Stem Cell Colonies Research Articles

Substrate elasticity dependent colony formation and cardiac differentiation of human induced pluripotent stem cells

Substrate elasticity regulates cell functions including cell aggregation and stem cell differentiation. The ability to manufacture substrates of desired elasticity over a broad range is therefore crucial for both fundamental research and advanced applications. In this work, we developed a method to fabricate dense elastomer pillars of different heights on a rigid substrate, providing an effective elasticity ranging from 3 to 168 kPa. Assisted with an elastomer stencil of honeycomb pattern for cell seeding, we obtained uniform colonies of human induced pluripotent stem cells (hiPSCs) and differentiated cardiomyocytes on the pillar substrates of different modulus. Our results showed that the elasticity of substrates significantly affected the cell colony formation via governing the colony edge propagation. More importantly, the results demonstrated that an intermediate substrate elasticity of about 9 kPa is preferable to reach an embryoid-like aggregation and optimal for cardiac differentiation of hiPSCs. Overall, this work sheds new insights on the importance of substrate modulus on cell aggregation and stem cell differentiation as well as the manufacturing of culture substrates with desired elasticity.

Hair-Follicle-Associated Pluripotent (HAP) Stem Cells Encapsulated on Polyvinylidene Fluoride Membranes (PFM) Promote Functional Recovery from Spinal Cord Injury.

Our previous studies showed that nestin-expressing hair follicle-associated-pluripotent (HAP) stem cells, which reside in the bulge area of the hair follicle, could restore injured nerve and spinal cord and differentiate into cardiac muscle cells. Here we transplanted mouse green fluorescent protein (GFP)-expressing HAP stem-cell colonies enclosed on polyvinylidene fluoride membranes (PFM) into the severed thoracic spinal cord of nude mice. After seven weeks of implantation, we found the differentiation of HAP stem cells into neurons and glial cells. Our results also showed that PFM-captured GFP-expressing HAP stem-cell colonies assisted complete reattachment of the thoracic spinal cord. Furthermore, our quantitative motor function analysis with the Basso Mouse Scale for Locomotion (BMS) score demonstrated a significant improvement in the implanted mice compared to non-implanted mice with a severed spinal cord. Our study also showed that it is easy to obtain HAP stem cells, they do not develop teratomas, and do not loose differentiation ability when cryopreserved. Collectively our results suggest that HAP stem cells could be abetter source compared to induced pluripotent stem cells (iPS) or embryonic stem (ES) cells for regenerative medicine, specifically for spinal cord repair.

Spatiotemporal mosaic self-patterning of pluripotent stem cells using CRISPR interference.

Morphogenesis involves interactions of asymmetric cell populations to form complex multicellular patterns and structures comprised of distinct cell types. However, current methods to model morphogenic events lack control over cell-type co-emergence and offer little capability to selectively perturb specific cell subpopulations. Our in vitro system interrogates cell-cell interactions and multicellular organization within human induced pluripotent stem cell (hiPSC) colonies. We examined effects of induced mosaic knockdown of molecular regulators of cortical tension (ROCK1) and cell-cell adhesion (CDH1) with CRISPR interference. Mosaic knockdown of ROCK1 or CDH1 resulted in differential patterning within hiPSC colonies due to cellular self-organization, while retaining an epithelial pluripotent phenotype. Knockdown induction stimulates a transient wave of differential gene expression within the mixed populations that stabilized in coordination with observed self-organization. Mosaic patterning enables genetic interrogation of emergent multicellular properties, which can facilitate better understanding of the molecular pathways that regulate symmetry-breaking during morphogenesis.

Influence of wideband visible light with an padding red component on the functional state of mice embryos and embryonic stem cells

