Determination of the Effect of Two Different Methods of Dental Anaesthesia on Pain Level in Pediatric Patients: A Cross-Over, Randomised Trial

Objective To compare intraosseous (IOA) and needle-free dental anaesthesia (NFA) methods that painless anaesthesia. Methods Twenty patients aged 8–10 years were included in this cross-over study. To determine the anxiety levels and pain experienced by the patients, Face, Legs, Activity, Cry, Consolability (FLACC) and Frankl Behavioural scales were used. The pulse rate (PR) and salivary opiorphin levels (SOL) determined. The Friedman and Wilcoxon signed-rank tests were used. p<0.01 was considered signicant. According to FLACC scores, IOA and NFA exhibited signicantly pain alteration patterns in during local and topical anaesthesia, respectively (p=0.004,0.001; p<0.01). Also, only NFA showed signicantly decreased SOL values in 5- and 10- minutes after local anaesthesia periods compared to the before levels (p=0.004, p=0.001; p<0.01). Patients feel similar pain perceptions during local anaesthesia application in both injection systems. According to the SOL values, NFA may provide more higher anaesthetic eciency than IOA.


Introduction
Dental treatments are often associated with pain but their patient acceptance has been facilitated by hypodermic injection of procaine and lidocaine amide as anesthetic agents. However, many patients associate pain with injection due to mechanical trauma of needle insertion or the sudden stretching of tissues by anesthesia during rapid release of the anesthetic [1].
According to the American Dental Association, fear of pain can prevent patients from visiting their dentists also local anesthetic injection is the most anxietyinducing procedure for patients [1]. Painless anaesthesia is a critical issue in terms of increasing the patients' trust in the physician during any procedure.
However, a child's feeling of anxiety during dental treatment may lead to maladaptive behaviours that may hinder and delay the treatment [2,3]. Therefore, especially in pediatric dentistry, ways to avoid the invasive and often painful nature of injection are being investigated [4]. Different measures to reduce discomfort associated with an injection have been tried like that using of topical anaesthetic, warming anaesthesia solution to body temperature using of lowlevel laser or another local anaesthetic delivery methods [1,5].
Computer-controlled local anaesthetic delivery systems (CCLAD), SleeperOne TM , Wand [6] and Comfort-In TM , which provides anaesthesia using pressure, can be regarded as new systems that have been developed to perform local analgesic injection while overcoming the shortcomings of traditional in ltration procedures. SleeperOne TM is a new type of CCLAD with a persistent resistance system analysis that regulates injection according to tissue density [4]. Unlike traditional needle syringe injection methods, it was indicated by the manufacturer that the in ltration anaesthesia injection of Comfort-In TM provides local anaesthesia by penetrating below the periosteum [7].
Although previous studies have generally compared traditional techniques and CCLAD systems using behavioural and pain scales, [8,9] a comparative study of Comfort-In TM , a new needle-free anaesthesia (NFA) method, and SleeperOne TM , an effective CCLAD method, is not available in the literature. In addition, in the literature review, no study was found in which the effect of anaesthesia methods on pain sensation was compared using a 'cross-over' research design by obtaining both biological, physiological and observational data from the patients with the aim of evaluating the effect of newly developed anaesthesia methods on pain levels in patients. Therefore, the project we designed is innovative compared to other studies.

Hypothesis:
No difference in pain levels of systemically healthy 8-10-year-old patients will be observed during local anaesthesia performed as the intraosseous anaesthesia (IOA) or NFA.

Sample size determination
The sample size was determined based on the study conducted by Campanella et al. [1] with an effect size=0.5, alpha error=0.05 and beta error=0.20, it was determined that a total of 35 samples were needed and considering possible losses, a total of 40 patients were found to be su cient. Since the study was designed as a 'cross-over' study, 20 patients were included in the study.

