An engineered multicellular stem cell niche for the 3D derivation of human myogenic progenitors from iPSCs.

Fate decisions in the embryo are controlled by a plethora of microenvironmental interactions in a three-dimensional niche. To investigate whether aspects of this microenvironmental complexity can be engineered to direct myogenic human-induced pluripotent stem cell (hiPSC) differentiation, we here screened murine cell types present in the developmental or adult stem cell niche in heterotypic suspension embryoids. We identified embryonic endothelial cells and fibroblasts as highly permissive for myogenic specification of hiPSCs. After two weeks of sequential Wnt and FGF pathway induction, these three-component embryoids are enriched in Pax7-positive embryonic-like myogenic progenitors that can be isolated by flow cytometry. Myogenic differentiation of hiPSCs in heterotypic embryoids relies on a specialized structural microenvironment and depends on MAPK, PI3K/AKT, and Notch signaling. After transplantation in a mouse model of Duchenne muscular dystrophy, embryonic-like myogenic progenitors repopulate the stem cell niche, reactivate after repeated injury, and, compared to adult human myoblasts, display enhanced fusion and lead to increased muscle function. Altogether, we provide a two-week protocol for efficient and scalable suspension-based 3D derivation of Pax7-positive myogenic progenitors from hiPSCs.

In vivo transcriptomic profiling using cell encapsulation identifies effector pathways of systemic aging.

Sustained exposure to a young systemic environment rejuvenates aged organisms and promotes cellular function. However, due to the intrinsic complexity of tissues it remains challenging to pinpoint niche-independent effects of circulating factors on specific cell populations. Here, we describe a method for the encapsulation of human and mouse skeletal muscle progenitors in diffusible polyethersulfone hollow fiber capsules that can be used to profile systemic aging in vivo independent of heterogeneous short-range tissue interactions. We observed that circulating long-range signaling factors in the old systemic environment lead to an activation of Myc and E2F transcription factors, induce senescence, and suppress myogenic differentiation. Importantly, in vitro profiling using young and old serum in 2D culture does not capture all pathways deregulated in encapsulated cells in aged mice. Thus, in vivo transcriptomic profiling using cell encapsulation allows for the characterization of effector pathways of systemic aging with unparalleled accuracy.

Nutritional Control of Intestinal Stem Cells in Homeostasis and Tumorigenesis.

Food and nutrition have a profound impact on organismal health and diseases, and tissue-specific adult stem cells play a crucial role in coordinating tissue maintenance by responding to dietary cues. Emerging evidence indicates that adult intestinal stem cells (ISCs) actively adjust their fate decisions in response to diets and nutritional states to drive intestinal adaptation. Here, we review the signaling mechanisms mediating the dietary responses imposed by caloric intake and nutritional composition (i.e., macronutrients and micronutrients), fasting-feeding patterns, diet-induced growth factors, and microbiota on ISCs and their relevance to the beginnings of intestinal tumors.

Neural priming of adipose-derived stem cells by cell-imprinted substrates.

Cell-imprinting technology is a novel method for directing stem cell fate using substrates molded from target cells. Here, we fabricated and studied cell-imprinted substrates for neural priming in human adipose-derived stem cells in the absence of chemical cues. We molded polydimethylsiloxane silicone substrates on fixed differentiated neural progenitor cells (ReNcellTMVM). The ReNcellTMcell line consists of immortalized human neural progenitor cells that are capable to differentiate into neural cells. The fabricated cell-imprinted silicone substrates represent the geometrical micro- and nanotopology of the target cell morphology. During the molding procedure, no transfer of cellular proteins was detectable. In the first test with undifferentiated ReNcellTMVM cells, the cell-imprinted substrates could accelerate neural differentiation. With adipose-derived stem cells cultivated on the imprinted substrates, we observed modifications of cell morphology, shifting from spread to elongated shape. Both immunofluorescence and quantitative gene expression analysis showed upregulation of neural stem cell and early neuronal markers. Our study, for the first time, demonstrated the effectiveness of cell-imprinted substrates for neural priming of adipose-derived stem cells for regenerative medicine applications.

