Induction of cleft palate in aniline hydrochloride-treated rats: Possible effect of maternal methemoglobinemic hypoxia

ABSTRACT  Aniline hydrochloride (AH), a methemoglobin formation-stimulating substance, at a dosage level of 520 mg/kg which does not induce apparent fetal death, was injected subcutaneously into pregnant rats once on day 14, 15 or 16 of gestation in order to assess the stage specificity of cleft palate induction. Also, doses of 260, 390, 520 and 650 mg/kg were administered to pregnant rats on day 15 of gestation, and the dose-response relationships with respect to fetal cleft palate and maternal methemo-globinemia induction were studied. In the stage-specificity study, paleness, decreased body weight gain and elevated methemoglobin concentration were noted in the dams treated with AH. Upon fetal examinations, although reduced body weight was noted in all AH-treated groups, cleft palate was observed only in fetuses from those dams treated on day 15 of gestation. In the dose-dependency study, AH induced maternal methemoglobinemia, decreased fetal body weight and increased the incidence of cleft palate dose dependently when administered at dosage levels of 260, 390, 520 and 650 mg/kg on day 15 of gestation. Additionally, administration of methylene blue, a methemoglobinemia-preventing substance, to the AH-treated dams ameliorated maternal methemoglobinemia and reduced the incidence of fetal cleft palate. In summation, it is considered that AH stage-specifically induces cleft palate in rats and that cleft palate is caused not by a direct teratogenic effect of AH but by maternal hypoxia due to methemoglobinemia.     

Cardiovascular malformations induced by prenatal exposure to phenobarbital in rats

The effects of prenatal exposure to phenobarbital (PB) on the cardiovascular system were examined in rat fetuses and pups. PB was administered at a dose of 80 or 120 mg/kg/day by gavage to Sprague Dawley (SD) rats on two consecutive gestational days (GD): 7-8, 8-9, 9-10, or 10-11. Fetuses were examined for cardiovascular malformations on GD 20. In addition, pups were examined for PB-induced cardiovascular malformations. Incidences of ventricular septal defect (VSD), overriding aorta, double outlet right ventricle and transposition of great arteries were significantly increased in the fetuses whose dams were administered PB at 120 mg/kg on GD 8-9, 9-10 or 10-11. GD 8-11 was the critical period for the cardiovascular malformations associated with administration of PB in rats. Various types of cardiovascular malformations were detected in pups from the PB-administered dam. Severe cardiovascular malformations induced by PB caused deaths on early postnatal days. However, slight malformations such as isolated VSD persisted until weaning, and did not affect postnatal viability.     

母鼠高脂饮食增加子代大鼠罹患代谢性疾病的危险性

目的观察母代高脂饮食和后天高脂饮食对子代大鼠代谢状态的不同影响,以探索母代高脂饮食与子代大鼠成年后代谢性疾病发生的关系。方法将成年SD大鼠分成对照组（C组）和高脂饮食组（H组）,至10周龄时交配怀孕生育子代大鼠,子代大鼠断乳后每组再分成高脂饮食组（HH组,CH组）和普通饮食组（HC组,CC组）,分别至子代大鼠3周龄和8周龄时观察其代谢相关性指标以及肝脏病理表现。结果H组母鼠孕前体质量、糖耐量曲线下面积、总胆固醇及三酰甘油均明显高于C组母鼠（P<0.05）;3周龄时H组母鼠所生育并喂养的子代大鼠其体质量明显高于C组母鼠生育的子代大鼠（P=0.002）,但两者的糖耐量曲线下面积差异无统计学意义（P＞0.05）;至8周龄时各组子代大鼠空腹血糖和胰岛素水平差异无统计学意义,而母鼠为H组的子代大鼠体质量和糖耐量曲线下面积大于母鼠为C组子代大鼠（母代饮食主效应P分别为0.024和0.013）,HH组和CH组子代大鼠糖耐量曲线下面积亦大于HC组和CC组的子代大鼠（子代饮食主效应P=0.041）;8周龄时HH组和CH组子代大鼠血清总胆固醇、三酰甘油和低密度脂蛋白水平均高于HC组和CC组（子代饮食主效应P分别为0.008,0.007和0.000）;8周龄时HH组、HC组和CH组子代大鼠表现出不同程度的脂肪肝;CC组子代大鼠肝脏镜下结构均显示正常。结论母代高脂饮食可以引起子代大鼠成年后体质量增加,糖耐量减退以及肝脏脂肪变,增加子代发生代谢性疾病的危险性。     

Cardiovascular malformations in rat fetuses with oesophageal atresia and tracheo-oesophageal fistula induced by adriamycin

