CoChat
Sign Up
Cited by 66Lymphatic System and DiseasesLymphatic Disorders and TreatmentsDevelopmental Biology and Gene Regulation

Notch signaling functions in lymphatic valve formation

Open access
Murtomaki, Aino, Uh, Minji K., Kitajewski, Chris, Zhao, Jin, Nagasaki, Takayuki, Shawber, Carrie J., Kitajewski, Jan·2014·Development
DOI 10.1242/dev.101188PMC PMC4050693
Open this paper in CoChat — free account

Abstract

Collecting lymphatic ducts contain intraluminal valves that prevent backflow. In mice, lymphatic valve morphogenesis begins at embryonic day 15.5 (E15.5). In the mesentery, Prox1 expression is high in valve-forming lymphatic endothelial cells, whereas cells of the lymphatic ducts express lower levels of Prox1. Integrin α9, fibronectin EIIIA, Foxc2, calcineurin and the gap junction protein Cx37 are required for lymphatic valve formation. We show that Notch1 is expressed throughout the developing mesenteric lymphatic vessels at E16.5, and that, by E18.5, Notch1 expression becomes highly enriched in the lymphatic valve endothelial cells. Using a Notch reporter mouse, Notch activity was detected in lymphatic valves at E17.5 and E18.5. The role of Notch in lymphatic valve morphogenesis was studied using a conditional lymphatic endothelial cell driver either to delete Notch1 or to express a dominant-negative Mastermind-like (DNMAML) transgene. Deletion of Notch1 led to an expansion of Prox1high cells, a defect in Prox1high cell reorientation and a decrease in integrin α9 expression at sites of valve formation. Expression of DNMAML, which blocks all Notch signaling, resulted in a more severe phenotype characterized by a decrease in valves, failure of Prox1high cells to cluster, and rounding of the nuclei and decreased fibronectin-EIIIA expression in the Prox1high cells found at valve sites. In human dermal lymphatic endothelial cells, activation of Notch1 or Notch4 induced integrin α9, fibronectin EIIIA and Cx37 expression. We conclude that Notch signaling is required for proper lymphatic valve formation and regulates integrin α9 and fibronectin EIIIA expression during valve morphogenesis.

AI snapshot

Written from the paper’s full text.

TL;DR

Notch1 expression and Notch activity become enriched in developing lymphatic valve endothelial cells, and lymphatic endothelial-specific loss of Notch1 or canonical Notch signaling disrupts valve morphogenesis. Notch activation in human dermal lymphatic endothelial cells induces integrin α9, fibronectin-EIIIA and Cx37, suggesting Notch regulates multiple steps of lymphatic valve formation.

Key findings

  • Notch1 was uniformly expressed in E16.5 mesenteric collecting lymphatic ductal endothelium, became highest in Prox1high LECs at putative valve sites by E17.5, and was enriched in valve-forming LECs by E18.5; the TNR Notch reporter detected Notch activity at E17.5 and E18.5 in valve sites and α9-positive valve LECs.
  • Prox1CreERT2;N1fl/fl (Notch1 loss-of-function) embryos induced at E15.5 and analyzed at E18.5 showed expanded Prox1high regions, a significant increase in Prox1high LECs normalized to ductal length, a significant decrease in reoriented Prox1high valve LECs, significantly lower α9 expression in Prox1high LECs that had migrated into the lumen, and poorly organized valve leaflets; total valve number was similar to controls.
  • Prox1CreERT2;DNMAMLfl/+ embryos, which block all canonical Notch signaling, had a more severe phenotype: significantly fewer Prox1high LECs in clusters, significantly reduced Prox1/α9/FNEIIIA triple-positive valves per duct, many lymphatics with no identifiable valves, abnormally rounded nuclei, and absent FNEIIIA expression in Prox1high valve LECs.
  • In HdLECs, N1IC or N4int3 induced α9 transcripts, but only N1IC increased α9/β1 surface expression; both increased FN-EIIIA+ and FN-EIIIA− transcripts and total fibronectin and FN-EIIIA+ proteins; both strongly induced Cx37 transcripts and protein; N1IC or N4int3 modestly induced Foxc2 transcripts 1.5-fold, while Cnb1 was unaffected.

