and, therefore, would enable the selection of personalized therapies for any patient at unprecedented precision. Meanwhile, the first human organoids-stem cell-derived complex three-dimensional organ models that expand and self-organize in vitro-have proven that in vitro self-assembly of minute premature human organ-like structures is feasible, once the respective stimuli of ontogenesis are provided to human stem cells. Such premature organoids can precisely reflect a number of distinct physiological and pathophysiological features of their respective counterparts in the human body. We now develop the human-on-a-chip concepts of the past into an organismoid theory. We describe the current concept and principles to create a series of organismoids-minute, mindless and emotion-free physiological in vitro equivalents of an individual's mature human body-by an artificially short process of morphogenetic self-assembly mimicking an individual's ontogenesis from egg cell to sexually mature organism. Subsequently, we provide the concept and principles to maintain such an individual's set of organismoids at a self-sustained functional healthy homeostasis over very long time frames in vitro. Principles how to perturb a subset of healthy organismoids by means of the natural or artificial induction of diseases are enrolled to emulate an individual's disease process. Finally, we discuss using such series of healthy and perturbed organismoids in predictively selecting, scheduling and dosing an individual patient's personalized therapy or medicine precisely. The potential impact of the organismoid theory on our healthcare system generally and the rapid adoption of disruptive personalized T-cell therapies particularly is highlighted.

Organ-on-a-chip: Determine feasibility of a human liver microphysiological model to assess long-term steroid metabolites in sports drug testing.

A fundamental challenge in preventive doping research is the study of metabolic pathways of substances banned in sport. However, the pharmacological predictions obtained by conventional in vitro or in vivo animal studies are occasionally of limited transferability to humans according to an inability of in vitro models to mimic higher order system physiology or due to various species-specific differences using animal models. A more recently established technology for simulating human physiology is the "organ-on-a-chip" principle. In a multichannel microfluidic cell culture chip, 3-dimensional tissue spheroids, which can constitute artificial and interconnected microscale organs, imitate principles of the human physiology. The objective of this study was to determine if the technology is suitable to adequately predict metabolic profiles of prohibited substances in sport. As model compounds, the frequently misused anabolic steroids, stanozolol and dehydrochloromethyltestosterone (DHCMT) were subjected to human liver spheroids in microfluidic cell culture chips. The metabolite patterns produced and circulating in the chip media were then assessed by LC-HRMS/(MS) at different time points of up to 14 days of incubation at 37°C. The overall profile of observed glucurono-conjugated stanozolol metabolites excellently matched the commonly found urinary pattern of metabolites, including 3'OH-stanozolol-glucuronide and stanozolol-N-glucuronides. Similarly, but to a lower extent, the DHCMT metabolic profile was in agreement with phase-I and phase-II biotransformation products regularly seen in postadministration urine specimens. In conclusion, this pilot study indicates that the "organ-on-a-chip" technology provides a high degree of conformity with traditional human oral administration studies, providing a promising approach for metabolic profiling in sports drug testing.

Supporting dataset of two integration-free induced pluripotent stem cell lines from related human donors.

Integration-free induced pluripotent stem cells from related human donors' exhibit great potential to the ongoing development of organ models. Blood cells from two different human donors were isolated, purified and reprogrammed into induced pluripotent stem cells. These induced pluripotent stem cell lines were characterized precisely for pluripotency markers (with the PluriTest and flow cytometry analysis) and their differentiation capacities into meso-, ecto- and endoderm. The induced pluripotent stem cell lines are available for commercial use and are therefore of high interest for many groups working in stem cell research. A normal karyotype of the induced pluripotent stem cells was proven with the KaryoStat assay. In total 6 human donors that belong to one family donated blood for induced pluripotent stem cell reprogramming. In this "Data in Brief" publication, we show the dataset for the two male iPSC lines HUMIMIC TISSUi006-A (StemUse106) and TISSUi007-A (StemUse107). The main characterisation was recently published by Ramme et al. in Stem Cell Research [1]. All iPSC lines were also examined negative for any mycoplasma or bacteria contamination.

