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1.
Tihomir Todorov Albena Todorova Bilyana Georgieva Vanyo Mitev 《Molecular biotechnology》2010,45(2):150-154
We report on a unified rapid betaine-based-PCR protocol for amplification of the (CAG)n region in Huntington disease (HD) and the (CGG)n region in Fragile X syndrome (FXS), followed by an electrophoretic separation on automated sequencer for precise determination
of the triplet numbers. The high betaine concentration (2.5 M betaine) permits precise amplification of the CAG and CGG repeats.
Ten HD affected patients and 10 healthy individuals from HD families were re-evaluated. For FXS the CGG region in normal individuals
and premutations of about 100 repeats were precisely amplified by this protocol. Ten unrelated FXS premutation carriers and
24 mentally retarded non-FXS affected boys were re-examined by this method. The results totally coincided with the previous
ones. This protocol is a good choice as a fast screening test. Within 24 h we can have preliminary information on the patient’s
genetic status. Normal individuals, CGG premutation carriers up to 100 repeats, as well as HD patients carrying an expansion
up to 50 CAG repeats can be easily clarified. This accounts for a relatively large proportion (about 90%) of the suspected
HD and FXS patients, referred to our laboratory for genetic analysis. The calculation of the repeat’s number is more accurate
for the correct interpretation of the results, screening tests and genetic counselling. 相似文献
2.
Tihomir Paul Obrenovitch Jutta Urenjak Douglas Arthur Richards † Yukihiko Ueda Gerald Curzon † Lindsay Symon 《Journal of neurochemistry》1993,61(1):178-186
Abstract: Changes in the extracellular levels of excitatory and inhibitory amino acid transmitters were studied in the rat striatum during penumbral ischaemia using intracerebral microdialysis. Effects of penumbral forebrain ischaemia were compared with those of ischaemia with sustained anoxic depolarisation and K+ (100 m M ). Comparisons were also made between different groups of animals at 2 and 24 h after dialysis probe implantation. The K+ stimulus did not provoke any release of excitatory amino acids in the 24-h group, probably reflecting a decrease of functional synapses adjacent to the probe. During 30 min of penumbral ischaemia, excitatory amino acids did not reach critical concentrations in the extracellular fluid, and increases in levels of inhibitory/modulatory amino acids were similar. On the other hand, severe transient ischaemia resulted in massive synchronous release of many neuroactive excitatory and inhibitory compounds, in both the 2- and 24-h groups. These and other data suggest that changes during severe ischaemia may arise from both neurotransmitter and metabolic pools. It is concluded that is- chaemic damage in the penumbra may not be related to extracellular neuroactive amino acid changes generated within this region. 相似文献
3.
Paula M. Nogueira Rafael R. Assis Ana C. Torrecilhas Elvira M. Saraiva Natália L. Pessoa Marco A. Campos Eric F. Marialva Cláudia M. Ríos-Velasquez Felipe A. Pessoa Nágila F. Secundino Jer?nimo N. Rugani Elsa Nieves Salvatore J. Turco Maria N. Melo Rodrigo P. Soares 《PLoS neglected tropical diseases》2016,10(8)
The immunomodulatory properties of lipophosphoglycans (LPG) from New World species of Leishmania have been assessed in Leishmania infantum and Leishmania braziliensis, the causative agents of visceral and cutaneous leishmaniasis, respectively. This glycoconjugate is highly polymorphic among species with variation in sugars that branch off the conserved Gal(β1,4)Man(α1)-PO4 backbone of repeat units. Here, the immunomodulatory activity of LPGs from Leishmania amazonensis, the causative agent of diffuse cutaneous leishmaniasis, was evaluated in two strains from Brazil. One strain (PH8) was originally isolated from the sand fly and the other (Josefa) was isolated from a human case. The ability of purified LPGs from both strains was investigated during in vitro interaction with peritoneal murine macrophages and CHO cells and in vivo infection with Lutzomyia migonei. In peritoneal murine macrophages, the LPGs from both strains activated TLR4. Both LPGs equally activate MAPKs and the NF-κB inhibitor p-IκBα, but were not able to translocate NF-κB. In vivo experiments with sand flies showed that both stains were able to sustain infection in L. migonei. A preliminary biochemical analysis indicates intraspecies variation in the LPG sugar moieties. However, they did not result in different activation profiles of the innate immune system. Also those polymorphisms did not affect infectivity to the sand fly. 相似文献
4.