It is known that visible light, including sunlight and laboratory lighting, adversely affect the development of embryos in vitro. In with article we present a technology for the synthesis of composite screens, capable to photoconvert UV and a part of the blue spectrum into red light with the maximum ~630 nm. It is established that the application of such transformed light with an evident red component raises the chances of embryos to survive and protects embryonic stem cells. To create photoconversion screens, the CdZn/Se quantum dots were obtained, the average size being about 7 nm. When the quantum dots are excited by electromagnetic waves of the UV and blue spectral range, photoluminescence is observed. The average photon energy for photoluminescence is of the order of 2 eV. On the basis of CdZn/Se quantum dots and methylphenylsiloxane polymer, light-transforming composite screens were made. In case of the light-transforming composite screen, the UV component disappeared from the energy spectrum, and the intensity of the blue region of the spectrum was reduced. On the contrary, in the red region (λmax = 630 nm) one can see a little more than two-fold increase of intensity. It is shown that when exposed to 2-cell embryos by transformed light, the proportion of normally developing embryos increases by 20%, the number of dead embryos decreases twice, and number of dead and apoptotic cells was lower in blastocysts, what's decreased by 70%, as compared to the control group. When blastocysts are transferred to the feeder substrate, colonies of embryonic stem cells are formed. Cells obtained from blastocysts irradiated with transformed visible light are in a normal state in 90% of cases and did not change expression levels, biochemistry and morphology for at least 20 passages. It is assumed that the data obtained can be used for the design of systems of efficient cultivation of embryonic cells for tissue engineering and cell therapy.

Enhanced photon collection enables four dimensional fluorescence nanoscopy of living systems

The theoretically unlimited spatial resolution of fluorescence nanoscopy often comes at the expense of time, contrast and increased dose of energy for recording. Here, we developed MoNaLISA, for Molecular Nanoscale Live Imaging with Sectioning Ability, a nanoscope capable of imaging structures at a scale of 45–65 nm within the entire cell volume at low light intensities (W-kW cm−2). Our approach, based on reversibly switchable fluorescent proteins, features three distinctly modulated illumination patterns crafted and combined to gain fluorescence ON–OFF switching cycles and image contrast. By maximizing the detected photon flux, MoNaLISA enables prolonged (40–50 frames) and large (50 × 50 µm2) recordings at 0.3–1.3 Hz with enhanced optical sectioning ability. We demonstrate the general use of our approach by 4D imaging of organelles and fine structures in epithelial human cells, colonies of mouse embryonic stem cells, brain cells, and organotypic tissues.

Stem cell colony interspacing effect on differentiation to neural cells.

Efforts to enhance the efficiency of neural differentiation of stem cells are primarily focused on exogenous modulation of physical niche parameters such as surface topography and extracellular matrix proteins, or addition of certain growth factors or small molecules to culture media. We report a novel neurogenic niche to enhance the neural differentiation of embryonic stem cells (ESCs) without any external intervention by micropatterning ESCs into spatially organized colonies of controlled size and interspacing. Using an aqueous two-phase system cell microprinting technology, we generated pairs of uniformly sized isolated ESC colonies at defined interspacing distances over a layer of differentiation-inducing stromal cells. Our comprehensive analysis of temporal expression of neural genes and proteins of cells in colony pairs showed that interspacing two colonies at approximately 0.66 times the colony diameter (0.66D) significantly enhanced neural differentiation of ESCs. Cells in these colonies displayed higher expression of neural genes and proteins and formed thick neurite bundles between the two colonies. A computational model of spatial distribution of soluble factors of cells in interspaced colony pairs showed that the enhanced neural differentiation is due to the presence of stable concentration gradients of soluble signalling factors between the two colonies. Our results indicate that culturing ESCs in colony pairs with defined interspacing is a promising approach to efficiently derive neural cells. Additionally, this approach provides a platform for quantitative studies of molecular mechanisms that regulate neurogenesis of stem cells.

A fast method to reprogram and CRISPR/Cas9 gene editing from erythroblasts

An efficient one-step procedure to reprogram fibroblasts into human induced pluripotent stem cells (hiPSC) and perform CRISPR/Cas9 gene editing simultaneously was recently reported. Here we show that such simultaneous reprogramming and gene editing can be efficiently done from erythroblasts. We successfully obtained human induced pluripotent stem cells colonies together with in frame and out of frame CAPN1 mutations in one or both alleles. We did not identify off-targets in edited cell lines. The entire process, from blood collection to mutated hiPSC took approximately 5 weeks, a much shorter period than standard multi-step methodologies using fibroblasts. Noteworthy, blood drawing is a less invasive procedure than a skin biopsy.

