Patterning Of Vasculature Research Articles

Altered renal vascular patterning reduces ischemic kidney injury and limits vascular loss associated with aging.

The kidney vasculature has a complex arrangement, which runs in both series and parallel to perfuse the renal tissue and appropriately filter plasma. Recent studies have demonstrated that the development of this vascular pattern is dependent on netrin-1 secreted by renal stromal progenitors. Mice lacking netrin-1 develop an arterial tree with stochastic branching, particularly of the large interlobar vessels. The current study investigated whether abnormalities in renal vascular pattern altered kidney function or response to injury. To examine this, we analyzed kidney function at baseline as well as in response to recovery from a model of bilateral ischemic injury and measured vascular dynamics in aged mice. We found no differences in kidney function or morphology at baseline between mice with an abnormal arterial pattern compared to control. Interestingly, male and female mutant mice with stochastic vascular patterning showed a reduction in tubular injury in response to ischemia. Similarly, mutant mice also had a preservation of perfused vasculature with aging compared to a reduction in the control group. These results suggest that guided and organized patterning of the renal vasculature may not be required for normal kidney function; thus, modulating renal vascular patterning may represent an effective therapeutic strategy. Understanding how patterning and maturation of the arterial tree affects physiology and response to injury or aging has important implications for enhancing kidney regeneration and tissue engineering strategies.

Sex Differences in renal ischemia-reperfusion injury following deletion of Netrin-1 in the kidney

Netrin-1, an angiogenic guidance cue, is heavily expressed in the kidney. Our lab has previously reported that deletion of Netrin-1 in Foxd1-positive stromal progenitors ( Foxd1 GC/+ Netrin1 fl/fl ) results in abnormal vascular patterning during kidney development, including a reduction in interlobar branching. It remains unknown how these changes in vascular patterning affect kidney function and physiology. In the current study, we hypothesized that changes in developmental patterning of the kidney vasculature would protect from renal ischemia-reperfusion injury in a sex-specific manner due to ischemic preconditioning to the kidney tubular cells. To test this hypothesis, we performed warm, bilateral ischemia-reperfusion by clamping the renal pedicles for 26-minutes in wild-type ( Netrin1 fl/fl or Netrin1 fl/+ )(3M, 3F), heterozygous ( Foxd1 GC/+ Netrin1 fl/+ ) (2M, 4F), and mutant ( Foxd1 GC/+ Netrin1 fl/fl ) (3M, 1F) male (M) and female (F) mice. Following 24-hours of reperfusion, blood and kidneys were collected for analysis of kidney injury markers. We found that plasma creatinine (p 2-wayANOVA =0.021) and blood urea nitrogen (p 2-wayANOVA =0.009) were decreased in female mice compared to male mice regardless of genotype. In both male and female mutant mice, there was a decrease in these markers of injury compared to heterozygous and wild-type control mice, although this did not reach statistical significance. In agreement, immunofluorescent staining of kidney injury marker 1 (KIM-1) indicated less tubular injury in female mice compared to males. These data suggest that female mice are less susceptible to ischemic injury than male mice despite both sexes having similar alterations in vascular patterning following deletion of Netrin-1. Further, our results indicate a protective effect of these developmental patterning changes due to Netrin-1 deletion. We suspect that changes in the vasculature in our mutant mice provide a preconditioning-like environment to mediate this effect. Future studies will investigate the underlying mechanisms which drive this injury phenotype between sexes. Understanding how renal vascular development impacts kidney injury is critical to the development of new therapeutic strategies to prevent the sustained injury following periods of ischemia. NIH R01DK121014 to LO This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

The transcription factor AtMYB12 is part of a feedback loop regulating cell division orientation in the root meristem vasculature.

