Pericyte Loss Research Articles

Morphine-mediated release of astrocyte-derived extracellular vesicle miR-23a induces loss of pericyte coverage at the blood-brain barrier: Implications for neuroinflammation.

Opioids such as morphine are the most potent and efficacious drugs currently available for pain management. Paradoxically, opioids have also been implicated in inducing neuroinflammation and associated neurocognitive decline. Pericytes, a critical component of the neurovascular unit (NVU), are centrally positioned between endothelial cells and astrocytes, maintaining function of the blood-brain barrier (BBB) nd regulating neuroinflammation by controlling monocyte influx under various pathological conditions. The role of pericytes in morphine-mediated neuroinflammation however, has received less attention, especially in the context of how pericytes crosstalk with other central nervous system (CNS) cells. The current study was undertaken to examine the effect of miRNAs released from morphine-stimulated human primary astrocyte-derived extracellular vesicles (morphine-ADEVs) in mediating pericyte loss at the blood-brain barrier, leading, in turn, to increased influx of peripheral monocytes. Our findings suggest that the heterogeneous nuclear ribonucleoprotein complex A2/B1 (hnRNP A2/B1) plays role in morphine-mediated upregulation and release of miR-23a in ADEVs, and through action of morphine via mu opioid receptor.We further demonstrated that miR-23a in morphine-ADEVs could be taken up by pericytes, resulting in downregulation of PTEN expression, ultimately leading to increased pericyte migration. Furthermore, both overexpression of PTEN and blocking the miR-23a target site at PTEN 3UTR (by transfecting miR-23a-PTEN target protector), attenuated morphine-ADEV-mediated pericyte migration. We also demonstrated that in the microvessels isolated from morphine-administered mice, there were fewer PDGFβR + pericytes co-localizing with CD31+ brain endothelial cells compared with those from saline mice. In line with these findings, we also observed increased loss of pericytes and a concomitantly increased influx of monocytes in the brains of morphine-administered pericyte-labeled NG2-DsRed mice compared with saline mice. In conclusion, our findings indicate morphine-ADEVs mediated loss of pericyte coverage at the brain endothelium, thereby increasing the influx of peripheral monocytes in the central nervous system, leading to neuroinflammation.

Mito-TEMPO, a Mitochondria-Targeted Antioxidant, Improves Cognitive Dysfunction due to Hypoglycemia: an Association with Reduced Pericyte Loss and Blood-Brain Barrier Leakage.

Hypoglycemia is associated with cognitive dysfunction, but the exact mechanisms have not been elucidated. Our previous study found that severe hypoglycemia could lead to cognitive dysfunction in a type 1 diabetes (T1D) mouse model. Thus, the aim of this study was to further investigate whether the mechanism of severe hypoglycemia leading to cognitive dysfunction is related to oxidative stress-mediated pericyte loss and blood-brain barrier (BBB) leakage. A streptozotocin T1D model (150 mg/kg, one-time intraperitoneal injection), using male C57BL/6J mice, was used to induce hypoglycemia. Brain tissue was extracted to examine for neuronal damage, permeability of BBB was investigated through Evans blue staining and electron microscopy, reactive oxygen species and adenosine triphosphate in brain tissue were assayed, and the functional changes of pericytes were determined. Cognitive function was tested using Morris water maze. Also, an in vitro glucose deprivation model was constructed. The results showed that BBB leakage after hypoglycemia is associated with excessive activation of oxidative stress and mitochondrial dysfunction due to glucose deprivation/reperfusion. Interventions using the mitochondria-targeted antioxidant Mito-TEMPO in both in vivo and in vitro models reduced mitochondrial oxidative stress, decreased pericyte loss and apoptosis, and attenuated BBB leakage and neuronal damage, ultimately leading to improved cognitive function.

CNS Pericytes Modulate Local T Cell Infiltration in EAE.

