Capillary Sprouts Research Articles

On the mathematical modeling of wound healing angiogenesis in skin as a reaction-transport process.

Over the last 30 years, numerous research groups have attempted to provide mathematical descriptions of the skin wound healing process. The development of theoretical models of the interlinked processes that underlie the healing mechanism has yielded considerable insight into aspects of this critical phenomenon that remain difficult to investigate empirically. In particular, the mathematical modeling of angiogenesis, i.e., capillary sprout growth, has offered new paradigms for the understanding of this highly complex and crucial step in the healing pathway. With the recent advances in imaging and cell tracking, the time is now ripe for an appraisal of the utility and importance of mathematical modeling in wound healing angiogenesis research. The purpose of this review is to pedagogically elucidate the conceptual principles that have underpinned the development of mathematical descriptions of wound healing angiogenesis, specifically those that have utilized a continuum reaction-transport framework, and highlight the contribution that such models have made toward the advancement of research in this field. We aim to draw attention to the common assumptions made when developing models of this nature, thereby bringing into focus the advantages and limitations of this approach. A deeper integration of mathematical modeling techniques into the practice of wound healing angiogenesis research promises new perspectives for advancing our knowledge in this area. To this end we detail several open problems related to the understanding of wound healing angiogenesis, and outline how these issues could be addressed through closer cross-disciplinary collaboration.

Agent-based model of angiogenesis simulates capillary sprout initiation in multicellular networks

Agent-based model of angiogenesis simulates capillary sprout initiation in multicellular networks

Role of angiogenesis and angiogenic factors in acute and chronic wound healing

Angiogenesis plays a crucial role in wound healing by forming new blood vessels from preexisting vessels by invading the wound clot and organizing into a microvascular network throughout the granulation tissue. This dynamic process is highly regulated by signals from both serum and the surrounding extracellular matrix environment. Vascular endothelial growth factor, angiopoietin, fibroblast growth factor and transforming growth factor-beta are among the potent angiogenic cytokines in wound angiogenesis. Specific endothelial cell ECM receptors are critical for morphogenetic changes in blood vessels during wound repair. In particular integrin (αvβ3) receptors for fibrin and fibronectin, appear to be required for wound angiogenesis: αvβ3 is focally expressed at the tips of angiogenic capillary sprouts invading the wound clot, and any functional inhibitors of αvβ3 such as monoclonal antibodies, cyclic RGD peptide antagonists, and peptidomimetics rapidly inhibit granulation tissue formation. In spite of clear knowledge about influence of many angiogenic factors on wound healing, little progress has been made in defining the source of these factors, the regulatory events involved in wound angiogenesis and in the clinical use of angiogenic stimulants to promote repair.

Agent-based model of angiogenesis simulates capillary sprout initiation in multicellular networks.

Many biological processes are controlled by both deterministic and stochastic influences. However, efforts to model these systems often rely on either purely stochastic or purely rule-based methods. To better understand the balance between stochasticity and determinism in biological processes a computational approach that incorporates both influences may afford additional insight into underlying biological mechanisms that give rise to emergent system properties. We apply a combined approach to the simulation and study of angiogenesis, the growth of new blood vessels from existing networks. This complex multicellular process begins with selection of an initiating endothelial cell, or tip cell, which sprouts from the parent vessels in response to stimulation by exogenous cues. We have constructed an agent-based model of sprouting angiogenesis to evaluate endothelial cell sprout initiation frequency and location, and we have experimentally validated it using high-resolution time-lapse confocal microscopy. ABM simulations were then compared to a Monte Carlo model, revealing that purely stochastic simulations could not generate sprout locations as accurately as the rule-informed agent-based model. These findings support the use of rule-based approaches for modeling the complex mechanisms underlying sprouting angiogenesis over purely stochastic methods.

