Extensive jejunal injury is repaired by migration and transdifferentiation of ileal enterocytes in zebrafish.

A major cause of intestinal failure (IF) is intestinal epithelium necrosis and massive loss of enterocytes, especially in the jejunum, the major intestinal segment in charge of nutrient absorption. However, mechanisms underlying jejunal epithelial regeneration after extensive loss of enterocytes remain elusive. Here, we apply a genetic ablation system to induce extensive damage to jejunal enterocytes in zebrafish, mimicking the jejunal epithelium necrosis that causes IF. In response to injury, proliferation and filopodia/lamellipodia drive anterior migration of the ileal enterocytes into the injured jejunum. The migrated fabp6+ ileal enterocytes transdifferentiate into fabp2+ jejunal enterocytes to fulfill the regeneration, consisting of dedifferentiation to precursor status followed by redifferentiation. The dedifferentiation is activated by the IL1ß-NFκB axis, whose agonist promotes regeneration. Extensive jejunal epithelial damage is repaired by the migration and transdifferentiation of ileal enterocytes, revealing an intersegmental migration mechanism of intestinal regeneration and providing potential therapeutic targets for IF caused by jejunal epithelium necrosis.

Intestinal precursors avoid being misinduced to liver cells by activating Cdx-Wnt inhibition cascade.

During coordinated development of two neighboring organs from the same germ layer, how precursors of one organ resist the inductive signals of the other to avoid being misinduced to wrong cell fate remains a general question in developmental biology. The liver and anterior intestinal precursors located in close proximity along the gut axis represent a typical example. Here we identify a zebrafish leberwurst (lbw) mutant with a unique hepatized intestine phenotype, exhibiting replacement of anterior intestinal cells by liver cells. lbw encodes the Cdx1b homeoprotein, which is specifically expressed in the intestine, and its precursor cells. Mechanistically, in the intestinal precursors, Cdx1b binds to genomic DNA at the regulatory region of secreted frizzled related protein 5 (sfrp5) to activate sfrp5 transcription. Sfrp5 blocks the mesoderm-derived, liver-inductive Wnt2bb signal, thus conferring intestinal precursor cells resistance to Wnt2bb. These results demonstrate that the intestinal precursors avoid being misinduced toward hepatic lineages through the activation of the Cdx1b-Sfrp5 cascade, implicating Cdx/Sfrp5 as a potential pharmacological target for the manipulation of intestinal-hepatic bifurcations, and shedding light on the general question of how precursor cells resist incorrect inductive signals during embryonic development.

Tel2 regulates redifferentiation of bipotential progenitor cells via Hhex during zebrafish liver regeneration.

Upon extensive hepatocyte loss or impaired hepatocyte proliferation, liver regeneration occurs via biliary epithelial cell (BEC) transdifferentiation, which includes dedifferentiation of BECs into bipotential progenitor cells (BP-PCs) and then redifferentiation of BP-PCs to nascent hepatocytes and BECs. This BEC-driven liver regeneration involves reactivation of hepatoblast markers, but the underpinning mechanisms and their effects on liver regeneration remain largely unknown. Using a zebrafish extensive hepatocyte ablation model, we perform an N-ethyl-N-nitrosourea (ENU) forward genetic screen and identify a liver regeneration mutant, liver logan (lvl), in which the telomere maintenance 2 (tel2) gene is mutated. During liver regeneration, the tel2 mutation specifically inhibits transcriptional activation of a hepatoblast marker, hematopoietically expressed homeobox (hhex), in BEC-derived cells, which blocks BP-PC redifferentiation. Mechanistic studies show that Tel2 associates with the hhex promoter region and promotes hhex transcription. Our results reveal roles of Tel2 in the BP-PC redifferentiation process of liver regeneration by activating hhex.

DNA methylation maintenance at the p53 locus initiates biliary-mediated liver regeneration.

