Inhibition of the replication of hepatitis B virus by the carbocyclic analogue of 2''-deoxyguanosine.

We report that treatment of 2.2.15, a human hepatoblastoma-derived cell line in which hepatitis B virus is actively replicating, with the carbocyclic analogue of 2'-deoxyguanosine [Shealy, Y. F., O'Dell, C. A., Shannon, W. M. & Arnett, G. (1984) J. Med. Chem. 27, 1416-1421] resulted in the nearly complete cessation of viral replication, as monitored by the absence of both intracellular episomal and secreted viral DNAs and by the absence of viral DNA polymerase activity. The drug was nontoxic in concentrations up to 200 times the minimum effective inhibitory concentration.     

Results of long-term leukocyte interferon (alpha IFN) therapy using an individually determined dose schedule in recurrent laryngeal papillomatosis

Out of 24 patients with laryngeal papillomatosis 6 (5 female, 1 male) suffered from repeated relapses and underwent long-term treatment with alpha-IFN-therapy. Age at onset of the disease: 1 5/12-16 2/12 years. Duration of illness: 1-7 years, with several relapses were treated surgically and with laser-coagulation. Three out of 6 patients had a tracheal cannula and were cauterized by podophylline at 2-4 week intervals. IFN was given in dosages of 5-20 X 10(4) U/kg 2 or 3 times a week. IFN-dosage for each patient was determined using the induction kinetics of (2'-5')-oligo(A)synthetase (OAS) in the mononuclear cells of the circulating blood of patients with laryngeal papillomatosis. A continuous effect could be achieved by the dose of IFN determined in the described way always before OAS activity decreased to its initial level. All 6 patients responded favorably to the alpha-IFN-therapy. Two patients treated only with IFN showed remission without relapses. In 2 cases IFN was successfully used to prevent relapses after surgical treatment and laser-coagulation. In 2 patients with papillomatosis extending into the main bronchi the disease could only be brought to a standstill, i.e. it was not necessary to remove the papillomas. Two out of 3 patients with laryngeal papillomatosis could be decannulated. Long-term therapy following the above described principles is efficient and without significant side-effects. Three patients are in treatment for more than 3 10/12 years.     

Mapping the active-site tyrosine of vaccinia virus DNA topoisomerase I.

Site-directed mutagenesis of the vaccinia virus gene encoding a type I DNA topoisomerase implicates Tyr-274 as the active-site residue that forms a covalent adduct with DNA during cycles of DNA-strand breakage and reunion. Replacement of Tyr-274 by phenylalanine results in loss of the ability of the enzyme to relax negatively supercoiled DNA as well as to form the covalent DNA-protein intermediate. Substitution of phenylalanine for tyrosine at nine other sites in the protein has no apparent effect on enzyme activity. Amino acid sequence alignment reveals Tyr-274 to be homologous to Tyr-727 and Tyr-771, respectively, of the type I topoisomerases from Saccharomyces cerevisiae and Saccharomyces pombe; Tyr-727 and Tyr-771 have been shown to represent the active-site tyrosines of those enzymes. Sequence comparison of the active-site regions defines a motif Ser-Lys-Xaa-Xaa-Tyr common to the viral and cellular type I topoisomerases, including the human enzyme.     

Stimulation of Alpha-Adrenoceptors Facilitates the Release of Growth Hormone-Releasing Hormone from Rat Hypothalamus in vitro

While extensive evidence suggests that adrenoceptors play an important role in the control of growth hormone in the rat, there are few studies involving the direct measurement of growth hormone-releasing hormone (GHRH). We have therefore developed a radioimmunoassay for rat GHRH, and used it to investigate the modulation of GHRH release by noradrenaline from incubated rat hypothalamus in vitro. The GHRH radioimmunoassay had no significant cross-reactivity with other hypothalamic or GHRH-related peptides, and was sensitive to 4 pg/tube; intra- and interassay coefficients of variation were 6% and 12% respectively. Single incubated rat hypothalami produced a stable and readily measurable output of GHRH in successive 20 min incubations after an initial 60 min preincubation; the release of GHRH was increased in the presence of 56 mM KCI, but did not respond to KCI-depolarization when calcium was excluded from the medium. Stimulated GHRH release was identical to synthetic rat GHRH(1–43) on high-performance liquid chromatography and Sephadex G-75 chromatography.
Noradrenaline stimulated GHRH secretion in a dose-dependent manner in the concentration range 10⁻¹⁰— 10⁻⁶M, with a plateau in response at 10⁻⁷M. Stimulation with noradrenaline 10⁻⁷M was blocked by idazoxan 10⁻⁵M and attenuated by thymoxamine 10⁻⁵M, but was unaffected by timolol 10⁻⁵M. Both the α₂-adrenoceptor agonist guanfacine, and the α₁-adrenoceptor agonist methoxamine, specifically stimulated GHRH secretion.
It is concluded that noradrenaline stimulates the release of GHRH at both α₁ and α₂-adrenoceptors.     

