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2008年  第29卷  第5期

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研究论文
Phylogenetic relationships of 36 Fringillidae species based on mitochondrial CoI gene sequences were reconstructed using the Bayesian (BI) maximum-likelihood (ML) and Neighbor-joining(NJ)methods. The results suggest that the relationship of Calcarius lapponicus and Emberiza is more closely related than other Fringillidae species.Our results support the view that Latoucheornis siemsseni is included in Genus Emberiza as E. siemsseni, and confirms that there is a close relationship between Mycerobas affinis and Eophona migratorius. This study also finds that Urocynchramus pylzowi is distantly related to Uragus sibiricus, but Uragus sibiricus and Carpodacus are closely related. The result also supports that fringillids and emberizids can be listed into two subfamilies under the family Fringillidae: Fringillinae and Emberizinae.
The cDNA encoding the luciferase from lantern mRNA of one diurnal firefly Pyrocoelia pygidialis Pic, 1926 has been cloned, sequenced and functionally expressed. The cDNA sequence of P. pygidialis luciferase is 1647 base pairs in length, coding a protein of 548 amino acid residues. Sequence analysis of the deduced amino acid sequence showed that this luciferase had 97.8% resemblance to luciferases from the fireflies Lampyris noctiluca, Lampyris turkestanicus and Nyctophila cf. caucasica. Phylogenetic analysis using deduced amino acid sequence showed that P. pygidialis located at the base of Lampyris+Nyctophila clade with robust support (BP=97%); but did not show a monophyletic relationship with its congeneric species P. pectoralis, P. rufa and P. miyako, all three are strong luminous and nocturnal species. The expression worked in recombinant Escherichia coli. Expression product had a 70 kDa band and emitted yellow-green luminescence in the presence of luciferin. Five loops in the P. pygidialis luciferase, L1 (N198-G208), L2 (T240-G247), L3 (G317-K322), L4 (L343-I350) and L5 (G522-D532), were found from the structure modeling analysis in the cleft, where it was considered the active site for the substrate compound entering and binding. Different amino acid residues between the luciferases of P. pygidialis and the three other known strong luminous species can not explain the situation of weak or strong luminescence. Future study of these loops, residues or crystal structure analysis may be helpful in understanding the real differences between the luciferases between diurnal and nocturnal species.
Myostatin (MST) is the cytokine negatively regulated skeletal muscle growth factor, and the loss of function caused by mutations is associated with increased skeletal muscle mass. The multiple sequence alignment of the available MST cDNA sequences and the related evolutionary analyses were performed; the quarternary structures of MST homodimer and the MST: ActRIIB complex was built using homology modelling technique. The phylogenetic tree revealed that there were four sub-families, i.e., mammalian MSTN, avian MSTN, and fish MSTN1, 2 in the MST gene (MSTN) family. MST orthologous genes derived from different species had a high sequence identity, which were probably caused by the purifying selection. Particularly, the high degree of conservation of C-terminal sequences of mammalian and avian MST mature peptides suggested they shared the very similar structure, function and signal transduction pathway. The analyses of electrostatic potential and hydrophobic amino acid distribution of the structural models revealed that the electrostatic force and hydrophobic interaction played an important role in receptor recognition by MST.
In vertebrates, non-lens bg-crystallins are widely expressed in various tissues and their functions are not well known. The molecular mechanisms of trefoil factors (TFFs), which involved in mucosal healing and tumorigenesis, have remained elusive. bg-CAT is a novel multifunctional protein complex of non-lens bg-crystallin and trefoil factor from frog skin secretions. Here we report that bg-CAT could induce sustained contraction of isolated rabbit aortic rings in dosage (2-35 nmol/L) and endothelium dependent manners (P<0.01). In addition, in situ immunofluorescence indicated that positive TNF-a signals were mainly detected at the endothelial cell layer of bg-CAT (25 nmol/L) treated rings. Furthermore,bg-CAT induced primary cultured rabbit thoracic aortic endothelial cells (RAECs) rapidly to release TNF-a. After bg-CAT (25 nmol/L) treated for 10 and 30 min, the levels of the endothelial cells released TNF-a were 34.17

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βγ-CAT is a naturally existing multifunctional protein complex of non-lens βγ-crystallin and trefoil factor from Bombina maxima skin secretions. In this paper, we first analysed the time-response curve of hemolysis and intracellular potassium efflux induced by βγ-CAT on human erythrocytes. After βγ-CAT (3 nmol/L) in human erythrocytes was treated at 37℃ for 5 min, we found that about 93.31±5.89% (P<0.01) of the intracellular potassium ions leaked, whereas the percentage of hemolysis was only13.12±1.92 % (P<0.05). We employed electron microscopy to observe the erythrocytes’ morphological changes and found erythrocytes were swollen and some had globular structures occurring on the cell surfaces, and fast releasing hemoglobins. These results indicated that the hemolytic effect of βγ-CAT on human erythrocyte membranes was due to intracellular potassium ion’s rapid efflux. These results demonstrate morphological proof in understanding the mechanism of βγ-CAT pore-forming effect on human erythrocytes.