In cases of extensive liver injury, biliary epithelial cells (BECs) dedifferentiate into bipotential progenitor cells (BPPCs), then redifferentiate into hepatocytes and BECs to accomplish liver regeneration. Whether epigenetic regulations, particularly DNA methylation maintenance enzymes, play a role in this biliary-mediated liver regeneration remains unknown. Here we show that in response to extensive hepatocyte damages, expression of dnmt1 is upregulated in BECs to methylate DNA at the p53 locus, which represses p53 transcription, and in turn, derepresses mTORC1 signaling to activate BEC dedifferentiation. After BEC dedifferentiation and BPPC formation, DNA methylation at the p53 locus maintains in BPPCs to continue blocking p53 transcription, which derepresses Bmp signaling to induce BPPC redifferentiation. Thus, this study reveals promotive roles and mechanisms of DNA methylation at the p53 locus in both dedifferentiation and redifferentiation stages of biliary-mediated liver regeneration, implicating DNA methylation and p53 as potential targets to stimulate regeneration after extensive liver injury.

Impact of the COVID-19 pandemic on neonatal admissions in a tertiary children's hospital in southwest China: An interrupted time-series study.

BACKGROUND: The unprecedented coronavirus disease 2019 (COVID-19) pandemic has caused millions of infections worldwide and represents a significant challenge facing modern health care systems. This study was conducted to investigate the impact of lockdown measures in a tertiary Children's Hospital in southwest China, which might be used to predict long-term effects related to health-seeking behavior of parents/caregivers. METHODS: This study included newborns enrolled over a span of 86 weeks between January 4, 2019, and August 27, 2020. We designated two time periods for analysis purposes: a stable pre-COVID period(55 weeks between January 4, 2019, and January 23, 2020) and a COVID-impacted period (31 weeks between January 24, 2020, and August 27, 2020). An interrupted time-series analysis was employed to compare changes and trends in hospital admissions and disease spectra before and after the period of nonpharmaceutical interventions (NPIs). Furthermore, this study was conducted to evaluate whether the health-seeking behavior of parents/caregivers was influenced by pandemic factors. RESULTS: Overall, 16,640 infants were admitted to the neonatology department during the pre-COVID period (n = 12,082) and the COVID-impacted period (n = 4,558). The per week neonatal admissions consistently decreased following the first days of NPIs (January 24, 2020). The average weekly admission rates of 220/week pre-COVID period and 147/week COVID-impacted period. There was an evident decrease in the volume of admissions for all disease spectra after the intervention, whereas the decrease of patients complaining about pathological jaundice-related conditions was statistically significant (p<0.05). In the COVID-impacted period, the percentage of patients who suffered from respiratory system diseases, neonatal encephalopathy, and infectious diseases decreased, while the percentage of pathological jaundice-related conditions and gastrointestinal system diseases increased. The neonatal mortality rates (NMRs) increased by 8.7% during the COVID-impacted period compared with the pre-COVID period. CONCLUSIONS: In summary, there was a significant decline in neonatal admissions in a tertiary care hospital during the COVID-19 Pandemic and the associated NPIs. Additionally, this situation had a remarkable impact on disease spectra and health-seeking behavior of parents/caregivers. We, therefore, advise continuing follow-ups and monitoring the main health indicators in vulnerable populations affected by this Pandemic over time.

Salivary Microbiome in Adenoid Cystic Carcinoma Detected by 16S rRNA Sequencing and Shotgun Metagenomics.

Microorganisms are confirmed to be closely related to the occurrence and development of cancers in human beings. However, there has been no published report detailing relationships between the oral microbiota and salivary adenoid cystic carcinoma (SACC). In this study, unstimulated saliva was collected from 13 SACC patients and 10 healthy controls. The microbial diversities, compositions and functions were comprehensively analyzed after 16S rRNA sequencing and whole-genome shotgun metagenomic sequencing. The alpha diversity showed no significant difference between SACC patients and healthy controls, while beta diversity showed a separation trend. The SACC patients showed higher abundances of Streptococcus and Rothia, while Prevotella and Alloprevotella were more abundant in healthy controls. The prevalent KEGG pathways, carbohydrate-active enzymes, antibiotic resistances and virulence factors as well as the biomarkers in SACC were determined by functional gene analysis. Our study preliminarily investigated the salivary microbiome of SACC patients compared with healthy controls and might be the basis for further studies on novel diagnostic and treatment strategies.

Formimidoyltransferase cyclodeaminase prevents the starvation-induced liver hepatomegaly and dysfunction through downregulating mTORC1.

