Blood Vessel Leakage Research Articles

Anti-VEGFR2 neutralising antibody slows the progression of multistep oral carcinogenesis.

Angiogenesis plays an important role in cancer growth and metastasis, and it is considered a therapeutic target to control tumour growth following anti-angiogenic therapy. However, it is still unclear when tissues initiate angiogenesis during malignant transformation from premalignant condition and whether this premalignant condition could be a therapeutic target of anti-angiogenic therapy. In this study, we aimed to analyse the onset of angiogenesis by evaluating morphological and functional alterations of microvessels during oral multistep carcinogenesis using a 4-nitroquinoline 1-oxide (4NQO)-induced oral carcinogenesis mouse model. In the study, we initially confirmed that with the use of 4NQO, oral lesions develop in a stepwise manner from normal mucosa through oral epithelial dysplasia (OED) to oral squamous cell carcinoma (OSCC). Evaluation of CD31-immunostained specimens revealed that microvessel density (MVD) increases in a stepwise manner from OEDs. Histological and functional analyses revealed the structural abnormalities and leakage of blood vessels had already taken place in OED. Then we evaluated the expression profiles of Hif1a and Vegfa along with hypoxic status and found that OED exhibited increased Vegfa expression under hypoxic conditions. Finally, we tested the possibility of OEDs as a target of anti-angiogenic therapy and found that anti-VEGFR2 neutralising antibody in OED slowed the disease progression from OED to OSCC. These data indicate that an angiogenic switch occurs at the premalignant stage and morphological, and functional alterations of microvessels already exist in OED. These findings also elucidate the tumour microenvironment, which gradually develops along with carcinogenic processes, and highlight usefulness of the 4NQO-induced carcinogenesis model in the study of epithelial and stromal components, which will support epithelial carcinogenesis. © 2024 The Author(s). The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

GGC repeat expansions in NOTCH2NLC cause uN2CpolyG cerebral amyloid angiopathy.

The expansion of GGC repeats within NOTCH2NLC leads to the translation of the uN2CpolyG protein, the primary pathogenic factor in neuronal intranuclear inclusion disease (NIID). This study aims to explore the deposition of uN2CpolyG as an amyloid in the vessel wall, leading to uN2CpolyG cerebral amyloid angiopathy (CAA)-related cerebral microbleeds (CMBs). A total of 97 patients with genetically confirmed NIID were enrolled in this study. We analyzed the presence of CMBs using susceptibility-weighted imaging sequences and compared general clinical information, cerebrovascular risk factors, stroke history, antiplatelet medication use, and MRI features between NIID patients with and without CMBs. We further performed hematoxylin and eosin (H&E), Perl's, Congo red, and Thioflavin S staining, ubiquitin, p62 and uN2CpolyG immunostaining on brain tissue obtained from four NIID patients. A total of 354 CMBs were detected among 41 patients with NIID, with nearly half located in the deep brain, one-third in the lobes, and approximately 20% in the infratentorial area. No significant differences in cerebrovascular disease risk factors or history of antiplatelet drug use were observed between patients with and without CMBs. However, patients with CMBs suffered a higher incidence of previous ischemic and hemorrhagic stroke events. This group also had a higher incidence of recent subcortical infarcts and a higher proportion of white matter lesions in the external capsule and temporal pole. Conversely, patients without CMBs showed higher detection of high signals at the corticomedullary junction on diffusion-weighted imaging and more pronounced brain atrophy. H&E staining showed blood vessel leakage and hemosiderin-laden macrophage clusters, and Prussian blue staining revealed brain tissue iron deposition. CMBs occurred more frequently in small vessels lacking intranuclear inclusions, and extensive degeneration of endothelial cells and smooth muscle fibres was observed mainly in vessels lacking inclusions. Congo red-positive amyloid deposition was observed in the cerebral vessels of NIID patients, with disordered filamentous fibres appearing under an electron microscope. Additionally, the co-localization of Thioflavin S-labeled amyloid and uN2CpolyG protein in the cerebral vascular walls of NIID patients further suggested that uN2CpolyG is the main pathogenic protein in this form of amyloid angiopathy. In conclusion, we reviewed patients with GGC repeat expansion of NOTCH2NLC from a novel perspective, providing initial clinical, neuroimaging, and pathological evidence suggesting that uN2CpolyG may contribute to a distinct type of CAA.

