RESUMEN

The delivery of bone marrow-derived cells (BMDCs) has been widely used to stimulate angiogenesis and arteriogenesis. We identified a progenitor-enriched subpopulation of BMDCs that is able to augment venular remodeling, a generally unexplored area in microvascular research. Two populations of BMDCs, whole bone marrow (WBM) and Lin(-)/Sca-1(+) progenitor cells, were encapsulated in sodium alginate and delivered to a mouse dorsal skinfold chamber model. Upon observation that encapsulated Sca-1(+) progenitor cells enhance venular remodeling, the cells and tissue were analyzed on structural and molecular levels. Venule walls were thickened and contained more nuclei after Sca-1(+) progenitor cell delivery. In addition, progenitors expressed mRNA transcript levels of chemokine (C-X-C motif) ligand 2 (CXCL2) and interferon gamma (IFNγ) that are over 5-fold higher compared to WBM. Tissues that received progenitors expressed significantly higher protein levels of vascular endothelial growth factor (VEGF), monocyte chemotactic protein-1 (MCP-1), and platelet derived growth factor-BB (PDGF-BB) compared to tissues that received an alginate control construct. Nine days following cell delivery, tissue from progenitor recipients contained 39% more CD45(+) leukocytes, suggesting that these cells may enhance venular remodeling through the modulation of the local immune environment. Results show that different BMDC populations elicit different microvascular responses. In this model, Sca-1(+) progenitor cell-derived CXCL2 and IFNγ may mediate venule enlargement via modulation of the local inflammatory environment.

Asunto(s)

Células de la Médula Ósea/citología , Células Madre/citología , Vénulas/citología , Animales , Diferenciación Celular/fisiología , Células Cultivadas , Perfilación de la Expresión Génica , Masculino , Ratones , Ratones Endogámicos C57BL , Neovascularización Fisiológica , Células Madre/metabolismo , Ingeniería de Tejidos

RESUMEN

Using eight newly generated models relevant to addiction, Alzheimer's disease, cancer, diabetes, HIV, heart disease, malaria, and tuberculosis, we show that systems analysis of small (4-25 species), bounded protein signaling modules rapidly generates new quantitative knowledge from published experimental research. For example, our models show that tumor sclerosis complex (TSC) inhibitors may be more effective than the rapamycin (mTOR) inhibitors currently used to treat cancer, that HIV infection could be more effectively blocked by increasing production of the human innate immune response protein APOBEC3G, rather than targeting HIV's viral infectivity factor (Vif), and how peroxisome proliferator-activated receptor alpha (PPARα) agonists used to treat dyslipidemia would most effectively stimulate PPARα signaling if drug design were to increase agonist nucleoplasmic concentration, as opposed to increasing agonist binding affinity for PPARα. Comparative analysis of system-level properties for all eight modules showed that a significantly higher proportion of concentration parameters fall in the top 15th percentile sensitivity ranking than binding affinity parameters. In infectious disease modules, host networks were significantly more sensitive to virulence factor concentration parameters compared to all other concentration parameters. This work supports the future use of this approach for informing the next generation of experimental roadmaps for known diseases.

Asunto(s)

Enfermedad , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Modelos Biológicos , Transducción de Señal , Simulación por Computador , Humanos , Análisis de Sistemas , Biología de Sistemas/métodos

RESUMEN

OBJECTIVE: Early alterations in the skeletal muscle microvasculature may contribute to the onset and progression of type 2 diabetes (DM2) by limiting insulin and glucose availability to skeletal muscle. Microvascular alterations reported with DM2 are numerous and include impaired endothelium-mediated vasodilation, increased arteriole wall stiffness, and decreased capillary density. Most previous analyses of skeletal muscle microvascular architecture have been limited to skeletal muscle cross sections and thus have not presented an integrated, quantitative analysis of the relative significance of observed alterations to elevated microvascular network resistance and decreased blood flow. In this work, we tested the hypothesis that the onset of diabetes would influence microvascular architecture in a manner that would significantly increase capillary network resistance and reduce blood flow. METHODS AND RESULTS: In whole-mount spinotrapezius muscle capillary networks from Zucker diabetic fatty (ZDF) rats before and after the onset of DM2, we found a significant 37% decrease in microvascular branching and a 19% decrease in microvessel length density associated with the onset of the disease. This was previously indiscernible in skeletal muscle cross-section data. Hemodynamic computational analysis revealed that the changes in DM2 capillary network connectivity result in a significant 44% decrease in computed capillary network flow compared to controls. A hemodynamic sensitivity analysis showed that DM2 networks were predicted to be less robust in their ability to maintain perfused network surface area in the event of upstream terminal arteriole constriction. CONCLUSIONS: This study illustrates that capillary network connectivity is altered by DM2 and this negatively impacts microvascular hemodynamics. This work can serve as a basis for a more quantitative approach to evaluating DM2 microvascular networks and their potential use as an early diagnostic aid and/or method for identifying therapeutic targets.

