Capillary electrophoresis reveals polyamine metabolism modulation in Leishmania (Leishmania) amazonensis wild-type and arginase-knockout mutants under arginine starvation.

l-Arginine is an essential amino acid in Leishmania (Leishmania) amazonensis metabolism. A key enzyme for parasite l-arginine metabolism is arginase (ARG) that uses arginine to produce urea and ornithine, a precursor of polyamine pathway guaranteeing parasite replication in both insect and mammal hosts. There is an alternative pathway to produce ornithine via l-proline and glutamate, but this mechanism is not described in Leishmania. In the mammal host, two enzymes can use l-arginine as substrate, the host ARG and the induced nitric oxide synthase that produces nitric oxide. The competition between induced nitric oxide synthase and both parasite and host ARG can favor the success of the infection or its control. Here, we established the metabolomics profile of the polyamine pathway of wild type (WT) L. (L.) amazonensis, submitted or not to l-arginine starvation, and compared to the ARG-knockout mutant (arg- ). Our results indicated that arginine starvation induces a decrease in arginine, ornithine, and putrescine, but we could not detect the significative level changes of spermidine, spermine, or agmatine. However, the absence of ARG on the arg- induced an increase of arginine and citrulline levels, but decreased the levels of ornithine and putrescine. Similarly to the WT arginine-starved parasites, the arg- parasites presented lower levels of proline when compared to the WT ones. This could be indicative of an alternative pathway to surpass the enzyme or its substrate absence.

A new approach to the studies of submicron particle suspensions based on the effect of pressure in capillary zone electrophoresis.

A new approach based on the effect of pressure in CZE is suggested for acceleration of particle migration in electrophoretic runs resulting in reduction of the analysis time. It provides conditions for studying fast processes in suspensions. The effect of pressure on the migration of silica spheres with average diameters of 100, 150, and 390 nm was studied by CZE at an applied voltage of 25 kV. The particle hydrodynamic behavior was also investigated under the same capillary dimensions and BGE composition. The total particle mobility (excluding the average flow rate) was found to increase with increasing the pressure applied and particle size. The particle migration mechanism explaining the effect of pressure on particle velocity was shown to be almost the same as in wide-bore hydrodynamic chromatography. It is based on changing radial distribution of particle concentration along the capillary cross section depending on particle diffusivity. On the basis of this mechanism appearance of a zone of negatively charged particles in electropherograms ahead of the marker peak can be explained.

Interactive control of purpose built analytical instruments with Forth on microcontrollers - A tutorial.

The use of the computer language Forth for controlling experimental analytical instruments built in laboratories is described. Forth runs on a microcontroller and as it is an interpreted language the user can directly communicate with it by employing a terminal emulator program running on a personal computer. Thus the user can test attached hardware, such as pumps, valves, electronic pressure regulators, detectors and chemical sensors, directly from the keyboard. This overcomes the lack of interactivity, a significant shortcoming, of the computer languages C and C++, the default on such microcontroller platforms as the Arduinos, which have become very popular in recent years for laboratory applications. Common examples of purpose built experimental analytical laboratory instruments are sequential injection analysis systems, microfluidic devices, or automated sample extraction systems. Application examples from our laboratory are given, namely the regulation of mass-flow controllers for gases, the sequencing of an experimental capillary electrophoresis instrument and the acquisition of a signal from an alcohol sensor.

Organ Specific Proteomic Dissection of Selaginella bryopteris Undergoing Dehydration and Rehydration.

To explore molecular mechanisms underlying the physiological response of Selaginella bryopteris, a comprehensive proteome analysis was carried out in roots and fronds undergoing dehydration and rehydration. Plants were dehydrated for 7 days followed by 2 and 24 h of rehydration. In roots out of 59 identified spots, 58 protein spots were found to be up-regulated during dehydration stress. The identified proteins were related to signaling, stress and defense, protein and nucleotide metabolism, carbohydrate and energy metabolism, storage and epigenetic control. Most of these proteins remained up-regulated on first rehydration, suggesting their role in recovery phase also. Among the 90 identified proteins in fronds, about 49% proteins were up-regulated during dehydration stress. Large number of ROS scavenging proteins was enhanced on dehydration. Many other proteins involved in energy, protein turnover and nucleotide metabolism, epigenetic control were also highly upregulated. Many photosynthesis related proteins were upregulated during stress. This would have helped plant to recover rapidly on rehydration. This study provides a comprehensive picture of different cellular responses elucidated by the proteome changes during dehydration and rehydration in roots and fronds as expected from a well-choreographed response from a resurrection plant.

Transferrina como marcador de maturação pulmonar em cordeiros nascidos a termo ou prematuros / Transferrin as a marker of pulmonary maturation in lambs born at term or premature

O objetivo do estudo foi procurar proteínas de fase aguda que possam indicar sinais de maturação no neonato prematuro, por meio da quantificação sérica delas. Identificou-se a imunoglobulina A, a ceruloplasmina, a haptoglobina, a glicoproteína ácida, a transferrina, a albumina e as imunoglobulinas G de cadeias leve e pesada, pela comparação do perfil dos proteinogramas de cordeiros nascidos a termo com os prematuros submetidos a diferentes protocolos terapêuticos, a fim de estimular a atividade respiratória. Constituíram-se seis grupos: PN (n= 9): nascidos de parto normal; CN (n= 7): nascidos de cesariana em tempo normal de gestação; CP (n= 6): nascidos de cesariana prematura sem nenhum tipo de tratamento; DEX (n= 9): prematuros cujas mães receberam dexametasona pré-parto; SURF (n= 6): prematuros tratados com surfactante; e DEXSURF (n= 6): prematuros tratados com surfactante cujas mães receberam dexametasona pré-parto. As avaliações foram realizadas nos momentos imediatamente após o nascimento (M0), após 24 (M24) e após 48 horas (M48). As amostras foram processadas por meio de eletroforese em gel de poliacrilamida contendo dodecil sulfato de sódio (SDS-PAGE). A albumina, as imunoglobulinas e a proteína total dos cordeiros tiveram elevação após a ingestão de colostro. Maiores valores séricos de transferrina são referentes a maior período gestacional, podendo essa proteína ser utilizada como marcador de maturação neonatal.(AU)

