Comparison of mesenchymal stem cells derived from arterial, venous, and Wharton’s jelly explants of human umbilical cord

We isolated mesenchymal stem cells (MSC) from arteries (UCA), veins (UCV), and Wharton’s jelly (UCWJ) of human umbilical cords (UC) and determined their relative capacities for sustained proliferation and multilineage differentiation. Individual UC components were dissected, diced into 1–2 mm3 fragments, and aligned in explant cultures from which migrating cells were isolated using trypsinization. Preparations from 13 UCs produced 13 UCWJ, 11 UCV, and 10 UCA cultures of fibroblast-like, spindle-shaped cells negative for CD31, CD34, CD45, CD271, and HLA-class II, but positive for CD13, CD29, CD44, CD73, CD90, CD105, and HLA-class I. UCV cells exhibited a significantly higher frequency of colony-forming units fibroblasts than did UCWJ and UCA cells. Individual MSCs could be selectively differentiated into osteoblasts, chondrocytes, and adipocytes. When compared for osteogenic potential, UCWJ cells were the least effective precursors, whereas UCA-derived cells developed alkaline phosphatase activity with or without an osteogenic stimulus. UC components, especially blood vessels, could provide a promising source of MSCs with important clinical applications.     

Safety and effect of umbilical cord blood –derived mesenchymal stem cells on apoptosis of human cardiomyocytes

Objective To study the safety and effect of the umbilical cord blood （UCB）-derived mesenchymal stem cells （MSCs） on apoptosis of human cardiomyocytes （HCM）. Methods UCB was collected at the time of delivery with informed consent obtained from 10 donors. The UCB-derived MSCs were treated with 5-azaserube （5-AZA） and were further induced to differentiate into cardiomyocytes. Telomerase activity, G-banding patterns of chromosomal karyotypes, tumor formation in nude mice, RT-PCR, and the effect of inhibiting apoptosis of HCM were investigated. Results MSCs derived from UCB were differentiated into cardiomyocytes in vitro, which possessed telomerase activity after 5-AZA induction, and no abnormal chromosomal karyotypes were observed. Expression of p53, cyclin A, cdk2, ~3 -actin, C-fos, h-TERT and c-myc were similar in MSCs before and after 5-AZA treatment. There was no tumor formation in nude mice after injection of UCB-derived MSCs. UCB-derived MSCs significantly inhibited apoptosis of HCM. Conclusion UCB-derived MSCs are a valuable, safe and effective source of cell-transplantation treatment .     

Hepatic differentiation capability of rat bone marrow-derived mesenchymal stem cells and hematopoietic stem cells

AIM:To investigate the different effects of mesenchymalstem cells(MSCs)and hematopoietic stem cells(HSCs)onhepatic differentiation.METHODS:MSCs from rat bone marrow were isolated andcultured by standard methods.HSCs from rat bone marrowwere isolated and purified by magnetic activated cell sorting.Both cell subsets were induced.Morphology,RT-PCR andimmunocytochemistry were used to identify the hepaticdifferentiation grade.RESULTS:MSCs exhibited round in shape after differentiation,instead of fibroblast-like morphology before differentiation.Albumin mRNA and protein were expressed positively in MSCs,without detection of alpha-fetoprotein(AFP).HSCs werepolygonal in shape after differentiation.The expression ofalbumin signal decreased and AFP signal increased.Theexpression of CK18 was continuous in MSCs and HSCs bothbefore and after induction.CONCLUSION:Both MSCs and HSCs have hepatic differentiationcapabilities.However,their capabilities are not the same.MSCs can differentiate into mature hepatocyte-like cells,never expressing early hepatic specific genes,while Thy-1.1~ cells are inclined to differentiate into hepatic stem cell-likecells,with an increasing AFP expression and a decreasingalbumin signal.CK18 mRNA is positive in Thy-1.1~ cellsand MSCs,negative in Thy-1.1 cells.It seems that CK18has some relationship with Thy-1.1 antigen,and CK18 maybe a predictive marker of hepatic differentiation capability.     