Asunto(s)

Capilares/fisiopatología , Diabetes Mellitus Experimental/fisiopatología , Diabetes Mellitus Tipo 2/fisiopatología , Microcirculación , Músculo Esquelético , Resistencia Vascular , Animales , Velocidad del Flujo Sanguíneo , Femenino , Masculino , Músculo Esquelético/irrigación sanguínea , Músculo Esquelético/fisiopatología , Ratas , Ratas Zucker

RESUMEN

OBJECTIVE: The canonical Wnt signaling pathway, heavily studied in development and cancer, has recently been implicated in microvascular growth with the use of developmental and in vitro models. To date, however, no study exists showing the effects of perturbing the canonical Wnt pathway in a complete microvascular network undergoing physiological remodeling in vivo. Our objective was to investigate the effects of canonical Wnt inhibition on the microvascular remodeling of adult rats. METHODS: Canonical Wnt inhibitor DKK-1, Wnt inhibitor sFRP-1, BSA or saline was superfused onto the exteriorized mesenteric windows of 300 g adult female Sprague-Dawley rats for 20 minutes. Three days following surgery, mesenteric windows were imaged intravitally and harvested for immunofluorescence staining with smooth muscle alpha-actin and BRDU. RESULTS: We observed prominent differences in the response of the mesenteric microvasculature amongst the various treatment groups. Significant increases in hemorrhage area, vascular density, and draining vessel diameter were observed in windows treated with Wnt inhibitors as compared to control-treated windows. Additionally, confocal imaging analysis showed significant increases in proliferating cells as well as evidence of proliferating smooth muscle cells along venules. CONCLUSIONS: Together, our results suggest that canonical Wnt inhibition plays an important role in microvascular remodeling, specifically venular remodeling.

Asunto(s)

Hemorragia/etiología , Microcirculación/fisiología , Proteínas Wnt/antagonistas & inhibidores , Animales , Proliferación Celular/efectos de los fármacos , Femenino , Hemorragia/patología , Hemorragia/fisiopatología , Péptidos y Proteínas de Señalización Intercelular/farmacología , Péptidos y Proteínas de Señalización Intercelular/fisiología , Proteínas de la Membrana/farmacología , Proteínas de la Membrana/fisiología , Venas Mesentéricas/efectos de los fármacos , Venas Mesentéricas/patología , Venas Mesentéricas/fisiopatología , Microcirculación/efectos de los fármacos , Modelos Cardiovasculares , Neovascularización Fisiológica/efectos de los fármacos , Ratas , Ratas Sprague-Dawley , Proteínas Recombinantes/farmacología , Transducción de Señal/efectos de los fármacos , Vénulas/efectos de los fármacos , Vénulas/patología , Vénulas/fisiopatología , Proteínas Wnt/fisiología

RESUMEN

OBJECTIVE: Bone marrow-derived cells (BMCs) and inflammatory chemokine receptors regulate arteriogenesis and angiogenesis. Here, we tested whether arteriolar remodeling in response to an inflammatory stimulus is dependent on BMC-specific chemokine (C-C motif) receptor 2 (CCR2) expression and whether this response involves BMC transdifferentiation into smooth muscle. METHODS AND RESULTS: Dorsal skinfold window chambers were implanted into C57Bl/6 wild-type (WT) mice, as well as the following bone marrow chimeras (donor-host): WT-WT, CCR2(-/-)-WT, WT-CCR2(-/-), and EGFP(+)-WT. One day after implantation, tissue MCP-1 levels rose from "undetectable" to 463 pg/mg, and the number of EGFP(+) cells increased more than 4-fold, indicating marked inflammation. A 66% (28 microm) increase in maximum arteriolar diameter was observed over 7 days in WT-WT mice. This arteriolar remodeling response was completely abolished in CCR2(-/-)-WT mice but largely rescued in WT-CCR2(-/-) mice. EGFP(+) BMCs were numerous throughout the tissue, but we found no evidence that EGFP(+) BMCs transdifferentiate into smooth muscle, based on examination of >800 arterioles and venules. CONCLUSIONS: BMC-specific CCR2 expression is required for injury/inflammation-associated arteriolar remodeling, but this response is not characterized by the differentiation of BMCs into smooth muscle.

