Brain Injury In Neonatal Rats Research Articles

Genistein-3′-sodium sulfonate enhances neurological function in neonatal rats with hypoxia-ischemia during the recovery period

Hypoxic-ischemic (HI) can cause neonatal brain damage leading to disability. Patients with HI experience long-term neurological issues impacting quality of life. Limited clinical treatments are available despite extensive research on HI's molecular mechanisms. Genistein-3′-sodium sulfonate (GSS), a phytoestrogen, has been found to improve acute brain injury in neonatal rats caused by hypoxic-ischemia, but its potential for chronic stage neurological recovery in HI is unknown. HI neonatal rats were treated with 1 mg/kg GSS once a day for 21 days. Then, a series of behavioral experiments was performed to evaluate the learning, memory, cognition, anxiety level and depression-like behaviors of the rats. GSS treatment reduced neuronal loss, enhanced learning, memory and cognitive function while also alleviated anxiety and depression-like behaviors in HI rats during the recovery period. These findings indicated that GSS exerted enhance neurological function in HI rats during the chronic stage, prompting further research on how it works to potentially develop new therapies.

Celastrol ameliorates hypoxic-ischemic brain injury in neonatal rats by reducing oxidative stress and inflammation.

Hypoxic-ischemic encephalopathy (HIE) is caused by perinatal hypoxia and subsequent reductions in cerebral blood flow and is one of the leading causes of severe disability or death in newborns. Despite its prevalence, we currently lack an effective drug therapy to combat HIE. Celastrol (Cel) is a pentacyclic triterpene extracted from Tripterygium Wilfordi that can protect against oxidative stress, inflammation, and cancer. However, whether Cel can alleviate neonatal hypoxic-ischemic (HI) brain damage remains unclear. Here, we established both in vitro and in vivo models of HI brain damage using CoCl2-treated PC12 cells and neonatal rats, respectively, and explored the neuroprotective effects of Cel in these models. Analyses revealed that Cel administration reduced brain infarction size, microglia activation, levels of inflammation factors, and levels of oxidative stress markers by upregulating levels of p-AMPKα, Nrf2, HO-1, and by downregulating levels of TXNIP and NLRP3. Conversely, these beneficial effects of Cel on HI brain damage were largely inhibited by AMPKα inhibitor Compound C and its siRNA. We present compelling evidence that Cel decreases inflammation and oxidative stress through the AMPKα/Nrf2/TXNIP signaling pathway, thereby alleviating neonatal HI brain injury. Cel therefore represents a promising therapeutic agent for treating HIE. We firstly report that celastrol can ameliorate neonatal hypoxic-ischemic brain injury both in in vivo and in vitro, which represents a promising therapeutic agent for treating related brain injuries. Celastrol activates the AMPKα/Nrf2/TXNIP signaling pathway to relieve oxidative stress and inflammation and thereby alleviates neonatal hypoxic-ischemic brain injury.

Docosahexaenoic Acid Supplementation for Neonatal Hyperbilirubinemia: A Double-Blind, Randomized Clinical Trial.

Docosahexaenoic acid (DHA) is an essential component for brain development during fetal and early postnatal life. Hyperbilirubinemia is characterized by abnormally high levels of bilirubin in the bloodstream, frequently leading to jaundice in newborns. In severe instances, this condition can progress to neurological damage or kernicterus, a form of brain damage. Initial cell-based experiments conducted by our research team revealed that DHA significantly enhances the survival rate of nerve cells treated with bilirubin and diminishes the oxidative stress indicated by reduced peroxide activity caused by unconjugated bilirubin (UCB). Further investigations through animal studies demonstrated that DHA effectively mitigates bilirubin-induced brain injury in neonatal rats. However, the potential of DHA to decrease the incidence of bilirubin-induced brain damage in clinical settings has not been previously explored or reported. Infants with neonatal hyperbilirubinemia (n = 30 per group) participated in a double-blind, randomized, placebo-controlled parallel study. They received either 100 mg/d DHA or placebo syrup immediately when they were diagnosed. The study found that the bilirubin level at 48 hours of treatment, serum neuron-specific enolase (NSE) levels, mean phototherapy duration, and abnormal rate of cranial magnetic resonance imaging (MRI) were lower in the DHA group than those in the control group (P < .05). These results suggested that DHA is effective as an adjuvant treatment for hyperbilirubinemia in children. It can reduce the incidence of neonatal hyperbilirubinemia brain injury and plays a certain protective role. Clinical study on protective effect of DHA on neonatal bilirubin injury is registered at Chinese Clinical Trial Registry as ChiCTR2300070250.

