Resveratrol potentially increased the tumoricidal effect of doxorubicin on SKOV3 cancer stem cells in vitro

Ovarian cancer is associated with a high percentage of recurrence of tumor and resistance to chemotherapy. Cancer stem cells (CSCs) form a rare population with a significant capacity to begin and expand malignant diseases. Eliminating the drug resistance of CSCs by factors that have fewer side effects to the patient is vital. To investigate the effect of resveratrol (RES) and doxorubicin (DOX) on drug resistance and apoptosis of CSCs; at the first, isolation of CSCs from SKOV3 ovarian carcinoma cells and their dosage adjustment (IC₅₀) with RES and DOX was performed. Then, isolated CSCs were treated with RES and DOX IC ₅₀ of 55 and 250 nM, respectively. Subsequently, their effects on drug resistance and cell death were evaluated using real-time polymerase chain reaction, rhodamine 123 uptakes. The results of the present study demonstrated that treatment of SKOV3 with 55 μM of RES and 250 nM of DOX simultaneously increased cell viability in CSCs to DOX after 24 and 48 hours by increasing the expression of Bcl-2-associated X protein (BAX) and caspase-3 genes, and decreased the expression and function of multidrug resistance protein 1 (MDR1) and multidrug resistance-associated protein 1 (MRP1) genes indicated by intracellular the rhodamine 123 content. Treatment of RES could increase the activity of DOX cell viability in CSCs originated from SKOV3 ovarian carcinoma and decrease drug resistance capacity to DOX.     

An analytical enrichment-based review of structural genetic studies on keratoconus

Keratoconus is a progressive bilateral corneal protrusion that leads to irregular astigmatism and impairment of vision. Keratoconus is an etiologically heterogeneous corneal dystrophy and both environmental and genetic factors play a role in its etiopathogenesis. In this analytical review, we have studied all the genes that are structurally associated with keratoconus and have tried to explain the function of each gene and its association with other eye disorders in a concise way. In addition, using gene set enrichment analysis, it was attempted to find the most important impaired metabolic pathways in keratoconus. Several genetic studies have been carried out on keratoconus and several genes have been identified as risk factors involved in the etiology of the disease. In the current study, 16 studies, including nine association studies, five genome-wide association studies, one linkage study, and one meta-analysis, were reviewed and based on the 19 genes found, enrichment was performed and the most important metabolic pathways involved in the disease were identified. The enrichment results indicated that the two pathways, interleukin 1 processing and assembly of collagen fibrils, are significantly associated with the disease. Obviously, the results of this study, in addition to providing information about the genes involved in the disease, can provide an integrated insight into the gene-based etiology of keratoconus and therapeutic opportunities thereof.     

MSLN (Mesothelin), ANTXR1 (TEM8), and MUC3A are the potent antigenic targets for CAR T cell therapy of gastric adenocarcinoma

Gastric adenocarcinoma is usually diagnosed in late stages, necessitating the use of different therapeutic modalities. Currently, antibody-based therapies have also been approved through with limited clinical efficacy. Reinforcing antibody-based immunotherapy by using chimeric antigen receptor (CAR) T cells may enhance the approach. However, the cells can cause severe on-target and off-tumor toxicities owing to their higher sensitivity to low-level antigen expressions. To address the need for safe and reliable targets, we made a bioinformatics pipeline by which we screened overexpressed genes in the disease for off-tumor sites in many normal tissues. Our inspection showed that MSLN (Mesothelin), ANTXR1 (TEM8), and MUC3A are the probable targets of CAR T cell therapy in gastric adenocarcinoma. The proposed antigenic targets might respond to the need to simultaneously target multiple antigens in a tumor matrix to prevent resistance.     

The reelin pathway modulates the structure and function of retinal synaptic circuitry

The formation of synaptic connections requires the coordination of specific guidance molecules and spontaneous neuronal activity. The visual system has provided a useful model for understanding the role of these cues in shaping the precise connections from the neural retina to the brain. Here, we demonstrate that two essential genes in the Reelin signaling pathway function during the patterning of synaptic connectivity in the retina. Physiological studies of mice deficient in either reelin or disabled-1 reveal an attenuation of rod-driven retinal responses. This defect is associated with a decrease in rod bipolar cell density and an abnormal distribution of processes in the inner plexiform layer. These results imply that, in addition to its essential role during neuronal migration, the Reelin pathway contributes to the formation of neuronal circuits in the central nervous system.     

When reverse genetics meets physiology: the use of site-specific recombinases in mice

The use of site-specific recombinases enables the precise introduction of defined genetic mutations into the mouse genome. In theory, any deletion, point mutation, inversion or translocation can be modeled in mice. Because gene targeting is controlled both spatially and temporally, the function of a given gene can be studied in the desired cell types and at a specific time point. This 'genetic dissection' allows to define gene function in development, physiology or behavior. In this review, we focus on the technical possibilities of Cre and other site-specific recombinases but also discuss their limitations.     

