Inhibition of hormone secretion in GH-secreting pituitary adenomas by receptor-subtype specific somatostatin analogues in vitro

The aim of the study was to determine the inhibitory effects of somatostatin analogues with relative specificity to somatostatin receptor subtype 2 (SSTR2) (BIM-23197), subtype 5 (SSTR5) (BIM-23268), and their combination on GH and PRL secretion in acromegalic adenomas in vitro. Three types of answer were observed: 1. In one resistant adenoma no inhibition was achieved. 2. The GH secretion in six adenomas was suppressed significantly more (p < 0.01 or p < 0.001 using Mann-Whitney U-test in concentration range of 10(-12) to 10(-8) mol/l) with SSTR2 specific analogue BIM-23197 with no additive effect of compounds combination. 3. In three adenomas the potency of BIM-23197 and BIM-23268 was almost equal and the combination of these SSTR2 and SSTR5 specific compounds had statistically significant additive effect (p < 0.05 or p < 0.01 in concentration range of 10(-12) to 10(-8) mol/l). PRL secretion of five adenomas was more suppressed with SSTR5 specific BIM-23268 (statistically significant in concentrations 10(-10) to 10(-8) mol/l). In conclusion the somatostatin analogue BIM-23268 had an additive effect on suppression of GH secretion in a subset of adenomas, where both SSTR2 and SSTR5 were involved. This effect was not observed in the majority of tumours, where the inhibitory effect seems to be mediated via SSTR2 only.     

A heterotrimeric PCNA in the hyperthermophilic archaeon Sulfolobus solfataricus

The sliding clamp, PCNA, of the archaeon Sulfolobus solfataricus P2 is a heterotrimer of three distinct subunits (PCNA1, 2, and 3) that assembles in a defined manner. The PCNA heterotrimer, but not individual subunits, stimulates the activities of the DNA polymerase, DNA ligase I, and the flap endonuclease (FEN1) of S. solfataricus. Distinct PCNA subunits contact DNA polymerase, DNA ligase, or FEN1, imposing a defined architecture at the lagging strand fork and suggesting the existence of a preformed scanning complex at the fork. This provides a mechanism to tightly couple DNA synthesis and Okazaki fragment maturation. Additionally, unique subunit-specific interactions between components of the clamp loader, RFC, suggest a model for clamp loading of PCNA.     

Pollen morphology of Carthamus L. species in Anatolian flora

In flora of Turkey, there are six species of Carthamus L: Carthamus dentatus (Forssk.) Vahl., C. glaucus M. Bieb. subsp. glaucus, C. lanatus L., C. tenuis (Boiss. & Blanche) Bornm., C. persicus Desf. ex Willd. and C. tinctorius L. Within these species, C. tinctorius L. is an alien species for the Turkish flora. The pollen grains belonging to six taxa collected from different locations of Anatolia and examined both under light microscope (LM) and under scanning electron microscope (SEM). As a result of these examinations and measurements, pollen grains of taxa are radially symmetrical, isopolar, oblate-spheroid, spheroid, tricolporate rarely tetracolporate, echinate. There are high similarities among taxa but some differences are recorded in size and spin length. According to cluster analyses of Carthamus species’ pollens, C. glaucus and C. tenuis have the most similar pollens. C. dentatus grouped with these two. C. persicus placed as the closest relative of C. tinctorius in the dendrogram.     

Individual genetic variations directly effect polarization of cytokine responses to superantigens associated with streptococcal sepsis: implications for customized patient care

Host immunogenetic variations strongly influence the severity of group A streptococcus sepsis by modulating responses to streptococcal superantigens (Strep-SAgs). Although HLA-II-DR15/DQ6 alleles strongly protect against severe sepsis, HLA-II-DR14/DR7/DQ5 alleles significantly increase the risk for toxic shock syndrome. We found that, regardless of individual variations in TCR-Vβ repertoires, the presentation of Strep-SAgs by the protective HLA-II-DR15/DQ6 alleles significantly attenuated proliferative responses to Strep-SAgs, whereas their presentation by the high-risk alleles augmented it. Importantly, HLA-II variations differentially polarized cytokine responses to Strep-SAgs: the presentation of Strep-SAgs by HLA-II-DR15/DQ6 alleles elicited significantly higher ratios of anti-inflammatory cytokines (e.g., IL-10) to proinflammatory cytokines (e.g., IFN-γ) than did their presentation by the high-risk HLA-II alleles. Adding exogenous rIL-10 significantly attenuated responses to Strep-SAgs presented by the high-risk HLA-II alleles but did not completely block the response; instead, it reduced it to a level comparable to that seen when these superantigens were presented by the protective HLA-II alleles. Furthermore, adding neutralizing anti-IL-10 Abs augmented Strep-SAg responses in the presence of protective HLA-II alleles to the same level as (but no higher than) that seen when the superantigens were presented by the high-risk alleles. Our findings provide a molecular basis for the role of HLA-II allelic variations in modulating streptococcal sepsis outcomes and suggest the presence of an internal control mechanism that maintains superantigen responses within a defined range, which helps to eradicate the infection while attenuating pathological inflammatory responses that can inflict more harm than the infection itself.     