Associated congenital anomalies have emerged as the most significant prognostic factor in babies born with oesophageal atresia and/or tracheo-oesophageal fistula (OA-TOF). The most frequently encountered groups of anomalies are cardiovascular (CV) and gastrointestinal, the former being more significant from a prognostic point of view. Some, such as a right-sided aortic arch (RAA), vascular ring, or major heart defects, may alter the timing and surgical approach for the repair of OA-TOF or adversely affect the prognosis. The rat fetal OA model induced by adriamycin (Adr) has been described previously. In the present experiments, information was sought regarding the incidence and type of CV abnormalities in fetal rats obtained from Adr-treated dams. OA-TOF was induced in 24 of 36 fetal rats from Adr-treated dams. DV abnormalities were found in 18 (75%) OA-TOF fetuses and 10 (83%) Adr-treated fetuses without OA-TOF. The difference was not significant (P > 0.05). The most frequently found anomalies were ventricular and atrial septal defects. A RAA was present in 8/36 fetuses and a double aortic arch in 2/36. A patent ductus arteriosus was present in all treated fetuses compared with two-thirds of controls. The findings in the present study emphasise the importance of CV anomalies in association with OA, and reinforce the value of the Adr-induced rat fetal OA model by adding to our knowledge of the basic embryogenesis of both OA and CV anomalies.     

缺氧缺血新生鼠脑内乙酰胆碱水平的变化

目的 探讨缺氧缺血对新生鼠中枢胆碱能系统的远期影响及治疗措施。方法 采用碱性羟胺比色法对缺氧缺血新生鼠（7日龄）损伤后即刻及其14d后左、右脑乙酰胆碱（Ach）含量进行测定,并观察了损伤后即刻及3d后腹腔内连续7d注射4µg碱性成纤维生长因子（bFGF）对脑内Ach含量的影响。结果 缺氧缺血损伤后即刻,在脑Ach含量显著降低,（P﹤0.05）,于14d后仍未恢复;bFGF治疗对损伤后脑内Ach含量无显著影响。结论 缺氧缺血可损伤新生鼠中枢胆碱系统功能,此影响可持续至21日龄。bFGF对胆碱能损伤无明显保护作用。     

未成熟大鼠出生后感染对脑白质中SOX10蛋白表达的影响

目的研究未成熟大鼠出生后感染对脑白质中SOX10蛋白表达的影响。方法将96只新生SD大鼠随机分为缺氧组、脂多糖组和对照组;脂多糖组及缺氧组新生大鼠于生后第3、6天予以腹腔注射脂多糖0.25 mg/kg,对照组注射等量的生理盐水;缺氧组在注射脂多糖同时,自生后第3天起置于常压缺氧舱中,连续37℃恒温水浴,以2 L/min的速度向缺氧舱中输入含有8%O2和92%N2的混合气体,通气2 h;分别于生后第7、10、14、21天,每次每组处死8只大鼠,取脑白质,HE染色后光镜下观察脑白质区病理变化;通过免疫组织化学方法及Western-Blot检测脑白质区SOX10蛋白的表达变化,用Western-Blot检测Toll样受体-4(TLR-4)的表达变化。结果脂多糖组、缺氧组的SOX10阳性细胞数、SOX10和TLR-4蛋白表达均在第7天开始增加,第10天时达高峰,之后逐渐下降,各时间点间两两比较差异均有统计学意义(P0.05);对照组在各时间点均只见少量阳性细胞以及SOX10和TLR-4的少量表达,差异无统计学意义(P0.05)。在不同时间点,脂多糖组、缺氧组和对照组三组之间SOX10阳性细胞数、SOX10和TLR-4蛋白表达的差异均有统计学意义(P均=0.000);均为缺氧组最高,其次为脂多糖组;各时间点两两比较,差异均有统计学意义(P0.05)。结论未成熟大鼠出生后感染可引起脑损伤,缺氧与感染双重因素同时存在可加重脑损伤程度,而高表达SOX10蛋白可能有保护作用。     

Syndactyly lethal: New mutation with multiple malformations occurring in Sprague Dawley rats

We previously found newborns exhibiting syndactyly of both fore‐ and hindlimbs in a litter from a pair of Sprague Dawley rats. Continuous breeding of the parental animals yielded pups with the same anomaly in following litters, suggesting that the syndactyly was genetic in origin. In the present study, as all the syndactylous pups died on postnatal day 0, we conducted genetic analyses using 30 phenotypically normal female progeny and the sire. The females were subjected to caesarean section on day 20 of gestation and the fetuses were examined for the phenotypes. The results of the mating experiments suggest that the mutant phenotype is caused by a single autosomal recessive gene at a homozygous condition. As homozygous mutants are lethal at the neonatal stage, the mutant gene was named syndactyly lethal, gene symbol syl. The mutant rats have multiple abnormalities, such as syndactyly, micrognathia, fused/absent/small lung lobes, absent kidney and ureter, small spleen, small uterus, fused phalanges, sternoschisis, absent/detached rib, and splitting/fused/absent/small thoracic vertebra, some of which must be the cause of death on postnatal day 0. This mutant is considered to be useful for investigating the mechanisms and/or pathogenesis of syndactyly, as well as the accompanying malformations.     