Pericytes at the blood-brain barrier (BBB) are located between the tight endothelial cell layer of the blood vessels and astrocytic endfeet. They contribute to central nervous system (CNS) homeostasis by regulating BBB development and maintenance. Loss of pericytes results in increased numbers of infiltrating immune cells in the CNS in experimental autoimmune encephalomyelitis (EAE), the mouse model for multiple sclerosis (MS). However, little is known about their competence to modulate immune cell activation or function in CNS autoimmunity. To evaluate the capacity of pericytes to directly interact with T cells in an antigen-specific fashion and potentially (re)shape their function, we depleted major histocompatibility complex (MHC) class II from pericytes in a cell type-specific fashion and performed T cell-pericyte cocultures and EAE experiments. We found that pericytes present antigen in vitro to induce T cell activation and proliferation. In an adoptive transfer EAE experiment, pericyte-specific MHC II KO resulted in locally enhanced T cell infiltration in the CNS; even though, overall disease course of mice was not affected. Thus, pericytes may serve as non-professional antigen-presenting cells affecting states of T cell activation, thereby locally shaping lesion formation in CNS inflammation but without modulating disease severity.

SARS-CoV-2 infection causes prolonged cardiomyocyte swelling and inhibition of HIF1α translocation in an animal model COVID-19.

Recovered COVID-19 patients often display cardiac dysfunction, even after a mild infection. Most current histological results come from patients that are hospitalized and therefore represent more severe outcomes than most COVID-19 patients face. To overcome this limitation, we investigated the cardiac effects of SARS-CoV-2 infection in a hamster model. SARS-CoV-2 infected hamsters developed diastolic dysfunction after recovering from COVID-19. Histologically, increased cardiomyocyte size was present at the peak of viral load and remained at all time points investigated. As this increase is too rapid for hypertrophic remodeling, we found instead that the heart was oedemic. Moreover, cardiomyocyte swelling is associated with the presence of ischemia. Fibrin-rich microthrombi and pericyte loss were observed at the peak of viral load, resulting in increased HIF1α in cardiomyocytes. Surprisingly, SARS-CoV-2 infection inhibited the translocation of HIF1α to the nucleus both in hamster hearts, in cultured cardiomyocytes, as well as in an epithelial cell line. We propose that the observed diastolic dysfunction is the consequence of cardiac oedema, downstream of microvascular cardiac ischemia. Additionally, our data suggest that inhibition of HIF1α translocation could contribute to an exaggerated response upon SARS-CoV-2 infection.

Intravitreally Administered Soluble VEGF Receptor-1 Variant Tested as a Potential Gene Therapeutic for Diabetic Retinopathy

In addition to laser photocoagulation, currently used therapeutic interventions for diabetic retinopathy (DR) include relatively short-lived anti-VEGF drugs targeting vascular endothelial growth factor (VEGF). The latter requires frequent administration via intravitreal injections to effect long-term VEGF suppression. However, due to the patient burden associated with this treatment modality, gene therapy may represent a preferable alternative, providing long-lasting yet patient-friendly effects. Here, we explore the therapeutic efficacy of rAAV2-sVEGFRv-1, a recombinant adeno-associated virus encoding a soluble variant of VEGF receptor-1, upon early DR processes. Bevacizumab, an anti-VEGF agent often prescribed off label to treat DR, was used as an experimental comparator. Administered by intravitreal injection to a streptozotocin-induced diabetic mouse model, rAAV2-sVEGFRv-1 was shown to effectively transduce the mouse retinas and express its transgene therein, leading to significant reductions in pericyte loss and retinal cell layer thinning, two processes that play major roles in DR progression. Acellular capillary formation, vascular permeability, and apoptotic activity, the latter being the cell death mechanism by which retinal neurodegeneration occurs, were also shown to be reduced by the therapeutic virus vector. Immunohistochemistry was used to visualize that rAAV2-sVEGFRv-1 has an effect on cell types important to DR pathophysiology, particularly the ganglion cell layer and glial cells. Combined with our previous work showing that the therapeutic virus vector reduces neovascularization, our current results reveal that rAAV2-sVEGFRv-1 addresses the early aspects of DR as well, thereby demonstrating its potential as a human gene therapeutic versus the condition as a whole.

