Ozone therapy for skin diseases: Cellular and molecular mechanisms.

Ozone is a highly reactive oxidant molecule consisting of triatomic oxygen atoms. Ozone therapy can be achieved using ozonated hydrotherapy, ozonated oil, ozone autohemotherapy, and other innovative dosage forms of ozone products. Ozone is frequently used as a complementary therapy for various cutaneous diseases, including infectious skin diseases, wound healing, eczema, dermatitis, psoriasis, axillary osmidrosis, diabetic foot, and pressure ulcers. In addition, several studies have reported the superior potential of ozone therapy for improving skin and gut microbiomes, as well as antitumour and antiaging treatment. Ozone therapy is an emerging treatment strategy that acts via complex mechanisms, including antioxidant effects, immunomodulatory capacity, and modulation of local microcirculation. Studies assessing the mechanism of ozone have gradually expanded in recent years. This review article aims to summarise and explore the possible molecular biological mechanisms of ozone in cutaneous diseases and provide compelling theoretical evidence for the application of ozone in cutaneous diseases.

Therapeutic efficacy of metronidazole by needle-free jet injection combined with blue light therapy in Moderate-to-Severe facial acne vulgaris.

BACKGROUND: Acne vulgaris is one of the most common dermatological diseases. Some topical treatments for acne used in combination, such as blue light and topical antibiotics (such as metronidazole) by needle-free jet injection (NFJI), are becoming prevalent in clinical practice, but the efficacy remains uncertain. METHODS: In order to investigate the effect of blue light combined with metronidazole by NFJI in the treatment of acne, the 251 enrolled patients were randomly assigned into the blue light group, metronidazole (MNZ) group, and MNZ + blue light group, and then received 6-weeks' treatment. A variety of objective and subjective methods such as clinical pictures, skin barrier physiological parameters (including trans-epidermal water loss (TEWL), stratum corneum hydration, facail surface sebum, erythema and pigmentation), the Investigator Global Assessment score, acne lesion count assessment, Patients' Self-Assessment, and VAS score were used to evaluate the efficacy and side effects of the treatments. RESULTS: Compared to the baseline, the MNZ + blue light group showed significant improvement in acne lesion count reduction, TEWL, straum corneum hydration, facial surface sebum and erythema (p < 0.05). The MNZ + blue light group showed significant differences compared with the MNZ group and blue light group in terms of acne lesion count reduction and erythema (p < 0.05) Compared to the MNZ group, the MNZ + blue light group demonstrated significant improvement in TEWL and sebum (p < 0.05). While compared to the blue light group, the MNZ + blue light group showed significant improvement in hydration (p < 0.05). There was no statistically significant difference among the three groups in pigmentation (p > 0.05). CONCLUSION: The combination of MNZ by NFJI and blue light has a synergistic effect and can relieve acne skin lesion within 6 weeks in the treatment of moderate and moderate-to-severe facial acne vulgaris, meanwhile, this method has a good safety.

Downregulated Serum Exosomal miR-451a Expression Correlates With Renal Damage and Its Intercellular Communication Role in Systemic Lupus Erythematosus.

Systemic lupus erythematosus (SLE) is a multi-system autoimmune disease characterized by continuous inflammation and the production of autoantibodies. Exosomes, acting as a critical tool for communication between cells, are involved in the pathogenesis of SLE, particularly in inflammation and immune imbalance. In this study, we aimed to extract and confirm the pro-inflammatory effect of serum exosomes in SLE. Then, we attempted to find differentially expressed exosomal microRNAs in the serum of healthy subjects and SLE patients via miRNA microarray analysis and validated the target exosomal microRNA, exosomal miR-451a, which expression level decreased in serum of SLE patients by RT-qPCR. Furtherly, we analyzed the correlation between exosomal miR-451a and disease activity, kidney damage and typing, and traditional medicine therapy. Finally, we investigated the intercellular communication role of exosomal miR-451a in SLE by co-culture assay in vitro. Taken together, our study demonstrated that downregulated serum exosomal miR-451a expression correlated with SLE disease activity and renal damage as well as its intercellular communication role in SLE which provided potential therapeutic strategies.

Ozonated autohemotherapy elevates PPAR-γ expression in CD4+ T cells and serum HDL-C levels, a potential immunomodulatory mechanism for treatment of psoriasis.