Asunto(s)

Arteriolas/fisiología , Células de la Médula Ósea/metabolismo , Miocitos del Músculo Liso/metabolismo , Receptores CCR2/metabolismo , Regeneración/fisiología , Análisis de Varianza , Animales , Células de la Médula Ósea/citología , Trasplante de Médula Ósea , Proliferación Celular , Células Cultivadas , Modelos Animales de Enfermedad , Regulación de la Expresión Génica , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Miocitos del Músculo Liso/citología , Neovascularización Fisiológica/fisiología , Probabilidad , Distribución Aleatoria , Receptores CCR2/genética

RESUMEN

External application of static magnetic fields (SMF), used specifically for the treatment of inflammatory conditions such as soft tissue injuries, has recently become popular as a complementary and/or alternative therapy with minimal investigation into efficacy or mechanism. Localized inflammation was induced via injection of inflammatory agents lambda-carrageenan (CA) or histamine into rat hindpaws, alone or in conjunction with pharmacological agents, resulting in a spatially and temporally defined inflammatory reaction. Application of a 10- or 70-mT, but not a 400-mT, SMF for 15 or 30 min immediately following histamine-induced edema resulted in a significant, 20-50% reduction in edema formation. In addition, a 2-h, 70-mT field application to CA-induced edema also resulted in significant (33-37%) edema reduction. Field application before injection or at the time of maximal edema did not influence edema formation or resolution, respectively. Together, these results suggest the existence of a therapeutic threshold of SMF strength (below 400 mT) and a temporal dependence of efficacy. Administration of pharmacological agents directed at nitric oxide signaling and L-type Ca(2+) channel dynamics in conjunction with SMF treatment and histamine-induced edema revealed that the potential mechanism of SMF action may be via modulation of vascular tone through effects on L-type Ca(2+) channels in vascular smooth muscle cells.

Asunto(s)

Edema/prevención & control , Campos Electromagnéticos , Inflamación/prevención & control , Músculo Liso Vascular/efectos de la radiación , Ácido 3-piridinacarboxílico, 1,4-dihidro-2,6-dimetil-5-nitro-4-(2-(trifluorometil)fenil)-, Éster Metílico/farmacología , Animales , Arginina/metabolismo , Calcio/metabolismo , Agonistas de los Canales de Calcio/farmacología , Canales de Calcio Tipo L/efectos de los fármacos , Canales de Calcio Tipo L/metabolismo , Carragenina , Modelos Animales de Enfermedad , Relación Dosis-Respuesta en la Radiación , Edema/inducido químicamente , Edema/metabolismo , Histamina , Inflamación/inducido químicamente , Inflamación/metabolismo , Masculino , Músculo Liso Vascular/efectos de los fármacos , Músculo Liso Vascular/enzimología , Músculo Liso Vascular/metabolismo , Óxido Nítrico/metabolismo , Óxido Nítrico Sintasa/metabolismo , Ratas , Ratas Sprague-Dawley , Índice de Severidad de la Enfermedad , Transducción de Señal/efectos de la radiación , Factores de Tiempo

