Pericyte Function Research Articles

A short term treatment with a Mek1/2 inhibitor promotes myocardial arteriogenesis and perfusion in vivo: focus on cardiac mural cells

Abstract Background Arteriogenesis is crucial for heart recovery after ischaemia, but cellular and molecular mechanisms able to foster this phenomenon are still poorly characterised. Purpose To discover novel pro-arteriogenic approaches by exploiting cardiac mural cells endowed with arteriogenic capacity: pericytes (PCs) and vascular smooth muscle cells (VSMCs). Methods and results We derived human and murine CD31neg CD34pos cardiac PCs (cPCs) from myocardial samples of adult subjects and confirmed the pericyte phenotype and function in vitro. We discovered that the withdrawal of EGF and bFGF from the culture media induces the differentiation of cPCs into contractile VSMCs. Molecular investigations of pathways associated with the two factors showed that the Mek1/2-Erk1/2 signalling exerts an inhibitory transcriptional control on contractile VSMC genes. Screening of compounds able to interfere with this pathway revealed that PD0325901 – a potent Mek1/2 inhibitor (MeKi) tested in clinical trials for the treatment of cancer – activates the VSMC phenotype in cPCs. We observed a similar effect on coronary artery VSMCs. Next, we interrogated the effect of PD0325901 on cardiac arteriogenesis in vivo. Adult C57BL6/J mice were given the MeKi 10 mg/kg/day or vehicle (DMSO), orally for 14 days (n=11/group). At the endpoint, echocardiographic evaluation of left ventricle (LV) function and dimensions (n=6/group) showed no difference in comparison with the respective baseline in both groups. Effective inhibition of Mek1/2 in the heart of PD-treated mice was confirmed by the reduced immunostaining for the phosphorylated form of Erk1/2. The MeKi cardiotoxicity was ruled out by assessment of cardiomyocytes and vascular cells apoptosis (Tunel) and plasmatic levels of cTn-I. Histological analyses of the hearts (n=5/group) showed an increase in small arterioles (diameter <20μm) density in the LV of PD-mice compared with the DMSO group (16.4 vs 11.7 art/mm2). No change was observed for the capillary density. The drug promoted the maturation of VSMCs within both small and large (>20μm) arterioles, as shown by the higher ratio between the areas of the vascular wall occupied by the mature contractile marker SM myosin heavy chain and the synthetic/early contractile marker alpha-SM actin (αSMA). The PD treatment reduced the fraction of small arterioles covered with a CD34pos layer (53% vs 70% of total arterioles), along with a lower ratio between the areas occupied by adventitial CD34pos cells and αSMApos VSMCs, suggesting a contribution of cPCs to the arteriolar remodelling. Last, the drug improved the LV myocardial perfusion in the PD- vs the DMSO-group (6.8 vs 5.3 ml/min/g of LV tissue, n=6/group). Conclusions We show that a short treatment with a Mek1/2 inhibitor stimulates myocardial arteriogenesis and perfusion without either inducing cardiotoxicity or deteriorating heart function. This may be a novel, intriguing approach to promote therapeutic arteriogenesis. Funding Acknowledgement Type of funding source: Public grant(s) – National budget only. Main funding source(s): British Heart Foundation Centre for Vascular Regeneration II

Identification and functional characterization of circular RNAs in pericytes

Abstract Background Circular RNAs (circRNAs) are generated by back-splicing. They are known to be robustly expressed in a variety of mammalian cell types and organism and have been reported to influence cell biology by acting e.g. as microRNA sponges or regulating host gene expression. Recently, our group reported functionally relevant circRNA expression in endothelial cells. Despite their important role in the cardiovascular system, the expression and function of circRNAs in pericytes is not well studied. Pericytes are perivascular mural cells, important for vessel maturation and endothelial barrier function. Their recruitment towards endothelial cells is mainly meditated by platelet-derived growth factor (PDGF) signaling. However, a more precise understanding of the regulation of pericyte differentiation and survival is necessary. Objective Here, we analyse circRNA expression in pericytes and demonstrate biological relevance of the hypoxia regulated circular RNA PLOD2 (cPLOD2). Methods and results Using RNA Sequencing in ribosomal depleted RNA we characterized the expression of circRNAs in human pericytes under normoxic and hypoxic (1% O2, 48h) conditions. We identified several circular RNAs being regulated upon hypoxia. The identified circular RNAs demonstrated resistance towards RNase-R digestion and lacking of poly-adenylation. Some of them were found to be localized and in the cytosol, whereas others also occur in the nucleus of the cells. Especially cPLOD2 raised our attention since it is significantly upregulated and robustly expressed upon hypoxia. Silencing cPLOD2 by siRNA resulted in significant de-differentiation of pericytes that went along with a loss of cell viability. Mechanistically, transcription factor screening assays revealed that silencing of cPLOD2 enhances the activity of the transcription factors ELK1/SRF, which have been documented to result in de-differentiation of smooth muscle cells. Conclusion Here we characterize the expression pattern of circRNAs in human primary pericytes. Among others, cPLOD2 significantly regulates pericyte function. Our results indicate hypoxia as a major regulator of circRNA expression in pericytes and show that circRNAs are capable of regulating pericyte function by modulating activity of transcription factors. Funding Acknowledgement Type of funding source: Foundation. Main funding source(s): Deutsche Forschungsgesellschaft (DFG) - SFB834; Deutsche Gesellschaft für Herz-Kreislaufforschung (DZHK)