Psoriasis is widely accepted as a metabolic syndrome with significantly abnormal lipid metabolism and high level of blood lipids that induce a persistent low level of inflammatory condition in patients. T cell mediated immune response plays a critical role in the occurrence and persistence of psoriasis lesions. Hyperlipidemia and associated inflammatory reaction are believed to be the major risk factors for the onset and recurrence of psoriasis. Peroxisome proliferator activated receptor-gamma (PPAR-γ) is known to effectively regulate the blood lipid level and inhibit inflammatory reaction. In this study, we examined the efficacy of ozonated autohemotherapy (OAHT) treatment on psoriatic patients by evaluating the Psoriasis Area and Severity Index (PASI) score and blood lipid level. In addition, PPAR-γ expression level and the correlation of PASI scores or blood lipid level with the PPAR-γ expression were also assessed to determine the psoriasis-associate targets of OAHT. We found that OAHT significantly decreased patients' PASI scores and increased blood HDL-C level. Furthermore, we found that PPAR-γ expression in CD4+ T cells from psoriasis patients was significantly lower than healthy controls, and OAHT treatment increased the expression of PPAR-γ. In conclusion, OAHT attenuates the psoriatic severity in patients and increased blood HDL-C level, which may be associated with increased PPAR-γ expression. Our data suggests that OAHT is an effective treatment in psoriasis and deserves further evaluations in clinical applications.

Ozone Therapy Attenuates NF-κB-Mediated Local Inflammatory Response and Activation of Th17 Cells in Treatment for Psoriasis.

Ozone therapy has been widely used to treat many skin diseases, including infections, allergic dermatosis, and skin ulcers. However, its efficacy as a treatment for psoriasis is unclear. In this study, we explored the clinical efficacy and the underlying molecular mechanisms of ozone therapy on psoriasis. We found that topical ozone treatment significantly decreased patients' psoriasis area and severity index (PASI) scores and the expression of psoriasis-associated cytokines in their peripheral blood CD4+ T cells. In the IMQ-induced psoriasis mouse model, topical ozone treatment significantly inhibited the formation of IMQ-induced psoriasis-like lesions and the expression of psoriasis-associated inflammatory factors. High-throughput sequencing confirmed that IMQ-induced activation of toll-like receptor 2 (TLR2)/ nuclear factor-κB (NF-κB) signaling pathway was significantly suppressed in psoriasis-like lesions after topical ozone treatment. Furthermore, the activation of spleen T helper (Th) 17 cells was blocked in the mouse model; this was associated with the downregulation of cytokines and NF-κB pathways upon topical ozone treatment. Ozone therapy can attenuate local inflammatory reactions and the activation of Th17 cells in psoriasis by inhibiting the NF-κB pathway. Our results show that ozone therapy is effective in treating psoriasis. We recommend further evaluations for its clinical applications.

Ozone therapy promotes the differentiation of basal keratinocytes via increasing Tp63-mediated transcription of KRT10 to improve psoriasis.

Psoriasis is a chronic immune-mediated inflammatory dermatosis. Recently, ozone therapy has been applicated to psoriasis treatment; however, the mechanism by which ozone therapy improves psoriasis remains unclear. The excessive proliferation and the differentiation of basal keratinocytes have been considered critical issues during pathological psoriasis process, in which keratin 6 (KRT6) and KRT10 might be involved. In the present study, KRT6, IL-17 and IL-22 protein within psoriasis lesions was decreased, while KRT10 and Tp63 protein in psoriasis lesions was increased by ozone treatment in both patient and IMQ mice psoriatic tissues. In the meantime, ozone treatment down-regulated KRT6 mRNA and protein expression while up-regulated KRT10 mRNA and protein expression within IL-22 treated primary KCs; the cell viability of KCs was suppressed by ozone treatment. Moreover, Tp63 bound to KRT10 promoter region to activate its transcription in basal keratinocytes; the promotive effects of ozone on Tp63 and KRT10 were significantly reversed by Tp63 silence. Both TP63 and KRT10 mRNA expression were significantly increased by ozone treatment in psoriasis lesions; there was a positive correlation between Tp63 and KRT10 expression within tissue samples, suggesting that ozone induces the expression of Tp63 to enhance the expression of KRT10 and the differentiation of keratinocytes, therefore improving the psoriasis. In conclusion, the application of ozonated oil could be an efficient and safe treatment for psoriasis; ozone promotes the differentiation of keratinocytes via increasing Tp63-mediated transcription of KRT10, therefore improving psoriasis.

