Alpha-pinene ameliorate behavioral deficit induced by early postnatal hypoxia in the rat: study the inflammatory mechanism.

Neonatal hypoxia has a negative impact on the developing brain during the sensitive period. Inflammation plays a key role in the physiological response to hypoxic stress. Considering the anti-inflammatory properties of alpha-pinene, which has received a lot of attention in recent years, in this research we focused on the impact of alpha-pinene on the behavioral responses and proinflammatory factors in rats subjected to the neonatal hypoxia. This study involved Wistar rats (7-day-old) that were divided into six experimental groups, including a control group, groups receiving different doses of alpha-pinene (5 and 10 mg/kg), a hypoxia group receiving 7% O2 and 93% N2, 90 min duration for 7 days, and groups receiving alpha-pinene 30 min before hypoxia. All injections were done intraperitoneally. The rats were evaluated for proinflammatory factors 24 h after exposure to hypoxia (PND14) and at the end of the behavioral test (PND54). The results showed that hypoxia led to decreased motor activity, coordination, and memory, as well as increased inflammation. However, the rats that received alpha-pinene showed improved behavioral responses and reduced inflammation compared to the hypoxia group (all cases p < 0.05). This suggests that alpha-pinene may have a protective effect via anti-inflammatory properties against the negative impacts of hypoxia on the developing brain.

Intestinal microbiota modulates neuroinflammatory response and brain injury after neonatal hypoxia-ischemia.

Premature infants lack a normal intestinal microbial community and also at risk of perinatal hypoxic-ischemic (HI) brain injury, which is considered to be one of the major factors for motor, sensory, and cognitive deficits. We hypothesized that neonatal gut microbiota composition modulated the immune reaction and severity of neonatal H-I brain injury. Neonatal C57BL/6J mouse pups were exposed to H-I protocol consisting of permanent left carotid artery ligation, followed by 8% hypoxia for 60 min. Microbial manipulation groups included 1) antibiotic treatment, E18 (maternal) to P5; 2) antibiotic treatment E18 to P5 + E. coli gavage; 3) antibiotic treatment E18 to P5 + B. infantis gavage; and 4) saline to pups with dams getting fresh water. The extent of brain injury and recovery was measured on MRI. Edematous injury volume was significantly higher in E. coli group than that in B. infantis group and in fresh water group. Gene expression in brains of pro-inflammatory cytokines (IL1ß, IL6, IL2, TNF-α and toll-like receptors 2-6) were elevated to a greater extent in the E. coli group at P10, no injury, and at P13, 72 hours after H-I relative to sham control and B. infantis groups. Significant effects of microbiome and brain injury and interaction of these factors were found in abundance of major phyla. The neuroinflammatory response and brain injury after neonatal hypoxia-ischemia are affected by intestinal microbiota, providing opportunities for therapeutic intervention through targeting the early colonization and development of the gut microbiota.

Epilepsy Incidence and Developmental Outcomes After Early Discontinuation of Antiseizure Medication in Neonatal Hypoxic-Ischemic Encephalopathy.

BACKGROUND: Neonatal seizures caused by hypoxic-ischemic encephalopathy (HIE) have significant morbidity and mortality. There is variability in clinical practice regarding treatment duration with antiseizure medication (ASM) after resolution of provoked neonatal seizures. We examined epilepsy incidence and developmental outcomes in post-HIE neonates discharged or not on ASM. METHODS: We conducted a retrospective chart review of all HIE-admitted neonates to the University of Iowa Hospitals & Clinics neonatal intensive care unit between January 2008 and February 2021 who presented with encephalopathy, underwent therapeutic hypothermia, and developed seizures. Neonates were divided into two groups depending on whether ASM was continued or discontinued on discharge. We evaluated the incidence of epilepsy and developmental outcomes on follow-up in these two cohorts up to 12 months. RESULTS: Sixty-nine neonates met the study criteria. ASM was continued on discharge in 41 neonates (59%) and discontinued before discharge in 28 (41%). At the 12-month follow-up, nine neonates (13%) had a diagnosis of epilepsy, out of which seven neonates had ASM continued on discharge (odds ratio [OR]: 2.84; 95% confidence interval [CI]: 0.48, 29.9)]. There was no statistical difference between the development of postneonatal epilepsy between the two groups (P value 0.29). There was no significant difference in developmental outcome between the two groups after adjusting for covariates like magnetic resonance imaging (MRI) brain abnormality and number of seizure days (OR: 0.68; 95% CI: 0.21, 2.22; P = 0.52). CONCLUSION: We found no significant risk of seizure recurrence by age 12 months in infants who had discontinued ASM before discharge compared with those who had continued ASM. There was no difference in developmental outcomes at the 12-month follow-up between groups after adjusting for brain MRI abnormality and the number of seizure days during admission. Our results support early discontinuation of ASM after resolution of acute provoked seizures in neonates with HIE.

