2020年Am J Case Rep；21：e925849 DOI：10.12659 / AJCR.925849
专科：感染性疾病 - 肺病学
Ozone (O3 ) is a gas composed of 3 atoms of oxygen, including a stable pair (O2 ) and a third, unstable, atom, which gives ozone its beneficial effects. Medical ozone generators produce 1–5% ozone gas in 95–99% oxygen from pure oxygen. For medical purposes, concentrations of 10–70 μg/mL are commonly used.Ozone therapy can be administered systemically by adding it to a sample of a patient’s own blood and then reinfusing it, in what is termed “ozonated autohemotherapy.” When blood is exposed to this gas mixture (O2 –O3 ), oxygen equilibrates with the extracellular and intraerythrocytic water before becoming bound to hemoglobin until it is fully oxygenated; in contrast, ozone, which is about 10 times more soluble than oxygen, allows its immediate reaction with any soluble compounds and biomolecules present in biological fluids, readily dissolves in water, and reacts instantaneously with several biomolecules such as ascorbic acid, urate, free cysteine, glutathione molecules, and albumin thiol groups, and then disappears. The compounds generated during the reactions (reactive oxygen species and lipid ozonation products) are the “ozone messengers” and are responsible for its biological and therapeutic effects. O2 –O3 therapy has many beneficial effects, including inactivation of microorganisms such as viruses, modulation of the immune system, improvement of microcirculation, anti-inflammatory action stimulation of oxygen metabolism, and promotion of tissue oxygenation.
三氧(O3)是一种由3个氧原子组成的气体，包括可稳定的对(O2)和第三个不稳定的原子，这使三氧具有有益的作用。医用臭氧发生器从95-99%的纯氧中产生1-5%的三氧气体。在医疗上，常用浓度为10-70μg/mL。三氧治疗可以通过将其加入到患者自身的血液样本中，然后再注入，即所谓的 "三氧自体血疗法"，进行系统性的治疗。当血液暴露在这种混合气体(O2 -O3 )中时，氧气与细胞外和红细胞内的水平衡，然后与血红蛋白结合，直到完全含氧。相反，三氧的溶解度约为氧气的10倍，可使其立即与生物液中存在的任何可溶性化合物和生物分子发生反应，易溶于水，并与抗坏血酸、尿酸盐、游离半胱氨酸、谷胱甘肽分子和白蛋白硫醇基等几种生物分子瞬间反应，然后消失。反应过程中产生的化合物(活性氧和脂质三氧化产物)是 "三氧信使"，是其生物和治疗作用的原因。 O2-O3疗法有许多有益的作用，包括灭活病毒等微生物，调节免疫系统，改善微循环，抗炎作用刺激氧代谢，促进组织氧合。
Three patients presenting with severe COVID-19 pneumonia had marked and rapid clinical improvement after O2 –O3 therapy. Informed consent was obtained from all 3 patients. Ozonated autohemotherapy involved intravenous infusion of ozonated autologous whole blood. Initially, 200 mL of autologous whole blood was drawn from the antecubital vein into a standard plastic disposable blood collection bag (certified SANO3 bag) containing 35 mL of anticoagulant citrate dextrose solution. The blood was then enriched with 200 mL of gas mixture O2 –O3 with an ozone concentration of 40 μg/mL obtained using an Ozonobaric P Sedecal device, which is an ozone generator with CE0120 certificate type IIb. The ozonized blood was then slowly reinfused into the same vein over a period of approximately 10 min.
3例出现重症COVID-19肺炎的患者，经过O2-O3治疗后，临床症状明显快速改善。3例患者均获得知情同意。起初，从肘中静脉抽取200 mL自体全血到标准的一次性塑料采血袋（抗氧化处理）中，内含35 mL抗凝剂(枸橼酸钠)。然后用200 mL的气体混合物O2 -O3富集血液，三氧浓度为40 μg/mL。充分混合后，将三氧化的血液缓慢地重新回输回同一静脉中。
A 49-year-old man (body mass index [BMI] 31 kg/m2 ) presented to the Emergency Department (ED) with abdominal discomfort for 1 week and progressive shortness of breath during the previous day. On examination, the abdomen was soft and lax, with no distension noted. Chest auscultation revealed bilateral crackles with reduced air entry. A computed tomography (CT) of the chest–abdomen revealed bilateral lung infiltrates compatible with COVID-19 pneumonia (Figure 1).