Cobra venom factor (CVF) depletes complement C3 and may therefore be important in preventing the complement-mediated damage and disease. In comparison with various CVF reported previously, Yunnan-cobra venom factor (Y-CVF) has higher anticomplement activity. The goal for this research is to investigate whether Y-CVF could induce the specific neutralized anti-Y-CVF antibody and xenoantibodies in non-human primates. Thus two cynomolgus monkeys were intravenously injected with Y-CVF (0.05 mg/kg) every two weeks for totally four times. The serum C3, CH50, anti-Y-CVF antibody and xenoantibody levels were measured at different time points before and after Y-CVF injection. The results revealed that, complement C3 was effectively depleted during the first two injections, and incompletely depleted during the third injection, but almost not depleted during the last injection. The results of Western blot and ELISA confirmed the production of anti-Y-CVF antibody, and its titer increased with the injection. Additionally no significant changes of anti-porcine endothelia cell xenoantibodies were found measured by flow cytometry. We concluded that multiple injection of Y-CVF stimulated anti-Y-CVF antibody production in monkeys, which resulted in the invalid of Y-CVF. The induction of anti-alpha -Gal xenoantibody by Y-CVF in primates was not observed.
Hepatocyte growth factor (HGF) is a pleiotropic factor, which plays an important role in the nervous system. Previously, we had produced highly enriched (88.3±8.1%) transplantable populations of neural progenitors from rhesus monkey embryonic stem cells (rESCs) using an embryoid body (EB) system containing HGF and G5 supplement. However, the exact role of HGF in the neural differentiation process as well as the mechanism of HGF is not clear. Here, on the basis of the ameliorated differentiation system, we efficiently produced populations of neural progenitors (81.66±4.37%) from rESCs in an adherent monoculture system containing HGF and basic fibroblast growth factor (bFGF) within two weeks. Moreover, we obtained similar results with the systems containing either HGF or bFGF alone, or neither of them, which implied that extrinsic HGF can not determine the neural fate of the rESCs in the differentiation process. Further results suggested HGF with bFGF synergistically promoted the proliferation of the neural progenitors derived from rESCs. In conclusion, through this study, a much simpler method of inducing rESCs differentiation towards neural cells has been established and extrinsic HGF did not function in neural induction from rESCs to neural progenitor cells. However, HGF with bFGF synergistically promoted the proliferation of the neural progenitors derived from rESCs.
In order to investigate the estrogen and estrogen receptor β changes after mating behavior of male mandarin vole (Microtus mandarinus), the radioimmunoassay (RIA) and immunohistochemistry methods were used to investigate changes of the serum estrogen (E) concentrations, estrogen immunoreactive neurons (E-IRs) and estrogen receptor β immunoreactive neurons (ERβ-IRs) in the relevant brain regions following mating behavior. Fifteen sexually matured male voles were randomly divided into three groups and treated differently: (1) control group: voles were exposed to clean hard-wood shavings (n=5), (2) exposure group: voles were exposed to the soiled bedding for more than 24 h on which estrous females had been placed (n=5), and (3) mating group: voles were placed with an estrous female for more than 24 h (n=5). The results showed circulating serum E concentrations were significantly higher in the mating group than in the exposure group and the control group, and there were no significant difference between the exposure group and the control group. E-IRs and ERβ-IRs were detected in the following brain regions related to mating behavior: the arcuate nucleus (ARC), bed nucleus of the stria terminalis (BST), lateral septal nucleus (LS), medial amygdaloid nucleus (ME), medial preoptic area (MPO) and ventromedial hypothalamic nucleus (VMH). The results showed that there were significantly more E-IRs in the six brain regions in the mating group than in the control group and the exposure group, and there were no significant difference between the exposure group and the control group except for LS. There was no significant difference in ERβ-IRs in the six brain regions among the three groups, and there were some lighter -stained ERβ-IRs in these brain regions. The results suggested that estrogen affect mating activity of male mandarin voles, but ERβ might not play an important role in mating behavior of male mandarin voles. Instead, it might be through other receptors.