Methods

  • Mouse embryonic mesenteric lymphatics were analyzed at E16.5–E18.5 using Prox1CreERT2 crossed to Notch1fl/fl or DNMAMLfl/fl, with tamoxifen (10 mg/40 g) at E13.5 or E15.5; TNR Notch reporter mice and immunohistochemistry/confocal imaging were used, with quantification of Prox1high cells normalized to duct length, clustering, reorientation at least 45°, and Prox1/α9/FNEIIIA triple-positive valves per duct; significance was determined by two-tailed Student’s t-test.
  • HdLECs were infected with adenovirus expressing constitutively active Notch1 (N1IC), Notch4 (N4int3), Hey1, Hey2, lacZ or GFP, and assessed 48 h after infection by qRT-PCR, western blot and flow cytometry (10,000 cells per group).

Limitations

  • Stated in the paper: a significant loss of α9 expression in Prox1CreERT2;DNMAMLfl/+ mutants was not achieved, largely because too few identifiable valves could be analyzed, suggesting valve initiation was disrupted.
  • Stated in the paper: the antibody used to detect total fibronectin recognizes both FN-EIIIA+ and FN-EIIIA− proteins, so increased total fibronectin may be due to increased FN-EIIIA+ alone or a combination of both isoforms.
  • Evident from the text: the in vitro conclusions rely on adenoviral overexpression in human dermal lymphatic endothelial cells, and the in vivo conclusions rely on mouse embryonic mesenteric and dermal lymphatics; no rescue experiment or direct transcriptional target assay is reported.

Model-written summary. Check it against the paper before relying on it.

Integrity signals

Registry

Crossref · 10.1242/dev.101188

Cites

22 of 22 references verified0 citation errors · 0 not checkable

Statistics

Nothing to recompute.

Claims the paper makes

18 of 25 cited sources checked · 6 supported · 0 flagged · accuracy not yet measured

  • Premisecitation
    § RESULTS AND DISCUSSION

    Formation of the lymphatic valve leaflets requires α9 and its ligand FNEIIIA.

    “Formation of the lymphatic valve leaflets requires α9 and its ligand FNEIIIA (Bazigou et al., 2009).”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites Bazigou et al., 2009 source
  • Premisecitation
    § RESULTS AND DISCUSSION

    α9 and its ligand FNEIIIA are indispensable for valve leaflet formation.

    “α9 and its ligand FNEIIIA are indispensable for valve leaflet formation (Bazigou et al., 2009).”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites Bazigou et al., 2009 source

      The Results (page not provided) show that loss of integrin-α9 disrupts leaflet formation and that FN-EIIIA deficiency causes underdeveloped or abnormal leaflets in many newborn mice. But FN-EIIIA-deficient mice still form some leaflets, and the Discussion says their defects diminish in adulthood. Thus, 'indispensable' overstates the evidence for FN-EIIIA. (LLM, conf=0.87)

      Experimental — accuracy not yet measured
  • Findingown data
    § RESULTS AND DISCUSSION

    Prox1CreERT2;DNMAMLfl/+ valve morphogenesis phenotype is more severe than the Prox1CreERT2;N1fl/fl phenotype.

    “Prox1CreERT2;DNMAMLfl/+ valve morphogenesis phenotype was more severe (Fig. 3) than the Prox1CreERT2;N1fl/fl phenotype (Fig. 2).”
    • Also stated in: § Abstract · plain
      “Expression of DNMAML, which blocks all Notch signaling, resulted in a more severe phenotype”
    Quote in the paperSubstantial claim
  • Findingown data
    § Abstract

    Notch activity is detected in lymphatic valves at E17.5 and E18.5.

