Identifying hypoxia in a newborn piglet model using urinary NMR metabolomic profiling.

Establishing the severity of hypoxic insult during the delivery of a neonate is key step in the determining the type of therapy administered. While successful therapy is present, current methods for assessing hypoxic injuries in the neonate are limited. Urine Nuclear Magnetic Resonance (NMR) metabolomics allows for the rapid non-invasive assessment of a multitude breakdown products of physiological processes. In a newborn piglet model of hypoxia, we used NMR spectroscopy to determine the levels of metabolites in urine samples, which were correlated with physiological measurements. Using PLS-DA analysis, we identified 13 urinary metabolites that differentiated hypoxic versus nonhypoxic animals (1-methylnicotinamide, 2-oxoglutarate, alanine, asparagine, betaine, citrate, creatine, fumarate, hippurate, lactate, N-acetylglycine, N-carbamoyl-β-alanine, and valine). Using this metabolomic profile, we then were able to blindly identify hypoxic animals correctly 84% of the time compared to nonhypoxic controls. This was better than using physiologic measures alone. Metabolomic profiling of urine has potential for identifying neonates that have undergone episodes of hypoxia

Physiological effects of hypoxia in newborn piglets.

<p>Temporal changes in (A) oxygen saturation (% O₂), (B) blood pH, (C) mean arterial pressure (mmHg), and (D) cardiac output between hypoxia (n = 7) and sham (n = 6) treated animals. *P<0.05 Sham vs. Hypoxia for corresponding time point.</p

The metabolomic model of hypoxic vs. sham treated animals.

<p>PLS-DA analysis of urine from hypoxic versus sham treated animals was based on differences in metabolites between groups shown as the Coefficient of Variation (CoV) plot (A). The importance of each metabolite within the model is shown as the Variability of Importance (VIP) plot (B).</p

Experimental timeline flowchart.

<p>Guinea pigs were sensitized to ovalbumin or given sham sensitizations via i.p. injections on Days 1 and 3. Sensitized and non-sensitized GPs were inoculated intranasally with parainfluenza virus (PIV) on Day 21. After recovery from virus or sham infection, all GPs were exposed to an aerosolized solution of ovalbumin on Day 45, with the exception of age-matched controls. Some GPs received dexamethasone on Days 61 to 65. Animals were inoculated intranasally with live-PIV, UV-PIV, or sham (NaCl 0.9%) on Day 70. On Day 75 all animals underwent airway hyperreactivity and inflammation assessment.</p

Repeat exposure to live virus induces airway hyperreactivity and airway inflammation in both non-sensitized and sensitized animals.

<p><b>(A)</b> Both non-sensitized and sensitized animals had significantly higher bronchoconstriction in response to histamine (i.v.) after re-infection with live PIV (closed square, n = 5 each), when compared to respective uninfected sham inoculated controls (open triangle, n = 3 each) or age-matched controls (n = 5 each; p<0.0001 each). <b>(B)</b> Non-sensitized animals re-exposed to live PIV (n = 6) have significantly higher total cell counts in the BAL compared to age-matched (n = 5) and sham controls (n = 3; p<0.001 for both). Sensitized animals re-infected with live PIV (n = 4) also developed higher total cell counts in the BAL compared to age-matched (n = 3) and sham controls (n = 4; p<0.001 for each). In the sensitized animals, macrophages and eosinophils were significantly more abundant. Error bars represent SEM.</p