Photodynamic antimicrobial chemotherapy (PACT) with methylene blue increases membrane permeability in Candida albicans

Photodynamic antimicrobial chemotherapy (PACT) is a potential antimicrobial therapy that combines light and a photosensitizing drug, promoting a phototoxic effect on the treated cells, in general via oxidative damage. In this work we studied the effect of PACT, using methylene blue (MB), on the permeability of Candida albicans membrane. Our results demonstrated that the combination of MB and laser (684 nm) promoted a decrease in Candida growth. The inhibition was more pronounced in the presence of 0.05 mg/ml MB and with an energy density of 28 J/cm². The decrease in Candida growth was associated with an increase in membrane permeabilization. Thus, we suggest that a PACT mechanism using MB can be related to damage in the plasma membranes of the cells.     

Methylene blue mediated photobiomodulation on human osteoblast cells

Photobiomodulation (PBM) and photodynamic therapy (PDT) are two major methods, which use light in medicine and dentistry. PBM uses low-level laser light to induce cell proliferation and activity. In contrast, PDT use laser light combined with a photosensitizer (PS) to cause cell death. Due to similar, not fully understood mechanisms and biphasic response of light, unexpected and complex outcomes may be observed. In the present study, the effect of 635 nm laser light, with power density 50 mW/cm², at three different energy densities (0.5, 1, and 2 J/cm² which last 10, 20, and 40 s, respectively) mediated by methylene blue (MB) on the human osteoblast cell line (ATCC-CRL-11372, Rockville, MD, USA) was investigated. Cell viability (MTT assay and acridine orange/propidium iodide staining) and proliferation (Alamar Blue assay) were assessed at 24, 48, and 72 h post irradiation. Alkaline phosphatase (ALP) activity, mineralization (Alizarin Red staining) and gene expressions (RT-PCR analysis) were analyzed at 7th and 14th days after treatment. Five groups were formed as the control group (no MB, no irradiation), MB (only 0.05 μM MB), MB + 0.5 J/cm², MB + 1 J/cm², and MB + 2 J/cm². Cell viability was decreased at 72 h (ANOVA; p < 0.05) for MB + 0.5 J/cm², MB + 1 J/cm², and MB + 2 J/cm² groups. Although proliferation does not seem to be effected by MB-mediated laser application, osteo-anabolic activity is altered. ALP activity was significantly increased at day 7 (ANOVA; p < 0.05) for MB-combined laser groups; on the other hand, mineralization was significantly decreased (ANOVA; p < 0.05) in all treatment groups. Alkaline phosphatase and collagen-I expressions were upregulated in MB + 2 J/cm² group at 7th and 14th days, respectively. These results may contribute to the low-dose PDT researches and understanding PBM effects on osteoblast behavior but further studies are needed since inappropriate conditions may lead to undesirable results for both therapies.     

Photodynamic therapy for the treatment of advanced gastrointestinal tumours

In the study, 120 patients with advanced gastrointestinal tumours were treated by PDT; 5 mg/kg of HpD was intravenously given 48–72 h prior to PDT. The light source was an argon dye laser with an output beam of 630 nm. The irradiation time varied from 15–25 min with a power of 100–350 mW cm^–2. The entire tumour was irradiated with a light dose of 100–250 J cm^–2. Of the 120 patients, 20 had cancer of esophagus, 72 had cancer of the gastric cardia, 18 had cancer of the stomach and 10 had cancer of the rectum. Eighty-eight patients (73.3%) had a response to PDT. Twelve patients with CR were followed up for one to five years, two patients died during the two years after PDT.     