HNF-4α determines hepatic differentiation of human mesenchymal stem cells from bone marrow

AIM: To investigate the differentiation status and key factors to facilitate hepatic differentiation of human bone-marrow-derived mesenchymal stem cells (MSCs). METHODS: Human MSCs derived from bone marrow were induced into hepatocyte-like cells following a previously published protocol. The differentiation status of the hepatocyte-like cells was compared with various human hepatoma cell lines. Overexpression of hepatocyte nuclear factor (HNF)-4α was mediated by adenovirus infection of these hepatocyte-like...     

Deficient primary cilia in obese adipose‐derived mesenchymal stem cells: obesity,a secondary ciliopathy?

Obesity alters the composition, structure and function of adipose tissue, characterized by chronic inflammation, insulin resistance and metabolic dysfunction. Adipose‐derived mesenchymal stem cells (ASCs) are responsible for cell renewal, spontaneous repair and immunomodulation in adipose tissue. Increasing evidence highlights that ASCs are deficient in obesity, and the underlying mechanisms are not well understood. We have recently shown that obese ASCs have defective primary cilia, which are shortened and unable to properly respond to stimuli. Impaired cilia compromise ASC functions. This work suggests an intertwined connection of obesity, defective cilia and dysfunctional ASCs. We have here discussed the current data regarding defective cilia in various cell types in obesity. Based on these observations, we hypothesize that obesity, a systemic chronic metainflammation, could impair cilia in diverse ciliated cells, like pancreatic islet cells, stem cells and hypothalamic neurons, making these critical cells dysfunctional by shutting down their signal sensors and transducers. In this context, obesity may represent a secondary form of ciliopathy induced by obesity‐related inflammation and metabolic dysfunction. Reactivation of ciliated cells might be an alternative strategy to combat obesity and its associated diseases.     

Small-molecule–directed,efficient generation of retinal pigment epithelium from human pluripotent stem cells

Age-related macular degeneration (AMD) is associated with dysfunction and death of retinal pigment epithelial (RPE) cells. Cell-based approaches using RPE-like cells derived from human pluripotent stem cells (hPSCs) are being developed for AMD treatment. However, most efficient RPE differentiation protocols rely on complex, stepwise treatments and addition of growth factors, whereas small-molecule–only approaches developed to date display reduced yields. To identify new compounds that promote RPE differentiation, we developed and performed a high-throughput quantitative PCR screen complemented by a novel orthogonal human induced pluripotent stem cell (hiPSC)-based RPE reporter assay. Chetomin, an inhibitor of hypoxia-inducible factors, was found to strongly increase RPE differentiation; combination with nicotinamide resulted in conversion of over one-half of the differentiating cells into RPE. Single passage of the whole culture yielded a highly pure hPSC-RPE cell population that displayed many of the morphological, molecular, and functional characteristics of native RPE.Age-related macular degeneration (AMD) is the leading cause of irreversible vision loss and blindness among the elderly in industrialized countries. Dysfunction of the retinal pigment epithelium (RPE) is an early event associated with AMD. The RPE, a monolayer of pigmented cells directly abutting the photoreceptor cell layer, plays many important roles in vision and in maintaining the health and integrity of the retina (1). As the RPE deteriorates, there is progressive degeneration of photoreceptor cells.Successful antiangiogenesis treatments have been developed for the neovascular, or “wet,” form of AMD. However, there are no Food and Drug Administration-approved treatment options available for the majority of AMD patients, who suffer from the more common nonneovascular, or “dry,” form of the disease. In the past few years, however, transplantation of human pluripotent stem cell-derived RPE (hPSC-RPE) has emerged as a promising new therapy for dry AMD. A Phase I clinical trial of human embryonic stem (hES)-derived RPE cells recently reported some preliminary encouraging results (2). Additionally, a Phase I trial that will use RPE cells generated from human induced pluripotent stem cells (hiPSCs) reprogrammed from the patients’ own skin cells recently injected their first patient (3).If the promise of hiPSC-based approaches for AMD is to be translated into the clinic, each patient would require individualized generation of RPE cells from his or her stem cells, thereby necessitating the development of simple, efficient, safe, and affordable protocols for RPE generation. Although highly efficient protocols have been established, they rely upon mixtures of growth factors (4–6) with the use of complex biologics derived from animal cells or bacteria, presenting potential clinical challenges. As an alternative approach, use of nonbiological products such as small molecules would limit the risk of infection or immune rejection (7). In addition, they offer a cost-effective alternative with less lot-to-lot variability. As a consequence, the development of small-molecule–based protocols has been the focus of extensive research (8). To date, however, the small-molecule–only protocols that have been developed for RPE differentiation have demonstrated only limited efficiency (9).We report here development of a highly efficient, small-molecule–based protocol for the production of RPE from hPSC that is suitable for clinical application. We performed a quantitative real-time PCR (qPCR)-based high-throughput screen (HTS) for molecules that promote RPE differentiation, and complemented the screen with a novel orthogonal hiPSC-based RPE reporter assay for compound validation. Using this strategy, we identified chetomin (CTM) as a potent promoter of RPE differentiation. Its use in combination with a previously known neural inducer, nicotinamide (NIC) (10, 11), provides a one-step treatment for differentiation of a wide range of hPSC lines. Following a single whole-dish passage, a pure and functional monolayer of RPE cells is obtained. This protocol should prove useful for the cost-efficient production of RPE cells.     

