Enzyme-catalysed non-oxidative decarboxylation of aromatic acids: I. Purification and spectroscopic properties of 2,3 dihydroxybenzoic acid decarboxylase from Aspergillus niger

In order to understand the molecular mechanism of non-oxidative decarboxylation of aromatic acids observed in microbial systems, 2,3 dihydroxybenzoic acid (DHBA) decarboxylase from Aspergillus niger was purified to homogeneity by affinity chromatography. The enzyme (Mr 120 kDa) had four identical subunits (28 kDa each) and was specific for DHBA. It had a pH optimum of 5.2 and Km was 0.34 mM. The decarboxylation did not require any cofactors, nor did the enzyme had any pyruvoyl group at the active site. The carboxyl group and hydroxyl group in the ortho-position were required for activity. The preliminary spectroscopic properties of the enzyme are also reported.     

Usefulness of the prevention of oxygen radical damage in the critical patient using the parenteral administration of reduced glutathione in high doses]

A hyperproduction of Oxygen Free Radicals (FRO) is frequently observed during stress, anoxia, hyperbarism and may worsen the clinical conditions of intensive care patients. The hyperproduction of FRO may be reduced by antioxidants. The glutathione (GSH) is frequently associated with organic antioxidant protective systems. Forty patients receiving a continuous infusion of 70 mg/Kg/die of GSH were compared with forty patients not receiving GSH; the patients in both groups were randomized for age, sex, and pathology. Some parameters which are indirect indexes of FRO hyperproduction were chosen: ethane in the expired air, plasma malondialdehyde, fibrinopeptide A and C5 activated complement fraction, also the erythrocyte membrane deformability was investigated. The results obtained in the group receiving GSH were compared with the control group and a significative difference was found indicating a reduced FRO production. These interesting results need more trials in order to confirm a real GSH involvement in the antioxidant organic protection. In any case the supplementation with antioxidants in the therapy of intensive care patients can be regarded as an interesting means to improve their clinical conditions.     

HLA antigens, blood groups and serum protein groups in patients with intermittent claudication

HLA (A and B) antigens, blood group systems (AB0, Rh, MNSs P, Kell, Lewis and Duffy) and serum group systems (Hp, Tf, Pi, C3 and C4) were studied in patients with intermittent claudication (IC) and controls. HLA antigen A 28 was significantly more common, and blood group 0 was significantly less common among the patients than among the controls. A comparison between patients with IC and those with abdominal aortic aneurysms showed a significant difference between these two groups concerning the MN blood groups.     

Correlation of heredity and environmental factors in the etiology of Vilyui encephalomyelitis. II. A population genetic study in districts of Vilyui encephalomyelitis distribution]

Population-genetic investigation was carried out in the regions endemic for Viljuisk encephalomyelitis (VE). The following indices were estimated: the relationship coefficient, the inbreeding coefficient, the intensity of migration, genetic structure of the population. The aim of the investigation was to explain causes of intrapopulational and, in some cases, of intragenus accumulation of the VE patients. No evidence of isolation or increased inbreeding were found in highly affected populations. The genetic structure of a group of VE patients tested for 9 polymorphic systems did not reveal any deviation from the control group. Several VE cases in healthy populations occured in some years after the immigration of a VE patient. These data confirm the hypothesis that VE is transmitted from a VE patient to healthy persons. Among these persons fall ill those who have a hereditary determined increased sensitivity to VE which results in intrapopulation and intragenus accumulation of VE cases.     

Genetic markers in patients with intracranial aneurysms

HLA antigens, blood group systems (ABO, Rh, MNSs, P, Kell, Lewis and Duffy) and serum group systems (Hp, Tf, Gc, Pi, Bf, C3 and C4) were studied in a series of patients with intracranial aneurysms. A significantly increased frequency of HLA antigen A28, a significantly decreased frequency of HLA antigen B40, and a significantly decreased frequency of complement factor C4 B2 was found among the patients when compared with controls from the same geographic area.     

Thirty-four year follow up of patients with breast cancer in clinical trial of postoperative radiotherapy.

An extended follow up to a maximum of 34 years was carried out on a series of 1461 patients who had been entered into a randomised clinical trial comparing immediate postoperative radiotherapy (treated group) with an optional delayed radiotherapy policy (watched group) after Halsted radical mastectomy for operable breast cancer. Long term survival was compared separately in patients with negative and positive axillary nodes and in two periods when different techniques of radiotherapy were used. No significant differences were noted in survival overall or in any subgroup. In patients with negative axillary nodes treated in the earlier period when the "quadrate" radiotherapy technique was used, however, the difference in survival was of borderline significance (p = 0.052), to the benefit of the watched group. Because of the large numbers of patients entered and the long follow up any possible harmful or beneficial effect of immediate postoperative radiotherapy would have been apparent in the results of this trial. The absence of any such effect implies that postoperative radiotherapy given to prevent or delay local recurrence of breast cancer is unlikely to be associated with a detectable effect on survival.     