Development of an alternative light source to lasers for photodynamic therapy: 2. Comparative in vivo tumour response characteristics

The performance of a low cost, table-top/portable light source was tested against an argon ion pumped dye laser for in vivo photodynamic therapy (PDT). The prototype delivers up to 1 W via a 4 mm flexible lightguide within a 30 nm bandwidth centred at any wavelength from 300 nm to 1200 nm at fluence rates of up to 8 W cm^–2. An in situ bioassay using regrowth delay of tumour T50/80 was used to quantify the relative efficacy of the prototype with a laser. The tumours were sensitized with haematoporphyrin derivative (HpD) and externally irradiated. There was no significant difference in the response of the tumour to treatment between the two light sources (p = 0.69). Mean growth delays ranged from 2 days (light dose 10 J cm^–2) to 20 days (light dose 100 J cm^–2). The estimate for the difference in means (laser minus prototype growth delay) was only 0.66 days and was not statistically significant. This in vivo study demonstrates that the prototype is equivalent to a laser in PDT effect. The device has low capital/running cost, is simple to use and is one of the most powerful, spectrally efficient non-laser PDT sources available.     

Study of photodynamic therapy in the control of isolated microorganisms from infected wounds—an in vitro study

The effective treatment of infected wounds continues to be a serious challenge, mainly due to the rise of antibiotic-resistant bacteria. Photodynamic therapy (PDT) refers to the topical or systemic administration of a non-toxic, photosensitizing agent (PS), followed by irradiation with visible light of a suitable wavelength. The possibility of applying the PDT locally is what makes it so favorable to the treatment of infected wounds. The goal of this study was to evaluate the action of the PDT in the inactivation in vitro of microorganisms coming from infected wounds, using methylene blue (MB) and photodithazine (PDZ) as the PS and comparing the efficacy of these two compounds for PDT on bacteria. For the application of PDT, isolated microorganisms identified from material collected from wounds were suspended in a saline solution containing 10⁶ viable cells/ml. Each isolated microorganism was submitted to PDT with MB and with PDZ in accordance with the following treatment groups: N/T—no treatment; T1—PDT with PDZ; T2—PDT with MB; T3—irradiation without PS; T4—treatment with PDZ without light; and T5—treatment with MB without light. As a light source, an LED-based device was used (Biopdi/Irrad-Lead 660), composed of 54 LEDs, each with 70 mW of power in the 660 nm region of the electromagnetic spectrum. Each tray of 96 wells was irradiated with an intensity of 25 mW/cm² and a dose of light of 50 J/cm³ for 33 min. All the tests were made in duplicate. It was then concluded that the PDT with PDZ was capable of inhibiting the growth of gram-positive bacteria samples, however it did not have the same effect on gram-negative bacteria, which showed growth greater than 100,000 CFU; the PDT with MB showed an effectiveness on gram-positive as well as gram-negative bacteria, for it was able to inhibit bacterial growth in both cases.     

Fluorescence biodistribution and photosensitising activity of toluidine blue o on rat buccal mucosa

. The antimicrobial activity of toluidine blue O (TBO) in the presence of red light has been demonstrated for a wide range of microorganisms. The response of tissues to TBO-induced photosensitisation is an important factor in assessing the clinical usefulness of this technique for the treatment of infectious diseases. The aims of this study were to determine the effect of TBO-mediated photosensitisation on rat buccal mucosa and the biodistribution of the photosensitiser in this tissue. An aqueous solution of TBO was applied to one side of the buccal mucosa of the animals. A 6 mm diameter area was then exposed to light (633 nm) from a copper vapour pumped-dye laser. The opposite, untreated, side of the buccal mucosa served as a control. TBO concentrations of 25, 50 and 200 μg/ml, laser light doses of 110, 170 and 340 J/cm² were assessed. Control groups of animals were subjected to 340 J/cm² laser light alone or to 200 μg/ml TBO alone. Serial sacrifices were performed after 72 h to obtain mucosal tissue samples for histological examination. For the determination of TBO biodistribution, additional groups received the same TBO doses and were sacrificed after 1 min or 10 min. Specimens were removed and frozen immediately for digital fluorescence imaging. No necrotic or inflammatory changes were found in the buccal mucosa of the animals with any of the treatments (using up to 200 μg/ml TBO and 340 J/cm² laser light). A high TBO fluorescence in the epithelium, particularly in the keratinised layer, with almost no fluorescence in the underlying connective tissue was demonstrated by the digital imaging. The results of this study suggest that TBO-mediated PDT (within the concentrations and light doses tested) could be a safe antimicrobial approach for the oral infections without damaging the adjacent normal tissue. Paper received 2 May 2001; accepted 23 July 2001.     