Molecular hydrogen promotes retinal vascular regeneration and attenuates neovascularization and neuroglial dysfunction in oxygen-induced retinopathy mice

BackgroundRetinopathy of Prematurity (ROP) is a proliferative retinal vascular disease occurring in the retina of premature infants and is the main cause of childhood blindness. Nowadays anti-VEGF and retinal photocoagulation are mainstream treatments for ROP, but they develop a variety of complications. Hydrogen (H2) is widely considered as a useful neuroprotective and antioxidative therapeutic method for hypoxic-ischemic disease without toxic effects. However, whether H2 provides physiological angiogenesis promotion, neovascularization suppression and glial protection in the progression of ROP is largely unknown.This study aims to investigate the effects of H2 on retinal angiogenesis, neovascularization and neuroglial dysfunction in the retinas of oxygen-induced retinopathy (OIR) mice.MethodsIn this study, mice that were seven days old and either wild-type (WT) or Nrf2-deficient (Nrf2−/−) were exposed to 75% oxygen for 5 days and then returned to normal air conditions. Different stages of hydrogen gas (H2) inhalation were administered. Vascular obliteration, neovascularization, and blood vessel leakage were analyzed and compared. To count the number of neovascularization endothelial nuclei, routine HE staining of retinal sections was conducted. Immunohistochemistry was performed using DyLight 594 labeled GSL I-isolectin B4 (IB4), as well as primary antibodies against proliferating cell nuclear antigen (PCNA), glial fibrillary acidic protein (GFAP), and Iba-1. Western blots were used to measure the expression of NF-E2-related factor 2 (Nrf2), vascular endothelial growth factor (VEGF), Notch1, Dll4, and HIF-1α. Additionally, the expression of target genes such as NQO1, HO-1, Notch1, Hey1, Hey2, and Dll4 was measured. Human umbilical vein endothelial cells (HUVECs) treated with H2 under hypoxia were used as an in vitro model. RT-PCR was used to evaluate the mRNA expression of Nrf2, Notch/Dll4, and the target genes. The expression of reactive oxygen species (ROS) was observed using immunofluorescence staining.ResultsOur results indicate that 3–4% H2 does not disturb retinal physiological angiogenesis, but ameliorates vaso-obliteration and neovascularization in OIR mice. Moreover, H2 prevents the decreased density and reverses the morphologic and functional changes in retinal astrocytes caused by oxygen-induced injury. In addition, H2 inhalation reduces microglial activation, especially in the area of neovascularization in OIR mice. H2 plays a protective role in vascular regeneration by promoting Nrf2 activation and suppressing the Dll4-induced Notch signaling pathway in vivo. Also, H2 promotes the proliferation of HUVECs under hypoxia by negatively regulating the Dll4/Notch pathway and reducing ROS levels through Nrf2 pathway aligning with our findings in vivo.Moreover, the retinal oxygen-sensing mechanisms (HIF-1α/VEGF) are also involved in hydrogen-mediated retinal revascularization and neovascularization suppression.ConclusionsCollectively, our results indicate that H2 could be a promising therapeutic agent for POR treatment and that its beneficial effect in human ROP might involve the activation of the Nrf2-Notch axis as well as HIF-1α/VEGF pathways.Graphical

A Focus on the Pathophysiology of Adrenomedullin Expression: Endothelitis and Organ Damage in Severe Viral and Bacterial Infections.

Adrenomedullin (ADM) is a peptide hormone produced primarily in the adrenal glands, playing a crucial role in various physiological processes. As well as improving vascular integrity and decreasing vascular permeability, ADM acts as a vasodilator, positive inotrope, diuretic, natriuretic and bronchodilator, antagonizing angiotensin II by inhibiting aldosterone secretion. ADM also has antihypertrophic, anti-apoptotic, antifibrotic, antioxidant, angiogenic and immunoregulatory effects and antimicrobial properties. ADM expression is upregulated by hypoxia, inflammation-inducing cytokines, viral or bacterial substances, strength of shear stress, and leakage of blood vessels. These pathological conditions are established during systemic inflammation that can result from infections, surgery, trauma/accidents or burns. The ability to rapidly identify infections and the prognostic, predictive power makes it a valuable tool in severe viral and bacterial infections burdened by high incidence and mortality. This review sheds light on the pathophysiological processes that in severe viral or bacterial infections cause endothelitis up to the development of organ damage, the resulting increase in ADM levels dosed through its more stable peptide mid-regional proadrenomedullin (MR-proADM), the most significant studies that attest to its diagnostic and prognostic accuracy in highlighting the severity of viral or bacterial infections and appropriate therapeutic insights.

