The enhancement of wound healing has been a goal of medical practitioners for thousands of years. The development of chronic, non-healing wounds is a persistent medical problem that drives patient morbidity and increases healthcare costs. A key aspect of many non-healing wounds is the reduced presence of vessel growth through the process of angiogenesis. This review surveys the creation of new treatments for healing cutaneous wounds through therapeutic angiogenesis. In particular, we discuss the challenges and advancement that have been made in delivering biologic, pharmaceutical and cell-based therapies as enhancers of wound vascularity and healing.

Peripheral ischemia as a result of occlusive vascular disease is a widespread problem in patients older than the age of 65. Angiogenic therapies that can induce microvascular growth have great potential for providing a long-lasting solution for patients with ischemia and would provide an appealing alternative to surgical and percutaneous interventions. However, many angiogenic therapies have seen poor efficacy in clinical trials, suggesting that patients with long-term peripheral ischemia have considerable therapeutic resistance to angiogenic stimuli. Glioblastoma is one of the most angiogenic tumor types, inducing robust vessel growth in the area surrounding the tumor. One major angiogenic mechanism used by the tumor cells to induce blood vessel growth is the production of exosomes and other extracellular vesicles that can carry pro-angiogenic and immunomodulatory signals. Here, we explored whether the pro-angiogenic aspects of glioblastoma-derived exosomes could be harnessed to promote angiogenesis and healing in the context of peripheral ischemic disease. We demonstrate that the exosomes derived from glioblastoma markedly enhance endothelial cell proliferation and increase endothelial tubule formation in vitro. An analysis of the microRNA expression using next generation sequencing identified that exosomes contained a high concentration of miR-221. In addition, we found that glioblastoma exosomes contained significant amounts of the proteoglycans glypican-1 and syndecan-4, which can serve as co-receptors for angiogenic factors, including fibroblast growth factor-2 (FGF-2). In a hindlimb ischemia model in mice, we found that the exosomes promoted enhanced revascularization in comparison to control alginate gels and FGF-2 treatment alone. Taken together, our results support the fact that glioblastoma-derived exosomes have powerful effects in increasing revascularization in the context of peripheral ischemia.

The metastatic spread of cancer is a major barrier to effective and curative therapies for cancer. During metastasis, tumor cells intravasate into the vascular system, survive in the shear forces and immunological environment of the circulation, and then extravasate into secondary tumor sites. Biophysical forces are potent regulators of cancer biology and are key in many of the steps of metastasis. In particular, the adhesion of circulating cells is highly dependent upon competing forces between cell adhesion receptors and the shear stresses due to fluid flow. Conventional in vitro assays for drug development and the mechanistic study of metastasis are often carried out in the absence of fluidic forces and, consequently, are poorly representative of the true biology of metastasis. Here, we present a novel high-throughput approach to studying cell adhesion under flow that uses a multi-well, mechanofluidic flow system to interrogate adhesion of cancer cell to endothelial cells, extracellular matrix and platelets under physiological shear stresses. We use this system to identify pathways and compounds that can potentially be used to inhibit cancer adhesion under flow by screening anti-inflammatory compounds, integrin inhibitors and a kinase inhibitor library. In particular, we identify several small molecule inhibitors of FLT-3 and AKT that are potent inhibitors of cancer cell adhesion to endothelial cells and platelets under flow. In addition, we found that many kinase inhibitors lead to increased adhesion of cancer cells in flow-based but not static assays. This finding suggests that even compounds that reduce cell proliferation might also enhance cancer cell adhesion during metastasis. Overall, our results validate a novel platform for investigating the mechanisms of cell adhesion under biophysical flow conditions and identify several potential inhibitors of cancer cell adhesion during metastasis.

Non-healing chronic ulcers are a serious complication of diabetes and are a major healthcare problem. While a host of treatments have been explored to heal or prevent these ulcers from forming, these treatments have not been found to be consistently effective in clinical trials.