Helium-neon laser: A contraindication for infected wounds? In vitro study on pseudomonas aeruginosa

The purpose of this investigation was to examine the susceptibility ofPseudomonas aeruginosa to helium-neon (He-Ne) laser in order to gain an insight into the clinical implications for this type of low-level laser therapy (LLLT) in the treatment of infected wounds. Suspensions, in the presence and the absence of methylene blue (MB, 0.001% w/v), were exposed to the light from a 9 mW He-Ne laser for 5 and 50 min. Exposure of samples without MB did not affect the viability of this organism based on the lack of significant differences in the number of colony forming units (CFU) between irradiated and control samples. In the presence of MB, statistically significant mean reductions in CFU of 39.5% (22.6 J cm^-2) and 58.2% (226 J cm^-2) were found, and growth-free zones in irradiated confluent platings could be observed. Since no logarithmic reductions were achieved, the clinical implementation of He-Ne laser as a bactericidal agent seems to be of no relevance yet. Otherwise, no stimulation of bacterial growth was observed. Therefore, infection as a contraindication for LLLT should be excluded.     

Photodynamic therapy: Optimal treatment parameters in murine fibrosarcoma

We have investigated the efficacy of photodynamic therapy (PDT) by using two preparations of haematoporphyrin derivative (HPD) that demonstrated different biological activities against experimental murine fibrosarcoma RIF-1 in C3H/He mice. We have been able to demonstrate a minimum clonogenic survival rate of 0.25% by using the more active HPD at 20 mg/kg with a 40-h retention time and a total laser light dose of 100 J/cm². Further, we noted that clonogenic survival rates of 21.6% and 25.0% respectively could be achieved by using the less active HPD (at 20 mg/kg) with a laser light dose of 150 J/cm², or the more active HPD (at 20 mg/kg) with a laser light dose of 20 J/cm². In both cases necrosis of the surrounding normal tissue was absent. Necrosis of the normal tissue surrounding the tumour was shown to be associated with high laser energy (100 J/cm² and higher) in conjunction with a high dose (20 mg/kg) of the more active HPD. A comparison of survival curves as a function of laser energy for the RIF-1 cells following PDT with the two different preparations of HPD showed a difference in the kinetics of cell death. A curve with a shoulder region and aD _o (mean lethal dose) of 41 J/cm² was obtained when the less active HPD was used, whereas PDT using the more active HPD resulted in a curve with no shoulder and aD _o of 16 J/cm².     

Photochemotherapy of experimental colonic tumours with intra-tumorally applied methylene blue

Introduction: Phototoxicity of intra-tumoral injected methylene blue (MB⁺) was studied in 48 experimental colonic tumours in comparison with photosan-3, Zn-phthalocyanine and tetrasulphanated ClAl-phthalocyanine. Methods: In mice, xenotransplanted subcutaneous tumours about 1 cm in diameter were treated photodynamically twice, with different sensitisers. The irradiation was performed at the sensitiser-specific wavelength, and a densitiy of 100 mW/cm² and a dose of 100 J/cm². Results: Light alone without sensitiser did not induce any effect in mice tumours. Surprisingly, Al-phthalocyanine could only be used for intratumoral injections because of toxic effects after intravenous applications in nude mice. Using MB⁺ (1%), 75% of the tumours were destroyed by a single photodynamic treatment (PDT). In addition, toxicity of MB⁺ was most intense when compared with Zn-phthalocyanine and photosan-3. However, after the second PDT, there was no statistically significant difference among these sensitisers. Dark toxicity of MB⁺ (1%) could be well demonstrated by sufficient sensitiser incorporation without irradiation, which led to a stationary tumour volume up to 3 weeks after injection. Conclusion: Intra-tumoral MB⁺ PDT is a potential treatment for inducing necrosis in vivo. With regard to tumour tissue, the selectivity of MB⁺ is high and depends on a precise local injection of the dye. Received: 25 May 1998     

Dosedependent photodynamic effects of 9-acetoxy-2,7,12,17-tetrakis(\-methoxyethyl)-porphycene in vitro

