Neuronal exosome proteins: novel biomarkers for predicting neonatal response to therapeutic hypothermia.

OBJECTIVE: Central nervous system (CNS) derived exosomes can be purified from peripheral blood and have been used widely in adult neurological disease. Application to neonatal neurological disease deserves investigation in the setting of hypoxic-ischaemic encephalopathy (HIE). DESIGN: Observational cohort. SETTING: Level III neonatal intensive care unit. PARTICIPANTS: Term/near-term neonates undergoing therapeutic hypothermia (TH) for HIE. INTERVENTIONS: Blood samples were collected at 0-6, 12, 24, 48 and 96 hours of life. MAIN OUTCOMES AND MEASURES: CNS exosomes were purified from serum using previously described methods. Biomarker protein levels were quantified using standard ELISA methods and normalised to exosome marker CD-81. The slope of change for biomarker levels was calculated for each time interval. Our primary outcome was MRI basal ganglia/watershed score of ≥3. RESULTS: 26 subjects were included (umbilical artery pH range 6.6-7.29; 35% seizures). An increasing MRI injury score was significantly associated with decreasing levels of synaptopodin between 0-6 and 12 hours (p=0.03) and increasing levels of lipocalin-2 (NGAL) between 12 and 48 hours (p<0.0001). Neuronal pentraxin was not significant. The negative predictive values for increasing synaptopodin and decreasing NGAL was 70.0% and 90.9%, respectively. CONCLUSIONS AND RELEVANCE: Our results indicate that CNS exosome cargo has the potential to act as biomarkers of the severity of brain injury and response to TH as well as quantify pharmacological response to neuroactive therapeutic/adjuvant agents. Rigorous prospective trials are critical to evaluate potential clinical use of exosome biomarkers.

Brain Hypoxia Is Associated With Neuroglial Injury in Humans Post-Cardiac Arrest.

[Figure: see text].

Serial blood cytokine and chemokine mRNA and microRNA over 48 h are insult specific in a piglet model of inflammation-sensitized hypoxia-ischaemia.

BACKGROUND: Exposure to inflammation exacerbates injury in neonatal encephalopathy (NE). We hypothesized that brain biomarker mRNA, cytokine mRNA and microRNA differentiate inflammation (E. coli LPS), hypoxia (Hypoxia), and inflammation-sensitized hypoxia (LPS+Hypoxia) in an NE piglet model. METHODS: Sixteen piglets were randomized: (i) LPS 2 µg/kg bolus; 1 µg/kg infusion (LPS; n = 5), (ii) Saline with hypoxia (Hypoxia; n = 6), (iii) LPS commencing 4 h pre-hypoxia (LPS+Hypoxia; n = 5). Total RNA was acquired at baseline, 4 h after LPS and 1, 3, 6, 12, 24, 48 h post-insult (animals euthanized at 48 h). Quantitative PCR was performed for cytokines (IL1A, IL6, CXCL8, IL10, TNFA) and brain biomarkers (ENO2, UCHL1, S100B, GFAP, CRP, BDNF, MAPT). MicroRNA was detected using GeneChip (Affymetrix) microarrays. Fold changes from baseline were compared between groups and correlated with cell death (TUNEL) at 48 h. RESULTS: Within 6 h post-insult, we observed increased IL1A, CXCL8, CCL2 and ENO2 mRNA in LPS+Hypoxia and LPS compared to Hypoxia. IL10 mRNA differentiated all groups. Four microRNAs differentiated LPS+Hypoxia and Hypoxia: hsa-miR-23a, 27a, 31-5p, 193-5p. Cell death correlated with TNFA (R = 0.69; p < 0.01) at 1-3 h and ENO2 (R = -0.69; p = 0.01) at 48 h. CONCLUSIONS: mRNA and miRNA differentiated hypoxia from inflammation-sensitized hypoxia within 6 h in a piglet model. This information may inform human studies to enable triage for tailored neuroprotection in NE. IMPACT: Early stratification of infants with neonatal encephalopathy is key to providing tailored neuroprotection. IL1A, CXCL8, IL10, CCL2 and NSE mRNA are promising biomarkers of inflammation-sensitized hypoxia. IL10 mRNA levels differentiated all three pathological states; fold changes from baseline was the highest in LPS+Hypoxia animals, followed by LPS and Hypoxia at 6 h. miR-23, -27, -31-5p and -193-5p were significantly upregulated within 6 h of a hypoxia insult. Functional analysis highlighted the diverse roles of miRNA in cellular processes.

