刀鲚肌间骨新类型的发现

文章对刀鲚(Coilia nasus)的肌间骨数目、形态、分布特征及组织学结构进行了系统研究。结果表明刀鲚肌间骨的形态种类与普通鲤科鱼类没有显著差别,但其分布类别却有很大不同,刀鲚不仅有髓弓小骨、椎体小骨和脉弓小骨,且在其背部上方和腹部下方仍存在大量肌间骨,这两类肌间骨的形态为简单的"1"形和"("形,分别位于背部上方和腹部下方肌膈的皱褶部;在短颌鲚(C. brachygnathus)背部上方和腹部下方也发现这两类肌间骨的存在,统称为肌骨杆(Myorhabdoi)。刀鲚个体肌间骨数目范围为492—543,其中髓弓小骨为114—142 (■=133),椎体小骨为28—51 (■=42),脉弓小骨为138—153 (■=142),背上肌骨杆为92—135 (=■114),腹下肌骨杆为66—98 (■=89)。采用茜素红与阿利新蓝组织染色观察不同类型肌间骨的组织结构特征,结果显示肌间骨由肌膈结缔组织包被,在髓弓小骨和脉弓小骨两种类型中,肌间骨之间通过结缔组织相互连接。研究在鱼类背上肌骨杆和腹下肌骨杆的发现,是对我国鱼类肌间骨系统进化研究的进一步补充。     

异育银鲫A+系和F系肌间骨的比较分析

研究采用常规测量法和解剖法, 并结合CT透视, 比较分析了异育银鲫主养品种“中科3号”(A+系)和候选新品系F系的肌间骨的数目、形态和分布。结果表明, 6月龄和18月龄异育银鲫F系平均总肌间骨数分别为71.8±2.9和83.3±1.4, 均极显著少于相应月龄银鲫A+系总肌间骨数目(78.6±3.9和87.0±1.5)(P<0.01)。并统计了2个品系的每一肌节的平均肌间骨数目, 6月龄和18月龄异育银鲫F系的每一肌节的平均肌间骨数目极显著少于相应月龄银鲫A+系(P<0.01)。银鲫A+系和F系均有髓弓小骨和脉弓小骨, 没有椎体小骨。6月龄和18月龄银鲫F系髓弓小骨的平均数目为48.2±1.1和55.8±0.52, 均极显著少于相应月龄银鲫A+系的髓弓小骨(53.7±1.6和58.7±0.5)(P<0.01)。异育银鲫两个品系的脉弓小骨数目差异不显著; 2个品系均具有“I”形、“卜”形、“Y”形、一端多叉形、两端两分叉形、两端多叉形和树枝形7种类型的肌间骨, 髓弓小骨比脉弓小骨数量多且形状复杂。随着月龄的增长, 2个品系具有的总肌间骨数目以及复杂肌间骨数目均增加。6月龄和18月龄银鲫F系躯轴上肌节间的复杂“Y”形髓弓小骨数目均比相应月龄的银鲫A+系少, 而简单的“I”形髓弓小骨数目比相应月龄的A+系多, 表现出一种有利于食用的优势。研究结果为异育银鲫候选新品系F系提供了一个品质评价指标, 同时也为后续进行异育银鲫肌间骨遗传改良提供了形态学基础资料。     

鲢肌间小骨发育的形态学观察

分别利用形态解剖和整体骨骼染色的方法,对鲢(Hypophthalmichthys molitrix)成鱼肌间小骨的形态、分布,以及仔、稚鱼肌间小骨的形态发生和出现进行观察.结果表明,鲢肌间小骨有I形、卜形、y形、一端多叉形、两端多叉形和树枝形6种类型,肌间小骨越靠前端形态越复杂.肌间小骨在43 dpf首先出现在尾部,然后往前依次出现,到55 dpf全部出现.出现一枚髓弓小骨的仔鱼最小体长是10.50 mm,出现一枚脉弓小骨的仔鱼最小体长是13.66 mm.各种复杂形态的肌问小骨均是从I形发展而来.本文结果为今后研究肌间小骨发生的分子机制提供了形态学基础.     