Generation of two additional integration-free iPSC lines from related human donors.

The integration-free iPSC lines TISSUi006-A and TISSUi007-A were generated by reprogramming blood cells with episomal vectors. The male human donors belong to a Caucasian family in which four additional family members donated and iPSC lines were generated. All iPSC lines within this family are approved for commercial use by donor consent. Those iPSC lines offer the opportunity to study the influence of affiliation within one family. In future, more iPSCs lines of many more family members can be created to understand the effects of relatives with different ages on the reprogramming into iPSCs and differentiation into specific cell types.

Generation of four integration-free iPSC lines from related human donors.

Four integration-free iPSC lines were generated by reprogramming peripheral blood mononuclear cells with episomal vectors. All four human donors (two male and two female donors) belong to one Caucasian family within three different generations with the age between 19-82 years. Additionally, all iPSC lines are approved for commercial use by donor consent. Those iPSC lines offer the opportunity to study the influence of affiliation within one family. In future, more iPSCs lines of many more family members can be created to understand the effects of relatives with different ages on the reprogramming into iPSCs and differentiation into specific cell types.

Autologous induced pluripotent stem cell-derived four-organ-chip.

Microphysiological systems play a pivotal role in progressing toward a global paradigm shift in drug development. Here, we designed a four-organ-chip interconnecting miniaturized human intestine, liver, brain and kidney equivalents. All four organ models were predifferentiated from induced pluripotent stem cells from the same healthy donor and integrated into the microphysiological system. The coculture of the four autologous tissue models in one common medium deprived of tissue specific growth factors was successful over 14-days. Although there were no added growth factors present in the coculture medium, the intestine, liver and neuronal model maintained defined marker expression. Only the renal model was overgrown by coexisting cells and did not further differentiate. This model platform will pave the way for autologous coculture cross-talk assays, disease induction and subsequent drug testing.

A multi-organ chip co-culture of neurospheres and liver equivalents for long-term substance testing.

Current in vitro and animal tests for drug development are failing to emulate the systemic organ complexity of the human body and, therefore, often do not accurately predict drug toxicity, leading to high attrition rates in clinical studies (Paul et al., 2010). The phylogenetic distance between humans and laboratory animals is enormous, this affects the transferability of animal data on the efficacy of neuroprotective drugs. Therefore, many neuroprotective treatments that have shown promise in animals have not been successful when transferred to humans (Dragunow, 2008; Gibbons and Dragunow, 2010). We present a multi-organ chip capable of maintaining 3D tissues derived from various cell sources in a combined media circuit which bridges the gap in systemic and human tests. A steady state co-culture of human artificial liver microtissues and human neurospheres exposed to fluid flow over two weeks in the multi-organ chip has successfully proven its long-term performance. Daily lactate dehydrogenase activity measurements of the medium and immunofluorescence end-point staining proved the viability of the tissues and the maintenance of differentiated cellular phenotypes. Moreover, the lactate production and glucose consumption values of the tissues cultured indicated that a stable steady-state was achieved after 6 days of co-cultivation. The neurospheres remained differentiated neurons over the two-week cultivation in the multi-organ chip, proven by qPCR and immunofluorescence of the neuronal markers ßIII-tubulin and microtubule-associated protein-2. Additionally, a two-week toxicity assay with a repeated substance exposure to the neurotoxic 2,5-hexanedione in two different concentrations induced high apoptosis within the neurospheres and liver microtissues, as shown by a strong increase of lactate dehydrogenase activity in the medium. The principal finding of the exposure of the co-culture to 2,5-hexanedione was that not only toxicity profiles of two different doses could be discriminated, but also that the co-cultures were more sensitive to the substance compared to respective single-tissue cultures in the multi-organ-chip. Thus, we provide here a new in vitro tool which might be utilized to predict the safety and efficacy of substances in clinical studies more accurately in the future.