Oxygen‐dependent delayed fluorescence of protoporphyrin IX measured in the stomach and duodenum during upper gastrointestinal endoscopy

Protoporphyrin IX‐triplet state lifetime technique (PpIX‐TSLT) is a method used to measure oxygen (PO₂) in human cells. The aim of this study was to assess the technical feasibility and safety of measuring oxygen‐dependent delayed fluorescence of 5‐aminolevulinic acid (ALA)‐induced PpIX during upper gastrointestinal (GI) endoscopy. Endoscopic delayed fluorescence measurements were performed 4 hours after oral administration of ALA in healthy volunteers. The ALA dose administered was 0, 1, 5 or 20 mg/kg. Measurements were performed at three mucosal spots in the gastric antrum, duodenal bulb and descending duodenum with the catheter above the mucosa and while applying pressure to induce local ischemia and monitor mitochondrial respiration. During two endoscopies, measurements were performed both before and after intravenous administration of butylscopolamine. Delayed fluorescence measurements were successfully performed during all 10 upper GI endoscopies. ALA dose of 5 mg/kg showed adequate signal‐to‐noise ratio (SNR) values >20 without side effects. All pressure measurements showed significant prolongation of delayed fluorescence lifetime compared to measurements performed without pressure (P < .001). Measurements before and after administration of butylscopolamine did not differ significantly in the duodenal bulb and descending duodenum. Measurements of oxygen‐dependent delayed fluorescence of ALA‐induced PpIX in the GI tract during upper GI endoscopy are technically feasible and safe.     

Adaptations of the antioxidant system in erythrocytes of trained adult rats: impact of intermittent hypobaric-hypoxia at two altitudes

We have investigated the effects of daily exposure to intermittent hypobaric-hypoxia to two simulated altitudes (5700 m and 6300 m) in adult male rats that had been regularly swim trained in normoxia at sea level prior to exposures. Superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSH-Px) along with the oxidative stress (OS) indices, malondialdehyde (MDA) and protein carbonyl content were measured in erythrocytes and their membranes. Hemoglobin increased in the trained animals exposed to 5700 m and in untrained rats exposed to 6300 m. Osmotic fragility in terms of hemolysis increased in altitude exposed animals. SOD increased in those exposed to 6300 m, while CAT increased in trained rats exposed to 5700 m and to 6300 m unlike in untrained rats where CAT increased only at 6300 m. GSH-Px showed varying degrees of elevation in all animals exposed to both altitudes. Erythrocyte membranes showed significant elevations in malondialdehyde (MDA) at 6300 m, while elevated protein carbonyls were noticeable at both altitudes in whole cells and membranes. These results suggest a positively associated elevation in protein oxidation with altitude in trained rats. At 5700 m, animals were less stressed, unlike at 6300 m, as seen from the magnitude of elevations in the OS indices and from the responses of the antioxidant enzymes.     

Translation of tyrosine aminotransferase mRNA in a modified reticulocyte system.

Tyrosine aminotransferase messenger RNA has been translated in a cell-free system derived from rabbit reticulocytes. Cytoplasmic poly(A)-containing RNA from rat liver was used as the source of the messenger RNA. The newly synthesized subunits of the enzyme were isolated by immunoprecipitation and identified and quantitated using polyacrylamide gel electrophoresis. These procedures were used to demonstrate that glucocorticoid induction is associated with increased cytoplasmic levels of functional tyrosine aminotransferase messenger RNA.     

Ultrasensitive green spectrofluorimetric approach for quantification of Hg(II) in environmental samples (water and fish samples) using cysteine@MnO2 dots