A hybrid mathematical model of tumor-induced angiogenesis with blood perfusion

Angiogenesis, the growth of new blood vessel from existing ones, is a pivotal stage in cancer development, and is an important target for cancer therapy. We develop a hybrid mathematical model to understand the mechanisms behind tumor-induced angiogenesis. This model describes uptake of Tumor Angiogenic Factor (TAF) at extracellular level, uses partial differential equation to describe the evolution of endothelial cell density including TAF induced proliferation, chemotaxis to TAF, and haptotaxis to extracellular matrix. In addition we also consider the phenomenon of blood perfusion in the micro-vessels. The model produces sprout formation with realistic morphological and dynamical features, including the so-called brush border effect, the dendritic branching and fusing of the capillary sprouts forming a vessel network. The model also demonstrates the effects of individual mechanisms in tumor angiogenesis: Chemotaxis to TAF is the key driving mechanisms for the extension of sprout cell; endothelial proliferation is not absolutely necessary for sprout extension; haptotaxis to ExtraCellular Matrix (ECM) gradient provides additional guidance to sprout extension, suggesting potential targets for anti-angiogenic therapies.

A hierarchical model for the development of cerebral arteriovenous malformations

ObjectiveCerebral arteriovenous malformations (AVMs) are vascular lesions whose pathogenesis, although not fully elucidated, is likely multifactorial. Recent research investigating vessel development suggests a potential hierarchical model in which capillary sprouts from higher-flow arteries give rise to lower-flow veins. It is possible that an embryologic structural vascular dysgenesis in this hierarchical development heavily contributes to the formation of AVMs. Subsequent genetic “second hits” may then allow development of a clinically significant cerebral AVM. We review this vascular developmental process and describe a novel proposal for the embryogenesis of AVMs and its implications in relation to recent research on polymorphisms and AVMs. MethodsA comprehensive literature search was performed using PubMed for recent research relative to cerebral AVMs, embryologic vascular development, and polymorphisms involved in AVM pathology. ResultsIt has recently been shown that both centrally, in the axial embryo, and peripherally, in the embryonic yolk sac, veins form via capillary sprouting from parent arteries. In developing intracranial vessels, a derangement in this embryonic process may lead to a primitive arteriovenous shunt. After this structural “first hit,” we suggest that single nucleotide polymorphisms (SNPs) are a major component in allowing AVM growth into symptomatic clinical lesions. ConclusionsThis is a novel theory for the embryologic formation of cerebral AVMs. Hierarchical vessel development, where higher-flow parent arteries give rise to lower-flow veins, provides a potential mechanism for the formation of primitive arteriovenous shunts that, with the influence of polymorphisms, allows AVMs to develop.

Intra-epithelially entrapped blood vessels in ameloblastoma.

The ameloblastoma is a benign but locally aggressive odontogenic neoplasm with a high recurrence rate. While significant progress has been made in our understanding regarding the role of tumoral vasculature relative to the diverse behavioral characteristics of this tumor, no attention has been paid to a distinct subset of blood vessels entrapped within its epithelial compartment. As vascular niches are known to influence tumoral growth, clarification of these vessels is important. The objectives of this study were to investigate the morphologic characteristics of intra-epithelially entrapped blood vessels (IEBVs) in ameloblastoma and to speculate on their relevance. Here, we evaluated the frequency, microvessel density (MVD), morphology, and distribution pattern of IEBVs in 77 ameloblastoma of different subtypes based on their immunoreactivity for endothelial markers (CD34, CD31, CD105), vascular tight junction protein (claudin-5), pericyte [α-smooth muscle actin (α-sma)], and vascular basement membrane (collagen IV). IEBVs were heterogeneously detected in ameloblastoma. Their mean MVD (CD34 = 15.46 ± 7.25; CD31 = 15.8 ± 5.04; CD105 = 0.82 ± 0.51) showed no significant correlation with different subtypes, and between primary and recurrent tumors (P > 0.05). These microvessels may occur as single/clusters of capillary sprouts, or formed compressed branching/non-branching slits entrapped within the epithelial compartment, and in direct apposition with polyhedral/granular neoplastic epithelial cells. They expressed proteins for endothelial tight junctions (claudin-5-positive) and pericytes (α-sma-positive) but had deficient basement membrane (collagen IV weak to absent). Aberrant expression for CD34, CD31, and CD105 in tumor epithelium was variably observed. Although rare in occurrence, identification of IEBVs in ameloblastoma could potentially represent a new paradigm for vascular assessment of this neoplasm.

