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Clinical utility of ozone therapy for musculoskeletal disorders

Omar Seyam1, Noel L. Smith2, Inefta Reid1, Jason Gandhi1, 3, Wendy Jiang1, Sardar Ali Khan1, 4, *1 Department of Physiology and Biophysics, Stony Brook University School of Medicine, Stony Brook, NY, USA 2 Foley Plaza Medical, New York, NY, USA
3 Medical Student Research Institute, St. George’s University School of Medicine, Grenada, West Indies
4 Department of Urology, Stony Brook University School of Medicine, Stony Brook, NY, USA

*Correspondence to: Sardar Ali Khan, MD, FRCS, FACS, skysalik@gmail.com. orcid: 0000-0002-4759-530X (Sardar Ali Khan)

Oxygen-ozone (O3) therapy serves as an alternative medical technique that increases the oxygen in the body along with the introduction of O3. O3 therapy has finally reached a level where the biological mechanisms of action have been understood, showing that they are in thedomain of physiology, biochemistry, and pharmacology. Few clinical applications have been reviewed here as well as exemplifying that O3therapy is particularly useful in musculoskeletal disorders. In the therapeutic range, O3 can be used as a more effective and safe substitute of standard medications. O3 therapy has been used for many years for its ability to inactivate various viruses, cancer, and acquired immunedeficiency syndrome but is now making strides in the treatment of musculoskeletal disorders such as rheumatoid arthritis, lumbar facetjoint syndrome, subacromial bursitis, carpal tunnel syndrome, osteoarthritis, hip bursitis, shoulder adhesive capsulitis, herniated disc, and temporomandibular joint disorder.

Key words: oxygen-ozone therapy; osteoarthritis; herniated disc; carpal tunnel syndrome; lumbar facet syndrome; muscle oxygenation,fibromyalgia; cervical disc herniation

doi: 10.4103/2045-9912.241075
How to cite this article: Seyam O, Smith NL, Reid I, Gandhi J, Jiang W, Khan SA. Clinical utility of ozone therapy for musculoskeletal disorders. Med Gas Res. 2018;8(3):103-110.

INtRODUCtiON
Ozone (O3) has great oxidizing activity as a soluble gas. Whenin contact with biological fluids, it forms reactive oxygenspecies as well as lipid oxidation products.1 Both of these products react with white blood cells initiating the formation of proteins, cytokines, and red blood cells which increases the tissue oxygen supply. O3 is used to treat many cases regarding the muscles, tendons, and joints. O3 therapy raises the pain thresholdas it works based on stimulating antinociceptive ap- paratus mediated by serotonin and endogenous opioids. Due to neoangiogenesis, O3 allows for vascularization caused from tissue hyperoxygenation. Therefore, the inhibitory capacity ofinflammatory metabolites is improved as well as local tissuetrophism. There is a common theme among many literature reports that O3 reduces local pain which favors the mobility lost during the painful state and recovery of joint function.2,3O3 can be injected by peri-articular, intra-articular, or percu- taneous means. It is considered a satisfactory treatment with a low risk of complications and high success rate.

MECHANiSM OF ACtiON
An endogenous cascade is started when beginning the use of O3 therapy. In response, a stress is induced from the bio- logically active substrates that are released. Because of O3’s ability to dissolve in the aqueous component of plasma, it can cause this oxidative stress.4 Hydrogen peroxide and a reac-

tive species (ROS) are formed when O
polyunsaturated fatty acids. When inhaled, the O3 reacts with polyunsaturated fatty acids that are found in the alveolar lin- ing layer. A mixture of lipid ozonation products (LOP) is also

formed simultaneously such as malonlydialdehyde, lipperoxyl- radicals, hydroperoxides, isoprostanes, 4-hydroxynonenal, and alkenals.5 The activation of the transcriptional factor mediating nuclear factor-erythroid 2-related factor 2 (NRF2) is increases with the moderate oxidative stress caused by O3. The role of NRF2 is to activate the transcription of antioxidant response elements. A variety of antioxidant enzymes attain an increased concentration level upon introduction of antioxidant response elements. Some of the antioxidants include glutathione S- transferase (GST), catalase (CAT), heme oxygenase (HO)-1, superoxide dismutase, glutathione peroxidase, heat shock proteins and quinone-oxidoreductase. most of these enzymes play a role as free radical scavengers in a range of diseases.5Depending on the cell’s redox status and the amount given, O3 and other medical gases such as nitric oxide and carbon monoxide have a twofold effect. O3 overexpresses HO-1 or heat shock proteins (HSPs) of 32 kPa which produces nitric oxide.6 The expression levels of Hsp70 are upregulated by O3which is related to HO-1. Therefore, there may be a develop- ing role in therapy of free radical-based diseases. Heme is enzymatically degraded by HO-1 and can be toxic depending on free iron, amount produced, and bilverdin. Biliverdin is a neutralizer of nitrosative and oxidative stress based on the ability to interact with reactive nitrogen species and NO.7,8 It was found recently that the heat shock response provided a cytoprotective state during aging, cancer, neurodegenerativedisorders and inflammation.9 Throughout the phylogenetic spectrum, HO isoforms are found to be as regulators of redox homeostasis and dynamic sensors of cellular oxidative stress. Hormesis is a defense mechanism for oxidative insults to

3

reacts with water and

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multiple organ systems.9 O3 can have a role in hormesis byregulating the proinflammatory and anti-inflammatory effectsof prostaglandin formation which is of similar nature to nitric oxide.10

