临床小儿外科杂志2018，Vol.17Issue(1)：54-59,6.DOI:10.3969/j.issn.1671-6353.2018.01.014
无神经节细胞动物模型的建立及鉴定
Establishment and identification of experimental rat model for aganglionosis
摘要
Abstract
Objective To establish a simple and valid rat model of aganglionosis.Methods Among 128 neonatal rats aged 1 week in 16 litters,8 rats per litter were randomly divided into control and treatment groups (n =4 each).Saline and benzalkonium chloride (BAC) were injected respectively into rectum.Such clinical manifestations as abdominal distention and defecation and rectal morphology were observed at Weeks 2,4,6,8 post-injection.The stains of hematoxylin-eosin (HE) and HuD protein immunofluorescence (IF) were used for counting the number of ganglion cells in mesenteric nervous plexus of rectum.And quantitative real time-polymerase chain reaction (qRT-PCR) was employed for detecting the expressions of glial fibrillary acidic protein (GFAP) and neuronal nitric oxide synthase (nNOS) in rectum.Results Survival status and rectal morphology:Varying degrees of diarrhea in two groups disappeared at Week 1.Treatment group showed mild abdominal distention and decreased defecation at Week 6.Abdominal distention worsened and defecation further improved at Week 8.There was an onset of mental depression.And control group had no obvious abnormalities.There were no obvious abnormal changes in neither groups at Weeks 2 & 4.In treatment group,mild rectal stricture occurred at Week 6,worsened at Week 8 and proximal intestine had obvious dilation with an accumulation of intestinal contents.Control group showed no change.HE staining:The mean numbers of ganglion cells of treatment and control groups were 6 vs 6 and 6 vs 6 at Weeks 2 & 4 and size or shape showed no difference (P ＞ 0.05).The mean numbers of ganglion cells of two groups were 3 to 6 at Week 6 (P ＜ 0.05).And mesenteric nervous plexus became smaller and more distorted in treatment group.At 8 weeks post-injection,mesenteric nervous plexus was absent and control group had no abnormities.Immunofluorescence (IF) stain:mesenteric nervous plexus was positive for yellow green fluorescence.Nucleus was obvious while cytoplasm remained obscured.Nerve fiber was non-stained and the boundary of staining was distinct.No inter-group difference existed in mesenteric nervous plexus at Weeks 2 & 4.Smaller size and more irregular shape existed in treatment group versus control group at Week 6 and yellow green fluorescence disappeared at Week 8 post-injection.Control group had no abnormities.Quantitative real time-polymerase chain reaction (qRT-PCR):The expressions of GDNF mRNA and nNOS mRNA showed no significant inter-group difference at Weeks 2 & 4 (P ＞ 0.05).At Weeks 6 & 8,the expressions of GDNF mRNA and nNOS mRNA were obviously lower in treatment group than those in control group (GDNF:0.06 ±0.03 vs 1.05 ±0.32,0.39 ±0.24 vs 1.02 ±0.22;nNOS:0.54 ±0.33 vs 1.14 ± 0.50,0.40 ±0.24 vs 1.03 ±0.26).In treatment group,it was obviously higher at Week 8 than that at Week 6.Conclusion Absence of ganglion cells in rectum may be successfully induced by an injection of BAC into rectum.And abnormal expressions of GDNF and nNOS are probably correlated with an occurrence of aganglionosis.关键词
Hirschsprung病/动物模型/大鼠/苯扎铵化合物Key words
Hirschsprung Disease/Models, Animal/Rats/Benzalkonium Compounds引用本文复制引用
金祝,尚卿,刘远梅..无神经节细胞动物模型的建立及鉴定[J].临床小儿外科杂志,2018,17(1):54-59,6.基金项目
贵州省科学技术基金项目(黔科合J字[2012] 2364号) （黔科合J字[2012] 2364号）
遵义市“15851”人才项目 （）
遵义医学院附属医院硕士科研启动基金(院字(2011)01号) （院字(2011）