Crystal structure of folliculin reveals a hidDENN function in genetically inherited renal cancer

Mutations in the renal tumour suppressor protein, folliculin, lead to proliferative skin lesions, lung complications and renal cell carcinoma. Folliculin has been reported to interact with AMP-activated kinase, a key component of the mammalian target of rapamycin pathway. Most cancer-causing mutations lead to a carboxy-terminal truncation of folliculin, pointing to a functional importance of this domain in tumour suppression. We present here the crystal structure of folliculin carboxy-terminal domain and demonstrate that it is distantly related to differentially expressed in normal cells and neoplasia (DENN) domain proteins, a family of Rab guanine nucleotide exchange factors (GEFs). Using biochemical analysis, we show that folliculin has GEF activity, indicating that folliculin is probably a distantly related member of this class of Rab GEFs.     

Structure-function relationships in human salivary <Emphasis Type="Italic">α</Emphasis>-amylase: role of aromatic residues in a secondary binding site

Human salivary α-amylase (HSAmy) has three distinct functions relevant to oral health: (i) hydrolysis of starch; (ii) binding to hydroxyapatite; and (iii) binding to bacteria (e.g. viridans streptococci). Oral bacteria utilize the starch hydrolyzing activity of HSAmy to derive their nutrients from dietary starch. Localized acid production by bacteria, through the metabolism of maltose generated by HSAmy, can lead to the dissolution of tooth enamel, a critical step in dental caries formation. HSAmy is a component of the acquired enamel pellicle and is used by Streptococcus gordonii to colonize the oral cavity. Although the active site of HSAmy for starch hydrolysis is well characterized, the regions responsible for the bacterial binding are yet to be defined. Since HSAmy possesses several secondary saccharide-binding sites in which aromatic residues are prominently located, we hypothesized that one of the secondary saccharide-binding sites harboring the aromatic residues W316 and W388, may play an important role in bacterial binding. To test this hypothesis, the aromatic residues W316 and W388 were mutated to alanine. The wild type and the mutant enzymes were characterized for their abilities to exhibit enzyme activity, starch binding and bacterial binding. Our results clearly showed that (i) the mutants W316A and W388A were not impaired in starch binding or bacterial binding; (ii) mutation of aromatic residues at these sites does not alter the overall conformation of the molecule; and (iii) the hydrolytic activity of the enzyme is unaffected against starch as substrates but reduced significantly against oligosaccharides.     

Immunologic analysis of human breast cancer progesterone receptors. 1. Immunoaffinity purification of transformed receptors and production of monoclonal antibodies

A monoclonal antibody (MAb), designated PR-6, produced against chick oviduct progesterone receptors [Sullivan, W. P., Beito, T. G., Proper, J., Krco, C. J., & Toft, D. O. (1986) Endocrinology (Baltimore) 119, 1549-1557] cross-reacts with the Mr 120,000 human B receptors. An immunomatrix prepared with PR-6 was used to purify progesterone receptors (PR) from T47D human breast cancer cells. Single-step immunoaffinity chromatography results in enrichment of B receptors (identified by immunoblot with PR-6 and by photoaffinity labeling with [3H]promegestone) to a specific activity of 1915 pmol/mg of protein (or 23% purity) and with 27% yield. Purity and yields as judged by gel electrophoresis and densitometric scanning of the B protein were approximately 1.7-fold higher due to partial loss in hormone binding activity at the elution step. A second purification step by diethylaminoethyl chromatography gives further enrichment to 3720 pmol/mg of protein (or 44% purity) to yield essentially two proteins, 120-kilodalton (kDa) B receptors and a 76-kDa non-steroid binding protein, each in approximately equivalent amounts. B receptors purified under these conditions are transformed and biologically active. They were maintained as undegraded 120-kDa doublets and retained both hormone and DNA binding activities. Isolated B receptors were free of the 90-kDa non-steroid binding protein observed to be associated with 8S untransformed receptors in other systems and were free also of the non-hormone binding 105-108-kDa B antigen described previously to copurify with chick PR. These purified B receptors were used as immunogen for production of four monoclonal antibodies against human PR. Three of the MAbs, designated as B-30 (IgG1), B-64 (IgG1), and B-11 (IgM), are specific for B receptors. The fourth MAb, A/B-52 (IgG1), reacts with both A and B receptors. The IgG MAbs are monospecific for human PR since they recognize and absorb native receptor-hormone complexes, displace the sedimentation of 4S receptors on salt containing sucrose gradients, and, by immunoblot assay of crude T47D cytosol, react only with receptor polypeptides. Although mice were injected with B receptors only, production of A/B-52 which recognized both A and B receptors provides evidence that these two proteins share regions of structural homology. These new MAbs are valuable reagents for further studies of human receptor structure and function and for clinical immunodetection of PR in breast tumors.     

Setaria digitata: identification and characterization of a hypodermally expressed SXP/RAL2 protein

SXP-1, an immunodominant filarial protein identified from Wuchereria bancrofti from our centre and previously exploited for diagnosis of human lymphatic filariasis, has been shown to be well conserved across several filarial species. In the present study, we describe the identification of SXP protein from the cattle filarid Setaria digitata using antiserum raised against recombinant WbSXP-1, and were able to detect 34 and 66kDa proteins from the crude protein extracts of S. digitata. These reactive proteins were found to be sheath proteins localized to the hypodermal region of the parasite.