Laser

Title

Adjunctive effects of a low-power laser on the healing of periodontal tissue

Author(s)

Lai, Man-lung, Stanley.; .

Citation

Issue Date

2003

URL

http://hdl.handle.net/10722/50883

Rights

The author retains all proprietary rights, (such as patent rights) and the right to use in future works.

ADJUNCTIVE EFFECTS OF A LOW-POWER LASER ON THE HEALING OF PERIODONTAL TISSUE

This dissertation is submitted to The University of Hong Kong in partial fulfillment of the requirements for the degree of

Master of Dental Surgery (Periodontology) by Lai Man Lung, Stanley 2003

Periodontology Faculty of Dentistry The University of Hong Kong Hong Kong

Adjunctive Effects of a Low-power Laser on the Healing of Periodontal Tissue

Lai Man Lung, Stanley 2003

Abstract

Abstract

The aim of this study was to evaluate the adjunctive effect of a low-power laser in non-surgical periodontal treatment of patients with moderate to advanced chronic periodontitis. The study was performed as an intra-individual, longitudinal trial of 9 months duration with a single-blinded, controlled and randomized design. A total of 14 patients, 13 females and 1 male, aged from 33 to 57 years (mean = 43.6) having at least two sites per quadrant with probing pocket depth (PPD) 5mm were recruited. Matched sites with PPD of 5mm or above and comparable radiographically detected bone defect were selected on each side of the mouth in each patient. Clinical parameters, including presence of supragingival plaque (PI%), bleeding on probing (BOP%), probing pocket depth (PPD) and probing attachment level (PAL) were recorded at baseline, 3 months, 6 months and 9 months after treatment. Gingival crevicular fluid (GCF) samples and standardized intra-oral radiographs for evaluation of alveolar bone changes by digital subtraction radiography were also taken at the baseline and after 1 month, 3 months, 6 months and 9 months. After initial examination, full-mouth supra- and sub-gingival debridement was performed and oral hygiene instructions were given. Test sites were selected randomly by a coin toss and irradiated by a low-power laser (He-Ne) operating at a wavelength of 632nm and an output power of 0.2mW while control sites received no additional treatment. Laser irradiation was applied directly to the buccal or palatal aspects of the selected sites, each time for ten minutes for a total of eight times in the first 3-month period by a trained operator. The whole-mouth PI% decreased from 83% at baseline to 15% at 9 months while the BOP% decreased

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Abstract

from 95% to 28%. Statistical significant changes in reductions of PPD and GCF volume, gain in PAL and increase in recession (REC) were seen in both test and control groups of sites at 3-, 6- and 9-month examination when compared to baseline (p< 0.05). The overall mean PPD reductions were 2.40.2mm, 2.50.2mm and 2.70.3mm for test sites and 2.30.3mm, 2.40.3mm and 2.80.4mm for control sites at 3-month, 6-month and 9-month examinations, respectively. The mean PAL gains were 2.00.3mm, 2.20.3mm and 1.60.4mm for test sites and 2.30.3mm, 2.30.3mm and 1.90.5mm for control sites at 3-month, 6-month and 9-month examinations, respectively. No significant differences in PPD reduction, PAL gain and REC change were revealed between test and control sites, except the GCF volume after 3 months which reduced from 69 to 12 in test sites and 66 to 18 in control sites and this difference was found to be statistically significant (p=0.02). Radiographic analysis by computer-assisted densitometric image analysis (CADIA) showed statistical significant increase in test sites (p=0.035) but not in control sites from baseline to 9-month. However, no statistical significant difference was found between test and control sites at any time point.

The findings from the present study suggest that the adjunctive use of low-power laser in patients with moderate to severe chronic periodontitis in non-surgical periodontal therapy did not provide additional clinical benefit.

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Acknowledgements

Acknowledgements

I would like to thank my supervisor, Dr. K. Y. Zee, for his guidance, critical appraisal, intellectual input and great effort throughout the study Thanks to Dr. Esmonde Corbet for his valuable suggestions and final revision of the script. Thanks to my brother, Mr. Simon Lai for his advice in statistical analysis and help in computer knowledge. In particular, I would like to thank Ms. Grace Wong, my dental surgery assistant, for her efficient support in all the clinical work. I would also like to express my sincere thankfulness to my parents, my girlfriend, Winnie, for their continuous spiritual support throughout the years. Last but not the least, thanks to my Lord, Jesus Christ, for His companionship and support throughout the years.