RESUMEN

Magnetic field therapy has recently become a widely used complementary/alternative medicine for the treatment of vascular, as well as other musculoskeletal pathologies, including soft tissue injuries. Recent studies in our laboratory and others have suggested that acute static magnetic field (SMF) exposure can have a modulatory influence on the microvasculature, acting to normalize vascular function; however, the effect of chronic SMF exposure has not been investigated. This study aimed to measure, for the first time, the adaptive microvascular response to a chronic 7-day continuous magnetic field exposure. Murine dorsal skinfold chambers were applied on day 0, and neodymium static magnets (or size and weight-matched shams) were affixed to the chambers at day 0, where they remained until day 7. Separate analysis of arteriolar and venular diameters revealed that chronic SMF application significantly abrogated the luminal diameter expansion observed in sham-treated networks. Magnet-treated venular diameters were significantly reduced at day 4 and day 7 (34.3 and 54.4%, respectively) compared with sham-treated vessels. Arteriolar diameters were also significantly reduced by magnet treatment at day 7 (50%), but not significantly at day 4 (31.6%), although the same trend was evident. Venular functional length density was also significantly reduced (60%) by chronic field application. These results suggest that chronic SMF exposure can alter the adaptive microvascular remodeling response to mechanical injury, thus supporting the further study of chronic application of SMFs for the treatment of vascular pathologies involving the dysregulation of microvascular structure.

Asunto(s)

Arteriolas/anatomía & histología , Magnetismo , Vénulas/anatomía & histología , Animales , Arteriolas/fisiología , Arteriolas/cirugía , Masculino , Ratones , Ratones Endogámicos C57BL , Microcirculación/fisiología , Flujo Sanguíneo Regional/fisiología , Piel/irrigación sanguínea , Vasodilatación/fisiología , Vénulas/fisiología , Vénulas/cirugía

RESUMEN

OBJECTIVE: Recently the authors have shown that neuron-glial antigen 2 (NG2) is expressed by perivascular cells along arterioles and capillaries, but not along venules in quiescent rat mesenteric microvascular networks. To investigate how the spatial distribution of this proteoglycan changes during microvascular remodeling, the objective of this study was to characterize the expression of NG2 in adult rat mesenteric microvascular networks undergoing active remodeling. METHODS: The distribution of NG2 expression was evaluated in adult rat mesenteric microvascular networks. Tissues were harvested from 250 g, female, Sprague-Dawley rats at 1, 3, and 5 days poststimulation and double immunolabeled for NG2 and CD31 (endothelial cell marker). RESULTS: After 1 day, NG2 expression was observed along 27 +/- 11% of network draining venules (14-55 microm) and after 3 days, 59 +/- 10% of draining venules (13-59 microm) stained positive for the proteoglycan. By 5 days poststimulation, the percentage of network draining venules (18-59 microm) staining positive for NG2 returned to 18 +/- 7%, indicating a downregulation of the proteoglycan toward quiescent levels along larger-sized venules. CONCLUSIONS: The results suggest that NG2 proteoglycan expression is transiently upregulated along venules during microvascular remodeling, implicating NG2 as a marker of activated venules.

Asunto(s)

Antígenos/genética , Pericitos/metabolismo , Proteoglicanos/genética , Regeneración , Vénulas/fisiología , Animales , Femenino , Microcirculación , Ratas , Ratas Sprague-Dawley , Circulación Esplácnica , Factores de Tiempo , Regulación hacia Arriba/genética

RESUMEN

Bone marrow-derived cells (BMCs) have been implicated as a modifiers of vascular growth either directly by transdifferentiation into endothelial cells (ECs) or indirectly through growth factor release. To examine these possibilities under physiological conditions, we developed a model of hypoxia-mediated angiogenesis in the mouse spinotrapezius muscle. This allows whole-mount analysis; therefore, the morphology and location of BMCs within the vascular network may be observed along with differentiation markers. We exposed bone marrow transplant chimeric mice to hypoxia and treated a subset with granulocyte macrophage colony-stimulating factor. Exposure to hypoxia caused an 13% increase in capillary density relative to control. Hypoxia did not increase the overall number of muscle-resident BMCs, but did increase the number of rounded BMCs by 25%. There was no discernable BMC contribution to the endothelium, although some BMCs assumed a pericyte morphology around capillaries. Granulocyte macrophage colony-stimulating factor treatment further increased the number of round BMCs within the muscle and caused a 23% increase in angiogenesis. The results of this study suggest a potentially beneficial action of BMCs during hypoxia through paracrine release of growth factors but not transdifferentiation into ECs.