Pericyte remodeling is deficient in the aged brain and contributes to impaired capillary flow and structure

Deterioration of brain capillary flow and architecture is a hallmark of aging and dementia. It remains unclear how loss of brain pericytes in these conditions contributes to capillary dysfunction. Here, we conduct cause-and-effect studies by optically ablating pericytes in adult and aged mice in vivo. Focal pericyte loss induces capillary dilation without blood-brain barrier disruption. These abnormal dilations are exacerbated in the aged brain, and result in increased flow heterogeneity in capillary networks. A subset of affected capillaries experience reduced perfusion due to flow steal. Some capillaries stall in flow and regress, leading to loss of capillary connectivity. Remodeling of neighboring pericytes restores endothelial coverage and vascular tone within days. Pericyte remodeling is slower in the aged brain, resulting in regions of persistent capillary dilation. These findings link pericyte loss to disruption of capillary flow and structure. They also identify pericyte remodeling as a therapeutic target to preserve capillary flow dynamics.

Markers of Proliferative Diabetic Retinopathy

The incidence of diabetes mellitus (DM) is steadily growing and today the World Health Organization (WHO) describes this situation as an epidemic. Diabetic retinopathy (DR) is one of the most common complications of DM, as well as one of the main causes of acquired irreversible vision loss. Nowadays the pathogenesis of proliferative DR remains completely unexplored, but many authors think about the important role of biologically significant mediators of this process cytokines and growth factors. Cytokines and growth factors are protein mediators that regulate various functions, both locally and systemically. They carry out the cells’ life cycle, the processes of proliferation, participate in the regulation of the protective inflammatory response of the body, control angiogenesis and other mechanisms. It is proved that the main links in the pathogenesis of DR are oxidative stress, thickening of the endothelial basement membrane in capillaries, loss of pericytes, end products of glycation and inflammation. Chorioretinal hypoxia and ischemia play a major role in the formation of newly formed vessels. Newly formed vessels are defective (with a thin wall devoid of pericytes), often lead to hemorrhages, hypoxia, which in turn closes the pathological circle and causes the production of cytokines and vasoproliferative factors. Frequent complications of DR are intraocular hemorrhages, retinal fibrosis and pathological changes in the posterior hyaloid membrane, traction retinal detachment, etc. This review examines some types of cytokines and growth factors and their role in the light in the pathogenesis of proliferative DR. Modern technologies make it possible to conduct effective studies of intraocular fluids to study the content of biologically active substances both in the moisture of the anterior chamber of the eye and in the vitreous body. To narrow the scope of the review on the subject attention is focused on the works that examined various markers locally in the intraocular fluids in patients with DM. It is worth noting that there are few such studies and their results often differ significantly from each other. This fact is a subject for discussion and encourages further study of this topic.

Intraocular Pressure-Induced Endothelial Dysfunction of Retinal Blood Vessels Is Persistent, but Does Not Trigger Retinal Ganglion Cell Loss.

Research has been conducted into vascular abnormalities in the pathogenesis of glaucoma, but conclusions remain controversial. Our aim was to test the hypothesis that retinal endothelial dysfunction induced by elevated intraocular pressure (IOP) persists after IOP normalization, further triggering retinal ganglion cell (RGC) loss. High intraocular pressure (HP) was induced in mice by episcleral vein occlusion (EVO). Retinal vascular function was measured via video microscopy in vitro. The IOP, RGC and their axons survival, levels of oxidative stress and inflammation as well as vascular pericytes coverage, were determined. EVO caused HP for two weeks, which returned to baseline afterwards. Mice with HP exhibited endothelial dysfunction in retinal arterioles, reduced density of RGC and their axons, and loss of pericytes in retinal arterioles. Notably, these values were similar to those of mice with recovered IOP (RP). Levels of oxidative stress and inflammation were increased in HP mice but went back to normal in the RP mice. Our data demonstrate that HP induces persistent endothelial dysfunction in retinal arterioles, which persists one month after RP. Oxidative stress, inflammation, and loss of pericytes appear to be involved in triggering vascular functional deficits. Our data also suggest that retinal endothelial dysfunction does not affect RGC and their axon survival.

Topical nerve growth factor prevents neurodegenerative and vascular stages of diabetic retinopathy.