Inhibiting the NLRP3 inflammasome with MCC950 ameliorates retinal neovascularization and leakage by reversing the IL-1\u03b2/IL-18 activation pattern in an oxygen-induced ischemic retinopathy mouse model

Activation of the nucleotide-binding domain leucine-rich repeat and pyrin domain containing receptor 3 (NLRP3) inflammasome plays an important role in ocular neovascularization. In our study, we found that the expression and activation levels of NLRP3 inflammasome components, including NLRP3, an apoptosis-associated speck-like protein (ASC) containing caspase activation and recruitment domain (CARD) and caspase-1 (CAS1), were significantly upregulated. In addition, we found interleukin (IL)-1β activity increased while IL-18 activity decreased in the retinas of oxygen-induced ischemic retinopathy (OIR) mice. MCC950, an inhibitor of NLRP3, reversed the IL-1β/IL-18 activation pattern, inhibited the formation of retinal neovascularization (RNV), decreased the number of acellular capillaries and reduced leakage of retinal vessels. Moreover, MCC950 could regulate the expression of endothelial cell- and pericyte function-associated molecules, such as vascular endothelial growth factor (VEGF), VEGF receptor (VEGFR)1, VEGFR2, matrix metalloproteinase (MMP)2, MMP9, tissue inhibitor of metalloproteinases (TIMP)1, TIMP2, platelet-derived growth factor receptor-β (PDGFR-β), platelet-derived growth factor-B (PDGF-B), and angiopoietin2 (Ang2). In vitro, recombinant human (r)IL-18 and rIL-1β regulated the expression of endothelial cell- and pericyte function-associated molecules and the proliferation and migration of endothelial cells and pericytes. We therefore determined that inhibiting the NLRP3 inflammasome with MCC950 can regulate the function of endothelial cells and pericytes by reversing the IL-1β/IL-18 activation pattern to ameliorate RNV and leakage; thereby opening new avenues to treat RNV-associated ocular diseases.

Vasculo-Neuronal Coupling and Neurovascular Coupling at the Neurovascular Unit: Impact of Hypertension.

Components of the neurovascular unit (NVU) establish dynamic crosstalk that regulates cerebral blood flow and maintain brain homeostasis. Here, we describe accumulating evidence for cellular elements of the NVU contributing to critical physiological processes such as cerebral autoregulation, neurovascular coupling, and vasculo-neuronal coupling. We discuss how alterations in the cellular mechanisms governing NVU homeostasis can lead to pathological changes in which vascular endothelial and smooth muscle cell, pericyte and astrocyte function may play a key role. Because hypertension is a modifiable risk factor for stroke and accelerated cognitive decline in aging, we focus on hypertension-associated changes on cerebral arteriole function and structure, and the molecular mechanisms through which these may contribute to cognitive decline. We gather recent emerging evidence concerning cognitive loss in hypertension and the link with vascular dementia and Alzheimer’s disease. Collectively, we summarize how vascular dysfunction, chronic hypoperfusion, oxidative stress, and inflammatory processes can uncouple communication at the NVU impairing cerebral perfusion and contributing to neurodegeneration.

Attenuation of Flightless I Increases Human Pericyte Proliferation, Migration and Angiogenic Functions and Improves Healing in Murine Diabetic Wounds.

