Tumor Xenograft Response to Redox-Active Therapies Assessed by Magnetic Resonance Imaging Using a Thiol-Bearing DOTA Complex of Gadolinium

Gd-LC6-SH is a thiol-bearing DOTA complex of gadolinium designed to bind plasma albumin at the conserved Cys(34) site. The binding of Gd-LC6-SH shows sensitivity to the presence of competing thiols. We hypothesized that Gd-LC6-SH could provide magnetic resonance imaging (MRI) enhancement that is sensitive to tumor redox state and that the prolonged retention of albumin-bound Gd-LC6-SH in vivo can be exploited to identify a saturating dose above which the shortening of MRI longitudinal relaxation time (T(1)) of tissue is insensitive to the injected gadolinium dose. In the Mia-PaCa-2 pancreatic tumor xenograft model in SCID mice, both the small-molecule Gd-DTPA-BMA and the macromolecule Galbumin MRI contrast agents produced dose-dependent decreases in tumor T(1). By contrast, the decreases in tumor T(1) provided by Gd-LC6-SH at 0.05 and 0.1 mmol/kg were not significantly different at longer times after injection. SCID mice bearing Mia-PaCa-2 or NCI-N87 tumor xenografts were treated with either the glutathione synthesis inhibitor buthionine sulfoximine or the thiol-oxidizing anticancer drug Imexon, respectively. In both models, there was a significantly greater increase in tumor R(1) (=1/T(1)) 60 minutes after injection of Gd-LC6-SH in drug-treated animals relative to saline-treated controls. In addition, Mercury Orange staining for nonprotein sulfhydryls was significantly decreased by drug treatment relative to controls in both tumor models. In summary, these studies show that thiol-bearing complexes of gadolinium such as Gd-LC6-SH can serve as redox-sensitive MRI contrast agents for detecting differences in tumor redox status and can be used to evaluate the effects of redox-active drugs.     

Genetic Diversity of Mycobacterium tuberculosis Isolates from Assam,India: Dominance of Beijing Family and Discovery of Two New Clades Related to CAS1_Delhi and EAI Family Based on Spoligotyping and MIRU-VNTR Typing

Tuberculosis (TB) is one of the major public health concerns in Assam, a remote state located in the northeastern (NE) region of India. The present study was undertaken to explore the circulating genotypes of Mycobacterium tuberculosis complex (MTBC) in this region. A total of 189 MTBC strains were collected from smear positive pulmonary tuberculosis cases from different designated microscopy centres (DMC) from various localities of Assam. All MTBC isolates were cultured on Lowenstein-Jensen (LJ) media and subsequently genotyped using spoligotyping and 24-loci mycobacterial interspersed repetitive units-variable number of tandem repeats (MIRU-VNTR) typing. Spoligotyping of MTBC isolates revealed 89 distinct spoligo patterns. The most dominant MTBC strain belonged to Beijing lineage and was represented by 35.45% (n = 67) of total isolates, followed by MTBC strains belonging to Central Asian-Delhi (CAS/Delhi) lineage and East African Indian (EAI5) lineage. In addition, in the present study 43 unknown spoligo patterns were detected. The discriminatory power of spoligotyping was found to be 0.8637 based on Hunter Gaston Discriminatory Index (HGDI). On the other hand, 24-loci MIRU-VNTR typing revealed that out of total 189 MTBC isolates from Assam 185 (97.9%) isolates had unique MIRU-VNTR profiles and 4 isolates grouped into 2 clusters. Phylogenetic analysis of 67 Beijing isolates based on 24-loci MIRU-VNTR typing revealed that Beijing isolates from Assam represent two major groups, each comprising of several subgroups. Neighbour-Joining (NJ) phylogenetic tree analysis based on combined spoligotyping and 24-loci MIRU-VNTR data of 78 Non-Beijing isolates was carried out for strain lineage identification as implemented by MIRU-VNTRplus database. The important lineages of MTBC identified were CAS/CAS1_Delhi (41.02%, n = 78) and East-African-Indian (EAI, 33.33%). Interestingly, phylogenetic analysis of orphan (23.28%) MTBC spoligotypes revealed that majority of these orphan isolates from Assam represent two new sub-clades Assam/EAI and Assam/CAS. The prevalence of multidrug resistance (MDR) in Beijing and Non-Beijing strains was found to be 10.44% and 9.01% respectively. In conclusion, the present study has shown the predominance of Beijing isolates in Assam which is a matter of great concern because Beijing strains are considered to be ecologically more fit enabling wider dissemination of M. tuberculosis. Other interesting finding of the present study is the discovery of two new clades of MTBC isolates circulating in Assam. More elaborate longitudinal studies are required to be undertaken in this region to understand the transmission dynamics of MTBC.     