Specific and effective preventive treatment for diabetic retinopathy (DR) is presently unavailable, mostly because the early stages of the complication have been, until recently, poorly understood. The recent demonstration that the vascular phase of DR is preceded and possibly caused by the neurodegeneration of retinal ganglion cells suggests that DR could, at least theoretically, be prevented through an early neuroprotective approach. The aims of our study were to clarify the natural history of diabetes-driven retinal neurodegeneration and to verify the possibility to prevent DR using topical nerve growth factor (NGF). The results of the study show that retinal neurodegeneration, characterized by the loss of retinal ganglion cells represents a relatively early phenomenon of diabetes (between 5 and 16 weeks of age), which tends to be self-limiting in the long run. Neurodegeneration is followed by the development of DR-related vascular dysfunctions, as confirmed by the development of acellular capillaries and the loss of retinal pericytes. Both retinal neurodegeneration and subsequent vascular dysfunction can be successfully prevented by topical NGF administration. These findings suggest that: 1) The first stage of DR consists in a self-limiting retinal neurodegeneration 2) The demonstrated effectiveness of topical NGF in the prevention of DR could be rapidly translated into clinical practice.

Leptin Promotes Angiogenesis via Pericyte STAT3 Pathway upon Intracerebral Hemorrhage.

Angiogenesis is a vital endogenous brain self-repair processes for neurological recovery after intracerebral hemorrhage (ICH). Increasing evidence suggests that leptin potentiates angiogenesis and plays a beneficial role in stroke. However, the proangiogenic effect of leptin on ICH has not been adequately explored. Moreover, leptin triggers post-ICH angiogenesis through pericyte, an important component of forming new blood vessels, which remains unclear. Here, we reported that exogenous leptin infusion dose-dependent promoted vascular endothelial cells survival and proliferation at chronic stage of ICH mice. Additionally, leptin robustly ameliorated pericytes loss, enhanced pericytes proliferation and migration in ICH mice in vivo, and in ICH human brain microvascular pericytes (HBVPC) in vitro. Notably, we showed that pericytes-derived pro-angiogenic factors were responsible for enhancing the survival, proliferation and tube formation followed leptin treatment in human brain microvascular endothelial cells (HCMEC/D3)/HBVPC co-culture models. Importantly, considerable improvements in neurobehavioral function and hostile microenvironment were observed in leptin treatment ICH mice, indicating that better vascular functionality post ICH improves outcome. Mechanistically, this study unveiled that leptin boost post-ICH angiogenesis potentially through modulation of leptin receptor (leptinR)/Signal Transducer and Activator of Transcription 3 (STAT3) signaling pathway in pericyte. Thus, leptin may be a lucrative option for the treatment of ICH.

A "multi-omics" analysis of blood-brain barrier and synaptic dysfunction in APOE4 mice.

Apolipoprotein E4 (APOE4), the main susceptibility gene for Alzheimer's disease, leads to blood-brain barrier (BBB) breakdown in humans and mice. Remarkably, BBB dysfunction predicts cognitive decline and precedes synaptic deficits in APOE4 human carriers. How APOE4 affects BBB and synaptic function at a molecular level, however, remains elusive. Using single-nucleus RNA-sequencing and phosphoproteome and proteome analysis, we show that APOE4 compared with APOE3 leads to an early disruption of the BBB transcriptome in 2-3-mo-old APOE4 knock-in mice, followed by dysregulation in protein signaling networks controlling cell junctions, cytoskeleton, clathrin-mediated transport, and translation in brain endothelium, as well as transcription and RNA splicing suggestive of DNA damage in pericytes. Changes in BBB signaling mechanisms paralleled an early, progressive BBB breakdown and loss of pericytes, which preceded postsynaptic interactome disruption and behavioral deficits that developed 2-5 mo later. Thus, dysregulated signaling mechanisms in endothelium and pericytes in APOE4 mice reflect a molecular signature of a progressive BBB failure preceding changes in synaptic function and behavior.

Calpain Inhibitor Calpeptin Improves Alzheimer's Disease-Like Cognitive Impairments and Pathologies in a Diabetes Mellitus Rat Model.