Pericytes are peri-vascular mural cells which have an important role in the homeostatic regulation of inflammatory and angiogenic processes. Flightless I (Flii) is a cytoskeletal protein involved in regulating cellular functions, but its involvement in pericyte activities during wound healing is unknown. Exacerbated inflammation and reduced angiogenesis are hallmarks of impaired diabetic healing responses, and strategies aimed at regulating these processes are vital for improving healing outcomes. To determine the effect of altering Flii expression on pericyte function, in vitro and in vivo studies were performed to assess the effect on healing, inflammation and angiogenesis in diabetic wounds. Here, we demonstrated that human diabetic wounds display upregulated expression of the Flii protein in conjunction with a depletion in the number of platelet derived growth factor receptor β (PDGFRβ) +/ neural glial antigen 2 (NG2) + pericytes present in the dermis. Human pericytes were found to be positive for Flii and attenuating its expression in vitro through siRNA knockdown led to enhanced proliferation, migration and angiogenic functions. Genetic knockdown of Flii in a streptozotocin-induced murine model of diabetes led to increased numbers of pericytes within the wound. This was associated with dampened inflammation, an increased rate of angiogenic repair and improved wound healing. Our findings show that Flii expression directly impacts pericyte functions, including proliferation, motility and angiogenic responses. This suggests that Flii regulation of pericyte function may be in part responsible for the changes in pericyte-related processes observed in diabetic wounds.

Dynamic Measurement and Imaging of Capillaries, Arterioles, and Pericytes in Mouse Heart.

Coronary arterial tone along with the opening or closing of the capillaries largely determine the blood flow to cardiomyocytes at constant perfusion pressure. However, it is difficult to monitor the dynamic changes of the coronary arterioles and the capillaries in the whole heart, primarily due to its motion and non-stop beating. Here we describe a method that enables monitoring of arterial perfusion rate, pressure and the diameter changes of the arterioles and capillaries in mouse right ventricular papillary muscles. The mouse septal artery is cannulated and perfused at a constant flow or pressure with the other dynamically measured. After perfusion with a fluorescently labeled lectin (e.g., Alexa Fluor-488 or -633 labeled Wheat-Germ Agglutinin, WGA), the arterioles and capillaries (and other vessels) in right ventricle papillary muscle and septum could be readily imaged. The vessel-diameter changes could then be measured in the presence or absence of heart contractions. When genetically encoded fluorescent proteins were expressed, specific features could be monitored. For examples, pericytes were visualized in mouse hearts that expressed NG2-DsRed. This method has provided a useful platform to study the physiological functions of capillary pericytes in heart. It is also suitable for studying the effect of reagents on the blood flow in heart by measuring the vascular/capillary diameter and the arterial luminal pressure simultaneously. This preparation, combined with a state-of-the-art optic imaging system, allows one to study the blood flow and its control at cellular and molecular level in the heart under near-physiological conditions.

Pericyte dysfunction due to Shb gene deficiency increases B16F10 melanoma lung metastasis

Intravasation, vascular dissemination and metastasis of malignant tumor cells require their passage through the vascular wall which is commonly composed of pericytes and endothelial cells. We currently decided to investigate the relative contribution of these cell types to B16F10 melanoma metastasis in mice using an experimental model of host Shb gene (Src homology 2 domain-containing protein B) inactivation. Conditional inactivation of Shb in endothelial cells using Cdh5-CreERt2 resulted in decreased tumor growth, reduced vascular leakage, increased hypoxia and no effect on pericyte coverage and lung metastasis. RNAseq of tumor endothelial cells from these mice revealed changes in cellular components such as adherens junctions and focal adhesions by gene ontology analysis that were in line with the observed effects on leakage and junction morphology. Conditional inactivation of Shb in pericytes using Pdgfrb-CreERt2 resulted in decreased pericyte coverage of small tumor vessels with lumen, increased leakage, aberrant platelet-derived growth factor receptor B (PDGFRB) signaling and a higher frequency of lung metastasis without concomitant effects on tumor growth or oxygenation. Flow cytometry failed to reveal immune cell alterations that could explain the metastatic phenotype in this genetic model of Shb deficiency. It is concluded that proper pericyte function plays a significant role in suppressing B16F10 lung metastasis.

Phosphoinositide 3-Kinase-Regulated Pericyte Maturation Governs Vascular Remodeling.

