| Acute ischemic stroke,one of the leading causes of mortality and adult disability worldwide,develops when BVs supplying oxygen and nutrients to certain brain areas are occluded.It results in damages to regional vascular network and brain tissues.Promoting angiogenesis and neurogenesis after cerebral ischemia can effectively reduce brain injury.Relief of the stroke-caused edema and prompt recovery of basic blood flows to supply minimal oxygen and nutrients serve as two of the major principles to alleviate symptoms and minimize lethality.Previous studies have found roles of meningeal lymphatic vessels(LVs)in anti-edema and guiding angiogenesis after cerebrovascular damages.Under physiological state,the lymphatic system is responsible for the maintenance of fluid homeostasis,fat absorption,and immune surveillance.During development,the lymphatic endothelial cells(LECs)are derived from blood vessel endothelial cells(BECs)in vertebrates.The lymphatics inside the skull,including mammalian meningeal lymphatic vessels(LVs)and leptomeningeal LECs(LLECs),as well as zebrafish meningeal LVs and brain LECs(BLECs),have recently been discovered.After cerebral vascular injury in zebrafish,meningeal lymphatic endothelial cells can quickly respond to injury and invade into the injured brain parenchyma,forming ingrown lymphatic vessels,which on one hand drain interstitial fluid to resolve brain edema,on the other hand act as "growing tracks" of nascent BVs to guide vascular regeneration.However,it is a time-consuming process for neovascularization to complete regeneration through lymphatic guidance,which cannot meet the urgent requirements of basic blood flows in the injured brain.It is an urgent problem of damaged brain prompt formation of early-regenerated BVs to restore basic blood supply.Previous studies have found that after the regeneration of the cerebral vascular network to restore blood flow,the cerebral parenchyma return to the physiological state without lymphatic vessels.Some lymphatic vessels entering the cerebral parenchyma finally undergo apoptosis and are cleared,while how the remaining lymphatic vessels disappear is still unclear.In order to explore the whereabouts of non-apoptotic lymphatic vessels,we found that meningeal lymphatic vessels could transdifferentiate into blood vessels after invading the damaged brain parenchyma by means of lineage tracing experiments and multiple independent high-resolution real-time imaging experiments.Compared with the slow regeneration of blood vessels along the lymphatic vessels,the direct transformation of these lymphatic vessels into functional vessels was rapid,and lymphatic transdifferentiation enabled the earliest cerebrovascular formation and restoration of the earliest blood flow.Further statistical analysis of the regularity of lymphatic transdifferentiation showed that only independent lymphatic vessels(no vessels migrated along the lymphatic surface)could activate Notch signaling pathway to produce transdifferentiation.However,track lymphatic vessels could't undergo transdifferentiation while guiding the growth of neovascularization,and were eventually cleared after apoptosis.To investigate the molecular mechanism of transdifferentiation,we found that ephrin B2 a ligands on neovascularization specifically bound to Eph B4 a receptors in track lymphatics.The combination of ephrin B2 a ligands activated Eph B4 a in track lymphatics,phosphorylated Eph B4 a and inhibited Notch signaling downstream,thus inhibited transdifferentiation of track lymphatic vessels.Without the binding of ephrin B2 a ligand,the Eph B4 a receptor on the independent lymphatic vessels were prevented from being activated,resulting in Notch signaling pathway de-inhibition and fateful transdifferentiation into vessels.The fate of lymphatic cells could be changed by adjusting the Notch signaling pathway,when systemic overexpression NICD or conditioned overexpression NICD in lymphatic endothelial cells activated Notch signaling pathways,lymphatic transdifferentiation proportion would greatly improve,not only independent lymphatic vessels could occur transdifferentiation,track lymphatic transdifferentiation wolud also occur.However,inhibition of Notch signaling pathway by inhibitor DAPT or systemic overexpression of dominant inactivation genes resulted in lymphatic transdifferentiation failure,and the effect of such inhibition of transdifferentiation was aggravated after the conditional inhibition of Notch signaling pathway in lymphatic endothelial cells.In addition,Juvenile fish that couldn't undergo transdifferentiation developed a lethal phenotype.These results suggested that the transdifferentiation of independent lymphatic vessels into blood vessels was not only necessary for the rapid restoration of basic blood supply,but also necessary for the survival of injured animals,which was a prerequisite for cerebrovascular regeneration and repair.The phenomenon of lymphatic transdifferentiation was not only found in the cerebrovascular injury model of juvenile fish,but also observed in the photochemical thrombosis cerebrovascular disease model of adult zebrafish constructed by mimicking human cerebral thrombosis model.These findings suggested that lymphatic transdifferentiation of blood vessels was an important mechanism for emergency restoration of minimum blood supply and maintenance of life after severe cerebral infarction.Early studies have shown that in mice ischemic stroke model,the lymphatic vessels could invade the damaged area after cerebral injury,and after middle cerebral artery obstruction,the lack of lymphatic mutation experience produced more severe phenotype,in view of this,we believe that the mammals of meningeal lymphatic or pia mater lymphatic endothelial cells also play a similar function of transdifferentiation.In the future,the Ephrin B2/Eph B4/Notch signaling pathway is expected to be used as a drug target to promote the differentiation of lymphatic vessels in mammals to facilitate cerebrovascular regeneration,providing a new breakthrough and idea for the treatment of ischemic stroke. |
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