How O helps repair musculoskeletal tissues (tissue repair)3

spine in which a tear in the fibrous ring of an intervertebraldisc allows the central portion to bulge out.19 The treatment of lumbar disc herniation is applied with a needle which is placed into the hernia between the inner margin of the facet joint and the lateral nerve root. This has been considered a minimally invasive treatment for nerve root treatment since the O3 is mini- mally invasive and the needle is thin. The mechanism of action is as follows: nucleur pulposus is oxidized by proteoglycan which then cause it to denaturize and reduce in volume. The local blood circulation will have reduced when the osmotic pressure is reduced. The doze of O3 administered is essential and should not pass the capacity of glutathione and antioxi- dant enzymes to prevent accumulation of hydrogen peroxide and superoxide anion which can possibly degradation of cell membranes.20 O3 in a medium of pH = 8 or higher will cause the formation of free radicals. While, in a medium of pH = 7.5 or lower the formation of peroxides occur due to the ozon- olysis mechanism. In oxygen-O3 therapy, O3 is administered at a nontoxic concentration of 1 to 40 μg of O3 per milliliter of oxygen. It was found through in studies that consisted ofin vitro on resected human disk specimens as well as in vivoon rabbits that the optimal concentration to administer is 27 μg. O3 has a direct effect on the disk’s nucleus pulposus spe-cifically in proteoglycans at this concentration which resultsin subsequent cell degradation of the matrix and release of water molecules.21 The matrix is replaced by fibrous tissuesand formation of new blood cells in approximately 5 weeks. A reduction in disk volume is the result of all these events. In a study conducted by Andreula and others,22 five histologicdisk specimens were removed during surgical microdiske- cotmy who received intradiscal injects of O3 at 27 μg/mL.Dehydration of the fibrillary matrix of the nucleus pulposus,signs of regression (fragmentation and vacuole formation), andrevealing collagen fibers were all noted in these specimens. Other findings of a herniated disk untreated with medicalO3 are proliferating and signs of new blood cell formationguided by lymphocyte inflammatory tissue and chondrocytehyperplasia. One of the main therapeutic causes of medical O3 is a reduction in disk size which can possibly reduce nerve root compression. Furthermore, disk shrinkage can improve local microcirculation and increase the supply of oxygen by reducing venous stasis caused by disk compression of vessels. O3 therapy also had analgesic and anti-inflammatory effects intreating disk herniation.23 The action ties into inhibiting release of bradykinin or release of algogenic compounds as well asinhibiting synthesis of proinflammatory prostaglandins. Proin-flammatory cytokines such as interleukin can be neutralized byincreasing release of antagonists. The results from this study was satisfactory compared to other percutaneous treatments for disk herniation such as enzymatic chemonucleolysis.24 The two procedures are similar, however oxygen-O3 therapy is less invasive due to the narrow nature of the needle and less trau- matic.25 Also, there were no anaphylactic or allergic reactions.25

Shoulder (Glenohumeral joint)

The shoulder allows for the role of orientating the hand and has a great degree of movement.26 Specifically, the glenohu- meral joint is not a stable ball-and-socket joint, but has high mobility.26 The rotator cuff muscles allow for joint motility.

When O3 gets in contact with organic fluids such as plasma,lymph, urine, and saliva, it interacts highly with all tissue components.11 It can react with antioxidants, glutathione, cys- teine, albumin, ribonucleic acid (RNA), and deoxyribonucleic acid (DNA).12 All these reagents, participate in the ozonation process and formation of lipid oxidation products and ROSs.11These two molecules function as biochemical regulators ofinflammation and physiological concentrations.11 ROS are described as being highly unstable, immediate action and a half-life less than one second.12 It is common that patients re- port the sensation of well-being during the course of O3 therapy and this is due to the LOPs which stimulate the central nervous system and endocrine system while also improving hormonal production, neurotransmitter release, and metabolism.11 It was proved that O3 was capable of promoting the preservation and increase of endogenous antioxidant systems through a study conducted by Re et al.13 Another study which dealt with the ozonation of platelet-rich plasma samples results demonstrated the increase of interleukin (IL)-8. The increase of IL-8 allows for the leukocytes to leave the circulation into the tissues to facilitate the phagocytosis of bacteria and necrotic tissue of ulcers.14 The results also showed an increase in growth factors such as platelet derived growth factor (PDGF), IL, and trans-forming growth factor beta (TGF-β) 1.14 Lim et al.15 that when dermal wounds were exposed to O3, it increased the activity of factor nuclear kappa B, an important immunomodulatory ofinflammation and the expression of TGF-β which is essentialfor remodeling tissue.

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temporomandibular joint disorder (myofascial pain syndrome)

The temporomandibular joints are the joints that connect the jawbone to the skull. Temporomandibular joint disorder also known as myofascial pain syndrome results in pain that limits chewing, talking, and other daily activities. It was found that the use of O3 therapy has been much more effective than medi- cation therapy in patients with high pain scores which relieves pain and increases the maximum voluntary interincisal mouth opening values. A proposed explanation of why O3 causes the joint to heal quicker than of the traditional therapy is due to the highly reactive nature of O3. It is able to stimulate the fibroblas- tic joint repairing abilities when injected into a joint capsule. It is also able to promote new cartilage growth as well as reducinginflammation. When split into separate oxygen atoms, O3 is able to react when in contact with a contaminant.16-18 In fact, it never fails to initiate this reactive activity. A recent study found that 87% patients had either improved or completely recovered from temporomandibular pain.18 It was concluded that this was a promising new treatment, but the mechanism of action is still being researched. A plausible mechanism can be due to a single oxygen atom oxidizing the contaminants. It is thought that O3 can stimulate the fibroblastic joint since it issuch a reactive molecule. O3 can react with the contaminant when it splits into single oxygen atoms.18

Herniated disc (back pain)

The herniated disc is characterized as a condition affecting the

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These muscles include the infraspinatus, subscapularis, su- praspinatus, and teres minor. O3 therapy for the glenohumeral joint includes injection with the posterior approach under the inferior margin laterally using a 22 G needle. In a case study, Benvenuti27 reported that a 58-year-old woman experiencing severely limited joint movement and pain underwent a session of O3 therapy with 10 mL gas mixture at a concentration of 15 μg/mL. Furthermore, it is helpful to puncture the long head of the bicep muscle and subacromial bursa with a 27 G needle in microdoses of 0.5 to 1 mL. After one week, the patient reported a reduction of nocturnal pain.27 The patient reported a recovery of active shoulder function and total reduction of pain.

Shoulder adhesive capsulitis

Shoulder adhesive capsulitis is a condition where there is a reduction in the arc of active motion.28 Adhesive capsulitis is characterized with an unknown etiology that has an onset of pain associated with it.28 The treatment with oxygen-O3 canhelp in the reduction of pain as well as inflammation activ- ity. It uses a mixture which is 95–96% oxygen and 4–5% O3.The reduction and modulation of inflammation activity and areduction of pain are a few of the many action accomplished with medical O3. The local injection of O3 destroys algogenic substances, alters serotonin, and inactivates bradykinin. All of these substances are altered to produce no pain. A releaseof soluble receptors or anatagonists neutralize proinflamma-tory cytokines such as interferon-a, tumor necrosis factor-α (TNF-α) and ILs. In addition, the denaturation of cellular pro- teins such as kallikrein, kininogen, and cyclooxygenase (COX) help form endorphins and modify the pain receptors. Another important activity is given by the muscle relaxant action bydirection action on the muscle fibers.12 Contraindication to this treatment include latent hypoglycemia, hyperthyroidism, favism, pregnancy, and sickle cell anemia. This study has many limitations since it was a case study. It can be a basis for future studies on a better statistical criterion as well as on more patients.

