Identification of a cDNA encoding a long form of prolactin receptor in human hepatoma and breast cancer cells

Human PRL receptor cDNA clones from hepatoma (Hep G2) and breast cancer (T-47D) libraries were isolated by using a rat PRL receptor cDNA probe. The nucleotide sequence predicts a mature protein of 598 amino acids with a much longer cytoplasmic domain than the rat liver PRL receptor. Although this extended region has additional segments of localized sequence identity with the human GH receptor, there is no identity with any consensus sequences known to be involved in hormonal signal transduction. This cDNA will be a valuable tool to better understand the role of PRL in the development and growth of human breast cancer.     

Stimulation of prostaglandin synthesis in cultured rat synovial cells by a factor derived from polymorphonuclear leukocytes

Stimulation of synovial cell prostaglandin production by a factor obtained from casein-induced peritoneal polymorphonuclear (PMN) cells has been investigated. Both the extract and short time cultured medium of rat peritoneal PMN cells stimulate prostaglandin (PG)E2 production as well as collagenase production in the culture of rat synovial cells. PGE2 production by the cells in the presence of the PMN factor is much faster (5 to 24 hr) than collagenase production (24 hr or later, Biomedical Res. 3, 506-516, 1982). This stimulating factor is confirmed to be derived from PMN cells, based on the purification of the cells from peritoneal exudate cells by the Ficoll-Urographin method. Elution profile of the factor on gel filtration has indicated that both PGE2 and collagenase productions by synovial cells are stimulated by the same effluent fractions corresponding to molecular weights of 15,000 - 20,000 daltons and 30,000 - 40,000 daltons. These results suggest that PMN cells are involved in PG production as well as collagenase production in the inflamed tissue by stimulating connective tissue cells such as synovial cells.     

The Prevalence and Characteristics of Metabolic Syndrome in Patients with Vertigo

Objectives/Hypothesis

Metabolic syndrome (MetS) is a condition that increases the risk of coronary artery disease and cerebral infarction. We determined the prevalence of MetS in vertigo patients and clinically investigated the association between MetS and vertigo.

Study Design

Case-control study

Methods

The subjects were 333 patients, including 107 males and 226 females, who presented with vertigo as a primary symptom. MetS was diagnosed according to the International Diabetes Federation definition, which is based on waist circumference, blood serum levels, and blood pressure.

Results

MetS was detected in 53 (15.9%) of 333 vertigo patients, including 24 males (22.4%) and 29 females (12.8%); i.e., the frequency of MetS was significantly higher among the male patients than the female patients. The overall prevalence of MetS (15.9%) among vertigo patients did not differ from that observed among general adults in previous Japanese surveillance studies; however, MetS was significantly more common among the vertigo patients in males than general adult males. The prevalence of MetS was also examined in five types of vertigo, Concomitant MetS was noted in many males with vertebrobasilar insufficiency (VBI) and isolated vertigo of unknown etiology.

Conclusion

It was suggested that MetS is involved in the development of vertigo in males. MetS might be a risk factor for vascular vertigo such as VBI in males. The high frequency of MetS among males with vertigo of unknown etiology suggested that the pathogenesis of metabolic syndrome is involved in this type of isolated vertigo.     

Ion Concentration- and Voltage-Dependent Push and Pull Mechanisms of Potassium Channel Ion Conduction

The mechanism of ion conduction by potassium channels is one of the central issues in physiology. In particular, it is still unclear how the ion concentration and the membrane voltage drive ion conduction. We have investigated the dynamics of the ion conduction processes in the Kv1.2 pore domain, by molecular dynamics (MD) simulations with several different voltages and ion concentrations. By focusing on the detailed ion movements through the pore including selectivity filter (SF) and cavity, we found two major conduction mechanisms, called the III-IV-III and III-II-III mechanisms, and the balance between the ion concentration and the voltage determines the mechanism preference. In the III-IV-III mechanism, the outermost ion in the pore is pushed out by a new ion coming from the intracellular fluid, and four-ion states were transiently observed. In the III-II-III mechanism, the outermost ion is pulled out first, without pushing by incoming ions. Increases in the ion concentration and voltage accelerated ion conductions, but their mechanisms were different. The increase in the ion concentrations facilitated the III-IV-III conductions, while the higher voltages increased the III-II-III conductions, indicating that the pore domain of potassium channels permeates ions by using two different driving forces: a push by intracellular ions and a pull by voltage.     

