Common Carotid Artery Ligation Research Articles

Abstract TMP111: Perivascular von Willebrand factor (VWF) promotes irregular remodeling of leptomeningeal collaterals in response to sustained increased luminal flow.

Introduction: VWF is an endothelial protein with known roles in hemostasis and thrombosis. We previously demonstrated abnormal deposition of VWF in the mural layer of leptomeningeal collateral arterioles (LMCs) during flow-induced outward remodeling. Additionally, LMC remodeling was attenuated in VWF KO mice, suggesting a modulating role for VWF. We now test the hypothesis that intramural VWF potentiates LMC remodeling in response to sustained increased luminal flow. Methods: VWF protein was infused into the cisterna magna (CM) of WT and VWF KO C57BL/6 mice (3-7 mos females) to deliver VWF to the perivascular space around the leptomeningeal arteries (2.5 mg/ml at 1 ul/min; 5-7ul total). To confirm successful delivery of VWF to the wall of the leptomeningeal vessels, brain surface preparations containing the cortical leptomeningeal vessels were “planed off” and evaluated for VWF and smooth muscle actin (SMA) immunofluorescence at one hour after VWF infusion (cohort 1). In a subsequent experiment, right common carotid artery ligation (rCCAL) was performed on VWF KO and WT mice to induce increased flow in collateral vessels supplying the right MCA territory (cohort 2). VWF or vehicle was injected via CM at 16 hours after the rCCAL. At 3 days post-rCCAL, the intact leptomeningeal vessel preparations were evaluated by immunofluorescence (Ki67&SMA) and morphometric analyses. Results: [Cohort 1] WT and VWF KO mice demonstrated exogenous VWF protein surrounding the leptomeningeal vessels and within the mural layer. VWF KO mice were used to distinguish between endogenous and exogenous VWF. [Cohort 2] At 3 days, rCCAL induced endothelial cell proliferation (Ki67+) and outward vascular remodeling of collateral vessels supplying rescue flow following carotid ligation. In the VWF infusion group, there was a significant increase in Ki67+ ECs, 14.1 ± 3.1 vs. 5.4 ± 1.2 per mm of vessel of the control group ( p = 0.042). In VWF KO mice, VWF infusion also increased the number of Ki67+ ECs, showing 18.3 ± 3.2 vs. 6.1 ± 2.8 per mm of vessel in the control group ( p < 0.0001). In both WT and VWF KO mice, VWF infusion promoted non-uniform LMC remodeling. Conclusions: Our study demonstrates that VWF in the vascular wall of LMCs potentiates the adaptive response to increased luminal flow, albeit with irregular resulting vascular morphology. These finding suggest that the presence of intramural VWF may contribute to abnormal vascular remodeling in the brain.

Human umbilical cord mesenchymal stem cell-derived exosomes combined with mouse nerve growth factor can more effectively ameliorate the motor disorder and brain pathological injury in mice with cerebral palsy.

Cerebral palsy (CP) is a neurodevelopmental disorder and motor disorder syndrome. It has been confirmed that mesenchymal stem cells (MSCs) and mouse nerve growth factor (mNGF) can repair brain tissue damage and nerve injury; however, exosomes derived from healthy cells may have a comparable therapeutic potential as the cells themselves. The purpose of this study was to explore the improvement effect of human umbilical cord mesenchymal stem cell (hUC-MSCs)-derived exosomes on a CP model and determine whether there is a synergistic effect when combined with mNGF. Exosomes were isolated from hUC-MSCs and examined using transmission electron microscopy (TEM), particle size and western blot (WB). A total of 38 BALB/c mice (male, postnatal day 6 (PND6)) were randomly divided into 5 groups: sham group, CP group, CP-exo group, CP-mNGF group, and CP-exo-mNGF group. Hypoxic induction after unilateral common carotid artery ligation combined with lipopolysaccharide (LPS) infection was used to construct the CP model. Pathological damage to neuron tissue and synaptic structures in the hippocampus was confirmed using light microscopy after hematoxylin-eosin (H&E) staining and TEM, respectively. Survival of neurons was evaluated using Nissl staining. Western blot was applied to monitor PSD-95 and synaptophysin (SYN) protein levels. This study indicated that exosomes released by hUC-MSCs ameliorated brain damage and synaptic structure destruction in CP mice induced by hypoxic ischemia and LPS infection. When combined with mNGF, there was more effective improvement. In the CP group, neuronal function was severely impaired; however, hUC-MSCs-derived exosomes and mNGF improved it. PSD-95 and SYN proteins were presynaptic and postsynaptic proteins, respectively. Interestingly, the PSD-95 and SYN protein levels were significantly lower in the CP mice, but with the addition of hUC-MSCs-exosomes or mNGF, they increased significantly, especially in the CP-exo-mNGF group. The nerve function injury in CP can be improved the most when hUC-MSCs-derived exosomes are combined with mNGF through intraperitoneal (ip.) administration.

