RESUMO
Corneal avascularity is important for optical clarity and normal vision. However, the molecular mechanisms that prevent angioblast migration and vascularization of the developing cornea are not clear. Previously we showed that periocular angioblasts and forming ocular blood vessels avoid the presumptive cornea despite dynamic ingression of neural crest cells. In the current study, we investigate the role of Semaphorin3A (Sema3A), a cell guidance chemorepellent, on angioblast migration and corneal avascularity during development. We show that Sema3A, Vegf, and Nrp1 are expressed in the anterior eye during cornea development. Sema3A mRNA transcripts are expressed at significantly higher levels than Vegf in the lens that is positioned adjacent to the presumptive cornea. Blockade of Sema3A signaling via lens removal or injection of a synthetic Sema3A inhibitor causes ectopic migration of angioblasts into the cornea and results in its subsequent vascularization. In addition, using bead implantation, we demonstrate that exogenous Sema3A protein inhibits Vegf-induced vascularization of the cornea. In agreement with these findings, loss of Sema/Nrp1 signaling in Nrp1(Sema-) mutant mice results in ectopic angioblasts and vascularization of the embryonic mouse corneas. Altogether, our results reveal Sema3A signaling as an important cue during the establishment of corneal avascularity in both chick and mouse embryos. Our study introduces cornea development as a new model for studying the mechanisms involved in vascular patterning during embryogenesis and it also provides new insights into therapeutic potential for Sema3A in neovascular diseases.
Assuntos
Córnea/irrigação sanguínea , Cristalino/irrigação sanguínea , Neuropilina-1/genética , Semaforina-3A/fisiologia , Fator A de Crescimento do Endotélio Vascular/metabolismo , Animais , Animais Geneticamente Modificados , Movimento Celular , Células Cultivadas , Embrião de Galinha , Córnea/embriologia , Células Endoteliais , Cristalino/embriologia , Camundongos , Neovascularização Fisiológica , Neuropilina-1/biossíntese , Codorniz/embriologia , RNA Mensageiro/biossíntese , Proteínas Recombinantes de Fusão/genética , Semaforina-3A/antagonistas & inibidores , Semaforina-3A/genética , Transdução de Sinais , Fator A de Crescimento do Endotélio Vascular/biossínteseRESUMO
JorRay, Blocker23, Nibbles, and OlgasClover are actinobacteriophages belonging to clusters G1, B2, CT, and DJ, respectively. JorRay and Blocker23 were identified in host bacterium Mycobacterium smegmatis mc2155. Nibbles and OlgasClover were identified in host bacterium Gordonia rubripertincta NRRL B-16540.
RESUMO
AnarQue and Figliar are bacteriophages identified from the host bacterium Gordonia rubripertincta NRRL B-16540. AnarQue is circularly permuted and has a length of 61,822 bp; it is assigned to cluster DR. Figliar has a 3' sticky overhang and a length of 61,147 bp; it is assigned to cluster DJ.
RESUMO
Jodelie19, BlingBling, and Burnsey are bacteriophages identified using host bacteria of the genus Gordonia Jodelie19 is a lytic phage found in Gordonia rubripertincta NRRL B-16540. The temperate phage BlingBling and lytic phage Burnsey were both isolated using the host bacterium Gordonia terrae 3612.
RESUMO
Jellybones and NHagos are bacteriophages that were identified in the host bacterium Gordonia rubripertincta NRRL B-16540. Jellybones has a direct terminal repeat and was assigned to the CS2 subcluster with a length of 77,514 bp. NHagos is circularly permuted and was assigned to the DR cluster with a length of 59,580 bp.
RESUMO
Trigeminal sensory innervation of the cornea is critical for protection and synthesis of neuropeptides required for normal vision. Little is known about axon guidance during mammalian corneal innervation. In contrast to the chick where a pericorneal nerve ring forms via Npn/Sema signaling, mouse corneal axons project directly into the presumptive cornea without initial formation of an analogous nerve ring. Here we show that during development of the mouse cornea, Npn1 is strongly expressed by the trigeminal ganglion whereas Npn2 is expressed at low levels. At the same time Sema3A and Sema3F are expressed in distinct patterns in the ocular tissues. Npn1(sema-/-) mutant corneas become precociously and aberrantly innervated by nerve bundles that project further into the corneal stroma. In contrast, stromal innervation was not affected in Npn2(-/-) mutants. The corneal epithelium was prematurely innervated in both Npn1(sema-/-) and Npn2(-/-) mutants. These defects were exacerbated in Npn1(sema-/-);Npn2(-/-) double mutants, which in addition showed ectopic innervation of the region between the optic cup and lens vesicle. Collectively, our data show that Sema3A/Npn1 and Sema3F/Npn2 signaling play distinct roles and both are required for proper innervation of the mouse cornea.
Assuntos
Substância Própria/fisiologia , Neuropilina-1/fisiologia , Neuropilina-2/fisiologia , Animais , Axônios/metabolismo , Substância Própria/metabolismo , Epitélio Corneano/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Mutação , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Neuropeptídeos/genética , Neuropeptídeos/metabolismo , Neuropeptídeos/fisiologia , Neuropilina-1/genética , Neuropilina-1/metabolismo , Neuropilina-2/genética , Neuropilina-2/metabolismo , Semaforina-3A/genética , Semaforina-3A/metabolismo , Transdução de Sinais , Gânglio Trigeminal/metabolismo , Gânglio Trigeminal/fisiologiaRESUMO
PURPOSE: Dense innervation of the cornea is important for maintaining its homeostasis and transparency. Although corneal nerves have been well studied in adults, little is known about mammalian corneal innervation during development. This study provides a detailed profile of nerves at various stages of mouse cornea development. METHODS: Mouse heads and corneas were collected at various stages of development including embryonic days (E)12.5 to E16.5, postnatal days (P)0, P10, three weeks after birth, and the adult. Corneas were immunostained with an anti-neuron-specific ß-tubulin antibody (TUJ1). Fluorescently labeled nerves in whole-mount tissues and sections were imaged and analyzed for their axonal projections during eye development. RESULTS: The first nerve bundles appear at the periphery of the anterior portion of the eye by E12.5. Initial projection into the stroma occurs at E13.5 without formation of a pericorneal nerve ring. Between E13.5 and E16.5, nerve bundles project directly into the periphery of the presumptive cornea stroma. They branch repeatedly as they extend toward the cornea center and epithelium. Concomitantly, nerve bundles originating from four quadrants of the eye bifurcate into smaller branches that innervate the entire stroma. The first epithelial innervation occurs at E16.5. Epithelial nerves arrange into patterns that project toward the center subsequently forming a swirl at three weeks after birth, which becomes more pronounced in adults. CONCLUSIONS: Nerve bundles that arise from four quadrants of the eye innervate the mouse cornea. The nerve bundles directly innervate the stroma without forming a pericorneal nerve ring. Radial arrangement of epithelial nerves gradually becomes centrally oriented, subsequently forming a swirl pattern.