Hip bursitis (inflammation)

Hip bursitis can be characterized as the swelling of bursae. Thebursae are fluid-filled sacs that cushion muscles and tendons.29The hip is similar to the shoulder since it is also a ball-and-socket joint. However, the hip has much more flexibility inmovement as compared to the shoulder where it is attached to the trunk. It is common that patients are not aware theyhave a hip inflammation since they might complain about thepain in the anterior knee and thigh rather than the hip itself. O3 therapy can be used to alleviate hip pain due to functional overload, trochanteric bursitis, pain caused by initial and late coxarthrosis, and hip tendonitis. It is also helpful to associ- ate oxygen-O3 therapy along with prescribed exercises and a period of rehabilitation. In a case report,27 a 54-year-old man was treated by a cycle of 5 therapy session of O3. The dosage consisted at a concentration of 25 μg/mL at 55 mL oxygen) through weekly intervals. It was injected at a lateral approachwhere the method of infiltration was called peritrochanteric. The patient referred difficulty walking with deambulationfor approximately 50 meters and nocturnal hip pain.27 Thejoint pain subsided after the first treatment session with an

improvement in trochanteric swelling. At the end of the treat- ment cycle daytime and nighttime pain had disappeared. By the subsequent follow-up visit which was approximately 5 weeks later, there were no signs of bursitis. Signs of limping stopped and the patient was able to take the stairs withoutany difficulty. This study demonstrates the versatility and ef-ficacy of O3 therapy when administered in small doses through weekly intervals.

Osteoarthritis (OA) (knee)

OA is a degenerative disease that affects function and produces pain.30 O3 infiltration accelerates anabolism and producesbetter vascularization on cartilage and bone. Regarding knee osterarticular disease, there is a great variability in terms of gas concentration of O3 as well as the side of injection. On the knee joint, it could be either periarticular, intra articular, and subcutaneous. Currently, there is no cure for knee OA. Because of this there is a focus on the amelioration of the symptoms caused by knee OA. O3 improves the range of motion (ROM) and slows down the degenerative process. It also is able toinhibit chondrocytes, stem cells, inflammatory cytokines, nitricoxide, and mineral metalloproteinases. The published studies on O3 regarding the knee are limited. Riva Sanseverino31 werethe first people to use O3 to treat the knee OA. Over the many years, intra articular O3 injections have found to be a costless procedure that is effective. Patients who have severe OA will improve at about the same rate as those with low grade OA. The pathophysiology of OA is characterized by the destruc- tion and softening of articular cartilage along with increased matrix degradation due to proteoglycanases and collagenases. Activated chondrocytes and monocytes release enzymes whichby releasing TNF-α and IL-1 multiply the inflammation. Thesynthesis of prostaglandins increases and there is an attempt to maintain biomechanical matrix. It is believed that the synthesis of O3 Messengers LOPs and ROS would act in two phasesin the synovial fluids. O3 would inhibit the proinflammatory cytokines such as prostaglandin E2 (PGE2) during the firstphase. During the second phase, O3 will act over inhibitorycytokines such as IL-10, TGF-β, and IL-4, antioxidant en- zymes, and neoangiogenesis. These are all working together in the process of repairing the articular joint by stimulatingfibroblasts, chondrocytes, and stem cells.

Carpal tunnel syndrome

Carpal tunnel syndrome is caused by a compressed nerve in a narrow passageway on the palm side of the wrist.32 Sec- ondary forms are associated with a variety of diseases such as hypothyroidism, diabetes mellitus, oestrogen therapy, rheumatoid arthritis, amyloid disease.33 The mechanism of oxygen O3 treatment is based on three mechanisms that are shared by the treatment of herniated disc in the spine. By increasing trans-tissue and intra-oxygenation along with reduced lymphatic stasis and hypoxia, there is an indirect vessel-mediated decompression of the nerve roots. Second, by inhibiting the release of polymorphonucleates through in- creasing immunosuppressive cytokines and proteinase through macrophages.34 Third, inhibiting the release of prostaglandinsand pro-inflammatory bradykinins, action on the cell-mediated inflammatory response would take place.35 Oxygen-O3 therapy

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seems to guarantee improvements of symptoms after one year compared to steroid injections. A study conducted by Zam- bello et al.,36 showed that 90% of patients had a significantimprovement after O3 injection. 17% had a good control of symptoms and 70% of patients no longer had symptoms after a one year follow up.36 However, further studies should be conducted to see if symptoms remain or not after two years or more to evaluate the long-term effects.

Partial tear of the supraspinatus tendon

The supraspinatus tendon is part of the rotator cuff.37 Oxygen-

relieve the symptoms. In a study,38 which consisted of 40patients, specifically 26 males and 14 females whom hadshoulder pain for approximately six months. In order for patients to be included in the study, the size of the tear of the supraspinatus had to be less than 1.5 cm. Ultrasound guidanceis recommended since the blind infiltration does not guaranteeto get properly in the shoulder joint.38 It was concluded thatthe ultrasound-guided infiltration of oxygen-O3 therapy proved to be an effective treatment method in partial tears of the supraspinatus tendon. However, it is necessary to do further research requiring a large sample size.

Subacromial bursitis

First degree spondylolisthesis and spondylolysis-spondylosis is a defect of part of the vertebral arch between the superior and inferior spinal processes. Spondylolisthesis is if the defect results in a forward shift of one of the vertebral body on one another.39 Eighty-three percent of the 18 patients in this had a complete recovery from pain immediately.40 The twofold ac- tion of O3 in the perganglionic region and in the lysis points of the neural arch or pars interarticularis region innervated by Luschka’s recurrent nerve is an explanation for such fast pain.21The use of the gas mixture directly next to the lysis points on Luschka’s nerve by exploiting the well-known analgesic andanti-inflammatory effects of the oxygen-ozone mixture. Thespine is innervated by Luschka’s recurrent nerve or posterior primary branch and vertebral plexus.41 The plexus innervates the end plates, longitudinal ligament, vertebral bodes, and rela- tive peridural tissues in the pars interarticularis region or the neural arch.41 The anti-inflammatory and analgesic effects ofthe oxygen-O3 mixture infiltrates directly proximal to the lysispoint on Luschka’s nerve.42 Prostaglandin levels and cytokine levels are normalized with a reduction of reactive oxidant species and an increase in superoxide dismutase production.A eutrophizing effect occurs when subsequent infiltration intothe periganglionic region takes place both adjacent to the nerve root compressed at the level of muscle spasm and injured by accompanying disc protrusion.43,44 A good final outcome isaccounted due to the combined action.