Hybrid Thoracic Endovascular Repair of a Large, Saccular Aortic Arch Aneurysm with Coil Embolization of the Left Subclavian Artery in a 31 Year Old Filipino Female with Takayasu Arteritis and Multiple Intracranial Aneurysms: A First in the Philippines

BACKGROUND: Takayasu Arteritis (TA) is a rare, primary large-vessel vasculitis frequently leading to stenosis and less commonly, aneurysm formation. Saccular aneurysms of the aortic arch in patients with TA are fatal, have rarely been reported and represent a significant technical challenge due to the difficult anatomical location and need for protection of the cerebral circulation. Concomitant intracerebral aneurysms in patients with TA are extremely uncommon and have mostly been documented in very few case reports in literature. CASE DISCUSSION: We present a case of a 31 year-old Filipino female with recurrent chest and neck pain radiating to the upper back. Computed tomographic (CT) angiography demonstrated a large saccular aortic arch aneurysm without branch stenosis. CTA of the cerebral circulation likewise demonstrated multiple, saccular, intra-cerebral aneurysms. She underwent hybrid thoracic arch repair with supra-aortic debranching via mini-sternotomy and proximal ligation of the left common carotid artery and staged endovascular aortic arch replacement with coil embolization of the ostial-to-proximal left subclavian artery segment. Post-operative aortogram showed optimal repair with thrombosed aneurysmal sac, optimal graft position, no endoleaks and preservation of cerebral circulation. Patient improved symptomatically post-procedure and remained symptom-free during follow-up after six months. Careful review of local literature suggests that this is the first Philippine TA case with a saccular aortic arch aneurysm successfully managed in this manner. CONCLUSION: Saccular aortic arch aneurysms in patients with Takayasu are unusual and presence of concomitant multiple cerebral saccular aneurysms have rarely been reported in literature. This case highlighted that hybrid endovascular arch repair in patients with TA is feasible, minimally invasive and effective. KEYWORDS: Takayasu arteritis, aortic arch aneurysm, cerebral aneurysm, hybrid endovascular arch repair, aortic de-branching

Gastrodin alleviates microglia-mediated inflammatory responses in neonatal mice with hypoxic-ischemic brain damage by regulating CCR5/AKT signaling

To investigate the mechanism behind the protective effects of gastrodin against microglia-mediated inflammatory responses following hypoxic-ischemic brain damage (HIBD) in neonatal mice. Thirty-six 10-day-old C57BL/6J mice were randomized into sham-operated group, HIBD (induced by ligation of the left common carotid artery followed by hypoxia for 40 min) group, and HIBD with gastrodin treatment groups (n=12). In gastrodin treatment group, 100 mg/kg gastrodin was injected intraperitoneally 1 h before and at 2 and 12 h after hypoxia. After the treatments, the expressions of CCR5, AKT, p-AKT, and TNF-α and the co-expression of IBA1 and CCR5 in the corpus callosum of the mice were detected with Western blotting and immunofluorescence double staining. In a BV2 microglial cell model of oxygen-glucose deprivation (OGD), the effects of pretreatment with gastrodin and Maraviroc (an CCR5 antagonist) on protein expressions of CCR5, AKT, p-AKT, TNF-α and IL-1β were evaluated using Western blotting and immunofluorescence double staining. The neonatal mice with HIBD showed significantly increased expressions of CCR5 and TNF-α with lowered p-AKT expression in the brain tissues, and GAS treatment obviously reversed these changes. HIBD also significantly increased the co-expression of IBA1 and CCR5 in the corpus callosum of the mice, which was obviously lowered by gastrodin treatment. In BV2 cells, OGD significantly increased the expressions of CCR5, TNF-α, and IL-1β and decreased the expression of p-AKT, and these changes were inhibited by treatment with gastrodin, Maraviroc or their combination; the inhibitory effect of the combined treatment did not differ significantly from that of gastrodin or Maraviroc alone. Gastrodin can produce neuroprotective effects in neonatal mice with HIBD by inhibiting inflammatory cytokine production and activate AKT phosphorylation via inhibiting CCR5.

Reliable infarction of the middle cerebral artery territory in C57BL/6 mice using pterygopalatine artery ligation and filament optimization - The PURE-MCAo model.

