Raised serum chondroitin sulfate epitope level in ovarian epithelial cancer

OBJECTIVE: To determine the value of serum chondroitin sulfate epitope WF6 and hyaluronan (HA) levels as a biomarker for early detection of ovarian epithelial cancer and other gynecological disorders. METHOD: Serum WF6 CS epitope and HA were measured in 91 patients with ovarian epithelial cancer, 39 patients with non-cancer gynecological disorders and 30 healthy women. Serum chondroitin sulfate (CS) WF6 epitope was determined by a competitive immunoassay with the monoclonal antibodies WF6, which specifically recognizes an epitope in native CS chains. In addition, serum HA concentration was measured by an ELISA-based assay with a biotinylated affinity HA-binding proteins. RESULTS: The serum concentration of CS (WF6) epitope was highly increased in epithelial types of ovarian cancer and at all stages of development (p < 0.005). Serum HA in ovarian cancer patients was significantly higher than normal controls (p < 0.05). CONCLUSION: These results reflect changes in ECM metabolism in progressive ovarian cancer, which cause an increase in serum CS epitopes and HA. Therefore, serum CS epitopes may provide useful biomarkers for cancers and other disorders of the ovary. Measurement of serum HA provided complementary information, which may be useful as a discriminator between benign ovarian disorders and malignant ovarian diseases.     

Karyotypic study of five Lutjanid species using conventional and Ag-NORs banding techniques

The first cytogenetic comparisons of five snapper species from Thailand were presented here. Renal cell samples were taken from blacktail snapper (Lutjanus fulvus), five lined snapper (L. quinquelineatus), dory snapper (L. fulviflamma), brownstripe red snapper (L. vitta), and mangrove red snapper (L. argentimaculatus). The mitotic chromosome preparation was prepared directly from kidney cells. Conventional staining and Ag-NOR banding techniques were applied to stain the chromosomes. The results exhibited that all five snapper species have the diploid chromosome numbers of 2n = 48 and the fundamental numbers (NF) of 48. The presences of large, medium, and small telocentric chromosomes were 22-24-2, 24-20-4, 36-10-2, 28-16-4 and 36-10-2, respectively. The Ag- NORs banding technique provides the pair of nucleolar organizer regions (NORs) at subcentromeric region of the long arm of the respective telocentric chromosome pairs 9, 1, 3, 4 and 9. Their karyotype formulas is as follows: L. fulvus (2n = 48): L ₂₂ ^t + M ₂₄ ^t + S ₂ ^t , L. quinquelineatus (2n = 48): L ₂₄ ^t + M ₂₀ ^t + S ₄ ^t , L. fulviflamma (2n = 48): Lt36 + Mt10 + St2, L. vitta (2n = 48): L ₂₈ ^t + M ₁₆ ^t + S ₄ ^t , and L. argentimaculatus (2n = 48): L ₃₆ ^t + M ₁₀ ^t + S ₂ ^t .     

Multiple bandshift assay: rapid identification and cloning of DNA fragments containing specific protein-binding sites

A new rapid method for the identification and cloning of DNA fragments containing specific protein-binding domains is based on the common bandshift assay. Cloned DNA is digested with a restriction endonuclease recognizing a particular 4-bp sequence, an aliquot of this digest is end-labelled and used in protein binding reactions with and without protein extract. The binding reactions are then loaded onto nondenaturing polyacrylamide gel. The main portion of the digest is run in a parallel lane and serves as a source of fragments for cloning. Autoradiography of the wet gel reveals loss in intensity of some bands from the restriction digest incubated with the protein extract. DNA fragments corresponding to these bands are cut out from the gel; DNA is eluted and cloned in the M13 vector, thus allowing rapid and simple sequencing of the inserts.

The method, terned multiple bandshift assay, is especially useful when screening relatively long DNA fragments (of several kb) for potential protein-binding domains. The procedure was used to study interaction of HeLa-cell nuclear proteins with a 5.2-kb downstream region of pseudorabies virus immediate-early gene ( . Vl ek, Z. Kozmik, V. Pa es, S. Schirm and M. Schwyzer, unpublished).     

Role of a <Emphasis Type="Italic">Burkholderia pseudomallei</Emphasis> polyphosphate kinase in an oxidative stress response,motilities, and biofilm formation

Burkholderia pseudomallei, a motile and rod Gram-negative bacterium, is the causative agent of melioidosis. The bacterium is an intracellular pathogen and that motility is generally crucial for their survival in a natural environment and for systemic infection inside a host. We report here a role of B. pseudomallei polyphosphate kinase in virulence, such as an oxidative stress response, motilities and biofilm formation. The polyphosphate kinase (ppk) mutant is susceptible to hydrogen peroxide in an oxidative stress condition, unable to perform swimming, swarming motilities, and has lower density biofilm forming capacity than the wild-type strain. We also demonstrated that both polyphosphate kinase and motile flagella are essential and independently involved in biofilm formation. The B. pseudomallei flagellin (fliC) mutant and B. mallei, a nonmotile species, are shown to produce higher density biofilm formation than the ppk mutant, but less than wild type B. pseudomallei.