Neuroprotective efficacy of hypothermia and Inter-alpha Inhibitor Proteins after hypoxic ischemic brain injury in neonatal rats

Therapeutic hypothermia is the standard of care for hypoxic-ischemic (HI) encephalopathy. Inter-alpha Inhibitor Proteins (IAIPs) attenuate brain injury after HI in neonatal rats. Human (h) IAIPs (60 ​mg/kg) or placebo (PL) were given 15 ​min, 24 and 48 ​h to postnatal (P) day-7 rats after carotid ligation and 8% oxygen for 90 ​min with (30 ​°C) and without (36 ​°C) exposure to hypothermia 1.5 ​h after HI for 3 ​h. Hemispheric volume atrophy (P14) and neurobehavioral tests including righting reflex (P8–P10), small open field (P13–P14), and negative geotaxis (P14) were determined. Hemispheric volume atrophy in males was reduced (P ​< ​0.05) by 41.9% in the normothermic-IAIP and 28.1% in the hypothermic-IAIP compared with the normothermic-PL group, and in females reduced (P ​< ​0.05) by 30.3% in the normothermic-IAIP, 45.7% in hypothermic-PL, and 55.2% in hypothermic-IAIP compared with the normothermic-PL group after HI. Hypothermia improved (P ​< ​0.05) the neuroprotective effects of hIAIPs in females. The neuroprotective efficacy of hIAIPs was comparable to hypothermia in female rats (P ​= ​0.183). Treatment with hIAIPs, hypothermia, and hIAIPs with hypothermia decreased (P ​< ​0.05) the latency to enter the peripheral zone in the small open field test in males. We conclude that hIAIPs provide neuroprotection from HI brain injury that is comparable to the protection by hypothermia, hypothermia increases the effects of hIAIPs in females, and hIAIPs and hypothermia exhibit some sex-related differential effects.

Blockade of connexin hemichannels with tonabersat protects against mild hypoxic ischemic brain injury in neonatal rats

Background and purposeThere is growing evidence that infants with mild hypoxic-ischemic (HI) encephalopathy have increased risk of brain injury and adverse neurodevelopmental outcomes. Currently, there is no approved treatment for these infants. It was previously shown that blocking connexin 43 hemichannels is neuroprotective in models of moderate to severe HI injury. However, it is yet to be established whether these channels play a role in the evolution of mild HI brain injury, and whether blocking these channels after mild HI is neuroprotective. MethodsHI was induced in postnatal day 10 rats of both sexes by right carotid artery ligation followed by 80 min of hypoxia in 8% oxygen. Pups receiving HI were randomised to receive intraperitoneal injections of either saline, vehicle (2-hydroxypropyl-beta-cyclodextrin polyethylene glycol-400), or tonabersat (2 mg/kg), at 60 min, 24 h, and 48 h after hypoxia. Seven days after HI, brains were harvested for measurement of volume loss and histological analysis. ResultsHI resulted in a significant reduction in hemispheric, hippocampal, and white matter volumes, which were significantly attenuated after treatment with tonabersat. HI was also associated with a significant reduction in numbers of neurons in the CA1 and CA3 hippocampal regions, a reduction in the numbers of oligodendrocytes in the corpus callosum, and an increase in the number of astrocytes in both regions, which were significantly attenuated by tonabersat treatment. There were no differences in rectal temperatures between tonabersat- and vehicle-treated rat pups. ConclusionsBlockade of connexin hemichannels with tonabersat significantly reduced mild HI injury in the hippocampus and white matter, without causing hypothermia.

Neuroserpin As an Adjuvant Therapy for Hypothermia on Brain Injury in Neonatal Hypoxic-Ischemic Rats.

