Agent-based computational model of retinal angiogenesis simulates microvascular network morphology as a function of pericyte coverage.

OBJECTIVE: Define a role for perivascular cells during developmental retinal angiogenesis in the context of EC Notch1-DLL4 signaling at the multicellular network level. METHODS: The retinal vasculature is highly sensitive to growth factor-mediated intercellular signaling. Although EC signaling has been explored in detail, it remains unclear how PC function to modulate these signals that lead to a diverse set of vascular network patterns in health and disease. We have developed an ABM of retinal angiogenesis that incorporates both ECs and PCs to investigate the formation of vascular network patterns as a function of pericyte coverage. We use our model to test the hypothesis that PC modulate Notch1-DLL4 signaling in endothelial cell-endothelial cell interactions. RESULTS: Agent-based model (ABM) simulations that include PCs more accurately predict experimentally observed vascular network morphologies than simulations that lack PCs, suggesting that PCs may influence sprouting behaviors through physical blockade of endothelial intercellular connections. CONCLUSIONS: This study supports a role for PCs as a physical buffer to signal propagation during vascular network formation-a barrier that may be important for generating healthy microvascular network patterns.

Flt-1 (VEGFR-1) coordinates discrete stages of blood vessel formation.

AIMS: In developing blood vessel networks, the overall level of vessel branching often correlates with angiogenic sprout initiations, but in some pathological situations, increased sprout initiations paradoxically lead to reduced vessel branching and impaired vascular function. We examine the hypothesis that defects in the discrete stages of angiogenesis can uniquely contribute to vessel branching outcomes. METHODS AND RESULTS: Time-lapse movies of mammalian blood vessel development were used to define and quantify the dynamics of angiogenic sprouting. We characterized the formation of new functional conduits by classifying discrete sequential stages-sprout initiation, extension, connection, and stability-that are differentially affected by manipulation of vascular endothelial growth factor-A (VEGF-A) signalling via genetic loss of the receptor flt-1 (vegfr1). In mouse embryonic stem cell-derived vessels genetically lacking flt-1, overall branching is significantly decreased while sprout initiations are significantly increased. Flt-1(-/-) mutant sprouts are less likely to retract, and they form increased numbers of connections with other vessels. However, loss of flt-1 also leads to vessel collapse, which reduces the number of new stable conduits. Computational simulations predict that loss of flt-1 results in ectopic Flk-1 signalling in connecting sprouts post-fusion, causing protrusion of cell processes into avascular gaps and collapse of branches. Thus, defects in stabilization of new vessel connections offset increased sprout initiations and connectivity in flt-1(-/-) vascular networks, with an overall outcome of reduced numbers of new conduits. CONCLUSIONS: These results show that VEGF-A signalling has stage-specific effects on vascular morphogenesis, and that understanding these effects on dynamic stages of angiogenesis and how they integrate to expand a vessel network may suggest new therapeutic strategies.

Multiscale computational models of complex biological systems.

Integration of data across spatial, temporal, and functional scales is a primary focus of biomedical engineering efforts. The advent of powerful computing platforms, coupled with quantitative data from high-throughput experimental methodologies, has allowed multiscale modeling to expand as a means to more comprehensively investigate biological phenomena in experimentally relevant ways. This review aims to highlight recently published multiscale models of biological systems, using their successes to propose the best practices for future model development. We demonstrate that coupling continuous and discrete systems best captures biological information across spatial scales by selecting modeling techniques that are suited to the task. Further, we suggest how to leverage these multiscale models to gain insight into biological systems using quantitative biomedical engineering methods to analyze data in nonintuitive ways. These topics are discussed with a focus on the future of the field, current challenges encountered, and opportunities yet to be realized.

Hypoxic culture and in vivo inflammatory environments affect the assumption of pericyte characteristics by human adipose and bone marrow progenitor cells.

Previous studies have shown that exposure to a hypoxic in vitro environment increases the secretion of pro-angiogenic growth factors by human adipose-derived stromal cells (hASCs) [Cao Y, et al., Biochem Biophys Res Commun 332: 370-379, 2005; Kokai LE, et al., Plast Reconstr Surg 116: 1453-1460, 2005; Park BS, et al., Biomed Res (Tokyo) 31: 27-34, 2010; Rasmussen JG, et al., Cytotherapy 13: 318-328, 2010; Rehman J, et al., Circulation 109: 1292-1298, 2004]. Previously, it has been demonstrated that hASCs can differentiate into pericytes and promote microvascular stability and maintenance during angiogenesis in vivo (Amos PJ, et al., Stem Cells 26: 2682-2690, 2008; Traktuev DO, et al., Circ Res 102: 77-85, 2008). In this study, we tested the hypotheses that angiogenic induction can be increased and pericyte differentiation decreased by pretreatment of hASCs with hypoxic culture and that hASCs are similar to human bone marrow-derived stromal cells (hBMSCs) in these regards. Our data confirms previous studies showing that hASCs: 1) secrete pro-angiogenic proteins, which are upregulated following culture in hypoxia, and 2) migrate up gradients of PDGF-BB in vitro, while showing for the first time that a rat mesenteric model of angiogenesis induced by 48/80 increases the propensity of both hASCs and hBMSCs to assume perivascular phenotypes following injection. Moreover, culture of both cell types in hypoxia before injection results in a biphasic vascular length density response in this model of inflammation-induced angiogenesis. The effects of hypoxia and inflammation on the phenotype of adult progenitor cells impacts both the therapeutic and the basic science applications of the cell types, as hypoxia and inflammation are common features of natural and pathological vascular compartments in vivo.

