Introduction 1 Table 1 2–5 6 Table 1. Enzyme systems involving glutathione Enzyme system Function Glutathione synthetase Gamma-glutamyl cycle. Riboflavin-containing glutathione reductase Catalyzes the conversion of oxidized glutathione (glutathione disulfide; GSSG) to its reduced form. GSH transferase isoenzymes Conjugation of GSH with fat-soluble substances for liver detoxification and the detoxification of environmental carcinogens, such as those found in tobacco smoke. Selenium-containing glutathione peroxidase (GPX) Protects cells from hydrogen peroxides and lipid hydroperoxides. If not neutralized, these peroxides will damage cellular membranes and other vital cellular components. 4 4 4 4 4 6 7 8–14 15 16 17 P Given that inhalation is the only known method that increases GSH levels in the ELF for a significant duration, a review of the literature was conducted to examine the clinical effectiveness of inhaled GSH as a treatment for various pulmonary diseases and respiratory-related conditions. Only reports involving human subjects were included in the analysis. The clinical and theoretical indications for GSH inhalation were summarized and potential concerns with this treatment reported. Other pertinent details such as its presumed mechanisms of action and optimal doses to be administered were compiled and evaluated. Methods Literature Search Computer searches were conducted of English and non-English language articles in the Biomedical Reference Collection (1984 to August 2006), CINAHL (1982 to August 2006), MEDLINE (1965 to August 2006) and Nursing and Allied Health Collection (1985 to August 2006) databases. Articles were searched with the key search terms ‘Nebulized Glutathione,’ and ‘Glutathione’ in combination with ‘Aerosol’ OR ‘Inhalation.’ These keywords were also searched with words related to pulmonary and/or respiratory disease. To supplement the search, references of the articles found from the initial search were reviewed. Hand searching of relevant journals was also completed as part of the search. Selection of Articles To be included in the final review, articles had to report on the use and administration of inhaled GSH for pulmonary diseases and respiratory-related conditions in human subjects. Only peer-reviewed articles were reviewed. Quality Assessment Table 2 18 Table 2. Grades of evidence A Systematic reviews of randomized controlled trials and/or randomized controlled trials with or without double-blind placebo control. B Systematic reviews of observational studies and/or high-quality observational studies including cohort and case-control studies and/or cohort ‘outcomes’ research and/or nonrandomized controlled trials. C Case-series, case-reports, and/or poor-quality cohort and case-control studies. D Expert opinion without explicit critical appraisal or based on physiology, bench research or ‘first principles.’ Results 9 10 17 19–27 17 9 10 19–27 Table 3 Table 3. Summary of articles demonstrating the effectiveness of inhaled glutathione for the treatment of pulmonary diseases and respiratory-related conditions Reference Condition N Dosages of inhaled GSH Outcome Evidence grade 21 Asthma Eight asthma patients [mean age, 29 ± 7 (standard deviation; SD) years] 600 mg once weekly for 3 months A subset of patients with clinically stable mild asthma experienced a bronchoconstrictor effect when treated with inhaled GSH. A: Randomized placebo-controlled trial 23 Chronic otitis media with effusion (chronic OME) 30 patients (3–12 years of age; mean age, 5.8 years) and 30 controls (3–12 years of age; mean age, 6.1 years) 600mg of GSH in 4 ml of saline subdivided into five 2-min sessions by nasal aerosol every 3–4 waking h for 2 weeks GSH should be considered for the nonsurgical management of chronic OME. A: Randomized placebo-controlled trial 24 Cystic fibrosis (CF) Nine patients [mean age, 16.1 ± 1.44 (SD) years] received the S-nitrosoglutathione (GSNO) and 11 patients [mean age, 19.9 ± 3.45 (SD) years] received the phosphate- buffered saline (PBS) solution 0.05 ml/kg of 10 mM GSNO The treatment group showed a modest improvement in oxygenation that was thought to be independent of the physiological effects of nitric oxide. A: Randomized placebo-controlled trial 27 CF 19 patients (6–19 years of age) were randomized to treatment [mean age, 13.3 ± 4.1 (SD) years] or placebo groups [mean age, 12.9 ± 4.9 (SD) years] Total daily dose administered to the patients in the treatment group was 66 mg/kg of body weight GSH can improve clinical parameters in CF patients, and that effective treatment should include the correction of GSH deficiency. A: Randomized placebo-controlled trial 9 Idiopathic pulmonary fibrosis (IPF) 10 patients with IPF [mean age, 46 ± 3 (SD) years] and 19 normal nonsmokers [mean age, 36 ± 3 (SD) years] 600 mg twice daily for 3 days Inhaled GSH might be beneficial among IPF patients by reversing the oxidant–antioxidant imbalance. B: Nonrandomized controlled trial 19 Human immunodeficiency virus (HIV) seropositive individuals 14 HIV seropositive individuals [mean age, 32 ± 2 (SD) years] 600 mg twice daily for 3 days