Topical ozone therapy restores microbiome diversity in atopic dermatitis.

BACKGROUND: Staphylococcus aureus (S. aureus) accounts for 90% of the microbiome in atopic dermatitis (AD) lesions and plays a role in disease flare-ups and worsens disease outcome. Ozone treatment can improve AD conditions by its bactericidal effect on S. aureus. OBJECTIVE: To study the effects of topical ozone therapy on microbiome diversity in AD lesions and explore potential probiotic pathogens correlated with AD progression. METHODS: Patients with moderate to severe bilateral skin lesions in AD were recruited. Randomized split sides were performed. One side was treated with ozone hydrotherapy followed by ozonated oil; while the contralateral side with tap water and basal oil. Patients' SCORAD scores and modified EASI were recorded before and after treatments. The microbiological compositions in targeting sites were determined using 16S rDNA sequencing. RESULTS: After three-day ozone therapy, patients showed a significant decrease in SCORAD scores and inflammatory cell infiltration in AD lesions. The micro-ecological diversity was higher in the non-lesional as compared with lesional areas (p < 0.05), which was also negatively correlated with the severity of AD (r = -0.499, p < 0.05). The proportion of S. aureus in AD lesions was positively correlated with the severity of AD (r = 0.564, p = 0.010), which was decreased after ozone treatment (p = 0.07). Ozone therapy showed an increase in microbiological diversity with a significant increase in the proportion of Acinetobacter (p < 0.05). CONCLUSION: Topical ozone therapy is highly effective for treatment for AD. It can change the proportional ratio of Staphylococcus and Acinetobacter, thereby restoring the microbiological diversity in AD lesions.

Microbiosis in pathogenesis and intervention of atopic dermatitis.

Atopic dermatitis (AD) is a chronic, non-contagious, inflammatory skin disorder characterized by relapsing eczematous lesions. Its pathogenesis remains incompletely understood. The current evidence has emerged to show that skin and gut microbiome play critical roles in the pathogenesis and progression of AD. Skin mircrobiome mainly refers to skin commensal organisms that promote normal immune system functions and prevent the colonization of pathogens; while gut microbiome can modulate immunologic, metabolic and neuroendocrine functions. With the current knowledge of microbiome effects on the onset of the disease, there are evolving multifarious interventions targeting microbiome for the treatment of AD. In this report, we have reviewed the critical roles of microbiosis in the pathogenesis of AD, summarized potential mechanisms mediated by microbiosis and aimed to enlighten a theoretical basis for its therapeutic applications in the treatment of AD.

[Bactericidal effect of ozonated camellia oil on Staphylococcus aureus in vitro].

OBJECTIVE: To explore a new method for detecting the bactericidal effect of oiling agent in vitro, and to determine the disinfectant effecacy of ozonated camellia oil on Staphylococcus aureus.
 Methods: Suspension of Staphylococcus aureus was prepared and innoculated on the LB plate by plate scribing method. After culture overnight, 21 bacterial monoclones with the same diameter were selected and divided into 3 groups: A negative control group, a baseoil (camellia oil) group and an ozonated camellia oil group. We used a ring to isolate the single clone and added oil inside the ring, cultured the whole plate over night, picked out each single clone (with gel) to 5 mL LB medium and cultured it for 12 h. The final concentration of the LB medium was detected by plate count method and turbidimetry.
 Results: According to the plate count method and turbidimetry, the bacterial concentration in the ozonated camellia oil group was lower than that in the negative control group and base oil group (P<0.001).
 Conclusion: Bacterial monoclone culture method shows that ozonated camellia oil can significantly kill Staphylococcus aureus, and this method is an effective method for evaluating the bactericidal function of the oiling agent in vitro.

[Safety evaluation for medical ozone oil on skin].

OBJECTIVE: To evaluate skin irritation, acute toxicity, and allergy of medical ozone oil for clinical application.
 Methods: In contrast to their left and right side irritation, one or more skin irritation tests were performed on the intact and damaged skins of guinea pigs. With the maximum concentration, acute skin toxicity test was applied on the intact and damaged skins of rats.Active cutaneous anaphylaxis was applied to the guinea pigs.
 Results: High concentration (ozone consumption: 150 g/L) of medical ozone oil showed a slight irritation on the broken skin of guinea pigs, while low concentrations did not show skin irritation.Medical ozone oil had no obvious acute toxicity to rats. The medical ozone oil and base oil showed mildallergy for the skin of guinea pig.
 Conclusion: The irritation of medical ozone oil is related to its concentration. With appropriateconcentration and duration of treatment, medical ozone oil is safe.