Catalpol alleviates hypoxia ischemia-induced brain damage by inhibiting ferroptosis through the PI3K/NRF2/system Xc-/GPX4 axis in neonatal rats.

Hypoxic-ischemic encephalopathy (HIE) is a brain damage caused by perinatal hypoxia and blood flow reduction. Severe HIE leads to death. Available treatments remain limited. Oxidative stress and nerve damage are major factors in brain injury caused by HIE. Catalpol, an iridoid glucoside found in the root of Rehmannia glutinosa, has antioxidant and neuroprotective effects. This study examined the neuroprotective effects of catalpol using a neonatal rat HIE model and found that catalpol might protect the brain through inhibiting neuronal ferroptosis and ameliorating oxidative stress. Behavior tests suggested that catalpol treatment improved functions of motor, learning, and memory abilities after hypoxic-ischemic injury. Catalpol treatment inhibited changes to several ferroptosis-related proteins, including p-PI3K, p-AKT, NRF2, GPX4, SLC7A11, SLC3A2, GCLC, and GSS in HIE neonatal rats. Catalpol also prevented changes to several ferroptosis-related proteins in PC12 cells after oxygen-glucose deprivation. The ferroptosis inducer erastin reversed the protective effects of catalpol both in vitro and in vivo. We concluded that catalpol protects against hypoxic-ischemic brain damage (HIBD) by inhibiting ferroptosis through the PI3K/NRF2/system Xc-/GPX4 axis.

Relationship of Neonatal Seizure Burden Before Treatment and Response to Initial Antiseizure Medication.

OBJECTIVE: To assess among a cohort of neonates with hypoxic-ischemic encephalopathy (HIE) the association of pretreatment maximal hourly seizure burden and total seizure duration with successful response to initial antiseizure medication (ASM). STUDY DESIGN: This was a retrospective review of data collected from infants enrolled in the HEAL Trial (NCT02811263) between January 25, 2017, and October 9, 2019. We evaluated a cohort of neonates born at ≥36 weeks of gestation with moderate-to-severe HIE who underwent continuous electroencephalogram monitoring and had acute symptomatic seizures. Poisson regression analyzed associations between (1) pretreatment maximal hourly seizure burden, (2) pretreatment total seizure duration, (3) time from first seizure to initial ASM, and (4) successful response to initial ASM. RESULTS: Among 39 neonates meeting inclusion criteria, greater pretreatment maximal hourly seizure burden was associated with lower chance of successful response to initial ASM (adjusted relative risk for each 5-minute increase in seizure burden 0.83, 95% CI 0.69-0.99). There was no association between pretreatment total seizure duration and chance of successful response. Shorter time-to-treatment was paradoxically associated with lower chance of successful response to treatment, although this difference was small in magnitude (relative risk 1.007, 95% CI 1.003-1.010). CONCLUSIONS: Maximal seizure burden may be more important than other, more commonly used measures in predicting response to acute seizure treatments.

High Doses of ANA12 Improve Phenobarbital Efficacy in a Model of Neonatal Post-Ischemic Seizures.

