维生素A缺乏导致器官发育异常的研究进展

综述了维生素A缺乏导致脑组织、免疫系统的损伤和胚胎发育异常的研究进展，并简要概述了可能的作用机制。     

维生素A缺乏引起的角膜软化症的研究

由于维生素Ａ（ＶＡ）缺乏导致营养不良性角膜软化症是一种严重的小儿致盲性眼病。多由全身营养紊乱引起。资料表明［１－３］ＶＡ在体内有多种功能,ＶＡ缺乏与眼病有重要关系。由于我国人民生活水平不断提高,此病发病率大大下降,但在边远山区、农村仍屡见不鲜。但目前...     

维生素A缺乏胎鼠作为先天性心脏病动物模型的实验研究

目的　探讨将维生素Ａ缺乏（ＶＡＤ）胎鼠作为先天性心脏病动物模型的可行性。方法取１１－１９ｄ不同胎龄正常及ＶＡＤ胎鼠心脏经石蜡包埋、切片及 ＨＥ染色观察其发育情况。结果 １．实验组饲料含维生素Ａ（ＶＡ）７μｇ／１００ｇ,经ＶＡＤ饮食喂养后实验组大鼠血清ＶＡ水平明显低于对照组［（０．１６８±０．０５９）μｍｏｌ／Ｌ Ｖｓ（２．１８±０．２３）μｍｏｌ／Ｌ,ｔ＝３２．８８, Ｐ＜０．００１］。 ２．大鼠死亡百分比：饲养于屏障系统的ＶＡＤ大鼠死亡百分比较饲养于开放系统中的要低４．６倍（１０％ Ｖｓ ４５．８３％．ｘ ２＝１６．６４, Ｐ＜０．００１）,对照组为０。 ３．实验组大鼠受孕百分比及每只孕鼠产仔数均低于对照组［５８．３３％ Ｖｓ ８１．５％, ｘ ２＝４．３７,Ｐ＜０．０５：（６．９７±２．７９） Ｖｓ（１３ ±１．０５）,ｔ＝７．１６, Ｐ＜０．００１］。 ４．经切片观察１１～１５ ｄ胎龄胎鼠实验组心脏出现明显发育延迟的占３６．６７％, １６～１９ ｄ胎龄胎鼠实验组心脏畸形占４１．４３％,血管异常占１８．５７％。结论ＶＡＤ胎鼠可用来作为先天性心脏病动物模型,但需改进饲养环境以减少异常死亡。     

维生素A缺乏对大鼠肺脏发育的影响

目的探讨维生素A缺乏（vitamin A deficiency,VAD）对大鼠肺脏发育的影响及其可能的机制。方法建立VAD大鼠模型,取出生后2周正常及VAD幼鼠肺脏,HE、Weigert染色分别观察其肺脏组织形态变化和弹力纤维变化;Image Pro Plus5．0软件做肺脏形态学定量分析,蛋白质免疫印记检测大鼠肺组织MMP-9及TIMP-1蛋白含量变化。结果①VAD组大鼠肺上皮细胞堆积,出现明显的肺气肿,并发弥散的间质性肺炎,肺组织弹力纤维显著减少;②VAD组大鼠肺MMP-9的蛋白质表达水平明显高于对照组,TIMP-1水平变化不大,MMP-9与TIMP-1的比值升高。结论VAD导致大鼠肺脏正常发育受损,肺脏组织形态异常,可能通过上调MMP-9的表达诱发大鼠肺气肿。     

原代肝细胞培养与动物实验对比研究维生素A缺乏对贫血的影响

目的：通过原代大鼠肝细胞培养和动物实验模型的对比来研究维生素A对贫血的影响。方法：将SD大鼠按体重随机分为4组，每组13只，分别为Fe+维生素A（Ⅰ组）、Fe+维生素A（Ⅱ组）、Fe+维生素A（Ⅲ组）、Fe+维生素A（Ⅳ组）。实验动物喂养10周后处死并分离肝脏；原代大鼠肝细胞按Seglent二步消化法分离后，分为严重缺乏组（A）、边缘缺乏组（B）、正常组（C）、治疗组（D），依次给予0、0.5、1.0、50 μmol/L 的视黄酸（全反式）培养96 h。用Western-Blot法测量转铁蛋白受体（TFR）、用ELISA方法测量铁蛋白（Fn）蛋白的表达，逆转录聚合酶链反应（RT-PCR法）检测肝TFR mRNA、Fn mRNA的表达。结果：当维生素A缺乏时，大鼠肝TFR mRNA表达上调，使TFR蛋白增加，而Fn mRNA表达下调，Fn蛋白表达下降，差异具有统计学意义（P<0.05）。结论：维生素A补给可显著减轻铁缺乏造成的利用障碍干扰。     