Pericytes regulate vessel stabilization and function, and their loss is associated with diseases such as diabetic retinopathy or cancer. Despite their physiological importance, pericyte function and molecular regulation during angiogenesis remain poorly understood. To decipher the transcriptomic programs of pericytes during angiogenesis, we crossed Pdgfrb(BAC)-CreERT2 mice into RiboTagflox/flox mice. Pericyte morphological changes were assessed in mural cell-specific R26-mTmG reporter mice, in which low doses of tamoxifen allowed labeling of single-cell pericytes at high resolution. To study the role of phosphoinositide 3-kinase (PI3K) signaling in pericyte biology during angiogenesis, we used genetic mouse models that allow selective inactivation of PI3Kα and PI3Kβ isoforms and their negative regulator phosphate and tensin homolog deleted on chromosome 10 (PTEN) in mural cells. At the onset of angiogenesis, pericytes exhibit molecular traits of cell proliferation and activated PI3K signaling, whereas during vascular remodeling, pericytes upregulate genes involved in mature pericyte cell function, together with a remarkable decrease in PI3K signaling. Immature pericytes showed stellate shape and high proliferation, and mature pericytes were quiescent and elongated. Unexpectedly, we demonstrate that PI3Kβ, but not PI3Kα, regulates pericyte proliferation and maturation during vessel formation. Genetic PI3Kβ inactivation in pericytes triggered early pericyte maturation. Conversely, unleashing PI3K signaling by means of PTEN deletion delayed pericyte maturation. Pericyte maturation was necessary to undergo vessel remodeling during angiogenesis. Our results identify new molecular and morphological traits associated with pericyte maturation and uncover PI3Kβ activity as a checkpoint to ensure appropriate vessel formation. In turn, our results may open new therapeutic opportunities to regulate angiogenesis in pathological processes through the manipulation of pericyte PI3Kβ activity.

Culture of Brain Capillary Pericytes for Cytosolic Calcium Measurements and Calcium Imaging Studies.

Pericytes are associated with endothelial cells and astrocytic endfeet in a structure known as the neurovascular unit (NVU). Brain capillary pericyte function is not fully known. Pericytes have been suggested to be involved in capillary development, regulation of endothelial barrier tightness and trancytosis activity, regulation of capillary tone and to play crucial roles in certain brain pathologies. Pericytes are challenging to investigate in the intact brain due to the difficulties in visualizing processes in the brain parenchyma, as well as the close proximity to the other cells of the NVU. The present protocol describes a method for isolation and culture of primary bovine brain capillary pericytes and their following usage in calcium imaging studies, where effects of agonists involved in brain signaling and pathologies can be investigated. Cortical capillary fragments are allowed to attach to the bottom of culture flasks and, after 6 days, endothelial cells and pericytes have grown out from the capillary fragments. The endothelial cells are removed by gentle trypsinization and pericytes are cultured for 5 additional days before passaging. Isolated pericytes are seeded in 96-well culture plates and loaded with the calcium indicator dye (Fura-2 acetoxymethyl (AM)) to allow for measurements of intracellular calcium levels in a plate reader setup. Alternatively, pericytes are seeded on coverslips and mounted in cell chambers. Following loading with the calcium indicator (Cal-520 AM), calcium live-imaging can be performed using confocal microscopy at an excitation wavelength of 488 nm and emission wavelength of 510-520 nm. The method described here has been used to obtain the first intracellular calcium measurements from primary brain capillary pericytes, demonstrating that pericytes are stimulated via ATP and are able to contract in vitro.

Interactions between Amyloid-Β Proteins and Human Brain Pericytes: Implications for the Pathobiology of Alzheimer's Disease.