Current techniques for inducing intraluminal filamentous middle cerebral artery occlusion (fMCAo) in mice produce highly variable results and often cause additional infarcts in the posterior cerebral artery (PCA) territory. The aim of the current study was to develop a novel procedure to overcome these shortcomings. Male C57BL/6 mice were subjected to 60 min of fMCAo with cerebral blood flow monitored by laser Doppler flowmetry. The influence of the length of the occlusion filament coating and the combination of common carotid artery (CCA) or pterygopalatine artery (PPA) ligation on lesion volume and functional outcome 24 h after reperfusion was evaluated. The use of appropriate filament and PPA ligation while maintaining CCA perfusion prevented the development of infarcts in the PCA area, resulted in pure MCA infarcts (68.3 ± 14.5 mm3) and reduced the variability of infarct volumes by more than half (from 26-38% to 14% standard deviation/mean). Using an improved fMCAo procedure, we were able to produce PCA area-unaffected reproducible (PURE) infarcts exclusively in the MCA territory. Thus PURE-MCAo reduced outcome variability by more than 50%. Our results may thus help to reduce the number of animals in preclinical stroke research and to increase the reproducibility of the fMCAo model.

Molecular profiling of a rat model of vascular dementia: Evidences from proteomics, metabolomics and experimental validations

Decrease of cerebral blood flow is the primary cause of vascular dementia (VD), but its pathophysiological mechanisms are still not known. This study aims to profile the molecular changes of a rat model of VD induced by bilateral common carotid artery ligation. The Morris water maze and new object recognition tasks were used to test the cognitive function of rats. Hematoxylin and Eosin (HE) staining was used to detect pathological changes in the hippocampus. After confirming the model, proteomics was used to detect differentially expressed proteins in the hippocampus, and metabolomics was used to detect differential metabolites in rat serum. Thereafter, bioinformatics were used to integrate and analyze the potential molecular profile. The results showed that compared with the sham control group, the spatial and recognition memory of the rats were significantly reduced, and pathological changes were observed in the hippocampal CA1 region of the model group. Proteomic analysis suggested 206 differentially expressed proteins in the hippocampus of VD rats, with 117 proteins upregulated and 89 downregulated. Protein-protein interaction network analysis suggested that those differentially expressed proteins might play crucial roles in lipid metabolism, cell adhesion, intracellular transport, and signal transduction. Metabolomics analysis identified 103 differential metabolites, and comparison with the human metabolome database revealed 22 common metabolites, which predicted 265 potential targets. Afterwards, by intersecting the predicted results from metabolomics with the differentially expressed proteins from proteomics, we identified five potential targets, namely ACE, GABBR1, Rock1, Abcc1 and Mapk10. Furthermore, western blotting confirmed that compared with control group, hippocampal GABBR1 and Rock1 were enhanced in the model group. Together, this study showed the molecular profile of VD rats through a combination of proteomics, metabolomics, and experimental confirmation methods, offering crucial molecular targets for the diagnosis and treatment of VD.

An Organic Fraction of Oenothera rosea L'Her Ex. Aiton Prevents Neuroinflammation in a Rat Ischemic Model.

Oenothera rosea L'Her Ex. Aiton, presenting antioxidant and anti-inflammatory activities, is traditionally used to treat bruises and headaches and as a healing agent. This study aimed to investigate whether its organic fraction (EAOr) has neuroprotective properties against neuroinflammation in the context of ischemia/reperfusion. The chemical composition of EAOr was determined using HPLC techniques, and its neuroprotective activities were evaluated in a common carotid-artery ligation model for the induction of ischemia/reperfusion (I/R). The animals were supplemented with EAOR for 15 days. On the last day, the animals were rested for one hour, following which the common carotid-artery ligation procedure was performed to induce I/R. The neurological deficit was evaluated at 24 h after I/R using Bederson's scale, and the relative expression of inflammatory genes and structure of hippocampal neurons were analyzed at 48 h. The chemical analysis revealed five major compounds in EAOr: gallic acid, rutin, ellagic acid, and glucoside and rhamnoside quercetin. EAOr prevented neurological deficit 24 h after I/R; led to the early activation of the AIF and GFAP genes; reduced Nfkb1, IL-1beta, Il-6 and Casp3 gene expression; and protected hippocampal neurons. Our findings demonstrate that EAOr contains polyphenol-type compounds, which could exert a therapeutic effect through the inhibition of neuroinflammation and neuronal death genes, thus maintaining hippocampal neurons.

BNIP3-mediated mitophagy attenuates hypoxic-ischemic brain damage in neonatal rats by inhibiting ferroptosis through P62-KEAP1-NRF2 pathway activation to maintain iron and redox homeostasis.