Transcriptional networks are crucial to integrate various internal and external signals into optimal responses during plant growth and development. In Arabidopsis thaliana, primary root vasculature patterning and proliferation are controlled by a network centred around the basic Helix-Loop-Helix transcription factor complex, formed by TARGET OF MONOPTEROS 5 (TMO5) and LONESOME HIGHWAY (LHW), which control cell proliferation and division orientation by modulating the cytokinin response and other downstream factors. Despite recent progress, many aspects of the TMO5/LHW pathway are not fully understood. In particular, the upstream regulators of TMO5/LHW activity remain unknown. Here, using a forward genetics approach to identify new factors of the TMO5/LHW pathway, we discovered a novel function of the MYB-type transcription factor, MYB12. MYB12 physically interacts with TMO5 and dampens the TMO5/LHW-mediated induction of direct target gene expression, as well as the periclinal/radial cell divisions. The expression of MYB12 is activated by the cytokinin response, downstream of TMO5/LHW, resulting in a novel MYB12-mediated negative feedback loop that restricts TMO5/LHW activity, to ensure optimal cell proliferation rates during root vascular development.

CsIVP Modulates Low Nitrogen and High-Temperature Resistance in Cucumber.

Crop plants experience various abiotic stresses that reduce yield and quality. Although several adaptative physiological and defense responses to single stress have been identified, the behavior and mechanisms of plant response to multiple stresses remain underexamined. Herein, we determined that the leaf and vascular changes in Cucumis sativus Irregular Vasculature Patterning (CsIVP)-RNAi cucumber plants can enhance resistance to nitrogen deficiency and high-temperature stress. CsIVP negatively regulated high nitrate affinity transporters (NRT2.1, NRT2.5) and reallocation transporters (NRT1.7, NRT1.9, NRT1.12) under low nitrogen stress. Furthermore, CsIVP-RNAi plants have high survival rate with low heat injury level under high-temperature condition. Several key high-temperature regulators, including Hsfs, Hsps, DREB2C, MBF1b and WRKY33 have significant expression in CsIVP-RNAi plants. CsIVP negatively mediated high-temperature responses by physically interacting with CsDREB2C. Altogether, these results indicated that CsIVP integrates innate programming of plant development, nutrient transport and high-temperature resistance, providing a potentially valuable target for breeding nutrient-efficient and heat-resistant crops.

BdERECTA controls vasculature patterning and phloem-xylem organization in Brachypodium distachyon

BackgroundThe vascular system of plants consists of two main tissue types, xylem and phloem. These tissues are organized into vascular bundles that are arranged into a complex network running through the plant that is essential for the viability of land plants. Despite their obvious importance, the genes involved in the organization of vascular tissues remain poorly understood in grasses.ResultsWe studied in detail the vascular network in stems from the model grass Brachypodium distachyon (Brachypodium) and identified a large set of genes differentially expressed in vascular bundles versus parenchyma tissues. To decipher the underlying molecular mechanisms of vascularization in grasses, we conducted a forward genetic screen for abnormal vasculature. We identified a mutation that severely affected the organization of vascular tissues. This mutant displayed defects in anastomosis of the vascular network and uncommon amphivasal vascular bundles. The causal mutation is a premature stop codon in ERECTA, a LRR receptor-like serine/threonine-protein kinase. Mutations in this gene are pleiotropic indicating that it serves multiple roles during plant development. This mutant also displayed changes in cell wall composition, gene expression and hormone homeostasis.ConclusionIn summary, ERECTA has a pleiotropic role in Brachypodium. We propose a major role of ERECTA in vasculature anastomosis and vascular tissue organization in Brachypodium.

CsIVP functions in vasculature development and downy mildew resistance in cucumber.

Domesticated crops with high yield and quality are frequently susceptible to pathogen attack, whereas enhancement of disease resistance generally compromises crop yield. The underlying mechanisms of how plant development and disease resistance are coordinately programed remain elusive. Here, we showed that the basic Helix-Loop-Helix (bHLH) transcription factor Cucumis sativus Irregular Vasculature Patterning (CsIVP) was highly expressed in cucumber vascular tissues. Knockdown of CsIVP caused severe vasculature disorganization and abnormal organ morphogenesis. CsIVP directly binds to vascular-related regulators YABBY5 (CsYAB5), BREVIPEDICELLUS (CsBP), and AUXIN/INDOLEACETIC ACIDS4 (CsAUX4) and promotes their expression. Knockdown of CsYAB5 resulted in similar phenotypes as CsIVP-RNA interference (RNAi) plants, including disturbed vascular configuration and abnormal organ morphology. Meanwhile, CsIVP-RNAi plants were more resistant to downy mildew and accumulated more salicylic acid (SA). CsIVP physically interacts with NIM1-INTERACTING1 (CsNIMIN1), a negative regulator in the SA signaling pathway. Thus, CsIVP is a novel vasculature regulator functioning in CsYAB5-mediated organ morphogenesis and SA-mediated downy mildew resistance in cucumber.

