Preparation and Evaluation of Gelatin/Sodium Carboxymethyl Cellulose Polyelectrolyte Complex Microparticles for Controlled Delivery of Isoniazid

The ratio of gelatin to sodium carboxymethyl cellulose (SCMC) at which maximum yield was obtained was optimized. This optimized ratio of gelatin to SCMC along with other parameters was used to prepare microparticles of different sizes. Vegetable oil was used as emulsion medium. Effect of various factors like amount of surfactant, concentration of polymer on the formation, and size of the microparticles was investigated. These microparticles were used as carrier for isoniazid. Among different cross-linkers, glutaraldehyde was found to be the most effective cross-linker at the temperature and pH at which the reaction was carried out. The loading efficiency and release behavior of loaded microparticles were found to be dependent on the amount of cross-linker used, concentration of drug, and time of immersion. Maximum drug loading efficiency was observed at higher immersion time. The release rate of isoniazid was more at higher pH compared to that of at lower pH. The sizes of the microparticles were investigated by scanning electron microscope. In all the cases, the microparticles formed were found spherical in shape except to those at low stirring speed where they were agglomerated. Fourier transform infrared study indicated the successful incorporation of isoniazid into the microparticles. Differential scanning calorimetry study showed a molecular level dispersion of isoniazid in the microparticles. X-ray diffraction study revealed the development of some crystallinity due to the encapsulation of isoniazid.     

Evaluation of possible mechanisms of protective role of Tamarix gallica against DEN initiated and 2-AAF promoted hepatocarcinogenesis in male Wistar rats

We have previously reported that Tamarix gallica caused a marked inhibition of thioacetamide-induced hepatotoxicity, oxidative damage and early tumor promotion related events in the liver. These results strongly indicates that T. gallica may have chemopreventive potential. Therefore, in the present study, we examined the inhibitory effects of T. gallica methanolic extract on diethylnitrosamine (DEN) initiated and 2-acetyl aminofluorene (2-AAF) promoted liver carcinogenesis in male Wistar rats. Interestingly, it was found that T. gallica (25 and 50 mg/kg body wt.) resulted in a marked reduction of the incidence of liver tumors. The study was further histologically confirmed. Furthermore to understand the underlying mechanisms of chemopreventive action by T. gallica we evaluated the levels activities of hepatic antioxidant defense enzymes, ornithine decarboxylase activity and hepatic DNA synthesis as a marker for tumor promotion since direct correlation between these marker parameters and carcinogenicity have been well documented. Treatment of male Wistar rats for five consecutive days with 2-AAF i.p. induced significant hepatic toxicity, oxidative stress and hyperproliferation. Pretreatment of T. gallica extract (25 and 50 mg/kg body wt.) prevented oxidative stress by restoring the levels of antioxidant enzymes and also prevented toxicity at both the doses. The promotion parameters induced (ornithine decarboxylase activity and DNA synthesis) by 2-AAF administration in diet with partial hepatectomy (PH) were also significantly suppressed dose-dependently by T. gallica. Therefore, we can conclude that ultimately the protection against liver carcinogenesis by T. gallica methanolic extract might be mediated by multiple actions, which include restoration of cellular antioxidant enzymes, detoxifying enzymes, ODC activity and DNA synthesis.     

Characterization of Sr9h,a wheat stem rust resistance allele effective to Ug99

Key message

Wheat stem rust resistance gene SrWeb is an allele at the Sr9 locus that confers resistance to Ug99.

Abstract

Race TTKSK (Ug99) of Puccinia graminis f. sp. tritici, the causal fungus of stem rust, threatens global wheat production because of its broad virulence to current wheat cultivars. A recently identified Ug99 resistance gene from cultivar Webster, temporarily designated as SrWeb, mapped near the stem rust resistance gene locus Sr9. We determined that SrWeb is also present in Ug99 resistant cultivar Gabo 56 by comparative mapping and an allelism test. Analysis of resistance in a population segregating for both Sr9e and SrWeb demonstrated that SrWeb is an allele at the Sr9 locus, which subsequently was designated as Sr9h. Webster and Gabo 56 were susceptible to the Ug99-related race TTKSF+ from South Africa. Race TTKSF+ possesses unique virulence to uncharacterized Ug99 resistance in cultivar Matlabas. This result validated that resistance to Ug99 in Webster and Gabo 56 is conferred by the same gene: Sr9h. The emergence of pathogen virulence to several resistance genes that are effective to the original Ug99 race TTKSK, including Sr9h, suggests that resistance genes should be used in combinations in order to increase resistance durability.     