The liver is a crucial center in the regulation of energy homeostasis under starvation. Although downregulation of mammalian target of rapamycin complex 1 (mTORC1) has been reported to play pivotal roles in the starvation responses, the underpinning mechanisms in particular upstream factors that downregulate mTORC1 remain largely unknown. To identify genetic variants that cause liver energy disorders during starvation, we conduct a zebrafish forward genetic screen. We identify a liver hulk (lvh) mutant with normal liver under feeding, but exhibiting liver hypertrophy under fasting. The hepatomegaly in lvh is caused by enlarged hepatocyte size and leads to liver dysfunction as well as limited tolerance to starvation. Positional cloning reveals that lvh phenotypes are caused by mutation in the ftcd gene, which encodes the formimidoyltransferase cyclodeaminase (FTCD). Further studies show that in response to starvation, the phosphorylated ribosomal S6 protein (p-RS6), a downstream effector of mTORC1, becomes downregulated in the wild-type liver, but remains at high level in lvh. Inhibition of mTORC1 by rapamycin rescues the hepatomegaly and liver dysfunction of lvh. Thus, we characterize the roles of FTCD in starvation response, which acts as an important upstream factor to downregulate mTORC1, thus preventing liver hypertrophy and dysfunction.

Acute brain vascular regeneration occurs via lymphatic transdifferentiation.

Acute ischemic stroke damages the regional brain blood vessel (BV) network. Acute recovery of basic blood flows, which is carried out by the earliest regenerated BVs, are critical to improve clinical outcomes and minimize lethality. Although the late-regenerated BVs form via growing along the meninge-derived ingrown lymphatic vessels (iLVs), mechanisms underlying the early, acute BV regeneration remain elusive. Using zebrafish cerebrovascular injury models, we show that the earliest regenerated BVs come from lymphatic transdifferentiation, a hitherto unappreciated process in vertebrates. Mechanistically, the LV-to-BV transdifferentiation occurs exclusively in the stand-alone iLVs through Notch activation. In the track iLVs adhered by late-regenerated BVs, transdifferentiation never occurs because the BV-expressing EphrinB2a paracellularly activates the iLV-expressing EphB4a to inhibit Notch activation. Suppression of LV-to-BV transdifferentiation blocks acute BV regeneration and becomes lethal. These results demonstrate that acute BV regeneration occurs via lymphatic transdifferentiation, suggesting this process and key regulatory molecules EphrinB2a/EphB4a/Notch as new postischemic therapeutic targets.

Zebrafish as a Model for Germ Cell Regeneration.

Germ cell acts as a link between transfer of genetic information and process of species evolution. Defects or malformations of germ cells can lead to infertility or tumors. Germ cell regeneration is one of the effective ways to treat the infertility. Therefore, it is of great scientific and clinical interests to dissect the cellular and molecular mechanisms underlying germ cell regeneration. Progress have already been achieved in germ cell regeneration using model organisms for decades. However, key open issues regarding the underpinning mechanisms still remain poorly understood. Zebrafish is well known for its powerful regenerative capacity to regenerate various tissues and organs. Recently, advances in genomics, genetics, microscopy, and single cell technologies have made zebrafish an attractive model to study germ cell development and regeneration. Here we review recent technologies for the study of germ cell regeneration in zebrafish, highlight the potential of germline stem cells (GSCs) in the contribution to reproductive system regeneration, and discuss the nanos. Wnt signaling and germ cell-specific factors involved in the regulation of germ cell regeneration.

Farnesoid X Receptor Is Required for the Redifferentiation of Bipotential Progenitor Cells During Biliary-Mediated Zebrafish Liver Regeneration.