    “Using a Notch reporter mouse, Notch activity was detected in lymphatic valves at E17.5 and E18.5.”
    • Also stated in: § RESULTS AND DISCUSSION · plain
      “At E17.5, Notch signal activation was seen in clusters of cells at the putative valve sites (Fig. 1B).”
    • Also stated in: § RESULTS AND DISCUSSION · plain
      “At E18.5, GFP expression was mainly detected in the α9-positive LECs of the lymphatic valves (Fig. 1B).”
    Quote in the paperSubstantial claim
  • Findingown data
    § RESULTS AND DISCUSSION

    Notch1 is expressed uniformly throughout the mesenteric collecting lymphatic ductal endothelium at E16.5.

    “At E16.5, Notch1 was uniformly expressed in the mesenteric collecting lymphatic ductal endothelium (Fig. 1A).”
    • Also stated in: § Abstract · plain
      “We show that Notch1 is expressed throughout the developing mesenteric lymphatic vessels at E16.5”
    Quote in the paperSubstantial claim
  • Classified as probably not a substantial claim. 21
  • May combine two claims. 15

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation
    § INTRODUCTION

    A valve leaflet ECM core consisting of laminin α5, collagen IV and fibronectin-EIIIA (FNEIIIA) is formed by the invaginating LECs, enabling leaflet elongation and maturation.

    “A valve leaflet ECM core consisting of laminin α5, collagen IV and fibronectin-EIIIA (FNEIIIA) is formed by the invaginating LECs, enabling leaflet elongation and maturation (Bazigou et al., 2009; Norrmen et al., 2009).”
    Quote in the paperNot a substantial claim2 of 2 sources checked
    • cites Bazigou et al., 2009 source

      The Results (page not available) place laminin-α5 in the developing valve matrix core and FN-EIIIA at the leaflet edges; the Discussion links FN-EIIIA matrix assembly to leaflet elongation. The passages do not identify collagen IV as a component of the core or establish that invaginating LECs form it. (LLM, conf=0.85)

      Experimental — accuracy not yet measured
    • cites Norrmen et al., 2009 source

      The Results section (‘Molecular characterization of developing collecting lymphatic vessels’; page unavailable) reports collagen IV accumulation in the basement membrane of developing collecting vessels. It does not identify a valve-leaflet ECM core containing laminin α5 and fibronectin-EIIIA, or describe its formation by invaginating LECs and role in leaflet elongation (searched: those components and invagination). (LLM, conf=0.78)

      Experimental — accuracy not yet measured

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation
    § RESULTS AND DISCUSSION

    FNEIIIA splice variant is expressed in the developing lymphatic valve ECM core.

    “FNEIIIA splice variant is expressed in the developing lymphatic valve ECM core (Bazigou et al., 2009).”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites Bazigou et al., 2009 source

  • May combine two claims.

    Premisecitation
    § Abstract

    Integrin α9, fibronectin EIIIA, Foxc2, calcineurin and the gap junction protein Cx37 are required for lymphatic valve formation.

    “Integrin α9, fibronectin EIIIA, Foxc2, calcineurin and the gap junction protein Cx37 are required for lymphatic valve formation.”
    Quote in the paperNot a substantial claim3 of 3 sources checked
    • cites Bazigou et al., 2009 source

      The Discussion identifies integrin-α9 and fibronectin-EIIIA as important for lymphatic valve development and reports valve-formation defects in FoxC2-deficient mice. The Results also document abnormal valves in fibronectin-EIIIA-deficient mice. Our copy does not substantiate the claim about calcineurin or Cx37 (searched those terms and “gap junction”), so the full list is not supported by this source. (LLM, conf=0.88)

      Experimental — accuracy not yet measured
    • cites Kanady et al., 2011 source

      The Abstract reports defective lymphatic valve formation with Cx37 deficiency, and the Discussion reports valve defects in integrin-α9 mutant mice and failure to initiate collecting-vessel development in Foxc2-deficient mice. The paper mentions a disorganized fibronectin matrix in integrin-α9-deficient valves, but does not establish that fibronectin EIIIA or calcineurin is required for lymphatic valve formation. (LLM, conf=0.88)

