Allantoinase: association with amphibian hepatic peroxisomes.

Differential and density-gradient centrifugation studies have estaiblished an associatiotn between allaintoinase and peroxisomes from the liver of the frog Rana pipiens. The presence of allantoincase in the peroxisome indicates. a iricolytic function for this orgainelle in the liver of amphibians.

A selective inhibitor of the osteoclastic V-H(+)-ATPase prevents bone loss in both thyroparathyroidectomized and ovariectomized rats.

A potent and selective inhibitor of the osteoclastic V-H(+)-ATPase, (2Z,4E)-5-(5,6-dichloro-2-indolyl)-2-methoxy-N-(1,2,2,6, 6-pentamethylpiperidin-4-yl)-2,4-pentadienamide (SB 242784), was evaluated in two animal models of bone resorption. SB 242784 completely prevented retinoid-induced hypercalcemia in thyroparathyroidectomized (TPTX) rats when administered orally at 10 mg/kg. SB 242784 was highly efficacious in the prevention of ovariectomy-induced bone loss in the rat when administered orally for 6 months at 10 mg/kg/d and was partially effective at 5 mg/kg/d. Its activity was demonstrated by measurement of bone mineral density (BMD), biochemical markers of bone resorption, and histomorphometry. SB 242784 was at least as effective in preventing bone loss as an optimal dose of estrogen. There were no adverse effects of compound administration and no effects on kidney function or urinary acidity. Selectivity of the inhibitor was further studied using an in situ cytochemical assay for bafilomycin-sensitive V-H(+)-ATPase using sections of osteoclastoma and numerous other tissues. SB 242784 inhibited the osteoclast enzyme at 1,000-fold lower concentrations than enzymes in any of the other tissues evaluated. SB 242784 demonstrates the utility of selective inhibition of the osteoclast V-H(+)-ATPase as a novel approach to the prevention of bone loss in humans.

Ion effects on protein-nucleic acid interactions: the disassembly of the 50-S ribosomal subunit from the halophilic bacterium, Halobacterium cutirubrum.

The 50-S ribosomal subunits from the extreme halophilic bacterium, Halo-bacterium cutirubrum, stable structurally and functionally in concentrated salt solutions were subjected to ionic environments depleted in either K+ or Mg2+ or both. Under these conditions specific classes of proteins are released from the subunit along with the 5 S RNA. Two-dimensional electrophoretic analysis of the resultant split protein fractions indicate some mutually exclusive effects of specific ions on the binding of specific proteins to the 23 S RNA as well as on the retention of 5 S RNA within the ribosomal macrostructure.

Cloning of the ColE3-CA38 colicin and immunity genes and identification of a plasmid region which enhances colicin production.

The colicin and immunity genes of plasmid ColE3-CA38 have been localized by characterization of bacteria carrying its cloned restriction fragments. They are within a 3.14-kb EcoRI segment, such that the immunity gene contains the KpnI site, and the colicin gene is adjacent to it within a 2.1-kb KpnI-HincII segment. The immunity gene and one end of the colicin gene are in the region of ColE3-CA38 which is not homologous to the closely related plasmid ColE2-P9. A 0.64-kb PvuI-EcoRI segment of the plasmid adjacent to that containing the colicin and immunity genes was found to augment colicin production on solid media, and also affected the morphology of clearing zones produced by the cells when used as indicators in overlays of stabs of colicin E2 or E7 producers. The 0.64-kb segment was required in its native orientation relative to the 3.14-kb EcoRI segment to cause its effects.

Restriction endonuclease mapping of ColE2-P9 and ColE3-CA38 plasmids.

Using single and double restriction-endonuclease digestions, 16 and 17 cleavage sites have been mapped for the ColE2-P9 and ColE3-CA38 plasmids, respectively. One or more sites for AvaI, BglI, EcoRI, HincII, PvuI, PvuII, SmaI and XhoI endonucleases were found in both plasmids, two BglII sites were found only in ColE2-P9, and one KpnI site was unique to ColE3-CA38. ColE2-P9 was found to be slightly smaller than ColE3-CA38, 4.4 Md compared to 4.6 Md. Eleven restriction sites are common to both plasmids in that they are identically placed relative to each other. These sites define a continuous DNA segment equal to over 60% of each plasmid. The remaining portions of the plasmids, which contain the non-homologous regions identified by Inselburg and Johns (1975) have no restriction sites in common, and differ in size by about 0.2 Md.