妊娠期糖尿病胎鼠心脏T-同源盒转录因子表达研究

目的 探讨T-同源盒转录因子5（TBX-5）在母鼠妊娠糖尿病（GDM）胎鼠心脏中的分布、表达规律及其在胎鼠心脏发育异常中的作用。方法 将成年SD雌鼠随机分成GDM组和对照组,通过阴道涂片确定怀孕后分别注射链脲佐菌素（STZ）及柠檬酸-柠檬酸钠缓冲液（PBS）制备GDM模型,于孕0、3、6、9、12、15、19天对母鼠进行尾静脉采血测血糖,并分别于孕后第12、15、19天进行胎鼠心脏取材,HE染色观察心脏组织病理变化,免疫组化检测TBX-5的表达并进行半定量检测。结果 与对照组相比,GDM组胎鼠出现液化吸收胎、死胎、畸心胎的例数增多（P〈0.01）。在大体观及高倍镜下,GDM组12、15、19天胎鼠心脏发育异常比例多于对照组（P〈0.01）。在正常对照组及GDM组各亚组中,TBX-5均在第12天时表达最强,明显高于15、19天（P〈0.01）。TBX-5在15天及19天时的表达无明显差别（P〉0.05）。孕12、15、19天GDM组TBX-5在心肌细胞表达显著增强,与相应时间点的对照组相比均明显增高（P〈0.01）。结论 TBX-5蛋白在妊娠糖尿病母鼠孕12、15、19天三个观察点胎鼠心脏的表达水平明显增高,提示其过度表达可能与妊娠期糖尿病致胎鼠心脏畸形的发生存在关联。     

缺氧缺血脑损伤时新生大鼠脑皮质神经细胞能量代谢的改变

目的　近年来的一些研究表明 ,线粒体能量代谢异常是新生儿缺氧缺血性脑病 (HIE)发病机制的重要环节之一。该实验研究新生大鼠缺氧缺血性脑损伤 (HIBD)时脑皮质神经细胞内磷酸肌酸 (PCr)、ATP和总腺苷酸池 (ATP +ADP +AMP)的变化。方法　 7日龄Wistar大鼠随机分为假手术对照组 (n =6 )和HIBD组 (n =6 0 )。HIBD组新生鼠分别于缺氧缺血 (HI)后 0 ,2 ,4 ,6 ,8,1 0 ,1 2 ,2 4 ,4 8h和 72h断头 (每个时间点 6只 ) ,取左侧顶枕部脑皮质 ;假手术对照组新生鼠于假手术后 4h断头。用高效液相色谱方法检测脑皮质高能磷酸物质的含量。结果HIBD组新生鼠HI后 0hPCr,ATP和ATP +ADP +AMP即低于正常对照组 (P <0 .0 1或 0 .0 5 ) ,并于缺氧缺血后0～ 6h出现第 1次低谷 ,分别为对照组的 5 1 % ,71 % ,5 0 % ,缺氧缺血后 8～ 1 2h分别恢复到对照组的 92 % ,83%和83% ,与对照组比较差异无显著性 (P >0 .0 5 ) ;以后又下降到低于正常对照组 (P <0 .0 1 ) ,并于缺氧缺血后 2 4～4 8h下降到第 2次低谷 ,分别为对照组的 5 8% ,6 1 %和 32 %。结论　新生大鼠HIBD后 2 4～ 4 8h皮质神经细胞出现第 2次能量代谢衰竭 ,能量代谢变化可作为判断缺氧缺血性脑损伤干预手段是否有效的观察指标。     

Impaired transient elevation of blood hemoglobin in response to acute hypoxia in neonates with asplenia