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Contents

ContentsAbstract........................................................................................................................i Acknowledgements................................................................................................... iii Contents .....................................................................................................................iv Part I............................................................................................................................1 Literature Review .......................................................................................................1 1. Literature Review ...................................................................................................1 1.1 Introduction.......................................................................................................1 1.2 Non-surgical periodontal therapy in treating periodontal disease ....................2 1.2.1 Clinical changes after therapy....................................................................2 1.2.2 Healing after treatment ..............................................................................4 1.2.3 Limitations with non-surgical mechanical periodontal therapy ................5 1.2.4 Surgical approach ......................................................................................6 1.3 Non-surgical mechanical periodontal therapy with adjunctive treatment ........7 1.4 Properties of laser .............................................................................................9 1.5 Types of laser..................................................................................................10 1.5.1 Surgical or hard laser ...............................................................................10 1.5.2 Low-power or soft laser...........................................................................11 1.6 Laser safety .....................................................................................................12 1.7 History of low-power laser (therapeutic laser) therapy ..................................12 1.8 Laboratory studies of low-power laser ...........................................................13 1.8.1 Fibrobalsts................................................................................................13 1.8.2 Osteoblasts ...............................................................................................14 1.8.3 Keratinocytes ...........................................................................................15 1.8.4 Bacteria ....................................................................................................15 1.9 In vivo studies of low-power laser ..................................................................16 1.9.1 Animal studies .........................................................................................16 1.9.2 Clinical studies in human.........................................................................18 1.10 Conclusion ....................................................................................................20 Part II ........................................................................................................................21 Materials and Methods..............................................................................................21 2. Materials and Methods..........................................................................................21 2.1 Aim of study ...................................................................................................21 2.2 Objectives .......................................................................................................21 2.3 Study Population.............................................................................................22 2.3.1 Subjects selection.....................................................................................22 2.3.2 Teeth/sites selection for laser irradiation .................................................23 2.4 Study Design...................................................................................................23 2.5 Ethical aspect ..................................................................................................24 2.6 Clinical protocol .............................................................................................24 2.6.1 Baseline examination...............................................................................24 2.6.2 Collection of GCF....................................................................................25 2.6.3 Alveolar bone change observed by Digital Subtraction Radiography (DSR) ................................................................................................................26 2.6.4 Non-surgical periodontal treatment .........................................................27 2.6.5 Laser irradiation.......................................................................................27 2.6.6 Recall appointment ..................................................................................28

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2.7 Analysis of alveolar bone changes..................................................................28 2.8 Calibration ......................................................................................................29 2.9. Data Analysis.................................................................................................30 Part III .......................................................................................................................31 Results.......................................................................................................................31 3. Results...................................................................................................................31 3.1 Subject population ..........................................................................................31 3.2 Sites selection .................................................................................................31 3.3 Full mouth clinical data ..................................................................................32 3.4 Comparison between test and control.............................................................35 3.4.1 Clinical parameters ..................................................................................35 3.4.2 CADIA value as obtained by DSR ..........................................................36 Part IV.......................................................................................................................39 Discussion & Conclusions ........................................................................................39 4. Discussion.............................................................................................................39 4.1 Effects of non-surgical periodontal therapy....................................................39 4.2 Adjunctive effects of low-power laser............................................................40 4.2.1 Clinical parameters ..................................................................................40 4.2.2 Alveolar bone changes.............................................................................42 4.3 Methodology and design.................................................................................46 4.3.1 Use of manual periodontal probing .........................................................46 4.3.2 Design of low-power laser research.........................................................47 4.3.3 Radiographic analysis ..............................................................................49 4.4 Conclusion ......................................................................................................52 Part V ........................................................................................................................53 References.................................................................................................................53 Part VI.......................................................................................................................67 Appendix...................................................................................................................67 Appendix IA. Explanatory notes for patients (English Version).........................67 Appendix IB. Explanatory notes for patients (Chinese Version) ........................69 Appendix II. Informed consent ..........................................................................70

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Literature Review

Part I Literature Review

Literature Review

1. Literature Review1.1 Introduction

Periodontal disease is an oral infectious disease involving inflammatory reactions which causes tissue destructions in tooth supporting structures, i.e., gingiva, cementum, periodontal ligament and alveolar bone (Lang & Brgger 1991). Based on the data from the early study of experimental gingivitis (Le et al. 1965) in the late 1960s, dental plaque is confirmed to be the aetiological factor triggering the inflammatory process in the soft tissues around the tooth. From some of the epidemiological studies, gingivitis was always found to precede periodontitis (Le et al. 1986). Since then, different contributing factors have been identified in the past forty years. Some of the researchers focused on the bacteria in dental plaque and believed that to be the primary cause of periodontal disease (Haffajee & Socransky 1994). They also suggested certain putative periodontal pathogens would cause the tissue destruction. Although there is quite strong evidence suggesting the association of specific microflora with periodontal destructions, it still remains arguable on whether specific microorganisms are the main cause of the disease. Other than the bacteria from dental plaque biofilms, pathogenesis of periodontal disease was believed to be caused by immune reactions of our body to plaque microbial challenge. Therefore, host responses were suggested to play an important role as an indirect mechanism (Page et al. 1997). Epidemiological studies(Albandar 1990, Le et al. 1986) also revealed that only small percentage of the population suffered from advanced periodontal destructions and accounted for most of the periodontally involved sites. These findings supported that host responses in an

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individual would result in different degree of destructions and further suggested that genetic and environmental factors contribute to the establishment of the disease (Hart & Kornman 1997, Kornman et al. 1997).

1.2 Non-surgical periodontal therapy in treating periodontal disease1.2.1 Clinical changes after therapy

Periodontal disease usually presents clinically as pocket formation and/or gingival recession as loss of attachment (Flemmig 1999) and it is agreed that chronic periodontitis is recognized as the most frequently occurring form. The disease can start at any age, but is most commonly found in adults. Although there is increasing knowledge in the understanding of the pathogenesis of periodontal disease, the sequences of the treatment in general still begin with the establishment of good plaque control, followed by a thorough professional supra- and sub-gingival debridement which usually termed as non-surgical periodontal therapy (Cobb 1996).

In the practice of dentistry nowadays, the removal of supra- and sub-gingival deposits is still the main and important part of periodontal therapy. Numerous studies have reported the successful outcome in treating periodontal disease by nonsurgical periodontal therapy (Badersten et al. 1981, Badersten et al. 1984a, Cercek et al. 1983, Claffey et al. 1988, Loos et al. 1989, Nordland et al. 1987, Westfelt et al. 1998). Clinical assessments after the therapy often include probing attachment level (PAL), probing pocket depth (PPD), gingival recession (REC) and bleeding on probing (BOP) as common parameters. All the literatures reported that there would

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Literature Review

be reduction of PPD following the therapy which results from a combination of gain in clinical attachment level and increase in gingival recession (Hughes & Caffesse 1978, Proye et al. 1982).

