真骨鱼类皮肤角质化衍生物：单细胞角质突起

皮肤是动物体表的器官系统,与周围环境有着直接的联系.动物皮肤可产生角质化衍生物,用以抵御外界环境条件所造成的伤害.在四足类动物适应陆地生存环境的过程中,皮肤角质化衍生物所起的作用显得至关重要.爬行类的角质鳞可以保护身体和阻止水分散失;鸟类的羽毛用于飞翔和保温;哺乳动物的毛发有保护身体和御寒等作用[1].     

记真盔甲鱼类两新属——兼论真盔甲鱼类系统发育关系

本文记述了早泥盆世真盔甲鱼类两新属:憨鱼属(Nochelaspis)和翼角鱼属(Pterogonaspis).真盔甲鱼类是盔甲鱼类的一个单系类群,现有七属十二种,文中运用分支系统学原理对其系统发育关系进行了初步探讨.     

人表皮生长因子基因转染原代角质形成细胞

目的:将带信号肽的人表皮生长因子基因转染原代成人角质形成细胞, 证实细胞活性及hEGF的有效表达, 为后期的皮肤修复研究打下基础.方法:先酶切验证pcDNA3.1-hEGF, 后消化成人皮肤组织, 以Defined Keratinocyte-SFM(DKSFM)传代培养角质形成细胞并鉴定, 脂质体转染质粒pcDNA3.1-hEGF入细胞, 转基因细胞培养48 h后作RT-PCR和Western-blot分析, 上清分别进行放射免疫测定和MTT测定.结果:质粒pcDNA3.1-hEGF上的hEGF序列经测序证实,双酶切后获得约230 bp和5.4 kb条带;成人角质形成细胞体外培养可快速稳定增殖, 转hEGF基因细胞经RT-PCR扩增出一条约230 bp的特异性条带, Western-blot检测到hEGF表达明显升高;放射免疫法和MTT实验证实转基因细胞有稳定的hEGF蛋白分泌.结论:质粒pcDNA3.1-hEGF在脂质体介导下成功转染成人皮肤角质形成细胞, 转基因细胞能分泌有生物活性的EGF;体外培养的成人皮肤角质形成细胞可见少量EGF分泌.     

TRPA1传导皮肤角质细胞温热信息的机制研究

目的 探究角质细胞中的温热信息是如何被传导至下游背根神经节（DRG）中的TRPA1离子通道。方法 采用单细胞钙离子成像技术,利用TRPA1过表达系统以及野生型和TRPA1基因敲除小鼠,联合TRPA1激动剂（BITC）和抑制剂（HC-030031）检测H₂O₂在DRG神经元温热感知中的作用及其与TRPA1的相关性。在不同温热条件下培养皮肤角质细胞,并提取角质细胞RNA;采用RNA-seq分析比较不同温度培养的角质细胞基因表达异同,筛选皮肤角质细胞传导温热信息的候选因子。结果 过表达TRPA1以及DRG神经元中的TRPA1均可在H₂O₂存在情况下被温热激活,而温热范围内不同温度可影响皮肤角质细胞中与H₂O₂产生有关的趋化因子配体2（CCL2）和核心蛋白聚糖（DCN）基因的表达。结论 H₂O₂相关因子可能参与TRPA1介导的温热传导过程。     

云南发现新的全骨鱼类化石

正据英国皇家学会《生物学报》报道,中国科学院古脊椎动物与古人类研究所、浙江自然博物馆和芝加哥大学组成的研究小组在云南省罗平县三叠纪地层中发现辐鳍鱼亚纲中一种新的全骨鱼类化石,命名为罗平强壮鱼(Robustichthys luopingensis)。该发现为研究全骨鱼类的早期演化和预言鱼目(Ionoscopiformes)的起源提供了关键性的证据。全骨鱼类是真骨鱼类(现生脊椎动物中最大的类群)的姐妹群,在研究辐鳍鱼亚纲新鳍鱼     

核糖体蛋白RPL15 cDNA序列在真骨鱼类系统进化研究中的价值 &

应用RT-PCR, 从真骨总目（Teleostei）5目15种鱼类中首次克隆了核糖体大亚基蛋白L15 (RPL15, ribosomal protein L15) 的完整cDNA序列。以海鲢形亚组（Elopomorpha）的鳗鲡作为外类群, 对这些真骨鱼类的核糖体蛋白L15 cDNA序列进行了系统发育分析, 结果表明: (1)RPL15基因在真骨鱼类等许多真核生物进化中高度保守; (2)系统树中各物种之间的关系与形态分类一致。RPL15编码区适合于真骨鱼类目以上分类阶元的分子系统学研究。      

胰酶对皮肤角质细胞分离和传代的影响

考察不同的胰酶浓度及其作用时间对角质细胞分离和传代的影响。实验发现在胰酶浓度 0.25%时作用5min可获得的总活细胞量和有克隆形成能力的细胞量均优于其它培养条件 ;而在胰酶浓度0.05%时作用 5min可获得最大的原代角质细胞贴壁率。随着胰酶浓度的提高 ,传代角质细胞的贴壁率和贴壁速率常数以及克隆形成率均随之增加 ,因此在传代培养时使用 0.25%胰酶浓度进行消化较为适宜.     