不同鱼类肌间骨的骨化模式研究

为研究鱼类肌间骨的骨化模式, 采用整体骨骼染色方法, 对黄鳝(Monopterus albus)和泥鳅(Misgurnus anguillicaudatus) 2种处于不同进化地位且游动模式不同的鱼类肌间骨发生发育进行了系统观察。结果显示合鳃鱼目合鳃鱼科且游动模式属于鳗鲡模式的黄鳝在孵出后30d (体长40 mm)时, 其头部开始出现椎体小骨; 随后依次向尾部骨化, 在孵出后55d (体长约100 mm)时, 肌间骨相继出现完毕, 所有椎体小骨均为“I”形。鲤形目鳅科且游动模式为鲹科模式的泥鳅在孵出后27d (体长17 mm)时, 肌间骨开始在尾部出现, 包括髓弓小骨和脉弓小骨; 随后依次向头部骨化, 在孵出后40d (体长35 mm)时, 肌间骨全部出现, 形态包括“I”、“Y”和“卜”形。此外, 研究利用成骨细胞特异性转录因子绿色转基因荧光蛋白(Osterix GFP)斑马鱼品系, 通过观察此osterix GFP活体斑马鱼, 可发现斑马鱼肌间骨从尾部向头部依次骨化的过程。研究结果揭示鱼类肌间骨的骨化规律与其游动模式密切相关, 且肌间骨的形态种类的多态性与其游动模式和体型有一定的关系。     

鱼类Ⅰ型与Ⅱ型胶原蛋白基因系统进化及其在有/无肌间骨代表鱼中的表达比较分析

研究在分析鱼类Ⅰ型与Ⅱ型胶原蛋白基因系统进化基础上, 以有肌间骨的团头鲂(Megalobrama amblycephala)和无肌间骨的尼罗罗非鱼(Oreochromis niloticus)为研究对象, 探究了Ⅰ型与Ⅱ型胶原蛋白基因在二者不同发育阶段及不同部位肌肉组织中的表达模式。系统进化分析结果显示, 胶原蛋白基因col1a1、col1a2和col2a1在有刺鱼和无刺鱼中均各自聚为一支, 团头鲂和罗非鱼3个基因的氨基酸同源性都在90%以下。不同部位肌肉组织(背部上方、尾部上方和尾部下方)的基因表达结果显示, 团头鲂col1a1a和col1a1b基因与罗非鱼该同源基因col1a1在不同肌肉组织中的表达模式存在明显差异。在团头鲂中, Ⅰ型与Ⅱ型胶原蛋白基因在背上肌肉中的表达量高于尾部; 而在罗非鱼中, 其表达模式则相反。团头鲂和罗非鱼不同发育时期的基因定量结果显示: 团头鲂col1a1a和col2a1b基因的表达在肌间骨出现以前(15 dph)和基本出现之后(50 dph)显著(P<0.01)增加, 且Ⅰ型胶原蛋白基因和col2a1b的相对表达量在不同时期差异明显, 其中col1a1a基因在50 dph的表达丰度极高; 与团头鲂相比, 罗非鱼中相应基因的表达量变化较小, 整体波动不大。研究揭示了Ⅰ型和Ⅱ型胶原蛋白在有刺鱼与无刺鱼肌肉中的表达模式, 结果表明col1a1在团头鲂和尼罗罗非鱼两种鱼类中表达模式显著不同(P<0.01), 推测其与肌间骨发育潜在相关。     

唇?硬骨蛋白基因克隆及其在肌间刺发生过程中的表达

为探究硬化蛋白(Sclerostin, SOST)与肌间刺发生之间的关系, 研究通过转录组测序和RT-PCR获得了唇?(Hemibarbus labeo)硬化蛋白基因cDNA序列。序列分析表明, 硬化蛋白由信号肽和成熟肽两部分组成, 成熟肽包含一个胱氨酸结样结构域。系统进化树分析表明, 唇?硬化蛋白与金鱼(Carassius auratus)硬化蛋白最为接近。通过荧光定量PCR检测发现唇?硬化蛋白基因mRNA在所有被检测的组织中均有表达, 在鳃中的表达量最高。RNA原位杂交结果表明, 硬化蛋白mRNA分布于肌膈, 且随着肌间刺的发生信号逐步减弱。RT-qPCR显示, 在肌间刺4个发育阶段中, 硬化蛋白基因mRNA的变化显著, 在阶段Ⅰ表达量最高, 随后逐步降低。综上, 唇?硬化蛋白与肌间刺骨化存在一定的相关性。研究结果将为进一步调查鱼类硬化蛋白功能及硬化蛋白在肌间刺形成过程中的作用提供理论依据。     