Alzheimer’s disease (AD) is a progressive neurodegenerative disease that is the most common cause of dementia, especially among aging populations. Despite advances in AD research, the underlying cause and the discovery of disease-modifying treatments have remained elusive. Two key features of AD pathology are the aberrant deposition of amyloid beta (amyloid-β or Aβ) proteins in the brain parenchyma and Aβ toxicity in brain pericytes of the neurovascular unit/blood–brain barrier (NVU/BBB). This toxicity induces oxidative stress in pericytes and leads to capillary constriction. The interaction between pericytes and Aβ proteins results in the release of endothelin-1 in the pericytes. Endothelin-1 interacts with ETA receptors to cause pericyte contraction. This pericyte-mediated constriction of brain capillaries can cause chronic hypoperfusion of the brain microvasculature, subsequently leading to the neurodegeneration and cognitive decline observed in AD patients. The interaction between Aβ proteins and brain pericytes is largely unknown and requires further investigation. This review provides an updated overview of the interaction between Aβ proteins with pericytes, one the most significant and often forgotten cellular components of the BBB and the inner blood–retinal barrier (IBRB). The IBRB has been shown to be a window into the central nervous system (CNS) that could allow the early diagnosis of AD pathology in the brain and the BBB using modern photonic imaging systems such as optical coherence tomography (OCT) and two-photon microscopy. In this review, I explore the regulation of Aβ proteins in the brain parenchyma, their role in AD pathobiology, and their association with pericyte function. This review discusses Aβ proteins and pericytes in the ocular compartment of AD patients as well as strategies to rescue or protect pericytes from the effects of Aβ proteins, or to replace them with healthy cells.

Effects of Gold Nanoparticles on Capillary Tube Formation with Special Reference to the Alteration of Pericyte Morphology and Function

Pericytes are perivascular cells that wrap around endothelial cells to form wall of capillaries and post‐capillary venules. These cells are primary importance in physiological and pathological processes, especially, involved in angiogenesis. At present, gold nanoparticles (AuNPs) were used for anti‐angiogenesis by inhibiting the activity of endothelial cell proliferation, migration, and tube formation. However, the effects of gold nanoparticles on the alteration of pericyte morphology and theirs function were not reported. Therefore, in this study, 20 nm‐AuNPs were applied to human placenta pericyte monoculture with concentration of 30 ppm. Morphology of pericytes was observed by transmission electron microscopy and cell migration was evaluated by transwell migration assay. Real‐time qRT‐PCR was used to determine Ki‐67, as a proliferation marker, and PDGFR‐β mRNA expression. Moreover, these AuNPs 30 ppm‐treated pericytes were co‐cultured with human umbilical cord endothelial cells on Matrigel® to investigate the 3D tube formation. The results revealed that AuNPs significantly suppressed migration, proliferation, and PDGFR‐β mRNA expression of pericytes. The ultrastructure of pericytes showed AuNPs in several organelles, i.e., swollen or damaged mitochondria, engulfed late endosomes and lysosomes. Moreover, the number of capillary tube formations was essentially reduced and we found that a number of pericytes on these capillary tubes were round shape and did not extend the processes along the tube, which in turn increased incomplete tube formation compared with the control. In conclusion, AuNPs could effect on pericyte biology including proliferation, migration, morphology, and tube formation property. Moreover, the results highlighted the potential application of AuNPs in anti‐angiogenesis by pericyte‐targeted therapy.Support or Funding InformationThe Ratchadaphiseksompotch Fund, Faculty of Medicine, Chulalongkorn University, grant number RA61/100 and Overseas Academic Presentation Scholarship for Graduate Students from the Graduate School, Chulalongkorn University

Metabolic Coordination of Pericyte Phenotypes: Therapeutic Implications.

Pericytes are mural vascular cells found predominantly on the abluminal wall of capillaries, where they contribute to the maintenance of capillary structural integrity and vascular permeability. Generally quiescent cells in the adult, pericyte activation and proliferation occur during both physiological and pathological vascular and tissue remodeling. A considerable body of research indicates that pericytes possess attributes of a multipotent adult stem cell, as they are capable of self-renewal as well as commitment and differentiation into multiple lineages. However, pericytes also display phenotypic heterogeneity and recent studies indicate that lineage potential differs between pericyte subpopulations. While numerous microenvironmental cues and cell signaling pathways are known to regulate pericyte functions, the roles that metabolic pathways play in pericyte quiescence, self-renewal or differentiation have been given limited consideration to date. This review will summarize existing data regarding pericyte metabolism and will discuss the coupling of signal pathways to shifts in metabolic pathway preferences that ultimately regulate pericyte quiescence, self-renewal and trans-differentiation. The association between dysregulated metabolic processes and development of pericyte pathologies will be highlighted. Despite ongoing debate regarding pericyte classification and their functional capacity for trans-differentiation in vivo, pericytes are increasingly exploited as a cell therapy tool to promote tissue healing and regeneration. Ultimately, the efficacy of therapeutic approaches hinges on the capacity to effectively control/optimize the fate of the implanted pericytes. Thus, we will identify knowledge gaps that need to be addressed to more effectively harness the opportunity for therapeutic manipulation of pericytes to control pathological outcomes in tissue remodeling.