lumbar facet joint syndrome

Lumbar facet joint syndrome can be described as pain at the joint between two vertebrae in your spine. It is a condition which affects about 80% of people who have lower back pain. The main mechanism of action may be considered as follows: there is an immediate oxidation by which the proteoglycan in the nucleus pulposus could be oxidized immediately and the osmotic pressure is reduced.45 The volume of the nucleus pulposus would decrease as well as necrotize, denaturize, and

atrophy. The local blood circulation would also be changed when the osmotic pressure is reduced. The symptoms may be improved when increasing the oxygen supply. Regarding theanti-inflammatory effect, as the vein, lymphoid tissue, and nerve root were compressed by the annulus fibrosus and herni-ated nucleus pulposus, the lymphatic backflow and venous wasobstructed which was accompanied by exudation and nerve edema. To induce an immune response, antigenic substances such as B-lipoprotein and glycoprotein could be released thatwould result in aseptic inflammation and adhesion. The pain ofdisc herniation could be caused by these factors. Lastly regard-ing the analgesic effects, enzyme products and inflammatorymediators stimulate the nerve endings near ligament and on the disc surface that causes the pain that is associated with disc herniation. In order to attain pain relief, the strong oxidative activity of O3 can inactivate the above inflammatory mediators.O3 injected into the middle of the disc through the conventional posteriorlateral route had produced the desired result for small or medium size disc herniations.22 Regarding the large disc herniation, the symptoms were not eliminated quickly and theefficacy was limited due to the hernia compressing the nerve root. The efficacy of O3 therapy is poor with patients who havelarge lumbar disc herniation and greatly significant with thosewho have a small or minimal disc herniation. Since the annulusfibrosus was either completely or partially ruptured, it wasfound that the O3 could diffuse through the tissues that were torn surround the vertebral or spinal disc. This would cause the hernia and nucleus pulposus to not be fully oxidized. In a study conducted by Lu et al.,46 the treatment of large lumbar disc herniation with percutaneous O3 injection was greatly effective. An improvement of local ablation and eliminatinglocalasepticinflammationwasachievedthroughO injection

through using percutaneous O3 injection with the inner margin of facet via the posterior-lateral was an effective method. It is important to note that the concentration of O3 should beacceptable in quantity and the rupture of the annulus fibrosusby high intradiscal pressure should be avoided in order toensure efficacy. Low pressure of repeated injection of O3 was suggested for large disc herniation. This can be accomplished by repeatedly pushing and pulling the syringe to allow the O3to completely oxidize and contact the nucleus pulposus. New O3 is then injected and the residual O3 is abandoned to avoidrupture of the annulus fibrosus.

Cervical disc herniation

Cervical disc herniation is wear and tear of a disc in the neck. It is suggested that the material from the nucleus pulposus may act as a chemical or immunologic irritant the nerve and thesemechanisms may produce an inflammatory effect.47 Studies have hypothesized that the injection of O3 induces overexpres- sion of antioxidant enzymes which then neutralizes excessive ROS formation. In the degenerated nucleus pulposus, the intradiscal injection of O3 can accelerate the degradation of proteoglycans.14 The biochemical modification of the mediumin the extraduralspace is one of the important aspects of it.48The cause of radicular pain is A2 phopholipase independentof a direct inflammatory process or immunological response.47A2 phospholipase is responsible for the prostaglandins and arachidonic acid liberation. It has been shown that there are

O therapy is one of the many methods that can be used to3

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in or around the hernia areas. Therefore, it was concluded

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high levels of A2 phospholipase in herniated discs. A power- ful stimulus to the activation of antioxidant defense is the result of O3 injected in the peridural space of the conjugation foramen and disc. Even in cases that had extruded cervical disc pathology, injections were still performed and had great results. This is most likely due to the fact that normal tissues and the isolate fragment are separated.48

Rheumatoid arthritis

Rheumatoid arthritis is characterized as an autoimmune dis- ease where the body’s immune system attacks the joints instead of bacteria and viruses.49 The syndromes include hyperplasiaof synovial cells, excess synovial fluid, and forming pannuswhich can damage joint deformities and articular cartilage.50,51The etiology for rheumatoid arthritis is still not understood. The common treatments are immunologic purging, advanced surgical treatment, and drug therapy.52 However, O3 therapy is a new treatment in treating rheumatoid arthritis can overcome these limitations at a certain level. The therapeutic mechanism of O3 still remains unclear. Previous research has showed that O3 can reduce the activity of TNF-α in the inflammatorytissues and suppress synovial hyperplasia and joint swell in rheumatoid arthritis in rats.53,54 Therefore, a study was con- ducted to treat bovine collagen II-induced RA in rats with intraarticular injection of O3 at various concentrations.55 Rats were injected with complete Fruend’s adjuvant bovine col- lagen II that successful established a rat model of rheumatoid arthritis.55 In the O3 treated groups, the optimal concentration for treating rheumatoid arthritis was 40 μg/mL.55 They also had higher TNF-receptor (TNF-R)1 indicating that O3 can reducesynovium injury in rats with RA and lower synovial TNF-αand TNF-R2 levels.55 A plausible mechanism is the reduction in TNF-R1 and rheumatoid arthritis TNF-a levels and increase the level of TNF-R1 which increase synovial cell apoptosis and preventing synovial cell proliferation.55

Systemic sclerosis

Systemic sclerosis is a chronic tissue disease characterized byvascular abnormalities in the joints, internal organs, and fibro- sis.56 The etiology and clinical manifestations of sclerodermaare still not understood; this is why it is difficult to treat sys- temic sclerosis.56 The inflammatory processes can be limited byO3’s potential in reducing the proliferation of neutrophils and mastocytes, increasing concentration of prostacyclin 6-keto-prostaglandin F1α (6-keto-PGF1α), impeding the release ofacute phase proteins, and decreasing the concentration ofprostacyclin (PGF)2α resulting from the oxygen radicals onarachidonic acid.57 The results from this study have proved to slow down the progression of the illness by limiting the activation of the immune system. It had also increased the movability of interphalanx joints and decreased the thickness of the skin.57 O3 is able to penetrate the epidermis water-fat barrier and also has a good solubility in serum which is why it has a good effect on the skin.57 The vasodilating effect of O3allows for the decrease of skin score index, decrease of arterial blood pressure, increase in angle of interphalangeal joints all through the synthesis of nitric oxide synthase.57

