SUMMARY: Damage to regional cerebrovascular network and neuronal tissues occurs during acute cerebrovascular diseases, such as ischemic stroke. The promotion of vascular regeneration is the most promising therapeutic approach. To understand cellular and molecular mechanisms underlying brain vascular regeneration, we developed two zebrafish cerebrovascular injury models using genetic ablation and photochemical thrombosis. Although brain parenchyma is physiologically devoid of lymphatic vasculature, we found that cerebrovascular injuries induce rapid ingrowth of meningeal lymphatics into the injured parenchyma. The ingrown lymphatics on one hand become lumenized drain interstitial fluid to resolve brain edema, on the other hand act as “growing tracks” for nascent blood vessels. The ingrown lymphatic vessels undergo apoptosis and clearance after cerebrovascular regeneration. This study reveals a pathological function of meningeal lymphatics, through previously unexpected ingrowth into brain parenchyma and a newly identified lymphatic function as vascular “growing tracks”. HIGHLIGHTS: Cerebrovascular injuries induce lymphatic ingrowth into the injured brain parenchyma The ingrown lymphatics drain interstitial fluid to resolve brain edema Nascent blood vessels use the ingrown lymphatic vessels as “growing tracks” The ingrown lymphatic vessels undergo apoptosis after vascular regeneration completes

The mechanisms that ensure fertilization of eggs by a single sperm are not fully understood. In all teleosts, a channel called the ‘micropyle’ is the only route of entry for sperm to enter and fertilize the egg. The micropyle forms by penetration of the developing vitelline envelope by a single specialized follicle cell, the micropylar cell, which subsequently degenerates. The mechanisms underlying micropylar cell specification and micropyle formation are poorly understood. Here, we show that an effector of the Hippo signaling pathway, the Transcriptional co-activator with a PDZ-binding domain (Taz), plays crucial roles in micropyle formation and fertilization in zebrafish. Genome editing mutants affecting taz can grow to adults, however, eggs from homozygous taz females are not fertilized even though oocytes in mutant females are histologically normal with intact animal-vegetal polarity, complete meiosis and proper ovulation. However, taz mutant eggs have no micropyle. We show that Taz protein is specifically enriched from mid-oogenesis onwards in two follicle cells located at the animal pole of the oocyte, and co-localizes with the actin and tubulin cytoskeleton. Taz protein and micropylar cell are not detected in taz mutant ovaries. Our work identifies a novel role for the Hippo/Taz pathway in micropylar cell specification in zebrafish, and uncovers the molecular basis of micropyle formation in teleosts.

The hepatopancreatic duct (HPD) system links the liver and pancreas to the intestinal tube and is composed of the extrahepatic biliary duct, gallbladder and pancreatic duct. Haematopoietically-expressed-homeobox (Hhex) protein plays an essential role in the establishment of HPD, however, the molecular mechanism remains elusive. Here we show that zebrafish hhex -null mutants fail to develop the HPD system characterized by lacking the biliary marker Annexin A4 and the HPD marker sox9b . The mutant HPD system is replaced by an intrahepatic intestinal tube characterized by expressing the intestinal marker fatty-acid-binding–protein 2a ( fabp2a ). Cell lineage analysis showed that this intrahepatic intestinal tube is not originated from hepatocytes or cholangiocytes. Further analysis revealed that cdx1b and pdx1 were expressed ectopically in the intrahepatic intestinal tube and knockdown of cdx1b and pdx1 restored the expression of sox9b in the mutant. Chromatin-immunoprecipitation analysis shows that Hhex binds to the promoters of pdx1 and cdx1b genes to repress their expression. We therefore propose that Hhex, Cdx1b and Pdx1 form a genetic network governing the patterning and morphogenesis of the HPD and digestive tract systems in zebrafish.