Abstract 39: GLP-1 Agonists Improve Cerebrovascular Integrity and Vascular Induced Cognitive Impairment and Dementia Beyond Glycemic Control via Restoration of Brain Pericytes Functions in Diabetic Mice

We have previously shown that diabetes causes pericyte-dysfunction that leads to loss of vascular integrity and vascular-induced cognitive impairment and dementia (VCID). Glucagon-like peptide-1(GLP-1), used in the management of type-2 diabetes mellitus, improves cognitive functions in diabetic patients beyond glycemic control, yet the mechanism is unknown. In the present study, we hypothesis that GLP-1 agonist improves VICD through prevention of diabetes-induced pericytes dysfunction in a non-glucose dependent way. Methods: Control and diabetic mice were randomly assigned for saline or Exendin-4 (GLP-1 agonist 30 ng/kg/day), delivered through osmotic pump over 28 days. Vascular integrity was assessed by measuring cerebrovascular neovascularization indices (Vascular density, tortuosity, and branching density). Cognitive functions were evaluated with Barnes maze and Morrison Water maze. Human brain microvascular pericytes, HBMPCs, were grown in high glucose 25 mM/ sodium palmitate 200 uM (HG/Pal) to mimic diabetic conditions. HBMPCs were treated with/out Exendin-4 and assessed for oxidative stress and angiogenic properties. Results: Diabetic mice treated with GLP-1 agonist showed a significant reduction in all cerebral pathological neovascularization indices (P<0.05). Exendin-4 vascular protective effects were accompanied by significant improvement of the learning and memory functions of diabetic mice (P<0.05). Our results showed that HBMPCs expressed the GLP-1 receptor. Treating HBMPCs exposed to diabetic conditions with GLP-1 agonist restored pericyte functions, decreased diabetes-induced inflammation, oxidative stress, and migration (P<0.05). Conclusion: Our results provide novel evidence that GLP-1 agonist produces neurovascular protective effects in part through targeting pericytes. Restoration of pericyte functions in diabetes represents a novel therapeutic target for diabetes-induced vascular remodeling and VCID.

Abstract WP134: 3K3A-APC Protects Pericyte-deficient Mice From Ischemic Brain Injury

Pericytes play a key role in maintaining the blood-brain barrier (BBB) integrity. BBB disruption occurs during early stages after ischemic stroke. However, the role of pericytes in the pathogenesis of ischemic stroke remains still understudied. 3K3A-APC, a recombinant variant of activated protein C, has shown benefits in preclinical models of ischemic stroke and has favorable safety profile and reduces hemorrhage in Phase 2 study in ischemic stroke patients (RHAPSODY). In the present study, we used PDGFRβ heterozygous knockout (PDGFRβ+/-) mice to investigate the effects of pericyte deficiency on ischemic brain injury using transient proximal middle cerebral artery occlusion (tMCAO). Additionally, we investigated the effects of 3K3A-APC therapy (0.2mg/kg i.v. 4h after stroke) in this model. Compared to controls, pericyte deficiency in PDGFRβ+/- mice resulted in ~35% increase in the infarct and edema volumes, reduction in pericyte coverage from 58% to 25%, and increased IgG and fibrin deposition suggesting accelerated BBB breakdown 24h after stroke. Additionally, PDGFRβ+/- mice showed by 36% more degenerating Fluoro-Jade+ neurons and exhibited accelerated neurobehavioral abnormalities. 3K3A-APC improved neuropathological changes and functional deficits. Our results suggest that pericyte deficiency worsens brain damage and functional outcome after ischemic stroke in mice suggesting that pericytes may play an important role in protecting brain from post-ischemic. We also suggests that 3K3A-APC protects pericyte function in stroked mice which could contribute to its overall neuroprotective effects.

Effects of complement dysfunction on diabetic retinopathy

Diabetic retinopathy (DR) is one of the common neuronal and microvascular complications of diabetes, and it is the key cause of blindness in the populations aged from 20 to 74.The pathogenesis of DR remains elusive, and the role of complement in DR has attracted intensive attentions in recent years.The dysregulation of complement factors and regulatory proteins could affect the fate and functions of pericytes, endothelial cells, neuronal cells and glia cells in retina, which may be novel targets for anti-DR therapy.Advances regarding the characteristics of complement in the eye, the relationship between complement dysfunction and DR, complement gene polymorphism in DR and recent therapy explorations targeting complement pathways were reviewed, so as to providing new clues for the related research in the future. Key words: Diabetic Retinopathy; Inflammation; Complement

Evaluation of serum platelet-derived growth factor receptor-ß and brain-derived neurotrophic factor levels in microvascular angina.