BACKGROUND AND AIMS: Liver regeneration after extreme hepatocyte loss occurs through transdifferentiation of biliary epithelial cells (BECs), which includes dedifferentiation of BECs into bipotential progenitor cells (BPPCs) and subsequent redifferentiation into nascent hepatocytes and BECs. Although multiple molecules and signaling pathways have been implicated to play roles in the BEC-mediated liver regeneration, mechanisms underlying the dedifferentiation-redifferentiation transition and the early phase of BPPC redifferentiation that is pivotal for both hepatocyte and BEC directions remain largely unknown. APPROACH AND RESULTS: The zebrafish extreme liver damage model, genetic mutation, pharmacological inhibition, transgenic lines, whole-mount and fluorescent in situ hybridizations and antibody staining, single-cell RNA sequencing, quantitative real-time PCR, and heat shock-inducible overexpression were used to investigate roles and mechanisms of farnesoid X receptor (FXR; encoded by nuclear receptor subfamily 1, group H, member 4 [nr1h4]) in regulating BPPC redifferentiation. The nr1h4 expression was significantly up-regulated in response to extreme liver injury. Genetic mutation or pharmacological inhibition of FXR was ineffective to BEC-to-BPPC dedifferentiation but blocked the redifferentiation of BPPCs to both hepatocytes and BECs, leading to accumulation of undifferentiated or less-differentiated BPPCs. Mechanistically, induced overexpression of extracellular signal-related kinase (ERK) 1 (encoded by mitogen-activated protein kinase 3) rescued the defective BPPC-to-hepatocyte redifferentiation in the nr1h4 mutant, and ERK1 itself was necessary for the BPPC-to-hepatocyte redifferentiation. The Notch activities in the regenerating liver of nr1h4 mutant attenuated, and induced Notch activation rescued the defective BPPC-to-BEC redifferentiation in the nr1h4 mutant. CONCLUSIONS: FXR regulates BPPC-to-hepatocyte and BPPC-to-BEC redifferentiations through ERK1 and Notch, respectively. Given recent applications of FXR agonists in the clinical trials for liver diseases, this study proposes potential underpinning mechanisms by characterizing roles of FXR in the stimulation of dedifferentiation-redifferentiation transition and BPPC redifferentiation.

A single-cell-resolution fate map of endoderm reveals demarcation of pancreatic progenitors by cell cycle.

A progenitor cell could generate a certain type or multiple types of descendant cells during embryonic development. To make all the descendant cell types and developmental trajectories of every single progenitor cell clear remains an ultimate goal in developmental biology. Characterizations of descendant cells produced by each uncommitted progenitor for a full germ layer represent a big step toward the goal. Here, we focus on early foregut endoderm, which generates foregut digestive organs, including the pancreas, liver, foregut, and ductal system, through distinct lineages. Using unbiased single-cell labeling techniques, we label every individual zebrafish foregut endodermal progenitor cell out of 216 cells to visibly trace the distribution and number of their descendant cells. Hence, single-cell-resolution fate and proliferation maps of early foregut endoderm are established, in which progenitor regions of each foregut digestive organ are precisely demarcated. The maps indicate that the pancreatic endocrine progenitors are featured by a cell cycle state with a long G1 phase. Manipulating durations of the G1 phase modulates pancreatic progenitor populations. This study illustrates foregut endodermal progenitor cell fate at single-cell resolution, precisely demarcates different progenitor populations, and sheds light on mechanistic insights into pancreatic fate determination.

Contributions of biliary epithelial cells to hepatocyte homeostasis and regeneration in zebrafish.

Whether transdifferentiation of the biliary epithelial cells (BECs) to hepatocytes occurs under physiological conditions and contributes to liver homeostasis remains under long-term debate. Similar questions have been raised under pathological circumstances if a fibrotic liver is suffered from severe injuries. To address these questions in zebrafish, we established a sensitive lineage tracing system specific for the detection of BEC-derived hepatocytes. The BEC-to-hepatocyte transdifferentiation occurred and became minor contributors to hepatocyte homeostasis in a portion of adult individuals. The BEC-derived hepatocytes distributed in clusters in the liver. When a fibrotic liver underwent extreme hepatocyte damages, BEC-to-hepatocyte transdifferentiation acted as the major origin of regenerating hepatocytes. In contrast, partial hepatectomy failed to induce the BEC-to-hepatocyte conversion. In conclusion, based on a sensitive lineage tracing system, our results suggest that BECs are able to transdifferentiate into hepatocytes and contribute to both physiological hepatocyte homeostasis and pathological regeneration.

Prognostic Value of Clinical Tests in Neonates With Hypoxic-Ischemic Encephalopathy Treated With Therapeutic Hypothermia: A Systematic Review and Meta-Analysis.