Photoreceptors inhibit pathological retinal angiogenesis through transcriptional regulation of Adam17 via c-Fos

Pathological retinal angiogenesis profoundly impacts visual function in vascular eye diseases, such as retinopathy of prematurity (ROP) in preterm infants and age-related macular degeneration in the elderly. While the involvement of photoreceptors in these diseases is recognized, the underlying mechanisms remain unclear. This study delved into the pivotal role of photoreceptors in regulating abnormal retinal blood vessel growth using an oxygen-induced retinopathy (OIR) mouse model through the c-Fos/A disintegrin and metalloprotease 17 (Adam17) axis. Our findings revealed a significant induction of c-Fos expression in rod photoreceptors, and c-Fos depletion in these cells inhibited pathological neovascularization and reduced blood vessel leakage in the OIR mouse model. Mechanistically, c-Fos directly regulated the transcription of Adam17 a shedding protease responsible for the production of bioactive molecules involved in inflammation, angiogenesis, and cell adhesion and migration. Furthermore, we demonstrated the therapeutic potential by using an adeno-associated virus carrying a rod photoreceptor-specific short hairpin RNA against c-fos which effectively mitigated abnormal retinal blood vessel overgrowth, restored retinal thickness, and improved electroretinographic (ERG) responses. In conclusion, this study highlights the significance of photoreceptor c-Fos in ROP pathology, offering a novel perspective for the treatment of this disease.

Preserving sight: Managing and preventing diabetic retinopathy

Abstract Preserving eyesight in the presence of diabetic retinopathy, a subtle complication of diabetes, requires a proactive and comprehensive approach. This condition, characterized by the gradual deterioration of the blood vessels in the retina, can quietly advance and pose a significant threat to vision. Effectively managing this issue involves careful control of blood glucose levels through medications, dietary choices, and physical activity. By preventing prolonged high blood sugar levels, the main culprit behind retinal damage, individuals can slow down the progression of diabetic retinopathy and minimize its impact on vision. Medical interventions, such as laser therapies and anti-vascular endothelial growth factor injections, offer targeted measures to address abnormal blood vessel growth and leakage. Regular eye assessments are crucial, providing early detection opportunities even in the absence of obvious symptoms and allowing for timely intervention before irreversible damage occurs. Prevention is equally important, emphasizing proactive measures to avoid diabetic retinopathy. Monitoring blood pressure and lipid levels, along with adopting a health-conscious lifestyle, strengthens cardiovascular health, indirectly protecting the fragile retinal blood vessels. By following diabetes management protocols, maintaining a balanced diet, engaging in regular physical activity, and avoiding tobacco use, individuals can significantly reduce their vulnerability to diabetic retinopathy. Combined with efforts to increase community awareness and educate about the importance of regular eye check-ups, this comprehensive approach empowers individuals to actively manage their eye health, ultimately preserving the precious gift of sight.

The Multi-Kinase Inhibitor RepSox Enforces Barrier Function in the Face of Both VEGF and Cytokines.

The therapeutic benefit provided by anti-vascular endothelial growth factor (VEGF) for patients with vision-threatening conditions such as diabetic retinopathy (DR) demonstrates the important role of VEGF in this affliction. Cytokines, which can be elevated in the vitreous of patients with DR, promote leakage of retinal blood vessels, and may also contribute to pathology, especially in those patients for whom anti-VEGF does not provide adequate benefit. In this in vitro study using primary human retinal endothelial cells, we compared anti-VEGF with the (transforming growth factor beta) TGFβ receptor inhibitor RepSox (RS) for their ability to enforce barrier function in the face of VEGF, cytokines, and the combination of both. RS was superior to anti-VEGF because it prevented permeability in response to VEGF, cytokines, and their combination, whereas anti-VEGF was effective against VEGF alone. The inhibitory effect of RS was associated with suppression of both agonist-induced pore formation and disorganization of adherens junctions. RS-mediated inhibition of the TGFβ pathway and increased expression of claudin-5 did not adequately explain how RS stabilized the endothelial cell barrier. Finally, RS not only prevented barrier relaxation, but also completely or partially reclosed a barrier relaxed with tumor necrosis factor α (TNF α) or VEGF, respectively. These studies demonstrate that RS stabilized the endothelial barrier in the face of both cytokines and VEGF, and thereby identify RS as a therapeutic that has the potential to overcome permeability driven by multiple agonists that play a role in the pathology of DR.