Objective:Microvascular angina (MVA) is a coronary microcirculation disease. Research on microcirculatory dysfunction has revealed several biomarkers involved in the etiopathogenesis of MVA. Platelet-derived growth factor receptor β (PDGFR-β) and brain-derived neurotrophic factor (BDNF) are 2 biomarkers associated with microcirculation, particularly pericytes function. The aim of this study was to investigate the role of PDGFR-β and BDNF in MVA.Methods:Ninety-one patients (median age, 56 y; age range, 40–79 y; 36 men) with MVA and 61 control group subjects (median age, 52 y; age range, 38–76 y; 29 men) were included in the study. Serum concentrations of PDGFR-β and BDNF were measured with commercially available enzyme-linked immunosorbent assay kits.Results:PDGFR-β [2.82 ng/ml; interquartile range (IQR), 0.57–7.79 ng/ml vs. 2.27 ng/ml; IQR, 0.41–7.16 ng/ml; p<0.0005] and BDNF (2.41 ng/ml; IQR, 0.97–7.97 ng/ml vs. 1.92 ng/ml; IQR, 1.07–6.67 ng/ml; p=0.023) concentrations were significantly higher in patients with MVA compared with the controls. PDGFR-β correlated positively with age (r=0.26, p=0.001), low-density lipoprotein (r=0.18; p=0.02), and BDNF (r=0.47; p<0.001), and BDNF showed a significant positive correlation with age (r=0.20; p=0.01). In binary logistic regression analysis, high-sensitivity C-reactive protein, uric acid, and PDGFR-β values were found to be independent predictors of MVA.Conclusion:MVA is associated with higher PDGFR-β and BDNF levels. This association may indicate an abnormality in microvascular function. Future studies are required to determine the role of these biomarkers in the pathogenesis of MVA. (Anatol J Cardiol 2020; 24: 397-404)

Fibroblast growth factor-2/platelet-derived growth factor enhances atherosclerotic plaque stability.

Increased immature neovessels contribute to plaque growth and instability. Here, we investigated a method to establish functional and stable neovessel networks to increase plaque stability. Rabbits underwent aortic balloon injury and were divided into six groups: sham, vector and lentiviral transfection with vascular endothelial growth factor‐A (VEGF)‐A, fibroblast growth factor (FGF)‐2, platelet‐derived growth factor (PDGF)‐BB and FGF‐2 + PDGF‐BB. Lentivirus was percutaneously injected into the media‐adventitia of the abdominal aorta by intravascular ultrasound guidance, and plaque‐rupture rate, plaque‐vulnerability index and plaque neovessel density at the injection site were evaluated. Confocal microscopy, Prussian Blue assay, Evans Blue, immunofluorescence and transmission electron microscopy were used to assess neovessel function and pericyte coverage. To evaluate the effect of FGF‐2/PDGF‐BB on pericyte migration, we used the mesenchymal progenitor cell line 10T1/2 as an in vitro model. VEGF‐A‐ and FGF‐2‐overexpression increased the number of immature neovessels, which caused intraplaque haemorrhage and inflammatory cell infiltration, eventually resulting in the plaque vulnerability; however, FGF‐2/PDGF‐BB induced mature and functional neovessels, through increased neovessel pericyte coverage. Additionally, in vitro analysis of 10T1/2 cells revealed that FGF‐2/PDGF‐BB induced epsin‐2 expression and enhanced the VEGF receptor‐2 degradation, which negatively regulated pericyte function consistent with the in vivo data. These results showed that the combination of FGF‐2 and PDGF‐BB promoted the function and maturation of plaque neovessels, thereby representing a novel potential treatment strategy for vulnerable plaques.

Glioblastoma ablates pericytes antitumor immune function through aberrant up-regulation of chaperone-mediated autophagy.