The aim of this study was to search for acute phase proteins that could indicate signs of maturation in the premature neonate by quantifying them in serum. Immunoglobulin A, ceruloplasmin, haptoglobin, acid glycoprotein, tranferrin, albumin, light and heavy chain immunoglobulin G were quantified, comparing the profile of proteinograms from term to preterm lambs submitted to different protocols that stimulate respiratory activity. Six groups were used: PN (n= 9): born from normal birth; CN (n= 7): born from caesarean section at normal time of gestation; CP (n= 6): born from premature cesarean without any type of treatment; DEX (n= 9) preterm whose mothers received prepartum dexamethasone; SURF (n= 6) preterm treated with surfactant; DEXSURF (n= 6): preterm treated with surfactant whose mothers received prepartum dexamethasone. The evaluations were performed immediately after birth (M 0), after 24 and 48 hours (M 24 and M 48). Samples were processed with sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Albumin, immunoglobulins, and serum total protein of the lambs were elevated, after colostrum ingestion. Higher serum transferrin values refer to a longer gestational period, and this protein may be used as a marker of neonatal maturation.(AU)

COMPARATIVE STUDIES OF THE ZYMOGRAMS OF ESTERASE AND LACTIC DEHYDROGENASE ISOZYMES FROM TWO SPECIES OF AEDES / 第二军医大学学报

The zymograms of esterase(Est)and lactic dehydrogenase (LDH) isozymes fromtwo species of Aedes were studied by polyacrylamide gel electrophoresis.The zy-mograms were divided into two groups of bands, Est Ⅰ and Ⅱ, and LDH ⅠandⅡ, respectively.They exhibited some similar characteristics of sibling species,On the basis of differences of Est Ⅱ phenotypes, the two species could be distinguished from each other

Genomic analysis of Mycobacterium foruitum by pulsed-filed gel electrophoresis / 영남의대학술지

Epidemiological studies are important in both the prevention and treatment of mycobacterial infections. This study was initiated to establish the pulsed-field gel electrophoresis (PFGE) method, which are not yet extensively studied. The most apprpriate restriction endonucleases included Dral, AsnI, and XbaI. The optimal PFGE condition was different according to the enzymes used. Two stage PFGE was performed, in case of DraI first stage was performed with 10 seconds of initial pulse and 15 seconds of findA pulse, while the second stage was performed with 60 seconds of initial pulse and 70 seconds of final pu',se. The electrophoresis time for DraI-PFGE was 14 hours for each stage. Electrophoresis was performed for 22 hours, in case of XbaI, with 3 seconds of initial pulse and 12 seconds of final pulse. Electrophoresis was performed for 22 hours, in case of AsnI, with 5 seconds of initial pulse and 25 seconds of final pulse. In all cases the voltage of the electrophoresis was maintained constantly at 200 voltage. Standard mycobacterial strains, which included Mycobacterium bovis BCG, M. tuberculosis, and M. fortuitum, could not be differentiated by PFGE analysis. PFGE analysis was performed to differentiate 9 clinically isolated M. fortuitum strains using AsnI. All M. fortuitum strains showed different genotypes except 2 strains. Cluster analysis divided M. fortuitum strains into 2 large groups. PFGE analysis was performed to further differentiate M. fortuitum isolates using XbaI. The undifferentiated 2 M. fortuitum strains showed different PFGE patterns with Xba I. Cluster analysis of the XbaI-PFGE patterns showed more complex grouping than AsnI-PFGE patterns, which showed that XbaI-PFGE analysis was better than AsnI-PFGE in M. fortuitum genotyping. The top dissimilarity values of AsnI-PFGE and XbaI-PFGE were 0.74 and 0.75, respectively. This value was higher than that of arbitrarily primed polymerase chain reaction (AP-PCR) analysis and lower than that of restriction fragment length polymorphism (RFLP) analysis. This suggested that PFGE can be used as a supportive or alternative genotyping method to RFLP analysis.

Effect of removing stasis medicinal herbs on proliferation and proteomics of THP-1 cell line induced by endotoxin / 中国免疫学杂志

Objective:To observe the proteomics of blood-activating and stasis-removing(BASR) herbs in cellular model of endotoxin infection in vitro.Methods:Cellular model of endotoxin was induced by LPS,and a technical system by two-dimensional electrophoresis was widely used,proteins in different expression were analyzed by PDQust 7.1.1.Results:There were 176 protein spots in model group,in which 26 protein spots with different expression were sieved.Compared the protein profiles of TanshinoneⅡA group in model group,proteins were found as two-way-regulated.Among them,50% proteins were observed down-regulated and 38.5% proteins were seen up-regulated in TanshinoneⅡA group.11.5% more up-regulated.Conclusion:Cellular model of endotoxin induce was applied to sieving traditional Chinese medicines in laboratory;The Tanshinone ⅡA can inhibit the proliferation of THP-1 cell line in infection model,and can modify expression of some proteins.