Androgen maintenance of erectile function in the rat penis.

Previous research has shown that the frequency and duration of penile erection is diminished after castration and that replacement with testosterone will restore the process. Using rats, the present study was designed to confirm that erection is androgen-dependent and to determine whether castration and androgen replacement affect the penile vascular smooth muscle responsiveness to vasoactive drugs. Blood pressure in the corpus cavernosum was measured directly during erections induced by electrical stimulation of the autonomic innervation of the penis. Maximal cavernosal pressure was markedly reduced after castration but was returned to normal levels if the castrated animals were treated with testosterone. Infusion of nitroglycerin (vasodilator) or phenylephrine (vasoconstrictor) resulted in a decline in cavernosal pressure in androgen-treated animals but not in castrated animals, even though the mean arterial blood pressure was strongly affected in all treatment groups by these drugs. When an inhibitor of nitric oxide synthesis was infused, cavernosal pressure was decreased in all groups, indicating that this substance is involved in penile erection. Taken together, these results show that androgens maintain the erectile process and may act specifically to support the responsiveness of the vascular smooth muscle to vasoactive drugs.     

Receptor-specific influence of endothelin-1 in the erectile response of the rat

Specific receptor antagonists were used to examine the role of endothelin-1 (ET-1) in the erectile response of the rat. In these studies, intact rats were cannulated to permit the continuous recording of mean arterial pressure (MAP) and intracavernosal pressure (CCP). Erection was induced by electrical stimulation of the autonomic ganglion, which regulates blood flow to the penis. The animals were subjected to intracavernosal injection with vehicle only (Cont) or with an antagonist to the endothelin-A receptor (ET(A)) or to the endothelin-B receptor (ET(B)). Blockade of the ET(A) or the ET(B) had no effect on the erectile response (CCP/MAP) during maximal ganglionic stimulation. When ET-1 was injected into Cont rats, there was a marked vasoconstriction with a sharp rise in MAP and a decline in CCP as the cavernosal arterioles constricted and limited inflow. The injection of the ET(A) antagonist prevented the vasoconstriction after ET-1 injection into Cont rats, whereas blockade of the ET(B) had no effect on the vasoconstrictive effect to ET-1. Similar results were obtained during submaximal ganglionic stimulation. With minimal levels of ganglionic stimulation, ET-1 injection led to a moderated degree of vasodilation in the presence of the ET(A) antagonist. The ET(B) antagonist failed to alter the CCP response during minimal stimulation, but it did have a marked effect on the MAP response to ET-1 injection. The results of these studies confirm that cavernosal tissue of the rat penis is highly responsive to ET-1. However, the failure of the ET-1 antagonists to affect penile erection in response to ganglionic stimulation reflects a minimal role of ET-1 in the erectile response in the rat.     

Survival Rate and Enzyme Activities ofLactobacillus acidophilusFollowing Frozen Storage

The ability of two strains of Lactobacillus acidophilus, CRL 640 and CRL 800, to survive and retain their biological activities under frozen storage was determined. Freezing and thawing, as well as frozen storage, damaged the cell membrane, rendering the microorganisms sensitive to sodium chloride and bile salts. Both lactic acid production and proteolytic activity were depressed after 21 days at -20 degreesC, whereas beta-galactosidase activity per cell unit was increased. Cell injury was partially overcome after repair in a salt-rich medium. Copyright 1998 Academic Press.     

Prevention of aneuploidy by S-adenosyl-methionine in human cells treated with sodium arsenite

Alterations in methyl group's metabolism affect availability of S-adenosyl-L-methionine (SAM); these modifications can be originated by enzyme polymorphisms, nutritional deficiencies, and exposure to chemical agents. There are several types of chemicals that interfere with methyl groups, among them is arsenic. It deserves special attention because it modifies a number of cell functions that influence the development of diseases such as cancer. Since part of arsenic's toxicity is influenced by changes on SAM availability, in a previous study we investigated whether exogenous addition of SAM to cells treated with sodium arsenite (NaAsO(2)) has an effect on its genotoxicity. Results demonstrated that SAM reduces the frequency of cells presenting micronuclei (MN) and tubulin-cytoskeleton defects after treatment with NaAsO(2). MN are fragments of the cell nucleus that may contain whole chromosomes or chromosome fragments depending on whether they derive either from the aneugenic or from the clastogenic action of chemicals. Therefore one question generated by these results was whether SAM reduced only the frequency MN resulting from aneugenic damage. To answer this question, in the present work we used an all-centromere DNA probe to distinguish the type of MN reduced by SAM after treatment with NaAsO(2) and vinblastine. In addition, the capacity of SAM to reduce clastogenicity was also evaluated. Results show that SAM decreases the frequency of cells with MN containing whole chromosomes in cultures treated either with NaAsO(2) or with vinblastine; however, induction of double-strand breaks by NaAsO(2) was not prevented by SAM.     