Phenobarbital (PB) remains the first-line medication for neonatal seizures. Yet, seizures in many newborns, particularly those associated with perinatal ischemia, are resistant to PB. Previous animal studies have shown that in postnatal day P7 mice pups with ischemic stroke induced by unilateral carotid ligation, the tyrosine receptor kinase B (TrkB) antagonist ANA12 (N-[2-[[(hexahydro-2-oxo-1H-azepin-3-yl)amino]carbonyl]phenyl]-benzo[b]thiophene-2-carboxamide, 5 mg/kg) improved the efficacy of PB in reducing seizure occurrence. To meet optimal standards of effectiveness, a wider range of ANA12 doses must be tested. Here, using the unilateral carotid ligation model, we tested the effectiveness of higher doses of ANA12 (10 and 20 mg/kg) on the ability of PB to reduce seizure burden, ameliorate cell death (assessed by Fluoro-Jade staining), and affect neurodevelopment (righting reflex, negative geotaxis test, open field test). We found that a single dose of ANA12 (10 or 20 mg/kg) given 1 h after unilateral carotid ligation in P7 pups reduced seizure burden and neocortical and striatal neuron death without impairing developmental reflexes. In conclusion, ANA12 at a range of doses (10-20 mg/kg) enhanced PB effectiveness for the treatment of perinatal ischemia-related seizures, suggesting that this agent might be a clinically safe and effective adjunctive agent for the treatment of pharmacoresistant neonatal seizures.

GSK-3ß inhibitor TWS119 promotes neuronal differentiation after hypoxic-ischemic brain damage in neonatal rats.

Brain injury in preterm infants is a major cause of disability and mortality in children. GSK-3ß is a common pathogenic factor for cognitive dysfunction and involves in neuronal proliferation and differentiation. However, GSK-3ß affected neuronal differentiation and its molecular pathogenesis after hypoxic-ischemic brain damage in neonatal rats remains unclear. This study investigated the effects of GSK-3ß inhibitor (TWS119) on cell cycle regulatory proteins, a neuronal differentiation factor (CEND1), maturation neurons, T-box brain transcription factor 1 (TBR1)-positive neurons to clarify the mechanisms of hypoxic-ischemic brain damage in neonatal rats. We used hypoxic-ischemic Sprague-Dawley neonatal rats with brain damage as models. These rats were used for investigating the effect of GSK-3ß on cell cycle regulatory proteins, neuronal differentiation factor (CEND1), maturation neurons, TBR1-positive neurons by western blot and immunofluorescence. Cyclin D1 (a positive cell cycle regulator) expression decreased, and p21 (a negative cell cycle regulator) expression increased in the TWS119 group compared to the hypoxia-ischemia (HI) group 7 days after HI. Additionally, compared to the HI group, TWS119 treatment up-regulated CEND1 expression and promoted neuronal differentiation and cortex development based on NeuN and TBR1 expression. Our study suggests that the GSK-3ß inhibitor TWS119 promotes neuronal differentiation after hypoxic-ischemic brain damage in neonatal rats by inhibiting cell cycle pathway.

Evaluating the Safety and Efficacy of Erythropoietin Therapy for Neonatal Hypoxic-Ischemic Encephalopathy: A Systematic Review and Meta-Analysis.

BACKGROUND: Erythropoietin (EPO) is a proposed drug for the treatment of neonatal hypoxic-ischemic encephalopathy (HIE). Multiple studies have linked its use, either as a monotherapy or in conjunction with therapeutic hypothermia (TH), with improved neonatal outcomes including death and neurodisability. However, there is also evidence in the literature that raises concerns about its efficacy and safety for the treatment of neonatal encephalopathy (NE). METHODS: We searched MEDLINE, Cochrane CENTRAL, and Embase for both observational studies and randomized controlled trials (RCTs) investigating the effectiveness of EPO in treating NE. Only studies in which at least 300 U/kg of EPO was used and reported any one of the following outcomes: death, death or neurodisability, and cerebral palsy, were included. RESULTS: Seven studies with 903 infants with the diagnosis of NE were included in our meta-analysis. EPO did not reduce the risk of death or neurodisability (risk ratio 0.68 [95% confidence interval [CI]: 0.43 to 1.09]) (P = 0.11). Similarly, the risk of cerebral palsy was not reduced by the administration of EPO (risk ratio 0.68 [95% CI: 0.33 to 1.40]) (P = 0.30). The risk of death was also not reduced at any dose of EPO regardless of the use of TH. CONCLUSIONS: The results of our meta-analysis do not support the use of EPO for the treatment of neonatal encephalopathy. However, future large-scale RCTs are needed to strengthen these findings.