We aimed to assess the effects of neuroserpin and its combination with hypothermia on hypoxic-ischemic (HI) brain injury in neonatal rats. Neuroserpin is an axon-secreted serine protease inhibitor and is important for brain development, neuronal survival, and synaptic plasticity. Male Wistar-Albino rats on postnatal day 7 (P7) were randomly divided into five groups: sham group (n = 10), (HI; n = 10), hypoxic-ischemic hypothermia (HIH; n = 10), hypoxic-ischemic neuroserpin (HIN; n = 10), and hypoxic-ischemic neuroserpin-hypothermia (HINH; n = 10). The P7 rat brain's maturation is similar to a late preterm human brain at 34 to 36 weeks of gestation. HI was induced in rats on P7 as previously described. A single dose of 0.2 µM neuroserpin (HINH and HIN) or an equivalent volume of phosphate-buffered saline (sham, HIH, and HI) was administered intraventricularly by a Hamilton syringe immediately after hypoxia. In the follow-up, pups were subjected to systemic hypothermia or normothermia for 2 hours. Euthanasia was performed for histopathological evaluation on P10. Apoptosis was detected by caspase-3 activity and terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) staining and was counted in the hippocampus. In comparison to the HI group, the TUNEL-positive and caspase-3-positive neurons in the sham, HIN, HIH, and HINH groups were considerably lower (13.4 ± 1.0 vs. 1.9 ± 0.9, 6.0 ± 0.9, 5.3 ± 1.6, and 4.0 ± 1.1; p < 0.001) and (13.5 ± 1.7 vs. 1.2 ± 0.7, 9.1 ± 2.7, 4.8 ± 1.0, and 3.9 ± 1.6; p < 0.001). HIN, HIH, and HINH, compared to the sham group, showed more TUNEL-positive and caspase-3-positive neurons (6.0 ± 0.9, 5.3 ± 1.6, 4.0 ± 1.1 vs. 1.9 ± 0.9 and 9.1 ± 2.7, 4.8 ± 1.0, 3.9 ± 1.6 vs. 1.2 ± 0.7; p < 0.001). The HINH group (synergistic effect) had significantly fewer TUNEL-positive neurons and caspase-3-positive neurons than the HIN group (4.0 ± 1.1 vs. 6.0 ± 0.9 and 3.9 ± 1.6 vs. 9.1 ± 2.7; p < 0.001). Our study showed that both neuroserpin alone and as an adjuvant treatment for hypothermia may have a neuroprotective effect on brain injury. · Neuroserpin decreased brain injury.. · Neuroserpin showed a synergistic effect when used as an adjuvant treatment for hypothermia.. · The neuroprotective effect of neuroserpine was related to its antiapoptotic properties..

Systemic administration of clinical-grade multilineage-differentiating stress-enduring cells ameliorates hypoxic–ischemic brain injury in neonatal rats

Multilineage-differentiating stress-enduring (Muse) cells are endogenous reparative pluripotent stem cells present in the bone marrow, peripheral blood, and organ connective tissues. We assessed the homing and therapeutic effects of systemically administered nafimestrocel, a clinical-grade human Muse cell-based product, without immunosuppressants in a neonatal hypoxic–ischemic (HI) rat model. HI injury was induced on postnatal day 7 (P7) and was confirmed by T2-weighted magnetic resonance imaging on P10. HI rats received a single dose nafimestrocel (1 × 106 cells/body) or Hank’s balanced salt solution (vehicle group) intravenously at either three days (on P10; M3 group) or seven days (on P14; M7 group) after HI insult. Radioisotope experiment demonstrated the homing of chromium-51-labeled nafimestrocel to the both cerebral hemispheres. The cylinder test (M3 and M7 groups) and open-field test (M7 group) showed significant amelioration of paralysis and hyperactivity at five weeks of age compared with those in the vehicle group. Nafimestrocel did not cause adverse events such as death or pathological changes in the lung at ten weeks in the both groups. Nafimestrocel attenuated the production of tumor necrosis factor-α and inducible nitric oxide synthase from activated cultured microglia in vitro. These results demonstrate the potential therapeutic benefits and safety of nafimestrocel.

Inter-Alpha Inhibitor Proteins Modify the Microvasculature after Exposure to Hypoxia-Ischemia and Hypoxia in Neonatal Rats.