Over expression of HIF-1α in human mesenchymal stem cells increases their supportive functions for hematopoietic stem cells in an experimental co-culture model

Introduction

Bone marrow transplantation is a critical approach for the treatment of many hematological disorders. Success of this approach is dependent on many factors the most important of which is the number of hematopoietic stem cells along with an efficient stroma. Co-transplantation of efficient mesenchymal stem cells can greatly improve the outcome of transplantations. Current researches assign a critical role for hypoxia inducible factor (HIF)-1α in protection of various cells and tissues probably through induction of cytokines. To make this feature applicable to human bone marrow-derived mesenchymal stem cells, we manipulated these cells to over express HIF-1α gene.

Materials and methods

Full-length cDNA of human HIF-1α was inserted into human bone marrow mesenchymal stem cells by pcDNA.3.1 non-viral plasmid vector, and the effect of this over expression on production of some hematopoietic growth factors was explored. Moreover, using a co-culture system, the interactive impact of HIF-1α-overexpressed mesenchymal stem cells on hematopoietic stem cells was evaluated.

Results

Over expression of HIF-1α in mesenchymal stem cells in normoxia increased production of one of the most important hematopoietic growth factors, Stem cell factor (also known as Steel factor or c-kit ligand). HIF-1α overexpression had no effect on production of other hematopoietic growth factors. In co-culture of mesenchymal stem cells-HIF-1α with hematopoietic stem cells, enhancement of colony formation and reduced differentiation of hematopoietic stem cells were observed.

Conclusion

Over expression of HIF-1α in human bone marrow-derived mesenchymal stem cells can augment the production of some hematopoietic growth factors, and we suggest this response of mesenchymal stem cells could help to improve the outcome of bone marrow transplantation.     

Differentiation of rat marrow mesenchymal stem cells into pancreatic islet beta-cells

