Yung Chieh Lin

Cerebral microvascular damage occurs early after hypoxia-ischemia via nNOS activation in the neonatal brain

Microvascular injury early after hypoxic ischemia (HI) may contribute to neonatal brain damage. N-methyl-D-aspartate receptor overstimulation activates neuronal nitric oxide synthases (nNOS). We hypothesized that microvascular damage occurs early post-HI via nNOS activation and contributes to brain injury. Postpartum day-7 rat pups were treated with 7-nitroindazole (7-NI) or aminoguanidine (AG) before or after HI. Electron microscopy was performed to measure neuronal and endothelial cell damage. There were vascular lumen narrowing at 1 hour, pyknotic neurons at 3 hours, and extensive neuronal damage and loss of vessels at 24 hours post HI. Early after reoxygenation, there were neurons with heterochromatic chromatin and endothelial cells with enlarged nuclei occluding the lumen. There was also increased 3-nitrotyrosin in the microvessels and decreased cerebral blood perfusion. 7-NI and AG treatment before hypoxia provided complete and partial neuroprotection, respectively. Early post-reoxygenation, the AG group showed significantly increased microvascular nitrosative stress, microvascular interruptions, swollen nuclei that narrowed the vascular lumen, and decreased cerebral perfusion. The 7-NI group showed significantly decreased microvascular nitrosative stress, patent vascular lumen, and increased cerebral perfusion. Our results indicate that microvascular damage occurs early and progressively post HI. Neuronal nitric oxide synthases activation contributes to microvascular damage and decreased cerebral perfusion early after reoxygenation and worsens brain damage.

The Role of Glucocorticoid Receptors in Dexamethasone-Induced Apoptosis of Neuroprogenitor Cells in the Hippocampus of Rat Pups

Background. Dexamethasone (Dex) has been used to reduce inflammation in preterm infants with assistive ventilation and to prevent chronic lung diseases. However, Dex treatment results in adverse effects on the brain. Since the hippocampus contains a high density of glucocorticoid receptors (GCRs), we hypothesized that Dex affects neurogenesis in the hippocampus through inflammatory mediators. Methods. Albino Wistar rat pups first received a single dose of Dex (0.5 mg/kg) on postnatal day 1 (P1) and were sacrificed on P2, P3, P5, and P7. One group of Dex-treated pups (Dex-treated D1D2) was given mifepristone (RU486, a GCR antagonist) on P1 and sacrificed on P2. Hippocampi were isolated for western blot analysis, TUNEL, cleaved-caspase 3 staining for cell counts, and morphological assessment. Control pups received normal saline (NS). Results. Dex reduced the developmental gain in body weight, but had no effect on brain weight. In the Dex-treated D1D2 group, apoptotic cells increased in number based on TUNEL and cleaved-caspase 3 staining. Most of the apoptotic cells expressed the neural progenitor cell marker nestin. Dex-induced apoptosis in P1 pups was markedly reduced (60%) by pretreatment with RU486, indicating the involvement of GCRs. Conclusion. Early administration of Dex results in apoptosis of neural progenitor cells in the hippocampus and this is mediated through GCRs.

Hypoxic/Ischemic and Infectious Events Have Cumulative Effects on the Risk of Cerebral Palsy in Very-Low-Birth-Weight Preterm Infants

Background: Hypoxia/ischemia and inflammation are two major mechanisms for cerebral palsy (CP) in preterm infants. Objective: To investigate whether hypoxia/ischemia- and infection-related events in the perinatal and neonatal periods had cumulative effects on CP risk in very-low-birth-weight (VLBW) premature infants. Methods: From 1995 to 2005, 5,807 VLBW preterm infants admitted to Taiwan hospitals were enrolled. The cumulative effects of hypoxic/ischemic and infectious events during the perinatal and neonatal periods on CP risk at corrected age 24 months were analyzed. Results: Of the 4,355 infants with 24-month follow-up, 457 (10.5%) had CP. The CP group had significantly higher incidences of hypoxia/ischemia-related events in the perinatal and neonatal periods, and sepsis in the neonatal period than the normal group. Three hypoxic/ischemic events, including birth cardiopulmonary resuscitation (OR 2.25; 95% CI 1.81-2.82), patent ductus arteriosus (PDA) ligation (2.94; 1.35-5.75) and chronic lung disease (3.14; 2.61-3.85) had the most significant contribution to CP. Relative to CP risk for infants with neither the three hypoxic/ischemic events nor sepsis, the CP odds increased 1.98-, 2.26- and 2.15-fold for infants with birth cardiopulmonary resuscitation, PDA ligation and chronic lung disease, respectively; while the combination with sepsis further increased the odds to 3.18-, 3.83- and 3.25-fold, respectively. Using the three hypoxic/ischemic events plus sepsis, CP rates were 10.0, 16.7, 26.7, 40.0 and 54.7% for infants with none, one, two, three and four events, respectively. Conclusions: Hypoxic/ischemic and infectious events across the perinatal and neonatal periods exerted cumulative effects on CP risk in VLBW premature infants.

