Differences in lectin binding patterns of normal human endometrium between proliferative and secretory phases

Lectin binding patterns in normal human endometrium were examined by light and electron microscopy using seven different lectins (ConA, WGA, RCA, PNA, UEA-1, DBA, and SBA). For light microscopic observations, criteria based on the incidence and intensity of cells positive for the lectin staining were adopted to evaluate the different staining patterns of the proliferative and secretory endometria obtained by the avidin-biotin-peroxidase complex (ABC) technique. At the light microscopic level, ConA, WGA, and RCA stained endometrial glandular cells in both phases. The number of PNA-positive cells with the binding sites entirely limited to the apical surface tended to be reduced slightly in the secretory phase. UEA-1 weakly stained the apical surface of glandular cells in the proliferative phase but not in the secretory phase. Among the lectins used in this study, DBA and SBA displayed remarkable changes between the phases. That is, in the proliferative phase they produced only a faint or slight positive stain at the apical surface, but the incidence and intensity of DBA- and the SBA-positive glandular cells increased in the secretory phase. By electron microscopy, the reaction product of ConA was observed in the plasma membrane, endoplasmic reticulum, nuclear envelope, and the Golgi apparatus, and the binding sites of RCA and DBA were observed in the plasma and Golgi membranes. Between both phases, the reactivity of ConA and RCA showed almost no change. However, the secretory endometrial cells containing the DBA-positive Golgi apparatus were markedly increased in number compared with the proliferative ones bearing the lectin-positive organelles.(ABSTRACT TRUNCATED AT 250 WORDS)     

Aberrant Glycogen Synthase Kinase 3β Is Involved in Pancreatic Cancer Cell Invasion and Resistance to Therapy

Background and Purpose

The major obstacles to treatment of pancreatic cancer are the highly invasive capacity and resistance to chemo- and radiotherapy. Glycogen synthase kinase 3β (GSK3β) regulates multiple cellular pathways and is implicated in various diseases including cancer. Here we investigate a pathological role for GSK3β in the invasive and treatment resistant phenotype of pancreatic cancer.

Methods

Pancreatic cancer cells were examined for GSK3β expression, phosphorylation and activity using Western blotting and in vitro kinase assay. The effects of GSK3β inhibition on cancer cell survival, proliferation, invasive ability and susceptibility to gemcitabine and radiation were examined following treatment with a pharmacological inhibitor or by RNA interference. Effects of GSK3β inhibition on cancer cell xenografts were also examined.

Results

Pancreatic cancer cells showed higher expression and activity of GSK3β than non-neoplastic cells, which were associated with changes in its differential phosphorylation. Inhibition of GSK3β significantly reduced the proliferation and survival of cancer cells, sensitized them to gemcitabine and ionizing radiation, and attenuated their migration and invasion. These effects were associated with decreases in cyclin D1 expression and Rb phosphorylation. Inhibition of GSK3β also altered the subcellular localization of Rac1 and F-actin and the cellular microarchitecture, including lamellipodia. Coincident with these changes were the reduced secretion of matrix metalloproteinase-2 (MMP-2) and decreased phosphorylation of focal adhesion kinase (FAK). The effects of GSK3β inhibition on tumor invasion, susceptibility to gemcitabine, MMP-2 expression and FAK phosphorylation were observed in tumor xenografts.

Conclusion

The targeting of GSK3β represents an effective strategy to overcome the dual challenges of invasiveness and treatment resistance in pancreatic cancer.     

Activation of protease by sol-gel entrapment into organically modified hybrid silicates

To prepare an immobilized protease with a high activity for transesterification of vinyl n-butyrate with 3-methyl-1-butanol (isoamyl alcohol) in organic media, a protease was entrapped into organic–inorganic hybrid silica gel on Celite 545 by the sol-gel method. When propyltrimethoxysilane was used as the organic silane precursor mixed with tetramethoxysilane at a molar ratio of 16:1, the hybrid gel-entrapped protease on Celite 545 had 8 times the activity of the protease deposited on Celite 545 from 35 to 85°C.     

