Cell Adhesion and Cellular Patterning on a Self‐Assembled Monolayer of Zeolite L Crystals

Chemically functionalized self‐assembled monolayers made by disk‐shaped zeolite L nanocrystals are used as models for biocompatible surfaces to study cell‐adhesion behavior. Different chemical groups lead to different cellular behavior and fluorescent‐molecule‐loaded zeolites allow the position of the cells to be determined. Furthermore, a patterned monolayer of asymmetrically functionalized zeolite L obtained by microcontact chemistry is used to grow cells. A spatial recognition of the cells, which proliferate only on the bioactive‐molecule‐functionalized stripes, is possible.     

MRI‐Visible siRNA Nanomedicine Directing Neuronal Differentiation of Neural Stem Cells in Stroke

A major challenge in stroke treatment is the restoration of neural circuit in which neuron function plays a central role. Although transplantation of exogenous neural stem cells (NSCs) is admittedly a promising therapeutical means, the treatment outcome is greatly affected due to the poor NSCs differentiation into neurons caused by myelin associated inhibitory factors binding to Nogo‐66 receptor (NgR). Herein, a nanoscale polymersome is developed to codeliver superparamagnetic iron oxide nanoparticles and siRNA targeting NgR gene (siNgR) into NSCs. This multifunctional nanomedicine directs neuronal differentiation of NSCs through silencing the NgR gene and meanwhile allows a noninvasive monitoring of NSC migration with magnetic resonance imaging. An improved recovery of neural function is achieved in rat ischemic stroke model. The results demonstrate the great potential of the multifunctional siRNA nanomedicine in stroke treatment based on stem cell transplantation.     

Dual Roles of Graphene Oxide in Chondrogenic Differentiation of Adult Stem Cells: Cell‐Adhesion Substrate and Growth Factor‐Delivery Carrier

Here, it is shown that graphene oxide (GO) can be utilized as both a cell‐adhesion substrate and a growth factor protein‐delivery carrier for the chondrogenic differentiation of adult stem cells. Conventionally, chondrogenic differentiation of stem cells is achieved by culturing cells in pellets and adding the protein transforming growth factor‐β3 (TGF‐β3), a chondrogenic factor, to the culture medium. However, pellets mainly provide cell‐cell interaction and diffusional limitation of TGF‐β3 may occur inside the pellet both of these factors may limit the chondrogenic differentiation of stem cells. In this study, GO sheets (size = 0.5–1 μm) were utilized to adsorb fibronectin (FN, a cell‐adhesion protein) and TGF‐β3 and were then incorporated in pellets of human adipose‐derived stem cells (hASCs). The hybrid pellets of hASC‐GO enhanced the chondrogenic differentiation of hASCs by adding the cell‐FN interaction and supplying TGF‐β3 effectively. This method may provide a new platform for stem cell culture for regenerative medicine.     

High‐Throughput Differentiation of Embryonic Stem Cells into Cardiomyocytes with a Microfabricated Magnetic Pattern and Cyclic Stimulation

Pluripotent stem cells are central tools to many regenerative medicine strategies due to their ability to differentiate toward the three embryonic germ layers. One challenge remains in providing control over their differentiation into specific lineages, such as cardiac commitment. Here, the possibility of directing cardiomyogenesis of embryonic stem cells using a microfabricated magnetic pattern is demonstrated. The stem cells are labeled with magnetic nanoparticles, aggregated into embryoid bodies (EBs) onto the pattern, and stimulated with a local magnetic force applied via the pattern. The EBs formed on this magnetic device experience the same differentiation profile as the ones created by the common hanging drop approach, while it allows high‐throughput production of hundreds of EBs. Further on/off cyclic magnetic force stimulation mediated by the same device is sufficient to enhance cardiomyogenesis in a way that almost all EBs develop spontaneous beating, confirmed by the overexpression of α‐actin and troponin proteins, and by the upregulation (twofold to fivefold) of genes involved in mesoderm differentiation (Nkx2.5, Gata4, and Gata6), and more specifically cardiac lineage (Tnnt2, Myh6, and Myl‐2). Beyond holding high application‐level potential, this work confirms that physical forces, and specifically on/off dynamic ones can be sufficient to govern cell function.     

2D and 3D Hybrid Systems for Enhancement of Chondrogenic Differentiation of Tonsil‐Derived Mesenchymal Stem Cells

2D/3D hybrid cell culture systems are constructed by increasing the temperature of the thermogelling poly(ethylene glycol)‐poly(l ‐alanine) diblock copolymer (PEG‐l ‐PA) aqueous solution in which tonsil tissue‐derived mesenchymal stem cells and graphene oxide (GO) or reduced graphene oxide (rGO) are suspended, to 37 °C. The cells exhibit spherical cell morphologies in 2D/3D hybrid culture systems of GO/PEG‐l ‐PA and rGO/PEG‐l ‐PA by using the growth medium. The cell proliferations are 30%–50% higher in the rGO/PEG‐l ‐PA hybrid system than in the GO/PEG‐l ‐PA hybrid system. When chondrogenic culture media enriched with TGF‐β3 is used in the 2D/3D hybrid systems, cells extensively aggregate, and the expression of chondrogenic biomarkers of SOX 9, COL II A1, COL II, and COL X significantly increases in the GO/PEG‐l ‐PA 2D/3D hybrid system as compared with the PEG‐l ‐PA 3D systems and rGO/PEG‐l ‐PA 2D/3D hybrid system, suggesting that the GO/PEG‐l ‐PA 2D/3D hybrid system can be an excellent candidate as a chondrogenic differentiation platform of the stem cell. This paper also suggests that a 2D/3D hybrid system prepared by incorporating 2D materials with various surface biofunctionalities in the in situ forming 3D hydrogel matrix can be a new cell culture system.     