Optimization of caprine embryo production in different media for generation of embryonic stem cell-like cells

The aim of the present study was to optimize the production of blastocyst for obtaining caprine embryonic stem cell-like cells. A total of 4372 cumulus oocyte complexes (COCs) were recovered by slicing the 1187 caprine ovaries and were matured in maturation media for 27 h in humidified atmosphere at 38.5°C with 5% CO2 in CO2 incubator. After 27 h of maturation, oocytes were denuded and were co-incubated with buck semen in fertilization medium (TALP medium + 8 mg/ml fatty acid free BSA and 50 μg/ml heparin) for 18 h. Good quality zygotes (2483) were selected and randomly divided into 2 groups (experiment 1), viz. Group 1 (1312) wherein the presumptive zygotes were cultured in RVCL while in Group 2 (1171) the presumptive zygotes were cultured in mCR2aa medium. The cleavage rate, blastocyst and hatched blastocyst production was significantly higher in Gr 1 (47.45±2.93, 10.13±1.31 and 3.90±1.13%) than Gr 2 (37.75±2.46, 4.20±0.93 and 1.66±0.72%). In experiment 2, after in-vitro fertilization, morula stage embryos and inner cell mass (ICM) from blastocyst and hatched blastocyst were used to isolate ES cell-like cells. Thus the results indicated that the RVCL medium is the best medium as far as the embryonic development up to blastocyst stage in comparison to mCR2aa media. Furthermore, the formation of putative embryonic stem cell colonies were higher from hatched blastocysts (91.6%) as compared to that of blastocysts (82.1%) and it was significantly higher than that from morulas (34.3%).

Self-organization of a human organizer by combined Wnt and Nodal signalling.

In amniotes, the development of the primitive streak (PS) and its accompanying “organizer” define the first stages of gastrulation. Despite detailed characterization in model organisms, the analogous human structures remain a mystery. We have previously shown that when stimulated with BMP4, micropatterned colonies of human embryonic stem cells (hESCs) self-organize to generate early embryonic germ layers1. Here we show that in the same type of colonies WNT signalling is sufficient to induce a PS, and WNT with ACTIVIN is sufficient to induce an organizer, as characterized by embryo-like sharp boundary formation, epithelial-to-mesenchymal transition (EMT) markers, and expression of the organizer specific transcription factor GSC. Moreover, when grafted into chick embryos, WNT and ACTIVIN treated human cells induce and contribute autonomously to a secondary axis while inducing neural fate in the host. This fulfills the most stringent functional criteria for an organizer, and its discovery represents a major milestone in human embryology.

Identification and Single-Cell Functional Characterization of an Endodermally Biased Pluripotent Substate in Human Embryonic Stem Cells.

SummaryHuman embryonic stem cells (hESCs) display substantial heterogeneity in gene expression, implying the existence of discrete substates within the stem cell compartment. To determine whether these substates impact fate decisions of hESCs we used a GFP reporter line to investigate the properties of fractions of putative undifferentiated cells defined by their differential expression of the endoderm transcription factor, GATA6, together with the hESC surface marker, SSEA3. By single-cell cloning, we confirmed that substates characterized by expression of GATA6 and SSEA3 include pluripotent stem cells capable of long-term self-renewal. When clonal stem cell colonies were formed from GATA6-positive and GATA6-negative cells, more of those derived from GATA6-positive cells contained spontaneously differentiated endoderm cells than similar colonies derived from the GATA6-negative cells. We characterized these discrete cellular states using single-cell transcriptomic analysis, identifying a potential role for SOX17 in the establishment of the endoderm-biased stem cell state.

What is the best protocol to cryopreserve immature mouse testicular cell suspensions?

From a clinical perspective, which parameters grant optimal cryopreservation of mouse testicular cell suspensions? We studied the effect of different cryopreservation rates, the addition of sugars, different vessels and the addition of an apoptotic inhibitor on the efficiency of testicular cell suspension cryopreservation. After thawing and warming, testicular cell suspensions were transplanted to recipient mice for further functional assay. After selecting the optimal cryopreservation procedure, a second experiment compared the transplantation efficiency between the selected freezing protocol and fresh testicular cell suspensions. Multiple- and single-step freezing did not differ significantly in terms of recovered viable cells (RVC) (33 ± 28% and 38 ± 25%). The addition of sucrose did not result in a higher RVC (33 ± 20%). Cells frozen in vials recovered better than those frozen in straws (52 ± 20% versus 33 ± 20%; P = 0.0049). The inclusion of an apoptosis inhibitor (z-VAD[Oe]-FMK) significantly increased the RVC after thawing (61 ± 18% versus 50 ± 17%; P = 0.0480). When comparing the optimal cryopreservation procedure with fresh testicular cell suspensions, a lower RVC (63 ± 11% versus 92 ± 4%; P < 0.0001) and number of donor-derived spermatogonial stem cell colonies per testis (34.04 ± 2.34 versus 16.78 ± 7.76; P = 0.0051) were observed. Upon freeze-thawing or vitrification-warming, and assessment of donor-derived spermatogenesis after transplantation, Dulbecco's modified Eagle's medium supplemented with 1.5M dimethyl-sulphoxide, 10% fetal calf serum and 60 µM of Z-VAD-(OMe)-FMK in vials at a freezing rate of -1°C/min was optimal.