Microvasculature develops during early brain development. Hypoxia-ischemia (HI) and hypoxia (H) predispose to brain injury in neonates. Inter-alpha inhibitor proteins (IAIPs) attenuate injury to the neonatal brain after exposure to HI. However, the effects of IAIPs on the brain microvasculature after exposure to HI have not been examined in neonates. Postnatal day-7 rats were exposed to sham treatment or right carotid artery ligation and 8% oxygen for 90 min. HI comprises hypoxia (H) and ischemia to the right hemisphere (HI-right) and hypoxia to the whole body, including the left hemisphere (H-left). Human IAIPs (hIAIPs, 30 mg/kg) or placebo were injected immediately, 24 and 48 h after HI/H. The brains were analyzed 72 h after HI/H to determine the effects of hIAIPs on the microvasculature by laminin immunohistochemistry and calculation of (1) the percentage area stained by laminin, (2) cumulative microvessel length, and (3) density of tunneling nanotubes (TNTs), which are sensitive indicators of the earliest phases of neo-vascularization/collateralization. hIAIPs mainly affected the percent of the laminin-stained area after HI/H, cumulative vessel length after H but not HI, and TNT density in females but not males. hIAIPs modify the effects of HI/H on the microvasculature after brain injury in neonatal rats and exhibit sex-related differential effects. Our findings suggest that treatment with hIAIPs after exposure to H and HI in neonatal rats affects the laminin content of the vessel basal lamina and angiogenic responses in a sex-related fashion.

Glutamine ameliorates hyperoxia-induced hippocampal damage by attenuating inflammation and apoptosis via the MKP-1/MAPK signaling pathway in neonatal rats.

Glutamine (Gln) is an immunomodulatory protein that mediates oxidative stress, inflammation, and apoptosis, but has not been reported in the treatment of hyperoxia (Hyp)-induced brain injury. The aim of this study was to determine whether Gln could improve hyp-induced brain injury in neonatal rats to and later learning and memory dysfunction, and to explore its possible mechanisms. We prepared a model of neonatal rat brain injury caused by normobaric hyperoxia while administered with Gln for 7days for evaluation. Learning memory function was assessed with the Morris water maze test. Histological analysis, protein expression analysis, oxidative stress and inflammation level analysis were performed using hippocampal tissue. Gln treatment significantly reduced brain tissue water content, oxidative stress levels, microglia activation and inflammatory factor expression, and attenuated tissue damage and apoptosis in the hippocampal region. Gln ameliorates hyp-induced learning, memory impairment in neonatal rats in water maze test. It also increased MKP-1 protein expression and decreased p-p38, p-ERK and p-JNK. Therefore, it is hypothesized that Gln may exert neuroprotective effects by increasing MKP-1 expression to negatively regulate MAPK signaling, with potential cognitive improvement in hyp-induced brain injury.

Reperfusion after hypoxia-ischemia exacerbates brain injury with compensatory activation of the anti- ferroptosis system: based on a novel rat model.

Hypoxic-ischemic encephalopathy, which predisposes to neonatal death and neurological sequelae, has a high morbidity, but there is still a lack of effective prevention and treatment in clinical practice. To better understand the pathophysiological mechanism underlying hypoxic-ischemic encephalopathy, in this study we compared hypoxic-ischemic reperfusion brain injury and simple hypoxic-ischemic brain injury in neonatal rats. First, based on the conventional Rice-Vannucci model of hypoxic-ischemic encephalopathy, we established a rat model of hypoxic-ischemic reperfusion brain injury by creating a common carotid artery muscle bridge. Then we performed tandem mass tag-based proteomic analysis to identify differentially expressed proteins between the hypoxic-ischemic reperfusion brain injury model and the conventional Rice-Vannucci model and found that the majority were mitochondrial proteins. We also performed transmission electron microscopy and found typical characteristics of ferroptosis, including mitochondrial shrinkage, ruptured mitochondrial membranes, and reduced or absent mitochondrial cristae. Further, both rat models showed high levels of glial fibrillary acidic protein and low levels of myelin basic protein, which are biological indicators of hypoxic-ischemic brain injury and indicate similar degrees of damage. Finally, we found that ferroptosis-related Ferritin (Fth1) and glutathione peroxidase 4 were expressed at higher levels in the brain tissue of rats with hypoxic-ischemic reperfusion brain injury than in rats with simple hypoxic-ischemic brain injury. Based on these results, it appears that the rat model of hypoxic-ischemic reperfusion brain injury is more closely related to the pathophysiology of clinical reperfusion. Reperfusion not only aggravates hypoxic-ischemic brain injury but also activates the anti-ferroptosis system.

Effects of Three Different Doses of Inter-Alpha Inhibitor Proteins on Severe Hypoxia-Ischemia-Related Brain Injury in Neonatal Rats.

