James C. Culver

Three‐Dimensional Biomimetic Patterning in Hydrogels to Guide Cellular Organization

An image-guided micropatterning method is demonstrated for generating biomimetic hydrogel scaffolds with two-photon laser scanning photolithography. This process utilizes computational methods to directly translate three-dimensional cytoarchitectural features from labeled tissues into material structures. We use this method to pattern hydrogels that guide cellular organization by structurally and biochemically recapitulating complex vascular niche microenvironments with high pattern fidelity at the microscale.

The effects of hemodynamic force on embryonic development.

Microcirculation (2010) 17, 164–178. doi: 10.1111/j.1549‐8719.2010.00025.x

Coronary Microvascular Pericytes Are the Cellular Target of Sunitinib Malate–Induced Cardiotoxicity

Sunitinib-induced cardiotoxicity is caused by depletion of coronary pericytes due to loss of PDGFR signaling; this side effect can be prevented by thalidomide. Saving Pericytes to Prevent a Broken Heart In the world of targeted cancer therapies, sunitinib is a versatile one, targeting a variety of tyrosine kinase receptors. The breadth of its activity allows it to be effective in multiple different types of cancer but also increases the chances of unintended adverse effects. One such side effect is cardiotoxicity, with frequent reports of left ventricular dysfunction and heart failure in patients treated with sunitinib. Chintalgattu and co-workers have now uncovered the mechanism for this toxicity and demonstrated a way to protect the heart from treatment-induced damage in a mouse model. Pericytes are contractile cells that wrap around small blood vessels and are essential to their function. After sunitinib treatment, pericytes were no longer coating the coronary microvasculature in a mouse model. The blood vessels depleted of pericytes were unusually leaky, and the hearts of treated animals showed clear evidence of cardiac dysfunction. The depletion of pericytes was caused by the inhibition of signaling through platelet-derived growth factor receptor (PDGFR), a known target of sunitinib. The authors also discovered that thalidomide, a small-molecule drug that is already used in humans for the treatment of some cancers, could protect pericytes and prevent sunitinib-induced cardiotoxicity without affecting the antitumor effects of sunitinib. Future studies will be needed to uncover additional mechanism explaining why coronary pericytes in particular are so sensitive to inhibition of PDGFR and how thalidomide can protect these cells from toxicity. Eventually, this research could enable the creation of more specific targeted drugs that inhibit the kinases driving cancer cell proliferation without injuring pericytes and other healthy cells. In the meantime, the current findings of Chintalgattu et al. provide a rationale for testing the combination of thalidomide and sunitinib in human cancer patients to protect the patients’ hearts from injury while continuing to effectively target cancer cells. Sunitinib malate is a multitargeted receptor tyrosine kinase inhibitor used in the treatment of human malignancies. A substantial number of sunitinib-treated patients develop cardiac dysfunction, but the mechanism of sunitinib-induced cardiotoxicity is poorly understood. We show that mice treated with sunitinib develop cardiac and coronary microvascular dysfunction and exhibit an impaired cardiac response to stress. The physiological changes caused by treatment with sunitinib are accompanied by a substantial depletion of coronary microvascular pericytes. Pericytes are a cell type that is dependent on intact platelet-derived growth factor receptor (PDGFR) signaling but whose role in the heart is poorly defined. Sunitinib-induced pericyte depletion and coronary microvascular dysfunction are recapitulated by CP-673451, a structurally distinct PDGFR inhibitor, confirming the role of PDGFR in pericyte survival. Thalidomide, an anticancer agent that is known to exert beneficial effects on pericyte survival and function, prevents sunitinib-induced pericyte cell death in vitro and prevents sunitinib-induced cardiotoxicity in vivo in a mouse model. Our findings suggest that pericytes are the primary cellular target of sunitinib-induced cardiotoxicity and reveal the pericyte as a cell type of concern in the regulation of coronary microvascular function. Furthermore, our data provide preliminary evidence that thalidomide may prevent cardiotoxicity in sunitinib-treated cancer patients.

Understanding vascular development.