The contractile perivascular cells, pericytes (PC), are hijacked by glioblastoma (GB) to facilitate tumor progression. PC's protumorigenic function requires direct interaction with tumor cells and contributes to the establishment of immunotolerance to tumor growth. Cancer cells up-regulate their own chaperone-mediated autophagy (CMA), a process that delivers selective cytosolic proteins to lysosomes for degradation, with pro-oncogenic effects. However, the possible impact that cancer cells may have on CMA of surrounding host cells has not been explored. We analyzed the contribution of CMA to the GB-induced changes in PC biology. We have found that CMA is markedly up-regulated in PC in response to the oxidative burst that follows PC-GB cell interaction. Genetic manipulations to block the GB-induced up-regulation of CMA in PC allows them to maintain their proinflammatory function and to support the induction of effective antitumor T cell responses required for GB clearance. GB-induced up-regulation of CMA activity in PC is essential for their effective interaction with GB cells that help tumor growth. We show that CMA inhibition in PC promotes GB cell death and the release of high immunogenic levels of granulocyte-macrophage colony stimulating factor (GM-CSF), through deregulation of the expression of cell-to-cell interaction proteins and protein secretion. A GB mouse model grafted in vivo with CMA-defective PC shows reduced GB proliferation and effective immune response compared to mice grafted with control PC. Our findings identify abnormal up-regulation of CMA as a mechanism by which GB cells elicit the immunosuppressive function of PC and stabilize GB-PC interactions necessary for tumor cell survival.

Activation of Sigma-1 Receptor Enhanced Pericyte Survival via the Interplay Between Apoptosis and Autophagy: Implications for Blood-Brain Barrier Integrity in Stroke.

Stroke is a cerebrovascular disorder that affects many people worldwide. Pericytes play an important role in stroke progression and recovery. The sigma-1 receptor (σ-1R) signaling pathway has been suggested as having promising neuroprotective potential in treating stroke; however, whether σ-1R activation regulates pericyte function remains unknown. The aim of this study was to elucidate the role of σ-1R and a novel σ-1R agonist in pericytes following ischemic stroke. An ischemic stroke animal model was induced by photothrombotic middle cerebral artery occlusion (pMCAO) in σ-1R knockout (KO) and wild-type (WT) mice. After pMCAO, there was significant pericyte loss and coverage in σ-1R KO mice compared with WT mice as determined using transmission electron microscopy, immunofluorescence staining, and western blot. Interestingly, a novel σ-1R agonist decreased infarct volume and blood-brain barrier damage with a concomitant amelioration of pericyte loss, as determined by western blot. Further studies indicated that cell apoptosis and autophagy were induced in an in vivo pMCAO ischemic stroke animal model and an in vitro oxygen glucose deprivation-treatment group. Inhibition of autophagy using a pharmacological approach significantly mitigated pericyte apoptosis, suggesting that autophagy was upstream of apoptosis in pericytes. Both in vivo and in vitro studies indicated that the σ-1R agonist significantly decreased cell apoptosis via inhibition of autophagy with a subsequent enhancement of pericyte survival. This study identified the unique roles for σ-1R in mediating pericyte survival via the regulation of the interplay between apoptosis and autophagy, suggesting that a novel σ-1R agonist may be a promising therapeutic agent for the treatment of stroke patients.

Pericytes and Neurovascular Function in the Healthy and Diseased Brain.

Pericytes are multi-functional cells embedded within the walls of capillaries throughout the body, including the brain. Pericytes were first identified in the 1870s, but little attention was paid to them during the following century. More recently, numerous vascular functions of pericytes have been identified including regulation of cerebral blood flow, maintenance of the blood-brain barrier (BBB), and control of vascular development and angiogenesis. Pericytes can also facilitate neuroinflammatory processes and possess stem cell-like properties. Pericytes form part of the neurovascular unit (NVU), a collection of cells that control interactions between neurons and the cerebral vasculature to meet the energy demands of the brain. Pericyte structure, expression profile, and function in the brain differ depending on their location along the vascular bed. Until recently, it has been difficult to accurately define the sub-types of pericytes, or to specifically target pericytes with pharmaceutical agents, but emerging techniques both in vitro and in vivo will improve investigation of pericytes and allow for the identification of their possible roles in diseases. Pericyte dysfunction is increasingly recognized as a contributor to the progression of vascular diseases such as stroke and neurodegenerative diseases such as Alzheimer’s disease. The therapeutic potential of pericytes to repair cerebral blood vessels and promote angiogenesis due to their ability to behave like stem cells has recently been brought to light. Here, we review the history of pericyte research, the present techniques used to study pericytes in the brain, and current research advancements to characterize and therapeutically target pericytes in the future.