维生素A缺乏对缺铁性贫血的影响——体内外实验对比研究

目的：研究维生素A缺乏对缺铁性贫血的影响。方法：体内实验将SD大鼠按体重随机分为4组,分别为Fe＋维生素A-Ⅰ组、Fe＋维生素A-Ⅱ组、Fe+维生素A－Ⅲ组、Fe－维生素A－Ⅳ组。实验动物喂饲8w后处死后取肝脏。体外实验将大鼠原代肝细胞按Seglen法分离后,随机分为严重缺乏组A、边缘性缺乏组B、正常生理组C、治疗剂量组D,分别给予0、0.5、1.0、50μmol/L 的全反式视黄酸培养72h。用逆转录—聚合酶链反应（RT-PCR法）检测肝脏的转铁蛋白受体（TFR）mRNA、铁蛋白（Fn）mRNA的表达。结果：体内外实验大鼠维生素A缺乏时,肝脏TFR mRNA表达增强,而Fn mRNA表达下降,差异具有统计学意义 （P<0.05）。结论：维生素A缺乏可以使大鼠肝脏 TFR mRNA表达增强而Fn mRNA表达下降,而补充维生素A后铁缺乏造成的很多不利影响都显著减轻了。     

维生素A缺乏大鼠致敏后免疫功能低下对肺炎的影响机制

目的探讨维生素A（VA）缺乏对致敏后免疫细胞发育、功能的影响机制。方法：低VA饲料喂养大鼠8周时用卵蛋白致敏并分组。VAS组隔天口服vA100u,治疗3次;VAD组与正常组口服大豆油。用免疫组织化学法观察细胞角蛋白19（CK19）、病理学检查肺与脾病理变化、ELISA法检查血清Th因子水平等。结果VAD组胸腺萎缩,胸腺上皮细胞（TEC）肥大、聚集、CK19少。血IL—10、IL-4、胸腺索a-1升高。肺感染病灶多,肺泡隔浸润细胞较少与红细胞渗出多。VAS组胸腺细胞增生,TEC内CK193上调,血IFN—γ水平较低、IL-4、IL-10明显下调。小气道单层纤毛柱状上皮正常,管壁淋巴细胞浸润较重,肺泡区巨噬细胞多、渗出红细胞少。结论VA缺乏并致敏后胸腺TEC、胸腺细胞等发育和功能异常。补充VA改善胸腺TEC结构、促进淋巴细胞增生与非特异免疫功能,保护呼吸道黏膜与肺泡上皮。     

β-连环素在维生素A缺乏及吸烟大鼠气道上皮鳞状化生中的作用

目的研究大鼠气道上皮鳞状化生时β-连环素(-βcatenin,-βcat)及其相关基因表达的变化,探讨-βcat与气道上皮鳞化及癌变的关系。方法采取缺乏VitA饲料饲养(12周)联合被动吸烟(4周)的方法建立支气管上皮鳞状化生模型,应用免疫组织化学(SABC法)和原位杂交技术研究-βcat、c-myc、增殖细胞核抗原(PCNA)以及-βcat mR-NA在气道上皮的表达。结果实验组大鼠支气管上皮细胞层次明显增多,其中4只大鼠支气管上皮出现明显的复层鳞状上皮化生。实验组支气管上皮-βcat在胞膜的表达减少(P<0.01),其中有6例在细胞质表达。实验组与对照组胞质中β-cat mRNA表达无明显差异(P>0.05)。实验组支气管上皮细胞核c-myc表达增加(P<0.01),PCNA阳性率明显增加(P<0.01)。结论-βcat、c-myc的表达异常参与了VitA缺乏与吸烟所致的大鼠支气管上皮鳞状化生的形成。     

