Genetic manipulation of antibiotic-producing Streptomyces

The genetic manipulation of Streptomyces species has been facilitated by the development of versatile cloning vectors, robust gene transfer systems and transposable elements. These molecular genetic tools have been used to construct antibiotic-producing strains with improved properties and recombinants for the production of hybrid glycopeptide and macrolide antibiotics, novel anthelminthic agents and novel cytotoxic anthracyclines.     

Real-time patch-cram detection of intracellular cGMP reveals long-term suppression of responses to NO and muscarinic agonists

Cyclic GMP (cGMP) is a crucial intracellular messenger in neuronal, muscle, and endocrine cells. The intracellular concentration of cGMP is regulated by various neurotransmitters, including acetylcholine (ACh) and nitric oxide (NO). While much is known about the biochemical steps leading to cGMP synthesis, little is known about cGMP kinetics in intact cells. Here, we use "patch-cramming," in which an excised, inside-out membrane patch containing cyclic nucleotide-gated ion channels is used as a biosensor, to obtain the first real-time measurements of cGMP in intact cells. Patch-cramming experiments on neuroblastoma cells show that both muscarinic agonists and NO rapidly elevate cGMP. NO elicits cGMP responses repeatedly without decrement, whereas responses to muscarinic agonists exhibit a profound and prolonged desensitization. Remarkably, muscarinic agonists also cause long-term (>30 min) suppression (LTS) of cGMP responses elicited by NO. Biochemical measurements reveal that rat sympathetic neurons also exhibit LTS of cGMP, suggesting that LTS is a widespread mechanism that may contribute to synaptic plasticity.     

Age incidence of meningococcal infection England and Wales, 1984-1991

Gene transfer with vectors derived from murine retroviruses is restricted to cells which are proliferating and synthesizing DNA at the time of infection. This suggests that retroviral-mediated gene transfer might permit targeting of gene integration into malignant cells in organs composed mainly of quiescent nonproliferating cells, such as in the brain. Accordingly, selective introduction of genes encoding for susceptibility to otherwise nontoxic drugs ("suicide" genes) into proliferating brain tumors may be used to treat this cancer. We investigated the efficacy and dynamics of in vivo transduction of growing brain tumors with the herpes simplex-thymidine kinase gene followed by administration of the antiviral drug ganciclovir. Ganciclovir is phosphorylated by thymidine kinase to toxic triphosphates that interfere with DNA synthesis, resulting in the preferential death of the transduced tumor cells. Rats inoculated with 4 x 10(4) 9L gliosarcoma cells into the frontal lobe were treated 7 days later with an intratumoral stereotaxic injection of murine fibroblasts (NIH 3T3 cells) that were producing a retroviral vector containing the herpes simplex-thymidine kinase gene. Controls received vector producer and nonproducer NIH 3T3 cell lines containing the Escherichia coli lacZ (beta-galactosidase) gene as well as nonproducer NIH 3T3 cells containing the thymidine kinase gene. The animals were rested for 7 days to allow time for in situ transduction of the proliferating tumor cells with the herpes-thymidine kinase retroviral vector. The animals were then treated with ganciclovir, 15 mg/kg i.p. twice a day for 14 days. Gliomas receiving an injection of 3-5 x 10(6) thymidine kinase producer cells regressed completely in 23 of 30 rats given ganciclovir therapy, while 25 of 26 control rats developed large tumors. Intratumoral injection of a lower concentration of thymidine kinase vector producer cells (1.8 x 10(6)) resulted in a lower frequency of tumor regression (5 of 13 rats). To estimate the efficiency of in vivo gene transfer, 9L brain tumors were given injections of 5 x 10(6) beta-galactosidase vector producer cells. 5-Bromo-4-chloro-3-indolyl-beta-D-galactopyranaside staining revealed maximal staining of beta-galactosidase within the tumor 7-14 days after injection of the vector producer cells. In vivo transduction rates in harvested tumors ranged from 10 to 70%. There was no evidence of transduction of the surrounding normal neural tissue. Occasional blood vessel endothelial cells within or adjacent to the tumor were observed to be 5-bromo-4- chloro-3-indolyl-beta-D-galactopyranaside positive.(ABSTRACT TRUNCATED AT 400 WORDS)     

