Stable overexpression of DNA fragmentation factor in T-47D cells: sensitization of breast cancer cells to apoptosis in response to acetazolamide and sulfabenzamide

Alterations in expression of the DFF40 gene have been reported in some cancers. This study is an in vitro study of the therapeutic effects of gene transfer that lead to elevation in DFF40 expression within T-47D cells in the presence of sulfonamide drugs. In this study, we have constructed a eukaryotic expression vector for DFF40 and transfected it into T-47D cancer cells. We used real time RT-PCR to detect the expression of DFF40 and the MTT assay to determine effects of the sulfonamide drugs acetazolamide, sulfabenzamide, sulfathiazole and sulfacetamide on cell viability in the presence of increased and normal DFF40 levels. Cell cycle distribution was assessed by propidium iodide (PI) staining and the rates of apoptosis by annexin V/PI staining. The DNA laddering analysis was employed to evaluate apoptosis. We observed that overexpression of DFF40 was only effective in decreasing viability in cells incubated with acetazolamide and sulfabenzamide. There was enhanced apoptosis in these groups, particularly with acetazolamide. The cell cycle distribution analysis showed that in the presence of sulfonamide drugs there were no substantial changes in empty-vector or DFF40-transfected cells, except for those cells treated with sulfabenzamide or sulfathiazole. There was no DNA laddering in cells that expressed the empty vector when incubated with sulfonamide drugs. In contrast, we observed DNA laddering in cells that expressed DFF40 in the presence of acetazolamide. Our results have demonstrated that combinatorial use of some sulfonamides such as acetazolamide along with increased expression of DFF40 can potently kill tumor cells via apoptosis and may be beneficial for treatment of some chemoresistant cancers.     

Relating Diseases by Integrating Gene Associations and Information Flow through Protein Interaction Network

Identifying similar diseases could potentially provide deeper understanding of their underlying causes, and may even hint at possible treatments. For this purpose, it is necessary to have a similarity measure that reflects the underpinning molecular interactions and biological pathways. We have thus devised a network-based measure that can partially fulfill this goal. Our method assigns weights to all proteins (and consequently their encoding genes) by using information flow from a disease to the protein interaction network and back. Similarity between two diseases is then defined as the cosine of the angle between their corresponding weight vectors. The proposed method also provides a way to suggest disease-pathway associations by using the weights assigned to the genes to perform enrichment analysis for each disease. By calculating pairwise similarities between 2534 diseases, we show that our disease similarity measure is strongly correlated with the probability of finding the diseases in the same disease family and, more importantly, sharing biological pathways. We have also compared our results to those of MimMiner, a text-mining method that assigns pairwise similarity scores to diseases. We find the results of the two methods to be complementary. It is also shown that clustering diseases based on their similarities and performing enrichment analysis for the cluster centers significantly increases the term association rate, suggesting that the cluster centers are better representatives for biological pathways than the diseases themselves. This lends support to the view that our similarity measure is a good indicator of relatedness of biological processes involved in causing the diseases. Although not needed for understanding this paper, the raw results are available for download for further study at ftp://ftp.ncbi.nlm.nih.gov/pub/qmbpmn/DiseaseRelations/.     

Validity of using recombinant melon profilin,Cuc m 2, for diagnosis of melon allergy

Background:

Allergy is a clinical disorder affecting humans worldwide. Allergenic extracts prepared from natural source materials remain heterogeneous in composition and content, but are regularly used for diagnosis and immunotherapy. Recombinant allergens are suitable candidates to use in place of natural allergens; however, the recombinant allergens should be assessed and compared with the natural ones. Cuc m 2 (profilin), one of the most important allergens of melon (Cucumis melo), has been cloned and was expressed in Escherichia coli (E. coli). We aimed to evaluate the validity of recombinant Cuc m 2 (rCuc m 2) in the diagnosis of melon allergy and investigate whether rCuc m 2 could be used as a replacement for natural Cuc m 2 (nCuc m 2).

Methods:

nCuc m 2 was purified by immuno-affinity chromatography and rCuc m 2 was purified by metal-affinity chromatography. SDS-PAGE and western blotting were carried out to evaluate the purification methods. Skin prick tests (SPT), and enzyme immunoassays to determine specific IgE, were performed with the natural and recombinant purified allergens on 53 patients with melon allergy.