On the mechanisms of development in monocot and eudicot leaves.

Contents Summary 706 I. Introduction 707 II. Leaf zones in monocot and eudicot leaves 707 III. Monocot and eudicot leaf initiation: differences in degree and timing, but not kind 710 IV. Reticulate and parallel venation: extending the model? 711 V. Flat laminar growth: patterning and coordination of adaxial-abaxial and mediolateral axes 713 VI. Stipules and ligules: ontogeny of primordial elaborations 715 VII. Leaf architecture 716 VIII. Stomatal development: shared and diverged mechanisms for making epidermal pores 717 IX. Conclusion 719 Acknowledgements 720 References 720 SUMMARY: Comparisons of concepts in monocot and eudicot leaf development are presented, with attention to the morphologies and mechanisms separating these angiosperm lineages. Monocot and eudicot leaves are distinguished by the differential elaborations of upper and lower leaf zones, the formation of sheathing/nonsheathing leaf bases and vasculature patterning. We propose that monocot and eudicot leaves undergo expansion of mediolateral domains at different times in ontogeny, directly impacting features such as venation and leaf bases. Furthermore, lineage-specific mechanisms in compound leaf development are discussed. Although models for the homologies of enigmatic tissues, such as ligules and stipules, are proposed, tests of these hypotheses are rare. Likewise, comparisons of stomatal development are limited to Arabidopsis and a few grasses. Future studies may investigate correlations in the ontogenies of parallel venation and linear stomatal files in monocots, and the reticulate patterning of veins and dispersed stoma in eudicots. Although many fundamental mechanisms of leaf development are shared in eudicots and monocots, variations in the timing, degree and duration of these ontogenetic events may contribute to key differences in morphology. We anticipate that the incorporation of an ever-expanding number of sequenced genomes will enrich our understanding of the developmental mechanisms generating eudicot and monocot leaves.

Hypoxia Supports Epicardial Cell Differentiation in Vascular Smooth Muscle Cells through the Activation of the TGFβ Pathway.

Epicardium-derived cells (EPDCs) are an important pool of multipotent cardiovascular progenitor cells. Through epithelial-to-mesenchymal-transition (EMT), EPDCs invade the subepicardium and myocardium and further differentiate into several cell types required for coronary vessel formation. We previously showed that epicardial hypoxia inducible factor (HIF) signaling mediates the invasion of vascular precursor cells critical for patterning the coronary vasculature. Here, we examine the regulatory role of hypoxia (1% oxygen) on EPDC differentiation into vascular smooth muscle cells (VSMCs). Results: Hypoxia stimulates EMT and enhances expression of several VSMC markers in mouse epicardial cell cultures. This stimulation is specifically blocked by inhibiting transforming growth factor-beta (TGFβ) receptor I. Further analyses indicated that hypoxia increases the expression level of TGFβ-1 ligand and phosphorylation of TGFβ receptor II, suggesting an indispensable role of the TGFβ pathway in hypoxia-stimulated VSMC differentiation. We further demonstrate that the non-canonical RhoA/Rho kinase (ROCK) pathway acts as the main downstream effector of TGFβ to modulate hypoxia’s effect on VSMC differentiation. Conclusion: Our results reveal a novel role of epicardial HIF in mediating coronary vasculogenesis by promoting their differentiation into VSMCs through noncanonical TGFβ signaling. These data elucidate that patterning of the coronary vasculature is influenced by epicardial hypoxic signals.

