OAR@UM Collection:

A study of the antioxidant status in Maltese patients suffering from bronchial asthma

Sun, 01 Jan 1995 00:00:00 GMT

Title: A study of the antioxidant status in Maltese patients suffering from bronchial asthma Abstract: Asthma is an inflammatory condition in which several mediators interact to bring about a narrowing of the airways. During such an inflammatory reaction, eosinophils, monocytes, macrophages, lymphocytes and neutrophils all release reactive oxygen species. A free radical cascade ensues, producing a variety of oxidative products, main amongst which are superoxide, hydrogen peroxide, hydroxyl radicals and a range of organic hydroperoxides. These species are known to induce smooth muscle contraction, damage the respiratory epithelium, stimulate arachidonic acid metabolism and enhance the degree of airway hyperresponsiveness (Kanazawa et al., 1991). Defence against this destructive potential is afforded by an array of antioxidants, whose role, is to provide a route for the detoxification of these highly reactive moieties. Superoxide dismutase is responsible for the dismutation of the superoxide ion to hydrogen peroxide, while selenium-dependent glutathione peroxidases catalyse the reduction of hydrogen peroxide and organic hydroperoxides. These enzymes therefore play a pivotal role in modulating the damaging effect of reactive oxygen species on the airways. Based on these considerations, this study aimed to establish the antioxidant status of Maltese patients suffering from bronchial asthma, and to compare it to baseline values obtained from healthy controls. The study also investigated the effects which inhaled and oral glucocorticoid therapy may have on these defence systems. Plasma selenium, erythrocyte glutathione peroxidase, plasma glutathione peroxidase and erythrocyte superoxide dismutase, were measured in mild and severe asthmatics, as well as healthy controls. All patients also underwent spirometry. Blood sampling and pulmonary function testing were always carried out between 9. 00 am and 11. 00 am, in order to eliminate possible diurnal variation effects. In addition, the antioxidant profile of a group of 10 patients suffering from severe asthma was established prior to and after four weeks treatment with inhaled beclomethasone dipropionate (750μg b.d.). Similar investigations were carried out on 14 severe asthmatics prior to and after two weeks of oral prednisolone therapy (10-15 mg daily taken as a single morning dose). There were no gender or age dependent variations in any of the measured parameters in healthy volunteers. Plasma selenium levels were similar in both control (114.4 ± 3.5ng/ml, n = 49), mild asthmatic (116.4 ± 4.8ng/ml, n = 22) and severe asthmatic (114.0 ± 3.3ng/ml, n = 29) groups, (mean± SEM). However, a significant increase was observed in a group of severe asthmatics (n = 14) who were given oral prednisolone therapy, with mean levels rising from 115.7 ± 4.5ng/ml pre-treatment to 141.0 ± 4.4ng/ml afterwards (p < 0.005). Erythrocyte superoxide dismutase levels were significantly reduced in both mild (62.9 ± 2.9 SOD525U/ml, n = 22) and severe (60.6 ± 1.9 SOD525U/ml, n = 29) asthmatic patients, irrespective of therapy, when compared to control values ( 68. 5 ± 1.0 SOD525U/ml, n = 49) (p < 0.005). However, no changes were observed in either erythrocyte or plasma glutathione peroxidase. Administration of inhaled beclomethasone dipropionate did not exert any demonstrable effect on the antioxidant profile. Within the severe asthmatic group, plasma glutathione peroxidase was found to correlate with the degree of disease severity as quantified by FEV 1, PEF and FVC (p < 0.05). The same enzyme correlated with plasma selenium levels, within the control subject group (p < 0.05). The correlation of selenium with plasma glutathione peroxidase in control subjects, suggests a non-saturated selenium-dependent synthesis, or a selenium-dependent compartmentalisation of the enzyme. Correlation of the plasma enzyme with spirometric parameters within patients, might be the result of an oxidative stress dependent increased enzyme demand in the airways. This work appears to be the first to report the effects of low dose oral, as well as high dose inhaled glucocorticoids, on the antioxidant profile of patients suffering from bronchial asthma. Likewise, the correlation between plasma glutathione peroxidase and lung function parameters in severe asthmatics, does not appear to be stated in the literature. Description: M.PHIL.