The vasculature of an organism has the daunting task of connecting all the organ systems to nourish tissue and sustain life. This complex network of vessels and associated cells must maintain blood flow, but constantly adapt to acute and chronic changes within tissues. While the vasculature has been studied for over a century, we are just beginning to understand the processes that regulate its formation and how genetic hierarchies are influenced by mechanical and metabolic cues to refine vessel structure and optimize efficiency. As we gain insights into the developmental mechanisms, it is clear that the processes that regulate blood vessel development can also enable the adult to adapt to changes in tissues that can be elicited by exercise, aging, injury, or pathology. Thus, research in vessel development has provided tremendous insights into therapies for vascular diseases and disorders, cancer interventions, wound repair and tissue engineering, and in turn, these models have clearly impacted our understanding of development. Here we provide an overview of the development of the vascular system, highlighting several areas of active investigation and key questions that remain to be answered. WIREs Dev Biol 2013, 2:327–346. doi: 10.1002/wdev.91

A Specialized Microvascular Domain in the Mouse Neural Stem Cell Niche

The microenvironment of the subependymal zone (SEZ) neural stem cell niche is necessary for regulating adult neurogenesis. In particular, signaling from the microvasculature is essential for adult neural stem cell maintenance, but microvascular structure and blood flow dynamics in the SEZ are not well understood. In this work, we show that the mouse SEZ constitutes a specialized microvascular domain defined by unique vessel architecture and reduced rates of blood flow. Additionally, we demonstrate that hypoxic conditions are detectable in the ependymal layer that lines the ventricle, and in a subpopulation of neurons throughout the SEZ and striatum. Together, these data highlight previously unidentified features of the SEZ neural stem cell niche, and further demonstrate the extent of microvascular specialization in the SEZ microenvironment.

Recapitulation and Modulation of the Cellular Architecture of a User-Chosen Cell of Interest Using Cell-Derived, Biomimetic Patterning.

Heterogeneity of cell populations can confound population-averaged measurements and obscure important findings or foster inaccurate conclusions. The ability to generate a homogeneous cell population, at least with respect to a chosen trait, could significantly aid basic biological research and development of high-throughput assays. Accordingly, we developed a high-resolution, image-based patterning strategy to produce arrays of single-cell patterns derived from the morphology or adhesion site arrangement of user-chosen cells of interest (COIs). Cells cultured on both cell-derived patterns displayed a cellular architecture defined by their morphology, adhesive state, cytoskeletal organization, and nuclear properties that quantitatively recapitulated the COIs that defined the patterns. Furthermore, slight modifications to pattern design allowed for suppression of specific actin stress fibers and direct modulation of adhesion site dynamics. This approach to patterning provides a strategy to produce a more homogeneous cell population, decouple the influences of cytoskeletal structure, adhesion dynamics, and intracellular tension on mechanotransduction-mediated processes, and a platform for high-throughput cellular assays.

Biomimetic Surface Patterning Promotes Mesenchymal Stem Cell Differentiation

Both chemical and mechanical stimuli can dramatically influence cell behavior. By optimizing the signals cells experience, it may be possible to control the behavior of therapeutic cell populations. In this work, biomimetic geometries of adhesive ligands, which recapitulate the morphology of mature cells, are used to direct human mesenchymal stem cell (HMSC) differentiation toward a desired lineage. Specifically, adipocytes cultured in 2D are imaged and used to develop biomimetic virtual masks used in laser scanning lithography to form patterned fibronectin surfaces. The impact of adipocyte-derived pattern geometry on HMSC differentiation is compared to the behavior of HMSCs cultured on square and circle geometries, as well as adipocyte-derived patterns modified to include high stress regions. HMSCs on adipocyte mimetic geometries demonstrate greater adipogenesis than HMSCs on the other patterns. Greater than 45% of all HMSCs cultured on adipocyte mimetic patterns underwent adipogenesis as compared to approximately 19% of cells on modified adipocyte patterns with higher stress regions. These results are attributed to variations in cytoskeletal tension experienced by cells on the different protein micropatterns. The effects of geometry on adipogenesis are mitigated by the incorporation of a cytoskeletal protein inhibitor; exposure to this inhibitor leads to increased adipogenesis on all patterns examined.