Human Umbilical Vein Endothelial Cells Protect Against Hypoxic-Ischemic Damage in Neonatal Brain via Stromal Cell-derived Factor 1/C-X-C Chemokine Receptor Type 4

Background and Purpose— Agents that protect against neurovascular damage provide a powerful neuroprotective strategy. Human umbilical vein endothelial cells (HUVECs) may be used to treat neonates with hypoxic-ischemia (HI) because of its autologous capability. We hypothesized that peripherally injected HUVECs entered the brain after HI, protected against neurovascular damage, and provided protection via stromal cell–derived factor 1/C-X-C chemokine receptor type 4 pathway in neonatal brain. Methods— Postpartum day 7 rat pups received intraperitoneal injections of low-passage HUVEC-P4, high-passage HUVEC-P8, or conditioned medium before and immediately after HI. HUVECs were transfected with adenovirus-green fluorescent protein for cell tracing. Oxygen–glucose deprivation was established by coculturing HUVEC-P4 with mouse neuroblastoma neuronal cells (Neuro-2a) and with mouse immortalized cerebral vascular endothelial cells (b.End3). Results— HUVEC-P4–treated group had more blood levels of green fluorescent protein–positive cells than HUVEC-P8–treated group 3 hours postinjection. Intraperitoneally injected HUVEC-P4, but not HUVEC-P8, entered the cortex after HI and positioned closed to the neurons and microvessels. Compared with the condition medium–treated group, the HUVEC-P4–treated but not the HUVEC-P8–treated group showed significantly less neuronal apoptosis and blood–brain barrier damage and more preservation of microvessels in the cortex 24 hours after HI. On postpartum day 14, the HUVEC-P4–treated group showed significant neuroprotection compared with the condition medium–treated group. Stromal cell–derived factor 1 was upregulated in the ipsilateral cortex 3 hours after HI, and inhibiting the stromal cell–derived factor 1/C-X-C chemokine receptor type 4 reduced the protective effect of HUVEC-P4. In vitro transwell coculturing of HUVEC-P4 also significantly protected against oxygen–glucose deprivation cell death in neurons and endothelial cells. Conclusions— Cell therapy using HUVECs may provide a powerful therapeutic strategy in treating neonates with HI.

A single postnatal dose of dexamethasone enhances memory of rat pups later in life

Postnatal dexamethasone (Dex) therapy is associated with adverse neurodevelopmental outcomes, which might be related to its timing of administration. We used time-dated pregnant Wistar albino rats, whose litters were divided into experimental (Dex) and control groups intraperitoneally administered one dose of Dex (0.5 mg/kg) or normal saline (NS), respectively, at either day 1 (P1) or 7 (P7). The magnitude of the contextual freezing response and performance on the Morris water maze were significantly higher in the Dex-P7 group than in those of the other groups at P56. Dendritic spine density, membranous expression of the N-methyl-d-aspartate receptor (NMDAR) subunit NR2A/2B, and postsynaptic density-95 (PSD-95) were significantly higher in the Dex-P7 group than in the other groups. Furthermore, cytosolic expression of nuclear factor kappa B (NF-κB) and phosphatidylinositol 3-kinase (PI3K) was significantly higher in the Dex group than in NS group. Moreover, Dex administration at P7 increased cell proliferation, neuronal differentiation, and the survival of newly born neurons in the dentate gyrus. These results suggest Dex at P7 enhances the acquisition of contextual fear and spatial memory later in life due to the modulation of the newly born neurons, increase in dendritic spine number, and NMDAR expression.