Identification of novel biomarkers for neonatal hypoxic-ischemic encephalopathy using iTRAQ.

BACKGROUND: A prompt diagnosis of HIE remains a challenge clinically. This study aimed to identify potential biomarkers of neonatal hypoxic-ischemic encephalopathy (HIE) via a novel proteomic approach, the isobaric tags for absolute and relative quantification (iTRAQ) method. METHODS: Blood samples were collected from neonates with mild (n = 4), moderate (n = 4), or severe (n = 4) HIE who were admitted to the neonatal intensive care unit of Children's Hospital of Soochow University between Oct 2015 and Oct 2017. iTRAQ was performed in HIE patients and healthy controls (n = 4). Bioinformatics analyses including Gene Ontology and KEGG pathway enrichment analysis were performed to evaluate the potential features and capabilities of the identified differentially expressed proteins. RESULTS: A total of 51 commonly differentially expressed proteins were identified among the comparisons between mild, moderate, and severe HIE as well as healthy controls. Haptoglobin (HP) and S100A8 were most significantly up-regulated in patients with HIE and further validated via real-time PCR and western blotting. The differentially expressed proteins represented multiple biological processes, cellular components and molecular functions and were markedly enriched in complement and coagulation cascades. CONCLUSIONS: HP and S100A8 may serve as a potential biomarker for neonatal HIE and reflects the severity of HIE. The complement and coagulation cascades play crucial roles in the development of neonatal HIE.

Role of miR-326 in neonatal hypoxic-ischemic brain damage pathogenesis through targeting of the δ-opioid receptor.

Hypoxic-ischemic brain damage (HIBD) is a relatively common malignant complication that occurs in newborn infants, but promising therapies remain limited. In this study, we focused on the role of miR-326 and its target gene δ-opioid receptor (DOR) in the pathogenesis of neonatal HIBD. The expression levels of miR-326 and DOR after hypoxic-ischemic injury were examined both in vivo and in vitro. The direct relationship between miR-326 and DOR was confirmed by a dual-luciferase reporter assay. Further, effects of miR-326 on cell viability and apoptosis levels under oxygen glucose deprivation (OGD) were analyzed. The expression levels of miR-326 were significantly lower and DOR levels were significantly higher in the HIBD group than the control group both in vivo and in vitro. Overexpression of miR-326 downregulated the expression of DOR, while suppression of miR-326 upregulated the expression of DOR. The dual-luciferase reporter assay further confirmed that DOR could be directly targeted and regulated by miR-326. MiR-326 knockdown improved cell survival and decreased cell apoptosis by decreasing the expression levels of Caspase-3 and Bax and increasing Bcl-2 expression in PC12 cells after exposure to OGD. Moreover, DOR knockdown rescued the effect of the improved cell survival and suppressed cell apoptosis induced by silencing miR-326. Our findings indicated that inhibition of miR-326 may improve cell survival and decrease cell apoptosis in neonatal HIBD through the target gene DOR.

Blocking lncRNA H19-miR-19a-Id2 axis attenuates hypoxia/ischemia induced neuronal injury.

Elevated expression of lncRNA H19 (H19) in the setting of hypoxia has been implicated as a promising therapeutic target for various cancers. However, little is known about the impact and underlying mechanism of H19 in ischemic brain stroke. This study found that H19 levels were elevated in the serum of stroke patients, as well as in the ischemic penumbra of rats with middle cerebral artery occlusion/reperfusion (MCAO/R) injury and neuronal cells with oxygen glucose deprivation (OGD). Further, knockdown of H19 with siRNA alleviated cell apoptosis in OGD neuronal cells, and inhibition of H19 in MCAO/R rats significantly decreased neurological deficit, brain infarct volume and neuronal apoptosis. Lastly, with gain and loss of function studies, dual luciferase reported assay, RNA immunoprecipitation (RIP) and pull-down experiments, we demonstrated the dual competitive interaction of miR-19a with H19 and the 3'-UTR of Id2 mRNA, resulting in the identification of the H19-miR-19a-Id2 axis. With biological studies, we also revealed that H19-miR-19a-Id2 axis modulated hypoxia induced neuronal apoptosis. This study demonstrates that the identified H19-miR-19a-Id2 axis plays a critical role in hypoxia induced neuronal apoptosis, and blocking this axis may serve as a novel therapeutic strategy for ischemic brain injury.