Background and Objective: There remains an unmet clinical need for markers that predict outcomes in the hypothermia-treated (HT) infants with HIE. The aim of this meta-analysis was to investigate the prognostic accuracy of currently available clinical tests performed in the immediate post-natal period for predicting neurological outcomes between 18 months and 3 years of age in HT near-term and term infants with perinatal asphyxia and HIE. Methods: A comprehensive review of the Embase, Cochrane library, and PubMed databases was performed to identify studies that evaluated the prognostic value of clinical tests for neurological outcomes in HT near-term and term infants with perinatal asphyxia and hypoxic-ischemic encephalopathy. Pooled sensitivity and specificity with corresponding 95% confidence intervals and area under the receiver operating characteristic (ROC) curve (AUC) were calculated. Results: Of the 1,144 relevant studies, 26 studies describing four clinical tests conducted in 1458 HT near-term or term infants were included. For predicting an unfavorable neurological outcome, of the imaging techniques, MRI within 2 weeks of birth performed best on sensitivity 0.85 (95% CI 0.79-0.89), specificity 0.72 (95% CI 0.66-0.77), and AUC 0.88; among the neurophysiological tests, multichannel EEG (Electroencephalogram) demonstrated the sensitivity 0.63 (95% CI 0.49-0.76), specificity 0.82 (95% CI 0.70-0.91), and AUC 0.88, and for aEEG (amplitude-integrated electroencephalography) background pattern pooled sensitivity, specificity and AUC were 0.90 (95% CI 0.86-0.94), 0.46 (95% CI 0.42-0.51), and 0.78 whereas for SEPs (Somatosensory evoked potentials), pooled sensitivity and specificity were 0.52 (95% CI 0.34-0.69), 0.76 (95% CI 0.63-0.87), and AUC 0.84, respectively. Conclusions: In the wake of the era of TH, MRI and neurophysiological tests (aEEG or EEG) were promising predictors of adverse outcomes, while SEPs need high-quality studies to confirm the findings. Continued follow-up of the children and well-designed large prospective studies are essential to determine whether these benefits are maintained in later childhood.

[Expression patterns of ectodysplasin and ectodysplasin receptor during early dental development in zebrafish].

OBJECTIVE: This study aims to study the expression patterns of ectodysplasin (EDA) and ectodysplasin receptor (EDAR) during the early development of zebrafish and provide a foundation for further research of the Eda signaling pathway in tooth development. METHODS: Total RNA was extracted from zebrafish embryos at 48 hours postfertilization (hpf) and then reverse transcribed for cDNA library generation. The corresponding RNA polymerase was selected for the synthesis of the digoxin-labeled antisense mRNA probe of zebrafish pharyngeal tooth specific marker dlx2b and Eda signaling-associated genes eda and edar in vitro. The three sequences were ligated into a pGEMT vector with a TA cloning kit, and polymerase chain reaction (PCR) was applied to linearize the plasmid. The resultant PCR sequences were used as templates for synthesizing Dig-labeled mRNA probe dlx2b, eda, and edar. Zebrafish embryos were collected at 36, 48, 56, 60, 72, and 84 hpf, then whole mount in situ hybridization was performed for the detection of eda and edar expression patterns. Then, their expression patterns at 72 hpf were compared with the expression pattern of dlx2b. RESULTS: The mRNA antisense probes of dlx2b, eda, and edar were successfully obtained. The positive signals of eda and edar were observed in zebrafish pharyngeal tooth region at 48-72 hpf and thus conform to the signals of dlx2b in the positive regions. CONCLUSIONS: The ligand eda and edar, which are associated with the Eda signaling pathway, are strongly expressed only at the pharyngeal tooth region in zebrafish from tooth initiation to the morphogenesis stage. Thus, the Eda signaling pathway may be involved in the regulation of the early development of zebrafish pharyngeal teeth.

[Oral microbiological diversity in patients with salivary adenoid cystic carcinoma].

OBJECTIVE: The aim of this study was to identify the differences in microbial diversity and community in patients with salivary adenoid cystic carcinoma (SACC). METHODS: Saliva was collected from 13 patients with SACC confirmed by histopathological diagnosis and 10 healthy control subjects. Total metagenomic DNA was extracted. The DNA amplicons of the V3-V4 hypervariable regions of the 16S rRNA gene were generated and subjected to high-throughput sequencing. Microbial diversity and community structure were analyzed with Mothur software. RESULTS: A total of 16 genera of dominant bacteria in the SACC group were found, including Streptococcus (36.68%), Neisseria (8.55%), Prevotella_7 (7.53%), and Veillonella (6.37%), whereas 15 dominant bacteria in the control group were found, including Streptococcus (18.41%), Neisseria (18.20%), Prevotella_7 (8.89%), Porphyromonas (6.20%), Fusobacterium (5.86%) and Veillonella (5.82%). The statistically different phyla between the two groups were Firmicutes, Proteobacteria and Fusobacterium (P<0.05). The statistically different genera between the two groups were Streptococcus, Neisseria and Porphyromonas (P<0.05), and Capnocytophaga was only detected in patients with SACC. CONCLUSIONS: Significant differences were observed in the oral microorganisms between the two groups.