Mercury (Hg²⁺) is a natural element present in foods such as fish, water and soil. Exposure to mercury leads to severe toxic effects on the nervous, digestive, and immune systems. Here, a novel, green, and environmentally friendly fluorescent probe decorated with cysteine/MnO₂ quantum dots (Cys@MnO₂ QDs) was synthesized. This synthesis was carried out using a simple ultrasound technique with the aid of cysteine for fabricating Cys@MnO₂ QDs to estimate Hg levels in fish and water samples. In this morphological study, Cys@MnO₂ QDs were fully characterized using high-resolution transmission electron microscopy, zeta potential analysis, fluorescence, ultraviolet–visible and infrared spectroscopy. The fluorescence of the synthesized Cys@MnO₂ QDs was significantly quenched by gradually increasing the Hg(II) concentration. The quenching mechanism based on the Hg–S bonds strengthened the utility of the Cys@MnO₂ QDs as a novel luminescent nanoprobe. The estimation of Hg was linear in the concentration range 0.7–100.0 ng mL⁻¹ with a limit of quantitation equal to 0.30 ng mL⁻¹. The Cys@MnO₂ QDs are fluorescent probes with various benefits such as speed, ease of use, cost- effective, and being environmentally friendly; they are easily applied in food manufacturing and for public health improvement.     

Long-term continuous monitoring of dissolved oxygen in cell culture medium for perfused bioreactors using optical oxygen sensors

For long-term growth of mammalian cells in perfused bioreactors, it is essential to monitor the concentration of dissolved oxygen (DO) present in the culture medium to ascertain the health of the cells. An optical oxygen sensor based on dynamic fluorescent quenching was developed for long-term continuous measurement of DO for NASA-designed rotating perfused bioreactors. Tris(4,7-diphenyl-1,10-phenanthroline) ruthenium(II) chloride is employed as the fluorescent dye indicator. A pulsed, blue LED was chosen as the excitation light source. The sensor can be sterilized using an autoclave. The sensors were tested in a perfused rotating bioreactor supporting a BHK-21 (baby hamster kidney) cell culture over one 28-day, one 43-day, and one 180-day cell runs. The sensors were initially calibrated in sterile phosphate-buffered saline (PBS) against a blood-gas analyzer (BGA), and then used continuously during the entire cell culture without recalibration. In the 180-day cell run, two oxygen sensors were employed; one interfaced at the outlet of the bioreactor and the other at the inlet of the bioreactor. The DO concentrations determined by both sensors were compared with those sampled and measured regularly with the BGA reference. The sensor outputs were found to correlate well with the BGA data throughout the experiment using a single calibration, where the DO of the culture medium varied between 25 and 60 mm Hg at the bioreactor outlet and 80-116 mm Hg at the bioreactor inlet. During all 180 days of culture, the precision and the bias were +/-5.1 mm Hg and -3.8 mm Hg at the bioreactor outlet, and +/- 19 mm Hg and -18 mm Hg at inlet. The sensor dynamic range is between 0 and 200 mm Hg and the response time is less than 1 minute. The resolution of the sensor is 0.1 mm Hg at 50 mm Hg, and 0.25 mm Hg at 130 mm Hg.     

生境不完全重叠的两种鲤科鱼类耐低氧及运动能力比较

以乌江流域亲缘关系近,但分布并不完全重叠的马口鱼(Opsariichthys bidens)和宽鳍鱲(Zacco platypus)作为实验对象,分别考察这两种实验鱼的低氧耐受及游泳运动能力。将野外采回的实验鱼置于(25±1)℃条件下,分别测定两种鱼的临界氧分压(Pcrit)、水面呼吸(ASR)、失去平衡点(LOE)以及在不同溶氧水平(8.0、4.0和2.0 mg/L)下的临界游泳速度(Ucrit)和活跃耗氧率(MO2active)。研究发现:马口鱼的Pcrit(2.44±0.20)mg/L显著高于宽鳍鱲(1.86±0.10)mg/L(P=0.031)。但马口鱼的50%ASR(1.23±0.16)mg/L显著低于宽鳍鱲(1.97±0.11)mg/L(P=0.023);马口鱼的50%LOE(0.84±0.01)mg/L同样显著低于宽鳍鱲(0.97±0.02)mg/L(P=0.004)。宽鳍鱲在8.0和4.0 mg/L下的游泳能力显著高于马口鱼,然而马口鱼和宽鳍鱲的Ucrit均随测定溶氧水平的下降而显著降低(P0.01);宽鳍鱲和马口鱼的运动耗氧率均随水流速度的增加而呈现指数增加,但随测定溶氧水平的降低运动耗氧曲线变得相对平缓,尤其是在溶氧为4.0 mg/L时马口鱼的运动耗氧曲线与宽鳍鱲相比越发平缓;两种实验鱼的MO2active随溶氧水平下降的变化趋势与Ucrit相似(P0.001)。结果表明:两种野外生存的实验鱼不仅在低氧耐受能力方面存在显著差异,而且两者的游泳运动能力也有不同表现,这很有可能与其遗传特征、生存环境及生态习性相关。     

