Redox regulation of type-I inositol trisphosphate receptors in intact mammalian cells.

A sensitization of inositol 1,4,5-trisphosphate receptor (IP3R)-mediated Ca2+ release is associated with oxidative stress in multiple cell types. These effects are thought to be mediated by alterations in the redox state of critical thiols in the IP3R, but this has not been directly demonstrated in intact cells. Here, we utilized a combination of gel-shift assays with MPEG-maleimides and LC-MS/MS to monitor the redox state of recombinant IP3R1 expressed in HEK293 cells. We found that under basal conditions, ∼5 of the 60 cysteines are oxidized in IP3R1. Cell treatment with 50 μm thimerosal altered gel shifts, indicating oxidation of ∼20 cysteines. By contrast, the shifts induced by 0.5 mm H2O2 or other oxidants were much smaller. Monitoring of biotin-maleimide attachment to IP3R1 by LC-MS/MS with 71% coverage of the receptor sequence revealed modification of two cytosolic (Cys-292 and Cys-1415) and two intraluminal cysteines (Cys-2496 and Cys-2533) under basal conditions. The thimerosal treatment modified an additional eleven cysteines, but only three (Cys-206, Cys-767, and Cys-1459) were consistently oxidized in multiple experiments. H2O2 also oxidized Cys-206 and additionally oxidized two residues not modified by thimerosal (Cys-214 and Cys-1397). Potentiation of IP3R channel function by oxidants was measured with cysteine variants transfected into a HEK293 IP3R triple-knockout cell line, indicating that the functionally relevant redox-sensitive cysteines are predominantly clustered within the N-terminal suppressor domain of IP3R. To our knowledge, this study is the first that has used proteomic methods to assess the redox state of individual thiols in IP3R in intact cells. © 2018 Joseph et al

Growth of Pseudomonas chloritidismutans AW-1T on n-alkanes with chlorate as electron acceptor

Microbial (per)chlorate reduction is a unique process in which molecular oxygen is formed during the dismutation of chlorite. The oxygen thus formed may be used to degrade hydrocarbons by means of oxygenases under seemingly anoxic conditions. Up to now, no bacterium has been described that grows on aliphatic hydrocarbons with chlorate. Here, we report that Pseudomonas chloritidismutans AW-1T grows on n-alkanes (ranging from C7 until C12) with chlorate as electron acceptor. Strain AW-1T also grows on the intermediates of the presumed n-alkane degradation pathway. The specific growth rates on n-decane and chlorate and n-decane and oxygen were 0.5 ± 0.1 and 0.4 ± 0.02 day−1, respectively. The key enzymes chlorate reductase and chlorite dismutase were assayed and found to be present. The oxygen-dependent alkane oxidation was demonstrated in whole-cell suspensions. The strain degrades n-alkanes with oxygen and chlorate but not with nitrate, thus suggesting that the strain employs oxygenase-dependent pathways for the breakdown of n-alkanes

Osmopriming combined with Boron-Tolerant Bacteria (Bacillus sp. MN54) improved the productivity of Desi Chickpea under rainfed and irrigated conditions