AIM:To explore the possibility of marrow mesenchymalstem cells(MSC)in vitro differentiating into functional islet-like cells and to test the diabetes therapeutic potency ofIslet-like cells.METHODS:Rat MSCs were isolated from Wistar rats andcultured.Passaged MSCs were induced to differentiate intoislet-like cells under following conditions:pre-induction withL-DMEM including 10 mmol/L nicotinamide l mmol/Lβ-mercaptoethanol 200 mL/L fetal calf serum(FSC)for 24 h,followed by induction with serum free H-DMEM solution including10 mmol/L nicotinamide l mmol/L,β-mercaptoethanol for10 h.Differentiated cells were observed under inversemicroscopy,insulin and nestin expressed in differentiatedcells were detected with immunocytochemistry.Insulinexcreted from differentiated cells was tested withradioimmunoassay.Rat diabetic models were made to testin vivo function of differentiated MSCs.RESULTS:Typical islet-like clustered cells were observed.Insulin mRNA and protein expressions were positive indifferentiated cells,and nestin could be detected in pre-differentiated cells.Insulin excreted from differentiatedMSCs(446.93±102.28 IU/L)was much higher than thatfrom pre-differentiated MSCs(2.45±0.81 IU/L(P<0.01).Injected differentiated MSCs cells could down-regulateglucose level in diabetic rats.CONCLUSION:Islet-like functional cells can be differentiatedfrom marrow mesenchymal stem cells,which may be anew procedure for clinical diabetes stem-cell therapy,thesecells can control blood glucose level in diabetic rats.MSCsmay play an important role in diabetes therapy by isletdifferentiation and transplantation.     

Functional analysis reveals angiogenic potential of human mesenchymal stem cells from Wharton’s jelly in dermal regeneration

Disorders in skin wound healing are a major health problem that requires the development of innovative treatments. The use of biomaterials as an alternative of skin replacement has become relevant, but its use is still limited due to poor vascularization inside the scaffolds, resulting in insufficient oxygen and growth factors at the wound site. In this study, we have developed a cell-based wound therapy consisting of the application of collagen-based dermal scaffolds containing mesenchymal stem cells from Wharton’s jelly (WJ-MSC) in an immunocompetent mouse model of angiogenesis. From our comparative study on the secretion profile between WJ-MSC and adipose tissue-derived MSC, we found a stronger expression of several well-characterized growth factors, such as VEGF-A, angiopoietin-1 and aFGF, which are directly linked to angiogenesis, in the culture supernatant of WJ-MSC, both on monolayer and 3D culture conditions. WJ-MSC proved to be angiogenic both in vitro and in vivo, through tubule formation and CAM assays, respectively. Moreover, WJ-MSC consistently improved the healing response in vivo in a mouse model of human-like dermal repair, by triggering angiogenesis and further providing a suitable matrix for wound repair, without altering the inflammatory response in the animals. Since these cells can be easily isolated, cultured with high expansion rates and cryopreserved, they represent an attractive stem cell source for their use in allogeneic cell transplant and tissue engineering.     

Derivation of na?ve human embryonic stem cells

The naïve pluripotent state has been shown in mice to lead to broad and more robust developmental potential relative to primed mouse epiblast cells. The human naïve ES cell state has eluded derivation without the use of transgenes, and forced expression of OCT4, KLF4, and KLF2 allows maintenance of human cells in a naïve state [Hanna J, et al. (2010) Proc Natl Acad Sci USA 107(20):9222–9227]. We describe two routes to generate nontransgenic naïve human ES cells (hESCs). The first is by reverse toggling of preexisting primed hESC lines by preculture in the histone deacetylase inhibitors butyrate and suberoylanilide hydroxamic acid, followed by culture in MEK/ERK and GSK3 inhibitors (2i) with FGF2. The second route is by direct derivation from a human embryo in 2i with FGF2. We show that human naïve cells meet mouse criteria for the naïve state by growth characteristics, antibody labeling profile, gene expression, X-inactivation profile, mitochondrial morphology, microRNA profile and development in the context of teratomas. hESCs can exist in a naïve state without the need for transgenes. Direct derivation is an elusive, but attainable, process, leading to cells at the earliest stage of in vitro pluripotency described for humans. Reverse toggling of primed cells to naïve is efficient and reproducible.It has become clear with the derivation of mouse epiblast stem cells (mEpiSCs) that pluripotency encompasses more than one stage of development (1, 2). The earlier “naïve” stage represents the preimplantation inner cell mass, typified by mouse ES cells (mESCs), and the “primed,” the postimplantation epiblast, typified by mEpiSCs and human ES cells (hESCs). The challenge in naïve cell maintenance has been protecting cells from differentiation stimuli. This has been achieved in mESCs through exposure to leukemia inhibitory factor (LIF), whereas addition of extracellular signal-regulated kinase (MEK) and glycogen synthase kinase 3 (GSK3) inhibitors (2i) in defined medium allows the cells to attain a homogeneous ground state (3). Defining characteristics of the naïve/ground vs. primed states are shown in Fig. S1A. In humans, the naïve stage has been difficult to capture as a stable in vitro state.There are practical advantages that come with a human naïve state. Among them is ease of trypsin passage and developmental capacity. Whole animals can be generated from good naïve mESCs through tetraploid complementation (4), and mEpiSCs cannot contribute to chimerism. Being more comparable to mESCs, naïve hESCs will likely allow increased developmental potential and a more accurate correlation to mESC data.It has been reported that human induced pluripotent cells (h-iPSCs) can be maintained in the naïve state if the pluripotency-inducing transgenes are not silenced (5). Only recently have hESCs been maintained in a naïve state without transgenes (6). Our primary aim was to generate naïve hESCs not dependent upon transgenes for stable culture. We toggled existing human ESC and mouse mEpiSC lines back from the primed state to grow under the influence of 2i without the need for Activin A. This helped us to define appropriate culture conditions for human naïve cells and allowed the de novo derivation of a naïve hESC line, Elf1. We report on the naïve state of human ESCs capable of unlimited culture in 2i.     