Generation of a transgene-free human induced pluripotent stem cell line (UNIPDi001-A) from oral mucosa epithelial stem cells

Human oral mucosa epithelial stem cells (hOMESCs) were obtained from a fresh oral biopsy collected from a healthy subject at the Fondazione Banca degli Occhi del Veneto (FBOV). An integration-free reprogramming protocol was applied exploiting episomal plasmids transfected into cells using a Nucleofector device. Around day 20 post transfection, several human induced pluripotent stem cell (hiPSC) colonies were manually picked and expanded. One of these (UNIPDi001-A-hiPSCs) expressed undifferentiated state marker alkaline phosphatase along with a panel of pluripotency state markers and was able to differentiate into the derivatives of all the three germ layers.

Critical texture pattern feature assessment for characterizing colonies of induced pluripotent stem cells through machine learning techniques

The objectives of this study are to assess various automated texture features obtained from the segmented colony regions of induced pluripotent stem cells (iPSCs) and confirm their potential for characterizing the colonies using different machine learning techniques. One hundred and fifty-one features quantified using shape-based, moment-based, statistical and spectral texture feature groups are extracted from phase-contrast microscopic colony images of iPSCs. The forward stepwise regression model is implemented to select the most appropriate features required for categorizing the colonies. Support vector machine (SVM), random forest (RF), multilayer perceptron (MLP), decision tree (DT), and adaptive boosting (Adaboost) classifiers are used with ten-fold cross-validation to evaluate the texture features within each texture feature group and fused-features group to characterize healthy and unhealthy colonies of iPSCs. Overall, based on the classification performances of the four texture feature groups using the five classifier models, statistical features always exhibit a high predictive capacity (>87.5%). However, the classification performance using fused texture patterns with statistical, shape-based, and moment-based features was found to be robust and reliable with fewer false positive and false negative values compared to the features when either one is used for the classification of colonies of iPSCs. Furthermore, the results showcase that the SVM, RF and Adaboost classifiers deliver better classification performances than DT and MLP. Our findings suggest that the proposed automated fused statistical, shape-based, and moment-based texture pattern features trained with machine learning techniques are potentially more appropriate and helpful to biologists for characterizing colonies of stem cells.

Induced Pluripotent Stem Cells from a Marsupial, the Tasmanian Devil (Sarcophilus harrisii): Insight into the Evolution of Mammalian Pluripotency.

We demonstrate the generation of Tasmanian devil (Sarcophilus harrisii) induced pluripotent stem cells (DeviPSCs) from dermal fibroblasts by lentiviral delivery of human transcription factors. DeviPSCs display characteristic pluripotent stem cell colony morphology, with individual cells having a high nuclear-to-cytoplasmic ratio and alkaline phosphatase activity. DeviPSCs are leukemia inhibitory factor dependent and have reactivated endogenous octamer-binding transcription factor 4 [OCT4, POU domain, class 5, transcription factor 1 (POU5F1)], POU2 [POU domain, class 5, transcription factor 3 (POU5F3)], sex determining region Y-box 2 (SOX2), Nanog homeobox (NANOG) and dosage-sensitive sex reversal, adrenal hypoplasia congenita critical region on the X chromosome, gene 1 (DAX1) genes, retained a normal karyotype, and concurrently silenced exogenous human transgenes. Notably, co-expression of both OCT4 and POU2 suggests that they are representative of cells of the epiblast, the marsupial equivalent of the inner cell mass. DeviPSCs readily form embryoid bodies and in vitro teratomas containing derivatives of all three embryonic germ layers. To date, DeviPSCs have been stably maintained for more than 45 passages. Our DeviPSCs provide an invaluable resource for studies into marsupial pluripotency and development, and they may also serve as an important tool in efforts to combat the threat of devil facial tumor disease.

