The theory of bifurcating vascular systems predicts vessel diameters that are related to optimality criteria like minimization of pumping energy or of building material. However, mechanisms for producing the postulated optimality have not been described so far, and quantitative data on bifurcation diameters during development are scarce. We used an embryonic vascular bed that rapidly grows and adapts to changing hemodynamic conditions, the chicken chorioallantoic membrane (CAM), and correlated vascular cast and tissue section morphology with in vivo time-lapse video monitoring. The bifurcation exponent ⌬ and associated parameters were quantitatively assessed in arterial and venous microvessels ranging in diameter from 30 to 100 m. We observed emergence of optimality by means of intussusception, i.e., formation of transvascular tissue pillars. In addition to intussusceptive microvascular growth (IMG ‫؍‬ expansion of capillary networks) and intussusceptive arborization (IAR ‫؍‬ formation of feeding vessels from capillaries) the observed intussusception at bifurcations represents a third variant of nonsprouting angiogenesis. We call it intussusceptive branching remodeling (IBR). IBR occurred in vessels of considerable diameter by means of two alternative mechanisms: either through pillars arising close to a bifurcation, which increased in girth until they merged with the connective tissue in the bifurcation angle; or through pillars arising at some distance from the bifurcation point, which then expanded by formation of ingrowing tissue folds until they became connected to the tissue of the bifurcation angle. Morphologic evidence suggests that IBR is a wide-spread phenomenon, taking place also in lung, intestinal, kidney, eye, etc., vasculature. Irrespective of the mode followed, IBR led to a branching pattern close to the predicted optimum, ⌬ ‫؍‬ 3.0. Significant differences were observed between ⌬ at arterial bifurcations (2.70 to 2.90) and ⌬ at venous bifurcations (2.93 to 3.75). IBR, by means of eccentric pillar formation and fusion, was also involved in vascular pruning. Experimental changes in CAM hemodynamics (by locally increasing blood flow) induced onset of IBR within less than 1 hr. Our study provides morphologic and quantitative evidence that a similar cellular machinery is used for all three variants of vascular intussusception, IMG, IAR, and IBR. It thus provides a mechanism of efficiently generating complex blood transport systems from limited genetic information. Differential quantitative outcome of IBR in arteries and veins, and the experimental induction of IBR strongly suggest that hemodynamic factors can instruct embryonic vascular remodeling toward optimality.

The origin of vascular pericytes (PCs) and smooth muscle cells (vSMCs) in the brain has hitherto remained an open question. In the present study, we used the quail-chick chimerization technique to elucidate the lineage of cranial PCs/vSMCs. We transplanted complete halves of brain anlagen, or dorsal (presumptive neural crest [NC]) or ventral cranial neural tube. Additional experiments included transplantations of neuroectoderm into limb mesenchyme, and of head mesoderm or limb mesenchyme into paraxial head mesoderm. After interspecific transplantation of quail brain rudiment, graft-derived vSMCs were found in the vessel walls of the grafted brain. Notably, transplanted ventral neural tube also gave rise to vSMCs. After grafting of quail head mesoderm, quail endothelial cells were found in the host brain, but no vSMCs of donor origin. Grafting of quail whole or ventral neural tube into the limb bud led to endowment of graft and host vessels with graft-derived vSMCs. Quail limb bud mesenchyme contributed to vSMCs in the ectopic neural graft, but, when transplanted into paraxial head mesenchyme, it did not form intraneural vSMCs. After orthotopic transplantation of cranial NC, graft-derived vSMCs were not only found in meninges and brain of the operated side, but also on the contralateral side. Our results show that 1) avian cranial neuroectoderm is able to differentiate into vSMCs of the brain; 2) this potential is not restricted to the prospective NC; and 3) neither cranial mesoderm nor cranially transplanted limb bud mesoderm can give rise to brain vSMC.

A spin-crossover coordination polymer [Fe(L1)(bipy)] (where L = a NO coordinating Schiff base-like ligand bearing a phenazine fluorophore and bipy = 4,4'-bipyridine) was synthesized and exhibits a 48 K wide thermal hysteresis above room temperature (T↑ = 371 K and T↓ = 323 K) that is stable for several cycles. The spin transition was characterized using magnetic measurements, Mössbauer spectroscopy, and DSC measurements. T-dependent X-ray powder diffraction reveals a structural phase transition coupled with the spin transition phenomenon. The dimeric excerpt {(μ-bipy)[FeL1(MeOH)]}·2MeOH of the coordination polymer chain crystallizes in the triclinic space group P1̅ and reveals that the packing of the molecules in the crystal is dominated by hydrogen bonds. Investigation of the emission properties of the complexes with regard to temperature shows that the spin crossover can be tracked by monitoring the emission spectra, since the emission color changes from greenish to a yellow color upon the low spin-to-high spin transition.

During most instances of angiogenesis, not only are the capillaries or terminal vessels generated and modified, but the supplying vascular system is subjected to remodeling as well. Intussusception, i.e., transluminal pillar formation, is one essential mechanism for growth, arborization, bifurcation remodeling, and pruning. Complex and efficient vascular beds can thus be generated by local interactions between vascular cells and hemodynamic conditions.

