Vascular Niche Controls Organ Regeneration

Introduction

Endothelial Cell-Derived Angiopoietin-2 Controls Liver Regeneration as a Spatiotemporal Rheostat

Hu et al

Science. 2014;343:416–419.

Vessels provide the conduits that deliver metabolites and oxygen to the tissue and export waste products. After ischemia or tissue injury, endothelial cells migrate and proliferate to re-establish the capillary network in a process termed angiogenesis to maintain oxygen supply. Besides these essential and well-established functions in oxygen delivery, recent studies suggest that endothelial cells contribute to the multicellular-crosstalk that balances regeneration and dysfunctional or maladaptive healing. Thus, endothelial cells not only seem important for oxygen delivery but act as paracrine source for signals that determine tissue regeneration versus fibrosis. The interaction of endothelial-derived signals with hepatocyte functions has already been shown in the development of model organisms such as zebrafish¹ and more recently during regeneration of lung and liver in adult mice.^2,3

Two recent studies provide novel important insights into the regulation and nature of the endothelial-derived signals that control liver regeneration.^4,5 The liver is a highly regenerative organ and as such well suited to defining the signals that control proliferation of hepatocytes to restore liver function and prevent maladaptive responses that cause liver fibrosis and finally liver failure. In a recent elegant study in Science, Hu et al⁵ report a critical role of angiopoietin-2 (Ang2) in liver regeneration. Ang2 acts as contextual antagonist of the vascular receptor tyrosine kinase Tie2 and was identified by Hu et al⁵ among the most profoundly downregulated genes during liver regeneration. To explore the function of Ang2 in liver regeneration, Ang2^−/− mice were partially hepatectomized and liver regeneration was time-dependently analyzed. Within the first days, Ang2^−/− mice showed significantly increased proliferation of hepatocytes demonstrating that Ang2 downregulation releases a paracrine growth regulatory brake on hepatocytes at the early inductive stages of liver regeneration. Extensive analysis of potential mechanisms that mediate the inhibitory effects of Ang2 revealed that this is likely mediated via an indirect effect on transforming growth factor (TGF)-β1, which was reduced in Ang2^−/− mice. Consistently, neutralizing antibodies directed against TGF-β increased the liver sinusoidal endothelial cell–mediated paracrine induction of hepatocyte proliferation in vitro. Together, these data demonstrate that at early stages of liver regeneration, inhibition of Ang2-dependent TGF-β production enhances hepatocyte proliferation and regeneration (Figure [A]).

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Figure.

The vascular niche in liver regeneration. A, Summary of proposed angiopoietin-2 (Ang2) effects in liver regeneration. Ang2 downregulation during early time points results in increased hepatocyte proliferation and regeneration. Ang2 upregulation at later stages is required for angiogenesis. B, Crosstalk of stromal-derived factor-1–dependent and Ang2-dependent signaling in endothelial cells with hepatocytes. A change in CXCR4/CXCR7 ratio toward increased CXCR7 activity promotes liver regeneration. EC indicates endothelial cell; HGF, hepatocyte growth factor; LSEC, liver sinusoidal endothelial cell; TGF, transforming growth factor; and VEGFR2, vascular endothelial growth factor receptor 2.

However, when the authors analyzed hepatocyte proliferation in Ang2^−/− mice at later time points after partial hepatectomy, a different picture emerged: whereas proliferation was increased during the inductive phase (days 0–3), after day 4 hepatocyte proliferation and liver weight were significantly reduced, implicating that Ang2 is required for liver regeneration at later time points.⁵ This is consistent with the timing of Ang2 expression, which was profoundly reduced at days 1 to 2 but is re-established 4 days after injury. To identify the mechanism underlying the late impairment of liver regeneration in Ang2^−/− mice, the authors determined an influence on the stromal cell population and showed that liver sinusoidal endothelial cell proliferation and vascular endothelial growth factor receptor 2 expression were markedly reduced in Ang2^−/− mice 4 days after partial hepatectomy. Indeed, inhibition of Ang2 reduced vascular endothelial growth factor receptor 2 expression and angiogenic functions of endothelial cells in vitro, suggesting that Ang2 is required for the angiogenic phase during liver regeneration. Together, these data strongly point to a critical influence of the timing of expression of the angiocrine factor Ang2 in liver regeneration.

