Establishment And Evaluation Of GMH-IVH Preterm Rabbit Model

Objective: Germinal matrix-intraventricular hemorrhage (GMH-IVH) isone of the most important brain damage in premature infants. It can lead toserious neurobehavioral sequela, cause a serious burden to the family and thecommunity. Nevertheless, there is no standard preterm animal GMH-IVHmodel. In order to carry on the studies of GMH-IVH in premature infants, thisresearch attempts to build a premature rabbit model to simulate GMH-IVH inhuman preterm infants, and finally by Cranial ultrasound, microscopy,immunohistochemistry, and neurobehavioral evaluation to comprehensivelyestimate the model.Methods1Premature rabbits were born by cesarean section and given incubated, milkpowder feeding after born. Then the premature rabbits were randomly dividedinto two groups, experimental group and control group, each group eighteen.Three hours later, the premature rabbits in experimental group were givenintraperitoneally injection of50%glycerol (10ml/kg), the preterm rabbits incontrol group were given intraperitoneally injected with the same dose ofsaline.2Cranial ultrasound scan was given to check the presence of GMH-IVH inpreterm rabbits at24h after born.3Premature rabbits were euthanized at24hours,48hours and72hours afterborn. Brain tissue was got in control group and experimental group. Each timepoint had six rabbits.The brain tissue was rapidly separated, the incidence ofGMH-IVH in both group were observed, and the result was compared withthat by cranial ultrasound.4The incidence of GMH-IVH was observed under microscope in thepremature rabbits between control group and experimental group at the24 hours,48hours and72hours after born.5The brains tissue structure of GMH-IVH premature rabbits was observed byelectron microscopy.615pups were recognized having GMH-IVH in experimental group(GMH-IVH model group) and an equal number of normal rabbits were alsochosen from control group (Normal control group). Within each group,15pups were randomly divided into three sub-groups: every time period had5pups (at24h,48h and72h after born).7Premature rabbits were accepted neurobehavioral assessment at the24hoursand72hours after born between the normal control group and the modelgroup by Derrick neurobehavioral score inorder to find the differencesbetween the normal control group and the model group on cranial nerves,consciousness, gait and to further observe the clinical similarities between theGMH-IVH premature rabbits and preterm infants with GMH-IVH.8The expression of glial fibrillary acidic protein（GFAP）of brains tissue wereobserved by immunohistochemistry method in normal control group andmodel group, the differences of GFAP positive cells expression rates werecompared in the model group between at24hours,48hours and72hours afterborn.9Statistical analysis SPSS17.0statistics software and Microsoft Excel2003were used to carry on statistics processing, Chi-Square was used to comparecategorical variables. α=0.05was size of test. P-value<0.05(double side) hadstatistical significance, highly significant when P-value<0.01.Results:1Cranial ultrasound: Premature rabbits were scanned by cranial ultrasound at24hours after born. In experimental group the incidence of GMH-IVH is83.33%(15/18).The incidence of GMH-IVH is11.11%(2/18) in control group.The incidence of GMH-IVH in experimental group was significantly higherthan control group. There were significant difference between them (P <0.01).2Specimens observed: There was no significant difference between theultrasound imaging results and the specimens. In experimental group the incidence of GMH-IVH is83.33%(15/18). The incidence of GMH-IVH is11.11%(2/18) in control group. The incidence of GMH-IVH in experimentalgroup was significantly higher than the control group. There were alsosignificant difference between them (P <0.01).3Microscope examinat was given to check the presence of GMH-IVH inpreterm rabbits between experimental group and control grou. The occurrenceof GMH-IVH in experimental group was88.89%(16/18). In control group theincidence of GMH-IVH was11.11%(2/18). The incidence of GMH-IVH incontrol group was significantly lower than that in experimental group. Therewere significant difference between them (P <0.01).4Electron microscope examination: The brain tissue of GMH-IVHperformance for the degeneration of the nerve cells, cytoplasmic vacuolization,axonal swelling, fracture myelin stripping, mitochondria disappeared and alarge number of vacuolar, all supported the GMH-IVH model was successfully.5Neurobehavioral testing: There was no significant difference in theneurobehavioral testing between model group and normal control group at24hours and72hours after born(P>0.05).6The expression of GFAP by immunohistochemical method: In normalcontrol group, GFAP-positive cells scattered distribution, lighter stain. Inmodel group, there was a significant increase in the number of GFAP.GFAP-positive cells staining deepened, irregular shape. GFAP-positive cells in model group was significantly higher than those in normal control group. There were significant difference between them (P<0.01). Within model group，GFAP positive cells express rate at48hours after bornwas higher than that at24hours after born (P <0.05). The expression at72hours after born was higher than that at48hours. There were significant difference between them within the model group (P<0.05).Conclusion:1Using the method of Georgiadis P, confirmed by cranial ultrasound, visualobservation of the specimens, microscope and electron microscope examination, the GMH-IVH preterm rabbit model was sucsessful established.The expression of GFAP in brain tissue by immunohistochemical analysissuggests that the established GMH-IVH model was similar to human preterminfant with GMH-IVH.2Through this study, we have established animal model closer to clinicalpreterm infant with GMH-IVH, and have also provided a more reliableexperimental evidence for future clinical research.