Comparative Effects of Monosialoganglioside versus Citicoline on Apoptotic Factor, Neurological Function and Oxidative Stress in Newborns with Hypoxic-Ischemic Encephalopathy.

OBJECTIVE: To determine the comparative effect of monosialoganglioside versus citicoline on the content changes of serum apoptotic factors (PDCD5, sFas and sFasL), neurological function indices (BDNF, NSE, S100-ß and NGF) and oxidative stress indices (SOD, MDA and GSH-PX) in newborns with hypoxic-ischemic encephalopathy (HIE). STUDY DESIGN: An experimental study. PLACE AND DURATION OF STUDY: Emergency Department, Affiliated Children's Hospital of Zhengzhou University, China, from October 2016 to February 2018. METHODOLOGY: A total of 90 newborns with HIE were randomly divided into a treatment group and a control group, with 45 cases in each group. In addition to the conventional treatment, the treatment group was given monosialoganglioside treatment, while the control group was given citicoline treatment. Both groups were treated for 10 days. After treatment, the content differences of serum apoptosis factors (PDCD5, sFas and sFasL), neurological function indices (BDNF, NSE, S100-ß and NGF) and oxidative stress indices (SOD, MDA and GSH-PX) were observed in the two groups. RESULTS: After treatment, the levels of serum PDCD5, sFas, sFasL, MDA, NSE and S100-ß in the treatment group were lower than those in the control group (all p<0.001). The contents of serum SOD, GSH-PX, BDNF and NGF in the treatment group were higher than those in the control group (all p<0.001). CONCLUSION: Monosialoganglioside can effectively improve the apoptotic factors, neurological function and oxidative stress indices in newborns and maintain the stability of the internal environment, so it is worthy of promotion and application.

Glial Fibrillary Acid Protein and Cerebral Oxygenation in Neonates Undergoing Cardiac Surgery.

BACKGROUND: Neonates undergoing surgery for complex congenital heart disease are at risk of developmental impairment. Hypoxic-ischemic brain injury might be a contributing factor. We aimed to investigate the perioperative release of the astrocyte cytoskeleton component glial fibrillary acid protein and its relation to cerebral oxygenation. METHODS: Serum glial fibrillary acid protein levels were measured before and 0, 12, 24, and 48 hours after surgery. Reference values were based on preoperative samples; concentrations above the 95th percentile were defined as elevated. Cerebral oxygenation was derived by near-infrared spectroscopy. RESULTS: Thirty-six neonates undergoing 38 surgeries utilizing cardiopulmonary bypass were enrolled (complete data available for 35 procedures). Glial fibrillary acid protein was elevated after 18 surgeries (arterial switch: 7/12; Norwood: 5/15; others: 6/8; p = 0.144). Age at surgery was higher in cases with elevated serum levels (6 [4-7] vs. 4 [2-5] days, p = 0.009) and intraoperative cerebral oxygen saturation was lower (70 ± 10% vs. 77 ± 7%, p = 0.029). In cases with elevated postoperative glial fibrillary acid protein, preoperative cerebral oxygen saturation was lower for neonates undergoing the arterial switch operation (55 ± 9% vs. 64 ± 4%, p = 0.048) and age at surgery was higher for neonates with a Norwood procedure (7 [6-8] vs. 5 [4-6] days, p = 0.028). CONCLUSIONS: Glial fibrillary acid protein was elevated after ∼50% of neonatal cardiac surgeries and was related to cerebral oxygenation and older age at surgery. The potential value as a biomarker for cerebral injury after neonatal cardiac surgery warrants further investigation; in particular, the association with neurodevelopmental outcome needs to be determined.

Inflammatory Responses are Sex Specific in Chronic Hypoxic-Ischemic Encephalopathy.