Particle inflow gun-mediated transformation of multiple-shoot clumps in rhodes grass (Chloris gayana)

Rhodes grass (Chloris gayana) is one of the most important warm-season forage grasses. It is cultivated in tropical and subtropical parts of the world and is mostly used for grazing and hay production. We have established a particle-bombardment transformation protocol for rhodes grass using multiple-shoot clumps (MSCs) as the target tissue. A vector pAHC25 containing a herbicide-resistance gene (bar) together with the beta-glucuronidase (GUS) gene was used in transformation experiments. The most efficient recovery of bialaphos-resistant tissue was achieved when the bombarded MSCs were first cultured for 15 d on bialaphos-free medium before being subjected to selection pressure. The resistant tissues regenerated transgenic plants that displayed GUS gene expression. Under optimized conditions, 251 target pieces yielded 46 transgenic plants from 4 independent transgenic lines.     

Variation in growth performance of Ryukyu-ayu, <Emphasis Type="Italic">Plecoglossus altivelis ryukyuensis</Emphasis>, inferred from otolith analysis

Ryukyu-ayu (Plecoglossus altivelis ryukyuensis) is an amphidromous fish species that migrates between the sea and rivers over its one-year life span. Although growth performance during the early marine stage may affect growth in the later riverine stage of this species’ life cycle, no studies have specifically examined this relationship in P. a. ryukyuensis. In the present study, we reconstructed the growth trajectories of P. a. ryukyuensis individuals collected from the Yakugachi River, Amami-Oshima Island, Japan in 2016 (n?=?47) throughout their growth period in both the sea and river by using otolith analysis. Using this, we determined the age and body size of individuals at the time of their upstream migration, as well as their growth rates during the marine and riverine stages. Results showed that body size at upstream migration significantly affected body size at the riverine stage, indicating that juveniles with larger body size in the sea had better growth performance in the river. Individuals with higher growth rates during the marine stage tended to enter the river younger and at larger body sizes than those with lower marine growth rates. Our results demonstrated the close linkage between the growth performance in the sea and in rivers of P. a. ryukyuensis. This information will contribute to better understanding variations in growth patterns of this endangered species and potentially aid in its conservation.     

Maintenance of meiotic arrest and developmental potential of porcine oocytes after parthenogenetic activation and somatic cell nuclear transfer

Several studies report that meiotic maturation of porcine oocytes can be reversibly preserved. The present study examined how long meiotic maturation can be suppressed. The first experiment determined the preservation medium suitable for reversibly suppressing meiotic maturation of porcine oocytes. The second experiment examined the in vitro developmental potential of oocytes maintained in meiotic arrest after parthenogenetic activation and nuclear transfer of somatic cells. Preservation of cumulus-oocyte complexes with NCSU-37 medium containing 10% follicular fluid, 1 mM dibutyryl cyclic AMP, and follicular shell pieces for 24-96 h at 39 degrees C did not affect oocyte maturation compared with controls (94-98% vs. 98%). The potential of parthenogenetically activated and nuclear-transferred oocytes maintained in meiotic arrest for 24-48 h to develop into blastocysts was not significantly different from that of controls (20-25% vs. 18% and 8-11% vs. 9%, respectively). The present study demonstrated that meiotic maturation of porcine oocytes can be suppressed after preservation for 48 h at 39 degrees C without decreasing oocyte maturation competence or the ability of oocytes to develop to at least the blastocyst stage.     

Purification of immunoglobulin heavy chain messenger RNA by immunoprecipitation from the mouse myeloma tumor, MOPC-31C.

Immunoglobulin heavy chain mRNA was purified from immunoprecipitated polysomes derived from the mouse myeloma tumor, MOPC-31C. The purified mRNA migrated predominantly as a single band upon polyacrylamide gel electrophoresis in 98% formamide and the molecular weight of this mRNA was calculated to be 700,000. This mRNA was as active as the purified light chain mRNA when it was employed as a template in a cell-free protein synthesizing system from wheat germ. The translation product had a molecular weight of 55,000 daltons, and migrated slightly faster than mature heavy chain upon polyacrylamide gel electrophoresis in sodium dodecylsulfate. The protein synthesized by the direction of this mRNA was shown to yield tryptic peptides corresponding to those derived from the mature heavy chain protein except that one missing peptide was replaced by another additional peptide. DNA complementary to the mRNA was synthesized by RNA-dependent DNA polymerase from avian myeloblastosis virus. Hybridization kinetic analysis between the heavy chain mRNA and its complementary DNA indicated that the RNA was essentially homogenous with rabbit globin mRNA as a standard.