Aging Disrupts Muscle Stem Cell Function by Impairing Matricellular WISP1 Secretion from Fibro-Adipogenic Progenitors.

Research on age-related regenerative failure of skeletal muscle has extensively focused on the phenotypes of muscle stem cells (MuSCs). In contrast, the impact of aging on regulatory cells in the MuSC niche remains largely unexplored. Here, we demonstrate that aging impairs the function of mouse fibro-adipogenic progenitors (FAPs) and thereby indirectly affects the myogenic potential of MuSCs. Using transcriptomic profiling, we identify WNT1 Inducible Signaling Pathway Protein 1 (WISP1) as a FAP-derived matricellular signal that is lost during aging. WISP1 is required for efficient muscle regeneration and controls the expansion and asymmetric commitment of MuSCs through Akt signaling. Transplantation of young FAPs or systemic treatment with WISP1 restores the myogenic capacity of MuSCs in aged mice and rescues skeletal muscle regeneration. Our work establishes that loss of WISP1 from FAPs contributes to MuSC dysfunction in aged skeletal muscles and demonstrates that this mechanism can be targeted to rejuvenate myogenesis.

The Muscle Stem Cell Niche in Health and Disease.

The regulation of stem cells that maintain and regenerate postnatal tissues depends on extrinsic signals originating from their microenvironment, commonly referred to as the stem cell niche. Complex higher-order regulatory interrelationships with the tissue and factors in the systemic circulation are integrated and propagated to the stem cells through the niche. The stem cell niche in skeletal muscle tissue is both a paradigm for a structurally and functionally relatively static niche that maintains stem cell quiescence during tissue homeostasis, and a highly dynamic regenerative niche that is subject to extensive structural remodeling and a flux of different support cell populations. Conditions ranging from aging to chronically degenerative skeletal muscle diseases affect the composition of the niche and thereby impair the regenerative potential of muscle stem cells. A holistic and integrative understanding of the extrinsic mechanisms regulating muscle stem cells in health and disease in a broad systemic context will be imperative for the identification of regulatory hubs in the niche interactome that can be targeted to maintain, restore, or enhance the regenerative capacity of muscle tissue. Here, we review the microenvironmental regulation of muscle stem cells, summarize how niche dysfunction can contribute to disease, and discuss emerging therapeutic implications.

Development of a Virtual Cell Model to Predict Cell Response to Substrate Topography.

Cells can sense and respond to changes in the topographical, chemical, and mechanical information in their environment. Engineered substrates are increasingly being developed that exploit these physical attributes to direct cell responses (most notably mesenchymal stem cells) and therefore control cell behavior toward desired applications. However, there are very few methods available for robust and accurate modeling that can predict cell behavior prior to experimental evaluations, and this typically means that many cell test iterations are needed to identify best material features. Here, we developed a unifying computational framework to create a multicomponent cell model, called the "virtual cell model" that has the capability to predict changes in whole cell and cell nucleus characteristics (in terms of shape, direction, and even chromatin conformation) on a range of cell substrates. Modeling data were correlated with cell culture experimental outcomes in order to confirm the applicability of the virtual cell model and demonstrating the ability to reflect the qualitative behavior of mesenchymal stem cells. This may provide a reliable, efficient, and fast high-throughput approach for the development of optimized substrates for a broad range of cellular applications including stem cell differentiation.

Engineering natural heart valves: possibilities and challenges.

Heart valve replacement is considered to be the most prevalent treatment approach for cardiac valve-related diseases. Among current solutions for heart valve replacement, e.g. mechanical and bioprosthetic valves, the main shortcoming is the lack of growth capability, repair and remodelling of the substitute valve. During the past three decades, tissue engineering-based approaches have shown tremendous potential to overcome these limitations by the development of a biodegradable scaffold, which provides biomechanical and biochemical properties of the native tissue. Among various scaffolds employed for tissue engineering, the decellularized heart valve (DHV) has attracted much attention, due to its native structure as well as comparable haemodynamic characteristics. Although the human DHV has shown optimal properties for valve replacement, the limitation of valve donors in terms of time and size is their main clinical issue. In this regard, xenogenic DHV can be a promising candidate for heart valve replacement. Xenogenic DHVs have similar composition to human valves, which will overcome the need for human DHVs. The main concern regarding xenogeneic DHV replacement is the immunological reaction and calcification following implantation, weak mechanical properties and insufficient recellularization capacity. In this review, we describe the essential steps required to address these impediments through novel engineering approaches. Copyright © 2016 John Wiley & Sons, Ltd.