T cells specific for alpha-beta interface regions of hemoglobin recognize the isolated subunit but not the tetramer and indicate presentation without processing.

Processing of a protein antigen into fragments is believed to be a prerequisite for its presentation by the antigen-presenting cell to the T cell. This model would predict that, in oligomeric proteins, T cells prepared with specificity for regions that are buried within subunit association surfaces should recognize the respective regions in vitro equally well on the isolated subunit or on the oligomer. Three hemoglobin (Hb) alpha-chain synthetic peptides, corresponding to areas that are situated either completely [alpha-(31-45)] or partially [alpha-(41-45) and alpha-(81-95)] within the interface between the alpha and beta subunits of Hb, and a fourth peptide representing a completely exposed area in tetrameric Hb were used as immunogens in SJL/J (H-2s) mice. Peptide-primed T cells were passaged in vitro with the respective peptide to obtain peptide-specific T-lymphocyte lines. T-cell clones were isolated from these lines by limiting dilution. T-cell lines and clones that were specific for buried regions in the subunit association surfaces recognized the free peptide and the isolated subunit but not the Hb tetramer. On the other hand, T cells with specificity against regions that are not involved in subunit interaction and are completely exposed in the tetramer recognized the peptide, the isolated subunit, and the oligomeric protein equally well. The responses of the T-cell lines and clones were major histocompatibility complex-restricted. Since the same x-irradiated antigen-presenting cells were employed, the results could not be attributed to differences or defects in Hb processing. The findings indicate that in vitro the native (unprocessed and undissociated) oligomeric protein was the trigger of major histocompatibility complex-restricted T-cell responses.     

The endoscopic bronchial picture of patients with pulmonary tuberculosis of various age groups

Endoscopic pictures of the bronchi were studied in 509 new cases of pulmonary tuberculosis. Tuberculosis of the bronchi and its aftereffects such as cicatricial lesions were detected in 31.1 per cent of the children and adolescents, 7.9 per cent of the adults at the age of 18 to 29 years, 9.0 per cent of the adults at the age of 30 to 55 years and 14.7 per cent of the adults at the age of 56 to 75 years. Nonspecific endobronchitis was diagnosed in 27.9 per cent of the children and adolescents, 44.9 per cent of the adults at the age of 18 to 29 years, 46.3 per cent at the age of 30 to 55 years and 44.1 per cent at the age of 56 to 75 years. The incidence and nature of tuberculosis of the bronchi and nonspecific endobronchitis were not related to the patients' age.     

von Hippel-Lindau disease affecting 43 members of a single kindred

We present a 6-generation kindred of over 221 members, 43 of whom were affected with von Hippel-Lindau (vHL) disease. Through a simple screening protocol, we diagnosed vHL retrospectively in 15 cases, and for the first time in 28, 11 of whom were presymptomatic. We found many complications of vHL in previously diagnosed relatives and in new cases. This study has demonstrated the utility and benefit of preventive surveillance in those known to have vHL, and of presymptomatic screening for affected relatives in families with vHL. The features of vHL were reviewed in our 43 cases and 511 cases from the medical literature. The patterns, frequencies, and ages of onset for each lesion were compared. Renal malignancies caused almost as much mortality in vHL as CNS malignancies. This family was exceptional for absence of pheochromocytoma and erythrocythemia, for more renal and pancreatic cysts and malignancies, and for slightly fewer eye or CNS lesions. Bilateral renal adenocarcinomata were found presymptomatically in five young subjects, who had bilateral nephrectomy and hemodialysis. Three survived long-term after renal transplants. Five relatives had pancreatic malignancies, which are definite although uncommon manifestations of vHL. Recommendations are made for family screening, which was economical and effective. Bayesian calculations help to predict risks for genetic counseling. The molecular basis of vHL may soon be found, since it has been linked to DNA markers on the short arm of chromosome 3.     

Prostaglandins with antiproliferative activity induce the synthesis of a heat shock protein in human cells.

Prostaglandins (PGs) A1 and J2 were found to potently suppress the proliferation of human K562 erythroleukemia cells and to induce the synthesis of a 74-kDa protein (p74) that was identified as a heat shock protein related to the major 70-kDa heat shock protein group. p74 synthesis was stimulated at doses of PGA1 and PGJ2 that inhibited cell replication, and its accumulation ceased upon removal of the PG-induced proliferation block. PGs that did not affect K562 cell replication did not induce p74 synthesis. p74 was found to be localized mainly in the cytoplasm of PG-treated cells, but moderate amounts were found also in dense areas of the nucleus after PGJ2 treatment. p74 synthesis was not necessarily associated with cytotoxicity or with inhibition of cell protein synthesis. The results described support the hypothesis that synthesis of the 70-kDa heat shock proteins is associated with changes in cell proliferation. The observation that PGs can induce the synthesis of heat shock proteins expands our understanding of the mechanism of action of these compounds whose regulatory role is well known in many physiological phenomena, including the control of fever production.