BACKGROUND: It has been shown that acute hypoxia induces the transient elevation of blood hemoglobin concentration ([Hb]) as a consequence of sympathetic-mediated splenic contraction to maintain the supply of oxygen, and splenectomy abolishes this phenomenon. The purpose of the present paper was to determine, retrospectively, whether the ability of transient elevation of [Hb] against acute hypoxia would be impaired in neonates with asplenia. METHOD: Eleven neonates who underwent surgery from 1998 to 2003 were enrolled in this retrospective study. They were divided into two groups: (i) five patients with asplenia syndrome with cyanotic congenital heart disease (asplenia group); and (ii) six patients with hypoplastic left heart syndrome who needed nitrogen gas inhalation therapy (HLHS group). In the asplenia group simultaneous data of arterial oxygen saturation (Sao(2)) and [Hb] were obtained before and after the temporary unexpected decrease of percutaneous arterial oxygen saturation. In the HLHS group they were obtained before and after nitrogen gas administration therapy. The arterial oxygen content (Cao(2)) changes and the ratio of Cao(2) change (Cao(2) after hypoxia divided by Cao(2) before hypoxia) were also calculated. RESULTS: In the asplenia group [Hb] was unchanged (12.9 +/- 1.6 g/dL to 12.8 +/- 1.4, n.s.) and Cao(2) was decreased (14.5 +/- 1.6 mL/dL to 11.9 +/- 1.1, P = 0.018). In the HLHS group [Hb] was increased (14.6 +/- 1.3 g/dL to 15.4 +/- 1.5, P = 0.028), but Cao(2) was changed (18.2 +/- 2.2 mL/dL to 16.7 +/- 3.0, P = 0.043). The ratio of Cao(2) change for the HLHS group was significantly different from that of the asplenia group (0.92 +/- 0.10, 0.83 +/- 0.10, respectively, P = 0.02). CONCLUSIONS: Patients with asplenia syndrome have some disadvantage regarding this protective mechanism against acute hypoxia.     

Are elevated cerebrospinal fluid levels of IL-6 in sudden unexplained deaths,infectious deaths and deaths due to heart/lung disease in infants and children due to hypoxia?

Many SIDS cases probably die after periods of hypoxia and it has been shown that hypoxia may stimulate IL-6 production. The purpose of this paper was to examine if there were any correlations between hypoxanthine in vitreous humour and II-6 in CSF. The concentration of IL-6, IL-1 β and TNF α in cerebrospinal fluid of 50 Sudden Infant Death syndrome (SIDS) cases, 9 borderline SIDS cases, 18 infectious deaths, 8 violent deaths and 22 cases with heart/lung disease were measured by ELISA. The hypoxanthine (Hx) vitreous humour concentrations in the same groups were determined by high performance liquid chromatography. The IL-6 levels in cases of infectious death, heart/lung disease and borderline cases were significantly higher than in the SIDS cases ( p < 0:01). The Hx levels were in the same range in cases of SIDS, borderline SIDS and infectious death, and they were significantly higher than the levels in cases of violent death and heart/lung disease ( p < 0:01). There was no correlation between hypoxanthine and IL-6 in any of the groups. In the cases studied IL-6 elevation is probably not induced by hypoxia, but is rather a result of immunological stimulation.     

First trimester diagnosis of iniencephaly associated with fetal malformations and trisomy 18: report of a new case and gene analysis on folate metabolism in parents

Iniencephaly is a rare congenital malformation consisting of a complex alteration of the embryonic development occurring around the third post-fertilization week and characterized by a hyper-retroflexion of the cephalic pole. We report a case of iniencephaly associated with acrania-encephalocele, spina bifida and abnormal ductus venosus in a fetus with trisomy 18 diagnosed at 12 week's gestation in a 41-year-old woman. A co-occurrence between aneuploidy and iniencephaly was documented and polymorphisms on folate metabolism-related genes were investigated in the parents to assess possible etiologic factors and recurrence risk for neural tube defects (NTD). An homozygous state for the MTRR polymorphism was diagnosed in the mother, identifying a clinical risk for NTD. Once iniencephaly or any other NTD are suspected, genetic analysis, second level ultrasound and fetal karyotype are recommended. Autopsy should also be performed in all cases of early ultrasound-based diagnosis of fetal malformations.     