大鼠维生素A和铁的相互作用关系对缺铁性贫血的影响

目的：通过研究维生素A缺乏和铁缺乏的相互作用来探讨缺铁性贫血的发生机制。方法：第一阶段，将44只SD大鼠随机分为4组，分别为Fe＋维生素A+Ⅰ组、Fe-维生素A+Ⅱ组、Fe-维生素A+Ⅲ组、Fe-维生素A-Ⅳ组；实验动物喂养2个月后处死，测定视黄醇（血清）、血红蛋白（HB）、维生素A、视黄基酯（肝脏），并采用RT-PCR法测定视黄醇结合蛋白mRNA（RBP mRNA）的表达。第二阶段，将SD大鼠按称重结果随机分4组，分别为Fe＋维生素A+Ⅴ组，Fe+维生素A-Ⅵ组，Fe+维生素A-Ⅶ组，Fe-维生素A-Ⅷ组。大鼠喂饲2个月后处死留取肝脏，用半定量PCR的方法测量铁调节蛋白2（IRP2）、铁蛋白（Fn）、转铁蛋白受体（TFR）转录mRNA的表达量。结果：第一阶段制造铁缺乏动物模型，造模后大鼠的血清视黄醇，肝脏维生素A储备量开始降低，同时RBP储备含量也有下降走势，经测肝脏RBP表达量下降明显；第二阶段实验原代肝细胞缺乏维生素A时，肝脏IRP2 mRNA、TFR mRNA表达同时增强，而Fn mRNA却表达下降，差异均具有统计学意义（P＜0.05）。结论：维生索A和铁相互缺乏时，可相互影响彼此的代谢和营养状况，在得到补充后，相互造成的很多利用障碍的影响（如贫血）都显著减轻。     

北京市6-13岁儿童维生素A营养水平及影响因素

目的：了解北京市6~13岁儿童维生素A营养水平及影响因素。方法：于2017年采用多阶段分层抽样对北京市982名6~13岁儿童进行调查,分析血清维生素A水平与食物种类、视黄醇活性当量摄入量、血生化指标、体格等指标的关联。结果：血清维生素A缺乏率和边缘缺乏率分别为0.10%、10.5%;膳食维生素A摄入量低于平均需要量的比例为71.6%（以视黄醇活性当量计）;血清维生素A水平与年龄、BMI、血红蛋白水平、铁蛋白水平、蔬菜摄入量存在显著正相关（P<0.05）,与超敏C反应蛋白存在显著负相关（P<0.05）。结论：北京市6~13岁儿童维生素A整体缺乏率较低,但仍有部分儿童处于边缘缺乏状态。膳食维生素A摄入量按视黄醇活性当量计算不能满足膳食营养素推荐供给量的风险高,蔬菜摄入量可能是血清维生素A水平的影响因素之一。     

Interaction between hindbrain and spinal networks during the development of locomotion in zebrafish

Little is known about the role of the hindbrain during development of spinal network activity. We set out to identify the activity patterns of reticulospinal (RS) neurons of the hindbrain in fictively swimming (paralyzed) zebrafish larvae. Simultaneous recordings of RS neurons and spinal motoneurons revealed that these were coactive during spontaneous fictive swim episodes. We characterized four types of RS activity patterns during fictive swimming: (i) a spontaneous pattern of discharges resembling evoked high-frequency spiking during startle responses to touch stimuli, (ii) a rhythmic pattern of excitatory postsynaptic potentials (EPSPs) whose frequency was similar to the motoneuron EPSP frequency during swim episodes, (iii) an arrhythmic pattern consisting of tonic firing throughout swim episodes, and (iv) RS cell activity uncorrelated with motoneuron activity. Despite lesions to the rostral spinal cord that prevented ascending spinal axons from entering the hindbrain (normally starting at approximately 20 h), RS neurons continued to display the aforementioned activity patterns at day 3. However, removal of the caudal portion of the hindbrain prior to the descent of RS axons left the spinal cord network unable to generate the rhythmic oscillations normally elicited by application of N-methyl-d-aspartate (NMDA), but in approximately 40% of cases chronic incubation in NMDA maintained rhythmic activity. We conclude that there is an autonomous embryonic hindbrain network that is necessary for proper development of the spinal central pattern generator, and that the hindbrain network can partially develop independently of ascending input.     

斑马鱼胚胎背腹轴建立的分子机制

脊椎动物胚胎发育起始于体轴的建立,是胚胎早期发育过程中最重要的事件之一。Wnt、BMP、Nodal和FGF等多个信号通路协同调控细胞分化和细胞运动,促进胚胎胚层的形成和空间上的分离,调控胚胎背腹轴、前后轴和左右轴线的分化,为胚胎进一步发育勾勒出蓝图。本文主要综述斑马鱼胚胎背腹轴建立的分子机制,包括背部组织中心简介;母源Wnt/β-catenin信号调控背部组织中心形成的分子机制;BMP信号调控背腹轴建立的分子机制。     

Retinoic acid causes abnormal development and segmental patterning of the anterior hindbrain in Xenopus embryos.