Efficacy of combination of ozonated water with oil for treatment of tinea pedis.

OBJECTIVE: To evaluate efficacy of combined therapy with ozonated water and oil on patients with tinea pedis.
 Methods: A total of 60 patients with tinea pedis were divided into 2 groups in a randomized and blinded test. Patients in a control group were treated with naftinfine hydrochloride and ketoconazole cream once a day. Patients in an ozone group were treated with ozonated water bath and then ozonated oil topical application once a day. Patients in the 2 groups were treated for 4 weeks. Clinical and laboratory data were collected for both groups at the end of the 1st week, the 2nd week, and the 4th week. The Pearson chi-square was performed to compare scores of the clinical signs and symptoms (CSS) and the mycological result between the 2 groups. Independent samples T-test was performed to compare the curative effect between the 2 groups.
 Results: After 4 weeks' treatment, 6 patients were positive in the control group determined by mycological examination while 1 patient was positive in the ozone group, with no significant difference between the 2 groups (P>0.05). Changes in CSS at the end of the 1st week, 2nd week, and 4th week were obtained and showed no significant difference between the 2 groups at the 3 different time points (P>0.05). No side effects were observed.
 Conclusion: Combination of ozonated water with oil is effective on treatment of tinea pedis and it shows no side effects.

[Effect of ozone on Staphylococcus aureus colonization in patients with atopic dermatitis].

OBJECTIVE: To verify the effect of ozone on Staphylococcus aureus (S. aureus) colonization in patients with atopic dermatitis (AD) and its correlation with the patient's status.
 Methods: A total of 12 patients with moderate or severe AD, aged from 6 to 65 years, were recruited from outpatient of the Third Xiangya Hospital. The treatment sides were showered with ozonated water and smeared with ozonated oil for 7 days (twice a day), while the control sides were washed with warm running water and smeared with base oil. At different time points, the severity scoring of atopic dermatitis (SCORAD) scores, sleep and pruritus scores were assessed and compared between the two sides. Meanwhile, plate cultivation was used to quantitatively detect the changes of S. aureus colonization in skin lesions.
 Results: After 7 days treatment, erythema and pimples were decreased in the treatment sides. The clear skin texture, smooth skin, improved skin lesions were also observed by dermoscopic examination. The results of reflectance confocal microscopy (RCM) demonstrated that the parakeratosis was improved, the structures were clearer, and the inflammatory cells infiltration was reduced after ozone treatment for 7 days. After ozone treatment for 3 and 7 days, the S. aureus colonization in the treatment sides decreased by (75.55±21.81)% and (97.24±2.64)% respectively. Compared to that of control sides, the percentage of S. aureus colony after ozone treatment for 7 days decreased significantly (P<0.01). After ozone treatment for 7 days, the SCORAD scores, sleep and pruritus scores were significantly decreased (all P<0.01). There was a linear correlation between the decreasing percentage of S. aureus colony and the declining percentage of SCORAD scores in AD patients.
 Conclusion: Topical ozone therapy can effectively reduce S. aureus colony in skin lesions and alleviate the severity of AD patients with moderate to severe degree.

[Topical ozone application: An innovative therapy for infantile atopic dermatitis].

OBJECTIVE: To evaluate the clinical efficacy and safety of the innovative topical ozone therapy for infantile atopic dermatitis.
 Methods: Sixty children with atopic dermatitis were divided into a treatment group and a control group. The treatment group was showered with ozonated water (3-5 times a week) and smeared with ozonated oil (twice a day), while the control group was washed with warm running water and smeared with base oil, adding moisturizer if necessary. The treatment course was 2 weeks. Efficacy and side effect were evaluated.
 Results: The skin exudation was reduced and erosion was healing after 3-5 days topical ozone therapy for infantile atopic dermatitis. The effective rates were 80.0% and 20.0% in the treatment group and control group for 1 week, and 89.6% and 30.7% for 2 weeks, respectively, with significant difference between the 2 groups (P<0. 001).
 Conclusion: Innovative treatment of infantile atopic dermatitis with topical ozone application is safe and effective, which is worth popularizing in clinic.

[Topical ozone therapy: An innovative solution to patients with herpes zoster].