      Experimental — accuracy not yet measured
    • cites Sabine et al., 2012 source

      The Introduction (page not available) says loss of Foxc2 prevents valve formation, while inactivation of integrin α9 or fibronectin EIIIA impairs leaflet elongation and produces nonfunctional valves—not an absence of valves. The Discussion and results sections (pages not available) support roles for Cx37 and calcineurin in valve development. The claim therefore overstates the role of integrin α9 and fibronectin EIIIA in formation. (LLM, conf=0.87)

      Experimental — accuracy not yet measured

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation
    § INTRODUCTION

    Connexin 37 (Cx37) is induced in clustered LECs and is required for proper valve morphogenesis.

    “Connexin 37 (Cx37; Gja4 – Mouse Genome Informatics) is induced in clustered LECs and is required for proper valve morphogenesis (Kanady et al., 2011; Sabine et al., 2012).”
    Quote in the paperNot a substantial claim2 of 2 sources checked
    • cites Kanady et al., 2011 source

      The Results section reports that Cx37 becomes enriched in lymphatic valves and that Cx37-deficient mice have fewer mesenteric valves, supporting a role in valve morphogenesis. It does not establish that Cx37 is induced specifically in clustered LECs: the Discussion says Cx37 is initially expressed throughout the developing collecting-vessel endothelium. Page numbers are unavailable in our copy. (LLM, conf=0.87)

      Experimental — accuracy not yet measured
    • cites Sabine et al., 2012 source

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation
    § INTRODUCTION

    Calcineurin signaling and Cx37 are required to specify the sites of valve formation in the duct wall.

    “Calcineurin signaling and Cx37 are required to specify the sites of valve formation in the duct wall (Sabine et al., 2012).”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites Sabine et al., 2012 source

      The sections on Cx37 and cell-autonomous calcineurin signaling show that both are needed for lymphatic-valve development: loss of Cx37 prevents early valve-forming clusters from becoming a circumferential territory, while loss of calcineurin disrupts its demarcation and condensation (page unavailable in our copy). The paper does not establish that either specifies where valve formation begins in the vessel wall; initial clusters are detected without Cx37. (LLM, conf=0.86)

      Experimental — accuracy not yet measured

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation
    § RESULTS AND DISCUSSION

    Mammalian fibronectin consists of various isoforms, created through alternative splicing.

    “Mammalian fibronectin consists of various isoforms, created through alternative splicing (Muro et al., 2003).”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites Muro et al., 2003 source

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation
    § RESULTS AND DISCUSSION

    Notch suppresses Prox1 and induces VEGFR3 in HdLECs.

    “it has been previously demonstrated that Notch suppresses Prox1 and induces VEGFR3 in HdLECs (Kang et al., 2010; Murtomaki et al., 2013).”
    Quote in the paperNot a substantial claim1 of 2 sources checked
    • cites Kang et al., 2010 source

      The section on VEGF-C-induced angiogenesis in embryos discusses VEGFR-3 in mouse endothelial cells, not Notch regulation in HdLECs (page not provided). Searches for Notch, Prox1, and human dermal lymphatic endothelial cells found no passage addressing the claimed suppression or induction. (LLM, conf=0.76)

      Experimental — accuracy not yet measured
    • cites Murtomaki et al., 2013 source

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation
    § INTRODUCTION

    Interaction of integrin α9 with its ligand FNEIIIA allows for proper valve leaflet ECM core assembly and leaflet elongation.

    “Interaction of integrin α9 with its ligand FNEIIIA allows for proper valve leaflet ECM core assembly and leaflet elongation.”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites Bazigou et al., 2009 source

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation
    § INTRODUCTION

    Genes encoding Prox1, Foxc2, integrin α9, FNEIIIA and Cx37 are all highly expressed in lymphatic valves.