唇?bmp2a基因克隆及其与肌间刺骨化相关性分析

骨形成蛋白(Bone morphogenetic protein, BMP)属于转化生长因子β(Transforming growth factor-beta, TGF-β)超家族。在骨形成蛋白家族中, BMP2属于分泌性多功能蛋白, 具有较强的诱导骨细胞形成的能力。为了探究BMP2与肌间刺骨化的相关性,研究通过转录组测序和RT-PCR获得了唇?(Hemibarbus labeo)bmp2a基因cDNA序列。氨基酸序列比对结果显示, 唇?BMP2A和其他鱼类BMP2保守性较高, 都具有一个高度保守的TGFβ结构域, 在该结构域中存在一个N-糖基化位点。此外, 包括唇?BMP2A在内, 鱼类BMP2都具有7个保守的半胱氨酸残基。系统进化树分析表明, 唇?BMP2A与团头鲂(Megalobrama amblycephala)BMP2A最为接近。通过荧光定量PCR检测发现唇?bmp2a基因在所有被检测的组织中均有表达, bmp2a基因在鳃中的表达量最高, 肝脏和肌肉中相对较高, 在心脏中表达量最低。免疫组化结果显示BMP2A分布于肌肉肌隔中。此外, bmp2a基因的表达与肌间刺骨化时机相吻合。综上, 唇?BMP2A与肌间刺骨化存在较高的相关性。研究结果将为进一步调查鱼类BMP2功能以及鱼类肌间刺形成分子机制提供理论依据。     

团头鲂肌间骨发育的形态学观察

研究利用整体骨骼染色、形态学解剖和X光透射的方法, 对团头鲂(Megalobrama amblycephala)仔稚鱼肌间骨的出现时期、形态以及成鱼肌间骨数目、形态、分布和长度变化进行了观察与分析。结果表明: 团头鲂的肌间骨在孵出后20d (体长为1.33 cm)骨化出现, 首先出现在尾部, 然后向头部方向依次出现, 到第40天(体长为2.36 cm)基本全部出现; 肌间骨出现与分化的时间受生长发育的影响大于日龄的影响。团头鲂肌间骨数目在108129, 平均为119 根, 其中躯干部轴上肌中的肌间骨数目最多(4045根), 尾部轴上肌与轴下肌中的肌间骨数目相近(3239根)。肌间骨形态包括1形、卜形、y形、一端多叉形、两端多叉形和(形6种类型, 各种形态的肌间骨均是从1形发展而来; 肌间骨越靠鱼体前端形态越复杂。团头鲂躯干轴上肌中的肌间骨显著长于尾部肌肉中的肌间骨(P 0.05), 躯干轴下肌中的肌间骨最短, 并且肌间骨长度与个体体重与体长呈正相关。研究结果为今后揭示团头鲂肌间骨发生与发育的分子机制, 抑制团头鲂肌间骨骨化, 培育无肌间骨的团头鲂提供了形态学基础。       