Results:

rCuc m 2 elicited no significantly different responses in skin compared with nCuc m 2. All patients'' sera showed similar ODs in ELISAs with natural and recombinant profilin.

Conclusion:

rCuc m 2 evoked strong immuno-reactivity equivalent to nCuc m 2, and has potential for diagnosis of melon allergy. Key Words: Allergy, Cuc m 2, Melon, Natural allergen, Recombinant allergen     

Gene copy number variation and its significance in cyanobacterial phylogeny

ABSTRACT: BACKGROUND: In eukaryotes, variation in gene copy numbers is often associated with deleterious effects, but may also have positive effects. For prokaryotes, studies on gene copy number variation are rare. Previous studies have suggested that high numbers of rRNA gene copies can be advantageous in environments with changing resource availability, but further association of gene copies and phenotypic traits are not documented. We used one of the morphologically most diverse prokaryotic phyla to test whether numbers of gene copies are associated with levels of cell differentiation. RESULTS: We implemented a search algorithm that identified 44 genes with highly conserved copies across 22 fully sequenced cyanobacterial taxa. For two very basal cyanobacterial species, Gloeobacter violaceus and a thermophilic Synechococcus species, distinct phylogenetic positions previously found were supported by identical protein coding gene copy numbers. Furthermore, we found that increased ribosomal gene copy numbers showed a strong correlation to cyanobacteria capable of terminal cell differentiation. Additionally, we detected extremely low variation of 16S rRNA sequence copies within the cyanobacteria. We compared our results for 16S rRNA to three other eubacterial phyla (Chroroflexi, Spirochaetes and Bacteroidetes). Based on Bayesian phylogenetic inference and the comparisons of genetic istances, we could confirm that cyanobacterial 16S rRNA paralogs and orthologs show significantly stronger conservation than found in other eubacterial phyla. Conclusions: A higher number of ribosomal operons could potentially provide an advantage to terminally differentiated cyanobacteria. Furthermore, we suggest that 16S rRNA gene copies in cyanobacteria are homogenized by both concerted evolution and purifying selection. In addition, the small ribosomal subunit in cyanobacteria appears to evolve at extraordinary slow evolutionary rates, an observation that has been made previously for morphological characteristics of cyanobacteria.     

Electronic sensor for sulfide dioxide based on AlN nanotubes: a computational study

Single-walled aluminum nitride nanotubes (AlNNTs) are introduced as an electronic sensor for detection of sulfur dioxide (SO(2)) molecules based on density functional theory calculations. The proposed sensor benefits from several advantages including high sensitivity: HOMO-LUMO energy gap of the AlNNT is appreciably sensitive toward the presence of SO(2) so that it decreases from 4.11?eV in the pristine tube to 1.01?eV in the SO(2)-adsorbed form, pristine application: this nanotube can detect the SO(2) molecule in its pristine type without manipulating its structure through doping, chemical functionalization, making defect, etc., short recovery time: the adsorption energy of SO(2) molecule is not so large to hinder the recovery of AlNNTs and therefore the sensor will possess short recovery times, and good selectivity: the tube can selectively detect the SO(2) molecule in the presence of several molecules such as H(2)O, CO, NH(3), HCOH, CO(2), N(2), and H(2).     

Amyloid Beta1-40-Induced Astrogliosis and the Effect of Genistein Treatment in Rat: A Three-Dimensional Confocal Morphometric and Proteomic Study

Astrocytes are highly involved in regulation and homeostasis of the extracellular environment in the healthy brain. In pathological conditions, these cells play a major role in the inflammatory response seen in CNS tissues, which is called reactive astrogliosis and includes hypertrophy and proliferation of astrocytes. Here, we performed 3D confocal microscopy to evaluate the morphological response of reactive astrocytes positive for glial fibrillary acidic protein (GFAP) in rats, to the presence of Aβ_1–40 in the rat brain before and after treatment with genistein. In 50 astrocytes per animal, we measured the volume and surface area for the nucleus, cell body, the entire cell, the tissue covered by single astrocytes and quantified the number and length of branches, the density of the astrocytes and the intensity of GFAP immunoreactivity. Injecting Aβ_1–40 into the brain of rats caused astrogliosis indicated by increased values for all measured parameters. Mass spectrometric analysis of hippocampal tissue in Aβ_1–40-injected brain showed decreased amounts of tubulins, enolases and myelin basic protein, and increased amounts of dihydropyrimidinase-related protein 2. In Aβ_1–40-injected rats pretreated with genistein, GFAP intensity was decreased to the sham-operated group level, and Aβ_1–40-induced astrogliosis was significantly ameliorated.     

