The endothelium is the single layer of cells lining all blood vessels, and it is a remarkable cardiovascular control centre. Each endothelial cell has only a small number (on average six) of interconnected neighbours. Yet this arrangement produces a large repertoire of behaviours, capable of controlling numerous cardiovascular functions in a flexible and dynamic way. The endothelium regulates the delivery of nutrients and removal of waste by regulating blood flow and vascular permeability. The endothelium regulates blood clotting, responses to infection and inflammation, the formation of new blood vessels, and remodelling of the blood vessel wall. To carry out these roles, the endothelium autonomously interprets a complex environment crammed with signals from hormones, neurotransmitters, pericytes, smooth muscle cells, various blood cells, viral or bacterial infection and proinflammatory cytokines. It is generally assumed that the endothelium responds to these instructions with coordinated responses in a homogeneous population of endothelial cells. Here, we highlight evidence that shows that neighbouring endothelial cells are highly heterogeneous and display different sensitivities to various activators. Cells with various sensitivities process different extracellular signals into distinct streams of information in parallel, like a vast switchboard. Communication occurs among cells and new ‘emergent’ signals are generated that are non-linear composites of the inputs. Emergent signals cannot be predicted or deduced from the properties of individual cells. Heterogeneity and emergent behaviour bestow capabilities on the endothelial collective that far exceed those of individual cells. The implications of heterogeneity and emergent behaviour for understanding vascular disease and drug discovery are discussed.

Endothelial cells line all blood vessels and are critical regulators of vascular tone. In hypertension, disruption of endothelial function alters the release of endothelial-derived vasoactive factors and results in increased vascular tone. Although the release of endothelial-derived vasodilators occurs in a Ca 2+ -dependent manner, little is known on how Ca 2+ signaling is altered in hypertension. A key element to endothelial control of vascular tone is Ca 2+ signals at specialized regions (myoendothelial projections) that connect endothelial cells and smooth muscle cells. This work describes disruption in the operation of this key Ca 2+ signaling pathway in hypertension. We show that vascular reactivity to phenylephrine is increased in hypertensive (spontaneously hypertensive rat) when compared with normotensive (Wistar Kyoto) rats. Basal endothelial Ca 2+ activity limits vascular contraction, but that Ca 2+ -dependent control is impaired in hypertension. When changes in endothelial Ca 2+ levels are buffered, vascular contraction to phenylephrine increased, resulting in similar responses in normotension and hypertension. Local endothelial IP 3 (inositol trisphosphate)-mediated Ca 2+ signals are smaller in amplitude, shorter in duration, occur less frequently, and arise from fewer sites in hypertension. Spatial control of endothelial Ca 2+ signaling is also disrupted in hypertension: local Ca 2+ signals occur further from myoendothelial projections in hypertension. The results demonstrate that the organization of local Ca 2+ signaling circuits occurring at myoendothelial projections is disrupted in hypertension, giving rise to increased contractile responses.

Pericytes, perivascular cells embedded in the microvascular wall, are crucial for vascular homeostasis. These cells also play diverse roles in tissue development and regeneration as multi-lineage progenitors, immunomodulatory cells and as sources of trophic factors. Here, we establish that pericytes are renin producing cells in the human kidney. Renin was localized by immunohistochemistry in CD146 and NG2 expressing pericytes, surrounding juxtaglomerular and afferent arterioles. Similar to pericytes from other organs, CD146+CD34–CD45–CD56– renal fetal pericytes, sorted by flow cytometry, exhibited tri-lineage mesodermal differentiation potential in vitro. Additionally, renin expression was triggered in cultured kidney pericytes by cyclic AMP as confirmed by immuno-electron microscopy, and secretion of enzymatically functional renin, capable of generating angiotensin I. Pericytes derived from second trimester human placenta also expressed renin in an inducible fashion although the renin activity was much lower than in renal pericytes. Thus, our results confirm and extend the recently discovered developmental plasticity of microvascular pericytes, and may open new perspectives to the therapeutic regulation of the renin-angiotensin system.

Three-dimensional fluorescence time-lapse imaging of the beating heart is extremely challenging, due to the heart’s constant motion and a need to avoid pharmacological or phototoxic damage. Although real-time triggered imaging can computationally “freeze” the heart for 3D imaging, no previous algorithm has been able to maintain phase-lock across developmental timescales. We report a new algorithm capable of maintaining day-long phase-lock, permitting routine acquisition of synchronised 3D + time video time-lapse datasets of the beating zebrafish heart. This approach has enabled us for the first time to directly observe detailed developmental and cellular processes in the beating heart, revealing the dynamics of the immune response to injury and witnessing intriguing proliferative events that challenge the established literature on cardiac trabeculation. Our approach opens up exciting new opportunities for direct time-lapse imaging studies over a 24-hour time course, to understand the cellular mechanisms underlying cardiac development, repair and regeneration.

Triptolide is a vine extract used in traditional Chinese medicines and associated with hepatotoxicity. In vitro data suggest that inhibition of RNA synthesis may be the mechanism of toxicity. For studying drug-induced liver injury the zebrafish has experimental, practical and financial advantages compared with rodents. The aim of this study was to explore the mechanism of triptolide toxicity using zebrafish as the model system. The effect of triptolide exposure on zebrafish larvae was determined with regard to mortality, histology, expression of liver specific microRNA-122 and liver volume. Fluorescent microscopy was used to track toxicity in the Tg(-2.8lfabp:GFP)as3 zebrafish line. Informed by microscopy, RNA-sequencing was used to explore the mechanism of toxicity. Triptolide exposure resulted in dose-dependent mortality, a reduction in the number of copies of microRNA-122 per larva, hepatocyte vacuolation, disarray and oncotic necrosis, and a reduction in liver volume. These findings were consistent across replicate experiments. Time-lapse imaging indicated the onset of injury was 6 h after the start of exposure, at which point, RNA-sequencing revealed that 88% of genes were down-regulated. Immune response associated genes were up-regulated in the triptolide-treated larvae including nitric oxide synthase. Inhibition of nitric oxide synthase increased mortality. Triptolide induces hepatotoxicity in zebrafish larvae. This represents a new model of drug-induced liver injury that complements rodents. RNA sequencing, guided by time-lapse microscopy, revealed early down-regulation of genes consistent with previous invitro studies, and facilitated the discovery of mechanistic inflammatory pathways.