甘油诱导的骨骼肌损伤修复过程肌间脂形态学及肌分泌因子表达变化

研究发现在甘油诱导的小鼠肌肉损伤修复过程中可能存在肌间脂的沉积,而肌肉分泌因子（myokines）作为特殊的蛋白参与了肌肉与脂肪的多种生理过程.为研究肌肉内注射甘油后对肌间脂生成的影响,以及注射后肌肉分泌因子在肌肉损伤后修复及肌间脂沉积过程中的表达趋势,本文选用三月龄C57BL/6品系小鼠,右腿胫骨前肌注射50% HBSS(V/V）甘油,左腿胫骨前肌注射等量的HBSS缓冲液作为对照.取注射后不同时期小鼠的胫骨前肌,冰冻切片技术检测肌肉再生及肌间脂沉积状况,实时定量PCR检测各分泌因子（IL-6、IL-15、MSTN、FNDC5、FGF21、myonectin和Insl6）的mRNA表达变化,酶联免疫分析（ELISA）检测分泌因子的蛋白表达变化.结果表明,在甘油诱导的肌肉损伤再生修复过程中存在肌间脂的生成,同时IL-6、Insl6、FGF21和IL-15的mRNA相对表达量在肌肉损伤修复过程中的前、中期变化明显,而MSTN和myonectin的mRNA相对表达量则在中、后期变化明显. IL-6、Insl6的蛋白表达量在前、中期明显升高.综上所述,甘油注射可引起肌肉损伤修复,并在这一过程中伴随着肌间脂的沉积,而肌肉分泌因子作为肌肉与脂肪之间的信息交换因子可能参与了肌肉损伤后的再生修复以及肌间脂的形成.     

鲌鲂F1、F2及其亲本肌间骨的比较分析

为研究新型鲌鲂杂交鱼的肌间骨,采用常规测量和解剖法对鲌鲂F1 (female Culter alburnus Basilewskymale Megalobrama amblycephala Yih)、鲌鲂F2 (self-crossing of F1 hybrid of female C. alburnusmale M. amblycephala)及其母本翘嘴鲌(C. alburnus)、父本团头鲂(M. amblycephala)肌间骨的数目、形态和分布进行统计分析,结果表明,翘嘴鲌肌间骨数目为134-139,平均为136.8根; 团头鲂的肌间骨数目为121-129,平均为124.2根; 鲌鲂F1肌间骨数目为129-134,平均为131.6根; 鲌鲂F2肌间骨数目为127-134,平均为130.1根; 鲌鲂F1、F2与翘嘴鲌、团头鲂之间肌间骨数目差异显著(P0.05); 鲌鲂F1每一肌节所含肌间骨数目最多,为0.8024; 鲌鲂F2最少,为0.7744; 翘嘴鲌和团头鲂介于鲌鲂F1和F2之间,分别为0.7953和0.7763。4种鱼均含有I形、卜形、Y形、一端多叉形、两端两分叉形、两端多叉形和树枝形7种类型肌间骨,髓弓小骨比脉弓小骨多且复杂; 鱼体左右两侧肌间骨的数目不完全相等,形态也不完全对称,但较为接近,且肌间骨越靠前端,形态越复杂; 研究获得的鲌鲂F1和F2在肌间骨总数、复杂型肌间骨数目和躯体轴下肌肌间骨数目均较母本有所减少,鲌鲂F2较F1还呈下降的趋势,且每一肌节所含肌间骨最少,表现出一种有利于提高食用品质和精深加工的优势。研究结果为鲌鲂属间远缘杂交培育少肌间骨新品种提供了基础资料。     

Prey capture in lizards: differences in jaw–neck–forelimb coordination

Fish body muscles are arranged along the vertebral column in three‐dimensional W‐shaped blocks, called myomeres. Each myomere is separated from its neighbours by a collagenous sheet, the myoseptum, and embedded in these myosepta and in positions that are conserved throughout gnathostome evolution are distinct tendons. Within teleosts these tendons often ossify. Ossification is usually intramembranous but cartilaginous structures within the tendons have also been reported. Ossified myoseptal tendons are homologous to intermuscular bones and appear only in teleosts. The phylogenetic signal of myoseptal tendon ossfication has not been tested previously, although the presence and morphology of intermuscular bones have been used to infer phylogenetic relationships. We sample over a broad phylogenetic range of teleost fishes to test for (1) the effects of phylogenetic history on the presence of intermuscular bones and (2) morphological correlations with the presence of intermuscular bones. Body shape and fin position as well as vertebral number and aspect ratio are characters that are likely to affect the distribution of stresses along myoseptal tendons, and are therefore good functional predictors of myoseptal tendon ossification. We use the summary information by Patterson & Johnson for a list of species with intermuscular bones and reanalyse the homology of intermuscular bones to myoseptal tendons. We find that there is a phylogenetic signal in the distribution of four out of six ossified tendons, but that after correcting for phylogenetic relationships there are still morphological predictors for the presence of all ossified tendons. © 2012 The Linnean Society of London, Biological Journal of the Linnean Society, 2012, 106 , 607–622.     