A protein recycling system for nuclear magnetic resonance-based screening of drug candidates

A sample-treating system for nuclear magnetic resonance (NMR)-based interaction screening between drug candidates (small molecules) and a protein of interest was developed by applying high-performance liquid chromatography (HPLC) technology. The system prepares a test solution by mixing a (15)N-labeled protein solution and a solution of each candidate compound, loads it to a flow cell-type NMR probe, and recycles the protein after the data acquisition. The system was designed to behave differently according to the information obtained in NMR measurements. In a test operation with a 100-compound library, the system could single out known interacting substances properly. Recovery values of the protein and one representative compound were 75 and 71%, respectively, and the recovered protein was found intact as intended.     

Hepatitis C virus (HCV) NS5A binds RNA-dependent RNA polymerase (RdRP) NS5B and modulates RNA-dependent RNA polymerase activity.

Hepatitis C virus (HCV) NS5B is RNA-dependent RNA polymerase (RdRP), the essential catalytic enzyme for HCV replication. Recently, NS5A has been reported to be important for the establishment of HCV replication in vitro by the adaptive mutations, although its role in viral replication remains uncertain. Here we report that purified bacterial recombinant NS5A and NS5B directly interact with each other in vitro, detected by glutathione S-transferase (GST) pull-down assay. Furthermore, complex formation of these proteins transiently coexpressed in mammalian cells was detected by coprecipitation. Using terminally and internally truncated NS5A, two discontinuous regions of NS5A (amino acids 105-162 and 277-334) outside of the adaptive mutations were identified to be independently essential for the binding both in vivo and in vitro (Yamashita, T., Kaneko, S., Shirota, Y., Qin, W., Nomura, T., Kobayashi, K., and Mkyrakami, S. (1998) J. Biol. Chem. 273, 15479-15486). We previously examined the effect of His-NS5A on RdRP activity of the soluble recombinant NS5Bt in vitro (see Yamashita et al. above). Wild NS5A weakly stimulated at first (when less than 0.1 molar ratio to NS5B) and then inhibited the NS5Bt RdRP activity in a dose-dependent manner. The internal deletion mutants defective in NS5B binding exhibited no inhibitory effect, indicating that the NS5B binding is necessary for the inhibition. Taken together, our results support the idea that NS5A modulates HCV replication as a component of replication complex.     

Novel roles of CpG oligodeoxynucleotides as a leader for the sampling and presentation of CpG-tagged antigen by dendritic cells.

Oligodeoxynucleotides containing CpG motifs have been highlighted as potent Th1 activators. We previously reported that Ag and CpG, when conjugated together, synergistically promoted the Ag-specific Th1 development and inhibited the Th2-mediated airway eosinophilia. In this study, we examined the mechanisms underlying the synergism of the covalent conjugation. The CpG-OVA conjugate enhanced the Th1 activation and development. These characteristic features of the conjugate could not be ascribed to the polymerization of OVA, but mirrored the augmented binding of the CpG-tagged Ag to dendritic cells (DCs) in a CpG-guided manner, because phycobiliprotein, R-PE, conjugated to CpG stained a higher proportion of DCs with higher intensity than the mixture. R-PE fluorescence was emitted from cytoplasmic portions of the DCs, which simultaneously expressed costimulatory molecules and IL-12. The CpG-conjugated R-PE trafficking described above actually served as a potent Ag. These results indicate that CpG conjugated to Ag exhibit novel joint properties as promoters of Ag uptake and DC activators, thereby potentiating the ability of DCs to generate Th1 cells. The DNA-mediated promotion of Ag uptake would be advantageous for evoking host immune responses against invading microorganisms.