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鹿茸软骨组织脱细胞基质材料的制备及生物相容性研究
Preparation of acellular matrix from antler cartilage and its biological compatibility
 [PDF]

付晶,张伟,张爱武,马丽娟,褚文辉,李春义
- , 2017, DOI: 10.7507/1002-1892.201612072
Abstract: 目的探讨鹿茸软骨制备脱细胞基质材料的可行性以及生物相容性,为软骨修复重建探索新材料。 方法取梅花鹿鹿茸生长中心间充质层,进行由 DNA 酶、RNA 酶、抑肽酶等介导的脱细胞处理;行组织学和 DNA 含量检测,评价脱细胞效果。取第 2 代鹿生茸区骨膜(antlerogenic periosteum,AP)细胞,行荧光干细胞标记明确其干细胞特性后,用 PKH26 荧光标记并与制备的间充质层脱细胞基质进行复合培养;7 d 后取材行HE染色观察以及荧光显微镜下观察 PKH26 标记的 AP 细胞在基质表面生长情况。以上观测均以未复合 AP 细胞的脱细胞基质作为对照。将复合培养 7 d 的样本移植至裸鼠一侧腹股沟(实验组),取空白培养样本移植于另一侧(对照组)。于移植后 7、21 d 取材行 HE 染色,同时对组织进行冰冻切片并在荧光显微镜下观察 PKH26 标记成功的 AP 细胞在脱细胞基质表面及内部的生长情况,评价含 AP 细胞的脱细胞基质在裸鼠体内的组织相容性。 结果HE 和 DAPI 染色显示脱细胞处理后材料中无细胞残留,DNA 含量为(19.367±5.254)ng/mg,较脱细胞处理前的(3 805.500±519.119)ng/mg 显著降低( t=12.630, P=0.000),提示成功制备间充质层脱细胞基质。AP 细胞与间充质层脱细胞基质复合培养 7 d 后,AP 细胞主要黏附于材料表面,部分进入脱细胞基质内部。植入裸鼠体内后,随观察时间延长,接种 AP 细胞可以在脱细胞基质材料中增殖并逐渐进入材料内部,并诱导血管生成。 结论实验成功制备鹿茸软骨脱细胞基质,该基质材料在离体和活体情况下适于种子细胞(AP 细胞)的黏附和增殖,并具有刺激血管生成的功能,为其用于软骨组织修复提供理论依据。
ObjectiveTo study the feasibility of acellular matrix materials prepared from deer antler cartilage and its biological compatibility so as to search for a new member of the extracellular matrix family for cartilage regeneration. MethodsThe deer antler mesenchymal (M) layer tissue was harvested and treated through decellular process to prepare M layer acellular matrix; histologic observation and detection of M layer acellular matrix DNA content were carried out. The antler stem cells [antlerogenic periosteum (AP) cells] at 2nd passage were labelled by fluorescent stains and by PKH26. Subsequently, the M layer acellular matrix and the AP cells at 2nd passage were co-cultured for 7 days; then the samples were transplanted into nude mice to study the tissue compatibility of M layer acellular matrix in the living animals. ResultsHE and DAPI staining confirmed that the M layer acellular matrix did not contain nucleus; the DNA content of the M layer acellular matrix was (19.367±5.254) ng/mg, which was significantly lower than that of the normal M layer tissue [(3 805.500±519.119) ng/mg]( t=12.630, P=0.000). In vitro co-culture experiments showed that AP cells could adhere to or even embedded in the M layer acellular matrix. Nude mice transplantation experiments showed that the introduced AP cells could proliferate and induce angiogenesis in the M layer acellular matrix. ConclusionThe deer antler cartilage acellular matrix is successfully prepared. The M layer acellular matrix is suitable for adhesion and proliferation of AP cells in vitro and in vivo, and it has the function of stimulating angiogenesis. This model for deer antler cartilage acellular matrix can be applied in cartilage tissue engineering in the future.