The effects of non-surgical periodontal therapy on PAL were related to initial PPD as documented by various clinical studies (Badersten et al. 1984a, Cercek et al. 1983, Claffey et al. 1988, Loos et al. 1988a, Nordland et al. 1987, Westfelt et al. 1998). In general, the mean reduction in PPD with an initial depth of 13mm has been reported to be 0.03mm with a net loss of 0.34mm in PAL. For those pockets initially measuring 46mm, the mean reduction in PPD was 1.29mm with a net gain in PAL of 0.55mm. Periodontal pockets of 7.0mm or more showed a mean reduction of 2.16mm and a gain in PAL of 1.19mm (Cobb 2002). The greatest change in PPD and gain or loss in PAL could be usually seen within 1-3 months post-treatment while the final healing and tissue remodelling would occur in 9-12 months afterwards (Badersten et al. 1981, Badersten et al. 1984a). Together with the changes of these two clinical parameters, the resolution of inflammation could also be observed as reflected by the reduction in percentage of BOP.

In summary, the favourable clinical results obtained by non-surgical periodontal therapy suggest there is no definite limitation in treating periodontal disease by such therapy with various initial PPD. In addition, most of the stable periodontal condition can be maintained after non-surgical periodontal therapy if

comprehensive supportive therapy is provided (Axelsson & Lindhe 1981).

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1.2.2 Healing after treatment

Beside clinical parameters, successful periodontal therapy will result in formation of a long junctional epithelial attachment covering the root surfaces but not the formation of new connective tissue attachment as observed in histological examination in human studies (Waerhaug 1978a, Waerhaug 1978b). These histological features can only be considered as repair but not regeneration. In an article by Bartold et al. (2000), four features should be observed before the healing could be confirmed as regeneration: Functional epithelial seal must be re-established at the most coronal portion of the tissues and be no more than 2mm in length. New connective tissue fibers (Sharpeys fibers) must be inserted into the previously exposed root surface to reproduce both the periodontal ligament and the dentogingival fiber complex. New acellular, extrinsic fiber cementum must be reformed on the previously exposed root surface. Alveolar bone height must be restored to within 2mm of the cementoenamel junction.

Since repair but not regeneration was observed in the healing mechanism, researchers have focused in achieving regeneration by various kinds of method in the past thirty years. Techniques such as guided tissue regeneration (Weltman et al. 1997), and using different growth factors (Heijl et al. 1997) and bone grafting materials (Bowen et al. 1989) have been developed and evaluated. However, not a single technique could produce predictable regeneration in all cases. Even though

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regeneration can be achieved in some cases, the amount is limited and the cost is very high.

1.2.3 Limitations with non-surgical mechanical periodontal therapy

Access to deep pockets using conventional instruments in non-surgical periodontal therapy is usually restricted. Waerhaug (1978a, 1978b) and Stambaugh (1981) noted that the chance of removing all sub-gingival plaque from tooth surfaces was fairly good if the probing depth was 3.0mm. At probing depths of 35mm, the chance of failure becomes greater. At probing depths 5.0mm, the chance of failure becomes significantly higher. In fact, Stambaugh (1981) also showed that removal of all subgingival plaque and calculus was not possible when mean PPD were 3.73mm. Dragoo (1992) reported that conventional instruments and ultrasonic tips could rarely approach the most apical region of the pocket. Besides that, numerous studies have also noted that removal of all calculus from all root surfaces was rarely achieved in vivo (Buchanan & Robertson 1987, Caffesse et al. 1986, Fleischer et al. 1989, Sherman et al. 1990a, Sherman et al. 1990b).

The limitation of non-surgical periodontal therapy in molar teeth could be illustrated more clearly in some studies (Kalkwarf et al. 1988, Loos et al. 1988b, Nordland et al. 1987). In these studies, they reported that deep molar furcation sites showed impaired healing response as compared to non-molar and molar flat surfaces. In addition, they also showed a higher frequency of continuous PAL loss. These differences were more noticeable for sites initially 7mm deep. This might be related to the fact that deep pockets on molar teeth often show furcation 5

Literature Review

involvement and hence, difficult to achieve adequate debridement. The contributing factors such as narrow furcation entrance diameter, long root trunk length, bifurcation ridges, concavities, cemental enamel projections and enamel pearls may act as predisposing factors and often increase the difficulties in mechanical cleaning (Al Shammari et al. 2001).

Long term prospective and retrospective studies have shown that molars with furcation involvement generally have a compromised prognosis (Becker et al. 1984, Goldman et al. 1986, Hirschfeld & Wasserman 1978, Ramfjord et al. 1987). Furcation areas present some of the greatest challenges to the success of periodontal therapy and reduce efficacy of this therapy, regardless of the treatment modalities employed. Ramfjord et al. (1987) reported that 16 of the 17 teeth with furcation involvement initially were lost during the maintenance phase in a longitudinal study.