It is a reasonable therapeutic strategy to augment the deficient GSH levels of the lower respiratory tracts of HIV seropositive individuals. B: Cohort ‘outcomes’ research 20 Chronic rhinitis 13 patients with chronic rhinitis and 13 healthy subjects (4–15 years of age for all subjects; mean age, 8.2 years) 600 mg daily for 14 days Statistically significant improvement in nasal obstruction, rhinorrhea and ear fullness. B: Nonrandomized controlled trial 10 CF Seven CF patients [mean age, 25 ± 1 (SD) years] 600 mg of GSH for 3 days Inhalation therapy with GSH does normalize the respiratory epithelial surface oxidant–antioxidant balance in CF patients. B: Cohort ‘outcomes’ research 25 CF 21 patients with CF (16–37 years of age for all subjects) 300 or 450 mg three times daily for 14 days Inhaled GSH can permeate the lower airways of the lungs and improve important parameters of lung function in CF patients despite not having any effect upon markers of oxidative injury. B: Cohort ‘outcomes’ research 26 CF 17 patients with CF (18–29 years of age for all subjects; mean age, 24 years) 450 mg three times daily for 14 days Inhaled GSH did not affect the oxidative status of the patients who were tested, but it did favorably modulate their immune responses. B: Cohort ‘outcomes’ research 22 Emphysema One (95 year-old male) 120 mg of GSH in office, then 120 mg twice daily for 3 days, and continuation of treatment (dose unknown) for 2 years When the patient returned for a follow-up visit, he no longer required the use of his wheelchair and oxygen. The striking results were unexpected and unlikely to be due to placebo alone. C: Case report Discussion Based exclusively on the published evidence included in this review, inhaled GSH is potentially indicated for the following clinical conditions: cystic fibrosis (CF), chronic otitis media with effusion (OME), HIV seropositive individuals, idiopathic pulmonary fibrosis (IPF) and chronic rhinitis. These conditions were chosen since the published studies were of good quality, received A and B evidence grades, and their respective results demonstrated benefits from the use of GSH inhalation. 22 in vitro 28 29 30 21 31 32 33 34 35 36 Future Research Directions Aspergillus 11 11 37 38 39 40 N d 40 41 40 41 42 43 44 45 14 46 47 48 49 50 51 48 52 Mechanism of Action Fig. 1 9 10 19 24–27 20 23 Figure 1. 1  Considerations Prior to Initiating GSH Inhalation 53 54 A random (fresh) urine sample is suitable, but a first morning void may be preferable due to its higher concentration. Once the test strip is dipped in the urine (for 1 s), the reaction zone changes color to indicate the concentration of sulfites present. After 30 s, the color on the test strip is compared to a color scale on the bottle indicating the concentrations of sulfites in the urine (can detect 10, 40, 80, 180 and 400 ppm of sulfites). The resultant concentration should be multiplied by a factor of 1.5 to provide the amount of free sulfites in mg/l (ppm). The strip will not detect below 10 ppm. The urine samples should be preservative free, and the urinary pH should also be tested with pH paper. If the urine pH is below 6, then the amount of sulfites might be underestimated by the test. In such cases, consider adding sodium acetate or sodium hydroxide to raise the pH to at least 7–10 (should not exceed a pH of 12), and then repeat with a new test strip. Method of Delivery, Recommended Daily Dosages and Side Effects Table 3 27 In terms of side effects, GSH inhalation is very safe. Minor side effects such as mild coughing and an unpleasant odor were reported in some of the studies included in this review. These minor side effects, better described as mild nuisance problems, were not severe enough to cause any of the study participants to discontinue treatment with inhaled GSH. The only worrisome or potentially life-threatening side effect to note is bronchoconstriction, which would be more likely to occur among sulfite-sensitive asthma and MCSD patients. However, if proper precautions such as sulfite testing are done prior to treatment, this serious side effect should be avoidable. Monitoring the Clinical Response to Inhaled GSH For pulmonary diseases or respiratory-related conditions, baseline pulmonary function testing with a spirometer or a simple peak flow meter is recommended prior to the first treatment. After a prescribed period of treatment time, pulmonary function tests should be repeated. This will help to establish if there are any clinical improvements from regular GSH inhalation. Conclusions GSH inhalation is an effective treatment for a variety of pulmonary diseases and respiratory-related conditions. Even very serious and difficult-to-treat diseases (e.g., CF, IPF) yielded benefits from this novel treatment. GSH inhalation is very safe, and rarely causes major or life-threatening side effects. The potential applications are numerous when one considers just how many pulmonary diseases and respiratory-related conditions are affected by deficient antioxidant status, poor oxygenation and/or impaired host defenses. More studies are clearly warranted.