室间隔缺损合并肺动脉高压肺血管床功能状态评估指标的研究

目的探讨室间隔缺损合并肺动脉高压肺血管床功能状态的评估指标。方法对室间隔缺损合并重度肺动脉高压的患儿，于心导管术中应用酚妥拉明，将轻度全肺循环阻力（ＴＰＲ）增加的２７例与重度全肺循环阻力增加的１２例患儿的试验结果进行比较。结果两组患儿的单一肺动脉压降幅（Ｐｐ降幅）分别为２．３ｋＰａ（１７ｍｍＨｇ，１ｋＰａ＝７．５ｍｍＨｇ）和２．２ｋＰａ，Ｐ＞０．０５，差异无显著意义；而肺动脉压降幅（Ｐｐ降幅）／体循环压降幅（Ｐｓ降幅）的比值，试验前后肺动脉血氧饱和度变化值这二项指标的差异有非常显著意义（Ｐ＜０．０１）。Ｐｐ降幅／Ｐｓ降幅比值与全肺循环阻力的相关性良好［ｒ＝－０．８９９），Ｙ（ＴＰＲ）＝２６６８－１８９２Ｘ（Ｐｐ降幅／Ｐｓ降幅）］。结论用酚妥拉明作扩血管降压试验时，单一的肺动脉压降幅不能完全反映肺血管床的功能状态；而Ｐｐ降幅／Ｐｓ降幅比值，试验前后肺动脉血氧饱和度变化值这二项指标对室间隔缺损合并肺动脉高压患儿的肺血管床功能状态的评估，对手术适应证的选择具有一定指导意义。     

3p25.3p25.2 染色体杂合缺失伴矮小及多发畸形1 例遗传学特征与临床表型分析

目的分析3号染色体p25.3p25.2(3p25.3p25.2)片段缺失的临床表型及分子遗传学特点。方法回顾分析1例3p25.3p25.2染色体片段缺失患儿的临床资料,分析其临床表型及分子遗传学特征。结果患儿,男,1岁4个月。宫内发育迟缓,重度矮小、全面生长发育落后、语言发育迟缓、特殊面容伴多发畸形(小头畸形、小下颌、长人中、低耳位、双侧耳前瘘管等)、先天性十二指肠闭锁、肠旋转不良,先天性心脏病、隐睾、龟头裸露、肌张力低下、婴儿期喂养困难、睡眠障碍、甲状腺功能减低。患儿染色体核型分析46,XY。基因芯片分析示3p25.3p25.2区域存在一段3 327 kb的杂合缺失,共39个基因缺失。结论 3p25.3p25.2区域3 327 kb杂合缺失,致SETD5、VHL、FANCD2基因缺失导致该患儿临床表型。     

PAF antagonist BN-52021 reduces intercellular adhesion molecule-1 expression and oxidative stress in rats with reperfusion damage due to unilateral testicular torsion

The aim of this study was to determine the effects of specific platelet-activating factor (PAF) antagonist BN-52021 on intercellular adhesion molecule-1 (ICAM) expression and oxidative stress in rats with reperfusion damage due to unilateral testicular torsion. Thirty male Sprague–Dawley rats were separated into three groups, each containing ten rats. A sham operation was performed in group 1 (control). In group 2 [ischemia-reperfusion (I-R)/untreated], 1-h detorsion of the testis was performed after 6 h of unilateral testicular torsion. In group 3 (I-R/BN-52021), after performing the same surgical procedures as in groups II, BN-52021 was given intravenously at the starting time of reperfusion. In all experimental rats, ipsilateral orchiectomies were performed for histological examination and measuring the tissue levels of malondialdehyde (MDA), superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSH-Px). MDA values and the testicular injury score decreased and SOD, CAT and GSH-Px values increased in the I-R/BN-52021 treated group compared to in the I-R/untreated group. Most of the specimens in the I-R/BN-52021 treated group showed grade-I testicular injury. However, the injuries in the I-R/untreated rats varied between grades III and IV. An ICAM-1 expression was intensive in the interstitial spaces and basement membrane of the tubuli seminiferi, of testicular tissue in the I-R/untreated group. However, an ICAM-1 expression was mild in the I-R/BN-52021 group. BN-52021 may play an important role in the immunohistochemical expression of adhesion molecule ICAM-1 and may reduce oxidative stress in rats with reperfusion damage due to unilateral testicular torsion.     

Successful treatment with eculizumab for posterior reversible encephalopathy syndrome due to underlying transplant‐associated thrombotic microangiopathy in patients transplanted for sickle cell disease

Preexisting endothelial dysfunction and vascular injury sustained during allogeneic hematopoietic cell transplantation (HCT) increases risk for endothelial injury‐related complications such as posterior reversible encephalopathy syndrome (PRES) and transplant‐associated thrombotic microangiopathy (TA‐TMA) in patients with sickle cell disease (SCD). We report two patients with SCD who developed PRES following allogeneic HCT. In both patients, PRES‐related symptoms resolved only after a diagnosis of TA‐TMA was established and eculizumab therapy was initiated. Renal manifestations at diagnosis included non‐nephrotic range proteinuria and hypertension. This report highlights the importance of screening PRES‐affected SCD HCT recipients for TA‐TMA as usual treatment strategies may be inadequate.