Anatomical study of the cervical sympathetic trunk and ganglia in the albino rat (Mus norvegicus albinus)

Nine adult albino rats of both sexes were studied. 16 sympathetic trunks and ganglia were dissected in detail in eight rats. The right and left superior cervical ganglion and the sympathetic trunk below the ganglion were removed from an additional rat. The cell bodies of these ganglia and the axons of the trunks were counted with the aid of light and electron microscopy. Considering the number and location of ganglia and patterns of branching, the rat's cervical sympathetic nervous system compares closely with man's. There appears to be a relationship between body size and myelination of preganglionic neurons in the cervical sympathetic trunks, with smaller animals having the least number of myelinated fibers.     

Vulvar histology after neutral red photoinactivation of herpes simplex virus

The use of photodynamic dye and light inactivation for the treatment of genital herpes simplex virus infections has been associated with the risk of potential oncogenesis. Sixteen patients treated with neutral red and fluorescent light for documented herpetic infections were studied at intervals ranging from 9 to 52 months following treatment. Four patients treated with other modalities were included in the study. Biopsies of the treated areas were obtained, and 3925 tissue sections were examined. Mild atypical epithelial changes were focally present in most specimens regardless of therapy. Histologically identifiable premalignant change could not be demonstrated.     

A twin study of dental dimension. II. Independent genetic determinants

To demonstrate the presence of independent genetic determinants of multiple correlated tooth dimensions from twin data, a multivariate analysis was performed on the covariance matrices of monozygotic and dizygotic within-pair differences for mesiodistal and buccolingual dimensions of 28 teeth of the secondary dentition. The results provided strong evidences that the correlation among tooth dimensions is primarily genetic in origin, probably attributable to the pleiotropic action of either independent genes or groups of genes. Among the genetic factors that were identified, one appeared to affect the maxillary teeth in general while a second influenced primarily the anterior mandibular teeth. There was a striking tendency for homologous measurements on the right and left sides to be associated with the same genetic factor. In contrast, genetic determination of the maxillary and mandibular dentition seemed to be independent of each other, and a wider range of genetic factors were found to influence the mandibular than the maxillary teeth, suggesting that a differential degree of evolutionary stability may have been achieved in the teeth of the two jaws.     

Evidence for the presence of components of the alternative (properdin) pathway of complement activation in respiratory secretions

Activation of the alternative complement pathway by respiratory secretory IgA was demonstrated by incubating purified, aggregated preparations of serum and secretory IgA with neat human serum. No depletion of the early components (C1-4) was observed, but 63 and 70% of C3-9, respectively, were consumed. The C3-9-consuming capacity of heat-aggregated nasal secretions from an IgA-deficient volunteer was compared with heat-aggregated nasal secretions from a normal volunteer known to have secretory IgA. The deficient secretions consumed C3-9, whereas the IgA deficient secretions did not. Reconstitution of the nasal secretions from the IgA-deficient volunteer with purified secretory IgA produced alternative pathway activation. Factor B of the alternative complement pathway was found to be present in 16 of 18 bronchoalveolar lavage samples (BALF) from normal volunteers. Simultaneous measurement of lavage and serum albumin and Factor B concentrations rendered it unlikely that Factor B was merely a transudative product from serum in half the samples but rather suggested that it may be a component of lower respiratory tract secretions. The presence of an intact alternative complement pathway in BALF was indicated by showing that cobra venom factor and endotoxin cleaved functionally pure human C3 when mixed with BALF, but had no effect on C3 in the absence of BALF.     

Comparison of human and hamster mitochondrial transfer RNA. Physical properties and methylation status

In a comparative study, the effective of intraventricularly or intraperitoneally injected p-chloroamphetamine (p-CA) and some chloroindoles on cerebral levels of serotonin was evaluated. 5-Chloroindole depressed 5-HT levels in the brainstem and telencephalon for three days, 6-chloro-2-methylindole (6-CMI) only during the first day. 5-Chloroindazole had no effect at all. p-CA was more toxic to guinea pigs than to rats. p-CA and 5-chloro-2-methylindole (5-CMI) had no effect on cerebral 5-HT in chicks. Apparently, none of these compounds represented or was converted to a metabolite possibly responsible for the neurotoxic effects of p-CA.