The cells of the vascular system are highly sensitive to biophysical cues from their local cellular microenvironment. To engineer improved materials for vascular devices and delivery of cell therapies, a key challenge is to understand the mechanisms that cells use to sense biophysical cues from their environment. Syndecans are heparan sulfate proteoglycans (HSPGs) that consist of a protein core modified with heparan sulfate glycosaminoglycan chains. Due to their presence on the cell surface and their interaction with cytoskeletal and focal adhesion associated molecules, cell surface proteoglycans are well poised to serve as mechanosensors of the cellular microenvironment. Nanotopological cues have become recognized as major regulators of cell growth, migration and phenotype. We hypothesized that syndecan-1 could serve as a mechanosensor for nanotopological cues and can mediate the responsiveness of vascular smooth muscle cells to nanoengineered materials. We created engineered substrates made of polyurethane acrylate with nanogrooves using ultraviolet-assisted capillary force lithography. We cultured vascular smooth muscle cells with knockout of syndecan-1 on engineered substrates with varying compliance and nanotopology. We found that knockout of syndecan-1 reduced alignment of vascular smooth muscle cells to the nanogrooves under inflammatory treatments. In addition, we found that loss of syndecan-1 increased nuclear localization of Yap/Taz and phospho-Smad2/3 in response to nanogrooves. Syndecan-1 knockout vascular smooth muscle cells also had elevated levels of Rho-associated protein kinase-1 (Rock1), leading to increased cell stiffness and an enhanced contractile state in the cells. Together, our findings support that syndecan-1 knockout leads to alterations in mechanosensing of nanotopographical cues through alterations of in rho-associated signaling pathways, cell mechanics and mediators of the Hippo and TGF-β signaling pathways.

The kinetics of receptor-mediated cell adhesion to extracellular matrix and adherent cell monolayers plays a key role in many physiological and pathological processes including cancer metastasis. Within this process the presence of fluidic shear forces is a key regulator of binding equilibrium and kinetics of cell adhesion. Current techniques to examine the kinetics of cell adhesion are either performed in the absence of flow or are low throughput, limiting their application to pharmacological compound screening or the high throughput investigation of biological mechanisms. We developed a high throughput flow device that applies flow in a multi-well format and interfaced this system with electric cell-substrate impedance sensing (ECIS) system to allow label free detection of cell adhesion. We demonstrate that this combined system is capable of making real time measurements of cancer cell adhesion to extracellular matrix and immobilized platelets. In addition, we examined the dependence of the kinetics of binding of cancer cells on the level of shear stress and in the presence of small molecule inhibitors to adhesion-related pathways. This versatile system is broadly adaptable to the high throughput study of cell adhesion kinetics for many applications including drug screening and the investigation of the mechanisms of cancer metastasis.

Physical activity has been consistently linked to decreased incidence of breast cancer and a substantial increase in the length of survival of patients with breast cancer. However, the understanding of how applied physical forces directly regulate breast cancer remains limited. We investigated the role of mechanical forces in altering the chemoresistance, proliferation and metastasis of breast cancer cells. We found that applied mechanical tension can dramatically alter gene expression in breast cancer cells, leading to decreased proliferation, increased resistance to chemotherapeutic treatment and enhanced adhesion to inflamed endothelial cells and collagen I under fluidic shear stress. A mechanistic analysis of the pathways involved in these effects supported a complex signaling network that included Abl1, Lck, Jak2 and PI3K to regulate pro-survival signaling and enhancement of adhesion under flow. Studies using mouse xenograft models demonstrated reduced proliferation of breast cancer cells with orthotopic implantation and increased metastasis to the skull when the cancer cells were treated with mechanical load. Using high throughput mechanobiological screens we identified pathways that could be targeted to reduce the effects of load on metastasis and found that the effects of mechanical load on bone colonization could be reduced through treatment with a PI3Kγ inhibitor.

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