As a major contributor to neonatal death and neurological sequelae, hypoxic-ischemic encephalopathy (HIE) lacks a viable medication for treatment. Oxidative stress induced by hypoxic-ischemic brain damage (HIBD) predisposes neurons to ferroptosis due to the fact that neonates accumulate high levels of polyunsaturated fatty acids for their brain developmental needs but their antioxidant capacity is immature. Ferroptosis is a form of cell death caused by excessive accumulation of iron-dependent lipid peroxidation and is closely associated with mitochondria. Mitophagy is a type of mitochondrial quality control mechanism that degrades damaged mitochondria and maintains cellular homeostasis. In this study we employed mitophagy agonists and inhibitors to explore the mechanisms by which mitophagy exerted ferroptosis resistance in a neonatal rat HIE model. Seven-days-old neonatal rats were subjected to ligation of the right common carotid artery, followed by exposure to hypoxia for 2 h. The neonatal rats were treated with a mitophagy activator Tat-SPK2 peptide (0.5, 1 mg/kg, i.p.) 1 h before hypoxia, or in combination with mitochondrial division inhibitor-1 (Mdivi-1, 20 mg/kg, i.p.), and ferroptosis inhibitor Ferrostatin-1 (Fer-1) (2 mg/kg, i.p.) at the end of the hypoxia period. The regulation of ferroptosis by mitophagy was also investigated in primary cortical neurons or PC12 cells in vitro subjected to 4 or 6 h of OGD followed by 24 h of reperfusion. We showed that HIBD induced mitochondrial damage, ROS overproduction, intracellular iron accumulation, lipid peroxidation and ferroptosis, which were significantly reduced by the pretreatment with Tat-SPK2 peptide, and aggravated by the treatment with Mdivi-1 or BNIP3 knockdown. Ferroptosis inhibitors Fer-1 and deferoxamine B (DFO) reversed the accumulation of iron and lipid peroxides caused by Mdivi-1, hence reducing ferroptosis triggered by HI. We demonstrated that Tat-SPK2 peptide-activated BNIP3-mediated mitophagy did not alleviate neuronal ferroptosis through the GPX4-GSH pathway. BNIP3-mediated mitophagy drove the P62-KEAP1-NRF2 pathway, which conferred ferroptosis resistance by maintaining iron and redox homeostasis via the regulation of FTH1, HO-1, and DHODH/FSP1-CoQ10-NADH. This study may provide a new perspective and a therapeutic drug for the treatment of neonatal HIE.

Phoenixin-14 maintains the contractile type of vascular smooth muscle cells in cerebral aneurysms rats.

The rupture of intracranial aneurysm (IA) is the primary reason contributing to the occurrence of life-threatening subarachnoid hemorrhages. The oxidative stress-induced phenotypic transformation from the contractile phenotype to the synthetic phenotype of vascular smooth muscle cells (VSMCs) plays a pivotal role in IA formation and rupture. Our study aimed to figure out the role of phoenixin-14 in VSMC phenotypic switching during the pathogenesis of IA by using both cellular and animal models. Primary rat VSMCs were isolated from the Willis circle of male Sprague-Dawley rats. VSMCs were stimulated by hydrogen peroxide (H2O2) to establish a cell oxidative damage model. After pretreatment with phoenixin-14 and exposure to H2O2, VSMC viability, migration, and invasion were examined through cell counting kit-8 (CCK-8), wound healing, and Transwell assays. Intracellular reactive oxygen species (ROS) production in VSMCs was evaluated by using 2',7'-Dichlorofluorescin diacetate (DCFH-DA) fluorescence probes and flow cytometry. Rat IA models were established by ligation of the left common carotid arteries and posterior branches of both renal arteries. The histopathological changes of rat intracranial blood vessels were observed through hematoxylin and eosin staining. The levels of contractile phenotype markers (alpha-smooth muscle actin [α-SMA] and smooth muscle 22 alpha [SM22α]) in VSMCs and rat arterial rings were determined through real-time quantitative polymerase chain reaction (RT-qPCR) and western blot analysis. Our results showed that H2O2 stimulated the production of intracellular ROS and induced oxidative stress in VSMCs, while phoenixin-14 pretreatment attenuated intracellular ROS levels in H2O2-exposed VSMCs. H2O2 exposure promoted VSMC migration and invasion, which, however, was reversed by phoenixin-14 pretreatment. Besides, phoenixin-14 administration inhibited IA formation and rupture in rat models. The decrease in α-SMA and SM22α levels in H2O2-exposed VSMCs and IA rat models was antagonized by phoenixin-14. Collectively, phoenixin-14 ameliorates the progression of IA through preventing the loss of the contractile phenotype of VSMCs.