1.2.4 Surgical approach

The usual alternative treatment for non-responding periodontal sites to non-surgical periodontal therapy would be access using surgical techniques. Numerous studies have shown that the mechanical therapy with open flap procedures can improve the access and result in considerably better healing (Becker et al. 1988, Isidor & Karring 1986, Kaldahl et al. 1996a, Kaldahl et al. 1996b, Knowles et al. 1979, Lindhe & Nyman 1985, Pihlstrom et al. 1983, Pihlstrom et al. 1984, Ramfjord et al. 1987, Westfelt et al. 1985). Recently, Serino et al. (2001) reported the initial outcome and long term effect of surgical and non-surgical mechanical treatment of advanced periodontal disease after 12 years. They showed that surgical therapy was 6

Literature Review

more effective than non-surgical therapy in reducing the overall mean PPD and in eliminating deep pockets. In addition, more subjects treated non-surgically exhibited signs of advanced disease progression in the 13 year period following active therapy than surgically-treated subjects. They concluded that in subjects with advanced periodontal disease, surgical therapy could provide better short- and longterm PPD reduction and lead to fewer subjects requiring additional adjunctive therapy.

1.3 Non-surgical mechanical periodontal therapy with adjunctive treatment

As discussed so far, it can be seen that there are some inherent limitations and shortcomings of non-surgical periodontal therapy in treating severe periodontal disease. Although surgical periodontal therapy in some way proved to be more effective in root debridement in periodontally involved sites, it should only be considered selectively in sites that are not responding well to non-surgical periodontal therapy (Sigurdsson et al. 1994). The success of non-surgical periodontal therapy in clinical studies still supports it to be the first phase and essential part of periodontal therapy. Therefore, in order to improve the success of non-surgical mechanical periodontal therapy, attempts are made to incorporate various adjunctive therapies.

One of the major adjunctive agents that has been investigated was using antimicrobial either locally as controlled release agents (Goodson et al. 1991,

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Literature Review

Jeffcoat et al. 1998, Wennstrm et al. 2001, Williams et al. 2001) or systemically (Berglundh et al. 1998, Winkel et al. 2001). Besides that, subgingival irrigations with antiseptic agents were also performed in clinical trial (Rosling et al. 2001).

Recently, host modulating drugs are being used as an adjunct in non-surgical periodontal therapy after the approval of the drug by Food and Drug Association in United States. In fact, the concept of host modulation is not new as it was already initiated in some studies in 1970s (Williams 1999). The drug recently used is a systemically administrated sub-antimicrobial dose of doxycycline which targets at preventing tissue breakdown by blocking bacterial and host-derived enzymes that are associated with loss of alveolar bone and connective tissues. More gain in PAL, more reduction in PPD and less percentage of having sites with 2m loss during the observation period was found in the test group when compared to placebo (Caton et al. 2000).

Although numerous studies have shown that the use of different adjunctive agents could provide additional benefits in clinical parameters, such benefits were quite minimal (not more than 1mm differences in mean reduction of PPD and PAL). Most of the adjunctive agents used with non-surgical periodontal therapy are antimicrobial in nature (often antibiotics). As there are also increasing concerns on the abuse of antibiotics together with their limited benefits during periodontal treatment, new adjunctive agents or therapies which are not antimicrobial should be sought.

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Literature Review

1.4 Properties of laser

Laser is one of the latest and advanced treatment modalities in dentistry. According to the European Standard IEC 601, the definition of laser is Any device which can be made to produce or amplify electromagnetic radiation in the wavelength range from 180nm to 1mm primarily by the process of controlled stimulated emission. The word LASER is an acronym of Light Amplification by Stimulated Emission of Radiation. Based on the famous scientist Albert Einstein in his theory Zur Quantum Theories der Strahlung which he started using the name stimulated emission in 1916, other scientists made forward steps on the development of laser theory. Maiman presented the first working laser, a ruby laser in 1960 in a press conference that marked a great step in the development of laser (Tunr & Hode 2002).

There are lots of different electromagnetic radiations from large variety of sources in the earth and the sun is the major natural source of radiation and energy. Light waves produced by laser have two essential characteristics to differentiate them from normal light waves, i.e., narrow bandwidth and high level of coherence. Coherence means that the stimulated photons are ordered and synchronized over a long distance while a narrow bandwidth is crucial for such a high degree of coherence. These two features are typical for laser and are also very important in laser therapy. Simply speaking, a laser was designed to include an energy source, which is the power supply and it may be an electric current, optical radiation from a flash lamp or another laser, radio waves or microwaves, or chemical reactions. The lasing medium is the material able to store the energy supplied and can assist in 9

Literature Review

transmitting the energy to the photons in an organized fashion with the same energy. The lasing medium can be solid, liquid or gas. A resonating cavity, which is also an integral part in laser design and is formed by mirrors fitted inside the lasing medium in order to increase the laser mediums amplification and make the light more coherent. There are also other features that many people believe to be typical; parallel beams and high intensity. These two features can be changed by altering the laser design and hence, they are not as typical characteristics as those mentioned before.

1.5 Types of laser1.5.1 Surgical or hard laser

In medical and dental field, the use of this high technological device can commonly be divided into two categories i.e., hard/surgical laser and soft/low-power laser. For surgical laser, they must have the power high enough to heat up tissues to temperatures over 50C. They produce vaporizing (1-5W), light cutting (5-20W) or deep cutting effects (20-100W) depending on the output power. One of the examples related to periodontal therapy is the use of Nd:YAG laser for soft tissue surgical procedures, such as pocket elimination and gingivectomy (Gold & Vilardi 1994, Pick & Colvard 1993, White et al. 1991). It is worthwhile to know that the use of this type of laser has recently been approved by the Food and Drug Administration in the United States. Removal of calculus and decontamination of root surfaces (Cobb et al. 1992, Schwarz et al. 2003), caries removal (Keller et al. 1998) and soft tissues operations (White et al. 1991) are the procedures that use

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laser technology instead of conventional instruments in the last decade. However, the potential for dental tissues injuries cause by surgical laser should not be overlooked.