Producing and evaluating a formulary for a community pharmacy

Sun, 01 Jan 1995 00:00:00 GMT

Title: Producing and evaluating a formulary for a community pharmacy Abstract: The formulary is set up in a local community pharmacy and aims to encourage rational, effective, safe prescribing. It limits extravagance by including drugs recommended by the specialities in a particular area. The formulary system is based on: • Flexibility by adapting immediately to ongoing needs of prescriber and patient • Willingness to accept change • Participation of the prescriber in the process of drug evaluation The formulary is set up for Regional Pharmacy, a typical pharmacy in Msida, Malta. The formulary is set up using the Database program Dbase 3 plus. Products included are medicines. diagnostic tests performed in the pharmacy, vaccines, baby care items, foot care items, medicated skin care items, wound management items, incontinency accessories, food supplements and dental care items. The formulary was first issued in November 1994 with 956 drugs and a total of 1094 items. The items were classified according to the shelf number and in order at trade name. The formulary includes the product formulation, shelf number and price of each item. Subsequently the formulary was issued in December, January and March. To date, the formulary is published every 3 months. Monthly Updates are produced whereby drugs included in the formulary, drugs deleted from the formulary and a list of drugs clue to expire in the next 3 months are brought to the attention of all pharmacists and prescribers. Description: M.PHIL.

Oxidative stress and expression of superoxide dismutase in escherichia coli and thermus aquaticus

Sun, 01 Jan 1995 00:00:00 GMT

Title: Oxidative stress and expression of superoxide dismutase in escherichia coli and thermus aquaticus Abstract: Escherichia coli is a facultative anaerobe and mesophile which produces two types of superoxide dismutase, iron (FeSOD) and manganese (MnSOD), the latter under the control of several global regulators of gene expression. Although much is known about the control ofMnSOD expression, FeSOD has always been assumed to be constitutively expressed. However, it has been suggested that the Fur (ferric uptake regulation) protein may be an inducer of this gene. Conflicting reports in the literature about the oxidative stress response of E.coli which overproduce plasmid-encoded FeSOD where addressed by assessing the types of plasmid used and, in particular, the type of promoter encoded by the plasmid. Strains which overproduced FeSOD unqer the control of the natural (wild-type) promoter were more sensitive to paraquat-induced oxidative stress than were wild type E. coli. Strains which overproduced F eSOD under the control of the artificial trc promoter were less sensitive to oxidative stress. Therefore the FeSOD promoter has a significant influence on the physiological response of the cell to oxidative stress and this result may be suggestive of a further mechanism of regulation. The gene encoding MnSOD from E. coli was cloned and expressed in several different E. coli strains. Tt was found that a manganese supplemented medium was required for the full activity of the enzyme, a result which explains other findings in the literature. In direct contrast, Thermus aquaticus is an obligate aerobe and thermophile which produces only MnSOD and although its gene has been cloned, nothing is known about the response of this important enzyme to oxidative stress in this organism. A bench-top 5 litre fermenter was installed as part of this work and facilitated experiments to investigate oxidative stress in Thermus aquaticus whereby pH and dissolved oxygen levels could be accurately monitored and controlled. A twofold increase in the expression and corresponding activity ofMnSOD was observed when Thermus aquaticus was subjected to oxidative stress using 200 µM paraquat. Surprisingly, no effect was observed when culture conditions included pure oxygen supplied at ambient pressure. A reduction in MnSOD activity was observed when using minimal broth which lacked manganese supplementation, an effect repeated when the iron concentration in the medium was increased. This is suggestive of an in vivo metal substitution in the MnSOD enzyme similar to that described for E. coli MnSOD. The fermentation technology employed to study T. aquaticus was used to investigate conditions for maximum expression and yield of recombinant human MnSOD which may in the near future gain pharmacological relevance. E. coli were used to express this protein under the control of the trc promoter and both maximal expression and cell yield were obtained using a glycerol and phosphate rich broth (terrific broth) Co-overexpression of the E. coli chaperonins GroES and GroEL from a second plasmid resulted in a slight (1.8 fold) increase in the measured specific activity of the SOD enzyme. This contrasts with the 3 fold increase previously observed with human copper-zinc SOD in the presence of these protein chaperones and may reflect a non-specific selectivity of these chaperones for β-sheet proteins (i.e. Copper-zinc SOD) over predominantly a-helical ones (i.e. MnSOD). During investigations on E. coli SOD expressiOn, a previously unreported band of SOD-like activity was observed only AFTER incubation of native polyacrylamide gels with hydrogen peroxide at 5 mM. Further investigation ruled out the possibility of an artefact due to buffers and solutions. Its significance remains enigmatic. Description: M.PHIL.