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Morphology of Nervous Lesion in Fetal Rats with Retinoic Acid-induced Myelomeningocele at Disparate Age

Author: ShenJian
Tutor: BiYunLi
School: Fudan University
Course: Pediatrics
Keywords: neurogenic bladder dysfunction myelomeningocele fetus retinoic acid α-actin neurotubulin-β-Ⅲ GFAP VAChT
CLC: R714.5
Type: Master's thesis
Year: 2011
Downloads: 22
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Abstract


BackgroundMyelomeningocele (MMC) is the main cause of neurogenic bladder dysfunction (NBD) in children, which can lead to lifelong physical disabilities, including paraplegia, urinary and fecal incontinence, hydrocephaly, sexual dysfunction, skeletal deformations and mental impairment. Early intervention by fetal surgery could be an effective approach to deal with MMC, although there are still some argument in the outcome of the surgery. Therefore, further study is needed by the means of animal models. While a number of surgically created or mutant animal models of MMC exist, they are unable to replicate the primary defect in neurulation, limiting their experimental relevance to the "secondary" injuries of mechanical or chemical trauma. Retinoic acid-induced MMC may be an ideal model. The relationship between spinal cord lesion of fetal MMC and the nervous and muscular modification of the bladder is so far unclear.Part1Optimization of retinoic acid-induced myelomeningocele in fetal ratsObjective:To create and optimize the model of retinoic acid-induced myelomeningocele in fetal rats with higher rate of MMC, lower rate of still birth/resorption and smaller range of lesion, and to approach the most optimal time and dose for administration of RA in our laboratory.Method:Time-dated pregnant SD rats were divided into there groups by the different time of being gavage fed with ATRA dissolved in2ml of olive oil:group E9, E9.5and E10, fed at9a.m.,11p.m. on the9th day after pregnancy,9a.m. on the10th day after pregnancy respectively. Control rats were fed with2ml of olive oil alone. Dosage in group E9, E9.5and E10ranged in60-110mg/kg,40-120mg/kg and80-100mg/kg respectively. Fetus were harvested at E16, E18or E20by cesarean section. Numbers of pregnant rats, fetal rats, rats with MMC and still birth/resorption were accounted. Rates of MMC and still birth/resorption in treatment group were calculated. Data were analyzed by Pearson χ2using Stata7. Significance was defined as P<0.05.Results:1. The rate of MMC was extremely low in group E9at any dosage, ranging from0to1.92%, while the rate of still birth/resorption was dramaticly high, ranging from90.91%to100%.2. In group E10, the rate of MMC at the dosage of120mg/kg was29.95%, which was the highest in this group.3. In group E9.5, the rate of MMC at the dosage of80mg/kg and100mg/kg were31.66%and29.41%respectively, and no significant difference was found. While the rate of still birth/resorption at80mg/kg was significantly lower than that at100mg/kg. At the dosage of80mg/kg, some fetus with fairly small MMC were gained.Part2Morphology of nervous lesion in the spinal cord and bladder of fetal rats with myelomeningocele at disparate ageObjective:To analyze the development and innervation to the bladder smooth muscle and nervous lesion in the spinal cord of the MMC fetus at disparate age, and to reveal the interaction between the lesion of spinal cord and bladder.Method:Fetus were divided into MMC group and control group. Each group was further divided into trere groups at disparate fetal age:E16, E18and E20. A-actin and neurotubulin-β-Ⅲ were analyzed in bladders, GFAP and VAChT were analyzed in lumbosacral levels spinal cords by immunohistochemisty. Photos were taken to determine the integrated optical density of each sample. Data were analyzed by independent t-test using SPSS16.0. Significance was defined as P<0.05.Results:1-Neurotubulin-β-Ⅲ was detected at E16in both MMC group and control group. Whether at E16, E18or E20, neurotubulin-β-Ⅲ in MMC group was significantly lower than that in control group.2. Numerous of a-actin was detected in both groups at all fetal age. No significant difference was found between MMC group and control group at any fetal age.3. At E16and E18, no apparente GFAP positive astrocyte in MMC group or control group was deteced. At E20, in the dorsal region of the spinal cord, numerous GFAP positive astrocytes were detected, with significant difference to the control group.4. VAChT in MMC group was less detected than that in control group at all fetal age with significant difference. At E20, a number of VAChT positive neurons were found in the dorsolateral nucleus in control group.ConclusionGavage feeding to SD rats with80mg/kg of ATRA at E9.5can found a successful model of MMC, with high rate of MMC, low rate of still birth/resorption, and high mimicry to human MMC. The model can be used in further study of fetal surgery of MMC.Differentiation and development of the bladder smooth muscle of MMC fetal rats is morphologically as good as that in normal fetus.Innervation to the bladder smooth muscle of fetal rats takes place no later than E16. Meanwhile, the degeneration of innervation to the bladder in MMC fetus has already occurred. This change synchronously accompanies with the reduction of cholinergic neuron in sacral parasympathetic nucleus and dorsolateral nucleus of the spinal cord.Numerous of astrocytes emerge in the dorsal region of spinal cord in MMC fetus at M20.

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CLC: > Medicine, health > Obstetrics and Gynaecology > Obstetrics > Fetus
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