Porphycenes are chemically pure photosensitizers for topical and systemic photodynamic therapy (PDT). Fast cellular uptake of 9-acetoxy-2,7,12,17-tetrakis-(sB-methoxyethyl)porphycene (ATMPn) has been shown previously. HaCaT human keratinocytes were incubated with ATMPn (1 nmol l^-1 to 1 μmol l^-1 in DMSO or DOPC liposomes). After 1 h, cells were irradiated with different light doses (0, 24, 48J cm^-2) using an incoherent light source (580—740 nm, 40 mW cm^-2). Cytotoxic effects were determined by assessing the mitochondrial activity using the MTT assay 24 h following irradiation. Cytotoxic effects were dependent on ATMPn concentration and light dose. Using 20 nmol 1^-1, a 50% decrease of mitochondrial activity (EC⁵⁰) after irradiation with 24 J cm^-2 was achieved. Lowering the ATMPn concentration (10nmol 1^-1) and increasing the light dose (48 J cm^-2) yielded the same effect (EC⁵⁰). Maximal decrease of mitochondrial activity (90%) was achieved using ATMPn concentrations of 50–100 nmol l^-1 and a light dose of 24 J cm^-2 or 25 nmol l^-1 ATMPn and 48 Jcm^-2. There was no difference regarding the dose-dependent cytotoxic effects using either ATMPn in DMSO or DOPC liposomes. In the control group (incubation with 1 nmol 1^-1 to 1μmol 1^-1 ATMPn, no irradiation), dark toxicity was not observed. Cell photosensitization with ATMPn was very efficient in vitro yielding the maximal cytotoxic effect at very low ATMPn concentrations as compared to other photosensitizers. Since ATMPn in DMSO and DOPC liposomes revealed the same cytotoxic effects without dark toxicity, theDMSO formulation, which is much easier to prepare, will be preferred in future studies.     

Toluidine blue-mediated photodynamic therapy of oral wound infections in rats

The purpose of this study was to examine the effect of toluidine blue (TB)-mediated photodynamic therapy (PDT) on oral wound infections in rats. The study called for a combination treatment of a 1mg/ml solution of TB with a red light at three intensity settings of 12 J/cm², 24 J/cm² and 48 J/cm². In the group that was given the highest light dose of 48 J/cm², an average kill rate of approximately 97% was achieved. A lesser killing effect was achieved in the group that was subjected to the lowest light dose of 12 J/cm², where an average of approximately 25% of the bacteria survived. After PDT, the lesions were allowed to develop, and the peak size of the lesions was larger in the control group than in the test groups, especially for the 48 J/cm² group. We also observed that in the 24 J/cm² and 48 J/cm² groups the lesions were of significantly smaller size. Our study demonstrated that combined TB-PDT therapy can successfully treat oral wound infections in rats. These promising results recommend the use of this treatment as a possible alternative to topical anti-microbials in future clinical applications.     

Antimicrobial photodynamic therapy—a promising treatment for prosthetic joint infections

Periprosthetic joint infection (PJI) is associated with high patient morbidity and a large financial cost. This study investigated Photodynamic Therapy (PDT) as a means of eradicating bacteria that cause PJI, using a laser with a 665-nm wavelength and methylene blue (MB) as the photosensitizer. The effectiveness of MB concentration on the growth inhibition of methicillin-sensitive Staphylococcus aureus (MSSA), methicillin-resistant Staphylococcus aureus (MRSA), Staphylococcus epidermidis, Pseudomonas aeruginosa and Acinetobacter baumannii was investigated. The effect of laser dose was also investigated and the optimized PDT method was used to investigate its bactericidal effect on species within planktonic culture and following the formation of a biofilm on polished titanium and hydroxyapatite coated titanium discs. Results showed that Staphylococci were eradicated at the lowest concentration of 0.1 mM methylene blue (MB). With P. aeruginosa and A. baumannii, increasing the MB concentration improved the bactericidal effect. When the laser dose was increased, results showed that the higher the power of the laser the more bacteria were eradicated with a laser power?≥?35 J/cm² and an irradiance of 35 mW/cm², eradicating all S. epidermidis. The optimized PDT method had a significant bactericidal effect against planktonic MRSA and S. epidermidis compared to MB alone, laser alone, or control (no treatment). When biofilms were formed, PDT treatment had a significantly higher bactericidal effect than MB alone and laser alone for all species of bacteria investigated on the polished disc surfaces. P. aeruginosa grown in a biofilm was shown to be less sensitive to PDT when compared to Staphylococci, and a HA-coated surface reduced the effectiveness of PDT. This study demonstrated that PDT is effective for killing bacteria that cause PJI.     