Anti-genotoxicity of coffee against N-methyl-N-nitro-N-nitrosoguanidine in mouse lymphoma cells

The main aim of this work was to test the hypothesis that instant coffee, a commonly consumed polyphenolic beverage with antioxidant activity, can protect mammalian cells against genotoxic effects in vitro. For this purpose, the L5178Y mouse lymphoma cell line was selected to assess modulatory effects of coffee on the genotoxicity of N-methyl-N-nitro-N-nitrosoguanidine (MNNG). We initiated the work with a set of preliminary experiments in which the cytokinesis-block micronucleus test was performed. Results obtained from these experiments demonstrated a dose-related decrease in genotoxicity following co-treatment of mouse lymphoma cells with three doses of caffeinated instant coffee. Both pre-treatment and co-treatment showed significant antigenotoxic effects against MNNG. Caffeinated and decaffeinated instant coffee samples inhibited genotoxicity. There was no significant change in the antigenotoxic effect of caffeinated instant coffee after filtration using a 0.2 microm filter. Similar in vitro experiments demonstrated antigenotoxic effects against MNNG when boiled coffee was used instead of instant coffee. On the basis of the findings from the above preliminary experiments, further work was carried out to evaluate the possible protective effects of caffeinated instant coffee against MNNG-induced DNA damage, mutation and chromosomal damage. Results from three or five independent experiments demonstrated significant protective effects of caffeinated instant coffee against MNNG-induced DNA damage in the comet assay, mutation at the Tk locus and chromosomal damage in the cytokinesis-block micronucleus test.     

Time-dependent effects of sodium arsenite on DNA breakage and apoptosis observed in the comet assay

To assess genotoxic effects of sodium arsenite (NaAsO2) the single-cell gel electrophoresis (comet assay) had been conducted in various studies indicating genotoxicity. However, DNA fragmentation due to NaAsO2-induced apoptosis may constitute a bias in the interpretation of the results. Apoptotic cells can show typically large and diffuse comets, which are usually excluded during genotoxicity analysis. It is controversial whether there is a time-window in which the apoptotic process generates comets that would falsely be interpreted to be the result of genotoxic DNA damage. Therefore, we evaluated frequency histograms for single-cell measures of tail DNA (% DNA in comet tail) in 30-min intervals after incubation of mouse lymphoma L5178Y cells with sodium arsenite (NaAsO2). In parallel, we evaluated apoptosis by measuring annexin V-positive cells with flow cytometry, and visualized apoptotic cells on slides by Hoechst bisbenzimide 33258 staining. The first observed effect at 30 min after treatment was an increase in annexin V-positive cells. At about 60 min the number of cells with moderate DNA migration increased in the comet-assay analysis. After 90 min, an increase in the number of cells with high levels of DNA migration was observed, which resulted in a bimodal distribution of cells with moderate and high levels of DNA migration. Hoechst-stained apoptotic cells could only be observed at later times (> or = 120 min). This means that the treatment would have been considered to be genotoxic if analysed at 120 min even if the cells with high levels of DNA migration would have been excluded. The occurrence of annexin V-positive cells preceded the appearance of cells with moderate levels of DNA migration. We hypothesize that these cells were early apoptotic cells and not indicative of genotoxic damage. We conclude that DNA-damaging effects of NaAsO2 cannot adequately be interpreted if the comet assay is not accompanied by separate analysis of early endpoints for induction of apoptosis.     

A DNA polymerase with unusual properties from the slime mold Physarum polycephalum

Two forms of a DNA polymerase have been purified from microplasmodia of Physarum polycephalum by poly(ethyleneimine) precipitation and chromatography on DEAE-Sephacel, phosphocellulose, heparin Sepharose, hydroxyapatite, DNA-agarose, blue-Sepharose. They were separated from DNA polymerase alpha on phosphocellulose and from each other on heparin-Sepharose. Form HS1 enzyme was 30-40% pure and form HS2 enzyme 60% with regard to protein contents of the preparations. Form HS2 enzyme was generated from form HS1 enzyme on prolonged standing of enzyme preparations. The DNA polymerases were obtained as complexes of a 60-kDa protein associated with either a 135-kDa (HS1) or a 110-kDa (HS2) DNA-polymerizing polypeptide in a 1:1 molar stoichiometry. The biochemical function of the 60-kDa protein remained unknown. The complexes tended to dissociate during gradient centrifugation and during partition chromatography as well as during polyacrylamide gradient gel electrophoresis under nondenaturing conditions at high dilutions of samples. Both forms existed in plasmodia extracts, their proportions depending on several factors including those which promoted proteolysis. The DNA polymerases resembled eucaryotic DNA polymerase beta by several criteria and were functionally indistinguishable from each other. It is suggested that lower eucaryotes contain repair DNA polymerases, which are similar to those of eubacteria on a molecular mass basis.     

Choosing the right path: enhancement of biologically relevant sets of genes or proteins using pathway structure

A method is proposed that finds enriched pathways relevant to a studied condition using the measured molecular data and also the structural information of the pathway viewed as a network of nodes and edges. Tests are performed using simulated data and genomic data sets and the method is compared to two existing approaches. The analysis provided demonstrates the method proposed is very competitive with the current approaches and also provides biologically relevant results.     

The intraovarian progesterone modulation of follicle development in the rabbit ovary

Intraovarian progesterone levels were manipulated by surgically adjusting the number of corpora lutea (CL) present in rabbit ovaries and this model was used to study the local effect of luteal progesterone on growth of follicles. The results show that when a single CL or several CL were present, follicle growth was inhibited. However, when all CL on one ovary were removed, increased numbers of follicles grew even when a single CL was present in the contralateral ovary. These findings show that progesterone inhibits follicle growth and that at least part of its action is local, i.e., exerted within the ovary. Additionally, ovarian blood vessels and periovarian lymph ducts were cannulated, and samples were collected and analyzed for steroid and protein content. The results show that when CL were present, ovarian vein progesterone levels were elevated 10-30-fold over levels in ovaries without CL; this high concentration points to the blood vascular system as the principal carrier of the steroid within the ovary. Analysis of lymph showed that protein content was consistently high and that the progesterone concentration was not significantly altered with the presence of CL; these two findings show that ovarian capillaries are extremely permeable to proteins, but the unexpectedly low concentrations of progesterone in lymph may signal an intraovarian countercurrent mechanism by which it is returned to the blood.