Hypoxia-ischemia (HI)-related brain injury is an important cause of morbidity and long-standing disability in newborns. We have previously shown that human plasma-derived inter-alpha inhibitor proteins (hIAIPs) attenuate HI-related brain injury in neonatal rats. The optimal dose of hIAIPs for their neuroprotective effects and improvement in behavioral outcomes remains to be determined. We examined the efficacy of 30, 60, or 90 mg/kg of hIAIPs administered to neonatal rats after exposure to HI for 2 h. Postnatal day 7 (P7) Wistar rats were exposed to either sham-surgery or unilateral HI (right carotid artery ligation, 2 h of 8% O2) brain injury. A placebo, 30, 60, or 90 mg/kg of hIAIPs were injected intraperitoneally at 0, 24 and 48 h after HI (n = 9-10/sex). We carried out the following behavioral analyses: P8 (righting reflex), P9 (negative geotaxis) and P10 (open-field task). Rats were humanely killed on P10 and their brains were stained with cresyl violet. Male extension/contraction responses and female righting reflex times were higher in the HI placebo groups than the sham groups. Female open-field exploration was lower in the HI placebo group than the sham group. hIAIPs attenuated these behavioral deficits. However, the magnitude of the responses did not vary by hIAIP dose. hIAIPs reduced male brain infarct volumes in a manner that correlated with improved behavioral outcomes. Increasing the hIAIP dose from 30 to 90 mg/kg did not further accentuate the hIAIP-related decreases in infarct volumes. We conclude that larger doses of hIAIPs did not provide additional benefits over the 30 mg/kg dose for behavior tasks or reductions in infarct volumes in neonatal rats after exposure to severe HI.

Effect of Hypoxic-Ischemic Brain Injury in Neonatal Rats on Behavioral Parameters and Expression of CDK8 in the Brain Tissue.

Behavioral changes in newborn 3-day-old rats (n=44) with modeled hypoxic-ischemic brain injury (HIBI) were observed, and the expression of CDK8 in brain tissues was detected to clarify the significance of CDK8. In 30 min, 3 h, and 3 days after HIBI, the left (ischemic) hemisphere was taken for examination. In 3 days after HIBI, the rat pups were examined in the behavioral tests. In rat pups with HIBI, changes of CDK8 expression were detected by Western blotting and real-time PCR and changes in the righting reflex and forelimb grip strength test (p<0.05) were revealed in comparison with sham-operated animals. The expression of CDK8 increased 30 min after HIBI and decreased in 3 h and 3 days. Hypoxia and ischemia of the left brain may affect locomotion, but not sensation. Since CDK8 is involved in the immune response after cerebral hypoxia and ischemia, this kinase can be used as an early diagnostic index.

Down-regulation of miR-204-5p ameliorates sevoflurane-induced brain injury in neonatal rats through targeting VCAM1

To confirm the regulation of miR-204-5p on VCAM1 and its effect on sevoflurane-induced brain injury in neonatal rats. We adopted the sevoflurane-induced brain injury model, and the double luciferase reporter gene assay was applied to explore the targeting relationship between vascular adhesion factor 1 (VCAM1) and miR-204-5p. RT-qPCR was applied to assess the levels of miR-204-5. VCAM1, LC3, P62 and cleaved-caspase 3 levels in the hippocampus were estimated by western blot. The number of autophagosomes in the cerebral cortex was assessed via Transmission electron microscopy (TEM), and histopathological changes within the hippocampus by HE staining. miR-204-5p levels were remarkably increased, but VCAM1 expression was decreased after neonatal rat brain injury. Furthermore, miR-204-5p directly targeted VCAM1. The escape latency, swimming distance, autophagosome number, neuronal apoptosis ratio, LC3 II and Cleaved-caspase 3 expression were reduced after miR-204-5p inhibition interference, whereas crossing times, and P62 expression increased in the sevoflurane-induced brain injury model. Furthermore, down-regulation of VCAM1 reversed the trend of these indices. These results suggest that down-regulation of miR-204-5p ameliorates sevoflurane-induced cognitive impairment and hippocampal pathology and inhibits neuronal autophagy and apoptosis by targeting VCAM1.

Antiapoptotic Effects of Hydroxychloroquine on Hypoxic-Ischemic Injury in Neonatal Rat Brain: May Hydroxychloroquine Be an Adjuvant Theraphy?