Fibromyalgia

Fibromyalgia is seen as a rheumatic disease which means that

it causes myofascial pain or soft tissue pain.58 The mechanism

of O3 is as follows. After the O3comes into contact with the

blood, it immediately reacts with various reducing molecules

such as antioxidizing agents, unsaturated fatty acids containing

double bonds to produce reactive oxygen species. Both lipid

peroxidation products and hydrogen peroxide are generated,

when O reacts with polyunsaturated fatty acid (PUFA).593

Enzymatic antioxidant systems such as aldehyde dehydro-

genase and glutathione-transferase neutralize the toxicity

of both molecules (LP and hydrogen peroxide). They act as

secondary messengers which stimulate further generation of

antioxidant enzymes.60 This can be done if O3 is administered

in quantities that are able to achieve a therapeutic effect which

can protect against radicals and is nontoxic. A case which a

45-year-old woman was administered oxygen-O3 therapy

biweekly sessions equating to a total of 12 sessions. Due to a

lowering of painful symptoms, the patients experience a sense

of well-being. An improvement in the asthenia was seen due to

a greater oxygenation of tissues because of O3. There were no

side effects seen in this case. The limitation from this study is

the small number of patients in which it was conducted. It was

foundthatO glycolysiswasspedupthroughtheactivationof

3 61,62
the mitochondrial respiratory chain. The O3 mixture will

cause an increase in oxygenation level due to the increasedefficiency of the antioxidant enzyme system, enhance sero- tonin production, and microcirculation. The production of endorphins was enhanced by the motor plate. So patients whohad fibromyalgia had improved their daily activities by 40%and sleeping disorders by 6%.63

Muscle oxygenation

The role of O3 therapy was observed for hypoxic tissues, those in which tissues were below-normal level of an adequate oxy- gen supply.64 A study has demonstrated that O3 therapy can change the level of oxygenation in resting muscles by measur- ing directly the pressure of oxygen.64 Through autohemotrans- fusion, O3 therapy avoids lung toxicity from oxidative stress. The effects of O3 are mediated by rapid oxidation of blood substances. Hydrogen peroxide and peroxidated lipoproteins both are reactive oxygen species that can activate the hexosemonophosphate shunt. Charge modification is done by the in- crease of malonyl aldehyde and lipid peroxidation as well as animprovement of blood rheology and flexibility of erythrocytemembrane. The collaboration of nitric oxide, adenosine, and prostaglandins can decrease vascular resistance. It is hypoth-esized that this will lead to blood flow redistribution. This issupported by the data collected in the study which shows the correlation between the change in pressure of oxygen post- ozone therapy and the initial oxygenation. Another possible mechanism to explain the results of this experiment, is the increase in production of 2,3-diphosphoglycerate in erythro- cytes and lipid peroxidation of red blood cell membranes can be achieved with the activation of the hexose monophosphate shunt.65 These would both cause a shift to the right in the oxyhemoglobin dissociation curve ultimately leading to an increase of release of oxygen to the tissues.65

Spinal muscle disorder (horse)

The harmonious movement of the spine and balance are the

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result of the muscles running along the spine to the dock of the tail. Muscle fatigue is caused by soft tissue spinal lesions. Impaired performance in athletic horses is mainly caused by changes in the thoracolumbar spine. One of the many patho- logic changes affecting the thoracolumbar spine is soft tissue spinal lesions. The muscles may suffer varying degrees ofinflammation after intense muscular stress. In the study con- ducted, all four horses had a positive response to O3 therapy.66They were all able to increase their trotting speed due to the stiffness that was relieved and increase in posterior muscle chain thrust. The mechanism of action of O3 is described as the O3 coming in contact with the blood on different targets. Since O3 is very active it reacts when it comes in to contactwith blood or any biological fluid. O3 first reacts with polyun- saturated fatty acids then with proteins, antioxidants (ascorbic acid and glutathione). When O3 reacts with bio molecules, it produces a molecule of ROS which is hydrogen peroxide and two molecules of lipid oxidation products. The ROS activates the pentose phosphate pathway. The lipid oxidation products that are produced as 4-hydroxynonenal and malonaldehyde.67Since they are toxic, they undergo a dilution in the circula- tion and get metabolized in the blood circulation. Overall, thebeneficial effects seen by O3 therapy is the increase in avail- ability and delivery of oxygen, adenosine triphosphate (ATP), and glucose within ischemic tissues, enhances implantation of bone marrow stem cells at the site of lesion which can provide neovascularization, tissue regeneration, and angiogenesis. Ballardini66 did not notice any short or long-term effects when administering different treatment cycles to the same horse; he always noticed a positive response from the horse.

Spinal pain

About 80% of the world’s population has a symptom of low back pain.68 In general, the pain one suffers from a herniateddisc is caused from inflammation and not compression. Theuse of O3 therapy is recommended to treat back pain before doing any surgical procedures. There are two techniques one can attempt: direct approach and the indirect approach. O3acting as a chemical reactant by needle insertion refers to the indirect approach. Whereas, the direct approach is done bydirect insufflation of the oxygen-O3 mixture of a concentration at approximately 30 μg/mL and preceded by needle insertion in the pathologic intersomatic space. Eighty percent of 63,000patients have shown good outcomes confirmed by magneticresonance imaging (MRI) controls and computed tomogra- phy.48,69 In the future, it is essential to conduct more studies to assess the role of variables such as place of needle, needle type, O3 concentration, and oxygen amount. The mechanism of action underlying O3 therapy for the direct method is: O3reacts with biomolecules when dissolved in interstitial water, which then results in the formation of reactive oxygen spe- cies such as hydrogen peroxide and hydroxyl radicals.70,71The matrix degenerates with disappearance of the herniated material, when ROS reacts with proteoglycans of the nucleus pulposus,72 thusleading to a lower of sensitivity of axons. Alternatively, nociceptors can be stimulated when algesic endogenous substances released during perineural ischemia. The indirect approach consists of one to four injection of 5–10 mL of O3. The disappearance of pain because of the complexseries and chemical and neurological reactions have defined

it as a chemical acupuncture. The O3 concentration must be between 18–25 μg/mL.73 If it is higher than 20 μg/mL, it can be too painful, and if it is too low, then it won’t be effective. Therefore, it is essential to maintain the right balance since itcan cause risky vasovagal reflex and lipothymia during initial treatments. Contrarily, the pain threshold rises after five toseven treatments therefore, the concentration of O3 increases,but must not exceed 30 μg/mL. The infiltration of O3 therapy uses the paravertebral muscles as a route. Regarding the indi- rect method, the mechanism of action underlying O3 therapy is described as O3 reacting with PUFA, LOPs, and anti-oxidants.The final therapeutic effect is achieved with these compoundsstimulating local C-nociceptors. Altogether, it is concluded that injection of O3 either into the paravertebral muscles or intradiscally has indicated long-term pain relief.16,74

lumbar spinal stenosis (lSS)

