Contact-dependent inhibition of angiogenesis by cardiac fibroblasts in three-dimensional fibrin gels in vitro: implications for microvascular network remodeling and coronary collateral formation

Angiogenesis and coronary artery collateral formation can improve blood flow and thereby prevent myocardial ischemia. The role of perivascular fibroblasts in neovascularization remains incompletely understood. Here we investigated the effects of epicardial and myocardial fibroblasts on angiogenesis in vitro by using a serum-free microcarrier-based fibrin gel angiogenesis system. To clearly distinguish between different cell types, we either stained endothelial cells or fibroblasts in the living with 1,1′-dioctadecyl-3,3,3′,3′-tetramethyl-indocarbocyanine-perchlorate (DiI). In cocultures, low numbers of heart fibroblasts stimulated endothelial sprouting, and capillary growth was also induced by fibroblast-conditioned media, indicating a paracrine mechanism. Capillary formation was decreased by increasing the density of fibroblasts in the cocultures, indicating contact-dependent inhibition. Using time-lapse studies, it turned out that close contacts between fibroblasts and endothelial cells resulted in rapid retraction of endothelial cells or, rarely, in cell death. Depending on the local ratio of fibroblasts to endothelial cell numbers, fibroblasts determined the location of capillary growth and the size of developing capillaries and thereby contributed to capillary network remodeling. In contrast to primary heart fibroblasts, NIH 3T3 fibroblasts did not display contact-dependent inhibition of endothelial sprouts. NIH fibroblasts were frequently seen in close association with endothelial capillaries, resembling pericytes. Contact-dependent inhibition of angiogenesis by epicardial fibroblasts could not be reversed by addition of neutralizing anti-TGF-β1 antibodies, by addition of serum, of medium conditioned by hypoxic tumor cells or myocardium, by various cytokines or by growing cocultures under hypoxic conditions. Our results implicate a pivotal role of periendothelial mesenchymal cells for the regulation of microvascular network remodeling and collateral formation. Received: 15 September 1997 / Accepted: 6 April 1998     

Models for pattern formation in somitogenesis: a marriage of cellular and molecular biology

Somitogenesis, the process by which a bilaterally symmetric pattern of cell aggregations is laid down in a cranio-caudal sequence in early vertebrate development, provides an excellent model study for the coupling of interactions at the molecular and cellular level. Here, we review some of the key experimental results and theoretical models related to this process. We extend a recent chemical pre-pattern model based on the cell cycle Journal of Theoretical Biology 207 (2000) 305-316, by including cell movement and show that the resultant model exhibits the correct spatio-temporal dynamics of cell aggregation. We also postulate a model to account for the recently observed spatio-temporal dynamics at the molecular level.     

Cell elongation is key to in silico replication of in vitro vasculogenesis and subsequent remodeling

Vasculogenesis, the de novo growth of the primary vascular network from initially dispersed endothelial cells, is the first step in the development of the circulatory system in vertebrates. In the first stages of vasculogenesis, endothelial cells elongate and form a network-like structure, called the primary capillary plexus, which subsequently remodels, with the size of the vacancies between ribbons of endothelial cells coarsening over time. To isolate such intrinsic morphogenetic ability of endothelial cells from its regulation by long-range guidance cues and additional cell types, we use an in vitro model of human umbilical vein endothelial cells (HUVEC) in Matrigel. This quasi-two-dimensional endothelial cell culture model would most closely correspond to vasculogenesis in flat areas of the embryo like the yolk sac. Several studies have used continuum mathematical models to explore in vitro vasculogenesis: such models describe cell ensembles but ignore the endothelial cells' shapes and active surface fluctuations. While these models initially reproduce vascular-like morphologies, they eventually stabilize into a disconnected pattern of vascular "islands." Also, they fail to reproduce temporally correct network coarsening. Using a cell-centered computational model, we show that the endothelial cells' elongated shape is key to correct spatiotemporal in silico replication of stable vascular network growth. We validate our simulation results against HUVEC cultures using time-resolved image analysis and find that our simulations quantitatively reproduce in vitro vasculogenesis and subsequent in vitro remodeling.     

New method of three-dimensional analysis of bipedal locomotion for the study of displacements of the body and body-parts centers of mass in man and non-human primates: Evolutionary framework

The current biomechanical interpretation of the chimpanzee's bipedal walking argues that larger lateral and vertical displacements of the body center of mass occur in the chimpanzee's “side-to-side” gait than in the human striding gait. The evolutionary hypothesis underlying this study is the following: during the evolution of human bipedalism one of the necessary changes could have been the progressive reduction of these displacements of the body center of mass. In order to quantitatively test this hypothesis, it is necessary to obtain simultaneously the trajectories of the centers of mass of the whole body and of the different body parts. To solve this problem, a new method of three-dimensional analysis of walking, associated with a volumetric modelling of the body, has been developed based on finite-element modelling. An orthogonal synchrophotographic device yielding four synchronous pictures of the walking subject allows a qualitative analysis of the photographic sequences together with the results of their quantitative analysis. This method was applied to an adult man, a 3-year-old girl and a 9-year-old male chimpanzee. Our results suggest that the trajectory of the body center of mass of the human is distinguished from that of the chimpanzee not by a lower movement amplitude but by the synchronization of the transverse and vertical displacements into two periodic curves in phase with one another. The non-human primate uses its repertoire of arboreal movements in its bipedal terrestrial gait, provisionally referred to as a “rope-walker” gait. We show that the interpretation of a “side-to-side” gait is not applicable to the chimpanzee. We argue that similarly this interpretation and the initial hypothesis presuppose a basic symmetric structure of the gait, in relation to the sagittal plane of progression, similar to the human one. This lateral symmetry of the right and left displacements of the center of gravity, in phase with the right and left single supports of walking, is probably a very derived feature of the human gait. We suggest that low lateral and vertical displacements of the body center of mass are not indicative of a progressive bipedal gait and we discuss the new evolutionary implications of our results. © 1993 Wiley-Liss, Inc.