Evidence for involvement of the electron transport system at a late step of anaerobic microbial heme synthesis

The penultimate step in heme biosynthesis, the oxidation of protoporphyrinogen to protoporphyrin, can be anaerobically coupled to the reduction of fumarate in extracts of anaerobically-grown Escherichia coli. This coupling is approximately 90% inhibited by 2-heptyl-4-hydroxy quinoline-N-oxide (HQNO), a known inhibitor of the electron transport chain. This observation suggests that the mechanism of the anaerobic oxidation of protoporphyrinogen in E. coli involves a coupling into the anaerobic electron transport system. In contrast, the aerobic oxidation of protoporphyrinogen, which occurs in mammalian and yeast mitochondria, is known to be linked directly to oxygen without the mediation of an electron transport system.     

利用微载体悬浮培养人脐带间充质干细胞

随着干细胞研究的不断深入,干细胞功能分化研究和临床应用转化的需求日益提升。人脐带间充质干细胞(human umbilical cord mesenchymal stem cells, hUCMSCs)来源广泛,不仅自我更新能力强、能够分化成多种类型的成体细胞,而且其自身具有免疫调节能力,不易引发免疫排斥反应,在干细胞功能分化研究和临床应用中具有巨大应用前景和应用潜力。目前,传统的细胞培养方式培养效率低、细胞活性较差,不能满足日益增长的研究和应用需求。本研究利用微载体结合旋转瓶的悬浮培养方法,通过优化细胞接种量及转速等影响因素,快速获得大量高质量的人脐带间充质干细胞。经悬浮培养总细胞量可高达到7×10 ⁸个细胞/L,而且细胞活性较高,MSC 特异性标记物表达良好,在恢复平面培养后仍能维持MSC的正常细胞形态和增殖能力。高效脐带间充质干细胞悬浮培养体系的初步建立,为未来的干细胞功能分化研究和临床应用奠定了基础。     

典型林区水分氢氧稳定同位素在土壤-植物-大气连续体中的分布特征

土壤-植物-大气连续体(SPAC)中水循环是水文学和生态学研究的重要内容,氢氧稳定同位素在不同水体中组成特征的差异可以指示水分循环过程.本研究通过分析成都平原区亚热带常绿阔叶林中降水、土壤水、植物水的同位素组成,探讨SPAC系统中水分的氢氧稳定同位素演化特征,揭示区域水循环不同界面过程.结果 表明:研究区雨季大气降水线...     

Chromosomal localization of HTLV-1 viral integration sites using in situ hybridization: detection of a novel IL2R fragment

Summary The presence of human T-cell leukemia virus (HTLV-1) in patients with adult T-cell leukemia (ATL) was investigated by Southern blotting and in situ hybridization. In all seven patients, HTLV-1 provirus was detected. A large and variable number of labeled restriction fragments were observed, indicating multiple integrations. Two of the patients analyzed by in situ hybridization had two, while the third patient had three, sites of viral integration on six different chromosomes, suggesting random integration. A single site of integration was shared by two patients, which was on chromosome 10 at bands p11p15. One of these sites was on an apparently normal chromosome 10 and the other was on a derivative chromosome 10,t(10;14)(p12;g32). The interleukin 2 receptor (IL2R) has previously been localized to this region (10p14p15). The -chain of the IL2R is continuously expressed on affected T-cells in this disease. Southern blotting with pIL2R showed the presence of a novel 3.5 kb fragment in five out of the seven patients. This novel fragment has not been previously reported. No direct correlation was found between the novel 3.5 kb fragment, present in patients both cytogenetically normal and abnormal, and viral integration in the 10p11p15 region in two patients. Therefore, it is suggested that the presence of the 3.5 kb fragment and the numerous chromosomal breaks associated with this disease may not be direct results of viral integration.     

