Expression of the adrenergic phenotype by dorsal root ganglion cells of the quail in culture in vitro

From the results of previous studies, we have suggested that "autonomic" cell precursors exist in latent form in sensory ganglia of avian embryos. The potentialities can be expressed when the ganglia are transplanted into a young embryo host. In the present study, we have observed a similar transformation in cultures of dissociated dorsal root ganglia taken from quail embryos of 7-15 days of incubation. From the 4th day of culture onward, numerous adrenergic cells appear. They display tyrosine hydroxylase immunoreactivity, synthesise and store catecholamines and generally differ in size and shape from primary sensory neurons. They and/or their precursors can actively proliferate in culture. The differentiation of these catecholaminergic cells, which can not be detected in quail dorsal root ganglia during normal development in vivo, is dependent on one or more factors present in 9-day chick embryo extract.     

A covalently constrained congener of the Saccharomyces cerevisiae tridecapeptide mating pheromone is an agonist

An analog of alpha-factor, the Saccharomyces cerevisiae tridecapeptide mating pheromone (Trp-His-Trp-Leu-Gln-Leu-Lys-Pro-Gly-Gln-Pro-Met-Tyr), in which the side chains of Lys7 and Gln10 were covalently linked, was synthesized using solid phase methodologies. The yield of the purified cyclic analog cyclo7,10[Nle12]alpha-factor was 30%, and its structure was verified by amino acid analysis, peptide sequencing, fast atom bombardment-mass spectrometry, and proton nuclear magnetic resonance spectroscopy. Cyclo7,10[Nle12]alpha-factor caused growth arrest and morphological alterations in S. cerevisiae MATa cells qualitatively identical to those induced by linear pheromone and was one-fourth to one-twentieth as active as the linear alpha-factor depending upon the S. cerevisiae strain tested. Consistent with the relative activities of the linear and cyclic peptides, binding competition studies indicated that cyclo7,10[Nle12]alpha-factor had approximately 20-40-fold less affinity for the alpha-factor receptor. Hydrolysis of the cyclic peptide by the target cells did not lead to opening of the ring and was less rapid than that of linear alpha-factor. The alpha-factor antagonist des-Trp1-[Ala3,Nle12]alpha-factor reversed the activity of the cyclic analog, and cyclo7,10[Nle12]alpha-factor was not active at the restrictive temperature in a temperature-sensitive receptor mutant. These results support the conclusion that the cyclic alpha-factor occupies the same binding site within the receptor as is occupied by the natural pheromone. The cyclic alpha-factor represents a rare example of an agonist among covalently constrained congeners of small linear peptide messengers.     

Triterpenoid saponins from Clinopodium polycephalum.

A new triterpenoid saponin, clinopodiside A, has been isolated from Clinopodium polycephalum. Its structure was established by spectroscopic methods and X-ray diffraction analysis as 3-O-beta-D-glucopyranosyl(1----6)-[ beta-D-glucopyranosyl (1----4)]-beta-D-glucopyranosyl-olean-11,13(18)-diene-3 beta,16 beta, 23,28-tetrol.     

体外重构STUB1介导α-1抗胰蛋白酶突变体Z蛋白泛素化修饰反应体系

α-1抗胰蛋白酶Z型突变体蛋白(α-1 antitrypsin Z-mutant protein, ATZ)是引发α-1抗胰蛋白酶缺陷症(α-1 antitrypsin deficiency, AATD)的主要原因,研究ATZ蛋白的泛素化修饰和降解对于治疗AATD具有重要意义。STUB1是一种重要的E3泛素连接酶,参与调节多种蛋白质的泛素化修饰。然而,STUB1是否参与ATZ的泛素化修饰尚未明确。本研究首先将ATZ和STUB1的编码基因克隆到pET28a质粒,构建了这2个蛋白的表达质粒。随后,将重组质粒转入大肠杆菌表达系统,在优化诱导条件实现了重组蛋白的异源表达。通过金属螯合亲和层析技术纯化得到目的蛋白,并通过蛋白质谱分析验证了其氨基酸序列的准确性。利用纯化的ATZ和STUB1重组蛋白,构建了一个体外泛素化修饰反应体系。实验结果显示,在ATP、E1泛素激活酶和E2泛素结合酶的协同作用下,STUB1成功催化了ATZ的泛素化修饰。本研究提供了一种体外获得Z型突变体ATZ纯化蛋白的方法,并确认了STUB1介导ATZ的泛素化修饰功能,推进了对α-1抗胰蛋白酶Z型突变体蛋白在细胞内降解过程的调控机制的理解。     

Hemin with Peroxidase Activity Can Inhibit the Oxidative Damage Induced by Ultraviolet A

Excessive reactive oxygen species (ROS), a highly reactive substance that contains oxygen, induced by ultraviolet A (UVA) cause oxidative damage to skin. We confirmed that hemin can catalyze the reaction of tyrosine (Tyr) and hydrogen peroxide (H₂O₂). Catalysis was found to effectively reduce or eliminate oxidative damage to cells induced by H₂O₂ or UVA. The scavenging effects of hemin for other free-radical ROS were also evaluated through pyrogallol autoxidation, 1,1-diphenyl-2-picrylhydrazyl radical (DPPH·)-scavenging assays, and phenanthroline–Fe²⁺ assays. The results show that a mixture of hemin and tyrosine exhibits strong scavenging activities for H₂O₂, superoxide anion (O₂⁻·), DPPH·, and the hydroxyl radical (·OH). Furthermore, the inhibition of oxidative damage to human skin keratinocyte (HaCaT) cells induced by H₂O₂ or UVA was evaluated. The results show that catalysis can significantly reduce the ratio of cell apoptosis and death and inhibit the release of lactate dehydrogenase (LDH), as well as accumulation of malondialdehyde (MDA). Furthermore, the resistance to apoptosis was found to be enhanced. These results show that the mixture of hemin and tyrosine has a significantly protective effect against oxidative damage to HaCaT cells caused by UVA, suggesting it as a protective agent for combating UVA damage.     