Long Non-Coding RNAs and Proliferative Retinal Diseases.

Retinopathy refers to disorders that affect the retina of the eye, which are frequently caused by damage to the retina's vascular system. This causes leakage, proliferation, or overgrowth of blood vessels through the retina, which can lead to retinal detachment or breakdown, resulting in vision loss and, in rare cases, blindness. In recent years, high-throughput sequencing has significantly hastened the discovery of new long non-coding RNAs (lncRNAs) and their biological functions. LncRNAs are rapidly becoming recognized as critical regulators of several key biological processes. Current breakthroughs in bioinformatics have resulted in the identification of several lncRNAs that may have a role in retinal disorders. Nevertheless, mechanistic investigations have yet to reveal the relevance of these lncRNAs in retinal disorders. Using lncRNA transcripts for diagnostic and/or therapeutic purposes may aid in the development of appropriate treatment regimens and long-term benefits for patients, as traditional medicines and antibody therapy only provide temporary benefits that must be repeated. In contrast, gene-based therapies can provide tailored, long-term treatment solutions. Here, we will discuss how different lncRNAs affect different retinopathies, including age-related macular degeneration (AMD), diabetic retinopathy (DR), central retinal vein occlusion (CRVO), proliferative vitreoretinopathy (PVR), and retinopathy of prematurity (ROP), which can cause visual impairment and blindness, and how these retinopathies can be identified and treated using lncRNAs.

The role and mechanism of tight junctions in the regulation of salivary gland secretion.

Tight junctions (TJs) are cell-cell interactions that localize at the most apical portion of epithelial/endothelial cells. One of the predominant functions of TJs is to regulate material transport through paracellular pathway, which serves as a selective barrier. In recent years, the expression and function of TJs in salivary glands has attracted great interest. The characteristics of multiple salivary gland TJ proteins have been identified. During salivation, the activation of muscarinic acetylcholine receptor and transient receptor potential vanilloid subtype 1, as well as other stimuli, promote the opening of acinar TJs by inducing internalization of TJs, thereby contributing to increased paracellular permeability. Besides, endothelial TJs are also redistributed with leakage of blood vessels in cholinergic-stimulated submandibular glands. Furthermore, under pathological conditions, such as Sjögren's syndrome, diabetes mellitus, immunoglobulin G4-related sialadenitis, and autotransplantation, the integrity and barrier function of TJ complex are impaired and may contribute to hyposalivation. Moreover, in submandibular glands of Sjögren's syndrome mouse model and patients, the endothelial barrier is disrupted and involved in hyposecretion and lymphocytic infiltration. These findings enrich our understanding of the secretory mechanisms that link the importance of epithelial and endothelial TJ functions to salivation under both physiological and pathophysiological conditions.

Mechanobiological Adaptation to Hyperosmolarity Enhances Barrier Function in Human Vascular Microphysiological System.

In infectious disease such as sepsis and COVID-19, blood vessel leakage treatment is critical to prevent fatal progression into multi-organ failure and ultimately death, but the existing effective therapeutic modalities that improve vascular barrier function are limited. Here, this study reports that osmolarity modulation can significantly improve vascular barrier function, even in an inflammatory condition. 3D human vascular microphysiological systems and automated permeability quantification processes for high-throughput analysis of vascular barrier function are utilized. Vascular barrier function is enhanced by >7-folds with 24-48h hyperosmotic exposure (time window of emergency care; >500mOsm L-1 ) but is disrupted after hypo-osmotic exposure (<200mOsm L-1 ). By integrating genetic and protein level analysis, it is shown that hyperosmolarity upregulates vascular endothelial-cadherin, cortical F-actin, and cell-cell junction tension, indicating that hyperosmotic adaptation mechanically stabilizes the vascular barrier. Importantly, improved vascular barrier function following hyperosmotic exposure is maintained even after chronic exposure to proinflammatory cytokines and iso-osmotic recovery via Yes-associated protein signaling pathways. This study suggests that osmolarity modulation may be a unique therapeutic strategy to proactively prevent infectious disease progression into severe stages via vascular barrier function protection.