    “Genes encoding the lymphatic proteins Prox1, Foxc2, integrin α9, FNEIIIA and Cx37 are all highly expressed in lymphatic valves (Bazigou et al., 2009; Kanady et al., 2011; Norrmen et al., 2009; Petrova et al., 2004; Sabine et al., 2012).”
    Quote in the paperNot a substantial claim3 of 3 sources checked
    • cites Bazigou et al., 2009 source

      The Results sections report high Prox1 and FoxC2 expression at developing valve sites, strong integrin-α9 staining in valves, and FN-EIIIA localization in the developing valve matrix. They do not establish the Cx37 part of the combined claim. The FN-EIIIA Results also say its staining weakened after birth and was absent from adult valves; page numbers are unavailable in our copy. (LLM, conf=0.86)

      Experimental — accuracy not yet measured
    • cites Kanady et al., 2011 source

      The Results (page not provided) report that Cx37 and integrin-α9 are concentrated in lymphatic valve leaflets; the Methods identify valves by locally elevated Prox1 staining. The Discussion also notes elevated Foxc2 expression in valves, but no passage found supports the claim about FNEIIIA. Thus, the paper does not support the full list attributed to it. (LLM, conf=0.88)

      Experimental — accuracy not yet measured
    • cites Norrmen et al., 2009 source

      The Discussion reports that Prox1 and Foxc2 levels remain high in developing and adult lymphatic valves, and the Results describe their expression in forming valves. Our copy does not establish high valve expression of integrin α9, FNEIIIA, or Cx37; the citing sentence also cites other papers for the combined list. (LLM, conf=0.88)

      Experimental — accuracy not yet measured

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation
    § INTRODUCTION

    The Prox1high LECs organize into a ring-like structure that constricts the collecting duct, and then valve LECs invaginate into the lumen.

    “The Prox1high LECs organize into a ring-like structure that constricts the collecting duct, and then valve LECs invaginate into the lumen (Sabine et al., 2012).”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites Sabine et al., 2012 source

  • May combine two claims.

    Findingown data
    § RESULTS AND DISCUSSION

    By E18.5, Notch1 expression is enriched in valve-forming LECs and weak in collecting duct LECs.

    “By E18.5, Notch1 expression was enriched in valve-forming LECs and weak Notch1 expression was seen in the collecting duct LECs (Fig. 1A, supplementary material Fig. S1).”
    • Also stated in: § Abstract · plain
      “by E18.5, Notch1 expression becomes highly enriched in the lymphatic valve endothelial cells”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data
    § RESULTS AND DISCUSSION

    When putative valves are observed in Prox1CreERT2;DNMAMLfl/+ lymphatics, they are disorganized and usually express only one of two valve markers, α9 or FNEIIIA.

    “When putative valves were observed in Prox1CreERT2;DNMAMLfl/+ lymphatics, the valves were disorganized and usually expressed only one of two valve markers, α9 or FNEIIIA (Fig. 3C).”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Findingown data
    § RESULTS AND DISCUSSION

    LECs in developing collecting ducts and blood vessels adjacent to the ducts also expressed GFP.

    “LECs in the developing collecting ducts and blood vessels adjacent to the ducts also expressed GFP (data not shown).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data
    § RESULTS AND DISCUSSION

    Expression of N1IC or N4int3 increased levels of FN-EIIIA+ and FN-EIIIA− transcripts, and of total fibronectin and FN-EIIIA+ proteins in HdLECs.

    “Expression of N1IC or N4int3 increased levels of FN-EIIIA+ and FN-EIIIA− transcripts, and of total fibronectin and FN-EIIIA+ proteins (Fig. 4B).”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Findingown data
    § RESULTS AND DISCUSSION

    Calcineurin subunit Cnb1 was unaffected by Notch activation in HdLECs.

    “whereas the calcineurin subunit Cnb1 was unaffected by Notch activation (supplementary material Fig. S6B).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data
    § RESULTS AND DISCUSSION

    A significant loss of α9 expression was not achieved in Prox1CreERT2;DNMAMLfl/+ mutants, largely owing to an insufficient number of identifiable valves that could be analyzed.