A direct-selection vector derived from pColE3-CA38 and adapted for foreign gene expression.

The construction of a plasmid vector, pVT25, which allows an efficient and direct selection for transformed cells carrying recombinant plasmids is described. In this vector, the replicon and ApR gene from plasmid pBR327 are fused to the colE3 gene of pColE3-CA38, whereby positive selection is based on the inactivation of the lethal colicin E3 by the insertion of a foreign DNA fragment. However, pVT25 can be maintained within the Escherichia coli cells when complemented with another plasmid, pVT26, which expresses the colicin E3 immunity (imm) and the TcR phenotypes. Furthermore, pVT25 was used to regulate the expression of the synthetic human proinsulin gene fused to the colE3 gene at the single ClaI site. The production of the characteristic C-peptide of proinsulin, monitored by radioimmunoassay, was shown to be under the control of the inducible promoter of the colE3 gene.

Characterization and nucleotide sequence of a colicin-release gene in the hic region of plasmid ColE3-CA38.

Downstream from its colicin and immunity genes (col. imm), Escherichia coli plasmid ColE3-CA38 contains a 0.81-kb DNA segment, the hic region, which is required for high colicin production. Characterization of derived plasmids, carrying the col-imm operon but varying in the hic region, showed that the latter functions in lacuna production, colicin release, cell death, and lysis. The hic gene expression after induction was shown to be dependent on the col gene promoter. The nucleotide sequence of the 0.81-kb region was determined and the hic gene localized to its imm-distal portion following an open reading frame (ORF) with no known function. There are two overlapping ORFs in that portion of the sequence, one of which was identified as the hic gene by its partial homology to lysis gene H of CloDF13. The 3' half of the hic gene is non-essential and contains a terminator-like DNA sequence. Preceding the gene, there are also inverted repeats which may attenuate its transcription.

Isolation and structure of the replicon of the promiscuous plasmid pCU1.

Evidence is presented to indicate that a PvuII fragment of approx. 2 kb isolated from the 39-kb IncN-group plasmid pCU-1 contains all plasmid-borne determinants for stable maintenance as an extrachromosomal element in Escherichia coli K-12. The fragment was sequenced. The features of this sequence include a group of 13 direct tandem repeats of 37 bp and a second group of two other direct repeats of 30 bp flanking a third partial member of this group. In addition, for a 19-bp sequence that overlaps a member of this second group, there are inverted repeats that straddle the members of the first group. There are three open reading frames within the fragment. We compare features of this sequence with that of other plasmid replicons and draw attention to similar and to dissimilar features.

Discovery of a novel class of substituted pyrrolooctahydroisoquinolines as potent and selective delta opioid agonists, based on an extension of the message-address concept.

This paper describes the design and synthesis of compounds belonging to a novel class of substituted pyrrolooctahydroisoquinolines which are potent and selective delta opioid agonists. Molecular modeling studies performed on known, selective delta ligands such as (+)-3 and the potent delta agonists SNC 80 led to the identification of the carboxamido moiety of the latter as a putative nonaromatic delta address. Insertion of this moiety onto the octahydroisoquinoline opioid message resulted in (+/-)-5b, a potent and selective delta ligand. The active enantiomer, (-)-5b, displayed nanomolar affinity for the delta receptor (Ki = 0.9 nM) with good mu/delta and kappa/delta binding selectivity ratios (140 and 1480, respectively). In addition, (-)-5b behaved as a full delta agonist in the mouse vas deferens bioassay having an IC50 = 25 nM and being antagonised in the presence of 30 nM naltrindole (NTI). These studies, based on the message-address concept, indicated that the nonaromatic (N,N-diethylamino)carbonyl moiety is a viable alternative to the classical benzene ring as a delta opioid address. Preliminary in vivo studies showed that (+/-)-5b produced a dose-related antinociception in the mouse abdominal constriction test after intracerebroventricular administration (ED50 = 1.6 micrograms/mouse).

Stable maintenance in chemostat-grown Escherichia coli of pBR322 and pACYC184 by disruption of the tetracycline resistance gene.

Plasmid stability was studied in antibiotic-free chemostat cultures. Disruption, either by deletion or insertion, of the tetracycline resistance gene in the EcoR1/EcoRV region of the cloning vector pBR322 or in the HindIII/BamH1 region of pACYC184 yields plasmids markedly more stable than the parent plasmids. Thus, at least for these two instances, cloning of a partitioning (par) locus is not prerequisite for plasmid maintenance.