Asunto(s)

Células de la Médula Ósea/fisiología , Diferenciación Celular , Células Endoteliales/citología , Movilización de Célula Madre Hematopoyética , Células Madre Hematopoyéticas/fisiología , Hipoxia/fisiopatología , Neovascularización Fisiológica , Animales , Células de la Médula Ósea/citología , Recuento de Células , Células Madre Hematopoyéticas/citología , Hipoxia/patología , Metaloproteinasa 9 de la Matriz/análisis , Ratones , Ratones Endogámicos C57BL , Músculo Esquelético/patología , Músculo Liso Vascular/citología , Factor A de Crecimiento Endotelial Vascular/análisis

RESUMEN

OBJECTIVE: Similar to other vascular pericyte markers, including smooth muscle (SM) alpha-actin, desmin, and PDGF-beta-receptor, NG2 proteoglycan is not pericyte specific. Therefore, the use of NG2 as a pericyte marker, especially in cell lineage studies, in comparison to other nonspecific pericyte markers requires an understanding of how its expression varies spatially within a microvascular network. The objective of this study was to characterize NG2 expression along vessels within rat microvascular networks and compare this to SM alpha-actin expression. METHODS: Mesenteric tissue, subcutaneous tissue, spinotrapezius muscle, and gracilis muscle were harvested from 250-g, female, Sprague-Dawley rats and stained for NG2 and SM alpha-actin. The distribution of NG2 expression was evaluated in mesenteric networks (n = 28) with complementary observations in subcutaneous tissue and skeletal muscle. RESULTS: Perivascular cells, including mature smooth muscle cells (SMCs), immature SMCs, and pericytes, expressed NG2. Most importantly, NG2 expression was primarily confined to perivascular cells along arterioles and capillaries, and continuous expression was not observed along venules beyond the immediate postcapillary vessels. The differential expression of NG2 along the arteriolar side of microvascular networks was also observed in rat subcutaneous and skeletal muscle. CONCLUSIONS: The results indicate that NG2 is expressed by all perivascular cells along arterioles, and its absence denotes a venule-specific phenotype. These results identify for the first time a marker that differentiates venous smooth muscle and pericytes from other capillary- and arteriole-associated perivascular cells.

Asunto(s)

Antígenos/genética , Arteriolas/citología , Regulación de la Expresión Génica , Pericitos/química , Proteoglicanos/genética , Vénulas/citología , Actinas/análisis , Actinas/genética , Animales , Antígenos/análisis , Femenino , Humanos , Proteoglicanos/análisis , Ratas , Ratas Sprague-Dawley , Circulación Esplácnica

RESUMEN

Angiogenesis and vessel remodeling determine the integrative control of the architectural structure and functional behaviors of the microcirculation over the lifetime of an organism. Vascular remodeling is the basis of promising therapeutic strategies, including vascularization of ischemic organs. The history of angiogenesis research is long-more than 250 years-and the Microcirculatory Society has been the birthplace of numerous techniques, assays, and scientific concepts that have stimulated massive research endeavors in the pharmaceutical and medical arena. At present, angiogenesis isa dynamic field in which the molecular genetic and proteomic components of the process are still being identified, while integrative systems approaches are once again being recognized as essential to understand microvascular assembly in vivo across multiple scales from cells to whole vessel networks. A short history of people and ideas in this field is presented, followed by discussion of emerging directions receiving intense attention today and major questions that remain unanswered. The primary conclusion is that the need for scientists trained in the integrative approaches nurtured by the Microcirculatory Society over the past 50 years has never been greater, as it is clear that a complete mechanistic understanding of vessel adaptation (based on genomic and proteomic supporting casts) will now require deeper studies of angiogenesis and microvascular remodeling in the exquisite complexity of the native microenvironment-the microcirculation.

Asunto(s)

Microcirculación/fisiología , Neovascularización Fisiológica , Animales , Tipificación del Cuerpo , Historia del Siglo XX , Historia del Siglo XXI , Humanos , Microcirculación/citología , Neovascularización Patológica/terapia , Investigación/historia