TrkB-mediated sustained neuroprotection is sex-specific and Erα-dependent in adult mice following neonatal hypoxia ischemia.

BACKGROUND: Neonatal hypoxia ischemia (HI) related brain injury is one of the major causes of life-long neurological morbidities that result in learning and memory impairments. Evidence suggests that male neonates are more susceptible to the detrimental effects of HI, yet the mechanisms mediating these sex-specific responses to neural injury in neonates remain poorly understood. We previously tested the effects of treatment with a small molecule agonist of the tyrosine kinase B receptor (TrkB), 7,8-dihydroxyflavone (DHF) following neonatal HI and determined that females, but not males exhibit increased phosphorylation of TrkB and reduced apoptosis in their hippocampi. Moreover, these female-specific effects of the TrkB agonist were found to be dependent upon the expression of Erα. These findings demonstrated that TrkB activation in the presence of Erα comprises one pathway by which neuroprotection may be conferred in a female-specific manner. The goal of this study was to determine the role of Erα-dependent TrkB-mediated neuroprotection in memory and anxiety in young adult mice exposed to HI during the neonatal period. METHODS: In this study, we used a unilateral hypoxic ischemic (HI) mouse model. Erα+/+ or Erα-/- mice were subjected to HI on postnatal day (P) 9 and mice were treated with either vehicle control or the TrkB agonist, DHF, for 7 days following HI. When mice reached young adulthood, we used the novel object recognition, novel object location and open field tests to assess long-term memory and anxiety-like behavior. The brains were then assessed for tissue damage using immunohistochemistry. RESULTS: Neonatal DHF treatment prevented HI-induced decrements in recognition and location memory in adulthood in females, but not in males. This protective effect was absent in female mice lacking Erα. The female-specific improved recognition and location memory outcomes in adulthood conferred by DHF therapy after neonatal HI tended to be or were Erα-dependent, respectively. Interestingly, DHF triggered anxiety-like behavior in both sexes only in the mice that lacked Erα. When we assessed the severity of injury, we found that DHF therapy did not decrease the percent tissue loss in proportion to functional recovery. We additionally observed that the presence of Erα significantly reduced overall HI-associated mortality in both sexes. CONCLUSIONS: These observations provide evidence for a therapeutic role for DHF in which TrkB-mediated sustained recovery of recognition and location memories in females are Erα-associated and dependent, respectively. However, the beneficial effects of DHF therapy did not include reduction of gross tissue loss but may be derived from the enhanced functioning of residual tissues in a cell-specific manner.

Periods of low oxygen delivery and blood flow to the brains of newborns are known to cause life-long impairments to their cognitive ability as adults. Interestingly, male newborns are more susceptible to this injury than females. The mechanisms causing this sex difference are poorly understood. Here we test the role of the nerve growth factor receptor tyrosine kinase B (TrkB) in providing long-term neuroprotection following neonatal hypoxia­ischemia (HI) in mice. We have previously shown that when mice are treated with the TrkB agonist 7,8-dihydroxyflavone (DHF) in the days following neonatal HI, the result is short-term neuroprotection only in females and this protection is dependent on the presence of the estrogen receptor alpha receptor ([Formula: see text]). In this study, we extend these observations by subjecting mice either with or without [Formula: see text] to HI. Some of the mice were then treated with DHF immediately after HI. As adults, we performed tests to assess the mice's memory and anxiety-like behavior. At the end of these tests, we assessed the brains for tissue loss. Our results show that as adults the DHF treatment following HI in neonatal mice preserved memory only in females and this effect was dependent on the presence of [Formula: see text]. In addition, DHF therapy triggered anxiety-like behavior in mice lacking [Formula: see text]. We also show that this neuroprotection is not dependent on preservation of brain tissue following the injury. These results provide insight into the mechanisms behind the female resistance to hypoxic ischemic episodes as newborns.

Calpain as a Therapeutic Target for Hypoxic-Ischemic Encephalopathy.

