Development and use of quantitative competitive PCR assays for relative quantifying rumen anaerobic fungal populations in both in vitro and in vivo systems

This paper describes the use of a quantitative competitive polymerase chain reaction (QC-PCR) assay; using PCR primers to the rRNA locus of rumen fungi and a standard-control DNA including design and validation. In order to test the efficiency of this method for quantifying anaerobic rumen fungi, it has been attempted to evaluate this method in in vitro conditions by comparing with an assay based on measuring cell wall chitin. The changes in fungal growth have been studied when they are grown in in vitro on either untreated (US) or sodium hydroxide treated wheat straw (TS). Results showed that rumen fungi growth was significantly higher in treated samples compared with untreated during the 12 d incubation (P < 0.05) and plotting the chitin assay's results against the competitive PCR's showed high positive correlation (R² ≥ 0.87). The low mean values of the coefficients of variance in repeatability in the QC-PCR method against the chitin assay demonstrated more reliability of this new approach. And finally, the efficiency of this method was investigated in in vivo conditions. Samples of rumen fluid were collected from four fistulated Holstein steers which were fed four different diets (basal diet, high starch, high sucrose and starch plus sucrose) in rotation. The results of QC-PCR showed that addition of these non-structural carbohydrates to the basal diets caused a significant decrease in rumen anaerobic fungi biomass. The QC-PCR method appears to be a reliable and can be used for rumen samples.     

Prognostic implication of CDC25A and cyclin E expression on primary breast cancer patients

Defect in cell cycle control is a hallmark character of cancer. We have investigated the association of Ki67 labeling index, cyclin E and CDC25A expressions with clinical follow-up data in breast carcinomas. Flow cytometry was used to detect gene amplification of cyclins in 44 tumor tissue with invasive breast carcinomas. Multivariate Cox proportional hazard ratio test was used to show the correlations. Cyclin E or CDC25A were upregulated in 34% of the tumors. Among the whole total material, expression of cyclin E and of CDC25A were found upregulated in 31.9% and 39.4% of cells, respectively. Both CDC25A and cyclin E protein expression levels were correlated with Ki67 expression level (p < 0.001). In addition, the expression of CDC25A was associated significantly with poor survival (P = 0.028), whereas no correlation was found with cyclin E. These findings suggest a possible prognostic value for CDC25A as a cell cycle marker and may imply in characteristic of high risk breast cancer patients.     

Role of Bach-1 in regulation of heme oxygenase-1 in human liver cells: insights from studies with small interfering RNAS

Heme oxygenase-1 is an antioxidant defense enzyme that converts heme to biliverdin, iron, and carbon monoxide. Bach-1 is a bZip protein that forms heterodimers with small Maf proteins and was reported recently to down-regulate the HO-1 gene in mice. Using small interfering RNAs targeted to human Bach-1 mRNA, we investigated whether modulation of human hepatic Bach-1 expression by small interfering (si)RNA technology influences heme oxygenase-1 gene expression. We found that Bach-1 siRNAs transfected into Huh-7 cells significantly reduced Bach-1 mRNA and protein levels approximately 80%, compared with non siRNA-treated cells. In contrast, transfection with the same amounts of nonspecific control duplexes or LaminB2-duplex did not reduce Bach-1 mRNA or protein levels, confirming the specificity of Bach-1 siRNA. Expression of the heme oxygenase-1 gene in Bach-1 siRNA-transfected cells was up-regulated 7-fold, compared with cells without Bach-1 siRNA. The effect of increasing concentrations of heme to up-regulate levels of heme oxygenase-1 was more pronounced when Bach-1 siRNA was present. Taken together, these results indicated that Bach-1 has a specific and selective ability to repress expression of human hepatic heme oxygenase-1. Silencing of Bach-1 by siRNAs is a useful method for up-regulating HO-1 gene expression. Exogenous heme produces additional up-regulation, beyond that produced by Bach-1 siRNAs, suggesting that heme does not act solely through its effects on Bach-1.     