中药固真方对成骨样细胞UMR106增殖分化的影响

 中药固真方具有补肾益精、延缓衰老的作用 .为了进一步探讨其作用机理 ,研究观察了固真方对成骨样细胞 UMR1 0 6DNA合成、原癌基因 c- fos,c- ki- ras m RNA表达以及骨钙素 (osteocal-cin)基因 m RNA表达的影响 .结果表明 :固真方可明显促进 DNA合成 ,且在 1 0 -5稀释度时达高峰 ;增殖相关的原癌基因 (c- fos,c- ki- ras) m RNA表达增强 ;成熟成骨细胞特征性标志蛋白——骨钙素基因 m RNA表达增强 .结果提示 :中药固真方可能通过影响一些与增殖相关的原癌基因及骨钙素基因表达而促成骨样细胞 UMR1 0 6的增殖和分化成熟     

紫外线A照射差异性损伤皮肤角质形成细胞和成纤维细胞

紫外线A(ultraviolet A,UVA)照射皮肤可产生红斑、色素沉着以及光敏性损伤,导致光老化甚至诱导皮肤癌。研究发现,UVA(320～400 nm)照射对皮肤角质形成细胞和成纤维细胞造成损伤的反应和损伤程度都有所不同。本文结合近年来的相关研究,对UVA照射导致的上述两种细胞的形态、细胞膜损伤、血红素氧合酶(heme oxygenase,HO)活性、HO-1表达、细胞内有害自由铁形成,以及其它抗氧化物水平的不同反应等进行阐述,总结分析了同等剂量UVA照射后对角质形成细胞和成纤维细胞损伤的差异性,这些分析可能为皮肤抗UVA照射损伤以及寻找新型防晒剂的开发提供新思路和实验依据。     

pcDNA3.1(+)-CTGF真核表达质粒构建及其在人成骨样细胞SaOS-2细胞中的表达*

目的:构建结缔组织生长因子(CTGF)的pcDNA3.1(+)真核表达质粒(pcDNA3.1(+)-CTGF),并检测其在人成骨样细胞SaOS-2中的表达,为进一步研究CTGF基因在骨发育和骨修复中的机制提供技术支撑。方法:采用PCR方法体外克隆CTGF基因全序列,将其用同源重组技术连接到线性pcDNA3.1(+)载体上,构建pcDNA3.1(+)-CTGF真核表达质粒,并对该质粒进行测序鉴定;鉴定无误后转染至SaOS-2细胞中,观察其48 h的表达情况。结果:基因测序证实pcDNA3.1(+)-CTGF真核表达重组质粒构建成功,与对照组相比,转染SaOS-2细胞48 h后的CTGF表达水平显著上调,达到对照组的4.8×10⁵倍(P＜0.01)。结论:成功构建了pcDNA3.1(+)-CTGF真核表达质粒,并能在人成骨样细胞SaOS-2中稳定表达,为深入研究CTGF基因对骨生成的调控机制奠定了基础。     

Horny cell formation in the epidermis of Rana pipiens

Well preserved transitional cells were found between differentiated cells and horny cells of the frog epidermis, thus facilitating the study of the sequential events involved in horny cell formation. Autolysosomes appear to play an important role in the formation of horny cells. These structures preferentially digest those cytoplasmic components which are not necessary constituents of the terminal horny cell. The release of the contents of the small mucous granules into the intercellular spaces is one of the initial events in horny cell formation. Filaments and large mucous granules seem to be resistant to the lytic digestion and contribute to the bulk of the horny cell. Loss of fluids through the plasma membrane and consolidation of the remaining constituents, results in a flattened horny cell. The appearance of a thickened membrane around the horny cell signifies the completion of the transformation process.     

Evolution of the Multicellular Animals

SYNOPSIS. Molecular sequence analysis is providing new insightsinto the study of metazoan relationships. The use of ribosomalRNA sequences is revising many of the metazoan phylogenies thathave been established traditionally with anatomical and embryologicaldata. Four new findings that seem to be well supported by moleculardata, both from the authors' laboratories and from others, aredescribed and discussed. First, the arthropods are members ofa deep primary clade within the protostomes and are not thesister taxa of either the annelids or the mollusks. Second,the lophophorate animals are clearly protostomes and are containedwithin a lophotrochozoan superclade including the mollusks,annelids, and many other phyla. Third, the arthropods togetherwith all other molting animals comprise a second monophyleticsuperclade within the protostomes, the ecdysozoa. Fourth, theplatyhelminthes are contained within the lophotrochozoan superclade.     