异育淇鲫及其亲本血清生化组成的比较研究

应用Beckman 42型临床生化分析仪测定异育淇鲫(淇鲫♀×兴国红鲤♂) 及其亲本血清的生化指标,包括总蛋白,白蛋白,尿素氮,血糖、乳酸脱氢酶,碱性磷酸酶,谷草转氨酶和羟丁酸脱氢酶,经t检验发现异育淇鲫与其母本的血清生化指标均无明显差异,而与其父本均有十分显著的差异。本文从代谢方面论证了淇鲫与兴国红鲤属间“杂交”,异源精子只起刺激作用,而并不产生生物学效应。结果与传统的雌核发育相吻合。     

Comparative analysis of intermuscular bones in fish of different ploidies

We documented the number, morphology, and distribution of intermuscular bones in five fishes of different ploidy: Carassius auratus (Abbr.WCC, 2n=100), Carassius auratus variety PengZe (Abbr.PZCC, 3n=150), improved triploid crucian carp (Abbr.ITCC, 3n=150), improved red crucian carp (Carassius auratus red var., Abbr.IRCC, ♀, 2n=100), and improved allotetraploids (Abbr.G×AT, ♂, 4n=200). The number of intermuscular bones in WCC, PZCC, and G×AT ranged from 78 to 83 ( $\bar x$ =81), 80 to 86 ( $\bar x$ =84), and 77 to 84 ( $\bar x$ =82), respectively. The numbers in ITCC and IRCC were significantly lower, ranging from 77 to 82 ( $\bar x$ =79) and 58 to 77 ( $\bar x$ =71), respectively. The average number of intermuscular bones in each sarcomere, ranked in order from highest to lowest, was 0.721 (WCC), 0.673 (PZCC), 0.653 (G×AT), 0.633 (ITCC), and 0.608 (IRCC). There was no difference between ITCC and G×AT or between G×AT and PZCC. However, the average number of intermuscular bones in the sarcomeres of ITCC, WCC, and PZCC differed significantly, as did that of IRCC and the four other kinds of fish. The intermuscular bone of these five fishes was divided into seven shape categories, non-forked (卜), one-end-unequal-bi-fork (卜), one-end-equal-bi-fork (Y), one-end-multi-fork, two-end-bi-fork, two-end-multi-fork, and tree-branch types. Generally, the morphological complexity was higher in the anterior intermuscular bones than in the posterior body. The number of intermuscular bones was similar but not equal between the left and right sides of the body. ITCC had significantly fewer intermuscular bones than either WCC or PZCC, making it of greater commercial value. Additionally, IRCC and ITCC had fewer intermuscular bones than WCC. Our observations are significant in both fish bone developmental biology and genetic breeding.     

禾花鲤与建鲤肌间骨miRNAs测序与分析比较

为了研究禾花鲤(Rice flower carp)肌间骨柔软的分子调控机制,采集禾花鲤和建鲤(Jian carp)的肌间骨进行micro RNAs (miRNAs)的高通量测序,并进行了生物信息学分析。从禾花鲤肌间骨的小RNA文库测序获得18 32 nt的高质量miRNA序列25474895条、鉴定出成熟miRNAs 595种,而从建鲤文库获得miRNA序列24625715条、成熟miRNAs 570种。表达差异分析结果显示,与建鲤相比,禾花鲤肌间骨中84个miRNAs呈上调表达、267个呈下调表达。禾花鲤下调表达的miRNAs中有7条为已报道的促进人类成骨的miRNAs,上调表达的miRNAs中有6条为已报道的抑制人类成骨的miRNAs。因此推测禾花鲤可能是通过下调表达促进成骨的miRNAs及上调表达抑制成骨的miRNAs来抑制成骨过程,从而维持其肌间骨细小及柔软的特性。研究为国内首次开展禾花鲤肌间骨发育的分子调控机制研究,为水产动物肌间骨发育的研究奠定了基础。     

