Antimicrobial photodynamic therapy: An overview.

Inflammatory periodontal disease caused by dental plaque is characterized by the clinical signs of inflammation and loss of periodontal tissue support. The mechanical removal of this biofilm and adjunctive use of antibacterial disinfectants and antibiotics have been the conventional methods of periodontal therapy. But the removal of plaque and the reduction in the number of infectious organisms can be impaired in sites with difficult access. The possibility of development of resistance to antibiotics by the target organism has led to the development of a new antimicrobial concept with fewer complications. Photodynamic therapy (PDT) involves the use of low power lasers with appropriate wavelength to kill micro organisms treated with a photosensitizer drug. PDT could be a useful adjunct to mechanical as well as antibiotics in eliminating periopathogenic bacteria.

INTRODUCTION

Periodontal disease results from inflammation of the supporting structure of the teeth and in response to chronic infection caused by various periodontopathic bacteria. In the treatment of periodontally involved teeth, current concepts are based on mechanical scaling and root planing to remove bacterial deposits, calculus, and cementum contaminated by bacteria and endotoxins.[12] However, removal of plaque and the reduction of the number of infectious cells can be impaired in sites with difficult access by mechanical scaling and root planing. Some therapeutic alternatives, such as systemic and local antibiotics, have been used in cases not responding to conventional treatments, although this therapy brings undesirable side effects and development of bacterial drug resistance.[34]

Photodynamic therapy (PDT) has emerged in recent years as a non – invasive therapeutic modality for the treatment of various infections by bacteria, fungi, and viruses.[5] This therapy is defined as an oxygen-dependent photochemical reaction that occurs upon light – mediated activation of a photosensitizing compound leading to the generation of cytotoxic reactive oxygen species, predominantly singlet oxygen.[6] PDT can be applied topically into a periodontal pocket avoiding overdoses and side effects associated with the systemic antimicrobial agent administration. It also minimizes the occurrence of bacterial resistance.[78]

Photodynamic antimicrobial chemotherapy represents an alternate antibacterial, antifungal, and antiviral treatment against drug - resistant organisms.[9] Applications of PDT in dentistry are growing rapidly. They are also used in the treatment of oral cancer, bacterial and fungal infections, and in the photodynamic diagnosis of the malignant transformation of oral lesions.[10] This review is aimed to discuss the role of PDT in periodontal and endodontic therapy.

Historical perspective of photodynamic therapy

The origin of light as a therapy in medicine and surgery are traced from antiquity to the modern day. Phototherapy began in ancient Greece, Egypt, and India, but disappeared for many centuries, only being rediscovered by the Western civilization at the beginning of the 20th century.[11] The use of contemporary photodynamic therapy was first reported by the Danish physician, Niels Finsen. He successfully demonstrated photodynamic therapy by employing heat – filtered light from a carbon – arc lamp (The Finsen Lamp) in the treatment of a tubercular condition of the skin known as Lupus Vulgaris.[11]

The concept of cell death induced by the interaction of light and chemicals was first reported by Osar Raab, a medical student working with Professor Herman Von Tappeiner in Munich. During the course of his study on the effects of acridine on paramecia cultures, he discovered that the combination of acridine red and light had a lethal effect on infusoria, a species of paramecium.[12] Subsequent work in the laboratory of Von Tappeiner coined the term “Photodynamic action” and showed that oxygen was essential. Much later, Thomas Dougherty and Co-workers at Roswell Park cancer institute, Buffalo, New York, clinically tested PDT. In 1978, they published striking results in which they treated 113 cutaneous or subcutaneous malignant tumors and observed a total or partial resolution of 111 tumors. The active photosensitizer used in this clinical PDT trial was called Hematoporphyrin Derivative. It was John Toth, who renamed it as PDT.[13]

PDT was approved by the Food and Drug Administration in 1999 to treat pre-cancerous skin lesions of the face or scalp.[14] PDT has emerged in recent years as a new non - invasive therapeutic option.