器官形态发生探索Ⅱ:独特的哺乳动物模型鹿茸  [PDF]
赵海平,褚文辉,陈广信,李春义
- , 2015, DOI: 10.3785/j.issn.1008-9209.2014.05.261
Abstract: 生物电编码器官形态发生在低等动物中已被证实。由于缺乏哺乳动物的形态发生研究模型,二者之间的关系是否同样适用于哺乳动物至今依然未知。鹿茸是一个复杂的哺乳动物附属器官,其形态发生信息存储于形态发生原胚(鹿前额未来生茸区的骨膜)中。该文1)综述了鹿茸的形态发生原胚及鹿茸发生与再生的影响因素,提出了鹿茸是哺乳动物器官形态发生研究的最佳模型的观点;2)分析了鹿茸形态发生信息的存储位点和复制方式、转移路径,并预测了通过生物电追踪形态发生信息来破解哺乳动物器官生物电密码的研究思路。总之,通过鹿茸这一模型的研究必将为人类器官形态发生的探索奠定理论基础。
Abstract: Morphogenesis is a subject to investigate morphological generation and shape maintenance of organisms. Regeneration can recapitulate the process of morphogenesis. Therefore, lower animals such as planaria were usually used to study morphogenesis, as they can regenerate some parts of their bodies. Correlation between bioelectricity and organ morphogenesis has been observed for lower animals such as amphibians. Whether this finding applies to the situation of mammals including humans is unknown due to lack of mammalian models. Deer antler is a unique mammalian organ that can regenerate annually. Deer antler development initiates from AP (antlerogenic periosteum, the periosteum overlying the frontal crest of prepubertal deer) tissue. Subsequent studies showed that the AP controls the morphogenesis of deer antler. 1) Ectopic antlers formed when APs were removed from normal position to anywhere of deer body. 2) Backward morphology of antlers developed when the APs were rotated 180° from original position. 3) Duplication of APs resulted in the growth of normal antlers. 4) Even minced APs can develop organized antlers. Morphogenetic potential of different regions of AP was also assessed. Antler morphogenetic information was mainly held in the anterior and medial halves of AP. The former is responsible for antler brow tine; and the latter is responsible for other branches of antler. All these features, such as full regeneration and a simple tissue-derivative, only exist in antlers comparing to other mammalian organs. Antlers morphogenesis can be studied by simply manipulating AP. Therefore, deer antler is believed to be an ideal model for organ morphogenesis study in mammals. Bioelectricity results on deer antler morphogenesis is a good beginning to understand the mystery of organ morphogenesis in mammals. In this review, the storage, duplication and transferring pathway of morphogenetic information for deer antlers were analyzed, and preliminary idea for cracking the secret codes of morphogenesis of mammalian organs through tracing bioelectricity was outlined. The findings from deer antler model are expected to be applied to artificial control of organ morphogenesis for mammal to serve for human health.
中国茸鹿品种(品系)的随机扩增多态dna(rapd)研究*  [PDF]
李和平,师守,李生
应用与环境生物学报 , 2000,
Abstract: 摘要采用随机扩增多态dna技术研究了5个茸鹿品种(品系)75个个体的遗传变异关系.在所使用的41个引物中,有40个引物扩增出多态谱带,共检测到395条扩增片段,其中259条(65.6%)出现变异.用shannon指数计算了5个品种(品系)的遗传变异及其遗传变异在群体内和群体间的分布.利用nei氏片段共享度计算了75个个体间的遗传距离,用upgma聚类法构建了75个个体的系统发育树状图,反映5个茸鹿品种(品系)的遗传关系.结果表明,遗传变异主要存在于群体内(53.97%),群体间的遗传变异居次要地位(46.05%);双阳品种群体内的遗传变异最低;5个品种(品系)的遗传多样性指数的大小依次为:西丰(1.1249)、长白山(1.0517)、乌兰坝(0.9908)、清原(0.9748)、双阳(0.7662).图1表3参14
Identification of Genetic Variants Within Androgen Receptor Gene of Sika Deer and its Association with Antler Production
Jiajun Xiong,Feifei Yang,Guohua Hua,Lei Shen,Bin Fan,Pu Zhang,Lijun Huo,Shujun Zhang,Liguo Yang
Journal of Animal and Veterinary Advances , 2012, DOI: 10.3923/javaa.2012.2059.2063
Abstract: Antler production is one of the most important economical traits of Sika deer. However, the genetic mechanism of antler growth and genetic markers associated with antler yield remain unclear. In the present study Androgen Receptor (AR) gene has been considered as a candidate gene to identify the polymorphisms. Besides, its effect on antler production was investigated in Chinese Sika deer. Genomic sequences of exons1-7 of Sika deer have been successfully obtained and showed high homogeneity with bovine. One SNP ss325995755 was identified in exons3. Genotyping of SNP and association analyses with antler yield were analyzed in two Chinese Sika deer populations (n = 215). The SNP ss325995755 was not significantly (p>0.05) associated with average antler yield.