1.5.2 Low-power or soft laser

In contrast to surgical laser, low-power laser usually operates in the milliwatt range, i.e., 1-500mW. There are several types of laser in this category with the well known ruby laser (Mester et al. 1971) and the Helium-Neon (He-Ne) laser. They generally operate in the visible or infra-red spectrum, 600 900nm wavelength and use gas or semiconductor as laser medium. Use of low-power laser in dentistry is not new and this has been widely used in Japan, China and Eastern Europe. With more than 30 years of knowledge in using this kind of laser, a wide variety of clinical procedures have been performed. Many therapeutic effects of the low-power laser have been claimed which included acceleration of wound healing (Medrado et al. 2003), enhanced remodelling and repair of bone (Barushka et al. 1995, Saito & Shimizu 1997, Trelles & Mayayo 1987), restoration of neural function after injuries (Miloro & Repasky 2000) and reduction of pain (Lim et al. 1995). Although positive results of the above procedures have been reported in the literature, it is still a dilemma to the clinician when choosing low-power laser therapy for their patients due to the lack of scientific evidence from the literature, improper clinical trial design and poor documentation. Apart from that, the exact mechanisms of low-power laser on various therapeutic effects are still not yet known.

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1.6 Laser safety

The concern in using laser as therapeutic regimen was the risk of eye injury. Even in the infancy of this technology, there was an awareness of the increased risk. Until now, very few accidents have been reported. When such an incident did occur, it was usually because of strong industrial lasers being used or protective measures ignored. Another supporting evidence of the safety in using laser is from the treatment of diabetic retinopathy. Vision corrections are all directed to the eye but without producing any injury although they are capable of burning tissues. Therefore, it seems that the risk is exaggerated and the use of low-power laser can be very safe if strict precautions are followed.

Another concern would be the cancerous change caused by laser. In principle, the shorter the wavelength, the higher ionizing power, no matter the intensity is low or high, the greater the chance of causing cancerous change. Theoretically, electromagnetic radiations have ionizing effects if their photons have enough energy level which is 3.91eV and wavelength 320nm. However, as low-power level laser always have wavelengths above 630nm, such photon energy is not capable to cause any ionizing effects and cancerous change.

1.7 History of low-power laser (therapeutic laser) therapy

Ruby laser was the oldest low-power laser reported and it emits at a wavelength of 694nm. In 1967, Mester et al. (1971) started to investigate the stimulatory effects of

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ruby laser in animals. They reported that there were better and faster reappearance of shaved fur in rats when the sites were irradiated and called this effect as biostimulation. However, such effect was dose dependent. When the dose is too low, there will be no effect and conversely, the results will be no or even suppressive effects in cell cultures if the dose is too high,

1.8 Laboratory studies of low-power laser

Various biostimulatory effects of low-power laser irradiation on different tissues have been reported, such as fibroblasts (Kreisler et al. 2003, Loevschall & ArenholtBindslev 1994, Mester et al. 1971), bone cells (Drtbudak et al. 2000, Ueda & Shimizu 2001) and epithelial cells (Haas et al. 1990, Rood et al. 1992). Most of the studies were performed on cell cultures and positive effects on cell proliferation, differentiation, collagen synthesis and release of growth factors have been shown.

1.8.1 Fibrobalsts

Fibroblast was the one type of cell that was investigated most. It has been shown that low-power laser could increase the proliferation rate (Kreisler et al. 2002, Kreisler et al. 2003) , cell numbers (Pereira et al. 2002) , enhance DNA synthesis (Loevschall & Arenholt-Bindslev 1994) and alter their characteristics (Noble et al. 1992, Pourreau-Schneider et al. 1990). In a study by Almeida-Lopes et al. (2001), they compared effects of low-power laser therapy on cultured human gingival fibroblasts proliferation using four different wavelengths: 670nm, 780nm, 692nm,

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and 786nm and the dosage was fixed in 2J/cm2. The cells were grown in two different mediums; ideal and nutrient deficient. Under normal condition, the cell proliferating rate was less in the nutrient deficient medium than the ideal medium. However, when irradiated, cells in nutrient deficient medium presented cell growth similar to or higher than that of cells grown in ideal culture condition. In addition, they also found that lasers of equal power output presented similar effect on cell growth independent of their wavelengths.

1.8.2 Osteoblasts

Other than fibroblasts, researchers were also interested in the effects of low-power laser on osteoblasts. Recently, Drtbudak et al. (Drtbudak et al. 2000) determined the effect of continuous wave diode laser irradiation (690nm) on osteoblasts derived from mesenchymal cells in vitro. The study showed that all lased cultures showed significantly more fluorescent bone deposits than the non-lased cultures. Ueda et al. (2001) investigated the effects of low-power Ga-Al-As laser (830nm, 500mW) in two different irradiation modes; continuous irradiation, and 1Hz pulsed irradiation on osteoblastic cells proliferation, bone nodule formation, alkaline phosphatase (ALP) activity and ALP gene expression. This study showed that both continuous and 1 Hz pulsed irradiation significantly stimulated the above parameters and the effect were dose-dependent in bone nodule formation. The low frequency pulsed laser irradiation produced better results than the continuous mode with the same total energy dose.

