Ozone in Medicine. The Low-Dose Ozone Concept and Its Basic Biochemical Mechanisms of Action in Chronic Inflammatory Diseases

Abstract

Low-dose ozone acts as a bioregulator in chronic inflammatory diseases, biochemically characterized by high oxidative stress and a blocked regulation. During systemic applications, "Ozone peroxides" are able to replace H₂O₂ in its specific function of regulation, restore redox signaling, and improve the antioxidant capacity. Two different mechanisms have to be understood. Firstly, there is the direct mechanism, used in topical treatments, mostly via radical reactions. In systemic treatments, the indirect, ionic mechanism is to be discussed: "ozone peroxide" will be directly reduced by the glutathione system, informing the nuclear factors to start the regulation. The GSH/GSSG balance outlines the ozone dose and concentration limiting factor. Antioxidants are regulated, and in the case of inflammatory diseases up-regulated; cytokines are modulated, here downregulated. Rheumatoid arthritis RA as a model for chronic inflammation: RA, in preclinical and clinical trials, reflects the pharmacology of ozone in a typical manner: SOD (superoxide dismutase), CAT (catalase) and finally GSH (reduced glutathione) increase, followed by a significant reduction of oxidative stress. Inflammatory cytokines are downregulated. Accordingly, the clinical status improves. The pharmacological background investigated in a remarkable number of cell experiments, preclinical and clinical trials is well documented and published in internationally peer reviewed journals. This should encourage clinicians to set up clinical trials with chronic inflammatory diseases integrating medical ozone as a complement.

Keywords: biological medicine; bioregulation; chronic inflammation; complementary medicine; ozone therapy; redox balance.

Figure 7

Pharmacological effects of ozone: signal transduction via glutathione and “ozone peroxides”. An intact GSH/GSSG balance is the limiting factor for the ozone concentration and the basis for the low-dose ozone concep revision from [23].

Figure 8

(a) The low-dose ozone concept and the regulation of antioxidants, started through the reduction of “ozone-peroxides” by the active (reduced) form of glutathione GSH, initating the signal transduction via the Keap 1/Nrf2–complex and translocation of Nrf2 into the nucleus. In close connection with proteins such as Maf and the ARE complex (antioxidant response elements) the transcription of genes, translation and finally the antioxidant synthesis proceed, see also [15,17,19]. (b). Schematic survey of cytokine induction through ozone in mononuclear cells. In healthy organisms, H₂O₂ acts as a messenger substance in chronic inflammation where the regulation is out of balance. It is most likely to be “ozone peroxide” replacing H₂O₂ and restarting the regulation acc. to [14,19].

Figure 1

(a) Hydrogen peroxide as key redox regulator in the biological system modified acc. to [1,2]. (b). Low-dose ozone as bioregulator.

Figure 2

(a) [9] Major autohemotherapy. Standardized system acc. to the Medical Device Directives MDD 93/42EC. 1. Ozone syringe (PP, siliconized). 2. Bacterial filter 0,2 µ (germstop). 3. Vacuum bottle (glass) with micro bubble system (PP) 4. Transfusion set (PE). 5. Latex-free plug. (b) [9] Set for rectal insufflation, standardized system acc. to the Medical Device Directives 93/42 EC. 1. Dosage pump (silicone). 2. storage bag. 3. Clamp. 4. one-way valve. 5. connection line (PE). 6. catheter (PE).

Figure 3

(a) Molecular structure. Radical and ionic reaction mechanisms of ozone modified after [9]. (b) Ozone reaction with isolated double bonds, i.e., ozonolysis according to Criegee 1953, 1975 [11,12]. The hydroxy hydroperoxides, here, “ozone peroxides” are understood as the pharmacologically active substances.

Figure 4

Direct and indirect ozone reactions in topical and systemic treatment within the corresponding concentration ranges.

Figure 5

The pharmacological effects of medical ozone via ozone-produced peroxides. The three effects: 1. improved oxygen release by the red blood cells (RBC) through activation of RBC metabolism. 2. Immunomodulation through activation of the white blood cells (WBC) and signal transduction via nuclear factors. 3. Regulation of cellular antioxidants via Nrf2 signalling [9].

Figure 6

Dose response: efficacy–concentration relationship of systemically administered medical ozone as hormetic substance (schematically), modified after [9].