Effect of photodynamic therapy on urinary bladder function: an experimental study with rats

Photodynamic therapy (PDT) produces localized necrosis with light after prior administration of a photosensitizing drug. The problems with laser light dosimetry and complications relating to bladder function appear to be important limiting factors of PDT in urology. Photodynamic therapy on urinary bladder with normal epithelium of rats was performed using an argon ion laser as an energy source, with aminolevulinic acid (ALA)-induced protoporphyrin IX (PpIX) photosensitizer. Four hours after ALA intravenous administration, the bladders were intravesically radiated with light doses 20, 40, or 80 J/cm². Animals in the control group did not receive ALA and were radiated with 20 J/cm² light dose. Three weeks prior to PDT, the bladder capacity and pressure changes during filling cystometry were assessed. Cystometrics were repeated 1, 3, 7, or 21 days after laser therapy. The light dose 20 J/cm² and 40 J/cm² together with the used ALA dose caused no reduction in bladder capacity, whereas 80 J/cm² light dose produced up to 50% reduction in the capacity at 3 weeks postoperatively. In control group without ALA, the animals did not regain more than 34% of the capacity of their control values at 3 weeks. The light dose of 20 J/cm² and 40 J/cm² with ALA induced functional changes that subsided after day 1. Our results indicate that with proper dosing of photosensitizing drug and light energy, the functional impairment of urinary bladder may be reduced as transient. These findings support the use of PDT as safe therapy of superficial bladder cancer. Received: 10 April 2000 / Accepted: 16 November 2000     

Wavelength-dependent effect of tetra(m-hydroxyphenyl)chlorin for photodynamic therapy in an ‘early’ squamous cell carcinoma model

The purpose of the present study was to correlate the wavelength of the irradiation source with the phototoxic activity of tetra(m-hydroxyphenyl)chlorin (mTHPC) in healthy and neoplastic mucosae. The hamster tumour model for early squamous cell carcinoma was used in these experiments. In vitro and in vivo studies have shown that mTHPC absorbs significantly at 652 nm (1, 2). This wavelength is used currently in clinical mTHPC photodynamic therapy (PDT) trials. In order to study the wavelength dependence of the phototoxic effect on normal and tumour tissues, irradiation tests were performed 4 days after injection of 0.5mg kg^-1 mTHPC. An argon-ion pumped dye laser was used as the light source. The light dose of 12 J cm^-2 was delivered at a light dose rate of 150 mW cm^-2. The wavelength was varied between 642.5 and 665 nm at 2.5-nm increments. The PDT damage was evaluated in serial Haematoxylin and Eosin stained sections using a tissue-damage scale. Light between 647.5 and 652.5 nm induced the highest damage to both the healthy and tumour mucosae. At wavelengths equal to or below 645 nm, and equal to or above 655 nm, tissue damage decreased. Wavelengths below 642 nm and above 660 nm did not induce any visible tissue damage. These results suggest that the in vivo optimal wavelength range for PDT with mTHPC is between 647 and 652 nm. This information is essential for selecting an appropriate light source.     

Photodynamic therapy inhibits transforming growth factor β activity associated with vascular smooth muscle cell injury