RESUMEN

Extremely low-magnitude (0.3 g), high-frequency (30-90 Hz), whole body vibrations can stimulate bone formation and are hypothesized to provide a surrogate for the oscillations of muscle during contraction. Little is known, however, about the potential of these mechanical signals to stimulate adaptive responses in other tissues. The objective of this study was to determine whether low-level mechanical signals produce structural adaptations in the vasculature of skeletal muscle. Eight-week-old male BALB/cByJ (BALB) mice were divided into two experimental groups: mice subjected to low-level, whole body vibrations (45 Hz, 0.3 g) superimposed on normal cage activities for 15 min/day (n = 6), and age-matched controls (n = 7). After the 6-wk experimental protocol, sections from end and mid regions of the soleus muscles were stained with lectin from Bandeiraea Simplicifolia, an endothelial cell marker, and smooth muscle (SM) alpha-actin, a perivascular cell marker. Six weeks of this low-level vibration caused a 29% decrease in the number of lectin-positive vessels per muscle fiber in the end region of the soleus muscle, indicating a significant reduction in the number of capillaries per muscle fibers. Similarly, these vibrations caused a 36% reduction in SM alpha-actin-positive vessels per muscle fiber, indicating a reduction in the number of arterioles and venules. The decreases in lectin- and SM alpha-actin-positive vessels per muscle fiber ratios were not significant in the mid muscle sections. These results demonstrate the sensitivity of the vasculature in mouse skeletal muscle to whole body, low-level mechanical signals.

Asunto(s)

Mecanotransducción Celular/fisiología , Microcirculación/fisiología , Fibras Musculares Esqueléticas/fisiología , Músculo Esquelético/irrigación sanguínea , Músculo Esquelético/fisiología , Neovascularización Fisiológica/fisiología , Estimulación Física/métodos , Adaptación Fisiológica/fisiología , Animales , Masculino , Ratones , Ratones Endogámicos BALB C , Microcirculación/citología , Fibras Musculares Esqueléticas/citología , Músculo Esquelético/citología , Vibración

RESUMEN

In the blastocoel roof (BCR) of the Xenopus laevis embryo, epibolic movements are driven by the radial intercalation of deep cell layers and the coordinate spreading of the overlying superficial cell layer. Thinning of the lateral margins of the BCR by radial intercalation requires fibronectin (FN), which is produced and assembled into fibrils by the inner deep cell layer of the BCR. A cellular automata (CA) computer model was developed to analyze the spatial and temporal movements of BCR cells during epiboly. Simulation parameters were defined based on published data and independent results detailing initial tissue geometry, cell numbers, cell intercalation rates, and migration rates. Hypotheses regarding differential cell adhesion and FN assembly were also considered in setting system parameters. A 2-dimensional model simulation was developed that predicts BCR thinning time of 4.8 h, which closely approximates the time required for the completion of gastrulation in vivo. Additionally, the model predicts a temporal increase in FN matrix assembly that parallels fibrillogenesis in the embryo. The model is capable of independent predictions of cell rearrangements during epiboly, and here was used to predict successfully the lateral dispersion of a patch of cells implanted in the BCR, and increased assembly of FN matrix following inhibition of radial intercalation by N-cadherin over-expression.

Asunto(s)

Simulación por Computador , Gástrula/fisiología , Modelos Biológicos , Xenopus laevis/embriología , Animales , Cadherinas/metabolismo , Adhesión Celular/fisiología , Movimiento Celular/fisiología , Fibronectinas/metabolismo , Gástrula/ultraestructura , Microscopía Fluorescente , Morfogénesis , Factores de Tiempo

RESUMEN

Remodeling of microvascular networks in mammals is critical for physiological adaptations and therapeutic revascularization. Cellular behaviors such as proliferation, differentiation, and migration are coordinated in these remodeling events via combinations of biochemical and biomechanical signals. We developed a cellular automata (CA) computational simulation that integrates epigenetic stimuli, molecular signals, and cellular behaviors to predict microvascular network patterning events. Over 50 rules obtained from published experimental data govern independent behaviors (including proliferation, differentiation, and migration) of thousands of interacting cells and diffusible growth factors in their tissue environment. From initial network patterns of in vivo blood vessel networks, the model predicts emergent patterning responses to two stimuli: 1) network-wide changes in hemodynamic mechanical stresses, and 2) exogenous focal delivery of an angiogenic growth factor. The CA model predicts comparable increases in vascular density (370+/-29 mm/mm3) 14 days after treatment with exogenous growth factor to that in vivo (480+/-41 mm/mm3) and approximately a twofold increase in contractile vessel lengths 5-10 days after 10% increase in circumferential wall strain, consistent with in vivo results. The CA simulation was thus able to identify a functional patterning module capable of quantitatively predicting vessel network remodeling in response to two important epigenetic stimuli.