Retinoic acid is a very potent teratogen and has also been implicated as an endogenous developmental signalling molecule in vertebrate embryos. One of the regions of the embryo reliably affected by exogenously applied RA is the hindbrain. In this paper, we describe in detail the hindbrain of Xenopus laevis embryos briefly treated with various levels of RA at gastrula stages. Such treatments lead to development of embryos with loss of anterior structures. In addition, RA has a general effect on rhombomere morphology and specific effects on the development of the anterior rhombomeres. This effect is demonstrated using neurofilament antibodies, HRP staining and in situ hybridisation using a probe for expression of the Xenopus Krox-20 gene. Anatomically it is evident that the development of the hindbrain normally anterior to the otocyst (rhombomeres 1-4) is abnormal following RA treatment. Sensory and motor axons of cranial nerves V and VII form a single root and the peripheral paths of V and VII and IX and X are also abnormal, as is the more anterior location of the otocyst. These anatomical changes are accompanied by changes in the pattern of expression for the gene XKrox-20, which normally expresses in rhombomeres 3 and 5, but is found in a single band in the anterior hindbrain of treated embryos which standardly fail to generate the normal external segmental appearance. The results are discussed in terms of both the teratogenic and possible endogenous roles of RA during normal development of the central nervous system. We conclude that low doses of RA applied during gastrulation have specific effects on the anterior Xenopus hindbrain which appear to be evolutionarily conserved in the light of similar recent findings in zebrafish.     

Vitamin A deficiency results in the dose-dependent acquisition of anterior character and shortening of the caudal hindbrain of the rat embryo

The developing nervous system is particularly vulnerable to vitamin A deficiency. Retinoid has been proposed to be a posteriorizing factor during hindbrain development, although direct evidence in the mammalian embryo is lacking. In the present study, pregnant vitamin A-deficient (VAD) rats were fed purified diets containing varying levels of all-trans-retinoic acid (atRA; 0, 0.5, 1.5, 6, 12, 25, 50, 125, or 250 microg/g diet) or were supplemented with retinol. Hindbrain development was studied from embryonic day 10 to 12.5 ( approximately 6 to 40 somites). Normal morphogenesis was observed in all embryos from groups fed 250 microg atRA/g diet or retinol. The most caudal region of the hindbrain was the most sensitive to retinoid insufficiency, as evidenced by a loss of the hypoglossal nerve (cranial nerve XII) in embryos from the 125 microg atRA/g diet group. Further reduction of atRA to 50 microg/g diet led to the loss of cranial nerves IX, X, XI, and XII and associated sensory ganglia IX and X in all embryos as well as the loss of hindbrain segmentation caudal to the rhombomere (r) 3/4 border in a subset of embryos. Dysmorphic orthotopic otic vesicles or immature otic-like vesicles in both orthotopic and caudally ectopic locations were also observed. As the level of atRA was reduced, a loss of caudal hindbrain segmentation was observed in all embryos and the incidence of otic vesicle abnormalities increased. Perturbations in hindbrain segmentation, cranial nerve formation, and otic vesicle development were associated with abnormal patterning of the posterior hindbrain. Embryos from VAD dams fed between 0.5 and 50 microg atRA/g diet exhibited Hoxb-1 protein expression along the entire neural tube caudal to the r3/r4 border at a time when it should be restricted to r4. Krox-20 protein expression was expanded in r3 but absent or reduced in presumptive r5. Hoxd-4 mRNA expression was absent in the posterior hindbrain, and the rostral limit of Hoxb-5 protein expression in the neural tube was anteriorized, suggesting that the most posterior hindbrain region (r7/r8) had been deleted and/or improperly patterned. Thus, when limiting amounts of atRA are provided to VAD dams, the caudal portion of the hindbrain is shortened and possesses r4/r5-like characteristics, with this region finally exhibiting r4-like gene expression when retinoid is restricted even more severely. Thus, regions of the anterior hindbrain (i.e., r3 and r4) appear to be greatly expanded, whereas the posterior hindbrain (r5-r8) is reduced or absent. This work shows that retinoid plays a critical role in patterning, segmentation, and neurogenesis of the caudal hindbrain and serves as an essential posteriorizing signal for this region of the central nervous system in the mammal.     

