Octapeptide analogs of somatostatin exhibiting greatly enhanced in vivo and in vitro inhibition of growth hormone secretion in the rat

Analogs of a superactive somatostatin (SRIF) octapeptide (code named SMS 201-995 (1)) were synthesized using solid-phase synthetic methodology and assayed for their ability to inhibit growth hormone release from cultured rat anterior pituitary cells and in sodium pentobarbital-anesthetized rats. One analog: (Formula: see text) exhibited greatly enhanced in vitro inhibitory activity (greater than 1,000x) relative to both the parent octapeptide molecule and to the 14 amino acid SRIF molecule. This analog which was also very potent in vivo contains a tyrosine residue and, given its high in vitro activity, may be of investigative importance as a radioiodinated ligand in receptor assays. An octapeptide retro-inverso analog also exhibited significant SRIF-like activity. Several very low activity octapeptide analogs were synthesized and were found to be devoid of SRIF-antagonist activity. A dodecapeptide analog previously shown to be superactive in vivo also demonstrated high in vitro activity.     

Selective release of follicle-stimulating hormone and luteinizing hormone by 5 alpha-dihydroprogesterone and 3 alpha, 5 alpha-tetrahydroprogesterone in pregnant mare's serum gonadotropin-primed immature rats exposed to constant light

The effects of 5 alpha-dihydroprogesterone (5 alpha-DHP) and 3 alpha, 5 alpha-tetrahydroprogesterone (3 alpha, 5 alpha-THP) on follicle-stimulating hormone (FSH) and luteinizing hormone (LH) release were examined in the pregnant mare's serum gonadotropin (PMSG)-primed immature female rat (8 IU PMSG at 28 days of age) maintained in constant light. Control rats kept in 14L:10D conditions exhibited proestrous-like surges of LH and FSH release with peak levels attained at 1800 h on the second day after PMSG treatment. In rats exposed to constant light, the PMSG-induced surges of LH and FSH were not only delayed until 1000 h on the third day after PMSG, resulting in a delay in ovulation, but were also significantly attenuated when compared to the gonadotropin surges that occurred on Day 2 in rats kept under normal light-dark conditions. The administration of 5 alpha-DHP significantly enhanced the release of FSH at 1000 h on Day 3 when compared to constant light-exposed controls, but had no effect on LH. Treatment with 3 alpha, 5 alpha-THP selectively potentiated the release of LH at 1000 h on Day 3 and had an attenuating effect on FSH release on Days 2 and 3. These observations confirm earlier findings in the immature ovariectomized estrogen-primed rat and suggest that 5 alpha-DHP and 3 alpha, 5 alpha-THP may have significant roles in the regulation of FSH and LH secretion.     

Nucleotide sequence of a spectinomycin adenyltransferase AAD(9) determinant from Staphylococcus aureus and its relationship to AAD(3") (9)

Summary The nucleotide sequence of the spc determinant of the Staphylococcus aureus transposon Tn554 has been determined. This gene encodes a spectinomycin adenyltransferase, AAD(9), that mediates resistance to spectinomycin but not to streptomycin. The sequence predicts a 260 amino acid protein of molecular weight 28,943. A spectinomycin-sensitive mutant (spc-1) contains a GA transition resulting in substitution of threonine (ACA) for alanine (GCA) at residue 165. The predicted amino acid sequence is 36% homologous to that of a widely distributed, gramnegative streptomycin/spectinomycin adenyltransferase, AAD(3) (9), specified by the aadA determinant (Holingshead and Vapnek 1985).     

Bicarbonate-Reversible and Irreversible Inhibition of Photosystem II by Monovalent Anions

We tested a number of inhibitory monovalent anions for their primary site of action on photosystem II(PSII) in chloroplasts. We find that the inhibitory effects of F⁻, HCO₂⁻, NO₂⁻, NO₃⁻, and CH₃CO₂⁻ are all reversed by addition of a high concentration of HCO₃⁻. This class of anions competitively inhibits H¹⁴CO₃⁻ binding to PSII. All of those anions tested reduced H¹⁴CO₃⁻ binding more in the light than in the dark. We conclude that the primary inhibitory site of action of a number of monovalent anions is at the HCO₃⁻ binding site(s) on the PSII complex. The carbonic anhydrase inhibitor gold cyanide, and also azide, inhibit PSII but at a site other than the HCO₃⁻ binding site. We suggest that the unique ability of HCO₃⁻ to reverse the effects of inhibitory anions reflects its singular ability to act as a proton donor/acceptor at the anion binding site. A similar role has been proposed for non-substrate-bound HCO₃⁻ on carbonic anhydrase by Yeagle et al. (1975 Proc Natl Acad Sci USA 72: 454-458).     

Biosynthesis of tissue inhibitor of metalloproteinases by human fibroblasts in culture. Stimulation by 12-O-tetradecanoylphorbol 13-acetate and interleukin 1 in parallel with collagenase

Biosynthesis of the glycoprotein tissue inhibitor of metalloproteinases (TIMP) by human fibroblasts in culture has been characterized by functional assays, immunoprecipitation, and immunocytochemistry with a monospecific antiserum. As determined by radiolabeling with [35S]methionine, immunoprecipitation, and analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the secreted form of TIMP had an Mr of 29,000, whereas the form associated with the cell layer had an Mr of 24,000. Unstimulated human lung fibroblasts (HFL-1) secreted TIMP at the rate of approximately 2 micrograms/10(6) cells/24 h, and normal foreskin fibroblasts (HS 27) and skin fibroblasts from a patient with Hurler's disease (GM 1391) secreted TIMP at 0.3 and 0.2 micrograms/10(6) cells/24 h, respectively. Secretion of TIMP was stimulated up to 10-fold by treating the cells with 20-100 ng/ml of 12-O-tetradecanoylphorbol 13-acetate or 10 units/ml of human interleukin 1. In the stimulated HFL-1 cells, TIMP accounted for 0.03-0.09% of the total [35S]methionine incorporated into protein, and 0.3-0.8% of the [35S]methionine in secreted protein. Although TIMP accounted for a relatively small proportion of total protein synthesis of the fibroblasts, greater than 80% of untreated and greater than 95% of stimulated fibroblasts synthesized TIMP, as determined by indirect immunofluorescence. The treatments of the human fibroblasts that increased TIMP secretion also induced synthesis and secretion of proenzyme forms of collagenase, indicating that degradative enzymes and their controlling inhibitors may be synthesized in parallel under certain conditions.     