Photothrombotic Middle Cerebral Artery Occlusion in Mice: A Novel Model of Ischemic Stroke.

Stroke is one of the main causes of death and disability worldwide. Over the past decades, several animal models of focal cerebral ischemia have been developed allowing to investigate pathophysiological mechanisms underlying stroke progression. Despite intense preclinical research efforts, the need for noninvasive mouse models of vascular occlusion targeting the middle cerebral artery yet avoiding mechanical intervention is still pressing. Here, by applying the photothrombotic stroke model to the distal branch of the middle cerebral artery, we developed a novel strategy to induce a targeted occlusion of a large blood vessel in mice. This approach induces unilateral damage encompassing most of the dorsal cortex from the motor up to the visual regions 1 week after stroke. Pronounced limb dystonia one day after the damage is partially recovered after one week. Furthermore, we observe the insurgence of blood vessel leakage and edema formation in the peri-infarct area. Finally, this model elicits a notable inflammatory response revealed as a strong increase in astrocyte density and morphologic complexity in the perilesional region of the cortex compared with both other regions of the ipsilesional and contralesional hemispheres, and in sham-operated mice. To conclude, the stroke model we developed induces in mice the light-mediated occlusion of one of the main targets of human ischemic stroke, the middle cerebral artery, free from the limitations of commonly used preclinical models.

Interacting with tumor cells weakens the intrinsic clockwise chirality of endothelial cells.

Endothelial cells (ECs) possess a strong intrinsic clockwise (CW, or rightward) chirality under normal conditions. Enervating this chirality of ECs significantly impairs the function of the endothelial barrier. Malignant tumor cells (TCs) undergo metastasis by playing upon the abnormal leakage of blood vessels. However, the impact of TCs on EC chirality is still poorly understood. Using a transwell model, we co-cultured the human umbilical vein endothelial cells or human lung microvascular endothelial cells and breast epithelial tumor cell lines to simulate the TC-EC interaction. Using a micropatterning method, we assessed the EC chirality changes induced by paracrine signaling of and physical contact with TCs. We found that the intrinsic clockwise chirality of ECs was significantly compromised by the TC's physical contact, while the paracrine signaling (i.e., without physical contact) of TCs causes minimal changes. In addition, ECs neighboring TCs tend to possess a left bias, while ECs spaced apart from TCs are more likely to preserve the intrinsic right bias. Finally, we found the chirality change of ECs could result from physical binding between CD44 and E-selectin, which activates protein kinase C alpha (PKCα) and induces pseudopodial movement of EC toward TC. Our findings together suggest the crucial role of EC-TC physical interaction in EC chirality and that weakening the EC chirality could potentially compromise the overall endothelial integrity which increases the probability of metastatic cancer spread.

Therapeutic Validation of GEF-H1 Using a De Novo Designed Inhibitor in Models of Retinal Disease.

Inflammation and fibrosis are important components of diseases that contribute to the malfunction of epithelia and endothelia. The Rho guanine nucleotide exchange factor (GEF) GEF-H1/ARHGEF-2 is induced in disease and stimulates inflammatory and fibrotic processes, cell migration, and metastasis. Here, we have generated peptide inhibitors to block the function of GEF-H1. Inhibitors were designed using a structural in silico approach or by isolating an inhibitory sequence from the autoregulatory C-terminal domain. Candidate inhibitors were tested for their ability to block RhoA/GEF-H1 binding in vitro, and their potency and specificity in cell-based assays. Successful inhibitors were then evaluated in models of TGFβ-induced fibrosis, LPS-stimulated endothelial cell-cell junction disruption, and cell migration. Finally, the most potent inhibitor was successfully tested in an experimental retinal disease mouse model, in which it inhibited blood vessel leakage and ameliorated retinal inflammation when treatment was initiated after disease diagnosis. Thus, an antagonist that blocks GEF-H1 signaling effectively inhibits disease features in in vitro and in vivo disease models, demonstrating that GEF-H1 is an effective therapeutic target and establishing a new therapeutic approach.

Retinal Pathophysiological Evaluation in a Rat Model.