Purpose: The multifunctional cytokine, transforming growth factor β1 (TGF-β), plays an important role in the development of injury-associated intimal hyperplasia (IH). Strategies to suppress local TGF-β activity may have a clinical potential to prevent restenosis caused by IH. Photodynamic therapy (PDT) involves the local generation of cytotoxic free radicals by light activation of photosensitizer dyes and has been shown to inhibit experimental IH. This study investigated whether PDT-generated free radicals can affect TGF-β activity in a biologic system using vascular smooth muscle cells (SMCs).Methods: The release and activation of TGF-β by injured SMCs in culture was compared between mechanical injury and PDT. Mechanical injury was induced with a rubber policeman, and PDT was performed with the photosensitizer chloroaluminum sulfonated phthalocyanine (5 μg/ml) and 675 nm laser light at subtherapeutic 10 J/cm² and the in vivo therapeutic dose of 100 J/cm². Cell viability was assessed by the tetrazolium salt conversion assay, and active and total (active + latent) TGF-β was determined by enzyme-linked immunosorbent assay in the conditioned media of SMCs 24 hours after treatment. Functional TGF-β activity was assessed by inhibition of endothelial cell mitogenesis.Results: Both forms of injury severely reduced (p < 0.0005) SMC viability to less than 15%. In untreated SMC conditioned media, only 14.5% of the total TGF-β was active (27.7 ± 8.7 pg per 1 × 10⁵ cells). However, after mechanical injury and PDT with 10 J/cm², there was a significant increase (p < 0.02) in active TGF-β (60.1 ± 10.1 pg and 48.6 ± 21.0 pg, respectively), despite a total reduction of approximately 50%. In contrast to this result, PDT with 100 J/cm² did not result in increased levels of active TGF-β (8.1 ± 3.5 pg), despite having similar levels of total TGF-β. Consequently, the conditioned media of SMCs that had 100 J/cm² PDT did not inhibit endothelial cell mitogenesis as compared with the conditioned media of SMCs with mechanical injury and 10 J/cm² PDT (p < 0.0002).Conclusions: This report describes two novel findings: (1) injury to SMCs in vitro induces the conversion of biologically latent TGF-β to active TGF-β; and (2) the therapeutic PDT dose interferes with this injury activation process. This study substantiates the concept of local cytokine inhibition by PDT in a biologic system and provides new insights into the mechanisms of PDT-mediated inhibition of experimental IH. (J Vasc Surg 1997;25:1033-43.)     

Synovial ablation in a rabbit rheumatoid arthritis model using photodynamic therapy

Background: At present there is no ideal minimally invasive method for ablating inflamed synovium in joints that has been unres­ponsive to optimal medical management in patients with rheumatoid arthritis. The aim of this study was to determine whether photo­dynamic therapy could be used for this purpose. Methods: In a rabbit knee model of rheumatoid arthritis the pharmacokinetics of the photosensitizer Haematoporphyrin Derivative (HpD) into periarticular tissues and blood was measured following intravenous injection of HpD. The second phase of the study was to determine the histological effect of HpD activation by 63 nm light delivered via an intra‐articular optic fibre using a dye pumped KTP‐YAG laser. The light dose was varied from 0?200 joule/cm². Results: Pharmacokinetic studies determined that inflamed synovium rapidly accumulated HpD, with peak levels being reached 12 h following intravenous injection. The ratio of HpD uptake into inflamed synovium versus peri‐articular quadriceps muscle was found to be 22.8. Histological examination of the treated knees indicated that selective destruction of inflamed synovium was achieved at light doses 100 joules/cm² and above. No significant effect was observed on normal intra‐articular tissues. Conclusion: We have demonstrated that the first generation photosensitizer HpD selectively accumulates within inflamed ­synovium. Activation of HpD by intra‐articular light administration resulted in selective ablation of the inflamed synovium. These findings indicate that PDT offers potential as a new selective, minimally invasive synovectomy technique.     

Comparison of the photodynamic fungicidal efficacy of methylene blue, toluidine blue, malachite green and low-power laser irradiation alone against Candida albicans

This study was to evaluate specific effects of photodynamic therapy (energy density 15.8 J/cm², 26.3 J/cm² and 39.5 J/cm²) using methylene blue, toluidine blue and malachite green as photosensitizers and low-power laser irradiation on the viability of Candida albicans. Suspensions of C. albicans containing 10⁶ cells/ml were standardized in a spectrophotometer. For each dye, 120 assays, divided into four groups according to the following experimental conditions, were carried out: laser irradiation in the presence of the photosensitizer; laser irradiation only; treatment with the photosensitizer only; no exposure to laser light or photosensitizer. Next, serial dilutions were prepared and seeded onto Sabouraud dextrose agar for the determination of the number of colony-forming units per milliliter (CFU/ml). The results were subjected to analysis of variance and the Tukey test (P < 0.05). Photodynamic therapy using the photosensitizers tested was effective in reducing the number of C. albicans.. The number of CFU/ml was reduced by between 0.54 log₁₀ and 3.07 log₁₀ and depended on the laser energy density used. Toluidine blue, methylene blue and malachite green were effective photosensitizers in antimicrobial photodynamic therapy against C. albicans, as was low-power laser irradiation alone.     