Protein kinase C delta null mice exhibit structural alterations in articular surface,intra-articular and subchondral compartments

IntroductionStructural alterations in intra-articular and subchondral compartments are hallmarks of osteoarthritis, a degenerative disease that causes pain and disability in the aging population. Protein kinase C delta (PKC-δ) plays versatile functions in cell growth and differentiation, but its role in the articular cartilage and subchondral bone is not known.MethodsHistological analysis including alcian blue, safranin O staining and fluorochrome labeling were used to reveal structural alterations at the articular cartilage surface and bone–cartilage interface in PKC-δ knockout (KO) mice. The morphology and organization of chondrocytes were studied using confocal microscopy. Glycosaminoglycan content was studied by micromass culture of chondrocytes of PKC-δ KO mice.ResultsWe uncovered atypical structural demarcation between articular cartilage and subchondral bone of PKC-δ KO mice. Histology analyses revealed a thickening of the articular cartilage and calcified bone–cartilage interface, and decreased safranin O staining accompanied by an increase in the number of hypertrophic chondrocytes in the articular cartilage of PKC-δ KO mice. Interestingly, loss of demarcation between articular cartilage and bone was concomitant with irregular chondrocyte morphology and arrangement. Consistently, in vivo calcein labeling assay showed an increased intensity of calcein labeling in the interface of the growth plate and metaphysis in PKC-δ KO mice. Furthermore, in vitro culture of chondrocyte micromass showed a decreased alcian blue staining of chondrocyte micromass in the PKC-δ KO mice, indicative of a reduced level of glycosaminoglycan production.ConclusionsOur data imply a role for PKC-δ in the osteochondral plasticity of the interface between articular cartilage and the osteochondral junction.

Electronic supplementary material

The online version of this article (doi:10.1186/s13075-015-0720-4) contains supplementary material, which is available to authorized users.     

中国野生动物调查存在的问题与改进建议

一百余年前,人类就开始了相对系统的野生动物调查。目前已经建立了成熟的方法体系,并制定了相应的调查规范。最近几十年来,我国科研人员进行了大量的野生动物调查。但是,当前我国的野外调查规范还不够细致,调查者在调查时缺乏必要的约束,导致调查数据不规范、不可靠,很多重要信息缺失。突出问题有：样线信息不全,只有起点和终点的经纬度,没有自动记录的详细样线信息（如每秒记录一次的连续点位信息,含经纬度和时间）;动物位点信息缺乏可信度指标（如距观测者的距离）;调查时间不合理;调查地点的空间取样不均衡;记录的标准化不够（如每个观测点的观测时长不确定）等等。对此,我们参考国际通用的调查规范,提出了一些简单易行的调查要点,以便提高野外调查数据的质量。另外,我们提倡野外记录的无纸化,充分利用现有的手机应用软件（APP）和模型工具提高野外记录以及后期数据处理的效率。最后,我们提议建立固定的中国生物多样性监测样线体系,以便消除每年调查时空间取样的不确定性,更好地量化野生动物的时间动态,为野生动物的保护和管理提供依据。     

Studies on Some Properties of the Apoplastic β-N-Acetyl-D-Hexosaminidase from Tomato Leaves

Some properties of the β-N-acetyl-D-hexosaminidase purified from intercellular fluid of tomato leaves after the plant was systematically infected by TMV (tobacco mosaic virus) were studied. When pNP β-D-GlcNAc (p nitrophenyl-N-aeetyl β-D-glucosaminide) or pNP β-D- GalNAc (p-nitrophenyl-N-acetyl-β-D galactosaminide) was used as the substrate, it showed the optical pH between 4. 8--5.0 and optical temperature between 44— 47℃. Studies of thermostabillty indicated that the enzyme had a biphasic denaturation curve. Using pNP-β-D-GIcNAc or pNP-β-D GalNAc as the substrate, the Km value of the enzyme was 0. 36 and 0. 67 mmol/L respectively. N acetyi-D glucosamine and N acetyl-D-galactosamine were competitive inhibitors of the enzyme activities. Ag+ and Hg2+ were sensitive inhibitors and Fe2+ . Fe3+ and Cu2+ were also inhibitors enzyme activities.     

Role of IKK/NF-κB signaling in extinction of conditioned place aversion memory in rats

The inhibitor κB protein kinase/nuclear factor κB (IKK/NF-κB) signaling pathway is critical for synaptic plasticity. However, the role of IKK/NF-κB in drug withdrawal-associated conditioned place aversion (CPA) memory is unknown. Here, we showed that inhibition of IKK/NF-κB by sulphasalazine (SSZ; 10 mM, i.c.v.) selectively blocked the extinction but not acquisition or expression of morphine-induced CPA in rats. The blockade of CPA extinction induced by SSZ was abolished by sodium butyrate, an inhibitor of histone deacetylase. Thus, the IKK/NF-κB signaling pathway might play a critical role in the extinction of morphine-induced CPA in rats and might be a potential pharmacotherapy target for opiate addiction.