Generation of functional hepatocyte-like cells from human bone marrow mesenchymal stem cells by overexpression of transcription factor HNF4α and FOXA2

Background: Our previous study showed that overexpression of hepatocyte nuclear factor 4α(HNF4α) could directly promote mesenchymal stem cells(MSCs) to differentiate into hepatocyte-like cells. However, the efficiency of hepatic differentiation remains low. The purpose of our study was to establish an MSC cell line that overexpressed HNF4α and FOXA2 genes to obtain an increased hepatic differentiation efficiency and hepatocyte-like cells with more mature hepatocyte functions. Methods: Successful establishment of high-level HNF4α and FOXA2 co-overexpression in human induced hepatocyte-like cells(hi Hep cells) was verified by flow cytometry, immunofluorescence and RT-PCR. Measurements of albumin(ALB), urea, glucose, indocyanine green(ICG) uptake and release, cytochrome P450(CYP) activity and gene expression were used to analyze mature hepatic functions of hi Hep cells. Results: hi Hep cells efficiently express HNF4α and FOXA2 genes and proteins, exhibit typical epithelial morphology and acquire mature hepatocyte-like cell functions, including ALB secretion, urea production, ICG uptake and release, and glycogen storage. hi Hep cells can be activated by CYP inducers. The percentage of both ALB and α-1-antitrypsin(AAT)-positive cells was approximately 72.6%. The expression levels of hepatocyte-specific genes( ALB, AAT, and CYP1A1) and liver drug transport-related genes( ABCB1, ABCG2, and SLC22A18) in hi Hep cells were significantly higher than those in MSCs-Vector cells. The hi Hep cells did not form tumors after subcutaneous xenograft in BALB/c nude mice after 2 months. Conclusion: This study provides an accessible, feasible and efficient strategy to generate hi Hep cells from MSCs.     

Measuring dynamic cell–material interactions and remodeling during 3D human mesenchymal stem cell migration in hydrogels