Prevention of local tumor recurrence following surgery using low-dose chemotherapeutic polymer films.

AIM: To evaluate the efficacy of a polymer film designed for prolonged paclitaxel release at surgical margins to prevent local recurrence in non-small-cell lung cancer (NSCLC) following complete surgical resection in a murine model. METHODS: Poly(glycerol monostearate co-epsilon-caprolactone) polymer films were prepared with or without 10% (w/w) paclitaxel and characterized for prolonged tumor cytotoxicity in vitro against several NSCLC cell lines including LLC, NCI-H460, and NCI-H292. Films were implanted following complete LLC tumor resection and assessed in vivo for prevention of local tumor recurrence, impact on wound healing, and extent of local drug delivery. Plasma and local tissue concentrations of paclitaxel were compared following systemic administration and film implantation. RESULTS: The flexible polymeric films eluted paclitaxel over several weeks and remained cytotoxic to LLC, NCI-H460, and NCI-H292 cells in vitro for 50 days, while unloaded films did not impair tumor cell growth. Implanted paclitaxel films prevented local tumor recurrence in vivo in 83.3% of animals, compared with unloaded films (12.5%), systemic (22.2%) or locally administered paclitaxel (0%) (P < 0.005). Although minimal paclitaxel remained in either plasma or tissue 10 days after systemic injection, local paclitaxel concentration at the site of surgical resection was significantly greater (3,000-fold) at 10 days when paclitaxel was locally delivered via films (P = 0.024). CONCLUSIONS: Local application of paclitaxel-loaded polymer films following surgical resection can prevent local tumor recurrence without impairing wound healing.

Expansile nanoparticles: synthesis, characterization, and in vivo efficacy of an acid-responsive polymeric drug delivery system.

Nanoparticles are finding increased uses in drug delivery applications as a means to increase treatment efficacy and improve patient care. Here, we report engineered polymeric nanoparticles that undergo a hydrophobic to hydrophilic transition at pH 5 to afford swelling and rapid release of their contents. As our clinical interest lies in the prevention of lung tumor recurrence following resection, the nanoparticles were evaluated in a model mimicking microscopic disease, akin to residual occult tumor that can remain at the resection margin following surgery. Expansile nanoparticles loaded with paclitaxel, a poorly water-soluble anticancer drug, prevent establishment of lung cancer in vivo and are superior to the conventional drug delivery method for paclitaxel using Cremophor EL/ethanol.

Prevention of local tumor growth with paclitaxel-loaded microspheres.

OBJECTIVES: Lung cancer is associated with a significant rate of locoregional recurrence after surgical resection, particularly when nonanatomic wedge resections are performed. The primary aim of this study was to assess the feasibility of a microsphere drug delivery system to locally deliver chemotherapy and prevent the establishment and growth of lung cancer cells and establish proof of concept for a potential future approach to target occult microscopic disease remaining at the surgical resection margin. METHODS: Poly-(D,L-lactic-co-glycolic acid) (PLGA) microspheres loaded with the antineoplastic agent paclitaxel were prepared and tested for antitumor efficacy in an in vitro cell proliferation assay for tumor inhibition and induction of apoptosis. The in vivo prevention of Lewis lung carcinoma cell establishment and growth in subcutaneous tissues of mice was also assessed by comparing 4 treatment groups: Lewis lung carcinoma cells alone, Lewis lung carcinoma cells combined with 100 x 10(6) unloaded (carrier alone) PLGA microspheres, and Lewis lung carcinoma cells combined with 50 x 10(6) or 100 x 10(6) paclitaxel-loaded PLGA microspheres. After the coinjection of Lewis lung carcinoma cells with or without microspheres, in vivo tumor growth was monitored, and tumor weight was recorded on death. RESULTS: Paclitaxel-loaded PLGA microspheres were found to effectively prevent growth of tumor cells in culture through the induction of apoptosis. Similarly, paclitaxel-loaded PLGA microspheres significantly inhibited tumor growth in vivo at both the 50 x 10(6) and 100 x 10(6) microsphere dose (0.497 +/- 0.183 and 0.187 +/- 0.083 g total tumor weight, respectively) compared with 2.91 +/- 0.411 g for Lewis lung carcinoma cells with unloaded microspheres and 3.37 +/- 0.433 g for untreated tumor (P < .001). Toxicity was not clinically apparent in any animal treated with paclitaxel-loaded PLGA microspheres. CONCLUSIONS: Paclitaxel-loaded PLGA microspheres induce tumor apoptosis and inhibit the establishment and growth of lung cancer cells both in vitro and in vivo without obvious systemic toxicity. By using models consistent with localized microscopic tumor burdens, these results suggest that local delivery of paclitaxel through a microsphere system might lead to an effective future method of decreasing local tumor recurrence in non-small cell lung cancer when applied to the surgical margins at risk for microscopic tumor foci. Such an approach might be particularly efficacious after wedge resection in the setting of poor pulmonary reserve or significant comorbidity, where local recurrence rates are increased and acceptable alternative treatment options are limited.