Gene Expression of Axon Growth Promoting Factors in the Deer Antler  [PDF]
Wolfgang Pita-Thomas,Carmen Fernández-Martos,Mónica Yunta,Rodrigo M. Maza,Rosa Navarro-Ruiz,Marcos Javier Lopez-Rodríguez,David Reigada,Manuel Nieto-Sampedro,Manuel Nieto-Diaz
PLOS ONE , 2012, DOI: 10.1371/journal.pone.0015706
Abstract: The annual regeneration cycle of deer (Cervidae, Artiodactyla) antlers represents a unique model of epimorphic regeneration and rapid growth in adult mammals. Regenerating antlers are innervated by trigeminal sensory axons growing through the velvet, the modified form of skin that envelopes the antler, at elongation velocities that reach one centimetre per day in the common deer (Cervus elaphus). Several axon growth promoters like NT-3, NGF or IGF-1 have been described in the antler. To increase the knowledge on the axon growth environment, we have combined different gene-expression techniques to identify and characterize the expression of promoting molecules not previously described in the antler velvet. Cross-species microarray analyses of deer samples on human arrays allowed us to build up a list of 90 extracellular or membrane molecules involved in axon growth that were potentially being expressed in the antler. Fifteen of these genes were analysed using PCR and sequencing techniques to confirm their expression in the velvet and to compare it with the expression in other antler and skin samples. Expression of 8 axon growth promoters was confirmed in the velvet, 5 of them not previously described in the antler. In conclusion, our work shows that antler velvet provides growing axons with a variety of promoters of axon growth, sharing many of them with deer's normal and pedicle skin.
Effect of Different Factors on Proliferation of Antler Cells, Cultured In Vitro  [PDF]
Erika Ku?mová,Luděk Barto?,Radim Kotrba,George A. Bubenik
PLOS ONE , 2012, DOI: 10.1371/journal.pone.0018053
Abstract: Antlers as a potential model for bone growth and development have become an object of rising interest. To elucidate processes explaining how antler growth is regulated, in vitro cultures have been established. However, until now, there has been no standard method to cultivate antler cells and in vitro results are often opposite to those reported in vivo. In addition, many factors which are often not taken into account under in vitro conditions may play an important role in the development of antler cells. In this study we investigated the effects of the antler growth stage, the male individuality, passaged versus primary cultures and the effect of foetal calf serum concentrations on proliferative potential of mixed antler cell cultures in vitro, derived from regenerating antlers of red deer males (Cervus elaphus). The proliferation potential of antler cells was measured by incorporation of 3H thymidine. Our results demonstrate that there is no significant effect of the antler growth stage, whereas male individuality and all other examined factors significantly affected antler cell proliferation. Furthermore, our results suggest that primary cultures may better represent in vivo conditions and processes occurring in regenerating antlers. In conclusion, before all main factors affecting antler cell proliferation in vitro will be satisfactorily investigated, results of in vitro studies focused on hormonal regulation of antler growth should be taken with extreme caution.
Nerve Growth Factor mRNA Expression in the Regenerating Antler Tip of Red Deer (Cervus elaphus)
Chunyi Li,Jo-Ann L. Stanton,Tracy M. Robertson,James M. Suttie,Philip W. Sheard,A. John Harris,Dawn E. Clark
PLOS ONE , 2012, DOI: 10.1371/journal.pone.0000148
Abstract: Deer antlers are the only mammalian organs that can fully regenerate each year. During their growth phase, antlers of red deer extend at a rate of approximately 10 mm/day, a growth rate matched by the antler nerves. It was demonstrated in a previous study that extracts from deer velvet antler can promote neurite outgrowth from neural explants, suggesting a possible role for Nerve Growth Factor (NGF) in antler innervation. Here we showed using the techniques of Northern blot analysis, denervation, immunohistochemistry and in situ hybridization that NGF mRNA was expressed in the regenerating antler, principally in the smooth muscle of the arteries and arterioles of the growing antler tip. Regenerating axons followed the route of the major blood vessels, located at the interface between the dermis and the reserve mesenchyme of the antler. Denervation experiments suggested a causal relationship exists between NGF mRNA expression in arterial smooth muscle and sensory axons in the antler tip. We hypothesize that NGF expressed in the smooth muscle of the arteries and arterioles promotes and maintains antler angiogenesis and this role positions NGF ahead of axons during antler growth. As a result, NGF can serve a second role, attracting sensory axons into the antler, and thus it can provide a guidance cue to define the nerve track. This would explain the phenomenon whereby re-innervation of the regenerating antler follows vascular ingrowth. The annual growth of deer antler presents a unique opportunity to better understand the factors involved in rapid nerve regeneration.