Hydroxychloroquine (HCQ) has immunomodulatory, antithrombotic, cardiovascular, antimicrobial, and antineoplastic effects. In this study, we aimed to investigate the antiapoptotic and immunomodulator effects of intraperitoneal HCQ on hypoxic-ischemic (HI) injury in newborn rats. Wistar albino rats, 7 to 10 days old, were randomly divided into three groups: hypoxic-ischemic encephalopathy (HIE) group, HIE treated with HCQ group, and Sham group. Left common carotid artery ligation and hypoxia model were performed in HIE and HCQ groups. The HCQ group was treated with 80 mg/kg intraperitoneal HCQ every 24 hours for 3 days, while Sham and HIE groups were given physiological saline. After 72 hours, rats were decapitated and brain tissues were stained with hematoxylin and eosin, TUNEL, and IL-1β for histopathological grading and neuronal cell injury. Neuronal apoptosis was statistically lower in all neuroanatomical areas in the HCQ group compared with the HIE group. IL-1β-stained areas were similar in both HCQ and HIE groups but significantly higher compared with the Sham group. Histopathological grading scores were found to be lower in the HCQ group on the left parietal cortex and hippocampus region. In this study, we have shown for the first time that HCQ treatment decreased apoptosis in HI newborn rat model in both hemispheres. HCQ may be a promising adjuvant therapy in neonatal HIE. · HCQ decreased neuronal apoptosis in the ischemic penumbra of the rat brain.. · HCQ attenuates hypoxia-ischemia-induced brain injury in neonatal rats.. · HCQ has no anti-inflammatory effect on HI injury..

Chlorogenic acid exerts neuroprotective effect against hypoxia-ischemia brain injury in neonatal rats by activating Sirt1 to regulate the Nrf2-NF-κB signaling pathway

BackgroundNeonatal hypoxic-ischemic brain injury (HIE) is caused by perinatal asphyxia, which is associated with various confounding factors. Although studies on the pathogenesis and treatment of HIE have matured, sub-hypothermia is the only clinical treatment available for HIE. Previous evidence indicates that chlorogenic acid (CGA) exerts a potential neuroprotective effect on brain injury. However, the role of CGA on neonatal HI brain damage and the exact mechanism remains elusive.Here, we investigate the effects of CGA on HI models in vivo and in vitro and explore the underlying mechanism.MethodsIn the in vivo experiment, we ligated the left common carotid artery of 7-day-old rats and placed the rats in a hypoxic box for 2 h. We did not ligate the common carotid artery of the pups in the sham group since they did not have hypoxia. Brain atrophy and infarct size were evaluated by Nissl staining, HE staining and 2,3,5-triphenyltetrazolium chloride monohydrate (TTC) staining. Morris Water Maze test (MWM) was used to evaluate neurobehavioral disorders. Western-blotting and immunofluorescence were used to detect the cell signaling pathway. Malondialdehyde (MDA) content test, catalase (CAT) activity detection and Elisa Assay was used to detect levels of inflammation and oxidative stress. in vitro experiments were performed on isolated primary neurons.ResultIn our study, pretreatment with CGA significantly decreased the infarct volume of neonatal rats after HI, alleviated brain edema, and improved tissue structure in vivo. Moreover, we used the Morris water maze to verify CGA’s effects on enhancing the learning and cognitive ability and helping to maintain the long-term spatial memory after HI injury. However, Sirt1 inhibitor EX-527 partially reversed these therapeutic effects. CGA pretreatment inhibited neuronal apoptosis induced by HI by reducing inflammation and oxidative stress. The findings suggest that CGA potentially activates Sirt1 to regulate the Nrf2-NF-κB signaling pathway by forming complexes thereby protecting primary neurons from oxygen-glucose deprivation (OGD) damage. Also, CGA treatment significantly suppresses HI-induced proliferation of glial.ConclusionCollectively, this study uncovered the underlying mechanism of CGA on neonatal HI brain damage. CGA holds promise as an effective neuroprotective agent to promote neonatal brain recovery from HI-induced injury.Graphical 8NePktn-RrRjrUAHAHCXyYVideo

Time Course of Changes in the Neurovascular Unit after Hypoxic-Ischemic Injury in Neonatal Rats.