Hypoxic-ischemic encephalopathy (HIE) is a complex pathophysiological process with multiple links and factors. It involves the interaction of inflammation, oxidative stress, and glucose metabolism, and results in acute and even long-term brain damage and impairment of brain function. Calpain is a family of Ca2+-dependent cysteine proteases that regulate cellular function. Calpain activation is involved in cerebral ischemic injury, and this involvement is achieved by the interaction among Ca2+, substrates, organelles, and multiple proteases in the neuronal necrosis and apoptosis pathways after cerebral ischemia. Many calpain inhibitors have been developed and tested in the biochemical and biomedical fields. This study reviewed the potential role of calpain in the treatment of HIE and related mechanism, providing new insights for future research on HIE.

Long-term effect of indomethacin on a rat model of neonatal hypoxia ischemic encephalopathy through behavioral tests.

BACKGROUND: Many medical experts prescribe indomethacin because of its anti-inflammatory, analgesic, tocolytic, and duct closure effects. This article presents an evaluation of the enduring impact of indomethacin on neonatal rats with hypoxic-ischemic (HI) insults, employing behavioral tests as a method of assessment. METHODS: The experiment was conducted on male Wistar-Albino rats weighing 10 to 15 g, aged between seven and 10 days. The rats were divided into three groups using a random allocation method as follows: hypoxic ischemic encephalopathy (HIE) group, HIE treated with indomethacin group (INDO), and Sham group. A left common carotid artery ligation and hypoxia model was applied in both the HIE and INDO groups. The INDO group was treated with 4 mg/kg intraperitoneal indomethacin every 24 h for 3 days, while the Sham and HIE groups were given dimethylsulfoxide (DMSO). After 72 h, five rats from each group were sacrificed and brain tissue samples were stained with 2,3,5-Triphenyltetrazolium chloride (TCC) for infarct-volume measurement. Seven rats from each group were taken to the behavioral laboratory in the sixth postnatal week (PND42) and six from each group were sacrificed for the Evans blue (EB) experiment for blood-brain barrier (BBB) integrity evaluation. The open field (OF) test and Morris water maze (MWM) tests were performed. After behavioral tests, brain tissue were obtained and stained with TCC to assess the infarct volume. RESULTS: The significant increase in the time spent in the central area and the frequency of crossing to the center in the INDO group compared with the HIE group indicated that indomethacin decreased anxiety-like behavior (p < 0.001, p < 0.05). However, the MWM test revealed that indomethacin did not positively affect learning and memory performance (p > 0.05). Additionally, indomethacin significantly reduced infarct volume and neuropathological grading in adolescence (p < 0.05), although not statistically significant in the early period. Moreover, the EB experiment demonstrated that indomethacin effectively increased BBB integrity (p < 0.05). CONCLUSIONS: In this study, we have shown for the first time that indomethacin treatment can reduce levels of anxiety-like behavior and enhance levels of exploratory behavior in a neonatal rat model with HIE. It is necessary to determine whether nonsteroidal anti-inflammatory agents, such as indomethacin, should be used for adjuvant therapy in newborns with HIE.

Dexmedetomidine alleviates cognitive impairment by promoting hippocampal neurogenesis via BDNF/TrkB/CREB signaling pathway in hypoxic-ischemic neonatal rats.