Neonatal hypoxic-ischemic encephalopathy (HIE) is increasingly recognized as a sexually dimorphic disease. Male infants are not only more vulnerable to ischemic insult; they also suffer more long-term cognitive deficits compared with females with comparable brain damage. The innate immune response plays a fundamental role in mediating acute neonatal HIE injury. However, the mechanism underlying the sex difference in chronic HIE is still elusive. The present study investigated the sex difference in HIE outcomes and inflammatory response in the chronic stage (30 days after HIE). Postnatal day 10 (P10) male and female C57BL/6 pups were subjected to 60-min Rice-Vanucci model (RVM) to induce HIE. Brain atrophy and behavioral deficits were analyzed to measure stroke outcomes at 30 days of HIE. Flow cytometry (FC) was performed to examine central (microglial activation) and peripheral immune responses. Serum levels of cytokines and sex hormones were determined by enzyme-linked immunosorbent assay (ELISA). Neurogenesis was quantified by 5-Bromo-2'-deoxyuridine (BrdU) incorporation with neurons. Results showed males had worse HIE outcomes than females at the endpoint. Female microglia exhibited a more robust anti-inflammatory response that was corresponding to an enhanced expression of CX3C chemokine receptor 1 (CX3CR1) than males. More infiltration of peripheral lymphocytes was seen in male vs. female HIE brains. Cytokine levels of tumor necrosis factor (TNF)-α and interleukin (IL)-10 were more upregulated in males and females respectively than their counterparts. Neurogenesis was more highly induced in females vs. males. No significant difference in circulating hormonal level was found between males and females after HIE. We conclude that a sex dichotomy in pro- and anti-inflammatory response underlies the sex-specific chronic HIE outcomes, and an enhanced neurogenesis in females also contribute to the sex difference.

Plasma Biomarkers of Brain Injury in Neonatal Hypoxic-Ischemic Encephalopathy.

OBJECTIVES: To evaluate plasma brain specific proteins and cytokines as biomarkers of brain injury in newborns with hypoxic-ischemic encephalopathy (HIE) and, secondarily, to assess the effect of erythropoietin (Epo) treatment on the relationship between biomarkers and outcomes. STUDY DESIGN: A study of candidate brain injury biomarkers was conducted in the context of a phase II multicenter randomized trial evaluating Epo for neuroprotection in HIE. Plasma was collected at baseline (<24 hours) and on day 5. Brain injury was assessed by magnetic resonance imaging (MRI) and neurodevelopmental assessments at 1 year. The relationships between Epo, brain-specific proteins (S100B, ubiquitin carboxy-terminal hydrolase-L1 [UCH-L1], total Tau, neuron specific enolase), cytokines (interleukin [IL]-1ß, IL-6, IL-8, IL-10, IL-12P70, IL-13, interferon-gamma [IFN-γ], tumor necrosis factor alpha [TNF-α], brain-derived neurotrophic factor [BDNF], monocyte chemoattractant protein-1), and brain injury were assessed. RESULTS: In 50 newborns with encephalopathy, elevated baseline S100B, Tau, UCH-L1, IL-1ß, IL-6, IL-8, IL-10, IL-13, TNF-α, and IFN-γ levels were associated with increasing brain injury severity by MRI. Higher baseline Tau and lower day 5 BDNF were associated with worse 1 year outcomes. No statistically significant evidence of Epo treatment modification on biomarkers was detected in this small cohort. CONCLUSIONS: Elevated plasma brain-specific proteins and cytokine levels in the first 24 hours of life are associated with worse brain injury by MRI in newborns with HIE. Only Tau and BDNF levels were found to be related to neurodevelopmental outcomes. The effect of Epo treatment on the relationships between biomarkers and brain injury in HIE requires further study. TRIAL REGISTRATION: ClinicalTrials.gov: 01913340.

The Effects of Intermittent Whole-Body Hypoxic Preconditioning on Patients with Carotid Artery Stenosis.

OBJECTIVE: To study the effects of intermittent whole-body hypoxic preconditioning on patients with carotid artery stenosis. METHODS: Fifty patients with carotid artery stenosis were selected and randomly divided into a hypoxic intervention group (HIG) and a control group (CG). Both groups were treated with a hypoxic respiration device for 7 days (HIG: 18% oxygen, CG: 21% oxygen). Venous blood samples were taken preoperatively and postoperatively. The subjects' vital signs were recorded during and after the intervention. After the completion of the trial, the concentrations of hemoglobin, hypoxia inducible factor-1α, erythropoietin, vascular endothelial growth factor, neuron-specific enolase, S100ß protein, brain-derived neurotrophic factor, serum aspartate transaminase, serum alanine aminotransferase, serum creatinine, and blood urea nitrogen were measured in the previously selected blood samples. RESULTS: During the intervention, the vital signs of the HIG were significantly different from those of the CG (P < 0.05). In the HIG, postoperative concentrations of hemoglobin, erythropoietin, hypoxia inducible factor-1α, and vascular endothelial growth factor were significantly more than the preoperative values (P < 0.05). In the CG, postoperative concentrations of neuron-specific enolase and S100ß protein were more than the preoperative values (P < 0.05). The concentrations of brain-derived neurotrophic factor, serum aspartate transaminase, serum alanine aminotransferase, serum creatinine, and blood urea nitrogen showed no significant differences between their preoperative and postoperative values in either the HIG or the CG (P > 0.05). CONCLUSIONS: Intermittent hypoxic preconditioning can change the vital signs and hematologic indexes of patients with carotid artery stenosis without causing new postoperative complications or organ damage.