Evaluation of instream habitat enhancement options using fish habitat simulations: case-studies in the river Pas (Spain)

In the northern coast of Spain there are rivers with Atlantic salmon populations. In the upper reaches of one of these streams, river Pas, the effectiveness of habitat enhancement measures was evaluated, under different instream flow conditions. By means of the Instream Flow Incremental Methodology and using a two dimensional hydraulic model (River2D, Steffler P (2000) Software River2D. Two Dimensional Depth Averaged Finite Element Hydrodynamic Model. University of Alberta, Canada), the potential value of stream habitat for different salmon development stages requirements was measured by Weighted Useable Area (WUA). This habitat evaluation was carried out for the unmodified stream reach, which represent the control or natural conditions. Habitat improvement measures (alternate deflectors and low dams) were simulated in the original riverbed topography. Over this modified base, habitat was estimated running River2D again. By comparing the salmon habitat evaluations in the control conditions with those obtained under those improvement conditions we have been able to assess the effectiveness of each one, and the instream flow environment at which maximum improvement is reached. The maximum habitat improvement was obtained around 10 m³/s for the adult salmon, and for the fry and parr it was around 6 m³/s. However, the habitat simulation results show that with both improvement measures, under a natural flow regime the mean annual habitat increases around 1% of the WUA in relation to the control conditions, which is not a significant improvement. A similar small WUA increase was obtained when changing the bed topography, considering geomorphological adjustments due to the new erosion and sedimentation areas caused by the presence of these structures. Therefore, these types of habitat improvement measures are not recommended in these stream reaches.     

Diversity and dynamics of bacterial species in a biofilm at the end of the Seoul water distribution system

To investigate changes in the bacterial species and hygienic safety of the biofilm at the end of the drinking water distribution system in Seoul (Korea), denaturing gradient gel electrophoresis (DGGE) and DNA sequencing were used to analyse the bacterial population in the biofilm of a semi-pilot galvanized iron pipe model. The presence of sequences from aerobic Sphingomonas sp., anaerobic Rhodobacter sp., and unculturable bacteria indicated that these organisms coexisted after 1 day of model operation, demonstrating the ease of biofilm formation on galvanized iron pipes in the end region of the water distribution system studied. Sequences similar to those of unculturable bacteria, E. coli, and anaerobic bacteria were detected during the course of succession on the biofilm. More complicated band patterns were observed after 70 days of operation. PCR-DGGE illustrated changes in the biofilm during succession as well as the possibilities of anaerobic conditions and faecal contamination of the drinking water system. PCR-DGGE and culture-dependent fatty acid methyl ester (FAME) analysis showed different patterns for the same samples (Lee & Kim 2003); however, PCR-DGGE showed less diversity than did FAME analysis. This study compared the culture-dependent FAME and culture-independent PCR-DGGE methods directly, and their use in promoting the hygienic safety of drinking water.     

Simulating the effects of localized red:far-red ratio on tillering in spring wheat (Triticum aestivum) using a three-dimensional virtual plant model

The outgrowth of tiller buds in Poaceae is influenced by the ratio of red to far-red light irradiance (R:FR). At each point in the plant canopy, R:FR is affected by light scattered by surrounding plant tissues. This paper presents a three-dimensional virtual plant modelling approach to simulate local effects of R:FR on tillering in spring wheat (Triticum aestivum). R:FR dependence of bud outgrowth was implemented in a wheat model, using three hypothetical responses of bud extension to R:FR (unit step, curvilinear and linear response). Bud break occurred when a threshold bud length was reached. Simulations were performed for three plant population densities. In accordance with experimental observations, fewer tillers per plant were simulated for higher plant population densities. The linear and curvilinear responses caused a delay in the increase in tiller number compared with experimental data. The unit step response approached experimental results best. It is suggested that a model based on relatively simple relations can be used to simulate degree of tillering. This study has shown that the virtual plant approach is a promising tool with which to address crop morphological and ecological research questions where the determining factors act at the level of the individual plant organ.     