Chickpeas are rich source of protein and predominantly grown in boron (B)-deficient sandy-loam soils in Pakistan. Boron-tolerant bacteria (BTB) could tolerate higher B levels in soil and increase B availability to the plants. Field trials were conducted under irrigated (district Layyah) and rainfed (district Chakwal) conditions to evaluate the interactive effects of pre-optimized B application methods and BTB (Bacillus sp. MN54) on the nodule’s population, grain quality, productivity, and grain-B concentration in desi chickpea during 2019–2020 and 2020–2021. Boron was applied as soil application (1 kg B ha−1), foliar application (0.025% B), osmopriming (0.001% B), and seed coating (1.5 g B kg−1 seed) with or without BTB inoculation. Untreated seeds receiving no B through any of the methods were regarded as control. The individual and interactive effects (up to three-way interaction of location × BTB inoculation × B application methods) of year, location, B application methods and BTB inoculation significantly altered the growth and yield-related traits of desi chickpea. The four-way interaction of year × location × BTB inoculation × B application methods was non-significant for all recorded growth and yield-related traits. Regarding individual effects, the higher values of growth and yield-related traits were noted for 2020–2021, rainfed location, BTB inoculation and B application through seed priming. Similarly, in two-way interactions 2020–2021 with rainfed location and BTB inoculation, rainfed location with BTB inoculation and osmopriming and osmopriming with BTB inoculation recorded higher values of the growth and yield-related traits. Osmopriming combined with BTB inoculation significantly improved dry matter accumulation and leaf area index in both locations. Boron application through all the methods significantly improved grain quality, yield grain B concentration. The highest grain and biological yields, and nodules’ population were recorded with osmopriming followed by soil application of B combined with BTB inoculation. The highest plant B concentration (75.05%) was recorded with foliar application of B followed by osmopriming (68.73%) combined with BTB inoculation. Moreover, the highest economic returns (USD 2068.5 ha−1) and benefit–cost ratio (3.7%) were recorded with osmopriming + BTB inoculation in 2020–2021 under rainfed conditions. Overall, B application through osmopriming and soil application combined with BTB inoculation could be used to increase productivity and profitability of desi chickpea, whereas foliar application is a better method to enhance grain and plant B concentration

Production of lignin peroxidase by Ganoderma leucidum using solid state fermentation

The main objectives of this study were to optimize the culture conditions for the production of lignin peroxidase by Ganoderma leucidum, economic utilization of waste corn cobs as inducers substrate by pollution free fermentation technology and to optimize the solid state fermentation (SSF) process for lignin peroxidase (LiP) production. Growth medium employed for the culture of G. leucidum for the production of LiP was supplied with fermentation conditions that were optimized before selection like incubation period, inoculum size, temperature, pH, substrate to water ratio, nitrogen source, yeast extract and cane molasses. Culture was harvested on the fourth day and diluted five times with distilled water and filtrate was stored in Eppendoff tubes for enzyme assay using 310 nm wave lengths in the spectrophotometer. Lignin peroxidase production was enhanced and maximum LiP activity (2807 U/ml) was found in the growth medium after 96 h, inoculum size 3 ml, pH 4.5 and temperature 35°C with substrate to water ratio of 20 ml/5 g, yeast extract 4%, (NH4)2SO4, 2% and cane molasses 3%. Results indicate the excellent scope of corn cobs as solid state substrate for the production of lignin peroxidase by G. leucidum.Key words: Lignin peroxidase (LiP), Ganoderma leucidum, fermentation, corn cobs, cane molasses

Soil-Applied boron combined with Boron-Tolerant Bacteria (Bacillus sp. MN54) improve root proliferation and nodulation, yield and agronomic grain biofortification of chickpea (Cicer arietinum L.)

Chickpea is widely cultivated on calcareous sandy soils in arid and semi-arid regions of Pakistan; however, widespread boron (B) deficiencies in these soils significantly decreases its productivity. Soil application of B could improve chickpea yield and grain-B concentration. However, optimizing suitable B level is necessary due to a narrow deficiency and toxicity range of B. Nonetheless, the co-application of B-tolerant bacteria (BTB) and synthetic B fertilizer could be helpful in obtaining higher chickpea yields and grain-B concentration. Therefore, this study optimized the level of soil applied B along with BTB, (i.e., Bacillus sp. MN54) to improve growth, yield and grain-B concentrations of chickpea. The B concentrations included in the study were 0.00 (control), 0.25, 0.50, 0.75 and 1.00 mg B kg−1 soil combined with or without Bacillus sp. MN54 inoculation. Soil application of B significantly improved root system, nodulation, yield and grain-B concentration, and Bacillus sp. MN54 inoculation further improved these traits. Moreover, B application at a lower dose (0.25 mg B kg−1 soil) with BTB inoculation recorded the highest improvements in root system (longer roots with more roots’ proliferation), growth, nodulation and grain yield. However, the highest grain-B concentration was recorded under a higher B level (0.75 mg B kg−1 soil) included in the study. Soil application of 0.25 mg B kg−1 with Bacillus sp. MN54 inoculation improved growth and yield-related traits, especially nodule population (81%), number of pods plant−1 (38%), number of grains plant−1 (65%) and grain yield (47%) compared with control treatment. However, the grain-B concentration was higher under the highest B level (1.00 mg kg−1 soil) with Bacillus sp. MN54 inoculation. In conclusion, soil application of 0.25 mg B kg−1 with Bacillus sp. MN54 inoculation is a pragmatic option to improve the root system, nodule population, seedling growth, yield and agronomic grain-B biofortification of chickpea