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1.8.3 Keratinocytes

Two other studies reported that He-Ne laser could enhance the motility of human epidermal keratinocytes to provide better wound healing and did not alter normal keratinocyte differentiation or the synthesis of keratins (Haas et al. 1990, Rood et al. 1992). The results suggested that the laser did not interfere with the formation of normal epidermis during the wound healing.

1.8.4 Bacteria

Recently, the use of low-power laser has been focused on the elimination of bacteria by photosensitizing effects. Some studies have demonstrated that lethal photosensitization of bacteria could be achieved in vitro without causing any damage on the treated surface (Dobson & Wilson 1992, Haas et al. 1997) and successfully reduced the bacterial counts in peri-implantitis patients using toluidine blue as the dye with low-power laser. Based on this report, it seems that this approach may be very useful in treating periodontal disease and peri-implantitis.

From the above in vitro studies, it can be seen that low-power laser can produce stimulatory effects on various cells and hence, may have potential beneficial effects in clinical situations. However, factors like optimal energy dose, laser spectrum, power density, irradiation phase and the pulse frequency, which are important in influencing the biological in vivo response, have not yet been clearly defined.

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1.9 In vivo studies of low-power laser1.9.1 Animal studies

Several animal studies reported the effects of low-power laser on wound healing and bone regeneration. Most of these studies used rats as the animal model. In an animal study investigating the effect of low-power laser in wound healing, Medrado et al. (2003) found that the extent of edema and number of inflammatory cells were reduced while the amount of collagen and elastic fibers were slightly increased. In contrary, different results were reported by Schlager et al. in two studies (2000b, 2000a). In the first study, they investigated the effects of two different low-power diode laser lights (635nm and 690nm) on the healing of burns in rats. Results showed that the diameter, redness, and edema of the wounds were similar throughout the entire observation period when comparing between or within irradiated wounds. In the second study, thirty rats were used and the flanks were burned on both sides. On each rat, one side was irradiated with 670nm laser light at 2J/cm2, whereas the other side remained untreated. They found that the irradiated wound showed no accelerated wound healing when compared with control wounds in histological examination. Conflicting results were reported in this aspect would probably be due to the different study protocols especially the dosage used.

In contrast to wound healing, studies on bone formation or regeneration seemed to provide more consistent and favourable results with the use of low-power laser. As early as in 1980s, Trelles et al. (1987) started an experimental study of bone fracture healing in the tibia of mice using He-Ne laser at 4mW, 632nm and a energy of 2.4J. An increase in vascularization and a faster formation of osseous tissue with a dense 16

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trabecular pattern was observed when compared to the control group, which presented with chondroid tissue and poor vascularization that correspond to an earlier stage of bone consolidation.

Two studies focused on the effect of low-power laser irradiation on bone response in rats. Barushka et al. (1995) found that the use of low-power laser could promote healing of fractures or acute defects in bones. The other study showed that lowpower laser could accelerate bone regeneration in a midpalatal suture during rapid palatal expansion (Saito & Shimizu 1997). Recently, computerized morphometric assessment of the effect of low-power laser therapy on bone repair was reported by Silva Jnior (2002). The amounts of newly formed bone after Ga-Al-As laser irradiation of surgical wounds created in the femur of rats and the results showed that there was an increase of bone repair at the early bone healing stage after irradiation.

Effect of low-power laser in implant sites were investigated and published recently. Drtbudak et al. (2002) examined the effects of low-power laser on osteocytes and bone resorption at bony implant sites in baboons with a 690nm laser at 100mW irradiated for 10 minutes immediately after drilling and insertion of implants. Results showed that higher osteocytes viability and similar bone resorption in irradiated sides.

Contradictory results were reported by David et al. (1996) where they investigated the radiological, biomechanical, and histological effects of He-Ne irradiation on fracture healing using rats. Radiological and histological examinations of the

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osteotomy sites failed to show any enhancing effect of He-Ne laser irradiation on the bone healing process. In addition, the irradiated bones of two of the six test groups were significantly weaker than the controls.

Unlike those in vitro studies, it seems that the effects of low-power laser did not provide consistent results in the animal studies we discussed so far. The possible reasons could be the more complex biological response in animals and also the different parameters used to investigate the effects in different studies.

1.9.2 Clinical studies in human

There has been an increase in research work with the application of laser in clinical situations during the last decade. Although the use of low-power laser in human started in late 1960s, it is still questioned by a lot of clinicians or scientists due to the lack of scientific reports written in English. Many studies of low-power laser therapy were conducted in Asian (Japan and China) and Eastern European countries and the results were reported in their own languages. For example, the Russian literatures related to low-power laser therapy appeared for 30 years but most of them were still not easy accessible by others in the world (Walsh 1997). There have been many therapeutic claims of the low-power laser in dentistry including management of aphthous ulcer (Colvard & Kuo 1991), reduction of pain and discomfort (Clokie et al. 1991, Lim et al. 1995), treatment of dentine hypersensitivity (Gerschman et al. 1994) and neurosensory recovery after surgical procedures in head and neck region (Khullar et al. 1996b, Khullar et al. 1996a, Miloro & Repasky 2000). Positive results were shown from the respective studies 18

Literature Review

but some negative results were also reported (Fernando et al. 1993, Roynesdal et al. 1993). In fact, it is not surprising to find conflicting results in clinical studies as the dose energy, time and frequency of treatment are different from different studies and more complex biological responses are involved.