Self‐Assembled Injectable Nanocomposite Hydrogels Stabilized by Bisphosphonate‐Magnesium (Mg2+) Coordination Regulates the Differentiation of Encapsulated Stem Cells via Dual Crosslinking

Nanocomposite hydrogels consist of a polymer matrix embedded with nanoparticles (NPs), which provide the hydrogels with unique bioactivities and mechanical properties. Incorporation of NPs via in situ precipitation in the polymer matrix further enhances these desirable hydrogel properties. However, the noncytocompatible pH, osmolality, and lengthy duration typically required for such in situ precipitation strategies preclude cell encapsulation in the resultant hydrogels. Bisphosphonate (BP) exhibits a variety of specific bioactivities and excellent binding affinity to multivalent cations such as magnesium ions (Mg²⁺). Here, the preparation of nanocomposite hydrogels via self‐assembly driven by bisphosphonate‐Mg²⁺ coordination is described. Upon mixing solutions of polymer bearing BPs, BP monomer (Ac‐BP), and Mg²⁺, this effective and dynamic coordination leads to the rapid self‐assembly of Ac‐BP‐Mg NPs which function as multivalent crosslinkers stabilize the resultant hydrogel structure at physiological pH. The obtained nanocomposite hydrogels are self‐healing and exhibit improved mechanical properties compared to hydrogels prepared by blending prefabricated NPs. Importantly, the hydrogels in this study allow the encapsulation of cells and subsequent injection without compromising the viability of seeded cells. Furthermore, the acrylate groups on the surface of Ac‐BP‐Mg NPs enable facile temporal control over the stiffness and crosslinking density of hydrogels via UV‐induced secondary crosslinking, and it is found that the delayed introduction of this secondary crosslinking enhances cell spreading and osteogenesis.     

Nanocarrier‐Mediated Codelivery of Small Molecular Drugs and siRNA to Enhance Chondrogenic Differentiation and Suppress Hypertrophy of Human Mesenchymal Stem Cells

Cartilage loss is a leading cause of disability among adults, and effective therapy remains elusive. Human mesenchymal stem cells (hMSCs), which have demonstrated self‐renewal and multipotential differentiation, are a promising cell source for cartilage repair. However, the hypertrophic differentiation of the chondrogenically induced MSCs and resulting tissue calcification hinders the clinical translation of MSCs for cartilage repair. Here, a multifunctional nanocarrier based on quantum dots (QDs) is developed to enhance chondrogenic differentiation and suppress hypertrophy of hMSCs simultaneously. Briefly, the QDs are modified with β‐cyclodextrin (β‐CD) and RGD peptide. The resulting nanocarrier is capable of carrying hydrophobic small molecules such as kartogenin in the hydrophobic pockets of conjugated β‐CD to induce chondrogenic differentiation of hMSCs. Meanwhile, via electrostatic interaction the conjugated RGD peptides bind the cargo siRNA targeting Runx2, which is a key regulator of hMSC hypertrophy. Furthermore, due to the excellent photostability of QDs, hMSCs labeled with the nanocarrier can be tracked for up to 14 d after implantation in nude mice. Overall, this work demonstrates the potential of our nanocarrier for inducing and maintaining the chondrogenic phenotype and tracking hMSCs in vivo.     

Black‐Phosphorus‐Incorporated Hydrogel as a Conductive and Biodegradable Platform for Enhancement of the Neural Differentiation of Mesenchymal Stem Cells

Conductive hydrogel scaffolds have important applications for electroactive tissue repairs. However, the development of conductive hydrogel scaffolds tends to incorporate nonbiodegradable conductive nanomaterials that will remain in the human body as foreign matters. Herein, a biodegradable conductive hybrid hydrogel is demonstrated based on the integration of black phosphorus (BP) nanosheets into the hydrogel matrix. To address the challenge of applying BP nanosheets in tissue engineering due to its intrinsic instability, a polydopamine (PDA) modification method is developed to improve the stability. Moreover, PDA modification also enhances interfacial bonding between pristine BP nanosheets and the hydrogel matrix. The incorporation of polydopamine‐modified black phosphorous (BP@PDA) nanosheets into the gelatin methacryloyl (GelMA) hydrogels significantly enhances the electrical conductivity of the hydrogels and improves the cell migration of mesenchymal stem cells (MSCs) within the 3D scaffolds. On the basis of the gene expression and protein level assessments, the BP@PDA‐incorporated GelMA scaffold can significantly promote the differentiation of MSCs into neural‐like cells under the synergistic electrical stimulation. This strategy of integrating biodegradable conductive BP nanomaterials within a biocompatible hydrogel provides a new insight into the design of biomaterials for broad applications in tissue engineering of electroactive tissues, such as neural, cardiac, and skeletal muscle tissues.     