Glutathione peroxidase3 of <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis> protects phospholipids during cadmium-induced oxidative stress

The present study was undertaken to determine the role of glutathione peroxidase3 (gpx3) in phospholipid protection in cells. Wild-type (WT) cells showed an overall increase in phospholipids upon 50 μM cadmium (Cd)-treatment, whereas an untreated gpx3Δ strain showed a drastic reduction in overall phospholipids which was further reduced with 50 μM Cd. In WT cells, Cd-exposure increased the short chain fatty acids and decreased the unsaturated fatty acids and the magnitude was high in Cd-treated gpx3Δ cells. Purified recombinant gpx3p showed higher activity with phospholipid hydroperoxides than shorter hydroperoxides. An increase in gpx activity was observed in Cd-treated WT cells and no such alteration was observed in gpx3Δ. WT cells treated with Cd showed an increase in MDA over untreated, while untreated gpx3Δ cells themselves showed a higher level of MDA which was further enhanced with Cd-treatment. Iron, zinc and calcium levels were significantly altered in WT and gpx3Δ cells during Cd-treatment.     

Development of transgenic strains for the biological control of the Mexican fruit fly, Anastrepha ludens

The Mexican fruit fly, Anastrepha ludens, is a highly significant agricultural pest species that has been genetically transformed with a piggyBac-based transposon vector system using independent vector and transposase helper plasmids. Minimum estimated germ-line transformation frequencies were approximately 13–21% per fertile G₀ individual, similar to previously reported frequencies using single vector-helper plasmids. Two vector constructs were tested with potential importance to transgenic strain development for mexfly biological control. The first allows post-integration stabilization of a transposon-vector by deletion of a terminal sequence necessary for mobilization. The complete pB[L1-EGFP-L2-DsRed-R1] vector was integrated into the Chiapas wild type strain with subsequent deletion of the L2-DsRed-R1 sub-vector carrying the piggyBac 3′ terminal sequence. Quality control tests for three of the stabilization vector lines (previous to stabilization) assessed viability at all life stages, fertility, adult flight ability, and adult male sexual competitiveness. All three transgenic lines were less fit compared to the wild strain by approximately 5–10% in most tests, however, there was no significant difference in sexual competitiveness which is the major prerequisite for optimal strain release. The second vector, pB[XL-EGFP, Asß2-tub-DsRed.T3], has the DsRed.T3 fluorescent protein reporter gene regulated by the A. suspensa Asß2-tubulin promoter, that resulted in testis and sperm-specific DsRed fluorescence in transgenic male mexflies. Fluorescent sperm bundles were unambiguously observed in the spermathecae of non-transgenic females mated to transgenic males. One transgenic line apparently had a male-specific Y-chromosome insertion, having potential use for sexing by fluorescent-embryo sorting. All transgenic lines expressed easily detectable and stable fluorescence in adults allowing their identification after trapping in the field.     

Potential novel candidate polymorphisms identified in genome-wide association study for breast cancer susceptibility

Previous genome-wide association studies (GWAS) have shown several risk alleles to be associated with breast cancer. However, the variants identified so far contribute to only a small proportion of disease risk. The objective of our GWAS was to identify additional novel breast cancer susceptibility variants and to replicate these findings in an independent cohort. We performed a two-stage association study in a cohort of 3,064 women from Alberta, Canada. In Stage I, we interrogated 906,600 single nucleotide polymorphisms (SNPs) on Affymetrix SNP 6.0 arrays using 348 breast cancer cases and 348 controls. We used single-locus association tests to determine statistical significance for the observed differences in allele frequencies between cases and controls. In Stage II, we attempted to replicate 35 significant markers identified in Stage I in an independent study of 1,153 cases and 1,215 controls. Genotyping of Stage II samples was done using Sequenom Mass-ARRAY iPlex platform. Six loci from four different gene regions (chromosomes 4, 5, 16 and 19) showed statistically significant differences between cases and controls in both Stage I and Stage II testing, and also in joint analysis. The identified variants were from EDNRA, ROPN1L, C16orf61 and ZNF577 gene regions. The presented joint analyses from the two-stage study design were not significant after genome-wide correction. The SNPs identified in this study may serve as potential candidate loci for breast cancer risk in a further replication study in Stage III from Alberta population or independent validation in Caucasian cohorts elsewhere.     

Influence of galectin-9/Tim-3 interaction on herpes simplex virus-1 latency

After HSV-1 infection, CD8(+) T cells accumulate in the trigeminal ganglion (TG) and participate in the maintenance of latency. However, the mechanisms underlying intermittent virus reactivation are poorly understood. In this study, we demonstrate the role of an inhibitory interaction between T cell Ig and mucin domain-containing molecule 3 (Tim-3)-expressing CD8(+) T cells and galectin 9 (Gal-9) that could influence HSV-1 latency and reactivation. Accordingly, we show that most K(b)-gB tetramer-specific CD8(+) T cells in the TG of HSV-1-infected mice express Tim-3, a molecule that delivers negative signals to CD8(+) T cells upon engagement of its ligand Gal-9. Gal-9 was also upregulated in the TG when replicating virus was present as well during latency. This could set the stage for Gal-9/Tim-3 interaction, and this inhibitory interaction was responsible for reduced CD8(+) T cell effector function in wild-type mice. Additionally, TG cell cultures exposed to recombinant Gal-9 in the latent phase caused apoptosis of most CD8(+) T cells. Furthermore, Gal-9 knockout TG cultures showed delayed and reduced viral reactivation as compared with wild-type cultures, demonstrating the greater efficiency of CD8(+) T cells to inhibit virus reactivation in the absence of Gal-9. Moreover, the addition of recombinant Gal-9 to ex vivo TG cultures induced enhanced viral reactivation compared with untreated controls. Our results demonstrate that the host homeostatic mechanism mediated by Gal-9/Tim-3 interaction on CD8(+) T cells can influence the outcome of HSV-1 latent infection, and manipulating Gal-9 signals might represent therapeutic means to inhibit HSV-1 reactivation from latency.     