Antimicrobial activities of Asafoetida and Shirazi thyme essential oils improve the vase life of gerbera cut flowers

This study was conducted to evaluate the chemical composition of asafoetida (Ferula assa-foetida) essential oil (FAEO) and Shirazi thyme (Zataria multiflora) EO (ZMEO) and their impact on vase life of gerbera cut flowers (Gerbera jamesonii cv. Rosalyn). Five concentrations of both, ZMEO and FAEO including 0, 100, 200, 300 and 400 mg L^?1 used as continuous vase solution for gerbera cut flowers. EOs used in this study were extracted by hydrodistillation method using Clevenger apparatus. They were analyzed by GC and GC–MS for determination of the active compounds. Thirty five compounds were identified in ZMEO, mainly including thymol (40.1%), p-cymene (15.5%) and carvacrol (6.5%). Also, thirty compounds were identified in FAEO. The main components were trans propenyl sec-butyl disulfide (21.7%), eudesmol (10-epu-γ) (19.2%) and cis propenyl sec-butyl disulfide (10.2%). The results showed that both ZMEO and FAEO at all concentrations could act as an effective antibacterial compounds and this property increased by increasing their concentration. The results of this research showed that ZMEO increased the vase life at all concentrations but high concentrations of FAEO increased mortality percentage and reduced the vase life of cut flowers. The relative fresh weight and vase solution uptake of gerbera cut flowers increased by the applied EOs treatments. ZMEO at 400 mg L^? 1 and FAEO at 300 and 400 mg L^? 1 resulted the least stem color change. Overall, 200 mg L^? 1 ZMEO and 100 mg L^? 1 FAEO were the best treatments for maintenance of gerbera cut flowers quality during vase life.     

Coupling of bioaugmentation and phytoremediation to improve PCBs removal from a transformer oil-contaminated soil

This study was carried out to assess the dissipation of 17 selected polychlorinated biphenyl (PCB_i) congeners in a transformer oil-contaminated soil using bioaugmentation with 2 PCB-degrading bacterial strains, i.e., Pseudomonas spp. S5 and Alcaligenes faecalis, assisted or not by the maize (Zea mays L.) plantation. After 5 and 10 weeks of treatment, the remaining concentrations of the target PCB_i congeners in the soil were extracted and measured using GC-MS. Results showed that the bacterial augmentation treatments with Pseudomonas spp. S5 and A. faecalis led to 21.4% and 20.4% reduction in the total concentration of the target PCBs (ΣPCB_i), respectively, compared to non-bioaugmented unplanted control soil. The ΣPCB_i decreased by 35.8% in the non-bioaugmented planted soil compared with the control. The greatest degradation of the PCB congeners was observed over a 10-week period in the soil inoculated with Pseudomonas spp. S5 and cultivated with maize. Under this treatment, the ΣPCB_i decreased from 357 to 119 ng g^?1 (66.7% lower) and from 1091 to 520 ng g^?1 (52.3% lower). Overall, the results suggested that the combined application of phytoremediation and bioaugmentation was an effective technique to remove PCBs and remediate transformer oil-contaminated soils.     

Assessing the ecological health status using macrobenthic communities of tropical coastal water

The relative contributions of natural and anthropogenic fluctuations are different in shaping habitat health status and natural benthic communities in tropical coastal water. Understanding responses of coastal benthic communities to these fluctuations are still equivocal and thus available data are inadequate. Here, multiple analytical approaches were used to address the significant risk factors and their impacts on coastal benthic habitat health through space and time. A total abundance of 1436 ± 612 individuals of 33 benthic species were counted and identified from 22 sampling stations across eight sampling periods over the two years of study. Bioassay results showed that the benthic community is moderately exposed to anthropogenic pollutants in Klang Strait coastal water. The results showed that there were no significant temporal changes of habitat health status and macrobenthic community structure; however, spatial changes were significant and synchronized with anthropogenic and natural fluctuations. This study demonstrates that Cd and Hg levels and sediment characteristics played the primary role in shaping the habitat health and macrobenthic assemblages, whereas the influence of other factors were insignificant.     