Asunto(s)

Biología Computacional/métodos , Microcirculación/anatomía & histología , Diferenciación Celular , División Celular , Movimiento Celular , Simulación por Computador , Sustancias de Crecimiento/farmacología , Hemodinámica , Humanos , Microcirculación/citología , Microcirculación/metabolismo , Modelos Cardiovasculares , Transducción de Señal , Estrés Mecánico , Ingeniería de Tejidos

RESUMEN

Microvascular networks undergo patterning changes that determine and reflect functional adaptations during tissue remodeling. Alterations in network architectures are a result of complex and integrated signaling events. To understand how two growth factor signals interact to stimulate angiogenesis and arterialization, we engineered spatially directed microvascular pattern changes in vivo by using combinations of focally delivered exogenous growth factors. We implanted microdelivery beads containing recombinant vascular endothelial growth factor-164 (VEGF(164)) and recombinant angiopoietin-1* (Ang-1*) into the dorsal subcutaneous tissue of fully anesthetized male Fischer 344 rats implanted with backpack window chambers, and we quantified vascular patterning changes by using intravital microscopy, a combination of architectural metrics, and immunohistochemistry. Focal delivery of VEGF(164) caused spatially directed increases in both the total number and the density of vessels with diameters <25 microm 7 days after microbead implantation. Increases were maintained out to 14 days but were reduced to control values by day 21. The addition of Ang-1* on day 7 maintained these increases out to day 21, induced vessel order ratios comparable to control levels, and was accompanied by increases in the length density of smooth muscle alpha-actin-positive vessels. We achieved spatial control of patterning changes in vivo by using multisignal stimulation via focal delivery of exogenous growth factor combinations and conclude that Ang-1* administered subsequent to VEGF(164) stimulation induces vascular growth while maintaining a network pattern consistent with native patterns that persist in the presence of vehicle control stimulation.

Asunto(s)

Angiopoyetina 1/farmacología , Neovascularización Fisiológica/efectos de los fármacos , Piel/irrigación sanguínea , Factor A de Crecimiento Endotelial Vascular/farmacología , Alginatos , Animales , Difusión , Sistemas de Liberación de Medicamentos , Ácido Glucurónico , Ácidos Hexurónicos , Radioisótopos de Yodo , Masculino , Microcirculación/efectos de los fármacos , Microesferas , Ratas , Ratas Endogámicas F344

RESUMEN

The objective of this study was to quantify the proliferation of existing vascular and perivascular cells during a specific form of microvascular remodeling characterized by increased coverage by smooth muscle cells (SMCs), in response to increased mechanical stress. Coordinated ligations of artery/vein pairs in the rat mesentery resulted in hemodynamic stress elevations within the targeted microvascular network. BRDU incorporation per unit length of smooth muscle (SM) alpha-actin positive vessel was evaluated following ligation at 2, 5, and 10 days. At 2 days, BRDU incorporation was significantly increased for both sham and ligated treatments, but the ligated response was not elevated over the sham response. After 5 days, proliferation for both groups returned to unstimulated levels. The results indicate that moderate elevations in hemodynamic stress do not cause perivascular cell proliferation along rat mesenteric microvessels, therefore, the increased coverage of differentiated SMCs along the same microvessels does not involve proliferation of vascular or perivascular cells.

Asunto(s)

Actinas/biosíntesis , Proliferación Celular , Arterias Mesentéricas/fisiología , Mesenterio/fisiología , Miocitos del Músculo Liso/metabolismo , Animales , Femenino , Arterias Mesentéricas/citología , Mesenterio/irrigación sanguínea , Ratas , Ratas Sprague-Dawley , Estrés Mecánico , Resistencia Vascular/fisiología

RESUMEN

Angiogenesis, the arterialization of capillaries, and arteriogenesis are specific manifestations of the complex continuum of blood vessel-remodeling processes that are produced by environmental stimuli. Together, they determine the integrative control of vascular assembly and pattern formation. Vascular assembly and pattern formation are critical elements of therapeutic vascular collateralization of progressively ischemic organs and in the tissue engineering or organogenesis of various tissue substitutes. An integrative systems approach is useful to measure the dynamics of vascular assembly in vivo across time scales from the embryo to the adult, and spanning spatial scales from cells to whole networks, to understand the complex interplay of multiple interacting cells and signal molecules. This requires in vivo observations, multiscale computer simulations, and tools for the genetic regulation of cell interactions. The new view of vascular remodeling as a continuum that can be manipulated in various tissues and in different size blood vessels, using appropriately coordinated multisignal stimuli, should open new therapeutic avenues.