UPARANT: A Urokinase Receptor–Derived Peptide Inhibitor of VEGF-Driven Angiogenesis with Enhanced Stability and In Vitro and In Vivo Potency

This work is based on previous evidence showing that chemotactic sequence of the urokinase receptor (uPAR(88-92)) drives angiogenesis in vitro and in vivo in a protease-independent manner, and that the peptide Ac-Arg-Glu-Arg-Phe-NH(2) (RERF) prevents both uPAR(88-92)- and VEGF-induced angiogenesis. New N-acetylated and C-amidated peptide analogues containing α-methyl α-amino acids were designed and synthesized to optimize the biochemical properties for therapeutic applications. Among these, Ac-L-Arg-Aib-L-Arg-D-Cα(Me)Phe-NH2, named UPARANT, adopts in solution a turned conformation similar to that found for RERF, is stable to sterilization in 3 mg/mL sealed vials in autoclave for 20 minutes at 120°C, is stable in blood, and displays a long-time resistance to enzymatic proteolysis. UPARANT competes with N-formyl-Met-Leu-Phe (fMLF) for binding to the formyl-peptide receptor, inhibits VEGF-directed endothelial cell migration, and prevents cytoskeletal organization and αvβ3 activation in endothelial cells exposed to VEGF. In vitro, UPARANT inhibits VEGF-dependent tube formation of endothelial cells at a 100× lower concentration than RERF. In vivo, UPARANT reduces to the basal level VEGF-dependent capillary sprouts originating from the host vessels that invaded Matrigel sponges implanted in mice, and completely prevents neovascularization induced by subcorneal implantation of pellets containing VEGF in rabbits. Both excellent stability and potency position UPARANT as a promising new therapeutic agent for the control of diseases fueled by excessive angiogenesis, such as cancer and inflammation.

The effect of cancer cells on angiogenesis: engineering the cancer niche (676.13)

Tumor growth requires uncontrolled angiogenesis, defined as the growth of new capillaries from existing blood vessels. Developing anti‐tumor therapies necessitates understanding how cancer cells and their microenvironment synergistically influence capillary sprouting from existing vascular networks. Recently, we demonstrated the use of the rat mesentery culture model for investigating angiogenesis in an intact microvascular network scenario. The objective of this study was to demonstrate the use of this model to determine the effects of cancer cells on angiogenesis. Mesenteric tissue windows were harvested from adult male Wistar rats and cultured for 5 days according to the following experimental groups: 1) Control (MEM + 10% DMEM + 1% FBS; n = 7 tissues) and 2) Cancer Cell (MEM + 10% DMEM + 1% FBS + cells; n = 6 tissues). For the cancer cell group, 2x10^5 RFP‐transfected metastatic MDA‐MB‐231 breast cancer cells were seeded onto tissues on day 0. On day 5, cancer cells remained viable, as confirmed by live/dead assay. More importantly, cells were observed to be in the same focal plane and region as vessels along intact microvascular networks. The presence of cancer cells decreased vascular length density and the number of capillary sprouts per vascular area compared to the Control (no cell) group. Our results suggest that cancer cells alone are not sufficient to induce angiogenesis. Application of the rat mesentery culture model as a multicellular breadboard system to engineer the cancer cell/vascular niche will offer a new tool for tumor angiogenesis research.Grant Funding Source: Supported by Tulane Center for Aging and NIH 5‐P20GM103629‐02

A novel ex vivo tissue culture assay for determining the effects of anti‐tumor drugs on angiogenesis (676.14)