Maines MD Miralem T Lerner-Marmarosh N Shen J Gibbs PE 《The Journal of biological chemistry》2007,282(11):8110-8122
Human biliverdin reductase (hBVR), a dual specificity kinase (Ser/Thr/Tyr) is, as protein kinase C (PKC) betaII, activated by insulin and free radicals (Miralem, T., Hu, Z., Torno, M. D., Lelli, K. M., and Maines, M. D. (2005) J. Biol. Chem. 280, 17084-17092; Lerner-Marmarosh, N., Shen, J., Torno, M. D., Kravets, A., Hu, Z., and Maines, M. D. (2005) Proc. Natl. Acad. Sci. U. S. A. 102, 7109-7114). Here, by using 293A cells co-transfected with pcDNA3-hBVR and PKC betaII plasmids, we report the co-immunoprecipitation of the proteins and co-purification in the glutathione S-transferase (GST) pulldown assay. hBVR and PKC betaII, but not the reductase and PKC zeta, transphosphorylated in assay systems supportive of activity of only one of the kinases. PKC betaII K371R mutant protein ("kinase-dead") was also a substrate for hBVR. The reductase increased the Vmax but not the apparent Km values of PKC betaII for myelin basic protein; activation was independent of phospholipids and extended to the phosphorylation of S2, a PKC-specific substrate. The increase in substrate phosphorylation was blocked by specific inhibitors of conventional PKCs and attenuated by sihBVR. The effect of the latter could be rescued by subsequent overexpression of hBVR. To a large extent, the activation was a function of the hBVR N-terminal chain of valines and intact ATP-binding site and the cysteine-rich C-terminal segment. The cobalt protoporphyrin-activated hBVR phosphorylated a threonine in a peptide corresponding to the Thr500 in the human PKC betaII activation loop. Neither serine nor threonine residues in peptides corresponding to other phosphorylation sites of the PKC betaII nor PKC zeta activation loop-derived peptides were substrates. The phosphorylation of Thr500 was confirmed by immunoblotting of hBVR.PKC betaII immunocomplex. The potential biological relevance of the hBVR activation of PKC betaII was suggested by the finding that in cells transfected with the PKC betaII, hBVR augmented phorbol myristate acetate-mediated c-fos expression, and infection with sihBVR attenuated the response. Also, in cells overexpressing hBVR and PKC betaII, as well as in untransfected cells, upon treatment with phorbol myristate acetate, the PKC translocated to the plasma membrane and co-localized with hBVR. hBVR activation of PKC betaII underscores its potential function in propagation of signals relayed through PKCs. 相似文献
5.
Purification and characterization of an extracellular lipase from a thermophilic Rhizopus oryzae strain isolated from palm fruit 总被引:2,自引:0,他引:2
Hiol A Jonzo MD Rugani N Druet D Sarda L Comeau LC 《Enzyme and microbial technology》2000,26(5-6):421-430
We have isolated a lipolytic strain from palm fruit that was identified as a Rhizopus oryzae. Culture conditions were optimized and highest lipase production amounting to 120 U/ml was achieved after 4 days of cultivation. The extracellular lipase was purified 1200-fold by ammonium sulfate precipitation, sulphopropyl-Sepharose chromatography, Sephadex G 75 gel filtration and a second sulphopropyl-Sepharose chromatography. The specific activity of the purified enzyme was 8800 U/mg. The lipolytic enzyme has a molecular mass of 32 kDa by SDS-polyacrylamide gel electrophoresis and gel filtration. The enzyme exhibited a single band in active polyacrylamide gel electrophoresis and its isoelectric point was 7.6. Analysis of Rhizopus oryzae lipase by RP-HPLC confirmed the homogeneity of the enzyme preparation. Determination of the N-terminal sequence over 19 amino acid residues showed a high homology with lipases of the same genus. The optimum pH for enzyme activity was 7.5. Lipase was stable in the pH range from 4.5 to 7.5. The optimum temperature for lipase activity was 35 degrees C and about 65% of its activity was retained after incubation at 45 degrees C for 30 min. The lipolytic enzyme was inhibited by Triton X100, SDS, and metal ions such as Fe(3+), Cu(2+), Hg(2+) and Fe(2+). Lipase activity against triolein was enhanced by sodium cholate or taurocholate. The purified lipase had a preference for the hydrolysis of saturated fatty acid chains (C(8)-C(18)) and a 1, 3-position specificity. It showed a good stability in organic solvents and especially in long chain-fatty alcohol. The enzyme poorly hydrolyzed triacylglycerols containing n-3 polyunsaturated fatty acids, and appeared as a suitable biocatalyst for selective esterification of sardine free fatty acids with hexanol as substrate. About 76% of sardine free fatty acids were esterified after 30 h reaction whereas 90% of docosahexaenoic acid (DHA) was recovered in the unesterified fatty acids. 相似文献
6.
