中医正骨
中醫正骨
중의정골
The Journal of Traditional Chinese Orthopedics and Traumatology
2015年
9期
3-8
,共6页
刘楚吟%麻昊宁%朱宝%董春科%颜勤华%谭明生
劉楚吟%痳昊寧%硃寶%董春科%顏勤華%譚明生
류초음%마호저%주보%동춘과%안근화%담명생
维生素 A 缺乏%枕骨%颈椎%先天畸形%Hoxd3 基因%动物实验
維生素 A 缺乏%枕骨%頸椎%先天畸形%Hoxd3 基因%動物實驗
유생소 A 결핍%침골%경추%선천기형%Hoxd3 기인%동물실험
vitamin A deficiency%occipital bone%cervical vertebrae%congenital abnormalities%Hoxd3 gene%animal experimentation
目的：探讨维生素 A 缺乏导致新生大鼠枕颈部畸形的作用机制。方法：将40只5周龄雌性健康 SPF 级 SD 大鼠随机分为2组，正常对照组20只，维生素 A 缺乏组20只；20只适龄雄性健康 SPF 级 SD 大鼠分笼饲养。正常对照组饲以 AIN －93M饲料，维生素 A 缺乏组饲以缺乏维生素 A 的 AIN －93M饲料，雄性大鼠饲以 AIN －93M饲料。饲养至第10周从雌性大鼠的目外眦取血，采用酶联免疫吸附法检测2组雌鼠血清维生素 A 含量，鉴定大鼠模型。造模成功后，将大鼠按雌2雄1的比例合笼交配，第2天清晨检查阴栓，有阴栓者记为 E0 d。于妊娠 E11．5 d 取2组半数的孕鼠，麻醉处死后剖腹，每只孕鼠取出2只胚胎，剥离被膜，置于液氮中，然后转移至－80℃的冰箱里保存待测。于妊娠 E21 d 取新生鼠，麻醉处死后剥离皮肤筋膜以及内脏，置于95％的酒精中固定待测。采用 RT －qPCR 方法检测胚胎 Hoxd3基因 mRNA 表达情况；采用双色染色法制作大鼠骨骼标本，观察新生鼠枕颈部骨骼发育情况。结果：①雌鼠血清维生素 A 含量。维生素 A 缺乏组大鼠血清维生素 A 含量明显低于正常对照组[（214±39）ng·mL －1，（432±54）ng·mL －1，t ＝14．636，P ＝0．000]。②胚胎 Hoxd3基因 mRNA 的表达量。维生素 A 缺乏组胚胎 Hoxd3基因表达量低于正常对照组（0．58±0．11，1．03±0．14，t ＝10．407，P ＝0．000）。③新生鼠枕颈部骨骼形态。正常对照组可见枕骨与寰枢椎发育正常，对位关系良好；维生素 A 缺乏组可见寰椎后弓与枕骨大孔后缘角度增大，寰枢椎对位关系异常，寰枢椎后弓角度增大，C2～C7椎弓及棘突高度降低、宽度变窄，且相邻椎体间隙增大。结论：维生素 A 缺乏可以导致新生大鼠枕颈部结构发育异常，这可能与其胚胎发育期 Hoxd3基因 mRNA 表达水平下降有关，但其具体的分子生物学机制还有待进一步研究。
目的：探討維生素 A 缺乏導緻新生大鼠枕頸部畸形的作用機製。方法：將40隻5週齡雌性健康 SPF 級 SD 大鼠隨機分為2組，正常對照組20隻，維生素 A 缺乏組20隻；20隻適齡雄性健康 SPF 級 SD 大鼠分籠飼養。正常對照組飼以 AIN －93M飼料，維生素 A 缺乏組飼以缺乏維生素 A 的 AIN －93M飼料，雄性大鼠飼以 AIN －93M飼料。飼養至第10週從雌性大鼠的目外眥取血，採用酶聯免疫吸附法檢測2組雌鼠血清維生素 A 含量，鑒定大鼠模型。造模成功後，將大鼠按雌2雄1的比例閤籠交配，第2天清晨檢查陰栓，有陰栓者記為 E0 d。于妊娠 E11．5 d 取2組半數的孕鼠，痳醉處死後剖腹，每隻孕鼠取齣2隻胚胎，剝離被膜，置于液氮中，然後轉移至－80℃的冰箱裏保存待測。于妊娠 E21 d 取新生鼠，痳醉處死後剝離皮膚觔膜以及內髒，置于95％的酒精中固定待測。採用 RT －qPCR 方法檢測胚胎 Hoxd3基因 mRNA 錶達情況；採用雙色染色法製作大鼠骨骼標本，觀察新生鼠枕頸部骨骼髮育情況。結果：①雌鼠血清維生素 A 含量。維生素 A 缺乏組大鼠血清維生素 A 含量明顯低于正常對照組[（214±39）ng·mL －1，（432±54）ng·mL －1，t ＝14．636，P ＝0．000]。②胚胎 Hoxd3基因 mRNA 的錶達量。維生素 A 缺乏組胚胎 Hoxd3基因錶達量低于正常對照組（0．58±0．11，1．03±0．14，t ＝10．407，P ＝0．000）。③新生鼠枕頸部骨骼形態。正常對照組可見枕骨與寰樞椎髮育正常，對位關繫良好；維生素 A 缺乏組可見寰椎後弓與枕骨大孔後緣角度增大，寰樞椎對位關繫異常，寰樞椎後弓角度增大，C2～C7椎弓及棘突高度降低、寬度變窄，且相鄰椎體間隙增大。結論：維生素 A 缺乏可以導緻新生大鼠枕頸部結構髮育異常，這可能與其胚胎髮育期 Hoxd3基因 mRNA 錶達水平下降有關，但其具體的分子生物學機製還有待進一步研究。
목적：탐토유생소 A 결핍도치신생대서침경부기형적작용궤제。방법：장40지5주령자성건강 SPF 급 SD 대서수궤분위2조，정상대조조20지，유생소 A 결핍조20지；20지괄령웅성건강 SPF 급 SD 대서분롱사양。정상대조조사이 AIN －93M사료，유생소 A 결핍조사이결핍유생소 A 적 AIN －93M사료，웅성대서사이 AIN －93M사료。사양지제10주종자성대서적목외자취혈，채용매련면역흡부법검측2조자서혈청유생소 A 함량，감정대서모형。조모성공후，장대서안자2웅1적비례합롱교배，제2천청신검사음전，유음전자기위 E0 d。우임신 E11．5 d 취2조반수적잉서，마취처사후부복，매지잉서취출2지배태，박리피막，치우액담중，연후전이지－80℃적빙상리보존대측。우임신 E21 d 취신생서，마취처사후박리피부근막이급내장，치우95％적주정중고정대측。채용 RT －qPCR 방법검측배태 Hoxd3기인 mRNA 표체정황；채용쌍색염색법제작대서골격표본，관찰신생서침경부골격발육정황。결과：①자서혈청유생소 A 함량。유생소 A 결핍조대서혈청유생소 A 함량명현저우정상대조조[（214±39）ng·mL －1，（432±54）ng·mL －1，t ＝14．636，P ＝0．000]。②배태 Hoxd3기인 mRNA 적표체량。유생소 A 결핍조배태 Hoxd3기인표체량저우정상대조조（0．58±0．11，1．03±0．14，t ＝10．407，P ＝0．000）。③신생서침경부골격형태。정상대조조가견침골여환추추발육정상，대위관계량호；유생소 A 결핍조가견환추후궁여침골대공후연각도증대，환추추대위관계이상，환추추후궁각도증대，C2～C7추궁급극돌고도강저、관도변착，차상린추체간극증대。결론：유생소 A 결핍가이도치신생대서침경부결구발육이상，저가능여기배태발육기 Hoxd3기인 mRNA 표체수평하강유관，단기구체적분자생물학궤제환유대진일보연구。