Toluidine blue-mediated photoinactivation of periodontal pathogens from supragingival plaques

This study aimed to assess the effect of toluidine blue (TB)-mediated photodynamic inactivation of periodontal pathogens (PP) from periodontopathic patients. Photodynamic therapy (PDT) was carried out using TB and 635 nm laser light irradiation. The bactericidal effect was evaluated, and important PDT parameters including light intensity, energy dose, and TB concentration were determined. Our findings suggest that TB-mediated lethal photosensitization of PP in vivo is possible. However, to obtain ideal bactericidal effect, higher doses of light and photosensitizer should be required in treatment in vivo than their planktonic counterparts. The best therapeutic effect was observed in treatment by 1 mg/ml TB combined with 12 J/cm² at 159 mW/cm² light irradiation. Moreover, because of the considerable interindividual differences of bacterial populations, TB-mediated PDT might not be equally effective among periodontopathic patients, and further studies on improvement of this therapeutic modality is needed.     

Photodynamic therapy to the oral cavity,tongue and larynx: a canine normal tissue tolerance study

Photodynamic therapy (PDT) has the potential to treat early carcinomas of the oral cavity and larynx while preserving normal tissue. However, normal tissues retain the photosensitizing agents and may be activated by high light fluence and dose rates resulting in normal tissue necrosis. The effects of varying dose rates of light delivery on various tissues in the upper aerodigestive tract have not been evaluated to date and are necessary to determine a therapeutic light dose range that will result in selective tumour necrosis. Thirty adult mongrel dogs received intravenous Photofrin, 2 mg kg^–1, 48 h prior to PDT treatment. Photodynamic therapy was administered to the tongue, buccal mucosa and larynx with a microlens fibre and implantable cylindrical diffuser at various dose rates from 20 to 125 J cm^–2 at 150 mW cm^–2. At the same dose rate of light delivery, the tongue was the most sensitive organ, followed by the buccal mucosa, and last by the larynx. The differential tissue effect of identical dose rates of therapy must be taken into account when administering PDT so that selective tumour necrosis with normal tissue preservation may be achieved. This study indicates the need to perform evaluations of the effect of PDT on other tissue types in an animal model with each new photosensitizer prior to administering PDT to those areas in humans.     

Interstitial photodynamic therapy in the Dunning R3327-AT6 prostatic carcinoma

Interstitial photodynamic therapy (PDT) could be an alternative radical treatment for prostate cancer. The ability to predict the depth of necrosis is necessary for light treatment planning using multiple optical fibres. The extent of PDT necrosis was studied in subcutaneously implanted R3327-AT6 Dunning prostate tumours which had similar optical characteristics to human prostate. Tumour-bearing subjects were given 20 mg kg^–1 Haematoporphyrin esters (HPE) and irradiated 24 h later with 630 nm laser light. Five subjects per group were treated with increasing light doses (50–450 J cm^–1) delivered interstitially via a single 2 cm long cylindrical diffuser. After 450 J cm^–1 of irradiation, 4.3±0.8 cm³ [standard error of the mean (s.e.m.)] of tumour tissue was necrosed to a depth of 10.5±0.8 mm around the diffuser. There was an approximately linear correlation between the volume of PDT necrosis around the fibre and prescribed light dose. The mean threshold light dose for PDT effect was 18±2 J cm^–2. In this tumour with a mean photosensitizer concentration of 16±1.5g g^–1, low light doses produced tumour necrosis. PDT using multiple diffusers could destroy a relatively large tumour volume and the diffusion theory model reliably predicted the depth of necrosis.