Safety Evaluation of Far-UV-C Irradiation to Epithelial Basal Cells in the Corneal Limbus.

Basal cells in the corneal limbus play an important role in the turnover cycle because they are the source of all cells that constitute the corneal epithelium. We examined the penetration depth of ultraviolet (UV) light in the corneal limbus and assessed the safety of Far-UV-C on stem cells in the basal area of the corneal limbus. Rats were irradiated with UV at peaks of 207, 222, 235, 254 and 311 nm while under anesthesia. The UV penetration depth in the rat corneal limbal epithelium was wavelength dependent: 311 nm UV-B and 254 nm UV-C reached the basal cells of the epithelium, and 235 nm radiation reached the middle area; however, 207 and 222 nm UV-C reached only the superficial layer of the epithelium. Porcine cornea, which is similar to the human eye in size and structure, were irradiated with 222 and 254 nm UV-C. As in rats, 222 nm UV-C reached only the superficial layer of the porcine corneal limbal epithelium. These results indicate that Far-UV-C, such as radiation of wavelengths of 207 and 222 nm, could not reach corneal epithelial stem cells, i.e. the cells remained intact. It is unlikely that the turnover of the corneal epithelium is obstructed or disrupted by exposure to Far-UV-C.

Biological Impact of Shorter Wavelength Ultraviolet Radiation-C†.

Life on earth has constantly coped with the impact of solar radiation, especially solar ultraviolet radiation (solar UV). Various biological mechanisms protect us from solar UV. New devices emitting shorter wavelengths UV-C, i.e. <254 nm emitted by conventional UV germicidal lamps, have emerged. These shorter wavelength UV-C emitting devices are useful for various purposes, including microorganism inactivation. However, as solar UV-C does not reach the earth surface, biological impacts of UV-C has been studied using 254 nm germicidal lamps, and those using shorter wavelength UV-C is rarely known. To balance the utility and risk of UV-C, the biological effect of these new UV-C emitting devices must be investigated. In addition, our knowledge of biological impacts of the wavelength-dependent entire UV (100-400 nm) must be enhanced. In this review, we briefly summarize the biological impacts of shorter wavelength UV-C. Mechanisms of UV-C-induced cellular damage and factors affecting the microorganism inactivation efficiency of UV-C have been discussed. In addition, we theoretically estimate the probable photocarcinogenic action spectrum of shorter wavelength UV-C. We propose that increasing the knowledge on UV-C will facilitate the adoption of shorter wavelength UV-C emitting new devices in an optimal and appropriate manner.

Effect of ultraviolet C emitted from KrCl excimer lamp with or without bandpass filter to mouse epidermis.

It has been reported that 222-nm ultraviolet C (UVC) exerts a germicidal effect on bacteria and viruses as well as UV radiation emitted from a conventional germicidal lamp but is less toxic to the mammalian cells than that from a germicidal lamp. An excimer lamp filled with krypton chloride (KrCl) gas principally emits 222-nm UVC. However, the lamp also emits a wide band of wavelengths other than 222 nm, especially UVC at a longer wavelength than 222 nm and ultraviolet B, which cause DNA damage. There are some reports on the critical role of bandpass filters in reducing the harmful effect of UVC emitted from a KrCl excimer lamp in a human skin model and human subjects. However, the effectiveness of a bandpass filter has not been demonstrated in animal experiments. In the present study, mice were irradiated with UVC emitted from a KrCl excimer lamp with or without a bandpass filter. UVC emitted from an unfiltered KrCl lamp at doses of 50, 150 and 300 mJ/cm2 induced cyclobutyl pyrimidine dimer (CPD)-positive cells, whereas UVC emitted from a filtered lamp did not significantly increase CPD-positive cells in the epidermis. The present study suggested that the bandpass filter serves a critical role in reducing the harmful effect of emission outside of 222 nm to mouse keratinocytes.

Safety of 222 nm UVC Irradiation to the Surgical Site in a Rabbit Model.

For the prevention of surgical site infection (SSI), continuous disinfection could be helpful. Short wavelength ultraviolet radiation C (UVC) is highly bactericidal but shows cytotoxicity. Radiation of UVC with a wavelength of 222 nm to the skin is considered to be safe because it only reaches the stratum corneum. However, the safety of 222 nm irradiation to the surgical field not covered with skin is unknown. The purpose of this study was to examine the safety of 222 nm UVC irradiation on a surgical field in a rabbit model. Five types of tissue were surgically exposed and irradiated with 222 or 254 nm UVC. Immunohistological assessment against cyclobutane pyrimidine dimer (CPD), an index of DNA damage by UVC, was performed. The CPD-positive cell rate was significantly higher in the 254 nm group than in the other groups in all tissues. A 222 nm group showed significantly more CPD than control in fat tissue, but no significant difference in all other tissues. In fat tissue collected 24 h after irradiation, the 254 nm group showed higher CPD than the other groups, while the 222 nm group had reduced to the control level. These data suggest that 222 nm UVC irradiation could be a new method to safely prevent SSI.