High levels of extra-group paternity in a population of Australian magpies Gymnorhina tibicen: evidence from microsatellite analysis

Breeding systems vary widely in birds, from monogamous pairs through to complex group systems where subordinates assist breeding individuals to rear young each season. The Australian magpie varies geographically both in plumage patterns and social organization. Some populations of both eastern and western plumage forms are plural breeders with group size varying from three to over 15 mature individuals. This study used variation at microsatellite loci to determine the level of extra-group paternity in a population of the western form near Perth in Western Australia. Extra-group paternity was the highest recorded for any bird species to date (82%) and indicates that few offspring within a territory are sired by the social partner of the female. In addition, the data indicated that nearly 10% of juveniles were not the genetic offspring of any female within their territory, suggesting some intraspecific brood parasitism. Taken together, these findings are remarkable considering the highly territorial nature of the species and the extent of territorial defence practised by all members of the group towards extra-group conspecifics during daylight hours.     

The humoral immune response and the productivity of laying hens kept on the ground or in cages

The effects of two different keeping systems on the humoral immune response and productivity were compared for 80 laying hens, divided into four groups. Two groups each of 20 hens were kept on the ground and two were kept in cages. All the birds were immunised subcutaneously with human serum immunoglobulin G (IgG) at a dose of 100(microg per injection. The immunisations were performed twice at 4-week intervals. The lipopeptide Pam(3)Cys-Ser-(Lys)(4) was used as an adjuvant at a dose of 0.25mg per injection in one group from each housing system. In the second group from each housing system, the hens were immunised without any adjuvant (antigen control groups). The mean egg yield was significantly higher in both the antigen control group and the adjuvant group, when laying hens were kept in cages. Total egg weight remained constant in both of the housing systems. Keeping hens in cages resulted in higher mean specific antibody titres and mean immunoglobulin Y concentrations in the egg yolk.     

Relevance of TSH receptor stimulating and blocking autoantibody measurement for the prediction of relapse in Graves' disease.

Recently, we demonstrated that higher levels of autoantibodies to the human TSH receptor (TBII) predict relapse of hyperthyroidism in Graves' disease (GD). The aim of this study was to extend this outcome prediction by dividing TBII into stimulating (TSAb) and blocking (TBAb) TSH receptor autoantibodies. Altogether, ninety patients (81 female, 9 male) were retrospectively analyzed; sixty-four patients (71 %) did not go into remission or relapsed, whereas twenty-six patients (29 %) went into remission (median follow-up: 17.5 months). TSAb and TBAb measurement was performed in a CHO cell bioassay with cAMP readout at the time of their first visit in our outpatient clinic (single point measurement in median 6.5 months after initial diagnosis). In the remission group, eighteen of twenty-six patients (69 %) were TSAb-positive, whereas fifty-three of sixty-four patients (83 %) were TSAb-positive in the relapse group (p = ns). The mean stimulation indices (SI) were 4.1 in the remission group and 12.9 in the relapse group, respectively (p = 0.015). By using a threshold of 10 SI, the specificity for relapse was 96.0 %, as only one in twenty patients with an SI above 10 went into remission during follow-up (PPV 95 %). Most TSAb-positive patients also had high levels of TBII. Neither group showed any difference with respect to blocking type autoantibodies, which were mostly negative in both groups. In summary, high TSAb levels are similar but not superior to TBII for predicting relapse in GD patients. In contrast, TBAb measurement does not add any valuable information in this context. In the clinical routine, TSAb/TBAb measurement may not play an important role for diagnosis or outcome prediction of GD, since sensitive 2 (nd) generation TBII assays are easier to perform and offer similar information to the clinician. Bioassays should be reserved for special clinical questions such as Graves' disease in pregnancy.     

Site-specific mutagenesis in cells with normal DNA repair systems: transitions produced from DNA carrying a single O6-alkylguanine.