Effects of Korean Red Ginseng extract on hepatic lipid accumulation in HepG2 cells

In this study, we investigated the effects of Korean red ginseng water extract (KRGE) on hepatic lipid accumulation in HepG2 cells. KRGE decreased hepatic triglyceride and cholesterol levels. Further, KRGE suppressed expression of fatty acid synthase (FAS) and 3-hydroxy-3-methyl glutaryl coenzyme A (HMG-CoA) reductase. These results suggest that KRGE may reduce hepatic lipid accumulation by inhibition of FAS and HMG-CoA reductase expression in HepG2 cells.     

High‐efficient and precise base editing of C•G to T•A in the allotetraploid cotton (Gossypium hirsutum) genome using a modified CRISPR/Cas9 system

The base‐editing technique using CRISPR/nCas9 (Cas9 nickase) or dCas9 (deactivated Cas9) fused with cytidine deaminase is a powerful tool to create point mutations. In this study, a novel G. hirsutum‐Base Editor 3 (GhBE3) base‐editing system has been developed to create single‐base mutations in the allotetraploid genome of cotton (Gossypium hirsutum). A cytidine deaminase sequence (APOBEC) fused with nCas9 and uracil glycosylase inhibitor (UGI) was inserted into our CRISPR/Cas9 plasmid (pRGEB32‐GhU6.7). Three target sites were chosen for two target genes, GhCLA and GhPEBP, to test the efficiency and accuracy of GhBE3. The editing efficiency ranged from 26.67 to 57.78% at the three target sites. Targeted deep sequencing revealed that the C→T substitution efficiency within an ‘editing window’, approximately six‐nucleotide windows of ?17 to ?12 bp from the PAM sequence, was up to 18.63% of the total sequences. The 27 most likely off‐target sites predicted by CRISPR‐P and Cas‐OFFinder tools were analysed by targeted deep sequencing, and it was found that rare C→T substitutions (average < 0.1%) were detected in the editing windows of these sites. Furthermore, whole‐genome sequencing analyses on two GhCLA‐edited and one wild‐type plants with about 100× depth showed that no bona fide off‐target mutations were detectable from 1500 predicted potential off‐target sites across the genome. In addition, the edited bases were inherited to T1 progeny. These results demonstrate that GhBE3 has high specificity and accuracy for the generation of targeted point mutations in allotetraploid cotton.     

The relationship between carbon dioxide fixation and chlorophyll a fluorescence during induction of photosynthesis in maize leaves at different temperatures and carbon dioxide concentrations

The rate of CO₂ fixation (F_c) and 680 nm chlorophyll fluorescence emission (F680) were measured simultaneously during induction of photosynthesis in Zea mays L. leaves under varying experimental conditions in order to assess the validity of fluorescence as an indicator of in vivo photosynthetic carbon assimilation. Z. mays leaves showed typical Kautsky fluorescence induction curves consisting of a fast rise in emission (O to P) followed by a slow quenching via a major transient (S-M) to a steady-state (T). After an initial lag, net CO₂ assimilation commenced at a point corresponding to the onset of the S-M transient on the F680 induction curve. Subsequently, F_c and F680 always arrived at a steady-state simultaneously. Decreasing the dark-adaption period increased the rate of induction of both parameters. Alteration of leaf temperature produced anti-parallel changes in induction characteristics of F_c and F680. Reducing the CO₂ level to below that required for saturation of photosynthesis also produced anti-parallel changes during induction, however, at CO₂ concentrations tenfold greater than the atmospheric level the rate of F680 quenching from P to T was appreciably reduced without a similar change in the induction of F_c. Removal of CO₂ at steady-state produced only a small increase in F680 and a correspondingly small decrease in F680 occurred when CO₂ was re-introduced. The complex relationship between chlorophyll fluorescence and carbon assimilation in vivo is discussed and the applicability of fluorescence as an indicator of carbon assimilation is considered.Abbreviations F_c rate of CO₂ fixation - F680 fluorescence emission at 680 nm     

Is a decreased water potential after withholding oxygen to roots the cause of the decline of leaf-elongation rates in Zea mays L. and Phaseolus vulgaris L.?