Inhibition of histone methyltransferase promotes cognition and mitochondrial function in vascular dementia model

Vascular dementia (VD) is one of the most common forms of dementia worldwide, characterized by problems with reasoning, planning, judgment, and memory. This study investigated the effect of a histone methyltransferase inhibitor on cognition and mitochondrial function in a rat model of VD, as well as its impact on H2O2-induced neurotoxicity in hippocampal neuronal cultures. In the in vivo experiments, VD was induced by bilateral occlusion of the common carotid artery (CCA) for one month. The histone methyltransferase inhibitor, BIX01294, was administered intracerebroventricularly for one month (22.5 µg.kg-1 three times/week). On day 30, behavioral tests, including the novel object recognition test and elevated plus maze test, were conducted. Mitochondrial enzyme activities, including aconitase, α-ketoglutarate dehydrogenase (α-KG), complex I, and complex IV, were evaluated in the hippocampus of rats following CCA ligation. In the in vitro experiments, the effect of BIX01294 (50-600 μM) on H2O2 (400 µM)-induced cytotoxicity in hippocampal neuronal cells was assessed using the MTT assay. Flow cytometry was performed to evaluate apoptosis. Our findings revealed that BIX01294 effectively improved memory function, Krebs cycle enzyme activity, and mitochondrial function in the rat model of VD. Moreover, in vitro results showed that BIX01294 at a concentration of 100 µM significantly reversed the cytotoxicity and apoptosis induced by H2O2 in neuronal cells. These findings suggest that BIX01294 may have the potential to improve VD complications by reducing oxidative stress and inhibiting histone methylation.

Intermittent theta-burst stimulation alleviates hypoxia-ischemia-caused myelin damage and neurologic disability

Neonatal hypoxia-ischemia (HI) results in behavioral deficits, characterized by neuronal injury and retarded myelin formation. To date, limited treatment methods are available to prevent or alleviate neurologic sequelae of HI. Intermittent theta-burst stimulation (iTBS), a non-invasive therapeutic procedure, is considered a promising therapeutic tool for treating some neurocognitive disorders and neuropsychiatric diseases. Hence, this study aims to investigate whether iTBS can prevent the negative behavioral manifestations of HI and explore the mechanisms for associations. We exposed postnatal day 10 Sprague-Dawley male and female rats to 2h of hypoxia (6% O2) following right common carotid artery ligation, resulting in oligodendrocyte (OL) dysfunction, including reduced proliferation and differentiation of oligodendrocyte precursor cells (OPCs), decreased OL survival, and compromised myelin in the corpus callosum (CC) and hippocampal dentate gyrus (DG). These alterations were concomitant with cognitive dysfunction and depression-like behaviors. Crucially, early iTBS treatment (15G, 190s, seven days, initiated one day post-HI) significantly alleviated HI-caused myelin damage and mitigated the neurologic sequelae both in male and female rats. However, the late iTBS treatment (initiated 18days after HI insult) could not significantly impact these behavioral deficits. In summary, our findings support that early iTBS treatment may be a promising strategy to improve HI-induced neurologic disability. The underlying mechanisms of iTBS treatment are associated with promoting the differentiation of OPCs and alleviating myelin damage.

Lacidipine, thiamine pyrophosphate and their combination on the ocular ischemic syndrome induced by bilateral common carotid artery ligation.

To investigate the effect of lacidipine, thiamine pyrophosphate (TPP) and the combination of lacidipine and TPP against oxidative and inflammatory eye damage induced by bilateral common carotid artery ligation in rats. Male albino Wistar rats were categorized as those who underwent sham surgery (SG), right and left common carotid cross-clamping and unclamping procedure (CCU), lacidipine+CCU (LCCU), TPP+CCU (TCCU), and combination of lacidipine and TPP (LTC)+CCU (LTCCU). One hour before anesthesia, the LCCU (n=6) received lacidipine (4 mg/kg, orally) and the TCCU (n=6) received TPP (20 mg/kg, intraperitoneally). The SG (n=6) and CCU (n=6) received the same volume of distilled water from the same route. After anesthesia (60 mg/kg ketamine, intraperitoneally), the necks of the rats were opened in the midline. Ischemia was created for 10min by placing clips on the right and left common carotid arteries. Rats in the SG only underwent subcutaneous incision. After 10min, the clips were removed and reperfusion was achieved for six days. Then, the animals were euthanized (120 mg/kg ketamine, intraperitoneally) and the levels of oxidant, antioxidant and proinflammatory cytokines in the eye tissues were determined. The retinal tissue of the eye was also examined histopathologically. Lacidipine, TPP, and LTC significantly prevent the increase in malondialdehyde, tumor necrosis factor-alpha, interleukin-1β (IL-1β), and IL-6 levels, decrease in total glutathione levels, superoxide dismutase and catalase activities and histopathological retinal damage in eye tissue induced by bilateral common carotid artery ligation in rats. The impact of these drugs on protection is determined to be LTC>lacidipine>TPP. As a result of the study, it is concluded that LTC may be more effective than lacidipine and TPP alone in treating ocular ischemic syndrome.