Laboratory tests showed increased levels of ferritin (1609 ng/mL), D-dimer (1900 ng/dL), C-reactive protein (CRP, 17.3 mg/dL), and lactate dehydrogenase (LDH, 536 IU/L). Real-time polymerase chain reaction (RT-PCR) analysis from a nasopharyngeal swab sample was positive for viral ribonucleic acid (RNA). The patient was admitted to the Intensive Care Unit (ICU), with clearly improved conditions and was discharged to the general ward after 24 h. Unfortunately, the patient experienced worsening hypoxemia and respiratory distress during the next 24 hours. The PaO2 /FiO2 ratio was 235. He was put on a non-rebreather face mask with oxygen on FiO2 of 0.8, but noninvasive ventilation (NIV) was not required. An X-ray showed diffuse bilateral infiltrates. Two sessions of ozone autohemotherapy were administered daily for 3 days, with 12 hours between each session, which was followed by rapid improvement in respiratory rate and increased PaO2 /FiO2 ratio, with decreased FiO2 to 0.31% (3 L) after only 1 day. Figure 2 shows the X-ray before and after only 2 sessions of ozone therapy. Figures 3 and 4 illustrate the trend of changes in ferritin and D-dimer levels before and after O2 –O3 therapy. The patient was discharged to home on day 4 after ozone therapy.
实验室检查显示铁蛋白（1609 ng/mL）、D-二聚体（1900 ng/dL）、C-反应蛋白（CRP，17.3 mg/dL）和乳酸脱氢酶（LDH，536 IU/L）水平升高。鼻咽拭子样本的实时聚合酶链反应（RT-PCR）分析对病毒核糖核酸（RNA）呈阳性。患者入住重症监护室(ICU)，病情明显好转，24 h后出院至普通病房，遗憾的是，在接下来的24小时内，患者出现了低氧血症恶化和呼吸困难。PaO2 /FiO2比值为235。他被戴上非再呼吸面罩，在FiO2为0.8时吸氧，但不需要无创通气（NIV）。X线检查显示双侧弥漫性浸润。每天进行两个疗程的三氧自体血治疗，共3天，每个疗程之间间隔12小时，随后呼吸频率迅速改善，PaO2 /FiO2比值升高，仅1天后FiO2降至0.31%（3L）。图2为仅2个疗程的三氧治疗前后的X线片。图3、4为O2-O3治疗前后铁蛋白和D-二聚体水平变化趋势。三氧治疗后第4天，患者出院回家。
图3. 三氧治疗开始后，铁蛋白水平从>2000 ng/mL下降到246 ng/mL。
图4：三氧治疗开始后，D-二聚体水平从1900 ng/mL下降到323 ng/mL。
A 61-year-old male patient (BMI 29 kg/m2 ), with a background of hypertension, presented to the ED with a week of fever (>39°C) and progressive shortness of breath over the last 48 hours. On examination, chest auscultation showed crackles with reduced air entry over the right hemithorax. A CT of the chest–abdomen revealed right upper infiltrates compatible with COVID-19 pneumonia. High levels of ferritin (2200 ng/mL),D-dimer (3660 ng/mL), CRP (10 mg/dL), and LDH (816 IU/L) were reported. Baseline PaO2/FiO2 was 253. Oxygen at a FiO2 of 0.6 was provided via face mask to the patient in the general ward. NIV was not required. Two sessions of ozone autohemotherapy were administered over a period of 24 hours for 2 days, which was followed by decreased FiO2 0.31% (3 L) with improved PaO2 to 90 mmHg, and decreased levels of laboratory markers. Figures 5 and 6 show the trend of changes in the LDH and CRP levels before and after O2 –O3 therapy. The patient received 4 sessions of O2 –O3 therapy in total and was discharged to home on day 3 after ozone therapy.
一名61岁的男性患者（BMI 29 kg/m2），有高血压背景，因发热一周（>39℃）和过去48小时内进行性气促而到急诊室就诊。检查时，胸部听诊显示右半身胸上方有裂纹，空气进入减少。胸腹CT显示右上浸润符合COVID-19肺炎。铁蛋白（2200 ng/mL）、D-二聚体（3660 ng/mL）、CRP（10 mg/dL）和LDH（816 IU/L）水平较高。基线PaO2/FiO2为253。在普通病房通过面罩为患者提供氧气，FiO2为0.6。不需要进行NIV。在24小时内进行两个疗程的三氧自体血疗，持续2天，随后FiO2下降0.31%（3L），PaO2改善至90mmHg，实验室标志物水平下降。图5和图6显示的是以下变化趋势。三氧治疗前后的LDH和CRP水平。患者共接受4次O2-O3治疗，三氧治疗后第3天出院回家。
图5：三氧治疗开始后，乳酸脱氢酶水平从816 U/L下降到469 U/L。
A 64-year-old woman (BMI 20 kg/m2 ) presented to the ED with progressive shortness of breath over 3 days. She was diagnosed with COVID pneumonia. Laboratory tests showed high levels of ferritin (656 ng/mL), D-dimer (657 ng/dL), CRP (5 mg/dL), and LDH (452 IU/L). RT-PCR analysis from a nasopharyngeal swab sample was positive for viral RNA. Baseline PaO2 /FiO2 was 243. Oxygen at an FiO2 of 0.8 was provided via face mask in the general ward. NIV was not required. O2 –O3 therapy was commenced immediately after admission to the ward, which was followed by rapid clinical and laboratory recovery profiles similar to those of the previous 2 cases. Figures 7 and 8 show the trend of changes in D-dimer and CRP before and after O2 –O3 therapy. The patient received 6 sessions of O2 –O3 therapy in total, and was discharged to home on day 4 after ozone therapy.