LSS can be characterized as the narrowing of the spinal ca- nal of the lumbar area.75 The three main symptoms that LSS gives rise to are radicular pain or discomfort, low back pain, and neurological intermittent claudication.O3 therapy blocks phospholipase A2 which is the same enzyme that epidural steroid injections target.76-78 Therefore, O3 can serve as a better substitute steroid since it has the same mechanism of action while being a much safer drug. The neurological pain in LSS can be improved by the microcirculation that O3 induces.79In a study conducted by Baeza-Noci,76 patients with spinal stenosis underwent O3 therapy for 10 biweekly sessions along with 5 weekly sessions. Each injection of O3 (10 mL) had aconcentration of 20 μg/mL. After 1 year, from the baseline,74% of patients improved with excellent results.76

Complications of O3 therapy in musculoskeletal disorders

The reactivity of O3 gives rise to a cascade of reactions such as the lipid ozonation products acting as signal transducer molecules, peroxidation of lipids leading to changes in mem- brane permeability.80 Endogenous mediators of inflammationare released by the activation of lipases through which LOP activates the lipases.81,82 It is the O3 which reacts with unsatu- rated fatty acids such as in the pulmonary cell bilayers andlung lining fluid. Enzyme inactivation occurs when there is aloss of functional groups in enzymes. Cell death or cell injury may occur from these reactions. Hazardous effects on lung alveoli can occur with the combination of nitrogen dioxide (NO2) and O3 that are in photochemical smog. These effects can be prevented by free radical scavengers such as vitamin E, C or dietary antioxidants. In an in vitro study, peroxides were found to be formed by the presence of O3 and oxidized arachidonic acid.83 The activity of prostaglandin endoperoxides are comprimable to that of arachidonic acid peroxides. The aggregation of human platelets in platelet-rich plasma was seen with arachidonic acid peroxides. While, the presence of vitamin E and indomethacin, presented no signs of aggrega- tion of human platelets in platelet-rich plasma.83 Therefore, this suggests that they can treat O3 toxicity.

cOncLUsIOn

O3 therapy is becoming an effective treatment option for musculoskeletal disorders as it promotes tissue hyperoxygen- ation as well as treating painful syndromes affecting muscles,

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tendons, and joints. Though O3 has indicated great success in most indications mentioned in this review, there still needs to be further research conducted to determine its activity for treatment of plantar fasciitis, costochondritis, and myofascial syndrome. In order to prevent the common side effects that O3 therapy causes, it is essential to continue researching the utility of O3 therapy in all indications.

7. Mancuso C, Capone C, Ranieri SC, et al. Bilirubin as an endog- enous modulator of neurotrophin redox signaling. J Neurosci Res. 2008;86:2235-2249.

8. Barone E, Trombino S, Cassano R, et al. Characterization of the S- denitrosylating activity of bilirubin. J Cell Mol Med. 2009;13:2365- 2375.

9. Dattilo S, Mancuso C, Koverech G, et al. Heat shock proteins and hormesis in the diagnosis and treatment of neurodegenerative dis- eases. Immun Ageing. 2015;12:20.

10. Mancuso C, Pistritto G, Tringali G, Grossman A, Preziosi P, Na- varra P. Evidence that carbon monoxide stimulates prostaglandin endoperoxide synthase activity in rat hypothalamic explants and in primary cultures of rat hypothalamic astrocytes.Brain Res Mol Brain Res. 1997;45:294-300.

11. Bocci V. Ozone as Janus: this controversial gas can be either toxic or medically useful. Mediators Inflamm. 2004;13:3-11.

12. Bocci VA. Scientific and medical aspects of ozone therapy. State ofthe art. Arch Med Res. 2006;37:425-435.

13. Re L, Mawsouf MN, Menendez S, Leon OS, Sanchez GM, Her- nandez F. Ozone therapy: clinical and basic evidence of its thera- peutic potential. Arch Med Res. 2008;39:17-26.

14. Bocci V. Biological and clinical effects of ozone. Has ozone thera- py a future in medicine? Br J Biomed Sci. 1999;56:270-279.
15. Lim Y, Phung AD, Corbacho AM, et al. Modulation of cutaneous

wound healing by ozone: differences between young and aged

mice. Toxicol Lett. 2006;160:127-134.
16. Rahimi-Movaghar V, Eslami V. The major efficient mechanisms of

ozone therapy are obtained in intradiscal procedures. Pain Physi-

cian. 2012;15:E1007-E1008.
17. Bocci V, Zanardi I, Travagli V. Has oxygen-ozonetherapy a future

in medicine.J Exp Integr Med. 2011;1:5-11.
18. Daif E.Role of intra-articular ozone gas injection in the manage-

ment of internal derangement of the temporomandibular joint.

Oral Surg Oral Med Oral Pathol Oral Radiol. 2012;113:e10-e14. 19. Jordan J, Konstantinou K, O’Dowd J. Herniated lumbar disc. BMJ

Clin Evid. 2009;2009:1118.
20. Bellomo G, Mirabelli F, Salis A, et al. Oxidative stress-induced

plasma membrane blebbing and cytoskeletal alterations in normal

and cancer cells. Ann N Y Acad Sci. 1988;551:128-130.
21. Iliakis E, Valadakis V, Vynios DH, Tsiganos CP, Agapitos E. Ratio- nalization of the activity of medical ozone on intervertebral disc a histological and biochemical study. Riv Neuroradiol. 2001;14:S23-

30.
22. Andreula CF, Simonetti L, De Santis F, Agati R, Ricci R, Leonardi

M. Minimally invasive oxygen-ozone therapy for lumbar disk her-

niation. AJNR Am J Neuroradiol. 2003;24:996-1000.
23. Bocci V, Luzzi E, Corradeschi F, Paulesu L, Di Stefano A. Studieson the biological effects of ozone: 3. An attempt to define con- ditions for optimal induction of cytokines. Lymphokine Cytokine

Res. 1993;12:121-126.
24. Matsui H, Terahata N, Tsuji H, Hirano N, Naruse Y. Familial pre-

disposition and clustering for juvenile lumbar disc herniation.