Keratinization in the epidermis of amphibians and the lungfish: comparison with amniote keratinization

Keratinization in the epidermis of amphibians and the lungfish has been studied by electron microscopy, autoradiography and immunocytochemistry to determine whether histidine-rich proteins, filaggrin and loricrin are present. In the lungfish and amphibian tadpoles, anti-keratin antibodies (AE1 and AE3) stain the whole epidermis but not the AE2 antibody, a marker for keratinization. In adult epidermis, the AE2 antibody mainly stains keratinized layers, AE1 mainly stained basal cells, less suprabasal cells and no pre-keratinized and keratinized layers, and AE3 stains all epidermal layers. This staining pattern resembles that of amniote epidermis. Little tritiated histidine is taken up in toad epidermis at 4-6 h post-injection but 24 h after injection the radioactivity is most concentrated in the replacement layer beneath the corneus. This indicates that protein synthesis takes place in the epidermis but, due to the metabolic conversion that takes place in 24 h, it is unlikely that histidine-rich proteins are formed. Neither filaggrin-like nor loricrine-like immunoreactivities are present in amphibian and lungfish epidermis. This indicates absence of histidine-rich matrix proteins and corneous cell envelope proteins and only mucus is present among keratin filaments. Filaggrine-like and loricrin-like proteins are characteristic of amniotes epidermis and might have originated in basic amniotes (cotylosaurs).     

Abnormal Epidermal Keratinization in the repeated epilation mutant mouse

Repeated epilation (Er) is a radiation-induced, autosomal, incomplete dominant mutation in mice which is expressed in heterozygotes but is lethal in the homozygous condition. Many effects of the mutation occur in skin: the epidermis in Er/Er mice is adhesive (oral and nasal orifices fuse, limbs adhere to the body wall), hyperplastic, and fails to undergo terminal differentiation. Skin from fetal +/+, Er/+ and Er/Er mice at ages pre- and postkeratinization examined by light, scanning, and transmission electron microscopy showed marked abnormalities in tissue architecture, differentiation, and cell structure; light and dark basal epidermal cells were separated by wide intercellular spaces, joined by few desmosomes, and contained phagolysomes. The numbers of spinous, granular, and superficial layers were highly variable within any given region and among various regions of the body. In some areas, 2-8 layers of granular cells, containing large or diminutive keratohyalin granules, extended to the epidermal surface; in others, the granular layers were covered by several layers of partially keratinized or nonkeratinized cells. In rare instances, a single or small group of cornified cells was present among the granular layers but was not associated with the epidermal surface. Both the granular and nonkeratinized/partially keratinized upper epidermal layers Er/Er skin gave positive immunofluorescence with antiserum to the histidine-rich, basic protein, filaggrin. Proteins in epidermal extracts from +/+, Er/+ and Er/Er mice were separated and identified by radio- and immunolabeling techniques. The Er/Er extract was missing a 26.5- kdalton protein and had an altered ratio of bands in the keratin region. The 26.5-kdalton band was histidine-rich and cross-reacted with the antiserum to rat filaggrin. Several high molecular weight bands present in both Er/Er and +/+ extracts also reacted with the antiserum. These are presumed to be the precursors of filaggrin and to account for the immunofluorescence om Er/Er epidermis even though the product protein is absent. The morphologic and biochemical data indicated that the genetic defect has a general and profound influence on epidermal differentiation, including alteration of two proteins (filaggrin and keratin) important in normal terminal differentiation, tissue architecture, and cytology. Identification of epidermal abnormalities at early stages of development (prekeratinization) and defective structure of other tissues and gross anatomy suggest that the mutation is responsible for a defect in same regulatory step important in many processes of differentiation and development.     

Retention of 14C-Labeled Tuberculin Purified Protein Derivative in the Skin of Sensitized and Nonsensitized Animals

Tuberculin purified protein derivative labeled with (14)C ([(14)C]PPD) with a biological potency equivalent to the International Standard for tuberculin PPD was used to study the retention of tuberculin PPD in the skin of sensitized and nonsensitized animals. We found that [(14)C]PPD was almost entirely cleared from the skin test site during the first 18 to 24 h after injection and that when approximately 5% of the initial concentration of [(14)C]PPD was present in the skin test site, the size of the tuberculin skin reaction in sensitized guinea pigs was at its maximum. Furthermore, the addition of 5 or 50 mug of Tween 80 per ml to a solution of PPD did not change either the rate of clearance of PPD from the skin test sites of sensitized guinea pigs or the size of the tuberculin skin reactions. There was no difference in the rate of clearance of [(14)C]PPD from the skin test sites between sensitized and nonsensitized guinea pigs and between guinea pigs of different age. However, there was a significant difference in the rate of clearance of [(14)C]PPD between the guinea pig and the mouse. Finally, the percentage of [(14)C]PPD retained in the site of injection at 24 h was in the neighborhood of 5% of the initial concentration of the solution of PPD injected. The significance of these phenomena is discussed.