[Iron metabolism and neonatal hypoxic ischemic brain damage].

Iron is an essential element for nervous system development, and maintaining a normal iron level in nervous system is controlled by multiple factors. Recent studies reported that iron dysregulation and the following iron metabolic pathways played an important role in hypoxic ischemic brain damage (HIBD) in neonates. Circulatory iron level is altered after hypoxia-ischemia exposure, which may cause abnormal iron deposition in the nervous system followed by neuronal injury. Finding the causing factors for abnormal iron metabolism after hypoxia-ischemia exposure, as well as understanding the mechanisms how iron metabolism contributes to HIBD, will shed lights on HIBD prevention and treatment. In this mini-review, we summarized changes in iron metabolism after neonatal hypoxia-ischemia exposure, its possible regulatory factors and how iron abnormalities contribute to HIBD.

A comparison of blood and cerebrospinal fluid cytokines (IL-1ß, IL-6, IL-18, TNF-α) in neonates with perinatal hypoxia.

Perinatal hypoxia-ischemia is a specific and important pathological event in neonatal care practice. The data on relationship between the concentrations of cytokines in blood and cerebrospinal fluid (CSF) and perinatal brain injury are scarce. The aim of this study is to evaluate changes in interleukin (IL-1ß, IL-6, and IL-18) and tumor necrosis factor alpha (TNF-α) levels in newborns with perinatal hypoxia (PNH). CSF and serum samples of 35 term and near-term (35-40 weeks) newborns with PNH, at the age of 3-96 hours, were analyzed using enzyme-linked immunosorbent assay. Control group consisted of 25 non-asphyxic/non-hypoxic infants of the same age sampled for clinically suspected perinatal meningitis, but proven negative and healthy otherwise. The cytokine values in CSF and serum samples were determined in relation to initial hypoxic-ischemic encephalopathy (HIE) staged according the Sarnat/Sarnat method, and compared with neurological outcome at 12 months of age estimated using Amiel-Tison procedure. The concentrations of IL-6 and TNF-α in serum of PNH patients were significantly higher compared to control group (p = 0.0407 and p = 0.023, respectively). No significant difference between average values of cytokines in relation to the stage of HIE was observed. Significantly higher levels of IL-6 and IL-18 corresponded to a mildly abnormal neurological outcome, while higher levels of IL-6 and TNF-α corresponded to a severely abnormal neurological outcome, at 12 months of age. Elevated serum levels of IL-6 and TNF-α better corresponded with hypoxia/ischemia compared to CSF values, within 96 hours of birth. Also, higher serum levels of IL-6, TNF-α, and IL-18 corresponded better with abnormal neurological outcome at 12 months of age, compared to CSF values.

Osteopontin Is a Blood Biomarker for Microglial Activation and Brain Injury in Experimental Hypoxic-Ischemic Encephalopathy.

Clinical management of neonatal hypoxic-ischemic encephalopathy (HIE) suffers from the lack of reliable surrogate marker tests. Proteomic analysis may identify such biomarkers in blood, but there has been no proof-of-principle evidence to support this approach. Here we performed in-gel trypsin digestion of plasma proteins from four groups of 10-d-old mice [untouched and 24 h after low-dose lipopolysaccharide (LPS) exposure, hypoxia-ischemia (HI), or LPS/HI injury; n = 3 in each group) followed by liquid chromatography-tandem mass spectrometry and bioinformatics analysis to search for HI- and LPS/HI-associated brain injury biomarkers. This analysis suggested the induction of plasma osteopontin (OPN) by HI and LPS/HI, but not by sham and injury-free LPS exposure. Immunoblot confirmed post-HI induction of OPN protein in brain and blood, whereas Opn mRNA was induced in brain but not in blood. This disparity suggests brain-derived plasma OPN after HI injury. Similarly, immunostaining showed the expression of OPN by Iba1+ microglia/macrophages in HI-injured brains. Further, intracerebroventricular injection of LPS activated microglia and up-regulated plasma OPN protein. Importantly, the induction of plasma OPN after HI was greater than that of matrix metalloproteinase 9 or glial fibrillary acid protein. Plasma OPN levels at 48 h post-HI also parallel the severity of brain damage at 7-d recovery. Together, these results suggest that OPN may be a prognostic blood biomarker in HIE through monitoring brain microglial activation.