We have studied the effect of VEGF(121) homodimer and VEGF(121/165) heterodimer on the chorioallantoic membrane (CAM) of 13-day-old chick embryos. The factors were applied in doses of 2-4 micrograms and the effects were evaluated macroscopically after 2 and 3 days. Histological studies were performed on semi- and ultrathin sections. Proliferation was studied according to the BrdU-anti-BrdU method on whole mounts and sections. The labeling density was quantified in whole mounts. The fractal dimension, D, of the vascular tree was assessed as a value for vascular bifurcation density. Both forms of VEGF induce brush-like vessel formation in the precapillary region. New capillaries are found in the stroma of the CAM, which normally does not contain capillaries. Our results show that VEGF(121) is a specific endothelial cell mitogen. A fourfold increase of BrdU-labeled endothelial cells is found after VEGF(121) application. The fractal dimension of the vascular tree increases from 1.26 in the controls to 1.44 (VEGF(121)) and 1.41 (VEGF(121/165)). The endothelial cells of the newly formed capillaries possess many mitochondria and micropinocytotic vesicles, but no fenestrations. These capillaries are obviously formed by intussusceptive microvascular growth. Signs of sprouting are almost absent. An effect on the lymphatic vessels of the CAM is not detectable. Compared to VEGF(165) and VEGF(121/165), VEGF(121) diffuses over a slightly greater distance. Using in situ hybridization, VEGF receptor-2 (flk-1/Quek1) and the homologous flt-4 (Quek2) receptor were studied in the CAM of normal quail embryos and after VEGF(121) application on the CAM of 11-day-old quail embryos. During normal development, flk-1 expression becomes restricted to vascular endothelial cells of large vessels in the stroma of the CAM. VEGF(121) application induces expression of flk-1 in capillaries that normally do not express the receptor. In the normal development of the CAM, flt-4 becomes restricted to endothelial cells of vessels that appear to be lymphatic vessels. Application of VEGF(121) does not alter flt-4 expression.

The purpose of this report is to introduce a new computer model for the simulation of microvascular growth and remodeling into arteries and veins that imitates angiogenesis and blood flow in real vascular plexuses. A C؉؉ computer program was developed based on geometric and biophysical initial and boundary conditions. Geometry was defined on a two-dimensional isometric grid by using defined sources and drains and elementary bifurcations that were able to proliferate or to regress under the influence of random and deterministic processes. Biophysics was defined by pressure, flow, and velocity distributions in the network by using the nodal-admittance-matrix-method, and accounting for hemodynamic peculiarities like Fahraeus-Lindqvist effect and exchange with extravascular tissue. The proposed model is the first to simulate interdigitation between the terminal branches of arterial and venous trees. This was achieved by inclusion of vessel regression and anastomosis in the capillary plexus and by remodeling in dependence from hemodynamics. The choice of regulatory properties influences the resulting vascular patterns. The model predicts interdigitating arteriovenous patterning if shear stress-dependent but not pressure-dependent remodeling was applied. By approximating the variability of natural vascular patterns, we hope to better understand homogeneity of transport, spatial distribution of hemodynamic properties and biomass allocation to the vascular wall or blood during development, or during evolution of circulatory systems.

Biological activities of vascular endothelial growth factor (VEGF) have been studied extensively in endothelial cells (ECs), but few data are available regarding its effects on pericytes. In murine embryoid body cultures, VEGF-induced expression of desmin and ␣-smooth muscle actin (␣-SMA) in CD-31 ϩ cells. The number of CD-31 ϩ /desmin ϩ vascular chords increased with VEGF treatment time and peaked during a differentiation window between 6 and 9 days after plating. In vivo, VEGF-induced elongation and migration of desmin-positive pericytes and coverage of angiogenic capillaries, as revealed by analysis of Sambucus nigra lectin-stained vascular beds of the chick chorioallantoic membrane. VEGF also caused significant decrease of intercapillary spaces, an indicator for intussusceptive vascular growth. These VEGF-mediated effects point at a more intricate interaction between ECs and pericytes cells than previously demonstrated and suggest that pericytes may be derived from EC progenitors in vitro and not only stabilize capillaries but also participate in vascular remodeling in vivo.

We studied the early pattern of neural tube (NT) vascularization in quail embryos and chick-quail chimeras. Angioblasts appeared first in the dorsal third at Hamburger and Hamilton (HH) stage 19 as single, migrating cells. Their distribution did not correspond to a segmental pattern. After this initial dorsal immigration, endothelial sprouts invaded the NT on either side of the floor plate (HH stage 21). These cells remained continuous with their arterial vascular sources, connected to the venous perineural vascular plexus at HH-stage 22, and formed the first perfused vessels of the NT at HH-stage 23. The same pattern of angiotrophic vascularization was observed in a craniocaudal sequence starting caudal to the rhombencephalic NT. Extremely long filopodia were observed on sprouting cells, extending toward the central canal and the mantle layer. The exclusively extraneuroectodermal origin of angioblastic cells was demonstrated with chick-quail chimeras. Following replacement of quail NT by chick NT graft, angioblast and sprout distribution in chimeras was the same as in controls. We conclude that the NT receives its first blood vessels by a combination of two different processes, dorsal immigration of isolated migrating angioblastic cells and ventral sprouting of endothelial cells, which derive from perfused vessels. The dorsal invasive angioblasts contribute to the developing intraneural vascular plexus after having traversed the neural tube. The initial distribution of blood vessels within the neuroepithelium corresponds to intrinsic random motility of angioblastic cells; a more regular pattern is seen later. The floor plate apparently prohibits connections between sprouts in both NT sides, whereas in the dorsal NT, such a separating effect on the migrating angioblasts does not exist.