Neglecting Rice Milling Yield and Quality Underestimates Economic Losses from High-Temperature Stress

Future increases in global surface temperature threaten those worldwide who depend on rice production for their livelihoods and food security. Past analyses of high-temperature stress on rice production have focused on paddy yield and have failed to account for the detrimental impact of high temperatures on milling quality outcomes, which ultimately determine edible (marketable) rice yield and market value. Using genotype specific rice yield and milling quality data on six common rice varieties from Arkansas, USA, combined with on-site, half-hourly and daily temperature observations, we show a nonlinear effect of high-temperature stress exposure on yield and milling quality. A 1°C increase in average growing season temperature reduces paddy yield by 6.2%, total milled rice yield by 7.1% to 8.0%, head rice yield by 9.0% to 13.8%, and total milling revenue by 8.1% to 11.0%, across genotypes. Our results indicate that failure to account for changes in milling quality leads to understatement of the impacts of high temperatures on rice production outcomes. These dramatic losses result from reduced paddy yield and increased percentages of chalky and broken kernels, which together decrease the quantity and market value of milled rice. Recently published estimates show paddy yield reductions of up to 10% across the major rice-producing regions of South and Southeast Asia due to rising temperatures. The results of our study suggest that the often-cited 10% figure underestimates the economic implications of climate change for rice producers, thus potentially threatening future food security for global rice producers and consumers.     

Modulating HIV-1 envelope glycoprotein conformation to decrease the HIV-1 reservoir

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Hypersensitive reaction and induced resistance in rice against the Asian rice gall midge Orseolia oryzae

Rice seedlings of the resistant variety Phalguna showed premature tillering, browning of central leaf, and tissue necrosis at the apical meristem following artificial infestation with avirulent biotype 1 of the Asian rice gall midge, Orseolia oryzae (Wood-Mason) (Diptera: Cecidomyiidae). Tissue necrosis representing a typical hypersensitive reaction (HR), accompanied by maggot mortality, was observed within 4 days after infestation. However, reinfestation of secondary tillers subsequent to HR in primary tiller, did not lead to HR in secondary tillers though maggot mortality was seen. Artificial infestation with the weed gall midge O. fluvialis did not result in HR either in gall midge susceptible TN 1 or resistant Phalguna rice varieties. Resistance in Phalguna against the virulent biotype 4 could be induced by either prior, simultaneous, or subsequent infestation with the avirulent biotype 1. The duration of effectiveness of such induced resistance varied with the sequence and time lag between infestations.     

Exposure to ionizing radiation during pregnancy: perception of teratogenic risk and outcome

We quantified the perception of teratogenic risk in women attending the Motherisk program for counseling about diagnostic radiation in pregnancy (n = 50) and compared it with a control group of women exposed to nonteratogenic drugs and chemicals (n = 48). Before receiving known information about the specific exposure, women exposed to radiation assigned themselves a significantly higher teratogenic risk compared with the control group (25.5 +/- 4.3% versus 15.7 +/- 3.0% for major malformations, P less than 0.01). The post-consultation perception of teratogenic risk did not differ between the two groups. Special consideration and attention should be given when counseling pregnant women exposed to low-dose ionizing radiation, as their misperception of teratogenic risk may lead them to unnecessary termination of their pregnancy.     

Soybean lipoxygenase inhibition by nonsteroidal antiinflammatory drugs

Eighteen known nonsteroidal antiinflammatory drugs (NSAID) were tested for their action against soybean lipoxygenase (E.C.1.13.11.12) using linoleic acid as substrate. It was found that the best inhibitors of lipoxygenase were naproxen, BW 755C, indomethacin and isoxicam. Drugs with intermediate potency were meclofenamic acid, phenyl-butazone and benoxaprofen. Other drugs such as ibuprofen and zomepirac were only weakly active in the test.     

The relationship between cell size and cell division in the yeast Saccharomyces cerevisiae.

Cell numbers in synchronous cultures of yeast cultured at fast growth rates increase from N to 2N after the first division and from 2N to 4N after the second division. At these fast growth rates, there are equal numbers of parents and daughters. In contrast, at slow growth rates the cell number increases from N to 2N after one division and from 2N to 3N rather than 4N after the second division. Moreover, the percentage of daughters increases with decreasing growth rate. Thus, slowly growing cultures actually consist of two sub-populations having different cell cycle transit times. These observations are predicted if a yeast cell requires a critical size before a particular cell cycle event can be completed and that after completion of this event cell division occurs following a period of time independent of growth rate.     

A method for site-directed transplacement of in vitro altered DNA sequences in Rhizobium

Summary A method that directs transplacement of in vitro altered DNA sequences to substitute the corresponding wild-type DNA sequences at the original location in the Rhizobium genome is described. The NPTII gene of transposon Tn5 which confers resistance to several antibiotics in a wide variety of organisms is used as a selectable marker on the vector. To generate DNA fragment substitution, it is not essential to place a selectable genetic marker directly linked to the in vitro altered region of the DNA fragment. The method utilizes the vector pSUP201 that replicates in E. coli but neither in Rhizobium nor in many other host systems. DNA transplacement occurs by the integration of the in vitro altered DNA fragment at its homologous site in the chromosome along with the vector. A duplicated target sequence is generated, which is then followed by a homologous recombination event between the duplicated DNA sequences leading to the excision of the vector carrying the selectable marker together with one of the duplicated copies of DNA. The excised DNA fragment is subsequently lost due to the inability of the vector pSUP201 to replicate in Rhizobium. Using this method, we have transplaced a nif DNA fragment containing a large deletion of approximately 9 Kb into its homologous site in the genome of cowpea Rhizobium IRc78. The IRc78 strain carrying the deletion forms nodules on host-plants but is unable to reduce atmospheric nitrogen. The method allows precise alterations of the genomic DNA in an organism that is genetically not well characterized, provided a vector carrying a selectable marker can be transferred.Dedicated to Professor Georg Melchers to celebrate his 50-year association with the journal