The purpose of this article is to detect sex and age difference in the structure of the olfactory bulb in dogs by histological methods. The thickness of the olfactory bulbs layers and its main cells were analyzed comparatively with the methods of HE-staining and statistics, through which we studied the development course of dogs’ olfactory bulb and the structural differences which affect the olfaction in both males and females. The results showed that between both male and female juveniles and adult males and females, the difference in thickness of each layer is not significant. But the difference in quantity of mitral cells between adult males and females was significant. Meanwhile, the structure of every layer in juvenile dogs was apparent while the volume and the weight of adult dogs’ olfactory bulb and each layer’s width increased significantly. On the other hand, the density of each layer’s cells decreased apparently. Our results demonstrated that the olfactory bulb developed with age, and the apparent differences in morphology and quantity of mitral cells between males and females may be one of the reasons leading to the sexual variations of olfactory sensitivity.
From 2000 to 2002,we investigated the distribution of red panda in 11 counties in Qionglai Mountains of Sichuan Province. After surveys on 2 332 line transects, tracks of the species were found in 130 line transects (5.57%), which distributed in nine counties and concentrated in Wenchuan County and Chongzhou City. The area of red panda habitat amounted to approximately 5107.82 km2, in which Baoxing (34.08%), Wenchuan (30.65%) and Tianquan (16.61%) counties were larger than the others and counted for 81.34% of the total. The highest density was in the Anzihe Nature Reserve of Chongzhou City. In the seven reserves of Qionglai Mountains, the area of red panda’s habitat covered 2 166.84 km2 and was 42.4% of the total. The distribution of red panda in Qionglai Mountains was discrete and existed many gaps and fragmentation trend. Thus, the protection of red panda in Qionglai Mountains should be further enhanced.
From October 2004 to May 2005, flock size and composition of wintering black-necked cranes (Grus nigricollis) were studied using spot scanning technique at Napahai Nature Reserve in Yunnan province. At night, the mean wintering population size was 67.9 (16-157, n=17). Composed by common crane (Grus grus) or not, roost cranes were divided into sing-species flock and mixed-species flock. The numbers of crane in sing-species flock were 65.3% of total black-necked cranes. In the day, feeding flocks were classified into three types: family group comprised of breeding pair and their offspring ranging between two to four; sub-adult flock composed mainly by sub-adults ranging between three to 65 with an average size of 16.1(n=1017); special flock composed by single cranes. Black-necked crane flocks were affected by season, temperature, breeding status, food abundance and availability, and their sizes changed significantly in daytime or between different months (P=0.000<0.05). After family breakup happened in late March, Juveniles were drove away by their parents and then they got together to join the sub-adult flocks.
The bird community of the middle-mountain moist evergreen broad-leaved forest, Xujiaba, Ailao Mountain, were directly observed from March to April in 2006. Throughout the observation, 14 345 times of free foraging behavior were observed. The study adopted the method of line transect without estimations. Partitioning occurs primarily by structure of forging behavior of birds. Through cluster (Furthest Neighbor) , the 62 birds species were divided into 11 vertical spatial perch guilds;(G1) ground gleaning foraging guild; (G2) ground scratching foraging guild; (G3) canopy aerial foraging/gleaning foraging guild; (G4) canopy aerial foraging guild; (G5) trunk barking foraging guild; (G6) brush of bamboo pole barking foraging guild; (G7) trunk bough searching foraging guild; (G8) canopy bough searching foraging guild; (G9) canopy gleaning foraging guild; (G10) underbrush foliage/trunk/ground gleaning foraging guild; and (G11) brush gleaning foraging guild. The results indicate that each guild partitions the fooding space and the food resource in this area because of the different perch substrate, foraging substrate and foraging methods. In each guild, bird species can be further divided mainly by foraging height, so that the fooding space and the food resource get further division in the guild, which minimizes the competition between species. Finally, all species in the community can reach dynamic balance. At the same time, the numbers and structures of the bird foraging guild are different in various vegetation types. There are more bird species in the climax community, and their ecology-like status are much more different.
The community structure and species diversity of litter-layer beetles in five habitats in Baijitan National Nature Reserve of Ningxia were surveyed from July to September in 2006. A total of 1896 beetles were collected by pitfall traps and other methods during field research. The result showed that all specimens belong to seventeen families. The individual of Tenebrionidae was the maximum, accounts for 25.58%; Chrysomelidae takes the second place, accounts for 20.99%; the individuals of Eumolpidae, Meloidae and Coccinellidae were also rich more than other families. These five families were considered as dominant groups. Among the five habitats, the individuals and species abundance of litter-layer beetles were the highest in the Baijitan plot, but the lowest in the Maanshan plot. The indices of species richness, diversity and evenness were the highest in the Changliushui plot, while the lowest in the Maanshan plot. Cluster analysis showed that the community structures and habitats were similar among the Daquan plot, Ciyaobao plot, Changliushui plot as well as Baijitan plot. It is particularly more similar between Changliushui plot and Ciyaobao plot. The community composition and diversity of litter-layer beetles were closely related to the spatial heterogeneity which was restored by the practice of desertification control.