Tumor growth is highly dependent upon angiogenesis and the interplay between the tumor stroma and the microvasculature. A challenge in evaluating anti‐tumor therapies is the inability to decouple treatment effects on cancer cells versus the microvascular network. Recently, our laboratory demonstrated that the “rat mesentery culture” model could be used for time‐lapse comparison of intact microvascular networks before and after angiogenesis induction. The objective of this study was to demonstrate that this model could be used to evaluate the anti‐angiogenic effects for a given drug treatment. Mesenteric windows were harvested from adult male Wistar rats and cultured for 3 days in serum containing medium (MEM + 10% Fetal Bovine Serum). Explants were exposed to increasing concentrations of Nelfinavir (Viracept™) (0 ‐ 4.5 μM) and Curcumin (0 ‐ 4.5 μM). Nelfinavir and Curcumin were selected for the study because Nelfinavir causes endothelial dysfunction and Curcumin, an anti‐oxidant and NF‐κB inhibitory dietary supplement, is known to have anti‐angiogenic effects. Preliminary studies also suggest that the combination of Nelfinavir/Curcumin produce synergistic anti‐cancer effects. Labeling tissues with FITC conjugated BSI‐lectin on Day 0 and Day 3 identified endothelial cells along all vessels. Results showed significant dose‐responsive effects of Nelfinavir/Curcumin combinations in suppressing both vessel segment density and the number of capillary sprouts per vascular area. Our findings establish the rat mesentery culture model as a valuable ex vivo tool for the rapid and reproducible screening of drug effects on angiogenesis in an intact network scenario.Grant Funding Source: Supported by Tulane Center for Aging and NIH 5‐P20GM103629‐02

Y-tocotrienol inhibits angiogenesis-dependent growth of human hepatocellular carcinoma through abrogation of AKT/mTOR pathway in an orthotopic mouse model.

Angiogenesis is one of the key hallmarks of cancer. In this study, we investigated whether γ-tocotrienol can abrogate angiogenesis-mediated tumor growth in hepatocellular carcinoma (HCC) and if so, through what molecular mechanisms. We observed that γ-tocotrienol inhibited vascular endothelial growth factor (VEGF)-induced migration, invasion, tube formation and viability of HUVECs in vitro. Moreover, γ-tocotrienol reduced the number of capillary sprouts from matrigel embedded rat thoracic aortic ring in a dose-dependent manner. Also, in chick chorioallantoic membrane assay, γ-tocotrienol significantly reduced the blood vessels formation. We further noticed that γ-tocotrienol blocked angiogenesis in an in vivo matrigel plug assay. Furthermore, γ-tocotrienol inhibited VEGF-induced autophosphorylation of VEGFR2 in HUVECs and also suppressed the constitutive activation of AKT/mammalian target of rapamycin (mTOR) signal transduction cascades in HUVECs as well as in HCC cells. Interestingly, γ-tocotrienol was also found to significantly reduce the tumor growth in an orthotopic HCC mouse model and inhibit tumor-induced angiogenesis in HCC patient xenografts through the suppression of various biomarkers of proliferation and angiogenesis. Taken together, our findings strongly suggest that γ-tocotrienol might be a promising anti-angiogenic drug with significant antitumor activity in HCC.

Integrated Effects of Matrix Mechanics and Vascular Endothelial Growth Factor (VEGF) on Capillary Sprouting

Angiogenesis is the growth of new capillaries from existing vasculature. Vascular network formation is known to be regulated by the biophysical and biochemical signals emanating from the microenvironment. However, it is not clear how endothelial cells integrate these signals to drive the capillary morphogenesis. In this study, human umbilical endothelial cells (HUVECs) were seeded on scaffolds with varying stiffness and capillary formation was quantified. Our study revealed that cells formed well defined networks on compliant gels. Increase in stiffness resulted in a decrease in sprouting. VEGF was encapsulated within the scaffolds at concentrations ranging from 0 to 100ng/mL to investigate the interplay between VEGF concentration profiles and matrix stiffness in guiding network formation. Quantitative analysis revealed that while VEGF disrupted sprout formation on compliant substrates, it elicited a sprouting response in cells seeded on scaffolds of intermediate rigidity. Additionally, we also observed that a minimum cell density is required for the formation of well-connected vascular networks.