AIMS: Dexmedetomidine (DEX) has been reported to alleviate hypoxic-ischemic brain damage (HIBD) in neonates. This study aimed to investigate whether DEX improves cognitive impairment by promoting hippocampal neurogenesis via the BDNF/TrkB/CREB signaling pathway in neonatal rats with HIBD. METHODS: HIBD was induced in postnatal day 7 rats using the Rice-Vannucci method, and DEX (25 µg/kg) was administered intraperitoneally immediately after the HIBD induction. The BDNF/TrkB/CREB pathway was regulated by administering the TrkB receptor antagonist ANA-12 through intraperitoneal injection or by delivering adeno-associated virus (AAV)-shRNA-BDNF via intrahippocampal injection. Western blot was performed to measure the levels of BDNF, TrkB, and CREB. Immunofluorescence staining was utilized to identify the polarization of astrocytes and evaluate the levels of neurogenesis in the dentate gyrus of the hippocampus. Nissl and TTC staining were performed to evaluate the extent of neuronal damage. The MWM test was conducted to evaluate spatial learning and memory ability. RESULTS: The levels of BDNF and neurogenesis exhibited a notable decrease in the hippocampus of neonatal rats after HIBD, as determined by RNA-sequencing technology. Our results demonstrated that treatment with DEX effectively increased the protein expression of BDNF and the phosphorylation of TrkB and CREB, promoting neurogenesis in the dentate gyrus of the hippocampus in neonatal rats with HIBD. Specifically, DEX treatment significantly augmented the expression of BDNF in hippocampal astrocytes, while decreasing the proportion of detrimental A1 astrocytes and increasing the proportion of beneficial A2 astrocytes in neonatal rats with HIBD. Furthermore, inhibiting the BDNF/TrkB/CREB pathway using either ANA-12 or AAV-shRNA-BDNF significantly counteracted the advantageous outcomes of DEX on hippocampal neurogenesis, neuronal survival, and cognitive improvement. CONCLUSIONS: DEX promoted neurogenesis in the hippocampus by activating the BDNF/TrkB/CREB pathway through the induction of polarization of A1 astrocytes toward A2 astrocytes, subsequently mitigating neuronal damage and cognitive impairment in neonates with HIBD.

Hypoxic ischemic brain injury: animal models reveal new mechanisms of melatonin-mediated neuroprotection.

Oxidative stress (OS) and inflammation play a key role in the development of hypoxic-ischemic (H-I) induced brain damage. Following H-I, rapid neuronal death occurs during the acute phase of inflammation, and activation of the oxidant-antioxidant system contributes to the brain damage by activated microglia. So far, in an animal model of perinatal H-I, it was showed that neuroprostanes are present in all brain damaged areas, including the cerebral cortex, hippocampus and striatum. Based on the interplay between inflammation and OS, it was demonstrated in the same model that inflammation reduced brain sirtuin-1 expression and affected the expression of specific miRNAs. Moreover, through proteomic approach, an increased expression of genes and proteins in cerebral cortex synaptosomes has been revealed after induction of neonatal H-I. Administration of melatonin in the experimental treatment of brain damage and neurodegenerative diseases has produced promising therapeutic results. Melatonin protects against OS, contributes to reduce the generation of pro-inflammatory factors and promotes tissue regeneration and repair. Starting from the above cited aspects, this educational review aims to discuss the inflammatory and OS main pathways in H-I brain injury, focusing on the role of melatonin as neuroprotectant and providing current and emerging evidence.

Traditional Chinese medicine for neonatal hypoxic-ischemic encephalopathy: A Bayesian network meta-analysis.

ETHNOPHARMACOLOGICAL RELEVANCE: Although therapeutic hypothermia is currently considered an effective treatment for neonatal hypoxic-ischemic encephalopathy (HIE), neonatal HIE is a disease requiring multiple therapeutic measures. Related Chinese herbal injections (CHIs) have been carried out in Chinese local hospitals for several years, and the outcomes all seem to show positive results. At the same time, other Traditional Chinese medicine (TCM) methods have also shown vigorous vitality. AIM OF THE STUDY: This study constructed a network meta-analysis (NMA) to investigate the efficacy of CHIs including Shenmai Injection (SMI), Compound musk injection (SXI), Ligustrazine injection (CXI), Compound danshen injection (DSI), Astragalus injection (HQI), Ginkgo biloba extract injection (YXI), and Puerarin injection (GGI) combined with traditional symptomatic treatment (TST) and TST alone in HIE. METHODS: A literature review was conducted in several databases from inception to 9 February 2023. The quality of the included studies was assessed by the Cochrane risk of bias tool. Data were analyzed by STATA 17.0 and R 4.2.2 software. Surface under the cumulative ranking curve (SUCRA) probability values were applied to rank the examined treatments. Bayesian network meta-analysis was designed to access the effectiveness of different CHIs. RESULTS: A total of 46 eligible randomized controlled trials involving 3,448 patients and 7 CHIs were included. The results of the NMA showed that SMI, SXI, CXI, DSI, HQI, YXI, and GGI combined with TST significantly improved treatment performance compared to TST alone. SMI + TST had obvious superiorities in the clinical effective rate and the original reflection recovery time. SXI + TST was the most advantageous in the Cure rate and the Neonatal Behavioral Neurological Assessment (NBNA). CXI + TST was shown to reduce the incidence of sequelae best. All articles reported that there were no obvious adverse drug reactions/adverse drug events (ADRs/ADEs). CONCLUSION: This NMA showed that using CHIs in combination with TST improved treatment performance and could be beneficial for patients with HIE compared to using TST alone. Thereinto, SXI + TST showed a preferable improvement in patients with HIE when unified considering the clinical effective rate and other outcomes. As for safety, more evidence is needed to support this hypothesis.

