4 research outputs found
Skeletal muscle differentiation of human iPSCs meets bioengineering strategies: perspectives and challenges
Although skeletal muscle repairs itself following small injuries, genetic diseases or severe damages may hamper its ability to do so. Induced pluripotent stem cells (iPSCs) can generate myogenic progenitors, but their use in combination with bioengineering strategies to modulate their phenotype has not been sufficiently investigated. This review highlights the potential of this combination aimed at pushing the boundaries of skeletal muscle tissue engineering. First, the overall organization and the key steps in the myogenic process occurring in vivo are described. Second, transgenic and non-transgenic approaches for the myogenic induction of human iPSCs are compared. Third, technologies to provide cells with biophysical stimuli, biomaterial cues, and biofabrication strategies are discussed in terms of recreating a biomimetic environment and thus helping to engineer a myogenic phenotype. The embryonic development process and the pro-myogenic role of the muscle-resident cell populations in co-cultures are also described, highlighting the possible clinical applications of iPSCs in the skeletal muscle tissue engineering field
Real-time bioimpedance measurements of stem cellbased disease models-on-a-chip
In vitro disease models are powerful platforms for the development of drugs and novel therapies. Stem-cell
based approaches have emerged as cutting-edge tools in disease modelling, allowing for deeper
insights into previously unknown disease mechanisms. Hence the significant role of these disease-in-a-dish
methods in therapeutics and translational medicine.
Impedance sensing is a non-invasive, quantitative technique that can monitor changes in cellular
behaviour and morphology in real-time. Bioimpedance measurements can be used to characterize and
evaluate the establishment of a valid disease model, without the need for invasive end-point biochemical
assays. In this work, two stem cell-based disease models-on-a-chip are proposed for acute liver failure
(ALF) and age-related macular degeneration (AMD).
The ALF disease model-on-a-chip integrates impedance sensing with the highly-differentiated HepaRG
cell line to monitor in real-time quantitative and dynamic response to various hepatotoxins.
Bioimpedance analysis and modelling has revealed an unknown mechanism of paracetamol
hepatotoxicity; a temporal, dose-dependent disruption of tight junctions (TJs) and cell-substrate
adhesion. This disruption has been validated using ultrastructural imaging and immunostaining of the
TJ-associated protein ZO-1.
Age-related macular degeneration (AMD) is the leading cause of blindness in the developed world with
a need for disease models for its currently incurable forms. Human induced pluripotent stem cells
(hiPSCs) technology offers a novel approach for disease modelling, with the potential to impact
translational retinal research and therapy. Recent developments enable the generation of Retinal
Pigment Epithelial cells from patients (hiPSC-RPE), thus allowing for human retinal disease in vitro
studies with great clinical and physiological relevance. In the current study, the development of a tissue-on-
a-chip AMD disease model has been established using RPE generated from a patient with an
inherited macular degeneration (case cell line) and from a healthy sibling (control cell line).
A reproducible Electric Cell-substrate Impedance Sensing (ECIS) electrical wounding assay was
conducted to mimic RPE damage in AMD. First, a robust and reproducible real-time quantitative
monitoring over a 25-day period demonstrated the establishment and maturation of RPE layers on
microelectrodes. A spatially-controlled RPE layer damage that mimicked cell loss in AMD was then
initiated. Post recovery, significant differences in migration rates were found between case and control
cell lines. Data analysis and modelling suggested this was due to the lower cell-substrate adhesion of
the control cell line. These findings were confirmed using cell adhesion biochemical assays. Moreover,
different-sized, individually-addressed square microelectrode arrays with high spatial resolution were
designed and fabricated in-house. ECIS wounding assays were performed on these chips to study
immortalized RPE migration. Migration rates comparable to those obtained with ECIS circular
microelectrodes were determined.
The two proposed disease-models-on-a-chip were then used to explore the therapeutic potential of the
antioxidant N-Acetyl-Cysteine (NAC) on hiPSC-RPE and HepaRG cell recovery. Addition of 10 mM
NAC at the end of a 24h paracetamol challenge caused a slight increase in the measured impedance,
suggesting partial cell recovery. On the other hand, no effect on case hiPSC-RPE migration has been
observed. More experiments are needed to examine the effect of different NAC concentrations and
incubation periods. The therapeutic potential of electrical stimulation has also been explored. A
preliminary study to evaluate the effect of electrical stimulation on RPE migration has been conducted.
An externally applied direct current electric field (DC EF) of 300 mV/mm was found to direct the
migration of the immortalized RPE cell line (hTERT-RPE1) perpendicular to the EF. The cells were
also observed to elongate and to realign their long axes perpendicular to the applied EF.
The proposed tissue-on-a-chip disease models are powerful platforms for translational studies. The
potential of such platforms has been demonstrated through revealing unknown effects of acetaminophen
on the liver as well as providing deeper insights into the underlying mechanisms of macular
degeneration. Combining stem cell technology with impedance sensing provides a high throughput
platform for studying patient-specific diseases and evaluating potential therapies
Engineering stem cells for future medicine
Despite their great potential in regenerative medicine applications, stem cells (especially pluripotent ones) currently show a limited clinical success, partly due to a lack of biological knowledge, but also due to a lack of specific and advanced technological instruments able to overcome the current boundaries of stem cell functional maturation and safe/effective therapeutic delivery. This paper aims at describing recent insights, current limitations, and future horizons related to therapeutic stem cells, by analyzing the potential of different bioengineering disciplines in bringing stem cells toward a safe clinical use. First, we clarify how and why stem cells should be properly engineered and which could be in a near future the challenges and the benefits connected with this process. Second, we identify different routes toward stem cell differentiation and functional maturation, relying on chemical, mechanical, topographical, and direct/indirect physical stimulation. Third, we highlight how multiscale modeling could strongly support and optimize stem cell engineering. Finally, we focus on future robotic tools that could provide an added value to the extent of translating basic biological knowledge into clinical applications, by developing ad hoc enabling technologies for stem cell delivery and control