Dll4-containing exosomes induce capillary sprout retraction in a 3D microenvironment

Delta-like 4 (Dll4), a membrane-bound Notch ligand, plays a fundamental role in vascular development and angiogenesis. Dll4 is highly expressed in capillary endothelial tip cells and is involved in suppressing neighboring stalk cells to become tip cells during angiogenesis. Dll4-Notch signaling is mediated either by direct cell-cell contact or by Dll4-containing exosomes from a distance. However, whether Dll4-containing exosomes influence tip cells of existing capillaries is unknown. Using a 3D microfluidic device and time-lapse confocal microscopy, we show here for the first time that Dll4-containing exosomes causes tip cells to lose their filopodia and trigger capillary sprout retraction in collagen matrix. We demonstrate that Dll4 exosomes can freely travel through 3D collagen matrix and transfer Dll4 protein to distant tip cells. Upon reaching endothelial sprout, it causes filopodia and tip cell retraction. Continuous application of Dll4 exosomes from a distance lead to significant reduction of sprout formation. This effect correlates with Notch signaling activation upon Dll4-containing exosome interaction with recipient endothelial cells. Furthermore, we show that Dll4-containing exosomes increase endothelial cell motility while suppressing their proliferation. These data revealed novel functions of Dll4 in angiogenesis through exosomes.

Mathematical model of gas plasma applied to chronic wounds

Chronic wounds are a major burden for worldwide health care systems, and patients suffer pain and discomfort from this type of wound. Recently gas plasmas have been shown to safely speed chronic wounds healing. In this paper, we develop a deterministic mathematical model formulated by eight-species reaction-diffusion equations, and use it to analyze the plasma treatment process. The model follows spatial and temporal concentration within the wound of oxygen, chemoattractants, capillary sprouts, blood vessels, fibroblasts, extracellular matrix material, nitric oxide (NO), and inflammatory cell. Two effects of plasma, increasing NO concentration and reducing bacteria load, are considered in this model. The plasma treatment decreases the complete healing time from 25 days (normal wound healing) to 17 days, and the contributions of increasing NO concentration and reducing bacteria load are about 1/4 and 3/4, respectively. Increasing plasma treatment frequency from twice to three times per day accelerates healing process. Finally, the response of chronic wounds of different etiologies to treatment with gas plasmas is analyzed.

A Urokinase Receptor–Derived Peptide Inhibiting VEGF-Dependent Directional Migration and Vascular Sprouting

The receptor for the urokinase-type plasminogen activator (uPAR) is a widely recognized master regulator of cell migration, and uPAR₈₈₋₉₂ is the minimal sequence required to induce cell motility. We previously showed that soluble forms of uPAR elicit angiogenic responses through their uPAR₈₈₋₉₂ chemotactic sequence and that the synthetic peptide SRSRY exerts similar effects. By a drug design approach, based on the conformational analysis of the uPAR₈₈₋₉₂ sequence, we developed peptides (pERERY, RERY, and RERF) that potently inhibit signaling triggered by uPAR₈₈₋₉₂. In this study, we present evidence that these peptides are endowed also with a clear-cut antiangiogenic activity, although to different extents. The most active, RERF, prevents tube formation by human endothelial cells exposed to SRSRY. RERF also inhibits VEGF-triggered endothelial cell migration and cord-like formation in a dose-dependent manner, starting in the femtomolar range. RERF prevents F-actin polymerization, recruitment of αvβ3 integrin at focal adhesions, and αvβ3/VEGFR2 complex formation in endothelial cells exposed to VEGF. At molecular level, the inhibitory effect of RERF on VEGF signaling is shown by the decreased amount of phospho-FAK and phospho-Akt in VEGF-treated cells. In vivo, RERF prevents VEGF-dependent capillary sprouts originating from the host vessels that invaded angioreactors implanted in mice and neovascularization induced by subcorneal implantation of pellets containing VEGF in rabbits. Consistently, RERF reduced the growth and vascularization rate of tumors formed by HT1080 cells injected subcutaneously in the flanks of nude mice, indicating that RERF is a promising therapeutic agent for the control of diseases fuelled by excessive angiogenesis such as cancer.