Genetic variation in the response of pea (Pisum sativum L.) to high soil concentrations of boron

In a greenhouse experiment nine current Australian cultivars of pea were grown to flowering time under five levels of soil boron (0, 10, 20, 30 and 40 mg kg^–1) applied to the soil. This study was conducted to identify the genetic range in tolerance to boron within the group and to identify specific responses which may be utilised as selection criteria in a breeding program. Significant differences in response to increasing levels of boron were found between cultivars for dry-weight yield, and boron concentrations were lowest in shoots of the most tolerant cultivars. Of the other parameters measured, emergence was not affected but plant height and the number of nodes were reduced and the severity of symptom expression increased at the higher boron treatments. Symptom expression was the most efficient observation for predicting the response of cultivars, as determined by dry-weight yield and concentration of boron in shoots, and it was found that the correlation coefficients between symptoms and the latter two measurements were r=–0.78 (p<0.01) and r=0.81 (p<0.01), respectively. Early Dun, Dundale, Alma and Maitland were the more tolerant of the cultivars and these happen to be the most widely grown cultivars in southern Australia.     

Continuous rearing on Ephestia kuehniella reshaped quality of the parasitoid wasp Trichogramma brassicae (Hymenoptera: Trichogrammatidae)

Continuous mass rearing of Trichogramma brassicae (Bezdenko) at commercial mass-rearing insectaries may affect both quality and performance of natural enemies. In the present study, we studied the quality and performance of a colony of T. brassicae reared for over 45 generations (G) on Ephestia kuehniella Zeller using two-sex life table parameters and parasitism capacity. Our results revealed that although different generations showed no significant difference in terms of female longevity or total life span until G35, G5 and G10 had the highest values of fecundity, gross reproductive rate (GRR), net reproductive rate (R₀), intrinsic rate of natural increase (r), and finite rate of increase (λ). No significant difference in male adult longevity was found among different generations. The longest and shortest mean generation times (T) were found in G10 (13.65 ± 2.31 d) and G45 (13.25 ± 3.37 d), respectively. The finite rate of parasitism (ω) ranged from 0.355 ± 2.332 host/parasitoid/day in G5 to 0.242 ± 0.017 host/parasitoid/day in G45. However, ω did not show any significant difference until G20. These results indicate that T. brassicae wasps held under continuous laboratory rearing declined in quality after 20 generations, and therefore periodical rejuvenation of the colony by adding feral parasitoids is strongly recommended.     

Angiotensinogen,angiotensine converting enzyme and plasminogen activator inhibitor-1 gene polymorphism in chronic allograft dysfunction

Despite dramatic improvements in first-year patient and graft survival rates, chronic allograft dysfunction (CAD) remains the leading cause of late renal allograft loss, while current immunologic strategies have little effect on this condition. The renin–angiotensin system (RAS) plays an important role in progression of chronic renal disease. It was shown that plasminogen activator inhibitor-1 (PAI-1) functions in the RAS. This study investigates the possible links between angiotensinogen (AGT M235T), angiotensin-converting enzyme (ACE) and PAI-1 genotypes with CAD. Assessments of polymorphism were performed in 127 renal allograft recipients (77 with CAD and 50 with normal renal function). Fifty healthy subjects were also considered for comparison. Genotypes were determined using polymerase chain reaction (PCR) sequence-specific primers and PCR followed by restriction fragment length polymorphism analysis. Kidney recipients with CAD had significantly higher frequencies of the TT than the recipients without CAD (P < 0.05). The transplant recipients with CAD also had significantly higher frequencies of the DD genotype than those without CAD (P < 0.05). No significant differences were observed between the allelic and genotypic distributions of PAI-1 polymorphisms. Therefore, determination of AGT M235T and ACE genotypes prior to transplantation may be useful to identify patients who are at risk for chronic renal transplant dysfunction.