The immunity genes of colicins E2 and E8 are closely related.

We have determined the nucleotide sequence of the newly characterized colicin E8 imm gene which exists in tandem with the colicin E3 imm gene in the ColE3-CA38 plasmid. Comparison of these immunity structures reveals considerable sequence divergence, but the ColE8 imm gene is markedly homologous to the colicin E2 imm gene from the ColE2-P9 plasmid.

[The psycho-affective behavior of pregnant women]. / Studio sul comportamento psico-affettivo della gravida.

The puerperal condition represents a quite new, physiological trial that every woman approaches, from a psycho-affective point of view, with her own cultural, cognitive and behavioural experiences. The strict individuality of these experiences makes the new affective maternal-foetal balances extremely unstable. It is suggested to establish the more common psycho-affective reactions of the pregnant woman by analysing the answers to a questionary proposed to 696 puerperae at the 2nd Obstetrics and Gynaecology Clinic, University of Turin, in the period january-september 1982. The main datum consists in the need of serenity during pregnancy and labour, that could be gained by means of better physician-patient relation.

Novel bone antiresorptive agents that selectively inhibit the osteoclast V-H+-ATPase.

The vacuolar proton pump (V-ATPase) located on the plasma membrane of the osteoclast is a potential molecular target for the discovery of novel bone antiresorptive agents useful for the treatment of osteoporosis. In order to design novel compounds able to selectively inhibit the osteoclast V-ATPase we firstly identified the minimal structural requirements of bafilomycin A1, a macrolide antibiotic which potently inhibits all V-ATPases. This information allowed the design of 2-(indole)pentadienamide derivatives whose optimization led to a novel class of potent inhibitors that demonstrated a high degree of selectivity for the osteoclast V-ATPase. The most interesting derivative, SB-242784, was able to inhibit bone resorption by human osteoclasts in vitro and to completely prevent ovariectomy-induced bone loss in rats when administered orally at 10 mg kg(-1) day(-1). Structure activity relationships of this class of compounds were investigated further by replacing the 2,4-pentadienoyl chain with suitable spacers able to maintain the correct orientation and distance between the indole ring and the amide moiety.

The tripeptide ZAMI-420 inhibits thromboxane-induced gastric vasoconstriction and ischaemia.

The tripeptide ZAMI-420 has been shown by Gervasi et al. (4) to be able to prevent experimentally-induced gastric damage, possibly by interfering with the synthesis and the action of thromboxane A2 (TXA2), a potent vasoconstrictor and platelet aggregator. Further studies on a canine stomach wedge preparation, supplied with a fixed flow of 10 ml/min-1 of arterial blood from the same dog have been designed to investigate this hypothesis. Bolus injection of arachidonic acid (AA) through a 30-sec incubation coil that allows the production of TXA2 resulted in a dose-related increase in resistance to flow in the stomach wedge vasculature and blanching of the gastric mucosa. This was progressively inhibited by ZAMI-420 perfused through the delay coil. Similar results were obtained with 1-benzylimidazole (BI). ZAMI-420, but not BI, produced a partial inhibition of TXA2-induced vasoconstriction when infused close to the stomach. Investigations of the antagonistic action of ZAMI-420 on the pharmacological effect of the formed TXA2 were carried out using strips of celiac and mesenteric artery from rabbits and gastric artery from dogs. Preincubation of these vascular preparations with ZAMI-420 led to progressive inhibition of the contraction induced either by the stable endoperoxide U-46619 or by CaCl2. Whittle et al. (3) reported that TXA2 may be involved in the pathogenesis of ulcerative disorders of the stomach; if so, both the inhibition of the synthesis and the antagonism of TXA2-induced effects could be of value in the prevention of experimentally-induced gastric disorders.

[Prevention of infection and improvement of cenesthesia with thymostimulin during chemotherapy following mastectomy]. / Prevenzione dei processi infettivi e miglioramento della cenestesi con timostimolina in pazienti mastectomizzate in corso di chemioterapia.