Reviews Neoangiogenesis in prostate cancer

Prostate cancer has the second highest incidence among malignant tumors in men in Poland, and the first in the USA. Currently, medical reports state that incidentally non-diagnosed prostate cancer is present in about 30–46% of men over the age of fifty.Tumor angiogenesis has significant implications in the diagnosis and treatment of various solid tumors. With fast, multi-slice CT scanners and their ability of qualitative and quantitative analysis of tumor angiogenesis, CT perfusion has been the subject of extensive investigation in the past twenty years.Tumor angiogenesis is characterized morphologically by an increase in the number of blood vessels including new capillaries, capillary sprouts, non-endothelialized capillaries and arterio-venous shunts.It is stated in the literature that pathological vessel density within prostate cancer is one of the factors determining the extent of the tumor – whether the tumor is confined to the prostate gland or extends beyond the prostate capsule. It was proved that the density of pathological vessels is higher in patients with tumors beyond the prostate capsule.Initial publications show validity of functional imaging, such as perfusion CT in prostate cancer staging before a surgical procedure. This examination can also show excessive density of pathological vessels within the prostate gland according to increased blood volume (BV) and blood flow (BF) and pathologically increased permeability of the vessels (PS). Vessel abnormalities in the examined area are also indicated by shorter mean transit time (MTT).

Wound healing modeling: investigating ambient gas plasma treatment efficacy

Chronic wounds are thought to be caused, in part, by the presence and persistence of aerobic microbes that deplete the local oxygen concentration and prevent or slow the rate of oxygen-dependent healing. Atmospheric-pressure gas plasmas have been shown to be strong bactericidal agents and there is evidence that plasma treatment can safely kill bacteria in wounds and speed wound healing. In this study, we adapted a six-species reaction–diffusion model of epithelial wound healing and used it to predict the efficacy of various plasma treatment protocols. We assume that the only effect of plasma application to the wound is to reduce the bacterial load and that this in turn reduces the bacterial oxygen consumption in the wound. The model follows the spatial and temporal concentration or density profiles within the wound of oxygen, chemoattractants, capillary sprouts, blood vessels, fibroblasts and extracellular matrix material. We highlight the importance of the effects of plasma application on the rate of bacterial regrowth in the wound. Even a relatively large initial reduction in the bacterial wound population may not be sufficient for improved healing if bacterial regrowth is not limited. Although it is clear that current efforts to model wound healing in general and the effects of plasma in particular are in their early stage, the present results suggest several important directions for coupling plasma models with models of tissue biochemical responses.

Attenuation of EphrinB2 Reverse Signaling Decreases Vascularized Area and Preretinal Vascular Tuft Formation in the Murine Model of Oxygen-Induced Retinopathy

EphB4 and ephrinB2 are known key regulators of retinal vascular development, but due to their capacity for bidirectional signaling, delineation of their individual roles in this process remains unclear. To better dissect out individual contributions, a model of proliferative retinopathy in mice with attenuated ephrinB2 reverse signaling was studied. It was hypothesized that endothelial ephrinB2 reverse signaling regulates hypoxia-induced capillary sprouting, as well as the pathologic formation of neovascular tufts in postnatal retinal microvascular networks. Genetically manipulated mice with attenuated ephrinB2 reverse signaling (ephrinB2(lacZ/+)), along with wild-type (WT) controls, were exposed to oxygen-induced retinopathy (OIR), a postnatal model of proliferative retinopathy. At peak disease (postnatal day 18), microvascular networks were analyzed to examine intraretinal revascularization, capillary sprouting, and pathologic neovascularization responses. EphB4 and phosphorylated ephrinB protein expression patterns along retinal microvessels were also assessed. EphrinB2(lacZ/+) mice exhibited reduced hypoxia-induced revascularization (P ≤ 0.04) and reduced formation of neovascular tufts (P < 0.001), as compared with WT controls. Corresponding to the observed inhibition of retinal angiogenesis, ephrinB2(lacZ/+) retinas displayed an increased number of blind-ended capillary sprout tips (P < 0.02) and endothelial filopodial processes (P = 0.001). In WT and ephrinB2(lacZ/+) OIR-exposed retinas, ephrinB was confined to endothelial cells, with expression detected along angiogenic vascular processes including neovascular tufts and blind-ended capillary sprouts. EphrinB2 reverse signaling is a regulator of key processes during retinal vascularization and controls pathologic retinal angiogenesis through direct effects on capillary sprouting and endothelial filopodia formation.