Erratum to: Morphometric analysis of Passiflora leaves: the relationship between landmarks of the vasculature and elliptical Fourier descriptors of the blade.

BackgroundLeaf shape among Passiflora species is spectacularly diverse. Underlying this diversity in leaf shape are profound changes in the patterning of the primary vasculature and laminar outgrowth. Each of these aspects of leaf morphology—vasculature and blade—provides different insights into leaf patterning.ResultsHere, we morphometrically analyze >3300 leaves from 40 different Passiflora species collected sequentially across the vine. Each leaf is measured in two different ways: using 1) 15 homologous Procrustes-adjusted landmarks of the vasculature, sinuses, and lobes; and 2) Elliptical Fourier Descriptors (EFDs), which quantify the outline of the leaf. The ability of landmarks, EFDs, and both datasets together are compared to determine their relative ability to predict species and node position within the vine. Pairwise correlation of x and y landmark coordinates and EFD harmonic coefficients reveals close associations between traits and insights into the relationship between vasculature and blade patterning.ConclusionsLandmarks, more reflective of the vasculature, and EFDs, more reflective of the blade contour, describe both similar and distinct features of leaf morphology. Landmarks and EFDs vary in ability to predict species identity and node position in the vine and exhibit a correlational structure (both within landmark or EFD traits and between the two data types) revealing constraints between vascular and blade patterning underlying natural variation in leaf morphology among Passiflora species.

Morphometric analysis of Passiflora leaves: the relationship between landmarks of the vasculature and elliptical Fourier descriptors of the blade.

Background: Leaf shape among Passiflora species is spectacularly diverse. Underlying this diversity in leaf shape are profound changes in the patterning of the primary vasculature and laminar outgrowth. Each of these aspects of leaf morphology—vasculature and blade—provides different insights into leaf patterning. Results: Here, we morphometrically analyze >3300 leaves from 40 different Passiflora species collected sequentially across the vine. Each leaf is measured in two different ways: using 1) 15 homologous Procrustes-adjusted landmarks of the vasculature, sinuses, and lobes; and 2) Elliptical Fourier Descriptors (EFDs), which quantify the outline of the leaf. The ability of landmarks, EFDs, and both datasets together are compared to determine their relative ability to predict species and node position within the vine. Pairwise correlation of x and y landmark coordinates and EFD harmonic coefficients reveals close associations between traits and insights into the relationship between vasculature and blade patterning. Conclusions: Landmarks, more reflective of the vasculature, and EFDs, more reflective of the blade contour, describe both similar and distinct features of leaf morphology. Landmarks and EFDs vary in ability to predict species identity and node position in the vine and exhibit a correlational structure (both within landmark or EFD traits and between the two data types) revealing constraints between vascular and blade patterning underlying natural variation in leaf morphology among Passiflora species.

Vascular cambium regeneration and vessel formation in wounded inflorescence stems of Arabidopsis.

Synchronized tissue polarization during regeneration or de novo vascular tissue formation is a plant-specific example of intercellular communication and coordinated development. According to the canalization hypothesis, the plant hormone auxin serves as polarizing signal that mediates directional channel formation underlying the spatio-temporal vasculature patterning. A necessary part of canalization is a positive feedback between auxin signaling and polarity of the intercellular auxin flow. The cellular and molecular mechanisms of this process are still poorly understood, not the least, because of a lack of a suitable model system. We show that the main genetic model plant, Arabidopsis (Arabidopsis thaliana) can be used to study the canalization during vascular cambium regeneration and new vasculature formation. We monitored localized auxin responses, directional auxin-transport channels formation, and establishment of new vascular cambium polarity during regenerative processes after stem wounding. The increased auxin response above and around the wound preceded the formation of PIN1 auxin transporter-marked channels from the primarily homogenous tissue and the transient, gradual changes in PIN1 localization preceded the polarity of newly formed vascular tissue. Thus, Arabidopsis is a useful model for studies of coordinated tissue polarization and vasculature formation after wounding allowing for genetic and mechanistic dissection of the canalization hypothesis.