In the periodontal context, effects of the low-power laser were investigated by Masse et al. (1993) and Rydn et al. (1994) in two different aspects. Masse et al. (1993) determined the post-surgical analgesic, anti-inflammatory and healing effects of low-power laser as an adjunctive treatment in periodontal patients. The laser used was a Ga-As laser with an output power of 2W combined with a He-Ne laser at 4mW. Results showed no significant differences in pain, oedema, gingival index and healing appearance when compared to the placebo operation. However, the dosage that was used in this study was not in full strength as the manufacturer specified. Such dosage was considered as very low as it was only about 0.0422J/cm2 and no therapeutic effects would be expected.

Rydn et al. (1994) investigated the effect of low-power on gingival inflammation by a stereophotographic technique. Lower incisors on one side were randomly allocated for irradiation while the contra-lateral sites served as control. The laser used was an infra-red pulsed Ga-As laser (IR CEB-UP MID Laser, Space Lase, Italy) and the total dose delivered to the sites was 1J/cm2. Results showed that there was no statistical significant difference between the test and control sides in plaque formation or gingival bleeding after 28 days.

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Literature Review

1.10 Conclusion

Non-surgical periodontal therapy is the conventional treatment aiming at removing plaque and calculus subgingivally which provides a biological acceptable root surface for healing (Corbet et al. 1993). Previous studies (Badersten et al. 1981, Badersten et al. 1984a, Claffey et al. 1988) already showed the effectiveness of this therapy in terms of clinical and microbiological changes and found that most of periodontal patients and periodontally involved sites responded well to this therapy. However, studies also showed the limitation of this therapy especially in treating deep periodontal pockets and sites with furcation involvement (Loos et al. 1988b, Nordland et al. 1987). Different adjunctive approaches have been tested, although most agents found to have minimal benefits. Majority of these materials in use nowadays are antimicrobials, except the use of sub-antimicrobial dose of doxycycline.

Hence, it would be useful to have an agent that can alter the host response during healing process or in other words, promote the healing response. Although there were data suggesting low-power laser could accelerate wound healing (Mester et al. 1971) and exert biostimulatory effects of different cells (Drtbudak et al. 2000, Kreisler et al. 2003) in vitro, results from clinical studies were contradictory. This may due to different protocols in using low-power laser that were attempted in various clinical studies or even improper use of low-power laser. Therefore, it is the attempt of this study to investigate the possible effect of low-power laser in nonsurgical periodontal therapy as an adjunct under an appropriate protocol.

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Materials and Methods

Part II Materials and Methods


2. Materials and Methods2.1 Aim of study

To evaluate the adjunctive effects of a low-power laser in non-surgical periodontal treatment of patients with moderate to advanced chronic periodontitis

2.2 Objectives

1. To investigate the adjunctive effects of a low-power laser in non-surgical periodontal treatment by comparing between low-power laser treated and untreated sites using the following parameters: a. Clinical parameters i.e., presence/absence of supragingival plaque and bleeding on probing, probing pocket depth, recession and probing attachment level b. Alveolar bone changes c. Volume of gingival crevicular fluid (GCF)

2. To compare the changes after non-surgical periodontal therapy at 1-month, 3months, 6-months, 9-months using the above parameters

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2.3 Study Population

16 Chinese patients were selected from the patient pool of Periodontology and those attending Primary Care Unit of the Prince Philip Dental Hospital, the University of Hong Kong, according to the following criteria:

2.3.1 Subjects selection

Chinese patients with moderate to advanced chronic periodontitis Age from 30 to 60 with more than 20 standing teeth With at least two teeth in each quadrant having a probing depth more than 5mm

With no systemic medical conditions known to be associated with the periodontal condition

No immunosuppressive drugs received before Non-smokers No previous antibiotic treatment within the last 3 months Not pregnant No periodontal treatment except oral hygiene instructions in at least 6 months Not wearing denture or receiving orthodontic treatment

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2.3.2 Teeth/sites selection for laser irradiation

Presence of sites with probing pocket depth of 5mm or above Presence of infra-bony defect No un-restorable carious lesions No obvious cracks involving the roots Responsive to Electric Pulp Test

Two sites on either side of a patient were chosen for laser irradiation. Preferably one site from the anterior region i.e. canine to canine and the other site from the premolars were selected. Molar teeth were excluded. As only the interdental bone can be assessed by the radiographs, all the sites that was chosen would be sites on the interdental region.

2.4 Study Design

The present clinical study was a single-blinded, longitudinal study lasting for 9 months. Matched sites with probing pocket depth of 5mm or above and comparable radiographically detected bone defect were selected on each side of the mouth. Laser irradiation was performed on the selected sites of the test side which was chosen randomly by tossing a coin. Clinical parameters were obtained at baseline, 3 months, 6 months and 9 months after completion of the non-surgical periodontal treatment. Gingival crevicular fluid (GCF) samples and standardized intra-oral radiographs for evaluation of alveolar bone changes were also taken at the baseline and after 1 month,

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3 months, 6 months and 9 months. All clinical data, radiographs and GCF samples were taken by a single calibrated operator, and all treatment procedures and reviews according to the clinical protocol were also carried out by the same operator who was unaware of the sites being irradiated

2.5 Ethical aspect

Detailed explanation about the study was given to suitable patients after screening (appendix IA & IB). Written consent (appendix II) was obtained from the patients willing to take part in the study. The study was approved by the Ethics Committee of the Faculty of Dentistry, The University of Hong Kong.