    “A significant loss of α9 expression was not achieved, largely owing to an insufficient number of identifiable valves that could be analyzed”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Findingown data
    § RESULTS AND DISCUSSION

    Notch1 expression is observed in adjacent blood vessels and capillaries.

    “Notch1 expression was observed in adjacent blood vessels and capillaries.”
    Quote in the paperNot a substantial claim

  • Classified as probably not a substantial claim.

    Premisecitation
    § INTRODUCTION

    Notch regulates lymphatic sprouting events in physiological and pathological settings.

    “Notch regulates lymphatic sprouting events in physiological and pathological settings (Niessen et al., 2011; Zheng et al., 2011).”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites Niessen et al., 2011 source
    • cites Zheng et al., 2011 source

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given
    § MATERIALS AND METHODS

    In vitro data represents three or more independent experiments.

    “In vitro data represents three or more independent experiments.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Premisecitation
    § RESULTS AND DISCUSSION

    DNMAML forms an inactive complex with Notch to inhibit canonical Notch/CSL signaling downstream of all Notch proteins without affecting CSL repressor function.

    “DNMAML forms an inactive complex with Notch to inhibit canonical Notch/CSL signaling downstream of all Notch proteins without affecting CSL repressor function.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Tu et al., 2005

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Interpretationown data
    § RESULTS AND DISCUSSION

    The absence of increased α9 surface in N4int3-expressing HdLECs may be due to its reduced activity relative to N1IC.

    “The absence of increased α9 surface in N4int3-expressing HdLECs may be due to its reduced activity relative to N1IC (supplementary material Fig. S5).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Study design and conductown data
    § RESULTS AND DISCUSSION

    GFP expression was detected in Prox1CreERT2;DNMAMLfl/+ mesenteric lymphatics but not in control mesenteries.

    “GFP expression was detected in Prox1CreERT2;DNMAMLfl/+ mesenteric lymphatics (supplementary material Fig. S4), but not in control (DNMAMLfl/+) mesenteries (data not shown).”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given
    § MATERIALS AND METHODS

    Number of embryos and images analyzed summarized in supplementary material Table S3.

    “Number of embryos and images analyzed summarized in supplementary material Table S3.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation
    § INTRODUCTION

    Notch signaling requires direct cell-cell contact between Notch ligand-presenting cells and receptor-expressing cells, and regulates cell-fate determination.

    “Notch signaling requires direct cell-cell contact between Notch ligand-presenting cells and receptor-expressing cells, and regulates cell-fate determination (Andersson et al., 2011).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Andersson et al., 2011 source

  • May combine two claims.

    Interpretationown data
    § RESULTS AND DISCUSSION

    Notch regulates the establishment of a valve LEC identity and subsequent clustering and/or valve leaflet formation.

    “the disruption of Prox1 expression pattern in Prox1CreERT2;DNMAMLfl/+mesenteries and failure of Prox1high cells to cluster at valve sites suggest that Notch regulates the establishment of a valve LEC identity and subsequent clustering and/or valve leaflet formation.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation
    § INTRODUCTION

    Ligand-induced proteolytic cleavage releases the Notch intracellular domain that translocates to the nucleus where it forms a transcriptional activation complex with CSL, Mastermind-like and co-activators.

    “Ligand-induced proteolytic cleavage releases the Notch intracellular domain that translocates to the nucleus where it forms a transcriptional activation complex with CSL, Mastermind-like and co-activators (Borggrefe and Liefeke, 2012).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Borggrefe and Liefeke, 2012 source

  • May combine two claims.

    Interpretationreasoning
    § RESULTS AND DISCUSSION

    α9 is a direct target of Notch transcriptional activation or regulated indirectly by other Notch-induced proteins.

    “suggesting that α9 is a direct target of Notch transcriptional activation or regulated indirectly by other Notch-induced proteins.”
    Quote in the paperNot a substantial claim

  • Classified as probably not a substantial claim.