Re-Evaluation of Rat Corneal Damage by Short-Wavelength UV Revealed Extremely Less Hazardous Property of Far-UV-C†.

Corneal damage-induced various wavelength UV (311, 254, 235, 222 and 207 nm) was evaluated in rats. For 207 and 222-UV-C, the threshold radiant exposure was between 10 000 and 15 000 mJ cm-2 at 207 nm and between 3500 and 5000 mJ cm-2 at 222 nm. Penetrate depth to the cornea indicated by cyclobutene pyrimidine dimer (CPD) localization immediately after irradiation was dependent on the wavelength. 311 and 254 nm UV penetrate to corneal endothelium, 235 nm UVC to the intermediate part of corneal stroma, 222 and 207 nm UVC only to the most outer layer of corneal epithelium. CPD observed in corneal epithelium irradiated by 222 nm UVC disappeared until 12 h after. The minimum dose to induce corneal damage of short-wavelength UV-C was considerably higher than the threshold limit value (TLV® ) promulgated by American Conference of Governmental Industrial Hygienists (ACGIH). The property that explains why UV-C radiation at 207 and 222 nm is extremely less hazardous than longer UV wavelengths is the fact that this radiation only penetrates to the outermost layer of the corneal epithelium. These cells typically peel off within 24 h during the physiological turnover cycle. Hence, short-wavelength UV-C might be less hazardous to the cornea than previously considered until today.

Evaluation of Acute Reactions on Mouse Skin Irradiated with 222 and 235 nm UV-C.

Biological response and DNA damage following irradiation with shorter wavelengths in the UV-C range were evaluated to investigate the safety at three wavelengths because of the recent emergence of germicidal equipment emitting short-wavelength UV-C for various purposes, including medical uses. To estimate an acceptable safety dose for human skin in the UV-C range, especially short UV-C, we studied the biological effects of 207, 222 and 235 nm UV-C using albino hairless mice and evaluated the inflammatory reactions in the skin. To explore an appropriate indicator to evaluate the biological response, we employed determination of the minimal perceptible response dose (MPRD), by which any subtle cutaneous response; erythema, edema and scale could be observed by visual inspection. Erythema was rarely observed, but edema and scale formation were evident for short UV-C wavelengths. The MPRD at 207, 222 and 235 nm was determined to be > 15, 15 and 2.0 kJ m-2 , respectively. These values could be thresholds and indicators for possible safety assessments. Our data suggest that the current human exposure limits for short UV-C wavelengths below 254 nm are overly restrictive and should be reconsidered for future disinfection lamps with short UV-C wavelengths.

Exploratory clinical trial on the safety and bactericidal effect of 222-nm ultraviolet C irradiation in healthy humans.

INTRODUCTION: Surgical site infection is one of the most severe complications of surgical treatments. However, the optimal procedure to prevent such infections remains uninvestigated. Ultraviolet radiation C (UVC) with a short wavelength has a high bactericidal effect; however, it is cytotoxic. Nonetheless, given that UVC with a wavelength of 222 nm reaches only the stratum corneum, it does not affect the skin cells. This study aimed to investigate the safety of 222-nm UVC irradiation and to examine its skin sterilization effect in healthy volunteers. METHODS: This trial was conducted on 20 healthy volunteers. The back of the subject was irradiated with 222-nm UVC at 50-500 mJ/cm2, and the induced erythema (redness of skin) was evaluated. Subsequently, the back was irradiated with a maximum amount of UVC not causing erythema, and the skin swabs before and after the irradiation were cultured. The number of colonies formed after 24 hours was measured. In addition, cyclobutene pyrimidine dimer (CPD) as an indicator of DNA damage was measured using skin tissues of the nonirradiated and irradiated regions. RESULTS: All subjects experienced no erythema at all doses. The back of the subject was irradiated at 500 mJ/cm2, and the number of bacterial colonies in the skin swab culture was significantly decreased by 222-nm UVC irradiation. The CPD amount produced in the irradiated region was slightly but significantly higher than that of the non-irradiated region. CONCLUSION: A 222-nm UVC at 500 mJ/cm2 was a safe irradiation dose and possessed bactericidal effects. In the future, 222-nm UVC irradiation is expected to contribute to the prevention of perioperative infection.