Exposure to hypoxic-ischemic (HI) insults in newborns can predispose them to severe neurological sequela. The mechanisms underlying HI-related brain injury have not been completely elucidated. The neurovascular unit (NVU) is a composite of structures that protect the brain from the influx of detrimental molecules. Changes in the NVU after HI are important because they could reveal endogenous neuroprotective pathways in the cerebral microvasculature. Furthermore, the time course of changes in the NVU after exposure to HI in the newborn remains to be determined. In this study, we examined the effects of severe HI on the time course of changes in the NVU in neonatal rats. Brains were collected from rats exposed to right carotid artery ligation and 2 h of hypoxia on postnatal day 7 with recovery for 6 or 48 h after exposure to sham treatment (Sham) or HI. The right HI and left hypoxic alone sides of the brains were examined by quantitative immunohistochemistry for vascular density (laminin), pericyte vascular coverage (PDGFRβ), astrocyte vascular coverage (GFAP), and claudin-5 expression in the microvasculature of the cerebral cortex, white matter, and hippocampus. HI-related brain injury in neonatal rats was associated with increases in vascular density in the cortex and hippocampus 48 h after HI as well as neurovascular remodeling, including loss of pericyte coverage in the cortex and increases in claudin-5 in the hippocampus 6 h after HI. Astrocyte coverage was not affected by HI injury. The time course of the responses in the different components of the NVU varied after exposure to HI. There were also differential regional responses in the elements of the NVU in response to HI and hypoxia alone.

Atorvastatin inhibits neuronal apoptosis via activating cAMP/PKA/p-CREB/BDNF pathway in hypoxic-ischemic neonatal rats.

Neuronal apoptosis is one of the main pathological processes of hypoxic-ischemic brain damage (HIBD) and is involved in the development of hypoxic-ischemic encephalopathy (HIE) in neonates. Atorvastatin has been found to have neuroprotective effects in some nervous system diseases, but its role in regulating the pathogenesis of neonatal HIBD remains elusive. Thus, this study aimed to explore the effects and related mechanisms of atorvastatin on the regulation of neuronal apoptosis after HIBD in newborn rats. The rat HIBD model and the neuronal oxygen glucose deprivation (OGD) model were established routinely. Atorvastatin, cAMP inhibitor (SQ22536), and BDNF inhibitor (ANA-12) were used to treat HIBD rats and OGD neurons. Cerebral infarction, learning and memory ability, cAMP/PKA/p-CREB/BDNF signaling molecules, and apoptosis-related indicators (TUNEL, cleaved caspase-3, and Bax/Bcl2) were then examined. In vivo, atorvastatin reduced cerebral infarction, improved learning and memory ability, decreased the number of TUNEL-positive neurons, inhibited the expression of cleaved caspase-3 and Bax/Bcl2, and activated the cAMP/PKA/p-CREB/BDNF pathway in the cerebral cortex after HIBD. In vitro, atorvastatin also decreased the apoptosis-related indicators and activated the cAMP/PKA/p-CREB/BDNF pathway in neurons after OGD. Furthermore, inhibition of cAMP or BDNF attenuated the effect of atorvastatin on the reduction of neuronal apoptosis, suggesting that atorvastatin inhibits HIBD-induced neuronal apoptosis and alleviates brain injury in neonatal rats mainly by activating the cAMP/PKA/p-CREB/BDNF pathway. In conclusion, atorvastatin may be developed as a potential drug for the treatment of neonatal HIE.

Inter-alpha Inhibitor Proteins Ameliorate Brain Injury and Improve Behavioral Outcomes in a Sex-Dependent Manner After Exposure to Neonatal Hypoxia Ischemia in Newborn and Young Adult Rats.