Perinatal Asphyxia and Erythropoietin and VEGF: Serial Serum and Cerebrospinal Fluid Responses.

BACKGROUND: Infants with neonatal encephalopathy (NE) of hypoxic-ischaemic origin are at risk of oxidative and ischaemia-reperfusion injury, which may induce abnormal inflammatory responses involving excessive cytokine production and release in serum and cerebrospinal fluid (CSF). Systemic inflammation is found in infants with NE, and we therefore were interested in cytokines associated with hypoxia, including vascular endothelial growth factor (VEGF) and erythropoietin (Epo). OBJECTIVE: To investigate the relationship between Epo, VEGF levels, brain injury and outcome in a group of term infants exposed to perinatal asphyxia (PA) compared to controls. METHODS: Serum and CSF biomarkers associated with hypoxia (VEGF, Epo) were serially measured using multiplex immunoassays over days 1-4 in term infants exposed to PA including infants with NE and controls. Results were compared to severity of encephalopathy, MR brain imaging and mortality. RESULTS: Ninety-four infants had 247 serum samples collected (n = 12 controls, 82 exposed to PA with 34 CSF samples), and 4 infants died. Controls had significantly lower serum Epo levels on days 1 and 2 compared to those exposed to PA (p = 0.02). Grade II/III NE was significantly associated with elevated day 2 Epo and decreased day 1 VEGF (p < 0.05; day 2 Epo AUC = 0.74, cut-off 10.05 IU/ml). Elevated serum Epo was associated with severely abnormal MRI. Mortality was associated with elevated day 3 Epo and decreased day 1 VEGF. CSF levels were all after hypothermia and were not significantly associated with outcome. CONCLUSION: Serum Epo and VEGF may be markers of severity of hypoxia-ischaemia and brain injury as they are closely related to hypoxic exposure.

Effect of GABAB receptor antagonists (CGP 35348 and CGP 55845) on serum interleukin 6 and 18 concentrations in albino mice following neonatal hypoxia ischemia insult.

Interleukin (IL) 6 and 18 plays an important role in inflammatory response following hypoxia ischemia encephalopathy (HIE). Present study was designed to demonstrate the effect of two GABAB receptor antagonists (CGP 35348 and 55845), respectively, on the serum IL6 and IL 18 concentrations in albino mice. Albino mice pups (of both genders) were subjected to Murine model of hypoxia-ischemia encephalopathy on postnatal day 10 (right common carotid artery was ligated followed by 8% hypoxia for 25 minutes). After neonatal brain damage and following weaning, mice were divided in three groups, in gender specific manner, and fed on normal rodent diet till they were 13 week old. At this time point, group 1 received intraperitonial saline solution (control group), group 2 was supplemented with CGP 35348 (1mg/ml solvent/Kg body weight) and group 3 with CGP 55845 (1mg/ml solvent/Kg body weight), intraperitonially, for 12 days and IL 6 and 18 concentrations were determined in serum by ELISA. It was observed that CGP 35348 supplementation resulted in reduced interlukin-6 and interlukin-18 concentrations in male albino mice. While CGP 55845 supplementation increased IL-6 and IL-18 concentrations in female albino mice following HIE. Our results are indicating that GABAB receptor antagonist's supplementation affects IL concentrations in albino mice in a gender specific manner following neonatal brain damage and can be further explored for the treatments of hypoxia ischemia associated neurological ailments.

Cytokine changes in newborns with therapeutic hypothermia after hypoxic ischemic encephalopathy.

OBJECTIVE: This study aimed to examine changes in cytokines according to therapeutic hypothermia (TH) for newborn hypoxic ischemic encephalopathy (HIE). STUDY DESIGN: We studied 20 neonates who were admitted with a diagnosis of HIE in the neonatal intensive care unit. Cytokine concentration assay was carried out for neonates (n=12) who received TH and neonates (n=8) who were not treated with hypothermia by collecting blood sample at 12, 48 and 120 h after birth. RESULTS: At 48 h after birth, interleukin (IL)-6 in the normothermia group was higher than that in the hypothermia group (P=0.010). At 48 h after birth, IL-10 was higher in the hypothermia group than in the normothermia group (P=0.038). CONCLUSION: This study confirmed that TH performs a role in the prevention of inflammatory process by way of maintaining proinflammatory cytokine IL-6 at low levels and anti-inflammatory cytokines IL-10 at high levels.