Viability evaluation of layered cell sheets after ultraviolet light irradiation of 222 nm.

INTRODUCTION: The objective of this study was to evaluate the cell viability of layered cell sheets, irradiated with 222 nm UV light. METHODS: UV transmittance of 222 nm and 254 nm was evaluated when the cell sheets of NCTC Clone 929 cells were irradiated UV light. Cell viability was evaluated after irradiation of 222 nm using 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyltetrazolium bromide (MTT) assay. Following irradiation of two layered cell sheets at 500 mJ/cm2, the cell damage of lower layers was evaluated by a colony formation and MTT assays. RESULTS: The UV transmittance of 222 nm was 10 times less than that of 254 nm. A MTT assay revealed that cells of cell sheets irradiated at 222 nm was less damaged than those at 254 nm, when irradiated at 5 mJ/cm2. Cell colonies were formed for cells of lower layers irradiated at 222 nm whereas no colony formation was observed for those irradiated at 254 nm. Significantly higher MTT activity was observed for cells of lower layers irradiated at 222 nm than at 254 nm. CONCLUSIONS: It is concluded that 222 nm irradiation is biologically safe for cell viability.

222-nm UVC inactivates a wide spectrum of microbial pathogens.

BACKGROUND: UVC has been used to inactivate several pathogens. Unlike the conventional 254-nm UVC, 222-nm UVC is harmless to mammalian cells. AIM: To investigate the disinfection efficacy of 222-nm UVC against human pathogens which are commonly found in the environment and healthcare facilities. METHODOLOGY: Staphylococcus aureus, Pseudomonas aeruginosa, Escherichia coli, Salmonella enterica subsp. serovar Typhimurium, Campylobacter jejuni, Bacillus cereus (vegetative cells and endospores), Clostridium sporogenes (vegetative cells and endospores), Clostoridioides difficile (endospores), Candida albicans (yeast), Aspergillus niger (hyphae and spores), Trichophyton rubrum (hyphae and spores), feline calicivirus and influenza A virus were irradiated with 222-nm UVC at various doses. The remaining live bacterial and fungal cells, and the viral infectivity were evaluated. The efficiency of 222-nm UVC germicidal effect was compared to that of the conventional 254-nm UVC. RESULTS: The 222-nm UVC showed potent germicidal effect to vegetative bacterial cells, yeast and viruses as efficient as the 245-nm UVC. The 222-nm UVC exhibited more potent germicidal effect to bacterial endospores, compared with the 254-nm UVC. The fungicidal effect of 222-nm UVC against the fungal spores and hyphae was weaker than that of 254-nm UVC. CONCLUSIONS: The 222-nm UVC is able to inactivate a wide spectrum of microbial pathogens. In comparison with the conventional 254-nm UVC, the germicidal effect of 222-nm UVC to the fungal hyphae and spores is low, but the 222-nm UVC exhibits strong germicidal effect to the bacterial endospores.

Long-term Effects of 222-nm ultraviolet radiation C Sterilizing Lamps on Mice Susceptible to Ultraviolet Radiation.

Germicidal lamps that emit primarily 254 nm ultraviolet radiation (UV) are routinely utilized for surface sterilization but cannot be used for human skin because they cause genotoxicity. As an alternative, 222-nm UVC has been reported to exert sterilizing ability comparable to that of 254-nm UVC without producing cyclobutane pyrimidine dimers (CPDs), the major DNA lesions caused by UV. However, there has been no clear evidence for safety in chronic exposure to skin, particularly with respect to carcinogenesis. We therefore investigated the long-term effects of 222-nm UVC on skin using a highly photocarcinogenic phenotype mice that lack xeroderma pigmentosum complementation group A (Xpa-) gene, which is involved in repairing of CPDs. CPDs formation was recognized only uppermost layer of epidermis even with high dose of 222-nm UVC exposure. No tumors were observed in Xpa-knockout mice and wild-type mice by repetitive irradiation with 222-nm UVC, using a protocol which had shown to produce tumor in Xpa-knockout mice irradiated with broad-band UVB. Furthermore, erythema and ear swelling were not observed in both genotype mice following 222-nm UVC exposure. Our data suggest that 222-nm UVC lamps can be safely used for sterilizing human skin as far as the perspective of skin cancer development.

Evaluation of acute corneal damage induced by 222-nm and 254-nm ultraviolet light in Sprague-Dawley rats.