茸鹿杂交试验  [PDF]
动物学杂志 , 1974,
Abstract:
鹿科动物声音通讯的研究进展
Research Development of Vocal Communication in Cervidae
 [PDF]

左智力,杨萍,陈渝倩,刘洋
- , 2018,
Abstract: 中文摘要:所有鹿科Cervidae动物都要依赖视觉、嗅觉和听觉线索进行交流,声音通讯作为一种高效的交流方式,在群体生活中起着非常重要的作用。本文列举了鹿科动物发情鸣叫、警戒鸣叫、交流鸣叫、求救鸣叫等叫声类型,总结了其发声机制,探讨了声音与鹿科动物自身信息的相关性、叫声对两性繁殖行为的影响,以及鹿科动物对种内和种间叫声的识别,并介绍了鹿科动物声音通讯的研究方法。最后,对鹿科动物声音通讯未来的研究方向进行了展望。
英文摘要:Deer rely on visual, olfactory and auditory cues to communicate. Vocal signals constitute a very efficient system of communication, and play a central role in connecting the members of the group. In this review, we listed various types of calls in Cervidae, such as rutting calls, alarm calls, contact calls and distress calls, and then the vocal mechanisms were summarized. We also discussed the relationship between acoustic parameters and signaler information, the effects of calls on reproductive behaviors of both sexes, as well as the intraspecific and interspecific discriminations of vocal signals. Additionally, we also introduced several research methods for the vocal communication in Cervidae. Finally, we proposed some potential study fields for the vocal communication in Cervidae and provided some positive suggestions. 2018,37(4): 474-480 收稿日期:2018-01-22 DOI:10.11984/j.issn.1000-7083.20180028 分类号:Q958.1 基金项目:成都动物园科研课题项目 作者简介:左智力(1982-),男,畜牧师,主要从事动物饲养与管理研究 *通讯作者:刘洋,E-mail:18782199355@163.com 参考文献: 范艳珠, 方光战. 2016. 蛙类声音通讯研究进展[J]. 动物学杂志, 51(6):1118-1128. 付义强, 胡锦矗, 朱欢兵, 等. 2008. 华南梅花鹿声音通讯行为的初步研究[J]. 四川动物, 27(2):266-268. 李春旺, 蒋志刚, 曾岩. 2001. 雄性麋鹿的吼叫行为、序位等级与成功繁殖[J]. 动物学研究, 22(6):449-453. 宁继祖, 郭延蜀, 郑慧珍. 2008. 四川梅花鹿发情期的几种发声行为[J]. 兽类学报, 28(2):187-193. 杨承忠, 张修月, 岳碧松, 等. 2012. 四川梅花鹿产仔换茸期声行为的研究[J]. 四川动物, 31(1):10-16. 杨德华, 马德惠. 1965. 云南西南部的豚鹿[J]. 生物学通报, 5:32-33. 曾治高, 宋延龄, 李善元, 等. 2001. 海南坡鹿雄鹿发情期的吼叫行为及其在繁殖中的意义[J]. 动物学报, 47(5):481-487. 张建军, 张知彬. 2003. 动物的婚配制度[J]. 动物学杂志, 38(2):84-89. Andersson M. 1994. Sexual selection[M]. Princeton:Princeton University Press. Atkeson TD, Marchinton RL, Miller KV. 1988. Vocalizations of white-tailed deer[J]. American Midland Naturalist, 120(1):194-200. Beckford NS, Schaid D, Rood SR, et al. 1985. Androgen stimulation and laryngeal development[J]. The Annals of Otology, Rhinology, and Laryngology, 94(6):634-640. Bradbury JW, Vehrencamp SL. 2011. Principles of animal communication 2nd ed[M]. Sunderland (MA):Sinauer Associates. Briefer EF. 2012. Vocal expression of emotions in mammals:mechanisms of production and evidence[J]. Journal of Zoology, 288(1):1-20. Cap H, Deleporte P, Joachim J, et al. 2008. Male vocal behavior and phylogeny in deer[J]. Cladistics, 24(6):917-931. Charlton BD, Keating JL, Kersey D, et al. 2011. Vocal cues to male androgen levels in giant pandas[J]. Biology Letters, 7(1):71-74. Clutton-Brock TH,
鹿筋胶原对大鼠佐剂性关节炎的治疗作用  [PDF]
孙晓迪,李银清,赵雨,曲毅,张鹤,曲晓波
中国中药杂志 , 2009,
Abstract: 目的:研究鹿筋胶原对大鼠佐剂性关节炎的治疗作用。方法:以完全弗氏佐剂(CFA)诱导的Wistar雄性大鼠佐剂性关节炎(AA)为研究对象,观察经鹿筋胶原治疗前后的足趾肿胀度、体重、血清中的IL-lβ和TNF-a炎性细胞因子、血清中的MDA含量和SOD活力。结果:鹿筋胶原可显著抑制AA大鼠原、继发性足肿胀度,抑制AA大鼠炎性细胞因子IL-lβ和TNF-a分泌,增强SOD活性,降低MDA含量。结论:鹿筋胶原对弗氏佐剂诱导的大鼠佐剂性关节炎有明显的治疗作用。
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