Detection and physical map of a omega tox+-related defective prophage in Corynebacterium diphtheriae Belfanti 1030(-)tox-.

A library of chromosomal DNA from Corynebacterium diphtheriae Belfanti 1030(-)tox- was cloned in the lambda phage vector EMBL4 and screened for sequences homologous to corynephage omega tox+ and the attB1-attB2 region of the C7(-)tox- chromosome. Two portions of the 1030(-)tox- chromosome, 35 and 30.5 kilobases long which contain, respectively, the entire region homologous to corynephage omega tox+ and the attB1-attB2 sites, were mapped with the restriction endonucleases BamHI and EcoRI. Chromosomal DNA from 1030(-)tox- was shown to contain a 15.5-kilobase region that was homologous to ca. 42% of the corynephage omega tox+ genome. These sequences were found to hybridize to three regions of the phage genome and do not contain either the diphtheria tox operon or the attP site. These sequences are distant from the chromosomal region that contains the attB1-attB2 sites. Moreover, unlike other known defective prophages, the physical map of this prophage starts at the cos site and is colinear with the vegetative phage map. The 30.5-kilobase region of the 1030(-)tox- chromosome, which contains the attB1-attB2 sites, has a central core region that is almost identical to the corresponding region of the C7(-)tox- chromosome; however, the flanking sequences in these two strains of C. diphtheriae are different.     

Nucleoprotein and membrane protein genes are associated with restriction of replication of influenza A/Mallard/NY/78 virus and its reassortants in squirrel monkey respiratory tract.

An avian influenza A virus, A/Mallard/NY/6750/78(H2N2), was restricted in in replication in the respiratory tract of squirrel monkeys. Avian-human influenza A reassortant viruses possessing the six RNA segments coding for nonsurface proteins (i.e., internal genes) of this avian virus were as restricted in replication in squirrel monkeys as their avian influenza parent. These findings indicated that restriction of replication of the avian influenza virus is a function of one or more of its internal genes. For an investigation of which of the avian influenza genes was responsible for restricted replication in the respiratory tract of primates, reassortant viruses were produced that contained human influenza virus surface antigens from the A/Udorn/72(H3N2) virus and one or more of the internal genes derived from the avian influenza virus parent. Avian-human reassortant influenza A viruses containing only the nucleoprotein or matrix protein RNA segment from the avian influenza virus parent were as restricted in their growth as an avian-human influenza reassortant virus containing each of the six avian influenza internal genes. In addition, an avian-human influenza reassortant virus possessing only the avian RNA 1 and nonstructural genes (which by themselves do not specify restricted replication) manifested a significant reduction of virus replication in squirrel monkey tracheas. Thus, the avian nucleoprotein and matrix genes appear to play a major role in the host range restriction exhibited by the A/Mallard/78 virus and its reassortants, but the combination of RNA 1 and nonstructural genes also contributes to restriction of replication.     

A 6-year experience with immediate reconstruction after mastectomy for cancer

A 6-year retrospective review is presented of 185 patients who underwent immediate reconstruction of the breast at the same operation as mastectomy for carcinoma. The patients were treated at two institutions under similar protocols of patient selection, surgical technique, and postoperative care. A detailed evaluation is presented from both the oncologic and surgical points of view. The data support the conclusion that immediate reconstruction of the breast does not alter survival or cancer recurrence rates and does not interfere with the treatment of primary or secondary disease. A low incidence of significant surgical complications is also detailed. Combined with previous reports answering psychological concerns about this mode and timing of reconstruction, this review offers significant reassurance about the overall safety of immediate reconstruction. The authors therefore recommend immediate reconstruction of the breast as a safe treatment option for the woman facing mastectomy.     

Quantitation of the transgenome size in chromosome-mediated gene transfer lines

Twenty-two HPRT-selected chromosome-mediated gene transfer lines were characterized by quantitative "dot" blotting. The range of human sequences in these lines extended from over 120,000 kb to less than 5,000 kb. One-half of these lines carried less than 16,000 kb.     

Changes in lung volume and deflation stability in hyaline membrane disease

Total lung capacity (TLC), inspiratory capacity, functional residual capacity, and deflation stability of prematurely delivered Macaca nemestrina primates were measured serially during development of, and recovery from, hyaline membrane disease (HMD) to relate changes in lung volumes to changes in deflation stability. Gestational age-matched primates that did not develop HMD served as controls. TLC, measured by N2 washout, fell at 2-12 h of age (P less than 0.0001) in animals with HMD and remained lower than controls for at least 48 h (P less than 0.005). However, deflation stability, defined as the fraction of TLC remaining upon deflation to 10 cm H2O, improved from 2 to 12 h of age (P less than 0.001). Postmortem studies confirm the measurements of TLC and deflation stability and provide evidence that interstitial thickening and obstruction of air spaces with debris may be partially responsible for the observed changes in TLC in primates that develop HMD. It has been assumed that TLC is reduced in HMD because of atelectasis from elevated alveolar surface tension, but the sequential measurements in these animals suggest that other mechanisms also contribute.