Cytotoxic T cell clones isolated from ovarian tumor-infiltrating lymphocytes recognize multiple antigenic epitopes on autologous tumor cells.

CTL clones were developed from tumor infiltrating lymphocytes (TIL) from the ascites of a patient with ovarian carcinoma by coculture of TIL with autologous tumor cells and subsequent cloning in the presence of autologous tumor cells. These CTL clones expressed preferential cytolytic activity against autologous tumor cells but not against allogeneic ovarian tumor cells and the NK-sensitive cell line K562. The cytolytic activity of these CTL against autologous tumors was inhibited by anti-TCR (WT31 mAb), anti-HLA class I, and anti-CD3 mAb but not by the NK function antibody Leu 11b. Cloning of the autologous tumor cells in vitro revealed that the CTL clones of the ovarian TIL expressed differential abilities to lyse autologous tumor cell clones. The specificity analysis of these autologous tumor specific CTL suggested that they recognize several antigenic determinants present on the ovarian tumor cells. Our results indicate the presence of at least three antigenic epitopes on the tumor cells (designated OVA-1A, OVA-1B, and OVA-1C), one of which (OVA-1C) is unstable. These determinants are present either simultaneously or separately, and six types of ovarian clones can be distinguished on the basis of their expression. These results indicate that CTL of the TIL detect intratumor antigenic heterogeneity. The novel heterogeneity identified within the ovarian tumor cells in this report may be of significance for understanding cellular immunity in ovarian cancer and developing adoptive specific immunotherapeutic approaches in ovarian cancer.     

The consequences of actin disruption at Sertoli ectoplasmic specialization sites facing spermatids after in vivo exposure of rat testis to cytochalasin D

Cytochalasin D (CD) was used to perturb actin filaments of the Sertoli ectoplasmic specialization (ES)--a cytoskeletal complex of the Sertoli cell related to spermatids. CD (500 microM for 6 h) produced a loss of 88% of the ES facing the head region of early (Step 8) elongating spermatids as compared to vehicle (dimethylsulfoxide:saline) controls. Nitrobenzoxadiazole-phallacidin staining of F-actin revealed a CD-related loss of uniform fluorescence over the head of elongated spermatids. To examine for a possible relationship between the presence of actin and cell attachment at ES sites, hypertonic fixatives were introduced to provoke cell shrinkage and stress ES-associated junctions. After osmotic stress, cell-to-cell adhesion at ES sites remained intact in vehicle-treated animals. CD treatment caused Sertoli cells to separate from elongating spermatids at sites where ES had been lost from the Sertoli cell surface. It is suggested that actin of the ES plays a role in cell-to-cell interaction analogous to its possible role at the Sertoli cell barrier. In CD-treated animals, structures resembling tubulobulbar complexes frequently developed at sites where ES was lost, suggesting that the loss of ES has a facilitatory role in tubulobulbar complex formation. It is hypothesized that tubulobulbar complexes are devices that rid the cells of ES-associated junctional links to effect dissociation of the spermatid from the Sertoli cell during spermiation. Spermatids at Step 8 of development are known to become oriented with their acrosomes facing the base of the Sertoli cell. After CD treatment, a 5.8-fold increase in malorientation of Step 8 spermatids was noted. A role for the ES cytoskeletal complex in orienting the spermatid acrosome toward the basal aspect of the Sertoli cell is also suggested.     

Comparative Chemical and Bioactivity Studies of Intra- and Extracellular Metabolites of Endophytic Bacteria,Bacillus subtilis NCIB 3610

International Journal of Peptide Research and Therapeutics - Endophytic bacteria are able to produce unique bioactive compounds for various biotechnological applications. The intracellular and...     

Towards using of new and safety material against tomato leafminer,Tuta absoluta (Mayrick)

New silica formulations offer expanded possibilities for use in horticultural crops. However, many crop pests are found on the leaf underside, and this is especially challenging when using silica because the substance must have direct contact with the insect to be effective. Silica a new and safety material was used against the second instar larvae of tomato leafminer, Tuta absoluta (Mayrick) alone and combined with other pesticides (chlorfenapyr, spinetoram and chlorantraniliprole). The results showed that when silica was used alone, the percents of mortalities were 51.7, 26.7 and 13.3%. The results showed also chlorantraniliprole was the most toxic compared with the other compounds. The per cents of mortalities were 90, 65 and 30% with the first, second and third concentrations, respectively. When silica was used in a combination with the tested pesticides, the mixture of silica and chlorfenapyr was the most toxic against the second instar larvae of T. absoluta, the percents of mortalities were 100, 76.7 and 60% with the first, second and third concentrations, respectively. The results recommended that silica was moderate toxic to the second instar larvae of T. absoluta when it used alone, so it can be used when this pest under the economic threshold or with integrated pest management programme. Silica has an activation action on the tested pesticides. Although silica was combined with one-fifth of the field rate of the tested pesticides, the toxicity of these pesticides was increased.     

Regulation of fatty acid synthetase activity. The 4'-phosphopantetheine hydrolase of rat liver.