Spine (Phila Pa 1976). 1992;17:1323-1328.
25. Leonardi M. Disc puncture under fluoroscopic guidance. Riv Ital

Ossigeno-Ozonoterapia. 2002;1:73-78.
26. Terry GC, Chopp TM. Functional anatomy of the shoulder. J Athl

Train. 2000;35:248-255.
27. Benvenuti P. Oxygen-ozone treatment of the knee, shoulder and

hip: A personal experience. Rivista Italiana di Ossigeno-Ozonote-

rapia. 2006;5:135-144.
28. Neviaser AS, Neviaser RJ. Adhesive capsulitis of the shoulder. J

Am Acad Orthop Surg. 2011;19:536-542.
29. Hirji Z, Hunjun JS, Choudur HN. Imaging of the bursae. J Clin

Imaging Sci. 2011;1:22.
30. Sinusas K. Osteoarthritis: diagnosis and treatment. Am Fam Physi-

cian. 2012;85:49-56.
31. Riva Sanseverino E. Intensive medical physical treatment of os-

teoporosis with the AID of oxygen-ozone therapy. Europa Medico

Physica. 1989;25:163-170.
32. Ibrahim I, Khan WS, Goddard N, Smitham P. Carpal tunnel syn-

drome: a review of the recent literature. Open Orthop J. 2012;6:69-

76.
33. Katz JN, Simmons BP. Clinical practice. Carpal tunnel syndrome.

N Engl J Med. 2002;346:1807-1812.

Seyam et al. / Med Gas Res

www.medgasres.com

RefeRences

1. Cardoso CC, Carvalho JC, Ovando EC, Macedo SB, Dall’Aglio R,Ferreira LR. Action of ozonized water in preclinical inflammatorymodels. Pharmacol Res. 2000;42:51-54.
2. Lopes de Jesus CC, Dos Santos FC, de Jesus LMOB, Monteiro I, Sant’Ana MSSC, Trevisani VFM. Comparison between intra-ar- ticular ozone and placebo in the treatment of knee osteoarthritis: A randomized, double-blinded, placebo-controlled study. PLoS One. 2017;12:e0179185.
3. Cardelli R, de Santis F, Dall’Olio M, Leonardi M. Osteoarthritisof the hip treated by intra-articular infiltration of oxygen-ozoneand hyaluronic acid (Hyalubrix®). Preliminary results. Int J Ozone Ther. 2008;7:66-69.
4. Bocci V, Larini A, Micheli V. Restoration of normoxia by ozone therapy may control neoplastic growth: a review and a working hypothesis. J Altern Complement Med. 2005;11:257-265.
5. Inal M, Dokumacioglu A, Özcelik E, Ucar O. The effects of ozonetherapy and coenzyme Q10 combination on oxidative stress mark- ers in healthy subjects. Ir J Med Sci. 2011;180:703-707.
6. Bocci V, Aldinucci C, Mosci F, Carraro F, Valacchi G. Ozona- tion of human blood induces a remarkable upregulation of heme oxygenase-1 and heat stress protein-70. Mediators Inflamm. 2007;2007:26785.

Medical Gas Research ¦ September ¦ Volume 8 ¦ Issue 3 109

[Downloaded free from http://www.medgasres.com on Monday, March 11, 2019, IP: 91.253.203.135]

34. Bocci V.Ozone as a bioregulator. Pharmacology and toxicology of ozonetherapy today. J Biol Regul Homeost Agents.1996;10:31-53.
35. Simonetti L, Raffi L, Cenni P, Agati R, Leonardi M. Pharmacologi-cal mechanisms underlying oxygen-ozone therapy for herniated

    disc. Rivista Italiana Di Ossigeno. 2003;16:S201-204.

36. Zambello A, Fumagalli L, Fara B, Bianchi MM. Oxygen-ozone treatment of carpal tunnel syndrome. Retrospective study and lit- erature review of conservative and surgical techniques. Int J OzoneTher. 2008;7:45-48.
37. Volk AG, Vangsness CT Jr. An anatomic study of the supraspinatusmuscle and tendon. Clin Orthop Relat Res. 2001:280-285.
38. Moretti M. Effect of treatment with O2-O3 and hyaluronic acid in partial tear of the supraspinatus tendon. Int J Ozone Ther.2012;11:98-100.
39. Donnally IC, Dulebohn SC. Lumbar Spondylolysis and Spondylo-listhesis. Treasure Island: StatPearls. 2018.
40. Bonetti M, Fontana A, Albertini F. CT-guided oxygen-ozone treat-ment for first degree spondylolisthesis and spondylolysis. Acta

    Neurochir Suppl. 2005;92:87-92.

41. R. Jinkins J. The pathoanatomic basis of somatic, autonomic andneurogenic syndromes originating in the lumbosacral spine. Rivis-

    ta di Neuroradiologia. 1995;8:S35-51.

42. Bocci V, Luzzi E, Corradeschi F, et al. Studies on the biologicaleffects of ozone: 4. Cytokine production and glutathione levels in

    human erythrocytes. J Biol Regul Homeost Agents. 1993;7:133-138.

43. Bocci V. Does ozone therapy normalize the cellular redox balance?Implications for therapy of human immunodeficiency virus infec-tion and several other diseases. Med Hypotheses. 1996;46:150-154.
44. Bocci V, Luzzi E, Corradeschi F, Silvestri S. Studies on the bio- logical effects of ozone: 6. Production of transforming growth fac- tor 1 by human blood after ozone treatment. J Biol Regul HomeostAgents. 1994;8:108-112.
45. Muto M, Andreula C, Leonardi M. Treatment of herniated lumbardisc by intradiscal and intraforaminal oxygen-ozone (O2-O3) injec-

    tion. J Neuroradiol. 2004;31:183-189.

46. Lu W, Li YH, He XF. Treatment of large lumbar disc herniation withpercutaneous ozone injection via the posterior-lateral route and in-

    ner margin of the facet joint. World J Radiol. 2010;2:109-112.

47. Saal JA, Saal JS, Herzog RJ. The natural history of lumbar inter- vertebral disc extrusions treated nonoperatively. Spine (Phila Pa1976). 1990;15:683-686.
48. Alexandre A, Coro L, Azuelos A, et al. Intradiscal injection ofoxygen-ozone gas mixture for the treatment of cervical disc her-

    niations. Acta Neurochir Suppl. 2005;92:79-82.