Two hundred twenty-two nanometres ultraviolet (UV) light produced by a krypton-chlorine excimer lamp is harmful to bacterial cells but not skin. However, the effects of 222-nm UV light exposure to the eye are not fully known. We evaluated acute corneal damage induced by 222- and 254-nm UV light in albino rats. Under deep anaesthesia, 6-week-old Sprague-Dawley albino rats were exposed to UV light. The exposure levels of corneal radiation were 30, 150, and 600 mJ/cm2. Epithelial defects were detected by staining with fluorescein. Superficial punctate keratitis developed in corneas exposed to more than 150 mJ/cm2 of UV light, and erosion was observed in corneas exposed to 600 mJ/cm2 of UV light. Haematoxylin and eosin staining also showed corneal epithelial defects in eyes exposed to 254-nm UV light. However, no damage developed in corneas exposed to 222-nm UV light. Cyclobutane pyrimidine dimer-positive cells were observed only in normal corneas and those exposed to 254-nm UV light. Although some epithelial cells were stained weakly in normal corneas, squamous epithelial cells were stained moderately, and the epithelial layer that was detached from the cornea exposed to 600 mJ/cm2 of light was stained intensely in corneas exposed to 254-nm UV light. In the current study, no corneal damage was induced by 222-nm UV light, which suggested that 222-nm UV light may not harm rat eyes within the energy range and may be useful for sterilising or preventing infection in the future.

Chronic irradiation with 222-nm UVC light induces neither DNA damage nor epidermal lesions in mouse skin, even at high doses.

Surgical site infections (SSIs) represent an important clinical problem associated with increased levels of surgical morbidity and mortality. UVC irradiation during surgery has been considered to represent a possible strategy to prevent the development of SSI. 254-nm UVC induces marked levels of DNA damage by generating cyclobutyl pyrimidine dimers (CPD) in microorganisms. However, this effect is elicited not only in microorganisms, but also in human cells, and chronic exposure to 254-nm UVC has been established to represent a human health hazard. In contrast, despite short wavelength-UVC light, especially 222-nm UVC, having been demonstrated to elicit a bactericidal effect, single irradiation with a high dose of 222-nm UVC energy has been reported to not induce mutagenic or cytotoxic DNA lesions in mammalian cells. However, the effect of chronic irradiation with a high dose of 222-nm UVC to mammalian cells has not been determined. In this study, it was demonstrated that large numbers of CPD-expressing cells were induced in the epidermis of mice following treatment with a small amount of single exposure 254-nm UVC, and then less than half of these cells reduced within 24 h. Chronic 254-nm UVC irradiation was revealed to induce sunburn and desquamation in mouse skin. Histological analysis demonstrated that small numbers of CPD-expressing cells were detected only in hyperkeratotic stratum corneum after chronic irradiation with a high dose of 254-nm UVC, and that significant hyperplasia and intercellular edema were also induced in the epidermis of mice. In contrast, chronic irradiation with 222-nm UVC light was revealed not to induce mutagenic or cytotoxic effects in the epidermis of mice. These results indicated that 222-nm UVC light emitted from the lamp apparatus (or device), which was designed to attenuate harmful light present in wavelengths of more than 230 nm, represents a promising tool for the reduction of SSI incidence in patients and hospital staff.

Disinfection and healing effects of 222-nm UVC light on methicillin-resistant Staphylococcus aureus infection in mouse wounds.

UVC radiation is known to be highly germicidal. However, exposure to 254-nm-UVC light causes DNA lesions such as cyclobutane pyrimidine dimers (CPD) in human cells, and can induce skin cancer after long-term repeated exposures. It has been reported that short wavelength UVC is absorbed by proteins in the membrane and cytosol, and fails to reach the nucleus of human cells. Hence, irradiation with 222-nm UVC might be an optimum combination of effective disinfection and biological safety to human cells. In this study, the biological effectiveness of 222-nm UVC was investigated using a mouse model of a skin wound infected with methicillin-resistant Staphylococcus aureus (MRSA). Irradiation with 222-nm UVC significantly reduced bacterial numbers on the skin surface compared with non-irradiated skin. Bacterial counts in wounds evaluated on days 3, 5, 8 and 12 after irradiation demonstrated that the bactericidal effect of 222-nm UVC was equal to or more effective than 254-nm UVC. Histological analysis revealed that migration of keratinocytes which is essential for the wound healing process was impaired in wounds irradiated with 254-nm UVC, but was unaffected in 222-nm UVC irradiated wounds. No CPD-expressing cells were detected in either epidermis or dermis of wounds irradiated with 222-nm UVC, whereas CPD-expressing cells were found in both epidermis and dermis irradiation with 254-nm UVC. These results suggest that 222-nm UVC light may be a safe and effective way to reduce the rate of surgical site and other wound infections.