Cerebrovascular Injuries Induce Lymphatic Invasion into Brain Parenchyma to Guide Vascular Regeneration in Zebrafish.

Damage to regional cerebrovascular networks 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 the 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 to drain interstitial fluid to resolve brain edema and 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."

The CCCH-type zinc finger transcription factor Zc3h8 represses NF-κB-mediated inflammation in digestive organs in zebrafish.

Degenerative diseases of organs lead to their impaired function. The cellular and molecular mechanisms underlying organ degeneration are therefore of great research and clinical interest but are currently incompletely characterized. Here, using a forward-genetic screen for genes regulating liver development and function in zebrafish, we identified a cq5 mutant that exhibited a liver-degeneration phenotype at 5 days postfertilization, the developmental stage at which a functional liver develops. Positional cloning revealed that the liver degeneration was caused by a single point mutation in the gene zc3h8 (zinc finger CCCH-type containing 8), changing a highly conserved histidine to glutamine at position 353 of the Zc3h8 protein. The zc3h8 mutation-induced liver degeneration in the mutant was accompanied by reduced proliferation, increased apoptosis, and macrophage phagocytosis of hepatocytes. Transcriptional profile analyses revealed up-regulation and activation of both proinflammatory cytokines and the NF-κB signaling pathway in the zc3h8 mutant. Suppression of NF-κB signaling activity efficiently rescued the proinflammatory cytokine response, as well as the inflammation-mediated liver degeneration phenotype of the mutant. Of note, the zc3h8 mutation-induced degeneration of several other organs, including the gut and exocrine pancreas, indicating that Zc3h8 is a general repressor of inflammation in zebrafish. Collectively, our findings demonstrate that Zc3h8 maintains organ homeostasis by inhibiting the NF-κB-mediated inflammatory response in zebrafish and that Zc3h8 dysfunction causes degeneration of multiple organs, including the liver, gut, and pancreas.

C1qr and C1qrl redundantly regulate angiogenesis in zebrafish through controlling endothelial Cdh5.

Angiogenesis plays central role in the formation of functional circulation system. Characterizations of the involved factors and signaling pathways remain to be the key interest in the angiogenesis research. In this report, we showed that c1qr/cd93 and c1qrl/clec14a are specifically expressed in the vascular endothelial cells during zebrafish development. Single mutation of c1qr or c1qrl is associated with slightly malformation of inter-segmental vessels (ISVs), whereas double mutant exhibits severe defects in the ISVs formation without affecting early vasculogenesis. Further studies reveal that the endothelial-endothelial junctional molecule Cdh5 becomes absent in the ISVs of the double mutant. Replenishment of Cdh5 efficiently rescue the impaired angiogenesis in the c1qr/c1qrl double mutant. These data demonstrate that c1qr and c1qrl redundantly regulate angiogenesis through controlling the expression of the endothelial junctional molecule Cdh5, thus playing an important role in angiogenesis.

Macrophages Mediate the Repair of Brain Vascular Rupture through Direct Physical Adhesion and Mechanical Traction.

Hemorrhagic stroke and brain microbleeds are caused by cerebrovascular ruptures. Fast repair of such ruptures is the most promising therapeutic approach. Due to a lack of high-resolution in vivo real-time studies, the dynamic cellular events involved in cerebrovascular repair remain unknown. Here, we have developed a cerebrovascular rupture system in zebrafish by using multi-photon laser, which generates a lesion with two endothelial ends. In vivo time-lapse imaging showed that a macrophage arrived at the lesion and extended filopodia or lamellipodia to physically adhere to both endothelial ends. This macrophage generated mechanical traction forces to pull the endothelial ends and facilitate their ligation, thus mediating the repair of the rupture. Both depolymerization of microfilaments and inhibition of phosphatidylinositide 3-kinase or Rac1 activity disrupted macrophage-endothelial adhesion and impaired cerebrovascular repair. Our study reveals a hitherto unexpected role for macrophages in mediating repair of cerebrovascular ruptures through direct physical adhesion and mechanical traction.