49. Ogrendik M. Rheumatoid arthritis is an autoimmune diseasecaused by periodontal pathogens. Int J Gen Med. 2013;6:383-386.
50. Rhee DK, Marcelino J, Baker M, et al. The secreted glycoprotein lubricin protects cartilage surfaces and inhibits synovial cell over-growth. J Clin Invest. 2005;115:622-631.
51. Cassim B, Shaw OM, Mazur M, et al. Kallikreins, kininogens andkinin receptors on circulating and synovial fluid neutrophils: rolein kinin generation in rheumatoid arthritis. Rheumatology (Ox- ford). 2009;48:490-496.
52. Ma MH, Kingsley GH, Scott DL. A systematic comparison of combination DMARD therapy and tumour necrosis inhibitor ther- apy with methotrexate in patients with early rheumatoid arthritis.Rheumatology (Oxford). 2010;49:91-98.
53. Cho HY, Morgan DL, Bauer AK, Kleeberger SR. Signal trans- duction pathways of tumor necrosis factor–mediated lung in- jury induced by ozone in mice. Am J Respir Crit Care Med. 2007;175:829-839.
54. Fakhrzadeh L, Laskin JD, Laskin DL. Ozone-induced produc- tion of nitric oxide and TNF-alpha and tissue injury are depen- dent on NF-kappaB p50. Am J Physiol Lung Cell Mol Physiol.2004;287:L279-285.
55. Chen H, Yu B, Lu C, Lin Q. The effect of intra-articular injection of different concentrations of ozone on the level of TNF-alpha, TNF-R1, and TNF-R2 in rats with rheumatoid arthritis. Rheumatol Int. 2013;33:1223-1227.
56. Hasan O, Jessar M, Ashar M, Noordin S, Ahmad T. Systemic scle- rosis: Clinical manifestations, anesthetic and orthopedic consider- ations in a patient. Int J Surg Case Rep. 2018;42:24-28.
57. Nowicka D. Thermography improves clinical assessment in pa- tients with systemic sclerosis treated with ozone therapy. BioMed Research International. 2017;2017:7.
58. Buskila D. Fibromyalgia, chronic fatigue syndrome, and myofas- cial pain syndrome. Curr Opin Rheumatol. 1999;11:119-126.

59. Stone JR, Yang S. Hydrogen peroxide: a signaling messenger. An- tioxid Redox Signal. 2006;8:243-270.

60. Bocci V, Borrelli E, Travagli V, Zanardi I. The ozone paradox: ozone is a strong oxidant as well as a medical drug. Med Res Rev. 2009;29:646-682.

61. Bocci V. Ozone: A New Medical Drug. Dordrecht, The Nether- lands: Springer. 2005.

62. Bocci V. Oxygen-Ozone Therapy: A Critical Evaluation. Springer Science & Business Media. 2013.

63. Vélez BPL. Ozone therapy, a supplement for patients with fibro- myalgia. Revista Española de Ozonoterapia. 2014;4:39-49.

64. Clavo B, Perez JL, Lopez L, et al. Effect of ozone therapy on mus- cle oxygenation. J Altern Complement Med. 2003;9:251-256.
65. Giunta R, Coppola A, Luongo C, et al. Ozonized autohemotransfu-

sion improves hemorheological parameters and oxygen delivery to tissues in patients with peripheral occlusive arterial disease. Ann Hematol. 2001;80:745-748.

66. Ballardini E. Oxygen-ozone therapy for spinal muscle disorders in the horse. Vol 42005.

67. Bhatt J, Bhat A, Dhama K, Amarpal A. An overview of ozone ther- apy in equine: an emerging healthcare solution. J Exp Biol Agric Sci. 2016;4:S203-210.

68. Freburger JK, Holmes GM, Agans RP, et al. The rising prevalence of chronic low back pain. Arch Intern Med. 2009;169:251-258.

69. Alexandre A, Buric J, Paradiso R. Intradiscal injection of O2-O3 totreat lumbar disc herniations: Results at five years. Rivista Italiana di Ossigeno-Ozonoterapia. 2002;1:165-169.

70. Ueno I, Hoshino M, Miura T, Shinriki N. Ozone exposure gener- ates free radicals in the blood samples in vitro. Detection by the ESR spin-trapping technique. Free Radic Res.1998;29:127-135.

71. Borrelli E. Mechanism of action of oxygen ozone therapy in the treatment of disc herniation and low back pain. Acta Neurochir Suppl. 2011;108:123-125.

72. Bocci VPR, Pogni R, Corradeschi F, et al. Oxygen-ozone in orthopaedics: EPR detection of hydroxyl free radicals in ozone-treated “nucleus pulposus” material. Neuroradiol J.doi:10.1177/197140090101400106.

73. Torri G, Grazia AD, Casadei C. Clinical experience in the treat- ment of lumbar disk disease, with a cycle of lumbar muscle in- jection of an oxygen + ozone mixture. http://www.biaccabi.com/ edocs/um_torri.html. Accessed at 2018-09-03.

74. Magalhaes FN, Dotta L, Sasse A, Teixera MJ, Fonoff ET.Ozone therapy as a treatment for low back pain secondary to herniated disc: a systematic review and meta-analysis of randomized con- trolled trials. Pain Physician. 2012;15:E115-129.

75. Lee SY, Kim TH, Oh JK, Lee SJ, Park MS. Lumbar stenosis: a re- cent update by review of literature. Asian Spine J. 2015;9:818-828. 76. Baeza-Noci J. Spinal ozone therapy in lumbar spinal stenosis. Int J

Ozone Ther. 2007;6:17-24.
77. Rosenberg SK, Grabinsky A, Kooser C, Boswell MV.Effectiveness

of transforaminal epidural steroid injections in low back pain: a

one year experience. Pain Physician. 2002;5:266-270.
78. Abdi S, Datta S, F Lucas L. Role of epidural steroids in the man- agement of chronic spinal pain: a systematic review of effective-

ness and complications. Pain Physician. 2005;8:127-143.
79. Bocci V, Paulesu L. Studies on the biological effects of ozone 1. Induction of interferon gamma on human leucocytes. Haemato-

logica. 1990;75:510-515.
80. Di Filippo C, Cervone C, Rossi C, et al. Antiarrhythmic effect

of acute oxygen-ozone administration to rats. Eur J Pharmacol.

2010;629:89-95.
81. Pryor WA, Squadrito GL, Friedman M. A new mechanism for the

toxicity of ozone. Toxicol Lett. 1995;82:287-293.
82. Pryor WA, Squadrito GL, Friedman M. The cascade mechanism to explain ozone toxicity: the role of lipid ozonation products. Free

Radic Biol Med. 1995;19:935-941.
83. Mustafa MG. Biochemical basis of ozone toxicity. Free Radic Biol

Med. 1990;9:245-265.Received: 2018-05-30

Accepted: 2018-08-09

C-Editor: Yang LJ, Zhao M; S-Editor: Yu J; L-Editor: Wang L; T-Editor: Jia Y

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