Heavy metal concentrations in the breast milk of Saudi women

Lead, cadmium, and mercury concentrations were determined in breast milk of Saudi lactating mothers from Riyadh and Al-Ehssa regions in Saudi Arabia who were not occupationally exposed. The mean levels for cadmium, lead, and mercury were 1.732 μg/L, 31.671 μg/L, and 3.100 μg/L, respectively. In contrast to mercury, mothers living in the Al-Ehssa region had significantly higher cadmium and lead concentrations in their breast milk than those in the Riyadh region. The estimated weekly intakes of cadmium, lead, and mercury of breast-fed infants in this study were in some cases higher than the Provisional Tolerance Weekly Intake (PTWI) recommended by FAO/WHO, which pose a threat to their health. This necessitates the urgent need to undertake a comprehensive study to determine the sources of exposure to these heavy metals. Breast-feeding is of great benefical value for the infant’s development; therefore, efforts should be made to prevent its contamination with environmental pollutants.     

Regeneration and transformation via Agrobacterium tumefaciens of the strawberry cultivar Chandler

The effects of growth regulator balance and culture conditions on the morphogenetic response of leaf disks from greenhouse grown plants of the strawberry cultivar Chandler, have been studied. Best results were obtained in the presence of 2.46 μM IBA and 8.88 μM BA, where 47% of the cultures regenerated after 16 weeks with 2.9 shoot colonies per regenerating leaf disk. Optimum incubation conditions included two weeks in the dark with subsequent transfer to light (40 μmol m-2 s-1, 16 h). The regeneration protocol was also valuable when leaf disks from in vitro grown plants were used as explants. Transformation was attempted using Agrobacterium tumefaciens carrying the plasmid pBI121. Leaf disks from in vitro cultures proliferating in the presence of 2.21 μM kinetin were best explants for transformation. A 4.22% of inoculated explants showed kanamycin resistance after 16 weeks in a medium containing 25 mg l-1 of this antibiotic. The transgenic nature of several shoots was also confirmed by the GUS assay and PCR analysis. This revised version was published online in June 2006 with corrections to the Cover Date.     

A Novel Pore-Forming Toxin in Type A Clostridium perfringens Is Associated with Both Fatal Canine Hemorrhagic Gastroenteritis and Fatal Foal Necrotizing Enterocolitis

A role for type A Clostridium perfringens in acute hemorrhagic and necrotizing gastroenteritis in dogs and in necrotizing enterocolitis of neonatal foals has long been suspected but incompletely characterized. The supernatants of an isolate made from a dog and from a foal that died from these diseases were both found to be highly cytotoxic for an equine ovarian (EO) cell line. Partial genome sequencing of the canine isolate revealed three novel putative toxin genes encoding proteins related to the pore-forming Leukocidin/Hemolysin Superfamily; these were designated netE, netF, and netG. netE and netF were located on one large conjugative plasmid, and netG was located with a cpe enterotoxin gene on a second large conjugative plasmid. Mutation and complementation showed that only netF was associated with the cytotoxicity. Although netE and netG were not associated with cytotoxicity, immunoblotting with specific antisera showed these proteins to be expressed in vitro. There was a highly significant association between the presence of netF with type A strains isolated from cases of canine acute hemorrhagic gastroenteritis and foal necrotizing enterocolitis. netE and netF were found in all cytotoxic isolates, as was cpe, but netG was less consistently present. Pulsed-field gel electrophoresis showed that netF-positive isolates belonged to a clonal population; some canine and equine netF-positive isolates were genetically indistinguishable. Equine antisera to recombinant Net proteins showed that only antiserum to rNetF had high supernatant cytotoxin neutralizing activity. The identifica-tion of this novel necrotizing toxin is an important advance in understanding the virulence of type A C. perfringens in specific enteric disease of animals.     

Intracellular Cytokine Staining and Flow Cytometry: Considerations for Application in Clinical Trials of Novel Tuberculosis Vaccines

Intracellular cytokine staining combined with flow cytometry is one of a number of assays designed to assess T-cell immune responses. It has the specific advantage of enabling the simultaneous assessment of multiple phenotypic, differentiation and functional parameters pertaining to responding T-cells, most notably, the expression of multiple effector cytokines. These attributes make the technique particularly suitable for the assessment of T-cell immune responses induced by novel tuberculosis vaccines in clinical trials. However, depending upon the particular nature of a given vaccine and trial setting, there are approaches that may be taken at different stages of the assay that are more suitable than other alternatives. In this paper, the Tuberculosis Vaccine Initiative (TBVI) TB Biomarker Working group reports on efforts to assess the conditions that will determine when particular assay approaches should be employed. We have found that choices relating to the use of fresh whole blood or peripheral blood mononuclear cells (PBMC) and frozen PBMC; use of serum-containing or serum-free medium; length of stimulation period and use of co-stimulatory antibodies can all affect the sensitivity of intracellular cytokine assays. In the case of sample material, frozen PBMC, despite some loss of sensitivity, may be more advantageous for batch analysis. We also recommend that for multi-site studies, common antibody panels, gating strategies and analysis approaches should be employed for better comparability.