Comparative analysis of intermuscular bones in three strains of common carp

The objective was to document the variations in the intermuscular bone counts among three carp strains: mirror carp, hybrid carp [Boshi carp (Cyprinus pellegrini) × Heilongjiang carp (Cyprinus carpio)], and a cold‐resistant strain of Hebao carp. The intermuscular bone counts, lengths, and weights were obtained from 146 fish; the bone count ranged from 55 to 110 (mean 92.85) among the three strains. Mirror carp had the lowest number of intermuscular bones and a higher coefficient of variation in counts relative to the other two species. There was no relationship between the intermuscular bone counts to standard length or body length. Similarly, there was no difference in the intermuscular bone count or shape between the left and right sides of the body. However, the count was significantly higher (P < 0.01) in the anterior region (snout to cloaca: 54% of the total count) than in the posterior region (cloaca to base of caudal fin: 46% of the total count). There was a significant difference between strains (P < 0.0001), but not between age classes; a significant difference was also observed in the posterior region counts among strains, but not in the anterior region (P < 0.0001). Similarly, the difference in counts in the posterior region was primarily related to differences in the counts of three of the six types: non‐forked (I), tree‐branch and two‐end‐bi‐fork type. Our results suggest that selective breeding protocols should target a reduction in the number of intermuscular bones in the posterior region, particularly in mirror carp.     

Seasonal development and pathological changes associated with the parasitic nematode Philometroides sanguineus in wild crucian carp Carassius carassius (L.) in England

Pathological changes associated with the parasitic nematode Philometroides sanguineus (Rudolphi, 1819) are described for the first time from observations of infections in wild crucian carp Carassius carassius (L.) in England. The damage caused by P. sanguineus was influenced strongly by host size, parasite development and the seasonal migrations of female nematodes within host tissues. Male and unfertilized female nematodes located on the swim-bladder wall caused only minor, localized changes. In contrast, the migration of gravid female nematodes into the fins during autumn provoked an acute inflammatory response comprising neutrophils, eosinophils and lymphocytes. This was accompanied by fin distortion, swelling of the dorsal and caudal tissues, degenerative changes and localized oedema. The encapsulation of female nematodes in connective tissue throughout winter limited additional tissue damage. The emergence of gravid nematodes from the fins in spring to facilitate larval dispersal caused necrosis, hyperplasia and loss of skin integrity. This activity was again accompanied by acute inflammatory reactions. Pathological changes were more severe in crucian carp measuring less than 60 mm in length, with no signs of debilitation in larger fish. These observations suggest that any impact of P. sanguineus is strictly seasonal and may be limited to fry. Lesions caused by this parasite, only recently recorded in Britain, may represent an additional pressure upon wild crucian carp populations already threatened by hybridization, competition and habitat loss.     

Zebrafish periostin is required for the adhesion of muscle fiber bundles to the myoseptum and for the differentiation of muscle fibers

The myoseptum of fishes, composed of dense collagen, is a connective tissue layer that forms in the embryo, dividing somites from the trunk, and its structure and function are similar to those of the mammalian tendon. Both the myoseptum and tendon serve as the transmitter of muscular contractility to bones and adjoining muscles, and their structure is indispensable for movement of vertebrate animals. We cloned the zebrafish periostin gene and examined its expression and function in the myoseptum. The expression in embryos started in the rostral part of each segmented somite in the early segmentation stage; and consequently, metameric stripes were observed. At the end of segmentation, the expression region shifted to the transverse myoseptum and the myotome-epidermis boundary, and each myotome was surrounded by periostin. Using a polyclonal antibody, we found that the periostin protein was localized to the transverse myoseptum. Consistently, periostin morpholino antisense oligonucleotide led to defects in myoseptum formation, a delay in the differentiation of myofibers, and disorder of connection between myofibrils and myoseptum. We demonstrated here that periostin is the first molecule involved in myoseptum formation and propose that periostin secretion on the surface of the myoseptum is required for the adhesion of muscle fiber bundles to the myoseptum and the differentiation of muscle fibers.