Diabetes mellitus (DM) has been considered an accelerator of Alzheimer's disease (AD), but the cellular and molecular mechanisms underlying this effect are not fully understood. Here, we attempted to determine the role and regulatory mechanism of calpain in the AD-like cognitive decline and pathological changes in rats caused by DM. In the initial stages, our results verified that DM model rats showed cognitive impairment, as well as a loss of neurons, decreased pericyte marker (PDGFR-β and α-SMA), and calpain-2 expression and amyloid-β (Aβ) deposition in the hippocampal tissues. In high glucose-induced primary pericytes, the cell apoptotic rate was increased, and cell proliferation was inhibited in a time-dependent manner. The protein level of calpain-2 was also upregulated by HG induction, but the level of calpain-1 did not change with HG treatment, which was also observed in DM model rats. Subsequently, some DM model rats were administered calpeptin, an inhibitor of calpain. Our data revealed that calpeptin treatment significantly suppressed calpain-1 and calpain-2 expression in the hippocampal tissues and effectively improved the cognitive impairments of DM model rats. Neuronal loss, Aβ accumulation, pericyte loss, inflammation, and oxidative stress injury in the hippocampal tissues of DM model rats were also partly rescued by calpeptin administration. Our work demonstrated that the calpain inhibitor calpeptin could alleviate DM-induced AD-like cognitive impairments and pathological changes in rats, and this effect may be associated with pericytes. Calpeptin may become a promising drug to treat the AD-like complications of DM.

The SARS-CoV-2 envelope protein disrupts barrier function in an in vitro human blood-brain barrier model.

Patients with coronavirus disease 2019 (COVID-19) have been frequently reported to exhibit neurological manifestations and disruption of the blood-brain barrier (BBB). Among the risk factors for BBB breakdown, the loss of endothelial cells and pericytes has caused widespread concern. Recent studies have revealed that severe acute respiratory syndrome coronavirus 2 envelope (S2E) protein caused cell death. We tested the hypothesis that the S2E protein alone could induce BBB dysfunction. The S2E protein bound to human BBB-related cells and inhibited cell viability in a dose- and time-dependent manner. Importantly, the S2E protein disrupted barrier function in an in vitro BBB model composed of HCMEC/D3 (brain endothelial cell line), HBVP (brain vascular pericyte), and U87MG (astrocyte cell line) cells and suppressed the expression of major genes involved in maintaining endothelial permeability and function. In addition, the S2E protein crossed the HCMEC/D3 monolayer. The S2E protein triggered inflammatory responses in HCMEC/D3 and U87MG cells. Taken together, these results show for the first time that the S2E protein has a negative impact on the BBB. Therapies targeting the S2E protein could protect against and treat central nervous system manifestations in COVID-19 patients.

Gene therapy targeting inflammatory pericytes corrects angiopathy during diabetic wound healing.

Wound healing is impaired in the diabetic status, largely attributable to diabetes-associated angiopathy. Pericytes play critical roles in the stabilization of the formed vessels. The loss and dysfunction of pericytes have been reported in inflammation during diabetes and associated with the pathology of diabetic angiopathy. However, a practical approach that targets inflammatory pericytes to improve diabetic wound healing is lacking. In the current study, we showed that the inflammatory pericytes from wound skin of diabetic patients were impaired in growth potential and underwent oxidative stress and apoptosis. Expression of antioxidant gene oxidation resistance protein 1 (OXR1) specifically in pericytes through an adenovirus carrying OXR1 under a pericyte-specific neuron glia antigen-2 (NG2) promoter (AV-NG2p-OXR1) relieved the oxidative stress, reduced the apoptosis, and recovered the growth potential in diabetic pericytes. Moreover, expression of OXR1 in diabetic pericytes retrieved their potential of both suppressing the migration of co-cultured HUVECs and inducing cell aggregates at the branching points, indicating a functional recovery. In vivo gene therapy using this AV-NG2p-OXR1 to DB/DB mice, the mouse model for type 2 diabetes, significantly improved wound healing, likely through enhancing blood flow at the wound rather than increasing vessel density. Together, our data suggest that gene therapy targeting inflammatory pericytes may improve diabetes-associated impaired wound healing.