Eighty-five breast cancer patients who had been operated for mastectomy and were currently undergoing adjuvant or therapeutic chemotherapy were included in the study. Patients were divided into two matching groups of 50 (A) and 35 (B) subjects; group A was treated with CT and thymostimulin Serono and group B with CT alone. The difference between the two groups in relation to first-degree (24% vs 57%) and second-degree leucopenia (8% vs 14%), in addition to the difference in relation to infections and the regularity of chemotherapy cycles confirmed the value of immunotherapy with thymostimulin during chemotherapy for breast cancer.

[Positive effects of oral contraceptives]. / Effetti positivi dei contraccettivi orali.

PIP: The usefulness of oral contraceptives (OCs) has been fully reappraised in recent years, and numerous beneficial effects on general health have been demonstrated over and above contraceptive action. Examination of several prospective and retrospective epidemiological studies has pointed to a reduced incidence of ovarian functional cysts and ovarian carcinoma in women taking OCs. Dysmenorrhea and premenstrual tension are also diminished while the risk of iron-deficiency anemia is decreased by 50% owing to a reduction in menstrual flow. There is approximately a 50% reduction in endometrial carcinoma risk, coupled with a significant reduction in the incidence of benign breast diseases. OCs also offer protection against rheumatoid arthritis and pelvic inflammation. Lastly, it is pointed that fears concerning augmented risk for cardiovascular disease while on OCs have proven to be false alarms. (author's modified)^ieng

[Antibiotic prophylaxis in obstetric surgery. Experience with a sulbactam-ampicillin combination]. / Profilassi antibiotica in chirurgia ostetrica. Esperienza con l'associazione sulbactam-ampicillina.

Antibiotic prophylaxis reduces the incidence of infections after some types of surgical interventions; in Obstetrics it can prevent infections in high risk situations. Infections can occur in particular situations, even in cesarean sections (CS) at low risk. The incidence of puerperal endometritis is variable in literature, while the incidence of pelvic or surgical wound infections is 3.8% in elective CS with respect to 7.5% in emergency CS. This study verifies the efficacy of the sulbactam-ampicillin association (Unasyn, Pfizer) in the prophylaxis of all cesarean sections, complicated or not. Unasyn was administrated one hour prior to CS and 8 and 16 hours after CS in 162 patients. Therapy was continued in 8 cases because of high risk for infection. The evaluation of the efficacy of the drug was based on clinical criteria. There were no complications or fever recorded and no toxic or allergic reactions occurred. Antibiotic prophylaxis is recommended for all patients undergoing CS.

Management of patients with intrauterine fetal death.

Fetal death incidence is 5-10 per 1,000 births. About 25% of the women who carry a dead fetus for more than 4 weeks will show significant alterations in their coagulation system. The treatment for a patient with endouterine fetal death depends on when the pregnancy is terminated, based on the ecographic fetus age. There were 15,070 births from January 1983 to December 1994 in Department B of the Institute of Obstetrics and Gynecology, University of Torino. We took into consideration the cases ofintrauterine fetal death between the 26th and 40th week before labour. This study is based on a cohort of 57 cases of intrauterine fetal demise from the 24th to the 40th week of pregnancy before spontaneous labour.

Treating rabies.

Rabies was first reported by Aristotle over 2,000 years ago and has plagued man ever since. Little has been done to provide adequate treatment once the symptoms have appeared. Therefore the battle against rabies involves health education and prophylaxis, although during the incubation period rabies is treatable. The incubation period of this communicable disease can range from ten days to six months but may be as long as two years. The length of the incubation period is determined by the proximity and severity of the bite, to the brain.

Selective cytotoxicity and apoptosis induction in glioma cell lines by 5-oxygenated-6,7-methylenedioxycoumarins from Pterocaulon species.

The coumarins 5-methoxy-6,7-methylenedioxycoumarin 1 5-(3-methyl-2-butenyloxy)-6,7-methylenedioxycoumarin 2 and 5-(2,3-dihydroxy-3-methylbutyloxy)-6,7-methylenedioxycoumarin 3 isolated from Pterocaulon species showed significant cytotoxicity against two glioma cells lines. Compound 1 presented IC(50) values of 34.6 µM and 31.6 µM against human (U138-MG) and rat (C6) glioma cells, respectively, and this compound was at least two times more potent than compounds 2 and 3. This result could be explained by the planar conformation adopted by 1 through a non-classical hydrogen bond between a hydrogen of the methoxy and the oxygen of the methylenedioxy groups. Another important finding was that the cytotoxic effect induced by 1 in glioma cells was not observed in organotypic cultures, indicating a selective cytotoxicity for tumor cells.