Mechanism of action

PDT involves three components: Light, a photosensitizer, and oxygen. The photosensitizer is administered to the patient, and upon irradiation with light of a specific wavelength, the photosensitizer undergoes a transition from a low energy ground state to an excited singlet state. Subsequently, the photosensitizer may decay back to its ground state with the emission of fluorescence or may undergo a transition to a higher energy triplet state.[15] The triplet state photosensitizer can react with biomolecules in two different pathways – type I and II.[16]

Type I reaction involves electron – transfer reactions between the excited state of the photosensitizer and an organic substrate molecule of the cells, producing free radicals. These free radical species are generally highly reactive and interact with endogenous molecular oxygen to produce highly reactive oxygen species, such as superoxide, hydroxyl radicals, and hydrogen peroxide, which are harmful to cell membrane integrity, causing irreparable biological damage.[117]

In type II reaction, the triplet state photosensitizer reacts with oxygen to produce an electronically excited and highly reactive state of oxygen, known as singlet oxygen (1O2) which can interact with a large number of biological substrates inducing oxidative damage on the cell membrane and cell wall. Microorganisms that are killed by singlet oxygen include viruses, bacteria, and fungi. Singlet oxygen has a short lifetime in biological systems and a very short radius of action (0.02 μm). Hence, the reaction takes place within a limited space, leading to a localized response; thus making it ideal for application to localized sites without affecting distant cells or organs. Thus, the type II reaction is accepted as the major pathway in microbial cell damage.[1718]

Light source

PDT requires a sources of light to activate the photosensitizer by exposure to low power visible light at a specific wavelength. Most photosensitizers are activated by red light between 630 and 700 nm, corresponding to a light penetration depth from 0.5 cm to 1.5 cm.[1920] This limits the depth of necrosis. The total light dose, dose rates, and the depth of destruction vary with each tissue treated and photosensitizer used.[2021] Currently, the light source applied in photodynamic therapy are those of helium – neon lasers (633 nm), gallium – aluminum – arsenide diode lasers (630-690, 830 or 906 nm), and argon laser (488-514 nm), the wavelength of which range from visible light to the blue of argon lasers, or from the red of helium-neon laser to the infra red area of diode lasers.[2223] Recently, non laser light source, such as light – emitting diodes (LED), has been used as new light activators in PDT. LED devices are more compact, portable, and cost effective compared to traditional lasers.[1]

Photosensitizers

An optimal photosensitizer must possess photo-physical, chemical, and biological characteristics. Most of the sensitizers used for medical purposes belong to the following basic structure.

Tricyclic dyes with different meso-atoms E.g.: Acridine orange, proflavine, riboflavin, methylene blue, fluorescein, and erythrosin.

Tetrapyrroles. E.g.: Porphyrins and derivatives, chlorophyll, phylloerythrin, and phthalocyanines.

Furocoumarins. E.g.: Psoralen and its methoxy-derivatives, xanthotoxin, and bergaptene.[24]

Photofrin and hematophyrin derivatives are referred to as first generation sensitizers. Second generation photosensitizers include 5–aminolevulinic acid (ALA), benzoporphyrin derivative, texaphyrin, and temoporfin (mTHPC). These photosensitizers have greater capability to generate singlet oxygen. Topical ALA have been used to treat pre-cancer conditions, and basal and squamous cell carcinoma of skin.[2526]

In antimicrobial PDT, photosensitizers used are toluidine blue O and methylene blue. Both have similar chemical and physicochemical characteristics. Toluidine blue O is a solution that is blue – violet in color. It stains granules within mast cells and proteoglycans/glycosaminoglycans within connective tissues. Methylene blue is a redox indicator that is blue in an oxidizing environment and becomes colorless upon reduction. Methylene blue combined with light has been reported to be beneficial in killing the influenza virus, Helicobacter pylori, and C. albicans.[2728] Methylene blue and toluidine blue O are very effective photosensitizing agents for the inactivation of both gram-positive and gram-negative periodontopathic bacteria. Tetracyclines used as antibiotics in periodontal diseases are also effective photosensitizers producing singlet oxygen.[29]

Application of photodynamic therapy in dentistry

Antimicrobial PDT can be considered as an adjunctive to conventional mechanical therapy. The liquid photosensitizer placed directly in the periodontal pocket can easily access the whole root surface before activation by the laser light through an optical fiber placed directly in the pocket.[30] As a result of the technical simplicity and the effective bacterial killing, the application of PDT in the treatment of periodontal diseases has been studied extensively.