Association of brain injury and neonatal cytokine response during therapeutic hypothermia in newborns with hypoxic-ischemic encephalopathy.

BACKGROUND: Cytokines have been proposed as mediators of neonatal brain injury via neuroinflammatory pathways triggered by hypoxia-ischemia. Limited data are available on cytokine profiles in larger cohorts of newborns with hypoxic-ischemic encephalopathy (HIE) undergoing therapeutic hypothermia (TH). METHODS: Serum cytokines interleukin (IL)-1ß, IL-2, IL-4, IL-6, IL-8, IL-10, IL-13, tumor necrosis factor-α, and interferon-γ were measured in newborns with HIE at 24 and 72 h of TH. Differences between infants with favorable (survivors with mild/no magnetic resonance imaging (MRI) injury) vs. adverse outcome (death or moderate/severe MRI injury) were compared using mixed models to adjust for covariates. RESULTS: Data from 36 term newborns with HIE (favorable outcome: n = 20, adverse outcome: n = 16) were evaluated. Cytokines IL-1ß, IL-2, IL-6, IL-8, IL-10, and IL-13 were elevated in the adverse relative to favorable outcome group at 24 h. IL-6 remained significantly elevated in the adverse outcome group at 72 h. IL-6 and IL-10 remained significantly associated with outcome group after controlling for covariates. CONCLUSION: Inflammatory cytokines are elevated in HIE newborns with brain injury by MRI. In particular, IL-6 and IL-10 were associated with adverse outcomes after controlling for baseline characteristics and severity of presentation. These data suggest that cytokine response may identify infants in need of additional neuroprotective interventions.

Effect of creatine monohydrate supplementation on relative serum level of IL-6 and IL-18 following neonatal hypoxia ischemia in male albino mouse.

IL-6 has been reported to have neuroprotective effects against cerebral ischemia while IL-8 is a pro inflammatory cytokine structurally related to interleukin-1 family. In the present study, we tried to determine whether 2% Creatine monohydrate supplementation for variable duration influence the IL-6 and 18 concentrations in the serum of male albino mouse following right common carotid artery ligation and hypoxia (8% oxygen) for 25 minutes. Our result revealed that serum concentration of IL6 (P=0.0001) as well as IL-18 (P=0.003) were significantly higher in mice supplemented with creatine monohydrate for 15 weeks than in male albino mice on normal rodent diet following hypoxic ischemic insult indicating that long term creatine monohydrate supplementation up regulates the IL-6 and IL-18 concentrations triggering the neuroinflammatory and neuroprotective responses.

Neuroprotective effect of escitalopram oxalate in rats with chronic hypoperfusion.

The neuroprotective effects of escitalopram oxalate in rats with chronic hypoperfusion and the possible mechanism were explored. Chronic hypoperfusion (2-VO) model was prepared and given escitalopram oxalate (experimental group) or PBS (control group) after 6 weeks. Eight weeks after the operation, Morris water maze test was carried out to evaluate the learning and memory ability of the rats. The cell proliferation, three-dimensional vascular distribution, cell morphological changes in ischemic area and the plasma vascular endothelial growth factor (VEGF) were detected to explore the possible mechanisms. (1) Morris water maze test showed that the escape latency in the experimental group was significantly shorter than in the control group, while the first quadrant swimming time in the experimental group was significantly longer than the control group (both P<0.01). (2) Cerebrovascular confocal detection results showed that the inside diameter of capillaries was significantly less in the experimental group than in the control group; the vascular density was significantly increased in the experimental group and the total area of capillaries was also significantly increased in the experimental group as compared with the control group. (3) There was statistically significant difference in BrdU-positive cells in the ischemic brain tissue between the experimental group and the control group (P=0.003<0.01). (4) VEGF concentrations in the plasma and the ischemic area were higher in the experimental group than in the control group (P<0.05). It was concluded that escitalopram oxalate could significantly improve the learning and memory ability of the rats with chronic cerebral ischemia probably by the VEGF-mediated angiogenesis.