Mitochondria-targeted cerium vanadate nanozyme suppressed hypoxia-ischemia injury in neonatal mice via intranasal administration.

Oxidative stress is a major obstacle for neurological functional recovery after hypoxia-ischemia (HI) brain damage. Nanozymes with robust anti-oxidative stress properties offer a therapeutic option for HI injury. However, insufficiency of nanozyme accumulation in the HI brain by noninvasive administration hinders their application. Herein, we reported a cerium vanadate (CeVO4) nanozyme to realize a noninvasive therapy for HI brain in neonatal mice by targeting brain neuron mitochondria. CeVO4 nanozyme with superoxide dismutase activity mainly co-located with neuronal mitochondria 1 h after administration. Pre- and post-HI administrations of CeVO4 nanozyme were able to attenuate acute brain injury, by inhibiting caspase-3 activation, microglia activation, and proinflammation cytokine production in the lesioned cortex 2 d after HI injury. Moreover, CeVO4 nanozyme administration led to short- and long-term functional recovery following HI insult without any potential toxicities in peripheral organs of mice even after prolonged delivery for 4 weeks. These beneficial effects of CeVO4 nanozyme were associated with suppressed oxidative stress and up-regulated nuclear factor erythroid-2-related factor 2 (Nrf2) expression. Finally, we found that Nrf2 inhibition with ML385 abolished the protective effects of CeVO4 nanozyme on HI injury. Collectively, this strategy may provide an applicative perspective for CeVO4 nanozyme therapy in HI brain damage via noninvasive delivery.

[The possibilities of Mexidol usage in neuropediatrics]. / Vozmozhnosti primeneniya Meksidola v neiropediatrii.

Mexidol (ethylmethylhydroxypyridine succinate) is a modern neurometabolic medication increasingly being used in neuropediatrics. The results of recent studies confirming the positive effects of Mexidol pharmacotherapy in children with attention deficit hyperactivity disorder (ADHD), perinatal damages of the central nervous system (hypoxic-ischemic encephalopathy) and their consequences, neurological disorders and neurodevelopmental delay after surgery for congenital heart defects, neuroinfections (meningitis, encephalitis), posttraumatic epilepsy. Taking into account the unique multimodal action of Mexidol, it seems promising to expand the range of indications for its application in neuropediatrics, based on the results of new clinical trials organized in accordance with modern principles of evidence-based medicine.

Correlation of sLOX-1 and CSF1 with the pathological progression of hypoxic-ischemic encephalopathy in neonatal rats.

To provide clinical evidence for the management of hypoxic-ischemic encephalopathy (HIE) by analyzing the role of soluble lectin-like oxidized low-density lipoprotein receptor-1 (sLOX-1) and colony-stimulating factor-1 (CSF1) in the disease. We purchased 15 Sprague-Dawley (SD) rat pups and randomized them into five groups (n=3), of which one group was untreated as the control group and the other four were modeled by HIE. After modeling, a group was treated as a model group without any treatment, another group was injected with sLOX-1-silencing lentiviral vector (sLOX-1-si group), and the third and fourth were injected with CSF1-silencing lentiviral vector (CSF1-si group) and an equal amount of normal saline (blank group), respectively. After the corresponding intervention, the rat tissue in each group was obtained to observe the pathological injury by HE and TUNEL staining. In addition, sLOX-1, CSF1, 5-hydroxytryptamine (5-HT), dopamine (DA), and norepinephrine (NE) levels in brain tissue of each group were determined. The model group showed more severe pathological damage of the hippocampus and higher neuronal apoptosis than the control group. Besides, higher sLOX-1 and CSF1 levels and lower 5-HT, DA and NE contents were identified in the model group versus the control group (P<0.05). Compared with the blank group, sLOX-1-si and CSF1-si groups showed significantly alleviated hippocampal damage, inhibited neuronal apoptosis, reduced 5-HT, DA, NE, Bax, and cl-caspase-3, and increased Bcl-2 (P<0.05). Silencing sLOX-1 and CSF1 expression ameliorated the pathological injury of HIE and inhibited neuronal apoptosis.