Figure 9

(a) Development of the total arthritis index (joint swelling) over 35 days in a preclinical study (n = 20) on rheumatoid arthritis RA in rats under intraarticular ozone treatment 3× per week, 20 µg/mL, 0.2 mL, starting on day 10. Control (n = 5): intraarticular needle stress 3× per week. PG/PS-induced rheumatoid arthritis (n = 15) (peptidoglycan/polysaccharide). PG/PS + ozone (n = 5): ozone treatment starting on day 10. PG/PS + oxygen (n = 5): oxygen (0.2 mL) treatment starting at day 10 [25]. (b) The antioxidant capacity, here as superoxide dismutase SOD in the spleen of RA-rats after 10 intraarticular ozone treatments compared to oxygen treatment and control as described in Figure 9a [25]. (c) Oxidative stress, measured as NO in the rat spleen following 10 intraarticular ozone treatments compared to oxygen and control as described in Figure 9a [25]. (d) Downregulation of inflammatory stress parameters TNF-α mRNA and IL-1β mRNA, measured in arbitrary units in the rat spleen following 10 intraarticular ozone treatments compared to oxygen as described in Figure 9a [25].

Figure 9

(a) Development of the total arthritis index (joint swelling) over 35 days in a preclinical study (n = 20) on rheumatoid arthritis RA in rats under intraarticular ozone treatment 3× per week, 20 µg/mL, 0.2 mL, starting on day 10. Control (n = 5): intraarticular needle stress 3× per week. PG/PS-induced rheumatoid arthritis (n = 15) (peptidoglycan/polysaccharide). PG/PS + ozone (n = 5): ozone treatment starting on day 10. PG/PS + oxygen (n = 5): oxygen (0.2 mL) treatment starting at day 10 [25]. (b) The antioxidant capacity, here as superoxide dismutase SOD in the spleen of RA-rats after 10 intraarticular ozone treatments compared to oxygen treatment and control as described in Figure 9a [25]. (c) Oxidative stress, measured as NO in the rat spleen following 10 intraarticular ozone treatments compared to oxygen and control as described in Figure 9a [25]. (d) Downregulation of inflammatory stress parameters TNF-α mRNA and IL-1β mRNA, measured in arbitrary units in the rat spleen following 10 intraarticular ozone treatments compared to oxygen as described in Figure 9a [25].

Figure 10

(a) Clinical trial in Rheumatoid arthritis RA (n = 60) following 20 rectal ozone insufflations with ozone conc.: 25–40 µg/mL and 5000–8000 µg per treatment during 4 weeks. MTX-group (n = 30): basic therapy MTX+Ibuprophen+Folic acid. Ozone-group (n = 30): basics as MTX-group plus ozone rectally. SOD: superoxide dismutase, CAT: catalase, GSH: reduced glutathione [26]. (b). Oxidative stress markers NO: nitrogen monoxide (µMol), AOOP: advanced oxidated proteins (µMol), TH: total hydroperoxide (µMol), MDA: malondialdehyde (µMol) [26]. (c). Disease activity score following the EUROPEAN LEAGE AGAINST RHEUMATISM in patients with rheumatoid arthritis treated with standard therapy (MTX-group, n = 30) and standard therapy plus rectal ozone insufflation (n = 30). DAS-28: low activity y ≤ 3.2; moderate activity 3.2 < y ≤ 5.1; high activity y ≤ 5.1 [26]. (d). Pain score in 60 RA-patients. MTX-group (n = 30) with basic treatment (see Figure 10a) and ozone group (n = 30) with standard therapy plus ozone at the beginning and at the end of the trial after 20 rectal insufflations [26].

Figure 10

(a) Clinical trial in Rheumatoid arthritis RA (n = 60) following 20 rectal ozone insufflations with ozone conc.: 25–40 µg/mL and 5000–8000 µg per treatment during 4 weeks. MTX-group (n = 30): basic therapy MTX+Ibuprophen+Folic acid. Ozone-group (n = 30): basics as MTX-group plus ozone rectally. SOD: superoxide dismutase, CAT: catalase, GSH: reduced glutathione [26]. (b). Oxidative stress markers NO: nitrogen monoxide (µMol), AOOP: advanced oxidated proteins (µMol), TH: total hydroperoxide (µMol), MDA: malondialdehyde (µMol) [26]. (c). Disease activity score following the EUROPEAN LEAGE AGAINST RHEUMATISM in patients with rheumatoid arthritis treated with standard therapy (MTX-group, n = 30) and standard therapy plus rectal ozone insufflation (n = 30). DAS-28: low activity y ≤ 3.2; moderate activity 3.2 < y ≤ 5.1; high activity y ≤ 5.1 [26]. (d). Pain score in 60 RA-patients. MTX-group (n = 30) with basic treatment (see Figure 10a) and ozone group (n = 30) with standard therapy plus ozone at the beginning and at the end of the trial after 20 rectal insufflations [26].

Figure 11

Liver toxicity of standard antirheumatic drugs is reduced by low-dose ozone, here shown as γ-GT (γ-glutamyl transferase) in a clinical trial with 100 Rheumatoid arthritis patients. MTX-group (n = 50): Metotrexate + Ibuprophen + Folic acid. Ozone-group (n = 50): MTX group + ozone. 20 rectal ozone insufflations 200 mL, concentrations 25–40 µg/mL during 4 weeks [27].