14-3-3epsilon controls multiple developmental processes in the mouse heart.

14-3-3ε plays an important role in the maturation of the compact ventricular myocardium by modulating the cardiomyocyte cell cycle via p27kip1 . However, additional cardiac defects are possible given the ubiquitous expression pattern of this protein. Germ line deletion of 14-3-3ε led to malalignment of both the outflow tract (OFT) and atrioventricular (AV) cushions, with resulting tricuspid stenosis and atresia, mitral valve abnormalities, and perimembranous ventricular septal defects (VSDs). We confirmed myocardial non-compaction and detected a spongy septum with muscular VSDs and blebbing of the epicardium. These defects were associated with abnormal patterning of p27kip1 expression in the subendocardial and possibly the epicardial cell populations. In addition to abnormal pharyngeal arch artery patterning, we found deep endocardial recesses and paucity of intramyocardial coronary vasculature as a result of defective coronary plexus remodeling. The malalignment of both endocardial cushions provides a new explanation for tricuspid and mitral valve defects, while myocardial non-compaction provides the basis for the abnormal coronary vasculature patterning. These abnormalities might arise from p27kip1 dysregulation and a resulting defect in epithelial-to-mesenchymal transformation. These data suggest that 14-3-3ε, in addition to left ventricular non-compaction (LVNC), might be linked to different forms of congenital heart disease (CHD). Developmental Dynamics 245:1107-1123, 2016. © 2016 Wiley Periodicals, Inc.

Coronary vasculature patterning requires a novel endothelial ErbB2 holoreceptor.

Organogenesis and regeneration require coordination of cellular proliferation, regulated in part by secreted growth factors and cognate receptors, with tissue nutrient supply provided by expansion and patterning of blood vessels. Here we reveal unexpected combinatorial integration of a growth factor co-receptor with a heterodimeric partner and ligand known to regulate angiogenesis and vascular patterning. We show that ErbB2, which can mediate epidermal growth factor (EGF) and neuregulin signalling in multiple tissues, is unexpectedly expressed by endothelial cells where it partners with neuropilin 1 (Nrp1) to form a functional receptor for the vascular guidance molecule semaphorin 3d (Sema3d). Loss of Sema3d leads to improper patterning of the coronary veins, a phenotype recapitulated by endothelial loss of ErbB2. These findings have implications for possible cardiovascular side-effects of anti-ErbB2 therapies commonly used for cancer, and provide an example of integration at the molecular level of pathways involved in tissue growth and vascular patterning.

The Arabidopsis Domain of Unknown Function 1218 (DUF1218) Containing Proteins, MODIFYING WALL LIGNIN-1 and 2 (At1g31720/MWL-1 and At4g19370/MWL-2) Function Redundantly to Alter Secondary Cell Wall Lignin Content.

DUF1218 is a land plant-specific innovation and has previously been shown to be associated with cell wall biology, vasculature patterning and abiotic/biotic stress response. The Arabidopsis genome encodes 15 members, two of which (At1g31720 and At4g27435) are preferentially expressed in the secondary cell wall depositing inflorescence stems. To further our understanding of the roles of DUF1218-containing proteins in secondary cell wall biology, we functionally characterized At1g31720 (herein referred to as MODIFYING WALL LIGNIN-1 or MWL-1). Since related gene family members may contribute to functional redundancy, we also characterized At4g19370 (MWL-2), the most closely related gene to MWL-1 in the protein family. Subcellular localization revealed that both Arabidopsis proteins are targeted to the cell periphery. The single T-DNA knockout lines, mwl-1 and mwl-2, and independent overexpression lines showed no significant differences in plant growth or changes in total lignin content relative to wild-type (WT) control plants. However, the double homozygous mutant, mwl-1/mwl-2, had smaller rosettes with a significant decrease in rosette fresh weight and stem height relative to the WT control at four weeks and six weeks, respectively. Moreover, mwl-1/mwl-2 showed a significant reduction in total lignin content (by ca. 11% relative to WT) and an increase in syringyl/guaiacyl (S/G) monomer ratio relative to the control plants. Our study has identified two additional members of the DUF1218 family in Arabidopsis as novel contributors to secondary cell wall biology, specifically lignin biosynthesis, and these proteins appear to function redundantly.