OBJECTIVE: To observe the clinical efficacy and safety of topical ozone therapy for patients with herpes zoster by reflectance confocal microscopy (RCM).
 Methods: A total of 60 patients with herpes zoster were divided into a control group and an ozone treatment group (n=30). In the control group, patients took oral valacyclovir tablets or granules (0.3 g per day, three times a day) and they were subjected to local weak laser irradiation treatment plus topical 2% mupirocin ointment twice a day. In the ozone group, the treatment is same as the control group except mupirocin ointment was replaced with topical ozone treatment (hydrotherapy every day plus ozonated oil twice a day). The clinical symptoms, discoid cell and adverse reactions were observed and taken records at day 0, 3, 7 and 14. Statistical analysis was performed to compare the clinical efficacy between the 2 groups. 
 Results: On the seventh day of treatment, the discoid cells of the ozone group disappeared, and the difference between the control group and the ozone group was statistically significant (P<0.05). The difference of decreased percentage of pain scores at each time point between the 2 groups was statistically significant (P<0.05). The clinical efficacy was 100% in the ozone group and 86.7% in the control group, with significant difference between the 2 groups (P<0.05).
 Conclusion: Topical ozone therapy in patients with herpes zoster is helpful in relieving pain, shortening the course as well as improving the clinical efficacy without obvious adverse reactions. It is worth to be popularized.

The antibacterial effect of topical ozone on the treatment of MRSA skin infection.

Skin can be infected by many types of microorganisms, most commonly by gram­positive strains of Staphylococcus and Streptococcus spp. Treatment of Staphylococcus aureus (S. aureus) infections, particularly that of methicillin resistant Staphylococcus aureus (MRSA), is a challenge in clinical practice. Ozone therapy has proven to be one of the strongest antiseptics against the majority of microorganisms involved in skin infections. The purpose of the present study was to evaluate the microbicidal effects of topical ozone therapy on S. aureus and MRSA, and determine the clinical efficacy of ozone therapy on patients with MRSA skin infection. Microbicidal effects of ozonated oil and ozonated water were determined by plating and Kirby Bauer methods. Clinical efficacy and safety of topical ozone were evaluated in two cases with skin MRSA infection. The killing rates of ozonated oil for S. aureus and MRSA were greater when compared with the control oil group. Almost 100% of S. aureus were eliminated by ozonated oil following 5 min. Almost 100% MRSA were eliminated by ozonated oil following 15 min. In addition, 100% S. aureus and 100% MRSA were eliminated by ozonated water in 1 min. The inhibition zone diameters of ozonated oil for S. aureus and MRSA were 17 and 13 mm, respectively, which were significantly larger than the control group. Both cases of skin MRSA infection were completely healed with ozone therapy. In conclusion, ozone therapy is a potential treatment for S. aureus and MRSA skin infection as it has great efficacy, few side effects and low­costs.

Ganoderma lucidum polysaccharides protect fibroblasts against UVB-induced photoaging.

Ganoderma lucidum has featured in traditional Chinese medicine for >1,000 years. Ganoderma polysaccharides (GL-PS), a major active ingredient in Ganoderma, confer immune regulation, antitumor effects and significant antioxidant effects. The aim of the present study was to investigate the efficacy and mechanism of GL­PS­associated inhibition of ultraviolet B (UVB)­induced photoaging in human fibroblasts in vitro. Primary human skin fibroblasts were cultured, and a fibroblast photoaging model was built through exposure to UVB. Cell viability was measured by MTT assay. Aged cells were stained using a senescence­associated ß-galactosidase staining (SA­ß­gal) kit. ELISA kits were used to analyze matrix metalloproteinase (MMP) ­1 and C­telopeptides of Type I collagen (CICP) protein levels in cellular supernatant. ROS levels were quantified by flow cytometry. Cells exposed to UVB had decreased cell viability, increased aged cells, decreased CICP protein expression, increased MMP­1 protein expression, and increased cellular ROS levels compared with non­exposed cells. However, cells exposed to UVB and treated with 10, 20 and 40 µg/ml GL­PS demonstrated increased cell viability, decreased aged cells, increased CICP protein expression, decreased MMP­1 protein expression, and decreased cellular ROS levels compared with UVB exposed/GL­PS untreated cells. These results demonstrate that GL­PS protects fibroblasts against photoaging by eliminating UVB­induced ROS. This finding indicates GL­PS treatment may serve as a novel strategy for antiphotoaging.