Cell-Imprinted Substrates Modulate Differentiation, Redifferentiation, and Transdifferentiation.

Differentiation of stem cells into mature cells through the use of physical approaches is of great interest. Here, we prepared smart nanoenvironments by cell-imprinted substrates based on chondrocytes, tenocytes, and semifibroblasts as templates and demonstrated their potential for differentiation, redifferentiation, and transdifferentiation. Analysis of shape and upregulation/downregulation of specific genes of stem cells, which were seeded on these cell-imprinted substrates, confirmed that imprinted substrates have the capability to induce specific shapes and molecular characteristics of the cell types that were used as templates for cell-imprinting. Interestingly, immunofluorescent staining of a specific protein in chondrocytes (i.e., collagen type II) confirmed that adipose-derived stem cells, semifibroblasts, and tenocytes can acquire the chondrocyte phenotype after a 14 day culture on chondrocyte-imprinted substrates. In summary, we propose that common polystyrene tissue culture plates can be replaced by this imprinting technique as an effective and promising way to regulate any cell phenotype in vitro with significant potential applications in regenerative medicine and cell-based therapies.

Osmolarity: A hidden factor in Nanotoxicology.

In the field of drug delivery, long circulating nanocarriers in the blood have many advantages such as targeted drug delivery and sustained release. Based on our current knowledge, evaluation of the effect of long circulating nanocarriers in the blood stream on osmolarity of plasma has not been reported before. In this study, osmotic pressure developed by some commercially available nanocarriers was estimated based on Van't Hoff equation. It is noteworthy that theoretically, nanocarriers do not have any significant effect on osmolarity of plasma. However, it is worth being evaluated experimentally in order to be taken into account in future studies.

Nanoelectromechanical Chip (NELMEC) Combination of Nanoelectronics and Microfluidics to Diagnose Epithelial and Mesenchymal Circulating Tumor Cells from Leukocytes.

An integrated nano-electromechanical chip (NELMEC) has been developed for the label-free distinguishing of both epithelial and mesenchymal circulating tumor cells (ECTCs and MCTCs, respectively) from white blood cells (WBCs). This nanoelectronic microfluidic chip fabricated by silicon micromachining can trap large single cells (>12 µm) at the opening of the analysis microchannel arrays. The nature of the captured cells is detected using silicon nanograss (SiNG) electrodes patterned at the entrance of the channels. There is an observable difference between the membrane capacitance of the ECTCs and MCTCs and that of WBCs (measured using SiNG electrodes), which is the key indication for our diagnosis. The NELMEC chip not only solves the problem of the size overlap between CTCs and WBCs but also detects MCTCs without the need for any markers or tagging processes, which has been an important problem in previously reported CTC detection systems. The great conductivity of the gold-coated SiNG nanocontacts as well as their safe penetration into the membrane of captured cells, facilitate a precise and direct signal extraction to distinguish the type of captured cell. The results achieved from epithelial (MCF-7) and mesenchymal (MDA-MB231) breast cancer cells circulated in unprocessed blood suggest the significant applications for these diagnostic abilities of NELMEC.

Harnessing the Cancer Radiation Therapy by Lanthanide-Doped Zinc Oxide Based Theranostic Nanoparticles.

In this paper, doping of europium (Eu) and gadolinium (Gd) as high-Z elements into zinc oxide (ZnO) nanoparticles (NPs) was designed to optimize restricted energy absorption from a conventional radiation therapy by X-ray. Gd/Eu-doped ZnO NPs with a size of 9 nm were synthesized by a chemical precipitation method. The cytotoxic effects of Eu/Gd-doped ZnO NPs were determined using MTT assay in L929, HeLa, and PC3 cell lines under dark conditions as well as exposure to ultraviolet, X-ray, and γ radiation. Doped NPs at 20 µg/mL concentration under an X-ray dose of 2 Gy were as efficient as 6 Gy X-ray radiation on untreated cells. It is thus suggested that the doped NPs may be used as photoinducers to increase the efficacy of X-rays within the cells, consequently, cancer cell death. The doped NPs also could reduce the received dose by normal cells around the tumor. Additionally, we evaluated the diagnostic efficacy of doped NPs as CT/MRI nanoprobes. Results showed an efficient theranostic nanoparticulate system for simultaneous CT/MR imaging and cancer treatment.