Interactions between mural cells and endothelial cells stabilize the developing zebrafish dorsal aorta.

Mural cells (vascular smooth muscle cells and pericytes) play an essential role in the development of the vasculature, promoting vascular quiescence and long-term vessel stabilization through their interactions with endothelial cells. However, the mechanistic details of how mural cells stabilize vessels are not fully understood. We have examined the emergence and functional role of mural cells investing the dorsal aorta during early development using the zebrafish. Consistent with previous literature, our data suggest that cells ensheathing the dorsal aorta emerge from a sub-population of cells in the adjacent sclerotome. Inhibition of mural cell recruitment to the dorsal aorta through disruption of pdgfr signaling leads to a reduced vascular basement membrane, which in turn results in enhanced dorsal aorta vessel elasticity and failure to restrict aortic diameter. Our results provide direct in vivo evidence for a functional role for mural cells in patterning and stabilization of the early vasculature through production and maintenance of the vascular basement membrane to prevent abnormal aortic expansion and elasticity.

The VASCULATURE COMPLEXITY AND CONNECTIVITY gene encodes a plant-specific protein required for embryo provasculature development.

The molecular mechanisms by which vascular tissues acquire their identities are largely unknown. Here, we report on the identification and characterization of VASCULATURE COMPLEXITY AND CONNECTIVITY (VCC), a member of a 15-member, plant-specific gene family in Arabidopsis (Arabidopsis thaliana) that encodes proteins of unknown function with four predicted transmembrane domains. Homozygous vcc mutants displayed cotyledon vein networks of reduced complexity and disconnected veins. Similar disconnections or gaps were observed in the provasculature of vcc embryos, indicating that defects in vein connectivity appear early in mutant embryo development. Consistently, the overexpression of VCC leads to an unusually high proportion of cotyledons with high-complexity vein networks. Neither auxin distribution nor the polar localization of the auxin efflux carrier were affected in vcc mutant embryos. Expression of VCC was detected in developing embryos and procambial, cambial, and vascular cells of cotyledons, leaves, roots, hypocotyls, and anthers. To evaluate possible genetic interactions with other genes that control vasculature patterning in embryos, we generated a double mutant for VCC and OCTOPUS (OPS). The vcc ops double mutant embryos showed a complete loss of high-complexity vascular networks in cotyledons and a drastic increase in both provascular and vascular disconnections. In addition, VCC and OPS interact physically, suggesting that VCC and OPS are part of a complex that controls cotyledon vascular complexity.

Ablation of the Renal Stroma Defines Its Critical Role in Nephron Progenitor and Vasculature Patterning

The renal stroma is an embryonic cell population located in the cortex that provides a structural framework as well as a source of endothelial progenitors for the developing kidney. The exact role of the renal stroma in normal kidney development hasn't been clearly defined. However, previous studies have shown that the genetic deletion of Foxd1, a renal stroma specific gene, leads to severe kidney malformations confirming the importance of stroma in normal kidney development. This study further investigates the role of renal stroma by ablating Foxd1-derived stroma cells themselves and observing the response of the remaining cell populations. A Foxd1cre (renal stroma specific) mouse was crossed with a diphtheria toxin mouse (DTA) to specifically induce apoptosis in stromal cells. Histological examination of kidneys at embryonic day 13.5–18.5 showed a lack of stromal tissue, mispatterning of renal structures, and dysplastic and/or fused horseshoe kidneys. Immunofluorescence staining of nephron progenitors, vasculature, ureteric epithelium, differentiated nephron progenitors, and vascular supportive cells revealed that mutants had thickened nephron progenitor caps, cortical regions devoid of nephron progenitors, aberrant vessel patterning and thickening, ureteric branching defects and migration of differentiated nephron structures into the medulla. The similarities between the renal deformities caused by Foxd1 genetic knockout and Foxd1DTA mouse models reveal the importance of Foxd1 in mediating and maintaining the functional integrity of the renal stroma.