Antimicrobial PDT not only kills the bacteria, but may also lead to the detoxification of endotoxins such as lipopolysaccharide. These lipopolysaccharides treated by PDT do not stimulate the production of pro-inflammatory cytokines by mononuclear cells. Thus, PDT inactivate endotoxins by decreasing their biological activity.[31]

It has been demonstrated that bacteria associated with periodontal disease can be killed through photosensitization with toulidine blue O by irradiating with helium – neon soft laser.[23] Data from an in vitro study indicated that PDT could kill bacteria organized in a biofilm.[32] In an animal study, it was found that PDT was useful in reducing the redness, bleeding on probing, and porphyromonas gingivalis levels.[33]

A randomized controlled clinical study compared the effects of PDT alone without sub gingival SRP to sub gingival SRP in subject with aggressive periodontitis. At three months following the therapy, both treatment yielded comparable outcomes in terms of reduction of bleeding on probing and probing depth (PD), gains in clinical attachment level (CAL), thus suggesting a potential clinical benefits of PDT.[34]

Christodoulides et al. evaluated the clinical and microbiologic effects of the adjunctive use of PDT to non - surgical periodontal treatment. Twenty four subjects with chronic periodontitis were randomly treated with scaling and root planing followed by a single episode of PDT. The additional application of a single episode of PDT to scaling and root planing failed to result an additional improvement in terms of pocket depth reduction and clinical attachment level gain, but it resulted in a significant reduction in bleeding scores compared to scaling and root planing alone.[35]

Bhatia et al. demonstrated that the optimal concentration of toluidine blue O to kill P. gingivalis was 12.5 μg/ml with helium-neon laser irradiations. This was caused by the disruption of outer membrane proteins of these bacteria.[3637] Chan and Lai showed that the presence of methylene blue at the wavelength of 632.8 nm (helium-neon laser) and 665 and 830 nm (diode laser) has a high bactericidal effect on periodontal pathogens.[38]

Yilmaz et al. randomly assigned a total of 10 patients to receive repeated application of scaling and root planing with photodynamic therapy the other groups were receiving only scaling and root planing, photodynamic therapy, and oral hygiene instructions. Methylene blue served as the photosensitizer and was used as a mouth rinse. Significant clinical and microbiological improvement was seen within groups receiving scaling and root planing with photodynamic therapy and the scaling and root planing alone. However, improvement in groups receiving photodynamic therapy alone, as well as those receiving only oral hygiene instructions, did not reach significant levels. The reduced effectiveness of PDT may be the due the application of PDT from the external surface of the gingiva.[39] Several studies have demonstrated bactericidal and detoxification effects on high-level lasers on contaminated dental implant surfaces. High-level lasers have been used successfully in the surgical management of peri - implantitis.[40] In an in vitro study, Hass et al. examined the efficacy of PDT in killing bacteria associated with peri - implantitis which adhered to titanium plates with different surface characteristics. Scanning electron microscopic analysis showed that antimicrobial photodynamic therapy led to bacterial cell destruction without damaging the titanium surface.[40] Similar antimicrobial results were obtained by Shibli et al. who reported that PDT could reduce the bacterial count of P. intermedia, P. nigrescens, Fusobacterium spp. in ligature induced peri - implantitis of dogs.[41]

On interpreting the data from the various controlled clinical studies, it becomes obvious that in patients with chronic periodontitis, aggressive periodontitis and peri implantitis, the adjunctive use of PDT to scaling and root planing may result in greater clinical attachment level gains, reduction in bleeding on probing and probing pocket depths. PDT has advantage such as reducing the treatment time, no need for anesthesia, destruction of bacteria, inactivation of endotoxins, and unlikely development of resistance by the target bacteria and no damage to the adjacent host tissues.[24]

