Urine Protein Biomarkers for the Diagnosis and Prognosis of Necrotizing Enterocolitis in Infants

Abstract

Objectives To test the hypothesis that an exploratory proteomics analysis of urine proteins with subsequent development of validated urine biomarker panels would produce molecular classifiers for both the diagnosis and prognosis of infants with necrotizing enterocolitis (NEC). Study design Urine samples were collected from 119 premature infants (85 NEC, 17 sepsis, 17 control) at the time of initial clinical concern for disease. The urine from 59 infants was used for candidate biomarker discovery by liquid chromatography/mass spectrometry. The remaining 60 samples were subject to enzyme-linked immunosorbent assay for quantitative biomarker validation. Results A panel of 7 biomarkers (alpha-2-macroglobulin-like protein 1, cluster of differentiation protein 14, cystatin 3, fibrinogen alpha chain, pigment epithelium-derived factor, retinol binding protein 4, and vasolin) was identified by liquid chromatography/mass spectrometry and subsequently validated by enzyme-linked immunosorbent assay. These proteins were consistently found to be either up- or down-regulated depending on the presence, absence, or severity of disease. Biomarker panel validation resulted in a receiver-operator characteristic area under the curve of 98.2% for NEC vs sepsis and an area under the curve of 98.4% for medical NEC vs surgical NEC. Conclusions We identified 7 urine proteins capable of providing highly accurate diagnostic and prognostic information for infants with suspected NEC. This work represents a novel approach to improving the efficiency with which we diagnose early NEC and identify those at risk for developing severe, or surgical, disease.

Current Research on the Epidemiology, Pathogenesis, and Management of Necrotizing Enterocolitis

“…Recently, biomarker discovery studies of the urinary proteome [16] and peptidome [17] have identified proteins and peptides associated with poor outcome (i.e., disease progression). Importantly, the biomarkers alone were not able to completely distinguish progressors from non-progressors, but were able to when clinical factors were incorporated into the prediction model.…”

Investigation of urine proteome of preterm newborns with respiratory pathologies

“…Non-invasive MS based methods for monitoring the status of children with ELBW and VLBW are based on the analysis of easily accessible biomaterials such as buccal scrapings [5], exhaled breath condensate [6], and urine [7][8][9][10]. Biomarkers in the urine samples of infants are widely used for predicting such pathologies as obstructive nephropathy [11,12], necrotizing enterocolitis [13], and for monitoring maturation and development of the urinary system [7]. The majority of throughput urine proteome studies are focused on the adult population (control and disease) [14][15][16][17].…”

“…Additional sample fractionation allowed Marimuthu and colleagues to add 671 proteins to this catalog [15]. Different strategies are applied for shotgun urinary proteomics, including semi-quantitative label-free HPLC-MS/MS, 2D-PAGE-MS/MS or CE-MS/MS [8][9][10][11][12][13]18], quantitative SRM/MRM HPLC-MS/MS [15][16][17].…”

Early Detection of Impending Necrotizing Enterocolitis with Urinary Intestinal Fatty Acid-Binding Protein

“…In a recent multicenter study, seven urinary biomarkers, including alpha-2-macroglobulin-like protein and retinol-binding protein 4, were accurate in differentiating NEC from sepsis [24]. Another study found that levels of interleukin-1 receptor in buccal swabs were lower in premature infants who developed NEC several weeks later than in those who did not [25].…”