Temperature and somitogenesis in embryos of the plaice (Pleuronectes platessa)

The rate of somitogenesis was observed in plaice embryos reared at 5 and 12°C. Somite formation occurred relatively later in development at 12 than at 5°C. At any given stage of development 5°C plaice embryos had significantly more somites than embryos reared at 12°C.     

Nodal coordinates the anterior-posterior patterning of germ layers and induces head formation in zebrafish explants

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The response of trout and zebrafish embryos to low and high boron concentrations is U-shaped

Fish in the embryo-larval stage of development have been shown to be sensitive to boron (B) at both ends of the dose-response curve (1,2). The present study evaluated the health effects of low and high B concentrations on rainbow trout (Oncorhynchus mykiss), a cold water species, and zebrafish (Danio rerio), a warm water species. Rainbow trout embryos were incubated from day 1 until 2 wk posthatch in Type 1 ASTM ultrapure-grade water (12.5°C) supplemented with only B (0-500 μM) as boric acid, or together with CaCO₃ (0–2 mM) to increase water hardness. Embryonic growth was stimulated by B in a dose-dependent manner at all Ca concentrations (p < 0.001). Chronic exposures below 9 μmol B/L impaired embryonic growth and above 10 mmol B/L caused death (p < 0.001). Thus, the safe range of exposure for the rainbow trout was between the adverse effect concentrations of 9 μmol B/L and 10 mmol B/L. Zebrafish were maintained for 6 mo in ultrapure water containing <0.2 μmol B/L to determine the effect of low-level exposure. High-level exposure was assessed by exposing zygotes, derived from parents maintained at 46 μmol B/L, to graded concentrations of boric acid up to a concentration of 75 mmol B/L from fertilization until they were free feeding (96 h). Fertilization occurred, but zygotes failed to survive when water contained <0.2 umol B/L (p < 0.001). Death occurred at and above 9.2 mmol B/L. Thus, the safe range of B exposure for zebrafish was between the adverse effect concentrations of 0.2 μmol B/L and 9.2 mmol B/L. The dose-response for both species was thus U-shaped. Part of this work was previously published in abstract form and presented at Experimental Biology 97, April 6–9, New Orleans, LA (Eckhert, C. [1997] Embryonic trout growth and boron exposure,FASEB J. 11, A406 [abstract]).     

Vitamin adeficiency and keratin biosynthesis in cultured hamster trachea

Summary Tracheas from vitamin A-deficient hamsters in organ culture in vitamin A-free medium developed squamous metaplasia. Addition of retinyl acetate to the medium prevented squamous metaplasia and a mucociliary epithelium was maintained. Indirect immunofluorescent staining with antikeratin antibodies AE1 and AE3 indicated positive reactions with epithelium of tracheas either cultured in vitamin A-free or retinyl acetate (RAc)-containing medium. The “stratum corneum”-like squames in metaplastic tracheas were strongly stained by AE3. Immunoprecipitation of cytoskeletal extracts from [³⁵S]methionine labeled tracheas with a multivalent keratin antiserum indicated that the concentration of keratins synthesized in tracheas cultured in vitamin A-free medium was greater than that observed in tracheas cultured in the presence of RAc. In addition, new species of keratin were expressed in tracheas cultured in RAc-free medium. Alterations in the program of keratin synthesis were clearly detectable after 1 d in vitamin A-free medium, even though squamous metaplasia was not yet obvious. Squamous tracheas were shown by immunoblot analysis to contain keratins of 50, 48, 46.5, and 45 kilodalton (kd) detected with AE1; and 58, 56, and 52 kd detected with AE3. Immunoblot analysis with monospecific antimouse keratin sera also demonstrated the presence of 60, 55, and 50 kd keratins in the metaplastic tracheas. All these various species of keratins were either absent or present in much reduced quantity in mucociliary tracheas in RAc-containing medium. Interestingly, the induction of squamous metaplasia in tracheal epithelium did not result in the expression of the 59 and 67 kd keratins which are characteristically expressed in the differentiated layers of the epidermis. Therefore, this study shows that squamous metaplasia of tracheas due to vitamin A-free cultivation is accompanied by an increase in keratin synthesis as well as by the appearance of keratin species not normally present in mucociliary tracheal epithelium.