Breast imaging reports for malignant lesions: Are we maintaining recommended BI-RADS® lexicon standards

Background: The purpose of this study was to evaluate mammography reports for diagnosed breast cancer cases in major government and private centers in Karachi, Pakistan, with respect to concordance with the Breast Imaging Reports And Data System (BI-RADS®) lexicon.Methods: A prospective, descriptive, multicenter study was conducted in the radiology sections of the Aga Khan University Hospital, Pakistan Naval Station Shifa Hospital, Advanced Radiology Clinic, Karachi Institute of Radiotherapy and Nuclear Medicine, and Civil Hospital Karachi between May and October 2010 after approval from the ethical review committee of Aga Khan University. Mammograms reported as BI-RADS category 4 and 5 were included in the study. Mammograms reported as BI-RADS category 0, 1, 2 and 3 were excluded. Fifty reports were collected from each center. Data were collected about the clinical indication, breast density, location and description of the lesion, calcification, and comments on axillary lymph nodes. This description was compared with the BI-RADS lexicon.Results: The mean age of the patients was 50 ± 12 years. The clinical indication, breast parenchymal density, lesion location, and presence of calcification were better described by the private centers, while description of lymph node status was better stated by the government centers. This difference was statistically significant, except for lesion description. The description of masses by the two reporting groups was comparable.Conclusion: Mammographic reporting of malignant breast lesions in the private sector is more in line with the BI-RADS lexicon, as compared with government sector hospitals in Karachi, Pakistan. Lymph node documentation was better in government sector reports

Synthesis and Characterization of Silver-Coated Polymeric Scaffolds for Bone Tissue Engineering: Antibacterial and In Vitro Evaluation of Cytotoxicity and Biocompatibility

In bone tissue engineering, multifunctional composite materials are very challenging. Bone tissue engineering is an innovative technique to develop biocompatible scaffolds with suitable orthopedic applications with enhanced antibacterial and mechanical properties. This research introduces a polymeric nanocomposite scaffold based on arabinoxylan-co-acrylic acid, nano-hydroxyapatite (nHAp), nano-aluminum oxide (nAl₂O₃), and graphene oxide (GO) by free-radical polymerization for the development of porous scaffolds using the freeze-drying technique. These polymeric nanocomposite scaffolds were coated with silver (Ag) nanoparticles to improve antibacterial activities. Together, nHAp, nAl₂O₃, and GO enhance the multifunctional properties of materials, which regulate their physicochemical and biomechanical properties. Results revealed that the Ag-coated polymeric nanocomposite scaffolds had excellent antibacterial properties and better microstructural properties. Regulated morphological properties and maximal antibacterial inhibition zones were found in the porous scaffolds with the increasing amount of GO. Moreover, the nanosystem and the polymeric matrix have improved the compressive strength (18.89 MPa) and Young’s modulus (198.61 MPa) of scaffolds upon increasing the amount of GO. The biological activities of the scaffolds were investigated against the mouse preosteoblast cell lines (MC3T3-E1) and increasing the quantities of GO helps cell adherence and proliferation. Therefore, our findings showed that these silver-coated polymeric nanocomposite scaffolds have the potential for engineering bone tissue