Spatiotemporal changes in the role of cytokinin during root development

The root is a dynamic system whose structure is regulated by a complex network of interactions between hormones. The primary root meristem is specified in the embryo. After germination, the primary root meristem grows and then reaches a final size that will be maintained during the life of the plant. Subsequently, secondary structures such as lateral roots and root nodules form via the re-specification of differentiated cells. Cytokinin plays key roles in the regulation of root development. Down-regulation of the cytokinin response is required for the specification of a new stem cell niche, during both embryo and lateral root development. In the root meristem, cytokinin signalling regulates the longitudinal zonation of the meristem by controlling cell differentiation. Moreover, cytokinin regulates radial patterning of root vasculature by promoting protophloem cell identity and by spatially inhibiting protoxylem formation. In this review, an effort is made to describe the known details of the role of cytokinin during root development, taking into account also the interactions between cytokinin and other hormones. Attention is given on the dynamicity of cytokinin signalling output during different developmental events. Indeed, there is much evidence that the effects of cytokinin change as organs grow, underlining the importance of the spatiotemporal specificity of cytokinin signalling.

Development of the Pulmonary Vasculature: Current Understanding and Concepts for the Future

The pulmonary circulation is a highly specialized vascular bed that physically and functionally connects the heart and the lungs. The interdependence of these two organs is illustrated in embryonic development, when the lung endoderm protrudes into the surrounding mesoderm as the heart tube elongates and folds into structurally distinct chambers. The pulmonary vascular precursors then undergo highly stereotyped cellular maturation and patterning to form a multilayered vascular network that parallels the airways and links the arterial and venous poles of the heart. Upon the first breath, the mature pulmonary circulation is poised to receive the entire cardiac output for efficient gas exchange, and deliver oxygenated blood to the systemic circulation. Disruption of this developmental process can result in congenital defects such as the syndrome tetralogy of Fallot, or differentiation defects leading to persistent pulmonary hypertension of the newborn. Prior studies into the role of angiogenesis and vasculogenesis in pulmonary vascular development have not clearly yielded the identity of pulmonary vascular precursors, or the signals coordinating vascular maturation. We outline key questions on pulmonary vascular development that consider the role of heart-lung interaction in promoting the differentiation and patterning of the pulmonary vasculature.

Conditional hypoxia inducible factor-1α induction in embryonic pulmonary epithelium impairs maturation and augments lymphangiogenesis

Hypoxia inducible factor (HIF) 1a, EPAS1 and NEPAS are expressed in the embryonic mouse lung and each isoform exhibits distinct spatiotemporal expression patterns throughout morphogenesis. To further assess the role of the HIF1a isoform in lung epithelial cell differentiation and homeostasis, we created transgenic mice that express a constitutively active isoform of human HIF-1a (HIF-1a three point mutant (TPM)), in a doxycycline-dependent manner. Expression of HIF1a TPM in the developing pulmonary epithelium resulted in lung hypoplasia characterized by defective branching morphogenesis, altered cellular energetics and impaired epithelial maturation, culminating in neonatal lethality at birth from severe respiratory distress. Histological and biochemical analyses revealed expanded glycogen pools in the pulmonary epithelial cells at E18.5, concomitant with decreased pulmonary surfactant, suggesting a delay or an arrest in maturation. Importantly, these defects occurred in the absence of apoptosis or necrosis. In addition, sub-pleural hemorrhaging was evident as early as E14.5 in HIF1a TPM lungs, despite normal patterning of the blood vasculature, consistent with defects in endothelial barrier function. Epithelial expression of HIF1a TPM also resulted in increased VEGFA and VEGFC production, an increase in the number of lymphatic vessels and indirect activation of the multiple Notch pathway components in endothelial precursor cells. Collectively, these data indicate that HIF-1a protein levels in the pulmonary epithelium must be tightly controlled for proper development of the epithelial and mesenchymal compartments.