2.6 Clinical protocol2.6.1 Baseline examination

Upper and lower study impressions were taken pre-baseline at screening for fabrication of occlusal stents for clinical data collection in future appointments. After comprehensive history taking, full-mouth clinical data, GCF samples and standardized radiographs were taken as follows: 1. GCF samples were taken from selected sites, one healthy and one gingivitis according to the procedure set at 2.62 2. Full-mouth clinical data were taken using the manual probe (PCP-UNC 15, HuFriedy Manufacturing Co., Chicago, IL) at six sites (mesio-buccal, mid-

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buccal, disto-buccal, mesio-lingual, mid-lingual, disto-lingual aspects) of each tooth, except third molars, in the following sequence: Plaque level assessment (Plaque percentage, PI%) according to presence or absence of plaque deposits determined by running the tip of a periodontal probe at each site Probing pocket depth (PPD) and probing attachment level (PAL) using a manual periodontal probe with a custom-made poly-ethylene occlusal stent as a reference guide Bleeding on probing (BOP%) was designated as positive if bleeding occurs within 10 seconds after probing 3. Standardized radiographs of selected sites were taken according to procedure as described in 2.63 4. Change of PAL was calculated by using the measurements of PAL at the baseline and at 3 months and 6 months 5. Change of recession (REC) was also calculated by the difference between the change of PPD and the change of PAL

2.6.2 Collection of GCF

After isolation of the selected sites with cotton roll and the removal of supragingival plaque, a standardized GCF strips (Periopaper GCF strips, IDE Interatate, Amityville, NY) were inserted into the gingival crevice or pockets until there was mild resistance and left for 30 seconds. GCF volumes were determined immediately using a GCF meter (Periotron 8000, IDE Interatate, Amityville, NY).

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2.6.3 Alveolar bone change observed by Digital Subtraction Radiography (DSR)

Two sites of test and control side as previously described were selected from each patient, one from anterior and one from premolars for the assessment of the bone changes. Comparison was made between the test and control sites. The control sites were chosen where the clinical and radiographical conditions were similar i.e. PPD +/ 1mm and comparable extent of infra-bony defect when compared to the test sites. Molar teeth were excluded for better detection of bone changsitees.

In order to have standardized periapical radiographs for subtraction, a custom-made acrylic bite block attached to Rinn XCP film holder (Rinn Corporation, Illinois) was made for each site. The X-ray long cone was modified in order to produce a rigid attachment of the bite block to the cone. As a consequence, the angulations between the X-ray source, the object and the film could be reproducible. All periapical radiographs were taken using paralleling technique with the same X-ray machine (GE 700, General Electric Co., U.S.A.) at the same setting (70kV, 15mA). The Size 1 radiographic films (Kodak Ektaspeed plus, Eastman Kodak Co., U.S.A.) were used for the anterior regions, while size 2 radiographic films (Kodak Ektaspeed plus) were used for the premolars. All the films were developed with an automatic developing machine (Periomat, Drr Dental, Germany).

All the radiographs were then scanned into the computer at 600dpi with a flatbed scanner (Agfa Studiostar, Agfa Gaevert, Belgium) and stored in the hard disk of a personal computer. The images were imported into an subtraction software based on the Linux system (Woo et al. 2003).

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2.6.4 Non-surgical periodontal treatment

Oral hygiene instruction was given before the commencement of the treatment. Nonsurgical periodontal treatments including supra-gingival and sub-gingival

debridement under local anaesthesia were provided in two appointments within two weeks time by one operator. The first appointment of non-surgical therapy performed on the test side where laser irradiation was assigned. Oral hygiene level was constantly re-assessed and reinforcement provided if necessary.

2.6.5 Laser irradiation

A low-power laser (He-Ne) operating at a wavelength of 632nm and an output power of 0.2mW was used in this study. It was applied onto two selected sites (test) through an optical fibre tube that attached to the laser machine. Laser irradiation was performed by another trained operator who was not involved in data analysis. It was applied directly onto the two selected sites where the fiber optic was put perpendicularly to the interdental papilla either on buccal or palatal sides as selected. This procedure was performed immediately after non-surgical therapy for a total of 10 minutes each and during the review appointments of the following weeks as shown in Figure 1. A total of 8 times of laser irradiation was performed in the selected sites.

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non-surgical periodontal treatment 2 weeks 2 weeks 1 wk 1 wk

Review

Maintenance Review

2 wks

2 wks

2 wks 1 month 3 months 3 months Laser 2

Laser 2 Laser 2 consent, examination, baseline records sampling OHI sampling Laser

clinical records sampling Laser

clinical records sampling

Figure 1 Time schedule of the clinical trial

2.6.6 Recall appointment

The clinical examination was carried out in the same sequence as in the baseline examination during the 3-, 6- and 9-month recall visits. Additional GCF samples and radiographs were taken during the 1-month visit. Oral hygiene reinforcement and professional prophylaxis were given in every appointment and maintenance visits were scheduled at monthly intervals. Subgingival debridement for pocket >5mm was performed as necessary.

2.7 Analysis of alveolar bone changes

The analysis of the alveolar bone changes was performed using a DSR system developed locally (Woo et al. 2003). The first step in the software was to align the

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paired images by selecting the same sets of two reference points. The software then compared the coordinates of the reference points and moved the subsequent image vertically, horizontally and rotationally until the pairs of images matched. Pixel-bypixel movement of the subsequent image could be performed manually whenever necessary. Gray-level normalization was performed nonparametrically using a cumulative density function (Rttimann et al. 1986). After normalization, the images were digitally subtracted. The selected sites were defined as region of interest (ROI) on the radiographs. The threshold value of the system in this study was determined by taking selected repeat radiographs of the same site on the same visit and a total of 7 pairs of radiographs were obtained for this purpose. They were then used for calculating the threshold value for this study in which noise level