A posterior segment eye disease like diabetic retinopathy alters the physiology of the retina. Diabetic retinopathy is characterized by a retinal detachment, breakdown of the blood-retinal barrier (BRB), and retinal angiogenesis. An in vivo rat model is a valuable experimental tool to examine the changes in the structure and function of the retina. We propose three different experimental techniques in the rat model to identify morphological changes of retinal cells, retinal vasculature, and compromised BRB. Retinal histology is used to study the morphology of various retinal cells. Also, quantitative measurement is performed by retinal cell count and thickness measurement of different retinal layers. A BRB breakdown assay is used to determine the leakage of extraocular proteins from the plasma to vitreous tissue due to the breakdown of BRB. Fluorescence angiography is used to study angiogenesis and leakage of blood vessels by visualizing retinal vasculature using FITC-dextran dye.

Handling cerebral malaria patient with limited resources: a case report

Cerebral malaria is an emergency condition. All patients with Plasmodium falciparum infection followed by neurological symptoms should be treated as cerebral malaria. The pathogenesis of cerebral malaria is caused by the damage of blood vessels endothelium due to parasites sequestration, production of pro-inflammatory cytokines and leakage of blood vessels which can cause brain hypoxia. The proper management is needed, however this become quiet challenging issue in the setting of limited resouces. We report a case of a 35 year old patient presenting with a loss of consciousness accompanied by shivering fever for 5 days. On examination of the peripheral blood smear, the Plasmodium falciparum was found. The patient was later diagnosed as cerebral malaria and treated with anti-malarial drugs. The fifth day of treatment the patient has fully alert. In the next day, the patient was allowed to go home. The management of cerebral malaria is challenging, particularly in the area with limited resources.

Tetrahedral framework nucleic acids-based delivery of microRNA-155 inhibits choroidal neovascularization by regulating the polarization of macrophages

Choroidal neovascularization (CNV) is a common pathological feature of various eye diseases and an important cause of visual impairment in middle-aged and elderly patients. In previous studies, tetrahedral framework nucleic acids (tFNAs) showed good carrier performance. In this experiment, we developed microRNA-155-equipped tFNAs (T-155) and explored its biological effects on CNV. Based on the results of in-vitro experiments, T-155 could regulate macrophages into the antiangiogenic M1 type. Then, we injected T-155 into the vitreous of laser-induced CNV model mice and found that T-155 significantly reduced the size and area of CNV, inhibited blood vessel leakage. In summary, we prove that T-155 could regulate the inflammatory process of CNV by polarizing macrophages, thereby improving the symptoms of CNV. Thus, T-155 might become a new DNA-based drug with great potential for treating CNV.

Ferritin nanocages for early theranostics of tumors via inflammation-enhanced active targeting.

Engineered nanocarriers have been widely developed for tumor theranostics. However, the delivery of imaging probes or therapeutic drugs to the tumor pre-formation site for early and accurate detection and therapy remains a major challenge. Here, by using tailor-functionalized human H-ferritin (HFn), we developed a triple-modality nanoprobe IRdye800-M-HFn and achieved the early imaging of tumor cells before the formation of solid tumor tissues. Then, we developed an HFn-doxorubicin (Dox) drug delivery system by loading Dox into the HFn protein cage and achieved early-stage tumor therapy. The intravenous injection of HFn nanoprobes enabled the imaging of tumor cells as early as two days after tumor implantation, and the triple-modality imaging techniques, namely, near-infrared fluorescence molecular imaging (NIR-FMI), magnetic resonance imaging (MRI), and photoacoustic imaging (PAI), ensured the accuracy of detection. Further exploration indicated that HFn could specifically penetrate into pre-solid tumor sites by tumor-associated inflammation-mediated blood vessel leakage, followed by effective accumulation in tumor cells by the specific targeting property of HFn to transferrin receptor 1. Thus, the HFn-Dox drug delivery system delivered Dox into the tumor pre-formation site and effectively killed tumor cells at early stage. IRDye800-M-HFn nanoprobes and HFn-Dox provide promising strategies for early-stage tumor diagnosis and constructive implications for early-stage tumor treatment.