This paper describes a systematic study of transition frequencies produced in vivo when a homologous series of O6-alkylguanine residues located at a preselected position in gene G of phi X174 form I' DNA (double-stranded, circular, covalently-closed, relaxed) is transfected into spheroplasts from two strains of Escherichia coli having normal DNA repair systems. Mutant frequencies were measured as percent of total phage produced by single bursts. The results are: (A) Synthetic DNA without any alkyl group gave a transition frequency of 0.02%. (B) In E. coli AB1157, the frequencies fall into two groups depending on the alkyl group: methyl and ethyl, 8-11%; n-propyl and n-butyl approximately 0.9%. (C) The average transition frequencies were higher in AB1157 than in C600. These data demonstrate that a single O6-alkylguanine residue can produce a specific transition at significant frequencies in cells with normal repair systems and that the mutant frequency depends upon the nature of the alkyl group and the cell type.     

Synthesis, antiviral and cytostatic activities, of carbocyclic nucleosides incorporating a modified cyclopentane ring. IV. Adenosine and uridine analogues

Eight new carbocyclic nucleosides were prepared by mounting a purine (compounds 8-10), 8-azapurine (12 and 13) or pyrimidine (15, 16 and 17b) on the amino group of (1S,3R)-3-aminomethyl-2,2,3-trimethylcyclopentylmethanol (6). All the compounds were evaluated as antiviral and antitumor agents in a variety of assay systems. Only compound 8 showed any cytostatic activity against the tumor cell lines examined.     

Effect of Electromagnetic Stimulation on Patients Suffering from Non-Union. A Retrospective Study with a Control Group

In this study we have reviewed a group of patients suffering from non-union treated at the same hospitals by the same surgeons. Twenty six patients (control group) received an orthopaedic treatment aimed at promoting bone healing, fourty one patients (experimental group) were stimulated with an electromagnetic field generator without any further change in the orthopaedic management of the non-union. Eighteen patients healed in the control group and thirty six in the experimental group. The average healing time was shorter among patients stimulated. No difference was observed in the success rate between the 2 groups of patients for non infected non-unions. Infection proved to be detrimental for the success of the treatment in the control group, while it did not negatively influence the outcome of stimulation. Provided that the orthopaedic treatment of the non-union is considered satisfactory, electromagnetic stimulation is a valid option in the treatment of non-unions.     

Impaired Glucose Tolerance: A Late Effect of Insulin Shock Treatment

Glucose tolerance tests were performed in a group of patients in a mental hospital who had been treated with insulin shock and in a matched control group. Five out of 31 (16%) patients in the insulin-treated group and 1 out of 22 (5%) controls had “diabetic” blood sugar curves. Median blood sugar values were significantly higher at 60 minutes and later in the insulin-treated group, as were mean blood sugar values when the “diabetic” patients were excluded. Small amounts of plasma insulin-binding antibody were found in two insulin-treated patients. In the absence of any other clear-cut explanation, it is suggested that in some patients massive doses of insulin by injection may leave diminished tissue responsiveness to insulin as a long-term after-effect.     

Plasma cholinesterase activity in patients with uterine cervical cancer during radiotherapy

The objectives of this study were to investigate: 1) the activity of pseudocholinesterase (PChE) in patients with uterine cervical cancer in different stages (uterine cervical carcinoma in stages II b and III and recurrent cervical carcinoma in stages III and IV a,b) and to compare it to the enzyme activity in patients with benign tumour of the uterus, and 2) the effects of radiotherapy on enzyme activity in those patients with uterine cervical carcinoma for which the chosen treatment was radical radiotherapy. Thirty patients with uterine cervical carcinoma in stages II b and III (Group A), sixteen patients with recurrent cervical carcinoma in stages III and IV a,b (Group B) and thirty-eight patients with benign tumours of the uterus (control, Group C) were evaluated and their PChE activity was determined prior to any treatment (pre-therapy enzyme activity). All eighty-four patients were free of any liver disease. The results have shown that the patients of Group A had the pre-therapy PChE activity practically identical to those in group C, but patients of Group B had significantly lower values of PChE with respect to enzyme activities of Groups A and C (p < 0.001). That is to say, PChE activity was influenced by the extent to which the malignancy had spread. Radical radiotherapy (up to 8 weeks in doses higher than 50 Gy into point A; average 80 Gy) which was the chosen treatment only for patients from group A did not cause a significant inhibition of PChE activity in any patients in comparison with their control values. With regard to the role of PChE in hydrolysis of succinylcholine, our results about the influence of the malignant disease and the radiotherapy on PChE activity are clinically significant.     