Leaf-elongation rates of Zea mays L. and Phaseolus vulgaris L. were measured in plants grown for 4 d in nutrient solution bubbled with N₂ and in soil-grown waterlogged Phaseolus plants. Leaf water potential in both species was lower 3–4h after replacing aeration by N₂-bubbling. In Zea, the water potential after 24 h or more was the same in control plants and plants with N₂ treatment. In Phaseolus, the water potential of inundated plants and plants with N₂ treatment was always lower than those of control plants. The leaf-elongation rate of both species was always lower in plants treated with N₂, especially during light periods. In Zea, the elongation rate was lowest in the first 24 h, whilst in Phaseolus it was lowest on the last (fourth) day of treatment. There was no difference between N₂ treatment and inundation experiments. It is concluded that during the first hours of treatment the leaf-elongation rate was reduced as a consequence of the lower water potential. Thereafter, however, elongation rates were lower than could be expected on the basis of the plant's water relations.Abbreviations LER leaf elongation rate - PEG-200 polyethylene-glycol 200 - RWC relative water content     

Assessment of grain quality in terms of functional group response to elevated [CO2], water,and nitrogen using a meta‐analysis: Grain protein,zinc, and iron under future climate

The increasing [CO₂] in the atmosphere increases crop productivity. However, grain quality of cereals and pulses are substantially decreased and consequently compromise human health. Meta‐analysis techniques were employed to investigate the effect of elevated [CO₂] (e[CO₂]) on protein, zinc (Zn), and iron (Fe) concentrations of major food crops (542 experimental observations from 135 studies) including wheat, rice, soybean, field peas, and corn considering different levels of water and nitrogen (N). Each crop, except soybean, had decreased protein, Zn, and Fe concentrations when grown at e[CO₂] concentration (≥550 μmol/mol) compared to ambient [CO₂] (a[CO₂]) concentration (≤380 μmol/mol). Grain protein, Zn, and Fe concentrations were reduced under e[CO₂]; however, the responses of protein, Zn, and Fe concentrations to e[CO₂] were modified by water stress and N. There was an increase in Fe concentration in soybean under medium N and wet conditions but nonsignificant. The reductions in protein concentrations for wheat and rice were ~5%–10%, and the reductions in Zn and Fe concentrations were ~3%–12%. For soybean, there was a small and nonsignificant increase of 0.37% in its protein concentration under medium N and dry water, while Zn and Fe concentrations were reduced by ~2%–5%. The protein concentration of field peas decreased by 1.7%, and the reductions in Zn and Fe concentrations were ~4%–10%. The reductions in protein, Zn, and Fe concentrations of corn were ~5%–10%. Bias in the dataset was assessed using a regression test and rank correlation. The analysis indicated that there are medium levels of bias within published meta‐analysis studies of crops responses to free‐air [CO₂] enrichment (FACE). However, the integration of the influence of reporting bias did not affect the significance or the direction of the [CO₂] effects.     

Analysis of fexofenadine in pharmaceutical formulations using tris(1,10‐phenanthroline)–ruthenium(II) peroxydisulphate chemiluminescence system in a multichip device

A simple, rapid and sensitive method has been developed for the analysis of fexofenadine (FEX) in pharmaceutical formulations, using a tris(1,10‐phenanthroline)–ruthenium(II) [Ru(phen)₃²⁺] peroxydisulphate chemiluminescence (CL) system in a multichip device. Various parameters that influence the CL signal intensity were optimized. These included pH, flow rates and concentration of reagents used. Under optimum conditions, a linear calibration curve in the range 0.05–5.0 µg/mL was obtained. The detection limit was found to be 0.001 µg/mL. The procedure was applied to the analysis of FEX in pharmaceutical products and was found to be free from interference from concomitants usually present in these preparations. Copyright © 2011 John Wiley & Sons, Ltd.