IMG-06. PREDICTING SURVIVAL FROM PERFUSION AND DIFFUSION MRI BY MACHINE LEARNING

INTRODUCTIONMagnetic Resonance Imaging (MRI) is routinely used in the assessment of children’s brain tumours. Reduced diffusion and increased perfusion on MRI are commonly associated with higher grade but there is a lack of quantitative data linking these parameters to survival. Machine learning is increasingly being used to develop diagnostic tools but its use in survival analysis is rare. In this study we combine quantitative parameters from diffusion and perfusion MRI with machine learning to develop a model of survival for paediatric brain tumours. METHOD: 69 children from 4 centres (Birmingham, Liverpool, Nottingham, Newcastle) underwent MRI with diffusion and perfusion (dynamic susceptibility contrast) at diagnosis. Images were processed to form ADC, cerebral blood volume (CBV) and vessel leakage correction (K2) parameter maps. Parameter mean, standard deviation and heterogeneity measures (skewness and kurtosis) were calculated from tumour and whole brain and used in iterative Bayesian survival analysis. The features selected were used for k-means clustering and differences in survival between clusters assessed by Kaplan-Meier and Cox-regression.RESULTSBayesian analysis revealed the 5 top features determining survival to be tumour volume, ADC kurtosis, CBV mean, K2 mean and whole brain CBV mean. K-means clustering using these features showed two distinct clusters (high- and low-risk) which bore significantly different survival characteristics (Hazard Ratio = 5.6).DISCUSSION AND CONCLUSIONDiffusion and perfusion MRI can be used to aid the prediction of survival in children’s brain tumours. Tumour perfusion played a particularly important role in predicting survival despite being less routinely measured than diffusion.

Olfactomedin-like 3 promotes PDGF-dependent pericyte proliferation and migration during embryonic blood vessel formation.

Pericytes promote vessel stability and their dysfunction causes pathologies due to blood vessel leakage. Previously, we reported that Olfactomedin-like 3 (Olfml3) is a matricellular protein with proangiogenic properties. Here, we explored the role of Olfml3 in a knockout mouse model engineered to suppress this protein. The mutant mice exhibited vascular defects in pericyte coverage, suggesting that pericytes influence blood vessel formation in an Olfml3-dependent manner. Olfml3-deficient mice exhibited abnormalities in the vasculature causing partial lethality of embryos and neonates. Reduced pericyte coverage was observed at embryonic day 12.5 and persisted throughout development, resulting in perinatal death of 35% of Olfml3-deficient mice. Cultured Olfml3-deficient pericytes exhibited aberrant motility and altered pericyte association to endothelial cells. Furthermore, the proliferative response of Olfml3-/- pericytes upon PDGF-B stimulation was significantly diminished. Subsequent experiments revealed that intact PDGF-B signaling, mediated via Olfml3 binding, is required for pericyte proliferation and activation of downstream kinase pathways. Our findings suggest a model wherein pericyte recruitment to endothelial cells requires Olfml3 to provide early instructive cue and retain PDGF-B along newly formed vessels to achieve optimal angiogenesis.

The Renin-Angiotensin-Aldosterone System (RAAS) Is One of the Effectors by Which Vascular Endothelial Growth Factor (VEGF)/Anti-VEGF Controls the Endothelial Cell Barrier

Leakage of retinal blood vessels, which is an essential element of diabetic retinopathy, is driven by chronic elevation of vascular endothelial growth factor (VEGF). VEGF quickly relaxes the endothelial cell barrier by triggering signaling events that post-translationally modify pre-existing components of intercellular junctions. VEGF also changes expression of genes that are known to regulate barrier function. Our goal was to identify effectors by which VEGF and anti-VEGF control the endothelial cell barrier in cells that were chronically exposed to VEGF (hours instead of minutes). The duration of VEGF exposure influenced both barrier relaxation and anti-VEGF-mediated closure. Most VEGF-induced changes in gene expression were not reversed by anti-VEGF. Those that were constitute VEGF effectors that are targets of anti-VEGF. Pursuit of such candidates revealed that VEGF used multiple, nonredundant effectors to relax the barrier in cells that were chronically exposed to VEGF. One such effector was angiotensin-converting enzyme, which is a member of the renin-angiotensin-aldosterone system (RAAS). Pharmacologically antagonizing either the angiotensin-converting enzyme or the receptor for angiotensin II attenuated VEGF-mediated relaxation of the barrier. Finally, activating the RAAS reduced the efficacy of anti-VEGF. These discoveries provide a plausible mechanistic explanation for the long-standing appreciation that RAAS inhibitors are beneficial for patients with diabetic retinopathy and suggest that antagonizing the RAAS improves patients' responsiveness to anti-VEGF.