一名64岁的女性（BMI 20 kg/m2）在3天内因进行性呼吸急促而到急诊室就诊，她被诊断为COVID肺炎。实验室检查显示铁蛋白（656 ng/mL）、D-二聚体（657 ng/dL）、CRP（5 mg/dL）和LDH（452 IU/L）水平较高。鼻咽拭子样本的RT-PCR分析为病毒RNA阳性。基线PaO2 /FiO2为243。在普通病房通过面罩提供0.8的氧气。不需要进行NIV。入院后立即开始O2-O3治疗，随后临床和实验室恢复情况迅速，与前2例相似。图7和图8显示了O2 -O3治疗前后D-二聚体和CRP的变化趋势。该患者共接受了6次O2 -O3治疗，三氧治疗后第4天出院回家。
图7.三氧治疗开始后，D-二聚体水平从657 ng/mL下降到>100 ng/mL
We report successful use of ozone therapy in 3 patients with COVID-19 pneumonia in terms of precluding the need for invasive ventilation and early discharge to home after 4–6 sessions of ozonated autohemotherapy. There is no currently available effective treatment for COVID-19 pneumonia. The pathogenesis of the virus is not fully understood, but the pathological picture in the lungs varies significantly in terms of diffuse alveolar damage and microcirculopathy leading to life-threatening hypoxia . Ozone has multiple beneficial properties that could be useful in treatment of COVID-19 pneumonia.
Ozone can deliver sufficient energy and oxygen to the tissues through activating the pentose phosphate pathway, elevating 2,3-diphosphoglyceric acid content in erythrocytes, and stimulating erythrocyte oxygen metabolism . Furthermore, it improves the rheology and capillary action of the blood , which has been reported to be helpful for patients with ischemic vascular diseases . Additionally, ozone has an antiplatelet effect and increases release of some prostacyclins such as PGI2 , which are beneficial for patients with microthrombosis . All of these effects can help decrease the hypercoagulation phenomena observed in COVID-19 patients. Another important role played by ozone in COVID-19 is its immunomodulatory effects. The inflammatory response is a hallmark of severe infection, and cytokine modulation is key to avoid patient deterioration. Ozone has potent anti-inflammatory properties through modulation of the NLRP3 inflammasome, which plays a crucial role in the initiation and persistence of inflammation in various diseases . Ozone can also confer renal protection; the rate of kidney damage in COVID patients is significant, and ozone modulates the accumulation of neutrophils locally, the expression of interleukin (IL)-6, tumor necrosis factor (TNF)-a, and albumin modified by ischemia in the kidneys, and increases local antioxidant capacity .
The effectiveness of ozone against pathogens is well known. Ozone can induce the release and modulation of interferons (IFNs) and related cytokines, such as IL-2, IFN-g, and TNF, and colony-stimulating factors , and can also modulate and stimulate phagocytic function , which can have a very positive effect in COVID-19 infection. Angiotensin-converting enzyme 2 (ACE-2) is an entry receptor for COVID ; the virus’s spike protein attaches to an ACE-2 receptor on the cell surface, mediating virus fusion with and entry into the host cell . This receptor can be blocked by control of the nuclear factor erythroid 2-related factor 2 (Nrf2), which regulates and blocks the activity of this receptor . Because ozone causes rapid Nrf2 activation , it seems very likely that this is an important physiological mechanism in blocking endogenous COVID-19 reduplication by preventing contact with this receptor.
O2 –O3 therapy appears to be effective in COVID-19 patients with severe respiratory failure; however, it is not yet an approved therapeutic technique. Large controlled clinical trials are required to study the efficacy and safety of using O2 –O3 therapy compared with standard supportive care in patients with COVID-19 in terms of the need for invasive ventilation and length of hospital and ICU stays.