Lung density standard deviations obtained using high-pitch dual-source computed tomography are valid predictors of bronchopulmonary dysplasia in preterm infants.

PURPOSE This study aimed to validate standard deviations of lung densities obtained using high-pitch dual-source computed tomography (DSCT) densitometry as indices of bronchopulmonary dysplasia (BPD) severity in premature infants. METHODS Data of preterm, late preterm group, and early term groups were evaluated. Mean and median standard deviations (SDmean, SDmedian) of CT lung density (CTLD) were calculated from CT images. RESULTS SDmean of CTLD in infants with severe BPD was significantly higher than that of infants without BPD (198.1 vs. 140.9, respectively; P=.002). CONCLUSIONS Study results support using high-pitch DSCT for BPD diagnosis and quantitative evaluation in prematurity.

Identifying Risk Factors Shared by Bronchopulmonary Dysplasia, Severe Retinopathy, and Cystic Periventricular Leukomalacia in Very Preterm Infants for Targeted Intervention

Background: Bronchopulmonary dysplasia (BPD), severe retinopathy of prematurity (sROP), and cystic periventricular leukomalacia (cPVL) are 3 major morbidities with long-term neurodevelopmental impairments in preterm infants. Objective: To investigate the strength of associations and identify key risk factors shared by BPD, sROP, and cPVL for targeted intervention. Methods: We studied the Taiwanese very-preterm-infant registry data on 3,507 infants admitted to neonatal intensive care units and discharged at postmenstrual age ≥36 weeks between 2008 and 2013. Results: Of 3,507 infants, 1,497 presented with at least 1 morbidity (26 [1.7%], 386 [25.8%], and 1,085 [72.5%] exhibited 3, 2, and 1 morbidities, respectively). BPD was strongly associated with sROP (odds ratio 5.93; 95% confidence interval 5.02–7.03), followed by cPVL (2.08; 1.63–2.64), but sROP and cPVL were weakly associated (1.59; 1.17–2.13). Most risk factors contributed to BPD, which shared risk factors with sROP and cPVL. A birth weight of < 1,000 g, male sex, and prolonged mechanical ventilation (MV) were shared by BPD and sROP, and chorioamnionitis, severe respiratory distress syndrome, and prolonged MV specifically contributed to BPD and cPVL. Prolonged MV was the single risk factor common to BPD, sROP, and cPVL. Avoiding prolonged MV reduced the risk of having at least 1 of the 3 morbidities by 37%. Conclusions: BPD and sROP were most strongly associated. Most risk factors contributed to BPD, with differentially shared effects on sROP and cPVL. Prolonged MV was the only risk factor shared by all 3 morbidities, and avoiding it potentially reduced the risk of having at least 1 of them.

PO-0694 Risk-adjusted Mortality Of Vlbw In Taiwan- A Population-based Study

Background and aims Preterm babies have higher mortality than terms. Risk-adjusted mortality (RAM) is useful for making comparisons among different NICUs. GA, BBW, sex, singleton birth and antenatal steroid have been used to estimate mortality (M) of preterm. The aims of this study are 1. To compare the performance of GA, BW, and Logistic Regression (LR) in predicting M of VLBW infants. 2. To compare the RAM in different areas and periods. Methods Cohort data from 2000 to 2011 were used. M is defined as death prior to discharge. Exclusion criteria included 1) Transferred after 24 h of age; 2) Death within 24 h of admission and 3) Lethal malformation. We developed a LR model to predict M [expected probability (Pro)]. ROC curves were used for assessing performance of predicting M. To compare the RAM, we calculated (O-E) Pro (observed Pro – expected Pro) values in each patient and used these values for comparisons. Results 9207 VLBWs were enrolled. The calculated probability of death by LR model was: P = 1/(1+e-Z), where e= natural logarithms and z = (-0.62•[prenatal steroid]) -(0.219•GA)- (0.004•BBW)- (0.327•[singleton]) + (0.286•[male]) + 8.438. Area under ROC were 0.858 for LR (95% CI: 0.847–0.869), 0.841 for BBW (95% CI: 0.829–0.853) and 0.827 for GA (95% CI: 0.815–0.839). Abstract PO-0694 Figure 1 There were significant differences of RAM in different locations and years (Figure 1). Conclusions The use of LR is better than GA and BW in predicting M of VLBWs. RAM can be used as a tool for quality improvement.