The alterations in the ethylene biosynthetic pathway provoked by cyclohexylamine (CHA) and/or methylglyoxal-bis(guanylhydrazone) (MGBG), inhibitors of the polyamine synthesis, have been studied in ...thermoinhibited seeds (delayed germination by supraoptimal temperatures) of the chick-pea (
Cicer arietinum). CHA and/or MGBG stimulated ethylene synthesis both at 25°C (control) and at 30°C (temperature at which thermoinhibition is induced, and reversed by both inhibitors). Moreover, both inhibitors caused: (1) an increase in ACC-synthase (EC 4.4.1.14) and free ACC; (2) a stimulation of in vivo ACC-oxidase; and (3) a stimulation of (malonyl)-1-aminocyclopropane-1-carboxylic acid synthesis at 25°C (since the supraoptimal temperatures had no effect). Compared with the control, both supraoptimal temperatures lead to a decrease the AdoMet-decarboxylase (AdoMetDC) (EC 4.1.1.50) activity. This was not affected by CHA. However, MGBG induced an increase of this activity at 25°C and at 30°C, which diminished strongly up to 35°C. The fact that the inhibition of spermidine and spermine synthesis by CHA and/or MGBG provoked an induction of the enzyme of ethylene biosynthesis and a concomitant increase in ethylene production and a reversion of thermoinhibition in chick-pea seeds suggests that the ethylene and polyamine pathways compete during germination, placing polyamines in the possible role of an endogenous germination regulator possibly by modulating ethylene biosynthesis.
To determine HCV and HGV replication sites in patients with chronic hepatitis C and to study interaction between these two viruses.
HGV RNA was studied in 272 patients with chronic hepatitis C. Of ...these, 35 were positive (group I). Twenty-three patients with chronic hepatitis C not co-infected with HGV were selected (group II).
Genomic and antigenomic chains of HCV were studied in both groups and those of HGV in group I in serum samples, peripheral blood mononuclear cells and liver tissue. In group I genomic chains of HCV and HGV were observed in 86 and 100%, respectively (ns), in serum samples (n = 35), and antigenomic chains in 17 and 23%, respectively (ns). In mononuclear cell samples (n = 15) 100% presented the genomic chain of HCV and 60% presented that of HGV (p < 0.05). Antigenomic chains were detected in 13 and 33%, respectively (ns). In liver tissue (n = 25) genomic chains were observed in 100 and 12%, respectively (p < 0.001); the antigenomic chain of HCV was detected in 76% while that of HGV was not present (p < 0.001). In group II genomic chains of HCV were found to be present in a very high percentage in all samples, while antigenomic chains appeared in 13% of serum and mononuclear cell samples and 89% of liver samples.
HCV and HGV have different sites of replication: whereas HCV replicates mainly in the liver, HGV is not hepatotropic. Mononuclear cells could represent a replication site for HGV but they are less important for HCV. Lastly, HGV does not modify the viral replication of HCV.
In the present work, certain biochemical characteristics of the enzyme 1-aminocyclopropane-1-carboxylate N-malonyltransferase (ACC N-MTase) which is responsible for the malonylation of ...1-aminocyclopropane-1-carboxylate (ACC) in chickpea (Cicer arietinum) are described. Phosphate buffer was the most appropriate buffer with regard to enzyme stability and, therefore, ACC N-MTase was extracted, assayed and purified in the presence of this buffer. ACC N-MTase was partially purified approximately 900-fold from embryonic axes of chick-pea seeds using ammonium sulphate precipitation, hydrophobic interaction and molecular filtration chromatography. By gel filtration chromatography on Superose-12, the molecular mass of the enzyme was estimated to be 54 ± 4 kDa. ACC N-MTase had an optimal pH and temperature of 7.5 and 40°C, respectively, as well as a Km for ACC and malonyl-CoA of 400 µM and 90 µM, respectively. D-Phenylalanine was a competitive inhibitor of ACC N-MTase with respect to ACC (Ki of 720 µM), whereas co-enzyme A was a competitive product inhibitor with respect to malonyl-CoA (Ki of 300 µM) and a non-competitive inhibitor with respect to ACC (Ki of 600 µM). Under optimal assay conditions, ACC N-MTase was strongly inhibited by (a)divalent Zn2+>Mg2+≫Co2+>Co2+>(NH4)2+>Fe2+ and monovalent metal cations (Li+>Na+>K+), without activity being detected in the presence of Hg2+, and (b) PCMB or mersalic acid, suggesting that sulphydryl group(s) are involved at the active site of the enzyme.
It is thought that the cytopathic effect of HGV is not important. Nevertheless, the cytopathic effect on liver is less known in the cases of co-infection with HCV. The aim was to study the prevalence ...of co-infection in patients with chronic hepatitis C (CHC) and to analyse the clinical-epidemiological and histological data and the interferon (IFN) response.
We included 180 patients with CHC and the HGV-RNA was determined.