Lara Saraiva Alanna Silva Reis Jeronimo Marteleto Nunes Rugani Agnes Ant?nia Sampaio Pereira Felipe Dutra Rêgo Ana Cristina Vianna Mariano da Rocha Lima Célia Maria Ferreira Gontijo José Dilermando Andrade Filho 《PloS one》2015,10(8)
Brazil is one of the most important endemic areas for leishmaniasis worldwide. Protected areas that are tourist attractions likely present an important risk of transmission of cutaneous leishmaniasis (CL). Furthermore, with the geographical expansion of visceral leishmaniasis (VL), several studies have recorded the occurrence of its vector, Lutzomyia longipalpis, and cases of human and canine VL in such tourist areas. The Parque Estadual do Sumidouro is an environmentally protected area located in the Brazilian Cerrado biome and in an important area endemic for leishmaniasis in the state of Minas Gerais. The purpose of this study was to monitor the sand fly fauna in areas of tourist activity in the park. Sampling was performed every month, from September 2011 to August 2013, using CDC light traps at six sites of differing environmental characteristics. Sampled specimens were identified following Galati (2003), and females were submitted to molecular techniques for the detection and identification of Leishmania DNA. A total of 4,675 sand fly specimens of 25 species belonging to nine genera were collected. The most abundant species were Micropygomyia quinquefer, Lutzomyia renei and Pintomyia pessoai, although only Pi. pessoai is implicated in the transmission of Leishmania braziliensis. The species accumulation curve reached saturation on the 16th sampling event. Species richness, diversity and evenness differed among the sampled areas. The seasonal curve was not determined by a single unique species, and no single species was the most abundant in all environments sampled. The main vector of Leishmania (Leishmania) infantum, Lutzomyia longipalpis, accounted for only 5.35% of the specimens collected. Proven or suspected vectors of Leishmania (Viannia) braziliensis were recorded, and one female of the cortellezzii complex tested positive for Le. braziliensis DNA. Even with a low infection rate (0.62%), these data indicate the circulation of the parasite and reinforce the need for entomological and epidemiological surveillance in the park and its surroundings. 相似文献
7.
Clastic sediments of Middle–Late Eocene age were studied on the Island of Rab (northern Adriatic Sea, Croatia) in order to
reconstruct their depositional history, depositional environments, and geometry of sandstone bodies. Detailed outcrop logging
and mapping revealed the response of depositional systems to frequent relative sea-level changes, which initiated significant
basinward and landward shifts of facies, respectively. Tidal sandstones are commonly underlain by shoreface sandstones, and
overlain by offshore sandy marls, whereas the latter are again overlain by shoreface sandstones. Major relative sea-level
falls initiated basinward shift of depositional systems and the incision of incised valleys or estuaries, and consequently
truncated the underlying shelf sediments. In some cases, the accelerated sea-level fall caused rapid shoreface progradation
which is interpreted as a forced regression. Relative sea-level rise caused flooding of the incised relief, and deposition
of tidal sandstone bodies which overlie type-I sequence boundaries. The coarse lag sediment of these sequence boundaries locally
disappears laterally, and the boundary is granulometrically less prominent. All of the major bounding surfaces have been recognized
in the sections studied, although the maximum flooding surface is recognized as a thin “zone” instead of a single surface.
Altogether, 28 complete sequences, and 15 parasequences are recognized in the informal unit of the Lopar sandstones, documenting
the depositional response to high-frequency relative sea-level oscillations. They have so far not been recognized in the Eocene
of the eastern Adriatic region. 相似文献
8.
Antitumor effect of Croatian propolis as a consequence of diverse sex-related dihydropyrimidine dehydrogenase (DPD) protein expression 总被引:1,自引:0,他引:1
9.
The potential environmental suitability and economic viability of growing two biodiesel crops in marginal regions of Australia were explored. Firstly, we used spatial analysis techniques to identify marginal agricultural regions suitable for growing pongam (Pongamia pinnata) and Indian mustard (Brassica juncea), and determined the base socioeconomic viability of investments for the production of biodiesel in the identified areas. Secondly, we used climate change projections (target years 2020 to 2070) from the Commonwealth Scientific, Industrial and Research Organization Mk3.0 global circulation model generated for two emission scenarios (A1B and A1FI) to determine the shift in potential areas for these crops. Under the climate change scenarios tested, the total area suitable for growing pongam between 2040 and 2070 is substantially different from the suitable area under current climate, indicating that long-term investments in this perennial tree crop may not be viable in all regions, especially in southern Australia. There is a greater variation in suitability projections for Indian mustard, although there is more flexibility for cropping options given that it is an annual crop. However, future economic viability is likely to depend on the ability to receive renewable energy certificates for both crops and, in the case of pongam, the certified emission reductions. Opportunities exist for sustainable pongam agroforestry to supply biodiesel to regional towns, cattle stations and mines in northern Australia. 相似文献
10.
Vukojević N Sikić J Curković T Juratovac Z Katusić D Sarić B Jukić T 《Collegium antropologicum》2005,29(Z1):25-27
Changes in the eye axial diameter were studied to assess the eye globe impact of conventional operation for retinal detachment. The study included 69 eyes in 69 patients operated on for rhegmatogenous retinal detachment. There were 46.4% of men and 53.6% of women, mean age 52.7 (+/- 15.21) years. Results of preoperative and postoperative ultrasonographic measurement of axial diameter are presented. The mean preoperative and postoperative eye axial diameter was 23.69 (+/- 1.84) mm and 24.43 (+/- 1.91) mm, respectively. Postoperative results showed the axial eye length to increase by a mean of 0.74 (+/- 0.44) mm, yielding a statistically significant difference from the preoperative measurement (p < 0.001). The mean myopia induced by this eyeball elongation was 1.77 D. The encircling band with and without segmental buckling used in surgical repair of retinal detachment creates circular and segmental indentation of the eyeball, thus increasing its axial length. The myopia induced by elongation of the eyeball results in considerable myopia, which requires appropriate correction in the early postoperative period to achieve favorable vision rehabilitation. 相似文献