CENP-A is essential for cardiac progenitor cell proliferation

Centromere protein A (CENP-A) is a homolog of histone H3 that epigenetically marks the heterochromatin of chromosomes. CENP-A is a critical component of the cell cycle machinery that is necessary for proper assembly of the mitotic spindle. However, the role of CENP-A in the heart and cardiac progenitor cells (CPCs) has not been previously studied. This study shows that CENP-A is expressed in CPCs and declines with age. Silencing CENP-A results in a decreased CPC growth rate, reduced cell number in phase G₂/M of the cell cycle, and increased senescence associated β-galactosidase activity. Lineage commitment is not affected by CENP-A silencing, suggesting that cell cycle arrest induced by loss of CENP-A is a consequence of senescence and not differentiation. CENP-A knockdown does not exacerbate cell death in undifferentiated CPCs, but increases apoptosis upon lineage commitment. Taken together, these results indicate that CPCs maintain relatively high levels of CENP-A early in life, which is necessary for sustaining proliferation, inhibiting senescence, and promoting survival following differentiation of CPCs.     

Gastroesophageal Reflux in Relation to Adenocarcinomas of the Esophagus: A Pooled Analysis from the Barrett’s and Esophageal Adenocarcinoma Consortium (BEACON)

Background

Previous studies have evidenced an association between gastroesophageal reflux and esophageal adenocarcinoma (EA). It is unknown to what extent these associations vary by population, age, sex, body mass index, and cigarette smoking, or whether duration and frequency of symptoms interact in predicting risk. The Barrett’s and Esophageal Adenocarcinoma Consortium (BEACON) allowed an in-depth assessment of these issues.

Methods

Detailed information on heartburn and regurgitation symptoms and covariates were available from five BEACON case-control studies of EA and esophagogastric junction adenocarcinoma (EGJA). We conducted single-study multivariable logistic regressions followed by random-effects meta-analysis. Stratified analyses, meta-regressions, and sensitivity analyses were also conducted.

Results

Five studies provided 1,128 EA cases, 1,229 EGJA cases, and 4,057 controls for analysis. All summary estimates indicated positive, significant associations between heartburn/regurgitation symptoms and EA. Increasing heartburn duration was associated with increasing EA risk; odds ratios were 2.80, 3.85, and 6.24 for symptom durations of <10 years, 10 to <20 years, and ≥20 years. Associations with EGJA were slighter weaker, but still statistically significant for those with the highest exposure. Both frequency and duration of heartburn/regurgitation symptoms were independently associated with higher risk. We observed similar strengths of associations when stratified by age, sex, cigarette smoking, and body mass index.

Conclusions

This analysis indicates that the association between heartburn/regurgitation symptoms and EA is strong, increases with increased duration and/or frequency, and is consistent across major risk factors. Weaker associations for EGJA suggest that this cancer site has a dissimilar pathogenesis or represents a mixed population of patients.     

Differential Regulation of Cellular Senescence and Differentiation by Prolyl Isomerase Pin1 in Cardiac Progenitor Cells

Autologous c-kit⁺ cardiac progenitor cells (CPCs) are currently used in the clinic to treat heart disease. CPC-based regeneration may be further augmented by better understanding molecular mechanisms of endogenous cardiac repair and enhancement of pro-survival signaling pathways that antagonize senescence while also increasing differentiation. The prolyl isomerase Pin1 regulates multiple signaling cascades by modulating protein folding and thereby activity and stability of phosphoproteins. In this study, we examine the heretofore unexplored role of Pin1 in CPCs. Pin1 is expressed in CPCs in vitro and in vivo and is associated with increased proliferation. Pin1 is required for cell cycle progression and loss of Pin1 causes cell cycle arrest in the G₁ phase in CPCs, concomitantly associated with decreased expression of Cyclins D and B and increased expression of cell cycle inhibitors p53 and retinoblastoma (Rb). Pin1 deletion increases cellular senescence but not differentiation or cell death of CPCs. Pin1 is required for endogenous CPC response as Pin1 knock-out mice have a reduced number of proliferating CPCs after ischemic challenge. Pin1 overexpression also impairs proliferation and causes G₂/M phase cell cycle arrest with concurrent down-regulation of Cyclin B, p53, and Rb. Additionally, Pin1 overexpression inhibits replicative senescence, increases differentiation, and inhibits cell death of CPCs, indicating that cell cycle arrest caused by Pin1 overexpression is a consequence of differentiation and not senescence or cell death. In conclusion, Pin1 has pleiotropic roles in CPCs and may be a molecular target to promote survival, enhance repair, improve differentiation, and antagonize senescence.