Neuroprotective Action of Tacrolimus before and after Onset of Neonatal Hypoxic-Ischaemic Brain Injury in Rats.

(1) Background: Neonatal brain injury can lead to permanent neurodevelopmental impairments. Notably, suppressing inflammatory pathways may reduce damage. To determine the role of neuroinflammation in the progression of neonatal brain injury, we investigated the effect of treating neonatal rat pups with the immunosuppressant tacrolimus at two time points: before and after hypoxic-ischaemic (HI)-induced injury. (2) Methods: To induce HI injury, postnatal day (PND) 10 rat pups underwent single carotid artery ligation followed by hypoxia (8% oxygen, 90 min). Pups received daily tacrolimus (or a vehicle) starting either 3 days before HI on PND 7 (pre-HI), or 12 h after HI (post-HI). Four doses were tested: 0.025, 0.05, 0.1 or 0.25 mg/kg/day. Pups were euthanised at PND 17 or PND 50. (3) Results: All tacrolimus doses administered pre-HI significantly reduced brain infarct size and neuronal loss, increased the number of resting microglia and reduced cellular apoptosis (p < 0.05 compared to control). In contrast, only the highest dose of tacrolimus administered post-HI (0.25 mg/kg/day) reduced brain infarct size (p < 0.05). All doses of tacrolimus reduced pup weight compared to the controls. (4) Conclusions: Tacrolimus administration 3 days pre-HI was neuroprotective, likely mediated through neuroinflammatory and cell death pathways. Tacrolimus post-HI may have limited capacity to reduce brain injury, with higher doses increasing rat pup mortality. This work highlights the benefits of targeting neuroinflammation during the acute injurious period. More specific targeting of neuroinflammation, e.g., via T-cells, warrants further investigation.

Recombinant human erythropoietin protects against immature brain damage induced by hypoxic/ischemia insult.

To investigate the neuroprotection of recombinant human erythropoietin (rhEPO) against hypoxic/ischemic (HI) insult in three-day-old rats. Postnatal day 3 (PD3) rats were randomly divided into three groups: Sham group, HI group and HI+rhEPO group. Ligation of the right common carotid artery and hypoxia to induce HI brain injury. After HI insult, the rats received intraperitoneal injection of rhEPO (5000 IU/Kg, qod) in HI+rhEPO group or equal saline in other groups. On PD10, damage of brain tissue was examined by hematoxylin-eosin (HE) staining, observation of neuronal apoptosis in the hippocampus and cortex using immunofluorescence assay (marker: TUNEL). Immunohistochemical staining or western blotting was performed to detect the expression of cyclooxygenase-2 (COX-2), Caspase-3 and phosphorylated Akt (p-Akt) protein. On PD28, cognitive ability of rats was assessed by Morris water maze test. HI injury causes brain pathological morphology and cognitive function damage in PD3 rats, which can be alleviated by rhEPO intervention. Compared with the HI group, the HI+rhEPO group showed an increase in platform discovery rate and cross platform frequency, while the search platform time was shortened (P < 0.05). The proportion of TUNEL positive neurons and the expression of COX-2 and Caspase-3 proteins in brain tissue in the hippocampus and cortex was decreased, while the expression of p-Akt protein was upregulated (P < 0.05). RhEPO could protect against the pathological and cognitive impairment of immature brain induced by HI insult. This neuroprotective activity may involve in inhibiting inflammatory and apoptosis by activation of PI3K/Akt signaling pathway.