The prevalence of co-infection was 12.2% (n = 22). No statistical differences were observed between the non co-infected and co-infected groups with regard to the age, sex, mechanism of transmission and alcohol abuse. Also, there were no differences in the hepatic biochemical, no organ-specific antibodies, histological lesions and Knodell index. The HCV biochemical response (BR) and virological response (VR) after 6 months post-IFN were the same in both groups (HGV negative: BR = 29%, VR = 12%; HGV positive: BR = 22%, VR = 18%). HGV was determined after 6 months posttreatment in the co-infected group (first cycle of IFN, n = 22; second cycle of IFN, n = 9): 12 (55%) were HGV-RNA negative and 5 (23%) HCV-RNA negative, (p = 0.021). When we compared the BR vs VR in this group, there were 12 HGV-RNA negative but only two had BR (NS). On the contrary, the BR was related to HCV-RNA negative (p = 0.023).
The prevalence of HGV co-infection is important in our area (12.8%). The HGV does not increase the pathogenicity of HCV and does not change the IFN response, although the HGV is more IFN sensible than HCV. The determination of HGV is not necessary in patients with HCV.
Recently the parenteral transmission of hepatitis G virus (HGV) has been shown. The aim of the study was to investigate the incidence of post-transfusion HGV.
HGV (RNA-HGV and anti-HGVE2) were ...retrospectively studied in 140 transfused patients.
12 (8.6%) were infected after transfusion: 9 of 12 (75%) the RNA-HGV remained detectable after 6 months and 3 (25%) seroconverted to anti-HGVE2. No patient had post-transfusional hepatitis criteria. In 5 (42%) the transaminases levels were slightly increased. The clinical evolution was favourable. No significant differences were found between patients with or without HGV infection.
HGV is an agent associated with transfusion but it carries a low pathogenic capability.
Treatment with cyclohexylamine, an inhibitor of spermidine synthase, accelerated radicle emergence in chick-pea (Cicer arietinum L. cv. Castellana) seeds. Stimulation in the growth of embryonic axis ...was correlated with: (a) a rise in putrescine (Put) and cadaverine (Cad); (b) a decrease in spermidine (Spd) and spermine (Spm), and (c) a concomitant acceleration of the transformation from S-adenosylmethionine (SAM) into ethylene with increases in the levels of l-aminocyclopropane-l-carboxylic acid (ACC), I-(malonylamino)cyclopropane-l-carboxylic acid (mACC) and ethylene and ACC synthase and ethylene-forming enzyme activities. Cyclohexylamine also stimulated the mitotic index in both apical and subapical zones of the radicle and the apical zone of the plumule. The sectional distribution of the ethylene pathway and polyamine content was studied in embryonic axes of seeds germinated for 65 h. Each axis was divided into 5 sections: radicle meristem, elongation zone, differentiation zone, hypocotyl and plumule. In the presence of cyclohexylamine, ACC synthase and ACC were strongly stimulated in both the differentiation and hypocotyl zones, whereas the mACC was stimulated in all sections of the embryonic axis. With respect to ethylene-forming enzyme activity and ethylene production, the hypocotyl and the zones of elongation and differentiation were affected most by cyclohexylamine. Cyclohexylamine also induced an accumulation of free Put and Spm in the differentiation, hypocotyl and radicle zones, whereas Put and Spm bound to small substances increased most in the hypocotyl and plumule. The Spd bound to small substances decreased in all sections in the presence of cyclohexylamine. With respect to polyamines bound to macromolecules, cyclohexylamine stimulated only the accumulation of Put since Spd and Spm were strongly inhibited in all sections.
For a deeper understanding of the germination of chick–pea (Cicer arietinum) seeds, which is dependent upon ethylene synthesis, a crude extract containing authentic ACC oxidase (ACCO) activity was ...isolated in soluble form from the embryonic axes of seeds germinated for 24 h. Under our optimal assay conditions (200 mM HEPES at pH 7.0, 4μM FeS04, 6 mM Na–ascorbate, 1 mM ACC, 20% 02, 3% CO2 , and 10%glycerol) this enzyme was 5–fold more active than under the conditions we used initially in the present work. The enzyme has the following Km: 28 μM for ACC (approximately 4–fold less than in vivo), 1.2% for O2 (in the presence of an optimal CO2 concentration of 3%), and 1% for CO2 in the presence of O2 (20%). The enzyme is inhibited by phenanthroline (PNT) (specific chelating agent of ferrous ion), and competitively inhibited (K1, =0.5 mM) by 2–aminoisobutyric acid (AIB), and the enzymatic activity was not detectable in the absence of CO2. Under optimal assay conditions, the enzyme has two optimum temperatures (28 °C and 35 °C) and is inhibited by divalent metal cations (Zn2+> CO2+>Ni2+>Cu2+>Mn2+ >Mg2+) and by salicylic acid, propylgallate, carbonyl cyanide m–chlorophenyl hydrazone (CCCP), dinitrophenol (DNP), and Na–benzoate. The in vitro ACCO activity which we recovered in soluble form is equivalent to approximately 80–85% of the apparent activity evaluated in vivo.
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