Asunto(s)

Capilares/fisiología , Neovascularización Fisiológica/fisiología , Animales , Arterias/crecimiento & desarrollo , Linaje de la Célula , Circulación Colateral/fisiología , Simulación por Computador , Humanos , Isquemia/fisiopatología , Modelos Biológicos , Morfogénesis , Neoplasias/irrigación sanguínea , Neovascularización Patológica/fisiopatología

RESUMEN

During vascular remodeling in adult organisms, new capillary growth is often coupled with the adaptation of arterioles and venules, a process that requires the recruitment and differentiation of precursor cells into smooth muscle. We studied the in vivo adaptation of microvessels in the presence of elevated pressure and circumferential wall stress using a ligation strategy for mesenteric microvascular networks. Acute pressure increases of 42.6+/-18% and 17.1+/-2.3% were respectively elicited in the 25- to 30-microm-diameter venules and arterioles supplying the networks. Wall shear rates were not significantly changed; however, diameters were increased in >10-microm-diameter venules and >20-microm-diameter arterioles. Smooth muscle cell contractile phenotype was determined in all microvessels by observing the expression of smooth muscle myosin heavy chain (SM-MHC; a marker of fully differentiated smooth muscle) and smooth muscle alpha-actin (a marker for all smooth muscle, including immature smooth muscle of fibroblast/pericyte lineage). The ratio of SM-MHC positive vessel length to smooth muscle alpha-actin-positive vessel length increased >2-fold after 5 and 10 days of the ligation treatment. Smooth muscle proliferation was studied by bromodeoxyuridine incorporation, and the increase in SM-MHC-labeled microvessel length density was accompanied by no measurable change in proliferation of SM-MHC-labeled cells 5 and 10 days after ligation. These results indicate that after a period of 5 or 10 days, mesenteric microvessels <40 microm in diameter exposed to elevated pressure and wall strain exhibit an enhanced coverage of mature, fully differentiated smooth muscle cells.

Asunto(s)

Vasos Sanguíneos/fisiología , Hemodinámica/fisiología , Músculo Liso Vascular/citología , Estrés Fisiológico/fisiopatología , Actinas/análisis , Animales , Presión Sanguínea/fisiología , División Celular/fisiología , Femenino , Inmunohistoquímica , Mesenterio/irrigación sanguínea , Músculo Liso Vascular/química , Cadenas Pesadas de Miosina/análisis , Ratas , Ratas Sprague-Dawley , Flujo Sanguíneo Regional , Estrés Mecánico

RESUMEN

With the advent of molecular embryology and exploitation of genetic models systems, many genes necessary for normal blood vessel formation during early development have been identified. These genes include soluble effectors and their receptors, as well as components of cell-cell junctions and mediators of cell-matrix interactions. In vitro model systems (2-D and 3-D) to study paracrine and autocrine interactions of vascular cells and their progenitors have also been created. These systems are being combined to study the behavior of genetically altered cells to dissect and define the cellular role(s) of specific genes and gene families in directing the migration, proliferation, and differentiation needed for blood vessel assembly. It is clear that a complex spatial and temporal interplay of signals, including both genetic and environmental, modulates the assembly process. The development of real-time imaging and image analysis will enable us to gain further insights into this process. Collaborative efforts among vascular biologists, biomedical engineers, mathematicians, and physicists will allow us to bridge the gap between understanding vessel assembly in vivo and assembling vessels ex vivo.

Asunto(s)

Vasos Sanguíneos/fisiología , Ingeniería de Tejidos/métodos , Adulto , Animales , Comunicación Celular , Endotelio Vascular/citología , Matriz Extracelular/metabolismo , Humanos , Recién Nacido , Modelos Biológicos , Músculo Liso/citología , Factores de Tiempo

Asunto(s)

Ingeniería Biomédica/métodos , Vasos Sanguíneos/fisiología , Animales , Vasos Sanguíneos/patología , Humanos , Programas Informáticos