Protecting effects of 4-octyl itaconate on neonatal hypoxic-ischemic encephalopathy via Nrf2 pathway in astrocytes

BackgroundNeonatal hypoxic-ischemic encephalopathy (HIE) is one of the most common neurological problems occurring in the perinatal period. However, there still is not a promising approach to reduce long-term neurodevelopmental outcomes of HIE. Recently, itaconate has been found to exhibit anti-oxidative and anti-inflammatory effects. However, the therapeutic efficacy of itaconate in HIE remains inconclusive. Therefore, this study attempts to explore the pathophysiological mechanisms of oxidative stress and inflammatory responses in HIE as well as the potential therapeutic role of a derivative of itaconate, 4-octyl itaconate (4OI).MethodsWe used 7-day-old mice to induce hypoxic-ischemic (HI) model by right common carotid artery ligation followed by 1 h of hypoxia. Behavioral experiments including the Y-maze and novel object recognition test were performed on HI mice at P60 to evaluate long-term neurodevelopmental outcomes. We employed an approach combining non-targeted metabolomics with transcriptomics to screen alterations in metabolic profiles and gene expression in the hippocampal tissue of the mice at 8 h after hypoxia. Immunofluorescence staining and RT-PCR were used to evaluate the pathological changes in brain tissue cells and the expression of mRNA and proteins. 4OI was intraperitoneally injected into HI model mice to assess its anti-inflammatory and antioxidant effects. BV2 and C8D1A cells were cultured in vitro to study the effect of 4OI on the expression and nuclear translocation of Nrf2. We also used Nrf2-siRNA to further validate 4OI-induced Nrf2 pathway in astrocytes.ResultsWe found that in the acute phase of HI, there was an accumulation of pyruvate and lactate in the hippocampal tissue, accompanied by oxidative stress and pro-inflammatory, as well as increased expression of antioxidative stress and anti-inflammatory genes. Treatment of 4OI could inhibit activation and proliferation of microglial cells and astrocytes, reduce neuronal death and relieve cognitive dysfunction in HI mice. Furthermore, 4OI enhanced nuclear factor erythroid-2-related factor (Nfe2l2; Nrf2) expression and nuclear translocation in astrocytes, reduced pro-inflammatory cytokine production, and increased antioxidant enzyme expression.ConclusionOur study demonstrates that 4OI has a potential therapeutic effect on neuronal damage and cognitive deficits in HIE, potentially through the modulation of inflammation and oxidative stress pathways by Nrf2 in astrocytes.

PHLDA1 contributes to hypoxic ischemic brain injury in neonatal rats via inhibiting FUNDC1-mediated mitophagy.

Hypoxia-ischemia (HI) is one of the main causes of neonatal brain injury. Mitophagy has been implicated in the degradation of damaged mitochondria and cell survival following neonatal brain HI injury. Pleckstrin homology-like domain family A member 1 (PHLDA1) plays vital roles in the progression of various disorders including the regulation of oxidative stress, the immune responses and apoptosis. In the present study we investigated the role of PHLDA1 in HI-induced neuronal injury and further explored the mechanisms underlying PHLDA1-regulated mitophagy in vivo and in vitro. HI model was established in newborn rats by ligation of the left common carotid artery plus exposure to an oxygen-deficient chamber with 8% O2 and 92% N2. In vitro studies were conducted in primary hippocampal neurons subjected to oxygen and glucose deprivation/-reoxygenation (OGD/R). We showed that the expression of PHLDA1 was significantly upregulated in the hippocampus of HI newborn rats and in OGD/R-treated primary neurons. Knockdown of PHLDA1 in neonatal rats via lentiviral vector not only significantly ameliorated HI-induced hippocampal neuronal injury but also markedly improved long-term cognitive function outcomes, whereas overexpression of PHLDA1 in neonatal rats via lentiviral vector aggravated these outcomes. PHLDA1 knockdown in primary neurons significantly reversed the reduction of cell viability and increase in intracellular reactive oxygen species (ROS) levels, and attenuated OGD-induced mitochondrial dysfunction, whereas overexpression of PHLDA1 decreased these parameters. In OGD/R-treated primary hippocampal neurons, we revealed that PHLDA1 knockdown enhanced mitophagy by activating FUNDC1, which was abolished by FUNDC1 knockdown or pretreatment with mitophagy inhibitor Mdivi-1 (25 μM). Notably, pretreatment with Mdivi-1 or the knockdown of FUNDC1 not only increased brain infarct volume, but also abolished the neuroprotective effect of PHLDA1 knockdown in HI newborn rats. Together, these results demonstrate that PHLDA1 contributes to neonatal HI-induced brain injury via inhibition of FUNDC1-mediated neuronal mitophagy.