Parentage analysis within a semi-free-ranging group of Barbary macaques Macaca sylvanus

This study of a group of semi-free-ranging Barbary macaques Macaca sylvanus aimed to determine paternity, to establish whether any individual male achieved prominent mating success and to assess genetic variability. Analyses involved electrophoresis of 15 blood protein systems and multilocus DNA fingerprinting (isotopic and nonisotopic). Genetic variability was low; only two blood protein systems were polymorphic. Although all DNA-fingerprints were individual-specific, they showed a high average band-sharing index value (0.67). Nevertheless, a combination of all methods permitted inference of paternity in 11 out of 15 (73%) cases tested. Several males from different age classes fathered infants.     

Polymorphisms of hemochromatosis, and alpha-1 antitrypsin genes in Egyptian HCV patients with and without hepatocellular carcinoma

Hereditary hemochromatosis and alpha-1antitrypsin deficiency are genetic diseases characterized by endoplasmic reticulum (ER) stress with subsequent development of liver disease. Our aim was to estimate the frequency of hemochromatosis gene (HFE) mutant alleles (C282Y and H63D) and alpha-1 antitrypsin S/Z variants among Egyptian HCV cirrhotic patients and in hepatocellular carcinoma patients and to evaluate their effects on disease progression. HFE and alpha-1 antitrypsin polymorphisms were characterized in 200 Egyptian patients with HCV infection (100 patients complicated with cirrhosis, 100 patients with HCC) and 100 healthy subjects who had no history of any malignancy. The frequencies of HD genotype of H63D mutation were significantly increased in HCC patients compared to control group and to cirrhosis group. Also, the frequencies of DD genotype were significantly increased In HCC group compared to control group and to cirrhosis group. Our results suggested that Carriers of the D allele of H63D mutation were significantly more likely to develop HCC.     

Effect of toremifene in breast cancer patients. Preliminary communication

Toremifene, an antiestrogenic drug administered at three dose levels (60, 120, and 300 mg/day) was investigated in 17 postmenopausal patients with advanced breast cancer previously treated with hormonal and/or cytostatic therapy. The drug proved to be well tolerated at all dose levels without any serious side effects even on prolonged administration. Neither response nor side effects have shown any dose dependency in this small group of patients.     

Cross sensitivity of mutator strains to physical and chemical mutagens.

Ten different mutator strains of Saccharomyces cerevisiae were tested for cross sensitivity to two alkylaitng agents, ethylmethanesulfonate (EMS) and methylmethanesulfonate (MMS), to determine if any of them are defective in the repair systems which normally deal with damage caused by these agents. For one of the mutators, namely mut2-1, it was shown by genetic analysis that mutator activity and MMS sensitivity are both controlled by the same gene. Two mutants, mut2-1 and mut7-1, were found to be sensitive to MMS but normal to ultraviolet and gamma-rays. Another group is represented by mut1, mut6 and mut8 which are not sensitive to any of the mutagens tested so far. Mutator strain mut2-1 was also shown not to be significantly altered for levels of UV-induced forward and reverse mutations. These observations lend support to the idea of multiple repair systems that deal with DNA damage caused by different agents and also show that mutator activity can often result from the loss of normal cellular repair systems.     

Biocompatibility and tissue interactions of a new filler material for medical use

Filler materials for medical use present limits, such as the induction of chronic inflammation and fibrosis. In the search for synthetic materials with improved biocompatible properties, a new polyacrylamide hydrogel, Aquamid (Contura SA, Montreux, Switzerland), has been investigated in preclinical systems. In cell cultures (endothelial cells and fibroblast), no or only transient biological effects were associated with 10% Aquamid exposure. The Aquamid-host interactions were examined in mice (10 mice per group) implanted subcutaneously or in the mammary fat pad with a very large volume (1.5 ml) of the material. Blood analysis, performed after 15 and 94 days (five mice per time for each group) to detect acute or late manifestations of toxicity, did not reveal relevant abnormalities in either group of Aquamid-bearing mice compared with control mice, except for a transient thrombocytopenia and a mild leukocytosis. Histological analysis of the pellet showed the presence of a thin, poorly vascularized cyst wall in implants. Only mild mesenchymal reparative and inflammatory processes were observed, even at longer observation times (more than 400 days). No alterations in any organ were detected. Despite the large volume implanted (approximately 5 percent of mouse body weight), the Aquamid pellet maintained its original size and shape without spreading or sticking to surrounding tissues. In conclusion, the study indicated a good tolerability of the new biopolymer in preclinical systems. The clinical utility of this new compound, if confirmed by clinical randomized trials showing its atoxic properties, could be in the field of aesthetic plastic surgery as a filler material for body contouring and in reconstructive surgery and above all in cancer patients to restore surgical defects.