Effect of rosiglitazone on capillary density and angiogenesis in adipose tissue of normoglycaemic humans in a randomised controlled trial.

Recent reports of decreased capillary density in the adipose tissue of obese individuals suggest that an imbalance of angiogenesis and adipogenesis may, in part, underlie insulin resistance. This study aimed to determine whether the insulin-sensitising peroxisome proliferator-activated receptor γ (PPARγ) activator rosiglitazone affects adipose tissue vascularisation in normal humans. A randomised, parallel-group, investigator-blinded placebo-controlled trial was conducted with normoglycaemic volunteers with BMI 27-43, recruited from the community at the University of Massachusetts Medical School, Worcester, MA, USA. Peri-umbilical adipose tissue biopsies were obtained before and after treatment for 6 weeks with rosiglitazone (8 mg once daily) or placebo, which were randomly allocated from a sequentially numbered list. The primary outcomes were adipocyte size and capillary density measured by immunohistochemistry, and angiogenic potential assessed by capillary sprout formation in Matrigel. Secondary outcomes were serum adiponectin, glycaemic, lipid and liver function variables. A total of 35 individuals fulfilling the inclusion criteria were randomised, and complete before-vs-after analyses were achieved in 30 participants (13 and 17, placebo and rosiglitazone, respectively). Significant differences, assessed by paired two-tailed Student t tests, were seen in response to rosiglitazone for adipocyte size (3,458 ± 202 vs 2,693 ± 223 μm(2), p = 0.0049), capillary density (5.6 ± 0.5 vs 7.5 ± 0.5 lumens/field, p = 0.0098), serum adiponectin (14.3 ± 1.5 vs 28.6 ± 3.0 ng/ml, p < 0.0001) and alkaline phosphatase (1.04 ± 0.07 vs 0.87 ± 0.05 μkat/l, p = 0.001). A difference in angiogenic potential before and after treatment between the placebo and rosiglitazone groups was also seen (-23.88 ± 14 vs 13.42 ± 13, p = 0.029, two-tailed Mann-Whitney test). Significant effects on adipose tissue vascular architecture occur after a short period of treatment with rosiglitazone in individuals with normal glucose tolerance. Improved adipose tissue vascularisation may, in part, mediate the therapeutic actions of this class of drugs. ClinicalTrials.gov NCT01150981 The study was funded by National Institutes of Health grant DK089101 to S. Corvera, and by pilot funding from the University of Massachusetts (UMASS) Center for Clinical Translational Sciences (M. Thompson, S. Malkani and S. Corvera). Morphology core services were supported by UMASS Diabetes Endocrine Research Center (DERC) grant DK32520.

Cell proliferation along vascular islands during microvascular network growth

BackgroundObservations in our laboratory provide evidence of vascular islands, defined as disconnected endothelial cell segments, in the adult microcirculation. The objective of this study was to determine if vascular islands are involved in angiogenesis during microvascular network growth.ResultsMesenteric tissues, which allow visualization of entire microvascular networks at a single cell level, were harvested from unstimulated adult male Wistar rats and Wistar rats 3 and 10 days post angiogenesis stimulation by mast cell degranulation with compound 48/80. Tissues were immunolabeled for PECAM and BRDU. Identification of vessel lumens via injection of FITC-dextran confirmed that endothelial cell segments were disconnected from nearby patent networks. Stimulated networks displayed increases in vascular area, length density, and capillary sprouting. On day 3, the percentage of islands with at least one BRDU-positive cell increased compared to the unstimulated level and was equal to the percentage of capillary sprouts with at least one BRDU-positive cell. At day 10, the number of vascular islands per vascular area dramatically decreased compared to unstimulated and day 3 levels.ConclusionsThese results show that vascular islands have the ability to proliferate and suggest that they are able to incorporate into the microcirculation during the initial stages of microvascular network growth.