NADH-Cytochrome B5 reductase 2 suppresses retinal vascular dysfunction through regulation of vascular endothelial growth factor A in diabetic retinopathy

Diabetic retinopathy (DR) is a progressive vascular complication of diabetes mellitus (DM) and is related to retinal vascular abnormalities. NADH-Cytochrome B5 Reductase 2 (CBR2) has been implicated in angiogenesis, but the effect of CBR2 on angiogenesis and endothelial cell biological behavior in DR remains unclear. Here, we aimed to explore the effect of CBR2 on retinal vascular dysfunction under diabetic conditions. The histological analyses were performed to explore the effect of CBR2 on pathological change in streptozotocin (STZ)-induced diabetic rat retinas. The effect of CBR2 on endothelial cell function was explored by CCK-8, scratch wound, transwell, tube formation, and immunofluorescence assays in high glucose (HG)-stimulated human retinal microvascular endothelial cells (HRMECs). CBR2 expression was significantly downregulated in DM rat retinas and HG-stimulated HRMECs. Intravitreal injection of CBR2-expressing lentivirus under diabetic conditions reduced retinal angiogenesis, acellular capillary formation, and pericyte loss, along with decreased expression of hypoxia-inducible factor-1α (HIF-1α), cluster of differentiation 31 (CD31), and vascular endothelial growth factor A (VEGFA) in vivo. Moreover, CBR2 overexpression inhibited cell growth and tube formation and led to decreased expression of HIF-1α and VEGFA in HG-induced HRMECs. Interestingly, the repressive effects of CBR2 on cell proliferation, migration, and tube formation under HG conditions were strongly reversed when VEGFA was overexpressed. Overall, the key findings of our study suggested that CBR2 might alleviate retinal vascular dysfunction and abnormal endothelial proliferation during the process of DR by regulating VEGFA, providing a piece of potent evidence for DR therapy.

Low-dose sodium-glucose cotransporter 2 inhibitor ameliorates ischemic brain injury in mice through pericyte protection without glucose-lowering effects

Antidiabetic sodium-glucose cotransporter 2 (SGLT2) inhibitors have attracted attention for their cardiorenal-protective properties beyond their glucose-lowering effect. However, their benefits in ischemic stroke remain controversial. Here we show the effects of luseogliflozin, a selective SGLT2 inhibitor, in acute ischemic stroke, using a permanent middle cerebral artery occlusion (pMCAO) model in non-diabetic mice. Pretreatment with low-dose luseogliflozin, which does not affect blood glucose levels, significantly attenuated infarct volume, blood-brain barrier disruption, and motor dysfunction after pMCAO. SGLT2 was expressed predominantly in brain pericytes and was upregulated in peri- and intra-infarct areas. Notably, luseogliflozin pretreatment reduced pericyte loss in ischemic areas. In cultured pericytes, luseogliflozin activated AMP-activated protein kinase α and increased mitochondrial transcription factor A expression and number of mitochondria, conferring resistance to oxygen-glucose deprivation. Collectively, pre-stroke inhibition of SGLT2 induces ischemic tolerance in brain pericytes independent of the glucose-lowering effect, contributing to the attenuation of ischemic brain injury.

Protective effects of omega-3 fatty acids in a blood–brain barrier-on-chip model and on postoperative delirium-like behaviour in mice

BackgroundPeripheral surgical trauma can trigger neuroinflammation and ensuing neurological complications, such as delirium. The mechanisms whereby surgery contributes to postoperative neuroinflammation remain unclear and without effective therapies. Here, we developed a microfluidic-assisted blood–brain barrier (BBB) device and tested the effects of omega-3 fatty acids on neuroimmune interactions after orthopaedic surgery. MethodsA microfluidic-assisted BBB device was established using primary human cells. Tight junction proteins, vascular cell adhesion molecule 1 (VCAM-1), BBB permeability, and astrocytic networks were assessed after stimulation with interleukin (IL)-1β and in the presence or absence of a clinically available omega-3 fatty acid emulsion (Omegaven®; Fresenius Kabi, Bad Homburg, Germany). Mice were treated 1 h before orthopaedic surgery with 10 μl g−1 body weight of omega-3 fatty acid emulsion i.v. or equal volumes of saline. Changes in pericytes, perivascular macrophages, BBB opening, microglial activation, and inattention were evaluated. ResultsOmega-3 fatty acids protected barrier permeability, endothelial tight junctions, and VCAM-1 after exposure to IL-1β in the BBB model. In vivo studies confirmed that omega-3 fatty acid treatment inhibited surgery-induced BBB impairment, microglial activation, and delirium-like behaviour. We identified a novel role for pericyte loss and perivascular macrophage activation in mice after surgery, which were rescued by prophylaxis with i.v. omega-3 fatty acids. ConclusionsWe present a new approach to study neuroimmune interactions relevant to perioperative recovery using a microphysiological BBB platform. Changes in barrier function, including dysregulation of pericytes and perivascular macrophages, provide new targets to reduce postoperative delirium.