Impaired ability to grow colonies of endothelial stem cells could be the mechanism explaining the high cardiovascular morbidity and mortality of patients with depression.

Background Subjects with depression are more prone to develop cardiovascular complications. Severity of depression is associated with higher rates of cardiovascular mortality and morbidity. Several mechanisms were suggested including accelerated atherosclerosis, alteration of the cardiac autonomic response with a decrease in heart rate variability. There is evidence that circulating endothelial progenitor cells (EPCs) are decreased in patients with major depression. Our hypothesis was that patients with depression would have an impaired ability to build colonies of EPCs. Methods A prospective study enrolled twenty women with a diagnosis of major. All were not treated before for depression. Thirteen healthy age-matched women served as controls. All signed a consent form before recruitment to the study. Peripheral blood was drawn to build colonies of EPCs within 5 days. ELISA methods were used to measure levels of vascular cell adhesion molecule-1 (VCAM-1) and vascular endothelial growth factor (VEGF). Results Twenty female patients with depression were recruited. The mean age was 43 ± 14 years (vs. controls 41 ± 11 years, P = 0.682), patients' average CFU-EPCs was 7 ± 8 colonies per well (controls 31 ± 11, P = 0.0001), VCAM-1 level was 121.7 ± 3.0 ng/ml (controls 119.3 ± 3.1 pg/ml, P = 0.037), VEGF level was 6.4 ± 0.2 pg/ml (controls 5.2 ± 0.5 pg/ml, P = 0.0001). An inverse correlation was found between VEGF level and EPCs' colonies (r = -0.547, P < 0.001) and between age and CFU-EPCs (r = -0.576, P = 0.008). Conclusions We found that patients with major depression had high levels of VCAM-1 and VEGF. They also had a significant inhibition of EPCs' colonies. An inverse correlation was found between levels of VEGF and the ability to grow colonies of EPCs in culture.

Deep vector-based convolutional neural network approach for automatic recognition of colonies of induced pluripotent stem cells.

Pluripotent stem cells can potentially be used in clinical applications as a model for studying disease progress. This tracking of disease-causing events in cells requires constant assessment of the quality of stem cells. Existing approaches are inadequate for robust and automated differentiation of stem cell colonies. In this study, we developed a new model of vector–based convolutional neural network (V-CNN) with respect to extracted features of the induced pluripotent stem cell (iPSC) colony for distinguishing colony characteristics. A transfer function from the feature vectors to the virtual image was generated at the front of the CNN in order for classification of feature vectors of healthy and unhealthy colonies. The robustness of the proposed V-CNN model in distinguishing colonies was compared with that of the competitive support vector machine (SVM) classifier based on morphological, textural, and combined features. Additionally, five-fold cross-validation was used to investigate the performance of the V-CNN model. The precision, recall, and F-measure values of the V-CNN model were comparatively higher than those of the SVM classifier, with a range of 87–93%, indicating fewer false positives and false negative rates. Furthermore, for determining the quality of colonies, the V-CNN model showed higher accuracy values based on morphological (95.5%), textural (91.0%), and combined (93.2%) features than those estimated with the SVM classifier (86.7, 83.3, and 83.4%, respectively). Similarly, the accuracy of the feature sets using five-fold cross-validation was above 90% for the V-CNN model, whereas that yielded by the SVM model was in the range of 75–77%. We thus concluded that the proposed V-CNN model outperforms the conventional SVM classifier, which strongly suggests that it as a reliable framework for robust colony classification of iPSCs. It can also serve as a cost-effective quality recognition tool during culture and other experimental procedures.

Granulocyte-macrophage progenitor cells response to magnetite nanoparticles in a static magnetic field