Recently, Ding et al⁴ highlighted the important role of another paracrine factor, the stromal-derived factor-1 (chemokine [C-X-C motif] ligand 12), and its receptors in liver regeneration. The group showed that the endothelial stromal-derived factor-1 receptors, C-X-C motif receptor 4 (CXCR4), C-X-C motif receptor 7 (CXCR7), modulate the balance between regeneration and liver fibrosis (Figure [B]). CXCR4 was the first identified receptor for the potent chemokine stromal-derived factor-1 and only recently a second receptor, namely CXCR7, gained increasing attention as it binds to stromal-derived factor-1 with even higher affinity.⁶ Both receptors are highly expressed in endothelial cells. Using endothelial-specific deletion strategies, Ding et al now made the surprising findings that only CXCR7 is required for hepatocyte proliferation and regeneration after acute carbon tetrachloride–induced liver injury. Moreover, pharmacological activation of CXCR7 prevented liver fibrosis in chronic injury models induced by iterative hepatotoxin injection and bile duct ligation. Interestingly, the CXCR4 receptor seems to counteract this proregenerative activity and its deletion in endothelial cells reduced TGF-β expression and prevented liver fibrosis in chronic models. The counteracting role of CXCR4 versus CXCR7 was mirrored by a striking time-dependent differential regulation of the receptors: CXCR7 was rapidly upregulated within 3 days after injury, whereas CXCR4 expression increased at >2 weeks after injury. Both studies identify TGF-β as profibrotic paracrine factor and it will be important to identify the specific proregenerative paracrine cocktail responsible for organ regeneration.

What do we learn from these studies? The endothelial response to injury critically regulates regeneration—at least in the liver. Thereby, the spatiotemporal control of endothelial cell intrinsic properties and paracrine activity is critical in modulating the different phases of regeneration. When considering the development of therapeutic strategies targeting growth factors to enhance regeneration and prevent maladaptive healing, the timing might be crucial. This may not be fully unexpected, considering, for example, the well-known biphasic activities of growth factors such as Wnts during differentiation⁷ or the double-edged role of inflammation in repair/regeneration.⁸ However, the recent studies now show that the endothelium independent of its function in oxygen delivery may act as either a proregenerative or a profibrotic niche, depending on the context.

Do these studies implicate a general role of endothelial cell paracrine activities in organ repair and regeneration? To date, the concept was validated for the liver, and to some extent to the lung, and several lines of evidence argue for a role of the vascular niche in the maintenance and regulation of hematopoietic stem cells in the bone marrow.⁹ Whether a similar concept may be relevant for the heart, which possesses a much lower endogenous regenerative activity, is unclear and deserves further studies. Several previous findings may suggest the existence of similar cell-to-cell communication scenarios in the heart as well: first, a disruption of the angiogenic response and angiocrine signal of endothelial cells was shown to lead to a transit from hypertrophy to heart failure in mice models.^10–12 Second, the paracrine activity of various progenitor and proangiogenic cells was shown to not only preserve cardiac myocyte survival but also contribute to myocyte replacement.^13,14 Because bone marrow–derived cell populations and cardiac stem cells have been shown to affect angiogenesis and endothelial cell function in mice and humans,^15,16 one may speculate that these activities may have contributed to the balance between regeneration/repair and dysfunctional healing leading to fibrosis. Third, studies in patients with dilative cardiomyopathy demonstrated that an impaired microcirculatory response is associated with a poor prognosis.¹⁷ Finally, conceptually, endothelial cells may be well suited to act as a paracrine reservoir because they contribute to all organs and can easily sense injuries. Of note, the well-known organ-specific features of endothelial cells may include a selective pattern of released paracrine factors that appropriately support organ healing and functions. Further insights into the multicellular-crosstalk of endothelial cells that balances regeneration versus maladaptive healing may not only provide a better understanding of the mechanisms but may help to develop better fine-tuned proangiogenic therapies.