Biomaterials that mimic aspects of the extracellular matrix by presenting a 3D microenvironment that cells can locally degrade and remodel are finding increased applications as wound-healing matrices, tissue engineering scaffolds, and even substrates for stem cell expansion. In vivo, cells do not simply reside in a static microenvironment, but instead, they dynamically reengineer their surroundings. For example, cells secrete proteases that degrade extracellular components, attach to the matrix through adhesive sites, and can exert traction forces on the local matrix, causing its spatial reorganization. Although biomaterials scaffolds provide initially well-defined microenvironments for 3D culture of cells, less is known about the changes that occur over time, especially local matrix remodeling that can play an integral role in directing cell behavior. Here, we use microrheology as a quantitative tool to characterize dynamic cellular remodeling of peptide-functionalized poly(ethylene glycol) (PEG) hydrogels that degrade in response to cell-secreted matrix metalloproteinases (MMPs). This technique allows measurement of spatial changes in material properties during migration of encapsulated cells and has a sensitivity that identifies regions where cells simply adhere to the matrix, as well as the extent of local cell remodeling of the material through MMP-mediated degradation. Collectively, these microrheological measurements provide insight into microscopic, cellular manipulation of the pericellular region that gives rise to macroscopic tracks created in scaffolds by migrating cells. This quantitative and predictable information should benefit the design of improved biomaterial scaffolds for medically relevant applications.Synthetic hydrogel scaffolds have been designed to serve as mimics of the native extracellular matrix (ECM) with the goal of promoting desired cell functions (e.g., proliferation, migration, differentiation), especially for applications in wound healing (1), tissue regeneration (2), and stem cell culture (3, 4). For example, poly(ethylene glycol) (PEG) hydrogels can serve as blank slates in which peptide cues can be systematically introduced in the scaffold to allow integrin binding (5, 6), proteolytic degradation (7, 8), and even local sequestering of growth factors (9). Furthermore, it is well known that cells respond to mechanical stimuli (e.g., stiffness) in their local microenvironment, the so-called pericellular region, and tuning of a scaffold’s mechanical properties can influence how a cell degrades and remodels its surroundings (10–12). The complex cell–matrix interactions that occur in the native ECM are often mimicked in peptide-functionalized hydrogels through the incorporation of adhesive binding peptides (e.g., RGDS, IKVAV) and enzymatically degradable peptide cross-linkers (e.g., GPQGIWGQ, GPLGLWAR), both of which are necessary for cell attachment, spreading (13), and motility (12, 14). However, changes in the local material properties as a result of this cell-mediated remodeling have largely remained a “black box,” limiting interpretation of data and confounding the design of more advanced biomaterials.Macroscopically, cells degrade micrometer-sized channels into scaffolds as they move, an event that begins with microscopic remodeling of their pericellular region and eventually permanently reengineering the scaffold architecture and material properties on a larger scale. If one seeks to design synthetic ECM environments to direct cellular processes, such as migration, it is important to better understand how these inputs are dynamically altered on the local length scale. Such information can help advance biomaterial design, especially for applications focused on the delivery or recruitment of cells, where directing cell–material interactions and migration can be critically important. At present, cell matrices are generally engineered to have certain initial material properties, but the resulting cell motility and cell–material interactions are often only empirically correlated with these design parameters (7, 15). To overcome this obstacle and provide an in situ measurement of scaffold degradation, microrheological measurements have been used to fingerprint and understand changes in material properties in the pericellular region during cell motility.Although real-time measurements of material properties near a cell are difficult, investigations have focused on developing techniques to access this information. In two dimensions, forces that cells exert when seeded on hydrogel surfaces have been measured using deflection of beds of microneedles (15) and deformation of gel surfaces (16). For example, Tan et al. (15) developed a measurement technique that exploits independent deflection of microneedles of varying lengths (and therefore stiffnesses) to measure the distribution of subcellular traction forces of both smooth muscle cells and fibroblasts. The main conclusion was that cellular spreading and morphology control the magnitude of the traction forces (15). The traction force of confluent cell sheets interacting with a gel surface was also analyzed, toward understanding how cellular processes are coordinated over large length scales. Using endothelial, epithelial, and breast cancer cell sheets, results showed that collective migration was due to a transmittance of normal stress across cell–cell junctions with migration orientated in the direction of the minimal intercellular shear stress (16).Cell-mediated degradation of the local microenvironment plays a critical role in permitting cellular migration and invasion in vivo. These processes are important during development, wound regeneration, and pathophysiological states facilitated by proteolytic events via cellular protease secretion. Previous work has begun to elucidate the length scales and spatial effects of secreted proteases in relation to migrating tumor cells during collagen matrix remodeling (17–19). For example, Packard et al. (20) used matrix metalloproteinase (MMP)-sensitive biosensors to visualize protease activity in the pericellular region of migrating tumor cells in collagen, finding increased activity at the polarized leading edge. These seminal works have elucidated the spatial presence and local activity of proteases in relation to individual migrating cells. However, how migrating cells temporally degrade and remodel the local microenvironment on larger length scales remains relatively unknown.Although 2D studies add to our understanding of cell–matrix interactions, 2D environments can unnaturally polarize cells, and some aspects of cell motility can be quite different in 2D versus 3D environments (21, 22). For these reasons, recent developments have focused on strategies to measure cell–material interactions in three dimensions (e.g., cell-laden hydrogels). Traction force microscopy measures spatial interfacial forces by quantifying the elastic deformation of a substrate (21). If the modulus of the material is known, this technique quantifies the forces cells exert in three dimensions calculated from embedded bead displacement. This approach has identified patterns of forces generated around distinct morphological regions during cellular invasion into a scaffold (21). Additionally, Bloom et al. (23) investigated the degradation of a collagen scaffold during the migration of a fibrosarcoma cell line (HT1080s) using embedded particle displacements. The authors showed that the hydrogel was reversibly deformed at the cell’s leading edge, but irreversibly remodeled at the trailing edge. Collectively, these pioneering investigations have provided insight into aspects of the complex interplay between cells and scaffold materials; however, complementary techniques that allow characterization of dynamic and local changes in mechanical properties, degradation, and scaffold erosion would be beneficial in further advancing our understanding of mechanotransduction, mechanisms of cell motility, and even biomaterials design.In this contribution, multiple particle tracking microrheology (MPT) is used to measure how human mesenchymal stem cells (hMSCs) remodel peptide cross-linked PEG hydrogels as they migrate. hMSC migration is characterized by significant remodeling of the local environment through attachment, enzymatic degradation, and cellular traction. Furthermore, hMSCs are observed to degrade the synthetic network through two pathways, MMP secretion that cleaves the peptide cross-linker and myosin II-regulated adhesion and reversible remodeling of the network. We find that MPT has the sensitivity to capture the temporal transition of the hydrogel from an elastic gel to a viscous liquid, during hMSC-mediated degradation. MPT simultaneously provides information about the spatial region, proximal to the cell, over which this matrix remodeling occurs. The technique and measurements enhance our understanding of cell–material interactions in three dimensions and enable visualization of dynamic cell-mediated matrix degradation, the so-called fourth dimension. On longer timescales, these microscopic changes give rise to the creation of macroscopic channels in the hydrogel that are important for hMSC motility. We believe that this approach and characterization can provide an important link for better understanding outside-in signaling experienced by cells when embedded in 3D environments.     