The 4'-phosphopantetheine hydrolase of rat liver, partially purified by ammonium sulfate precipitation, catalyzes the hydrolysis of the prosthetic group 4'-phosphopantetheine from the holo-fatty acid synthetase. The two products of the action of this enzyme, 4'-phosphopantetheine and apo-fatty acid synthetase, were isolated by DEAE-cellulose chromatography and by chromatography on a Sepharose epsilon-aminocaproyl pantetheine column, respectively. The resultant apo-fatty acid synthetase was quantitated by immunoprecipitation and it was also converted to the holoprotein with a crude preparation of rat liver 4'-phosphopantetheine transferase. Quantitative determination of the hydrolase reaction product, 4'-phosphopantetheine, by amino acid analysis and microbiological assays confirmed the presence of 1 mol of this compound/mol of holo-fatty acid synthetase.     

ATP enhances cyclic AMP accumulation by intact P388 mouse lymphoma cells

One of the characteristics of malignant cells is a poor response to hormones and a low level of cyclic AMP. Whilst this is true of intact P388 mouse lymphoma cells, high levels of adenylate cyclase (ATP pyrophosphate-lyase (cyclizing), EC 4.6.1.1) activity can be measured in particulate preparations of these cells. When ATP is added to the incubation medium of intact lymphoma cells, the cyclic AMP level is enhanced. This effect of ATP is not mediated by adenosine, nor is it enhanced by NaF. The ATP content of the lymphoma cells is much lower than that of CH23 Chinese hamster fibroblast and PCM3 hybrid cells, whose cyclic AMP levels are not affected by the presence of ATP. This suggests that adenylate cyclase, in the lymphoma cells, is bathed in a pool which is deficient in substrate. The substrate concentration of this pool is thought to be elevated by addition of ATP to the incubation medium with ATP, itself, crossing the plasma membrane.     

Pathotypic and molecular evolution of contemporary population of Puccinia striiformis f. sp. tritici in Egypt during 2016–2018

The contemporary races of Puccinia striiformis f. sp. tritici (Pst) in Egypt during 2016–2018 were differentiated based on virulence and molecular patterns. Virulence patterns based on the reaction of the 17 World/European differential sets carrying stripe rust resistance genes (Yr genes) resulted in ten races including four new (first recorded in Egypt) and six old (previously recorded in Egypt). The new races were identified as 64E0 (virulence [V] Yr4, Su), 0E16 (V Yr8, 19), 66E0 (V Yr4, 7, 22, 23, Su) and 4E130 (V Yr2, 6, 7, 25, HVII), while the old were 0E0 (avirulence), 2E0 (V Yr7, 22, 23), 2E16 (V Yr7, 8, 19, 22, 23), 4E0 (V Yr2, 6), 6E4 (V Yr2, 6, 7, 22, 23, 25) and 70E4 (V Yr2, 4, 6, 7, 22, 23, 25, Su). Cluster analysis differentiated Pst races based on virulence frequency to Yr genes. Simple sequence repeat (SSR) markers were used to detect the molecular polymorphism of the Pst races. Clustering separated the old and new races into two groups, indicating their common ancestry since the new races were very distinct from the old races. Although clustering based on virulence revealed some evolutionary patterns, where the new races 64E0 and 66E0 may have probably evolved from the old races (2E16, 2E0, 6E4, 70E4) and the new race 4E130 may be evolved from the joint race 4E0. However, clustering based on molecular patterns indicated that the new races appear to be genetically distinct and may represent an exotic introduction rather than a mutation in isolates of the old races. A weak association between virulence and molecular patterns revealed that they are independent of each other. The SSR markers did not correspond to the virulences in the pathogen. Further studies on the potential virulence genes of the detected Pst virulences are needed.     

Biotin attenuation of oxidative stress,mitochondrial dysfunction,lipid metabolism alteration and 7β-hydroxycholesterol-induced cell death in 158N murine oligodendrocytes

Mitochondrial dysfunction and oxidative stress are involved in neurodegenerative diseases associated with an enhancement of lipid peroxidation products such as 7β-hydroxycholesterol (7β-OHC). It is, therefore, important to study the ability of 7β-OHC to trigger mitochondrial defects, oxidative stress, metabolic dysfunctions and cell death, which are hallmarks of neurodegeneration, and to identify cytoprotective molecules. The effects of biotin were evaluated on 158N murine oligodendrocytes, which are myelin synthesizing cells, exposed to 7β-OHC (50?µM) with or without biotin (10 and 100?nM) or α-tocopherol (positive control of cytoprotection). The effects of biotin on 7β-OHC activities were determined using different criteria: cell adhesion; plasma membrane integrity; redox status. The impact on mitochondria was characterized by the measurement of transmembrane mitochondrial potential (ΔΨm), reactive oxygen species (ROS) overproduction, mitochondrial mass, quantification of cardiolipins and organic acids. Sterols and fatty acids were also quantified. Cell death (apoptosis, autophagy) was characterized by the enumeration of apoptotic cells, caspase-3 activation, identification of autophagic vesicles, and activation of LC3-I into LC3-II. Biotin attenuates 7β-OHC-induced cytotoxicity: loss of cell adhesion was reduced; antioxidant activities were normalized. ROS overproduction, protein and lipid oxidation products were decreased. Biotin partially restores mitochondrial functions: attenuation of the loss of ΔΨm; reduced levels of mitochondrial O₂^?? overproduction; normalization of cardiolipins and organic acid levels. Biotin also normalizes cholesterol and fatty acid synthesis, and prevents apoptosis and autophagy (oxiapoptophagy). Our data support that biotin, which prevents oligodendrocytes damages, could be useful in the treatment of neurodegeneration and demyelination.