Single versus double pass technique for preparation of ultrathin Descemet’s stripping automated endothelial keratoplasty tissues from donated whole eyes

This study was conducted to analyze the preoperative thickness profile and endothelial rating of ultrathin Descemet’s stripping automated endothelial keratoplasty (UT-DSAEK) tissues prepared with a single versus double microkeratome pass from donated whole eyes and corresponding eye bank postoperative results. Microkeratome-assisted UT-DSAEK tissues were prepared from freshly donated whole eyes with single-pass (SP) and double-pass (DP) technique in the Central Eye Bank of Iran. Preoperative thickness profiles and endothelial cell densities of UT-DSAEK tissues were obtained from optical coherence tomography and specular microscopy, respectively, and compared between groups. Corneal perforation rates during the eye bank preparation and postoperative reports of transplanted UT-DSAEK tissues were also compared. Over a 15-month period, 342 UT-DSAEK tissues were prepared: 248 via SP and 94 with DP technique. Mean donor corneal central thickness was 610?±?58 µm with SP and 790?±?100 µm with DP technique. Mean central thickness of UT-DSAEK tissues was not statistically different between the groups (84.8?±?11.0 µm with SP and 85.1?±?10.5 µm with DP technique, P?=?0.857). Mean increase of UT-DSAEK thickness from central to pericentral and peripheral cornea was not significantly different with both techniques. Mean differences between thicknesses of 2 pericentral locations and between those of 2 peripheral locations were not statistically different in the study groups. Corneal perforation of 1.6 and 1.1% occurred in SP and DP groups, respectively. Failed graft was reported 6 months postoperatively in 4 (1.6%) cases with SP and in 1 (1.1%) case with DP technique. Preoperative thickness profiles of UT-DSAEK tissues prepared from donated whole eyes via SP technique were not significantly different from those prepared with DP, showing a symmetric increase of thickness towards peripheral locations.     

Evaluation of application of EPID for rapid QC testing of linear accelerator

Aim

Evaluation of application of EPID for rapid QC testing of linear accelerator.

Thermal dissection of lentil seedling amine oxidase domains by differential scanning calorimetry

The relationships between the structural and energetic domains of lentil seedling amine oxidase (LSAO) were investigated using modifiers that target the active site and the carbohydrate moiety of the enzyme. An irreversible inhibitor, aminoguanidine, specifically modified the active site of the lentil enzyme, whereas sodium metaperiodate cleaves carbohydrate moieties covalently bound to the native enzyme. Differential scanning calorimetry (DSC) measurements were made on the modified LSAOs. Deconvolution of the reversible thermal DSC profiles of the modified enzyme gave three subpeaks (energetic domains), each of which was assigned to one of the three structural domains of the native protein. Our results led us to conclude that deglycosylation of LSAO has no effect on thermal stability, whereas binding of the inhibitor imparts more stability to the enzyme.     

Poly (3-hydroxybutyrate-co-3-hydroxyvalerate) improved osteogenic differentiation of the human induced pluripotent stem cells while considered as an artificial extracellular matrix

Cocell polymers can be the best implants for replacing bone defects in patients. The pluripotent stem cells produced from the patient and the nanofibrous polymeric scaffold that can be completely degraded in the body and its produced monomers could be also usable are the best options for this implant. In this study, electrospun poly (3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) nanofibers were fabricated and characterized and then osteogenic differentiation of the human-induced pluripotent stem cells (iPSCs) was investigated while cultured on PHBV scaffold. MTT results showed that cultured iPSCs on PHBV proliferation were increased compared to those cultured on tissue culture polystyrene (TCPS) as the control. Alkaline phosphatase (ALP) activity and calcium content were also significantly increased in iPSCs cultured on PHBV compared to the cultured on TCPS under osteogenic medium. Gene expression evaluation demonstrated that Runx2, collagen type I, ALP, osteonectin, and osteocalcin were upregulated in iPSCs cultured on PHBV scaffold in comparison with those cultured on TCPS for 2 weeks. Western blot analysis have shown that osteocalcin and osteopontin expression as two major osteogenic markers were increased in iPSCs cultured on PHBV scaffold. According to the results, nanofiber-based PHBV has a promising potential to increase osteogenic differentiation of the stem cells and iPSCs-PHBV as a cell-co-polymer construct demonstrated that has a great efficiency for use as a bone tissue engineered bioimplant.