Possibilities in Germ Cell Research: An Engineering Insight.

Germ cells (GCs) are responsible for fertility and disruptions in their development or function cause infertility. However, current knowledge about the diverse mechanisms involved in GC development and function is still in its infancy. This is mainly because there are low numbers of GCs, especially during embryonic development. A deeper understanding of GCs would enhance our ability to produce them from stem cells. In addition, such information would enable the production of healthy gametes for infertile couples. In this regard, pluripotent stem cells (PSCs) demonstrated a promising ability to produce GCs in vitro. In this review, we highlight recent advances in the field of tissue engineering that suggest novel strategies to enhance GC research.

Personalized disease-specific protein corona influences the therapeutic impact of graphene oxide.

The hard corona, the protein shell that is strongly attached to the surface of nano-objects in biological fluids, is recognized as the first layer that interacts with biological objects (e.g., cells and tissues). The decoration of the hard corona (i.e., the type, amount, and conformation of the attached proteins) can define the biological fate of the nanomaterial. Recent developments have revealed that corona decoration strongly depends on the type of disease in human patients from which the plasma is obtained as a protein source for corona formation (referred to as the 'personalized protein corona'). In this study, we demonstrate that graphene oxide (GO) sheets can trigger different biological responses in the presence of coronas obtained from various types of diseases. GO sheets were incubated with plasma from human subjects with different diseases/conditions, including hypofibrinogenemia, blood cancer, thalassemia major, thalassemia minor, rheumatism, fauvism, hypercholesterolemia, diabetes, and pregnancy. Identical sheets coated with varying protein corona decorations exhibited significantly different cellular toxicity, apoptosis, and uptake, reactive oxygen species production, lipid peroxidation and nitrogen oxide levels. The results of this report will help researchers design efficient and safe, patient-specific nano biomaterials in a disease type-specific manner for clinical and biological applications.

Regulation of stem cell fate by nanomaterial substrates.

Stem cells are increasingly studied because of their potential to underpin a range of novel therapies, including regenerative strategies, cell type-specific therapy and tissue repair, among others. Bionanomaterials can mimic the stem cell environment and modulate stem cell differentiation and proliferation. New advances in these fields are presented in this review. This work highlights the importance of topography and elasticity of the nano-/micro-environment, or niche, for the initiation and induction of stem cell differentiation and proliferation.

A single-cell correlative nanoelectromechanosensing approach to detect cancerous transformation: monitoring the function of F-actin microfilaments in the modulation of the ion channel activity.

Cancerous transformation may be dependent on correlation between electrical disruptions in the cell membrane and mechanical disruptions of cytoskeleton structures. Silicon nanotube (SiNT)-based electrical probes, as ultra-accurate signal recorders with subcellular resolution, may create many opportunities for fundamental biological research and biomedical applications. Here, we used this technology to electrically monitor cellular mechanosensing. The SiNT probe was combined with an electrically activated glass micropipette aspiration system to achieve a new cancer diagnostic technique that is based on real-time correlation between mechanical and electrical behaviour of single cells. Our studies demonstrated marked changes in the electrical response following increases in the mechanical aspiration force in healthy cells. In contrast, such responses were extremely weak for malignant cells. Confocal microscopy results showed the impact of actin microfilament remodelling on the reduction of the electrical response for aspirated cancer cells due to the significant role of actin in modulating the ion channel activity in the cell membrane.

Impacts of quantum dots in molecular detection and bioimaging of cancer.