Objective:To explore the mechanism of occipito -cervical deformity induced by vitamin A deficiency in neonatal rats. Methods:Forty healthy 5 -week -old female SPF -grade SD rats were randomly divided into 2 groups,20 rats in each group.The female rats were respectively fed with normal AIN -93Mstoyer(normal control group)and AIN -93Mstoyer lacking in vitamin A(vitamin A defi-ciency group,VAD group).Meanwhile,20 healthy male SPF -grade SD rats were recruited and fed with AIN -93Mstoyer in separate ca-ges.Ten weeks later,serum content of vitamin A were detected in female rats by enzyme -linked immunosorbent assay to identify the rat models.After successful modeling,the rats were mated according to the female -male ratio of 2:1 .In the next morning,the female rats were examined in vaginal plug and rats which had vaginal plug were recorded as E0 d.At 1 1 .5 days after gestation,half of the pregnant rats were anesthetized and sacrificed.The belly of each pregnant rat were cut open and two embryos were fetched out and their capsules were stripped.The embryos were placed in liquid nitrogen and then were transferred to refrigerator and reserved at -80 ℃.On 21 st gestational day,neonatal rats were anesthetized and sacrificed.Their skins and fascias were stripped and their internal organs were removed.Then the neonatal rats were reserved in 95% alcohol.The qRT -PCR assays was used for detecting the Hoxd3 mRNA expression in embryos and the bicolor staining method was used for observing the development of occipito -cervical bone of neonatal rats.Results:The serum contents of vitamin A were obviously lower in the VAD group compared to the control group(21 4 +/-39 vs 432 +/-54 ng/mL,t =1 4.636,P =0.000).The Hoxd3 mRNA expression in embryos was obviously lower in the VAD group compared to the control group(0.58 +/-0.1 1 vs 1 .03 +/-0.1 4,t =1 0.407,P =0.000).Normally developed occipital bone,atlas and axis and right contraposition of atlanto -axial joint were found in the control group.However,in the VAD group the angle betwwen posterior arch of atlas and posterior margin of great occipital foramen increased ,and contraposition of atlanto -axial joint was abnormal.Atlantoaxial angle and intervertebral space increased,while the height and width of vertebral arch and spinous process of C2 -C7 decreased.Conclusion:Vitamin A deficiency can lead to occipito -cervi-cal dysplasia in neonatal rats,which may be related to decrease in Hoxd3 mRNA expression in embryonic development period.However,the specific molecular mechanism needs to be further studied.