Urotensin II receptor deficiency ameliorates ligation-induced carotid intimal hyperplasia partially through the RhoA-YAP1 pathway

Intimal hyperplasia (IH) is a common pathological feature of vascular proliferative diseases, such as atherosclerosis and restenosis after angioplasty. Urotensin II (UII) and its receptor (UTR) are widely expressed in cardiovascular tissues. However, it remains unclear whether the UII/UTR system is involved in IH. Right unilateral common carotid artery ligation was performed and maintained for 21days to induce IH in UTR knockout (UTR-/-) and wild-type (WT) mice. Histological analysis revealed that compared with WT mice, UTR-deficient mice exhibited a decreased neointimal area, angiostenosis and intima-media ratio. Immunostaining revealed fewer smooth muscle cells (SMCs), endothelial cells and macrophages in the lesions of UTR-/- mice than in those of WT mice. Protein interaction analysis suggested that the UTR may affect cell proliferation by regulating YAP and its downstream target genes. In vitro experiments revealed that UII can promote the proliferation and migration of SMCs, and western blotting also revealed that UII increased the protein expression of RhoA, CTGF, Cyclin D1 and PCNA and downregulated p-YAP protein expression, while these effects could be partly reversed by urantide. To evaluate the translational value of UTRs in IH management, WT mice were also treated with two doses of urantide, a UTR antagonist, to confirm the benefit of UTR blockade in IH progression. A high dose of urantide (600μg/kg/day), rather than a low dose (60μg/kg/day), successfully improved ligation-induced IH compared with that in mice receiving vehicle. The results of the present study suggested that the UII/UTR system may regulate IH partly through the RhoA-YAP signaling pathway.

The role of the sensory input intervention in recovery of the motor function in hypoxic ischemic encephalopathy rat model.

Motor disturbances predominantly characterize hypoxic-ischemic encephalopathy (HIE). Among its intervention methods, environmental enrichment (EE) is strictly considered a form of sensory intervention. However, limited research uses EE as a single sensory input intervention to validate outcomes postintervention. A Sprague-Dawley rat model subjected to left common carotid artery ligation and exposure to oxygen-hypoxic conditions is used in this study. EE was achieved by enhancing the recreational and stress-relief items within the cage, increasing the duration of sunlight, colorful items exposure, and introducing background music. JZL184 (JZL) was administered as neuroprotective drugs. EE was performed 21 days postoperatively and the rats were randomly assigned to the standard environment and EE groups, the two groups were redivided into control, JZL, and vehicle injection subgroups. The Western blotting and behavior test indicated that EE and JZL injections were efficacious in promoting cognitive function in rats following HIE. In addition, the motor function performance in the EE-alone intervention group and the JZL-alone group after HIE was significantly improved compared with the control group. The combined EE and JZL intervention group exhibited even more pronounced improvements in these performances. EE may enhance motor function through sensory input different from the direct neuroprotective effect of pharmacological treatment.NEW & NOTEWORTHY Rarely does literature assess motor function, even though it is common after hypoxia ischemic encephalopathy (HIE). Previously used environmental enrichment (EE) components have not been solely used as sensory inputs. Physical factors were minimized in our study to observe the effects of purely sensory inputs.

Evaluation of PET imaging as a tool for detecting neonatal hypoxic-ischemic encephalopathy in a preclinical animal model

A preclinical model of neonatal hypoxic-ischemic brain injury was made in 9-d-old rat pups by permanent ligation of the left common carotid artery followed by hypoxia (8% oxygen and 92% nitrogen) for 120min. In vivo PET imaging was performed immediately after injury induction or at different timepoints up to 21 d later. After imaging, ex vivo brain autoradiography was performed. Brain sections were stained with cresyl violet to evaluate the extent of the brain injury and to correlate it with [18F]FDG uptake. PET imaging revealed that all three of the radiotracers tested had significant uptake in the injured brain hemisphere. Ex vivo autoradiography revealed high [18F]EF5 uptake in the hypoxic hemisphere immediately after the injury (P<0.0001), decreasing to baseline even 1 d postinjury. [18F]FDG uptake was highest in the injured hemisphere on the day of injury (P<0.0001), whereas [18F]F-DPA uptake was evident after 4 d (P=0.029), peaking 7 d postinjury (P<0.0001), and remained significant 21 d after the injury. Targeted evaluation demonstrated that [18F]FDG uptake measured by in vivo imaging 1 d postinjury correlated positively with the brain volume loss detected 21 d later (r=0.72, P=0.028). Neonatal hypoxic-ischemic brain injury can be detected using PET imaging. Different types of radiotracers illustrate distinct phases of hypoxic brain damage. PET may be a new useful technique, worthy of being explored for clinical use, to predict and evaluate the course of the injury.