    Study design and conductnone given
    § MATERIALS AND METHODS

    Prox1high cell threshold was determined for control ducts and applied to all images analyzed.

    “Prox1high cell threshold was determined for control ducts and applied to all images analyzed.”
    Quote in the paperSubstantial claim

  • Classified as probably not a substantial claim.

    Study design and conductnone given
    § MATERIALS AND METHODS

    Significance was determined using two-tailed Student's t-test.

    “Significance determined using two-tailed Student's t-test.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Interpretationreasoning
    § RESULTS AND DISCUSSION

    The increase in total fibronectin levels may be due to increased FN-EIIIA+ levels or to a combination of increased FN-EIIIA+ and FN-EIIIA− levels.

    “thus the increase in total fibronectin levels may be due to increased FN-EIIIA+ levels or to a combination of increased FN-EIIIA+ and FN-EIIIA− levels.”
    Quote in the paperNot a substantial claim
30 of 30

  • The paper relies on this without support.

    In mice, lymphatic valve morphogenesis begins at embryonic day 15.5 (E15.5).

    “In mice, lymphatic valve morphogenesis begins at embryonic day 15.5 (E15.5).”
    “In mice, lymphatic valve formation begins at embryonic day 15.5 (E15.5).”

  • The paper relies on this without support.

    Valve LECs must be specified from ductal LECs.

    “Valve LECs must be specified from ductal LECs.”

  • The paper relies on this without support.

    Notch mediates bi-potential cell fate decisions in multiple biological settings.

    “Notch mediates bi-potential cell fate decisions in multiple biological settings.”
3 of 3

Cites

Flagged first, then by citations

Cited by

Sorted by citations

Related

Authors
Murtomaki, A., Uh, M. K., Kitajewski, C., Zhao, J., Nagasaki, T., Shawber, C. J., & Kitajewski, J.
Affiliations
  • Department of OB/GYN, Columbia University Medical Center, New York, NY 10032, USA
  • Columbia University Irving Medical Center (US)
Corresponding author
Kitajewski, Jan
ORCID
Venue
Development
Publisher
The Company of Biologists
Published
2014-06-15
Volume
141
Issue
12
Pages
2446-2451
ISSN
  • 0950-1991 (ISSN-L)
  • 1477-9129
DOI
10.1242/dev.101188
PMC
PMC4050693
OpenAlex
W2137343175
Semantic Scholar
e138617b265e5b775baf8f1974834a6a74128266
Cited by
58
References
18
Citations by year
2026: 5 · 2025: 2 · 2024: 3 · 2023: 3 · 2022: 3 · 2021: 11 · 2020: 4 · 2019: 7 · 2018: 3 · 2017: 8 · 2016: 5 · 2015: 2 · 2014: 1
FWCI
2.32
Type
journal article
Language
en
Open access
Locations
  • Development (publishedVersion)
  • PubMed (publishedVersion)
  • PubMed Central (submittedVersion)
  • HighWire Press Open Archive (submittedVersion)
Topics
  • Health Sciences › Medicine › Oncology › Lymphatic System and Diseases
  • Health Sciences › Medicine › Surgery › Lymphatic Disorders and Treatments
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Developmental Biology and Gene Regulation
Keywords
Notch signaling, Notch1, Notch4, Prox1, integrin alpha9, Cx37, lymphatic endothelial cells, mesenteric lymphatics
MeSH terms
Receptor, Notch4, Gap Junction alpha-4 Protein, Animals, Cells, Cultured, Fibronectins, Genes, Dominant, Humans

About these checks

Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.

Claims read by deepseek-v4.1-flash under definition version 3 (extract-v3) on 2026-10-07.

Classified by jev-1.13-20260917 (gold-guideline-v0.2) on 2026-10-07.

Cited sources are checked by gpt-6-sol.

Statistics and arithmetic are recomputed, not judged.

Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.

What counts as a claim ›How we measure confidence ›How we check this ›

Are you an author, or do you see a problem with this page? Request a review