Hypoxic-ischemic (HI) brain injury is a major contributor to neurodevelopmental morbidities. Inter-alpha inhibitor proteins (IAIPs) have neuroprotective effects on HI-related brain injury in neonatal rats. However, the effects of treatment with IAIPs on sequential behavioral, MRI, and histopathological abnormalities in the young adult brain after treatment with IAIPs in neonates remain to be determined. The objective of this study was to examine the neuroprotective effects of IAIPs at different neurodevelopmental stages from newborn to young adults after exposure of neonates to HI injury. IAIPs were given as 11-sequential 30-mg/kg doses to postnatal (P) day 7-21 rats after right common carotid artery ligation and exposure to 90min of 8% oxygen. The resulting brain edema and injury were examined by T2-weighted magnetic resonance imaging (MRI) and cresyl violet staining, respectively. The mean T2 values of the ipsilateral hemisphere from MRI slices 6 to 10 were reduced in IAIP-treated HI males + females on P8, P9, and P10 and females on P8, P9, P10, and P14. IAIP treatment reduced hemispheric volume atrophy by 44.5 ± 29.7% in adult male + female P42 rats and improved general locomotor abilities measured by the righting reflex over time at P7.5, P8, and P9 in males + females and males and muscle strength/endurance measured by wire hang on P16 in males + females and females. IAIPs provided beneficial effects during the learning phase of the Morris water maze with females exhibiting beneficial effects. IAIPs confer neuroprotection from HI-related brain injury in neonates and even in adult rats and beneficial MRI and behavioral benefits in a sex-dependent manner.

Asiatic acid exerts neuroprotective effect against hypoxicischemic brain injury in neonatal rats via inhibition of oxidative damage

Purpose: To investigate the effect of asiatic acid on hypoxic ischemia-induced injury in neonatal rats, and the underlying mechanism of action.Methods: Hypoxic-ischemia (HI) neonatal rat model was established via permanent ligation of the carotid artery, followed by hypoxia (exposure to 8 % oxygen and 92 % nitrogen) for 24 h. Immunofluorescence, using fluorescence microscope, was used for the determination of expressions of p-TAK1, NeuN and GFAP. Western blotting was used for assaying protein expression levels, while TUNEL assay was employed for the measurement of apoptosis.Results: Treatment of rats with asiatic acid prior to HI effectively prevented up-regulation of pTAK1 and decreased the count of p-TAK1-containing astrocytes. The proportion of NeuN containing p-TAK1 in HI rat brain cortex was significantly reduced by asiatic acid (p &lt; 0.05). Treatment of rats with asiatic acid suppressed HI- induced up-regulation of pJNK expression. The HI-induced increase in the expression levels of caspase-3, p53 and p-c-Jun in rat brain cortex were reversed by asiatic acid (p &lt; 0.05). The HImediated up-regulation of expressions of p- JNK, caspase-3, p53 and p-c-Jun in rat brain cortex were inhibited significantly by NG25. Asiatic acid treatment also significantly alleviated HI-mediated increase in apoptosis of neurons in rat brain cortex, when compared to model group (p &lt; 0.05).Conclusion: These findings suggest that asiatic acid prevents HI-induced brain injury in neonatal rats via inhibition of neuronal apoptosis. Moreover, it inhibits TAK1 activation, suppresses p-JNK expression and targets pro-apoptotic factors in brain cortex. Therefore, asiatic acid may be a therapeutic agent for the management of HI-induced brain injury.

Azithromycin reduces inflammation-amplified hypoxic-ischemic brain injury in neonatal rats.

Background:Systemic inflammation amplifies neonatal hypoxic-ischemic (HI) brain injury. Azithromycin (AZ), an antibiotic with anti-inflammatory properties, improves sensorimotor function and reduces tissue damage after neonatal rat HI brain injury.Objective:To determine if AZ is neuroprotective in two neonatal rat models of inflammation-amplified HI brain injury.Design/Methods:Seven-day-old (P7) rats received injections of toll-like receptor agonists lipopolysaccharide (LPS) or Pam3Cys-Ser-(Lys)4 (PAM) prior to right carotid ligation followed by 50 min (LPS+HI) or 60 min (PAM+HI) in 8% oxygen. Outcomes included contralateral forelimb function (forepaw placing; grip strength), survival, %intact right hemisphere (brain damage), and a Composite Score incorporating these measures. We compared P35 outcomes in controls and groups treated with 3 or 5 AZ doses. Then, we compared P21 outcomes when the first (of 5) AZ doses was administered 1, 2, or 4 hours after HI.Results:In both LPS+HI and PAM+HI models, AZ improved sensorimotor function, survival, brain tissue preservation, and Composite Scores. Benefits increased with 5 vs. 3-dose AZ and declined with longer initiation delay.Conclusion:Perinatal systemic infection is a common comorbidity of neonatal asphyxia brain injury and contributes to adverse outcomes. These data support further evaluation of AZ as a candidate treatment for neonatal neuroprotection.