Антиоксидантные эффекты синтетического аналога тиронамина при экспериментальной ишемии головного мозга

The oxidative stress associated with ischemic stroke is a major factor damaging the nervous tissue. Thyroid hormones have a significant effect on the body’s redox status, however, the impact of their derivatives, thyronamines, considered as potential neuroprotectors, on the characteristics of lipid peroxidation (LP) is not clearly understood. The study was aimed to assess the impact of the Т0АМ thyronamine synthetic analogue on the main LP indicators in the model of acute cerebral ischemia. Permanent ligation of the right common carotid artery was performed to simulate acute cerebral ischemia in white rats. The animals were divided into two groups: the control group receiving no treatment and the experimental group, to which the Т0АМ thyronamine synthetic analogue was intraperitoneally administrated (75 mg/kg of the rat’s body weight). After 24 h the rat was decapitated, and the cerebral cortex tissue was extracted for biochemical analysis. The following LP indicators were determined by spectrophotometry: malondialdehyde (MDA), superoxide dismutase (SOD), glutathione peroxidase (GPx). When administering the Т0АМ thyronamine synthetic analogue, a significant (2-fold) decrease in MDA levels was observed in the ischemic hemisphere (р = 0.022), along with the 2.49-fold increase in the GPx activity in the brain tissue (р = 0.004) of the intact hemisphere and the 2.65-fold increase in its activity (р = 0.021) in the ischemic hemisphere, as well as the 1.23-fold increase in SOD activity in the ischemic hemisphere (р = 0.042). The Т0АМ thyronamine synthetic analogue has a great potential in terms of activation of the antioxidant protection mechanisms in the cerebral cortex of white laboratory rats under conditions of acute hemispheric ischemia.

Human amniotic mesenchymal stromal cell-derived exosomes promote neuronal function by inhibiting excessive apoptosis in a hypoxia/ischemia-induced cerebral palsy model: A preclinical study

Cerebral palsy (CP) is a condition resulting from perinatal brain injury and can lead to physical disabilities. Exosomes derived from human amniotic mesenchymal stromal cells (hAMSC-Exos) hold promise as potential therapeutic options. This study aimed to investigate the impact of hAMSC-Exos on neuronal cells and their role in regulating apoptosis both in vitro and in vivo. hAMSC-Exos were isolated via ultracentrifugation and characterized via transmission electron microscopy, particle size analysis, and flow cytometry. In vitro, neuronal damage was induced by lipopolysaccharide (LPS). CP rat models were established via left common carotid artery ligation. Apoptosis levels in cells and CP rats were assessed using flow cytometry, quantitative reverse transcription polymerase chain reaction (RT-qPCR), Western blotting, and TUNEL analysis. The results demonstrated successful isolation of hAMSC-Exos via ultracentrifugation, as the isolated cells were positive for CD9 (79.7%) and CD63 (80.2%). Treatment with hAMSC-Exos significantly mitigated the reduction in cell viability induced by LPS. Flow cytometry revealed that LPS-induced damage promoted apoptosis, but this effect was attenuated by treatment with hAMSC-Exos. Additionally, the expression of caspase-3 and caspase-9 and the Bcl-2/Bax ratio indicated that excessive apoptosis could be attenuated by treatment with hAMSC-Exos. Furthermore, tail vein injection of hAMSC-Exos improved the neurobehavioral function of CP rats. Histological analysis via HE and TUNEL staining showed that apoptosis-related damage was attenuated following hAMSC-Exo treatment. In conclusion, hAMSC-Exos effectively promote neuronal cell survival by regulating apoptosis, indicating their potential as a promising therapeutic option for CP that merits further investigation.

Veno-arterial extracorporeal membrane oxygenation for respiratory and cardiac support in neonates: a single center experience.

Extracorporeal membrane oxygenation (ECMO) is an advanced life support that has been utilized in the neonate for refractory respiratory and circulatory failure. Striving for the best outcomes and understanding optimal surgical techniques continue to be at the forefront of discussion and research. This study presents a single-center experience of cervically cannulated neonatal patients on V-A ECMO, a description of our cannulation/decannulation techniques and our patient outcomes. Single center retrospective review of neonates who received neck V-A ECMO support from January 2012 to December 2022. The data and outcomes of the patients were retrospectively analyzed. A total of 78 neonates received V-A ECMO support. There were 66 patients that received ECMO for respiratory support, the other 12 patients that received ECMO for cardiac support. The median duration of ECMO support was 109 (32-293) hours for all patients. During ECMO support, 20 patients died and 5 patients discontinued treatment due to poor outcome or the cost. A total of 53 (68%) patients were successfully weaned from ECMO, but 3 of them died in the subsequent treatment. Overall 50 (64%) patients survived to hospital discharge. In this study, 48 patients were cannulated using the vessel sparing technique, the other 30 patients were cannulated using the ligation technique. We found no significant difference in the rates of normal cranial MRI at discharge between survivors with and without common carotid artery ligation. We achieved satisfactory outcomes of neonatal ECMO in 11-year experience. This study found no significant difference in early neuroimaging between survivors with and without common carotid artery ligation. The long-term neurological function of ECMO survivors warranted further follow-up and study.