Danzer Enrico, Kiddoo Darcie A, Redden Robert A, Robinson Lauren, Radu Antoneta, Zderic Steve A, Doolin Edward J, Adzick N Scott, Flake Alan W
The Center for Fetal Research, The Children's Hospital of Philadelphia, The University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania 19104-4318, USA.
Am J Physiol Renal Physiol. 2007 Jan;292(1):F197-206. doi: 10.1152/ajprenal.00001.2006. Epub 2006 Aug 29.
Myelomeningocele (MMC) is the most common cause of neurogenic bladder dysfunction (NBD). We recently developed a novel retinoic acid (RA)-induced MMC model in fetal rats. The objective of this study was to use this model to assess functional and structural characteristics of the detrusor muscle in MMC-associated NBD. Time-dated pregnant Sprague-Dawley rats were gavage fed 60 mg/kg RA dissolved in olive oil or olive oil alone [embryonic day 10 (E10)]. Bladder specimens from olive oil-exposed fetuses (OIL; n = 71), MMC (n = 79), and RA-exposed-no MMC (RA, n = 62) were randomly assigned for functional and histopathological evaluation and protein analysis. Contractility responses to field and agonist-mediated stimulation (KCl and bethanecol) were analyzed. The expression patterns of alpha-smooth muscle actin, myosin, desmin, vimentin, and collagen III and I were analyzed by immunohistochemistry and Western blotting. Spatial and temporal distribution of nerve fibers within the detrusor muscle was monitored by neurotubulin-beta-III throughout gestation. Neither OIL, MMC, nor RA detrusor responded to field stimulation. MMC bladder strips showed a significant decrease in contractility after KCl and bethanechol stimulation compared with OIL and RA bladders. Bladder detrusor morphology and expression patterns of smooth muscle markers were similar between groups. Detrusor muscles in OIL and RA fetuses were densely innervated, possessing abundant intramural ganglia and nerve trunks that branch to supply smooth muscle bundles. In MMC bladders, neurotubulin-beta-III-positive nerve fibers were markedly decreased with advancing gestational age and were almost completely absent at term (E22). We conclude that the biomechanical properties of fetal rat MMC bladders are analogous to that seen in humans with MMC-associated NBD. Decreased nerve density indicates loss of peripheral neural innervation throughout gestation. The early observation of decreased innervation and decreased contractility in the absence of morphologic abnormalities in muscle structure or extracellular matrix supports a pathophysiological hypothesis that denervation is the primary insult preceding the observed alterations in bladder muscle structure and function.
脊髓脊膜膨出（MMC）是神经源性膀胱功能障碍（NBD）最常见的病因。我们最近在胎鼠中建立了一种新型视黄酸（RA）诱导的MMC模型。本研究的目的是利用该模型评估MMC相关NBD中逼尿肌的功能和结构特征。对处于特定孕期的Sprague-Dawley孕鼠在胚胎第10天（E10）经口灌胃给予溶解于橄榄油中的60 mg/kg RA或仅给予橄榄油。将来自暴露于橄榄油的胎儿（OIL；n = 71）、MMC（n = 79）和暴露于RA但无MMC（RA，n = 62）的膀胱标本随机分配用于功能、组织病理学评估和蛋白质分析。分析了对场刺激和激动剂介导刺激（氯化钾和氨甲酰甲胆碱）的收缩反应。通过免疫组织化学和蛋白质印迹分析α-平滑肌肌动蛋白、肌球蛋白、结蛋白、波形蛋白以及胶原蛋白III和I的表达模式。在整个妊娠期通过β-III微管蛋白监测逼尿肌内神经纤维的空间和时间分布。OIL、MMC和RA组的逼尿肌均对场刺激无反应。与OIL和RA组膀胱相比，MMC膀胱条在氯化钾和氨甲酰甲胆碱刺激后收缩性显著降低。各组间膀胱逼尿肌形态和平滑肌标志物的表达模式相似。OIL和RA组胎儿的逼尿肌神经支配密集，具有丰富的壁内神经节和神经干，这些神经干分支以供应平滑肌束。在MMC膀胱中，随着胎龄增加，β-III微管蛋白阳性神经纤维明显减少，足月时（E22）几乎完全消失。我们得出结论，胎鼠MMC膀胱的生物力学特性与MMC相关NBD患者所见相似。神经密度降低表明在整个妊娠期外周神经支配丧失。在肌肉结构或细胞外基质无形态学异常的情况下，早期观察到神经支配减少和收缩性降低，支持了一种病理生理学假说，即去神经支配是膀胱肌肉结构和功能发生观察到的改变之前的主要损伤。