Adult bone marrow–derived cells: Regenerative potential, plasticity, and tissue commitment

Reconstitution of infarcted myocardium with functional new cardiomyocytes and vessels, a goal that only a few years ago would have been regarded as extravagant, is now actively pursued in numerous laboratories and clinical centers. Several recent studies in animals as well as humans have shown that transplantation of adult bone marrow-derived cells (BMCs) can improve left ventricular function and halt adverse remodeling after myocardial infarction. Differentiation of adult BMCs into cells of cardiac and vascular lineages has been proposed as a mechanism underlying these benefits and, indeed, differentiation of adult BMCs into cells of non-hematopoietic lineages, including cells of brain, skeletal muscle, heart, liver, and other organs, has been documented repeatedly both in vitro and in vivo. These results are in contrast with conventional definitions and dogma, according to which adult tissue-specific stem cells exhibit only restricted differentiation potential. Thus, these recent studies have sparked intense debate over the ability of adult BMCs to differentiate into non-hematopoietic tissues, and the regeneration of myocardium by differentiation of adult BMCs remains highly controversial. Because of the enormous clinical implications of BMC-mediated cardiac repair, numerous laboratories are currently addressing the feasibility of cardiac regeneration with BMCs and deciphering the mechanism underlying the beneficial effects. The purpose of this review is to critically examine the available evidence regarding the ability of adult BMCs to regenerate non-hematopoietic tissues and their utility in therapeutic cardiac regeneration.