Intrabursal administration of protein kinase or proteinase inhibitors: effects on ovulation in the rat.

OBJECTIVE: To test the hypothesis that inhibition of protein kinase (PK) activity or proteolysis inhibits ovulation. DESIGN: Rats were injected intrabursally with protein kinase (H9 or staurosporin) or proteinase (alpha 2-macroglobulin) inhibitors and oocyte release was evaluated. SETTING: Clinical Research Laboratory, Center for Reproductive Medicine, University of Kentucky Medical Center. PARTICIPANTS: Immature rats stimulated with pregnant mare serum gonadotropin. INTERVENTIONS: Staurosporin (1 or 10 microM), H9 (1 mM), alpha 2-macroglobulin (835 microIU of activity); or vehicle was injected into the right ovarian bursa. The left ovarian bursa remained intact. Animals immediately received human chorionic gonadotropin (hCG). MAIN OUTCOME MEASURES: Analysis of oocyte release and ovarian morphology. RESULTS: Oocyte release from the inhibitor-treated side decreased for the H9 group (8.1 +/- 1.9 fewer oocytes released, P less than 0.001) and the 10 microM staurosporin group (5.5 +/- 0.6, P less than 0.001). No change in oocyte release was observed in the 1 microM staurosporin, alpha 2-macroglobulin, or vehicle injected groups. Histologic examination of vehicle treated ovaries demonstrated numerous developing corpora lutea (CL; 20.5 +/- 2.1 CL/ovary) and a lack of preovulatory follicles. In contrast, ovaries treated with PK inhibitors contained unruptured preovulatory follicles coincident with fewer forming CL (11.5 +/- 3.5 CL/ovary). CONCLUSIONS: Inhibition of PK activity in vivo suppresses ovulation, demonstrating that protein phosphorylation plays an important intermediary role in the ovulatory process.     

Parasitism by Virulent Treponema pallidum of Host Cell Surfaces

The interaction between virulent Treponema pallidum extracted from infected rabbit testes and animal cells in culture was examined. The extent of treponemal attachment to monolayers of normal rabbit testicular and HEp-2 cells was dependent upon the incubation temperature and retained motility of the spirochetes. The specific orientation of treponemes to host cell surfaces was demonstrated by dark-field microscopic examination of wet-mount preparations and scanning and transmission electron microscopy. Once attached, T. pallidum organisms remained actively motile yet anchored in place by their terminal tapered structures. After several hours of co-incubation, maximal attachment was attained, and the degree of parasitism seemed regulated not only by available surface sites on individual host cells but also by the proposed membrane response of parasitized cells to continued exposure to treponemes. The avirulent strain, Treponema phagedenis biotype Reiter, did not adhere to monolayer cultures. Characterization of host cell determinants that permitted surface colonization by T. pallidum was attempted. Also, properties of virulent treponemes that enabled surface parasitism were monitored by measuring the effects of enzymes, detergents, and metabolic inhibitors on the host-parasite interaction. Results reinforced the specific nature of the treponemal attachment mechanism. Furthermore, the ability of convalescent rabbit sera to reduce attachment of treponemes to host cells suggested that surface structures on T. pallidum could be masked or inactivated by host components, thus providing a potentially effective research approach for investigating the pathogenesis of syphilis and screening appropriate vaccine candidates.     

Development of an immuno-lectin-enzymatic assay for the detection of serum cancer-associated glycoproteins bearing Tn determinant

We report the development of an immuno-lectin-enzymatic assay (CA83.4) with the purpose of quantifying serum glycoproteins bearing Tn determinant (GalNAc alpha-O-Ser/Thr). An anti-Tn monoclonal antibody (83D4) is bound to the solid phase in order to capture glycoproteins. After the addition of a test sample, we used biotinylated isolectin B4 from Vicia villosa and avidin-peroxydase to act as a detection system. The linear relationship between CA83.4 determinations and the serum dilutions, the reproducibility of the dosage in intra- and inter-assay, and the specificity of the test for the N-acetylgalactosamine residue in a-glycosidic O-linkages, demonstrated the reliability of this trial. Self-recognition of Vicia villosa B4 molecules (K-D: 0.73x10(-6) M determined using biosensor technology) could determine an additional step of signal amplification in this assay. Using 0.25 units/ml of CA83.4 antigen as the cut-off level, higher values were found in 25/49 patients with breast cancer, 8/13 with colorectal carcinoma, 3/11 with lung cancer, but in none of the 49 patients with non-malignant diseases nor in 97 healthy controls. This first report on soluble Tn-glycoprotein detection assays suggests that Tn-glycoproteins are specific serological tumor markers and we believe that they could represent a valuable tool in the diagnosis of cancer.     

Randomized,double-blind,placebo-controlled,safety and immunogenicity study of 4 formulations of Anthrax Vaccine Adsorbed plus CPG 7909 (AV7909) in healthy adult volunteers

A new anthrax vaccine that could accelerate the immune response and possibly reduce the number of injections needed for protection would be desirable in a post-exposure setting.     

One-Year Follow-up of Duodenal Ulcers after 1-Wk Triple Therapy for Helicobacter pylori

Objective : to study the ulcer recurrence rate of Helicobacter pylori-positive duodenal ulcers at 1 yr after eradication of the bacteria by triple therapy. Method : Patients with H. pylori-positive duodenal ulcers were randomized to receive either triple therapy for 1 wk plus omeprazole for 4 wk (THple+OMP) (n = 78), or omeprazole alone (OMP) for 4 wk (N = 77). Patients were followed up every 3 months for symptom enquiry. At 1 yr, all asymptomatic patients were invited to attend for gastroscopy. Results : At 8 wk, 16 patients in the OMP group and four in the Triple+OMP group had an ulcer. During the 1-yr period, 12 patients in the OMP group and no patient in the Triple+OMP group developed symptomatic ulcers. At follow-up endoscopy at 1 yr, another 10 ulcers were detected in the OMP group and two in the Triple+OMP group. Fifteen patients in the OMP group and 13 in the Triple+OMP group were lost to follow-up. In total, ulcers were de-tected in 39 of 61 (64%) assessahle patients in the OMP group, and in six of 65 (97o) assessahle patients in the Triple+OMP group after I yr (χ² test: p < 0.001). Of the patients whose H, pytori were successfully eradicated hy Triple+OMP at 8 wk, 90% remained H. pylori negative at 1 yr. Conclusion : Triple therapy for 1 wk eradicates H, pylori infection and significantly reduces duodenal ulcer relapses.