In operative dentistry, it is proved that the antimicrobial photodynamic therapy technique is effective for the treatment and prevention of dental caries.[42] Endodontic failures are caused by the proliferation of residual bacteria that are left behind within the root canal due to the complexity of the root canal system that makes complete debridement with instrumentation and irrigation alone almost impossible. Antimicrobial photodynamic therapy has been reported to be effective as an adjunct to conventional endodontic disinfection treatment to destroy the bacteria that remain even after irrigation with sodium hypo chlorite.[43]

Soukos et al. conducted a study to investigate the effect of PDT on endodontic pathogens in planktonic phase as well as on Enterococcus faecalis biofilms in experimentally infected root canals of extracted teeth. Strains of microorganisms were sensitized with methylene blue (25 μg/ml) for five minutes followed by exposure to red light of 665 nm with an energy florescence of 30 J/cm2. Methylene blue fully eliminated all bacterial species with the exception of E. faecalis (53% killed). The same concentration of methylene blue in combination with red light (222 J/cm2) was able to eliminate 97% of E. faecalis biofilm bacteria in root canals using an optical fiber with multiple cylindrical diffusers that uniformly distributed light at 360°. Hence, PDT may be developed as an adjunctive procedure to kill residual bacteria in the root canal system after standard endodontic treatment.[44]

The study by Garcez et al. reported the antimicrobial effect of PDT combined with endodontic treatment in patients with necrotic pulp infected with microflora resistant to a previous antibiotic therapy. The use of PDT added to conventional endodontic treatment leads to a further major reduction of microbial load. PDT is an efficient treatment to kill multi-drug resistant microorganisms.[45]

Adverse effects

The risk and side effects of antimicrobial PDT are basically classified into two categories.

Relates to the effect of light energy.

Relates to the photosensitizer and the photo chemical reaction.[1]

The potential inadvertent irradiation of the patients eyes must be strictly avoided during treatment, even though the laser power employed is very low. The use of protective glasses by the patient, the operator and the assistant is recommended. During treatment with high level lasers, thermogenesis occurs as a result of the interaction of the laser with the tissues. PDT as a low level therapy, using a diode laser with short irradiation time, does not produce any thermal changes within the gingival tissues and root surfaces. With regard to photosensitizers and photochemical reactions, it is important to apply antimicrobial photodynamic therapy to stain and kill selectively the targeted bacteria without adversely affecting the surrounding periodontal tissues.[137]

Perspectives and future directions

The numerous studies cited in this review indicate that PDT appears to be most efficient for treatment of localized and superficial infections. Thus, infections in the oral cavity such as mucosal and endodontic infections, periodontal diseases, caries, and peri - implantitis are potential targets. PDT will not replace antimicrobial chemotherapy, but may improve the treatment of oral infections, accelerating and lowering the cost of the treatment. Development of new photosensitizers, more efficient light delivery systems, and further studies are required to establish the optimum treatment parameters.

CONCLUSION

Antimicrobial PDT seems to be a unique and interesting therapeutic approach towards periodontal and endodontic therapy. The numerous in vitro studies have clearly demonstrated the effective and efficient bactericidal effect of PDT. However, the superior effects of the adjunctive use of PDT have not been demonstrated clinically or in vivo in either periodontal or endodontic therapy. There is a great need to develop an evidence based approach to the use of PDT for the treatment of periodontitis, peri-implantitis, and endodontic therapy. It would be prudent to say that there is an insufficient evidence to suggest that PDT is superior to the traditional modalities of periodontal and endodontic therapy. Further, randomized long term clinical studies and meta analyses are necessary to demonstrate the beneficial effect of antimicrobial photodynamic therapy, and in comparison with conventional methods. Antimicrobial photodynamic therapy may hold promise as a substitute for currently available chemotherapy in the treatment of periodontal and peri - implant diseases, and in endodontic therapy.