The ability to modulate the life process of stem cells conjugated with MNPs via a magnetic field is promising as multifunctional tool in biomedicine. Experiment on mice demonstrates the high accumulation of iron ions measured via stripping voltammetry in the bone marrow in respect to bone, liver or kidney. Therefore, the potential cytotoxic effects of iron oxide magnetic nanoparticles (MNPs) on bone marrow cells become more predictable in comparison with the other examined cells. This study examined in vitro responses of mouse bone marrow-derived granulocyte–macrophage committed stem cells (colony-forming units of granulocytes and macrophages, CFU-GM) to a controlled amount of MNPs prepared by the exploding wire method; and to a moderate static magnetic field (SMF) of about 160 Oe. The moderate SMF did not affect CFU-GM capacity. Two different regimes of bone marrow cells cultivation with MNPs were investigated. Remarkably, adding MNPs to cells either decreased (1st cultivation regime, 6 pg MNPs per 1 cell) or enhanced (2nd regime, 20 pg MNPs per 1 cell) the colony-forming activity of CFU-GM, depending on the regimes of cultivation. The action of a moderate SMF on cells cultivated with MNPs inverts their effects on stem cell colony formation. Possible mechanisms of these phenomena are discussed. The main pathway was explained by a change in the presence of iron, either inside or outside of the cell, following the formation of free radicals susceptible to SMFs. Nanoscale magnetite activity within the pool of bipotent hemopoietic stem cells supports its proposed usage as supplement in cell technologies, theranostics, bone marrow repair and regenerative medicine. Further study is necessary to examine the intra- and intercellular mechanisms of the SMF’s modulating effect on stem cell activity caused by magnetite MNPs.

A Machine Learning Assisted, Label-free, Non-invasive Approach for Somatic Reprogramming in Induced Pluripotent Stem Cell Colony Formation Detection and Prediction

During cellular reprogramming, the mesenchymal-to-epithelial transition is accompanied by changes in morphology, which occur prior to iPSC colony formation. The current approach for detecting morphological changes associated with reprogramming purely relies on human experiences, which involve intensive amounts of upfront training, human error with limited quality control and batch-to-batch variations. Here, we report a time-lapse-based bright-field imaging analysis system that allows us to implement a label-free, non-invasive approach to measure morphological dynamics. To automatically analyse and determine iPSC colony formation, a machine learning-based classification, segmentation, and statistical modelling system was developed to guide colony selection. The system can detect and monitor the earliest cellular texture changes after the induction of reprogramming in human somatic cells on day 7 from the 20–24 day process. Moreover, after determining the reprogramming process and iPSC colony formation quantitatively, a mathematical model was developed to statistically predict the best iPSC selection phase independent of any other resources. All the computational detection and prediction experiments were evaluated using a validation dataset, and biological verification was performed. These algorithm-detected colonies show no significant differences (Pearson Coefficient) in terms of their biological features compared to the manually processed colonies using standard molecular approaches.

Physical-chemical mechanisms of pattern formation during gastrulation.

Gastrulation is a fundamental phase during the biological development of most animals when a single layer of identical embryo cells is transformed into a three-layer structure, from which the organs start to develop. Despite a remarkable progress in quantifying the gastrulation processes, molecular mechanisms of these processes remain not well understood. Here we theoretically investigate early spatial patterning in a geometrically confined colony of embryonic stem cells. Using a reaction-diffusion model, a role of Bone-Morphogenetic Protein 4 (BMP4) signaling pathway in gastrulation is specifically analyzed. Our results show that for slow diffusion rates of BMP4 molecules, a new length scale appears, which is independent of the size of the system. This length scale separates the central region of the colony with uniform low concentrations of BMP molecules from the region near the colony edge where the concentration of signaling molecules is elevated. The roles of different components of the signaling pathway are also explained. Theoretical results are consistent with recent in vitro experiments, providing microscopic explanations for some features of early embryonic spatial patterning. Physical-chemical mechanisms of these processes are discussed.

Surface Topography Guides Morphology and Spatial Patterning of Induced Pluripotent Stem Cell Colonies.

SummaryThe relevance of topographic cues for commitment of induced pluripotent stem cells (iPSCs) is largely unknown. In this study, we demonstrate that groove-ridge structures with a periodicity in the submicrometer range induce elongation of iPSC colonies, guide the orientation of apical actin fibers, and direct the polarity of cell division. Elongation of iPSC colonies impacts also on their intrinsic molecular patterning, which seems to be orchestrated from the rim of the colonies. BMP4-induced differentiation is enhanced in elongated colonies, and the submicron grooves impact on the spatial modulation of YAP activity upon induction with this morphogen. Interestingly, TAZ, a YAP paralog, shows distinct cytoskeletal localization in iPSCs. These findings demonstrate that topography can guide orientation and organization of iPSC colonies, which may affect the interaction between mechanosensors and mechanotransducers in iPSCs.