An NIR‐II Fluorescence/Dual Bioluminescence Multiplexed Imaging for In Vivo Visualizing the Location,Survival, and Differentiation of Transplanted Stem Cells

The in vivo distribution, viability, and differentiation capability of transplanted stem cells are vital for the therapeutic efficacy of stem cell–based therapy. Herein, an NIR‐II fluorescence/dual bioluminescence multiplexed imaging method covering the visible and the second near‐infrared window from 400 to 1700 nm is successfully developed for in vivo monitoring the location, survival, and osteogenic differentiation of transplanted human mesenchymal stem cells (hMSCs) in a calvarial defect mouse model. The exogenous Ag₂S quantum dot–based fluorescence imaging in the second near‐infrared window is applied for visualizing the long‐term biodistribution of transplanted hMSCs. Endogenous red firefly luciferase (RFLuc)‐based bioluminescence imaging (BLI) and the collagen type 1 promoter–driven Gaussia luciferase (GLuc)‐based BLI are employed to report the survival and osteogenic differentiation statuses of the transplanted hMSCs. Meanwhile, by integrating the three imaging channels, multiple dynamic biological behaviors of transplanted hMSCs and the promotion effects of immunosuppression and the bone morphogenetic protein 2 on the survival and osteogenic differentiation of transplanted hMSCs are directly observed. The novel multiplexed imaging method can greatly expand the capability for multifunctional analysis of the fates and therapeutic capabilities of the transplanted stem cells, and aid in the improvement of stem cell–based regeneration therapies and their clinical translation.     

Multivalent Aptamer Functionalized Ag2S Nanodots/Hybrid Cell Membrane‐Coated Magnetic Nanobioprobe for the Ultrasensitive Isolation and Detection of Circulating Tumor Cells

Circulating tumor cells (CTCs) play key roles in the development of tumor metastasis. It remains a significant challenge to capture and detect CTCs with high purity and sensitivity from blood samples. Herein, a nanoplatform is developed for the efficient isolation and ultrasensitive detection of CTCs by combining near‐infrared (NIR) multivalent aptamer functionalized Ag₂S nanodots with hybrid cell membrane‐coated magnetic nanoparticles. Multivalent aptamer functionalized Ag₂S nanodots are synthesized using a one‐pot method under mild conditions (60 °C). White blood cell and tumor cell membranes are fused as the hybrid membrane and coated with magnetic nanoparticles, which are further modified with streptavidin (SA). Through the specific interaction of SA‐biotin, the multivalent aptamer‐Ag₂S nanodots are grafted with hybrid cell membrane‐magnetic nanoparticles. Due to the features of hybrid cell membrane modification, multivalent aptamer functionalization, magnetic separation, and NIR fluorescence measurements, the nanoplatform shows sensitive recognition, efficient capture, easy isolation, and sensitive detection of CTCs due to its great enhancement in anti‐interference from background and improvement on binding ability toward CTCs. The capture efficiency and purity for CTCs is as high as 97.63% and 96.96%, respectively. Furthermore, the nanoplatform is successfully applied to the detection of CTCs in blood samples.     

MRI/SPECT/Fluorescent Tri‐Modal Probe for Evaluating the Homing and Therapeutic Efficacy of Transplanted Mesenchymal Stem Cells in a Rat Ischemic Stroke Model

Quantitatively tracking engraftment of intracerebrally or intravenously transplanted stem cells and evaluating their concomitant therapeutic efficacy for stroke has been a challenge in the field of stem cell therapy. In this study, first, an MRI/SPECT/fluorescent tri‐modal probe (¹²⁵I‐fSiO4@SPIOs) is synthesized for quantitatively tracking mesenchymal stem cells (MSCs) transplanted intracerebrally or intravenously into stroke rats, and then the therapeutic efficacy of MSCs delivered by both routes and the possible mechanism of the therapy are evaluated. It is demonstrated that ⁽¹²⁵⁾I‐fSiO4@SPIOs have high efficiency for labeling MSCs without affecting their viability, differentiation, and proliferation capacity , and found that 35% of intracerebrally injected MSCs migrate along the corpus callosum to the lesion area, while 90% of intravenously injected MSCs remain trapped in the lung at 14 days after MSC transplantation. However, neurobehavioral outcomes are significantly improved in both transplantation groups, which are accompanied by increases of vascular endothelial growth factor, basic fibroblast growth factor, and tissue inhibitor of metalloproteinases‐3 in blood, lung, and brain tissue (p < 0.05). The study demonstrates that ¹²⁵I‐fSiO4@SPIOs are robust probe for long‐term tracking of MSCs in the treatment of ischemic brain and MSCs delivered via both routes improve neurobehavioral outcomes in ischemic rats.