INTRODUCTION: A number of assays have so far been exploited for detection of cancer biomarkers in various malignancies. However, the expression of cancer biomarker(s) appears to be extremely low, therefore accurate detection demands sensitive optical imaging probes. While optical detection using conventional fluorophores often fail due to photobleaching problems, quantum dots (QDs) offer stable optical imaging in vitro and in vivo. METHODS: In this review, we briefly overview the impacts of QDs in biology and its applications in bioimaging of malignancies. We will also delineate the existing obstacles for early detection of cancer and the intensifying use of QDs in advancement of diagnostic devices. RESULTS: Of the QDs, unlike the II-VI type QDs (e.g., cadmium (Cd), selenium (Se) or tellurium (Te)) that possess inherent cytotoxicity, the I-III-VI 2 type QDs (e.g., AgInS2, CuInS2, ZnS-AgInS2) appear to be less toxic bioimaging agents with better control of band-gap energies. As highly-sensitive bioimaging probes, advanced hybrid QDs (e.g., QD-QD, fluorochrome-QD conjugates used for sensing through fluorescence resonance energy transfer (FRET), quenching, and barcoding techniques) have also been harnessed for the detection of biomarkers and the monitoring of delivery of drugs/genes to the target sites. Antibody-QD (Ab-QD) and aptamer- QD (Ap-QD) bioconjugates, once target the relevant biomarker, can provide highly stable photoluminescence (PL) at the target sites. In addition to their potential as nanobiosensors, the bioconjugates of QDs with homing devices have successfully been used for the development of smart nanosystems (NSs) providing targeted bioimaging and photodynamic therapy (PDT). CONCLUSION: Having possessed great deal of photonic characteristics, QDs can be used for development of seamless multifunctional nanomedicines, theranostics and nanobiosensors.

Cell-imprinted substrates act as an artificial niche for skin regeneration.

Bioinspired materials can mimic the stem cell environment and modulate stem cell differentiation and proliferation. In this study, biomimetic micro/nanoenvironments were fabricated by cell-imprinted substrates based on mature human keratinocyte morphological templates. The data obtained from atomic force microscopy and field emission scanning electron microscopy revealed that the keratinocyte-cell-imprinted poly(dimethylsiloxane) casting procedure could imitate the surface morphology of the plasma membrane, ranging from the nanoscale to the macroscale, which may provide the required topographical cell fingerprints to induce differentiation. Gene expression levels of the genes analyzed (involucrin, collagen type I, and keratin 10) together with protein expression data showed that human adipose-derived stem cells (ADSCs) seeded on these cell-imprinted substrates were driven to adopt the specific shape and characteristics of keratinocytes. The observed morphology of the ADSCs grown on the keratinocyte casts was noticeably different from that of stem cells cultivated on the stem-cell-imprinted substrates. Since the shape and geometry of the nucleus could potentially alter the gene expression, we used molecular dynamics to probe the effect of the confining geometry on the chain arrangement of simulated chromatin fibers in the nuclei. The results obtained suggested that induction of mature cell shapes onto stem cells can influence nucleus deformation of the stem cells followed by regulation of target genes. This might pave the way for a reliable, efficient, and cheap approach of controlling stem cell differentiation toward skin cells for wound healing applications.

Electrochemical impedance spectroscopic sensing of methamphetamine by a specific aptamer.

INTRODUCTION: Electrochemical impedance spectroscopy (EIS) is a simple and highly sensitive technique that can be used for evaluation of the aptamer-target interaction even in a label-free approach. METHODS: To pursue the effectiveness of EIS, in the current study, the folding properties of specific aptamer for methamphetamine (METH) (i.e., aptaMETH) were evaluated in the presence of METH and amphetamine (Amph). Folded and unfolded aptaMETH was mounted on the gold electrode surface and the electron charge transfer was measured by EIS. RESULTS: The Ret of methamphetamine-aptaMETH was significantly increased in comparison with other folding conditions, indicating specific detection of METH by aptaMETH. CONCLUSION: Based on these findings, methamphetamine-aptaMETH on the gold electrode surface displayed the most interfacial electrode resistance and thus the most folding situation. This clearly indicates that the aptaMETH can profoundly and specifically pinpoint METH; as a result we suggest utilization of this methodology for fast and cost-effective identification of METH.