Table 1.
Commonly used and relevant animal models of NEC.
| Model (Abbreviation) | Animal (Age) | Protocol Time | Key Points | Ref. |
|---|---|---|---|---|
| Hypoxia Hypothermia Formula (HHF) | Rats (E20–21) | 3–4 days | Commonly used model, basis of several current models across species. Adaptations typically include gavage feeds with hyperosmolar formula creating impaired barrier/dysbiosis and exposure to bacterial pathogen (either colonized or administered via orogastric tube) ± LPS following exposure to periods of hypothermia, hypoxia ( ± hyperoxia to stimulate clinical conditions in certain variations). Activates TLR-pathway. | (9, 32, 40, 42–45) |
| Mouse (P0–10) | 3–7 days | Widely used adaptation of rat HHF model with vast number of variations that often include exposure to bacterial pathogens (Klebsiella, C. sakazakii, E. coli, “NECteria”) ± LPS for increased disease manifestation. Commonly applied NEC induction protocol in transgenic mice testing specific genes/mechanisms (TLR4, VEGF, eNOS) ± exposure to various reagents/antibodies/modulators (amniotic fluid, HIF1α, IGF) to test therapeutic potential. | (9, 27, 33, 46–50) | |
| Piglet (Term, E115) | 3–4 days | High rate of complications including 36% with pulmonary hemorrhage, 24% rectal perforations partly associated to technique (intubation, tonometer applied transanally), with severe manifestation of NEC. | (51) | |
| Hypoxia Formula (HF) | Rats (E20–21, P0–3) | 4–7 days | Requires exogenous or catheter colonization of bacteria to induce consistent NEC-like injury. Many variations utilized with varying exposures to hypoxia (decreased FiO2, 100% Nitrogen, etc) and different formula types (RMS, Ebsilac, human formula) | (8, 31, 52–56) |
| Mouse (P0-P14) | 3–4 days | Requires bacterial challenge ± LPS to trigger bacterial/inflammatory signaling; Important model for TLR4 signaling mechanisms. | (57, 58) | |
| Formula Feeds (FF) | Mouse: SSC/Elecare (P8) | 3 days | Use of hyperosmotic preterm human formula to induce NEC-like injury without exposure to hypoxia/hypothermia, has not yet been validated/replicated | (59) |
| Mouse: Maltodextrin (P5–6 and P9–10) | 10 days | Important model to study specific components (maltodextrin) of formula that create susceptibility to NEC. Consistent pattern of impairment with addition of maltodextrin combined Klebsiella (K) and/or hypoxia (H), worse with M/H. High survival rate after NEC induction protocol with milder severity. | (60) | |
| Piglet: Formula variations (E105–108) | 1–4 days | Induction of NEC with formula feeding alone without exogenous hypoxic or hypothermic conditions. Prematurity of piglet (with transitional hypoxia and similarly impaired microvasculature to human premature infants), and presence bacterial colonization (no impact on germ-free piglets) necessary to induce intestinal injury. | (38, 48, 49, 61, 62) | |
| Piglet: Parenteral Nutrition (PN) (E105–109) | 5 days | Gut dysfunction worsens with PN followed by FF, PN results in delayed gut maturation worsened by dysbiosis-induced FF. Several management changes in initiation of feeds, rate of feeding advancements, type of enteral feed, and PN-related applications derived from this model. Low true-NEC manifestation, wider distribution of disease to entire GI tract (including stomach, colon). | (51, 63–66) | |
| Paneth Cell Disruption (PCD) | Mouse (P14–16) | 16 h | Dithizone- or Diphtheria toxin-induced PC depletion. Model to study role of PCs in NEC, intestinal development closer to human development, model further simplified with either exposure to hyperosmotic formula (RMS) or bacterial challenge (Klebsiella, NECteria); short protocol time, TLR4-independent pathway | (10, 67–70) |
| Microvasculature Maldevelopment (MM) | Mouse: -VEGF/-IGF (E16–20, P0-1) | 3 days | Models used to study intestinal vascular development and function using HHF NEC induction protocol in neonatal pups, using inhibition, deletion or down regulation on VEGF-related pathways. Fetal exposure to inflammation (using LPS) in utero (E16–20) followed by NEC induction at P0-1 also explored to determine chorioamnionitis impact on vasculature development and susceptibility to NEC. Addition of TNF shown to worsen NEC severity via decreased VEGF/VEGFR2 activity but prevented by DMOG (via HIF1a). | (27, 71, 72) |
| Mouse: eNOS (P5) | 4 days | Loss of eNOS in transgenic mice leads to greater gut and lung injury with altered inflammatory cascade, NO protective | (73) | |
| Phlebotomy-induced anemia (PIA) | Mouse (P2) | 10–12 days | Model investigating whether severe anemia ± RBC transfusion contributes to development and severity of NEC, activating TLR4-signalling mechanisms to drive inflammation and injury; Can be used to evaluate risk factor of iatrogenic anemia and gut perfusion. | (74, 75) |
| Antibiotic Exposure/Dysbiosis (ABT) | Mouse (P1–14) | 14 days | Model to study role of prolonged antibiotic exposure leading to increased susceptibility to NEC when challenged at P14. | (76) |
| Piglet (E 106) | 5 days | Model to study effects of enteral vs. parenteral antibiotics in immediate post-natal period suggesting that enteral but not parenteral exposure protected from NEC, short duration (<5 days) of IV antibiotics with mild injury noted, no NEC. | (77) | |
| Trinitrobenzene sulfonic acid (TNBS) | Mouse (P10) | 1 day | Model using non-specific immunologic stimulant (TNBS) to induce NEC-like injury, highlighted critical role of gut microbiome with absence of injury in germ-free mice. | (78) |
| Dextran Sulfate Sodium (DSS) | Mouse (P3) | 6 days | Adaptation of DSS model of IBD in adult mice, applied to neonatal mice to induce intestinal injury in the absence of hypoxia, hypothermia or LPS driven by humoral and cellular immune responses. | (79) |
| Anti-CD3 mAb | Mouse (P0) | 2 days | Novel model illustrating the effect of T-cell inhibition to explore role of adaptive immunity in severity of NEC-like injury combined with dysbiosis from formula feeds (injury attenuated with antibiotics) | (80) |