MTOR inhibition as a novel gene therapeutic strategy for diabetic retinopathy.

In addition to laser photocoagulation, therapeutic interventions for diabetic retinopathy (DR) have heretofore consisted of anti-VEGF drugs, which, besides drawbacks inherent to the treatments themselves, are limited in scope and may not fully address the condition’s complex pathophysiology. This is because DR is a multifactorial condition, meaning a gene therapy focused on a target with broader effects, such as the mechanistic target of rapamycin (mTOR), may prove to be the solution in overcoming these concerns. Having previously demonstrated the potential of a mTOR-inhibiting shRNA packaged in a recombinant adeno-associated virus to address a variety of angiogenic retinal diseases, here we explore the effects of rAAV2-shmTOR-SD in a streptozotocin-induced diabetic mouse model. Delivered via intravitreal injection, the therapeutic efficacy of the virus vector upon early DR processes was examined. rAAV2-shmTOR-SD effectively transduced mouse retinas and therein downregulated mTOR expression, which was elevated in sham-treated and control shRNA-injected (rAAV2-shCon-SD) control groups. mTOR inhibition additionally led to marked reductions in pericyte loss, acellular capillary formation, vascular permeability, and retinal cell layer thinning, processes that contribute to DR progression. Immunohistochemistry showed that rAAV2-shmTOR-SD decreased ganglion cell loss and pathogenic Müller cell activation and proliferation, while also having anti-apoptotic activity, with these effects suggesting the therapeutic virus vector may be neuroprotective. Taken together, these results build upon our previous work to demonstrate the broad ability of rAAV2-shmTOR-SD to address aspects of DR pathophysiology further evidencing its potential as a human gene therapeutic strategy for DR.

Transcription factor FOXP1 mediates vascular endothelial dysfunction in diabetic retinopathy.

Diabetic retinopathy (DR) is still the fastest growing cause of blindness in working aged adults, and its typical characteristics are endothelial cell dysfunction and pericytes loss. Transcription factor fork head box P1 (FOXP1) is a member of FOX family involved in diabetes progression and is expressed in endothelial cells. The purpose of this study was to investigate the role and mechanism of FOXP1 in DR. The vitreous of DR patients and non-DR patients were collected, and the expression of FOXP1 was detected by real-time polymerase chain reaction (RT-qPCR) and enzyme-linked immunosorbent assay (ELISA). Human umbilical vein endothelial cells (HUVECs) cultured in high glucose simulated DR environment, and the expressions of FOXP1, vascular endothelial growth factor (VEGF), and pigment epithelium derived factor (PEDF) were detected by RT-qPCR and western blot (WB) after transfection of small interfering RNA (siRNA) to knock out FOXP1. At the same time, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay (MTT), 5-ethynyl-2'-deoxyuridine assay (EDU), flow cytometry, Transwell assay, and tube-forming experiment were performed to determine cell proliferation, migration, and tube-forming ability. We found that FOXP1 was highly expressed in the vitreous of DR patients and HUVECs under high glucose condition. After FOXP1 was decreased, the activation of VEGF expression and inhibition of PEDF expression in HUVECs induced by high glucose were reversed; meanwhile, cell proliferation, migration, and tube formation decreased, and apoptosis was promoted. Generally, FOXP1 is highly expressed in the vitreous of DR patients, and its silence prevented VEGF/PEDF signaling pathway stimulated by high glucose and also reduced the proliferation, migration, and tube formation of endothelial cell, thus improving vascular endothelial dysfunction caused by DR. The results indicate that FOXP1 may be a therapeutic target of DR.

Mito-TEMPO, a mitochondria-targeted antioxidant, improves cognitive dysfunction due to hypoglycemia: An association with reduced pericyte loss and blood-brain barrier leakage

Mito-TEMPO, a mitochondria-targeted antioxidant, improves cognitive dysfunction due to hypoglycemia: An association with reduced pericyte loss and blood-brain barrier leakage