13 - Effects of Caffeine on Enolase as a Biomarker in a Rat Model for Preterm Hypoxic Ischemic Encephalopathy

Approximately 5-9 of 1000 preterm infants (< 35 weeks gestational age) experience hypoxic ischemic encephalopathy (HIE). Yet, there is no diagnostic biomarkers or approved treatment therapy for them. MRI imaging is used to diagnose HIE in term infants, but not in preterm infants. Neuronal specific enolase (NSE) has a positive correlation to neuronal damage and can be detected in plasma; but its relevance in detection of HIE is yet to be determined. Caffeine, a non-selective adenosine receptor antagonist, is used to treat apnea of prematurity in infants. In animal models of term HIE, caffeine shows anti-inflammatory properties and improves neurobehavioral sequelae. It remains unknown if caffeine has similar neuroprotective effects in preterm HIE model.


Objective: Determine neuroprotective effects of caffeine therapy and investigate NSE as a potential biomarker for both detecting HIE and testing the efficacy of caffeine in an established rat model of preterm HIE.

Design/Methods:

Rat litters were culled to 4 males & 4 females. Pups were randomly assigned within litters to sham+saline (S), sham+caffeine (C), hypoxic-ischemic (HI)+S, and HI+C. At postnatal day (P)7, (~34 weeks GA preterm infant), HI was induced by carotid ligation + 8% O2 (90 min hypoxia). Control (sham) pups underwent similar surgical procedure without ligation or hypoxia. S or C was given (IP injection) at 0, 24 & 48 hours (h) post-HI.  Then at 1, 24 & 72 h post-HI, both brain (cortex) and plasma samples were collected and assayed for IL1b (known marker for brain injury) and NSE levels using ELISA. Spearman correlation was computed for IL1b and NSE. 2-way ANOVA with post hoc Tukey comparison tests were used to analyze changes among treatment groups.


Results:

HI increased plasma NSE levels compared to sham groups at 24 h (p= 0.017), but not at 72 h post-HI (p=0.200). Caffeine treatment for HI shows no effect on plasma NSE levels at 24 h (p=0.495) and 72 h (p=0.683). IL1b is increased post-HI with statistical significance at 24 h (p=0.009), but not at 72 h (p=0.560) compared to sham groups. No effects on IL1B between HI+S and HI+C groups were seen (24 h, p=0.65 & 72 h, p=0.84).  There were no sex differences among treatment groups.


Conclusion(s):

The findings suggest NSE as a potential diagnostic biomarker for preterm HIE. More studies are ongoing to validate NSE as a potential objective marker of preterm HIE injury. Given the neuroprotective effects of a combined caffeine and melatonin treatment in term HIE, we will test this drug combo in future studies.