Patient selection
Patients aged 8-10 years, who were systemically healthy, without any physical or psychological disorders, did not use any medication, had undergone dental treatment (except for extraction) previously but with at least 1 week passed since then, had never tried IOA and NFA anaesthesia techniques before, were not undergoing any orthodontic or ongoing dental treatment (e.g. Root canal therapy), were compatible enough to undergo therapy in the clinic in a sitting position, had infection between the roots with a rate over 1/3, had 2/3 root formation, had been indicated for extraction of the maxillary primary molar teeth (55/65 No. teeth), [10] and did not have acute and/or subacute infection of the related or other teeth or mouth and surrounding tissues were included in the study. Further, patients who scored above 41 in the state anxiety subscale and above 44 in the trait anxiety subscale [11] of the Spielberger State-Trait Anxiety Scale (SSAS-SAAS) were not included in the study.
In this study, fourty patients' legal representatives that suitable for including criteria and coming university dental hospital paediatric clinic between September 2020 and January 2020 were interviewed. Among them nine patients did not accept to attend study. After the procedures were started, ve patients were withdrawn from the study voluntarily, while four patients were excluded due to cooperation problems. Addition, although the patients were excluded from the study due to data loss in the video recordings of 2 patients, the study was completed patients with a number suitable for the sample size (n=20) (Fig. 1).

Randomisation
Twenty patients who presented to the clinic, met the inclusion criteria and agreed to participate in the study were assigned numbers from 1 to 20, respectively. Subsequently, patient numbers were randomly distributed using a computer-assisted randomization [12] programme. According to the sequence number of the patients, envelopes containing details about the application group (i.e., which anaesthesia method would be applied rst) were chosen by each patient in a blinded manner. Accordingly, the rst procedure to be applied to that patient was decided.

Study design
A cross-over design that in which two or more treatments are applied to the same patient within the framework of a plan was used in this study [13]. An informed consent form was obtained from all patients included in the study and their parents before the procedure. Patients included in the study were taken to isolated single rooms around 9-10 am. The patients were asked to feed and brush their teeth approximately 1 hour before the dental procedure.
SSAS-SAAS was administered by the principal investigator to determine the patients' state and trait anxiety levels before the procedure. The SSAS-SAAS scale consisted of two separate scales included of 20 questions each, state and trait, and each question was scored from 1 (not at all) to 4 (completely). The scales included direct and reversed statements like "I am currently calm" and "I usually get tired quickly" [14].
Subsequently, like in every patient who presented to the pediatric dentist clinic, the tell-show-do behaviour guidance technique was used and unstimulated saliva samples were passively taken three times from the oor of the mouth of each patient using a Pasteur pipette: 15 minutes (min) before dental anaesthesia, 5 min after dental anaesthesia and 10 min after dental anaesthesia [15,16]. A pulse metre (Yonker Fingertip Pulse Oximeter, Prolinx GmbH, Germany) was attached to the patient's right index nger 15 min before the procedure as an indicator showing the relationship of heart rate (measured as pulse rate (PR)) with pain.
During dental anaesthesia with two systems, 4% articaine 1/100.000 (Ultracain DS Forte Ampul, Articaine HCl 40 mg/ml Epinephrine HCl 0.012 mg/ml, PharmaVision San. ve Tic. A.Ş., Turkey) [17] in ampoule and carpule forms were used. No topical anaesthetic was applied before anaesthesia. All anaesthesia procedures and tooth extraction treatments were performed by the same researcher who had 1 month of experience using Comfort-In™ (Mika Medical; Busan, Korea) and SleeperOne TM (DHT, Cholet, France) injection systems.
The Frankl and the FLACC scores were determined by the two calibrated pediatric dentists, who did not perform the procedure, to objectively evaluate the patient's behaviours and pain levels before the treatment, during the administration of local anaesthesia, and during and after the treatment, based on the video recordings of the patients. Before the study these two pediatric dentists evaluated FLACC and Frankl scales on 10 patients that independent of the study group, and the interobserver agreement was found to be good (κ = 0.729). In addition, the Wong-Baker Faces Pain Rating Scale (WB) that included faces describing pain from 0 (no pain) to 10 (unbearable pain) was used before the procedure, immediately after anaesthesia, immediately after extraction and 15 min after the procedure to analyse the subjective pain felt by the children [18]. The FLACC scale graded ve pain-related behaviours with 0, 1, or 2 points, and an increase in the score indicated the presence of pain. The Frankl Behavioural Scale rated children's behaviour in dental treatments from 1 (absolutely negative) to 4 (absolutely positive). However, all scales (SSAS-SAAS, FLACC, Frankl, WB) were used in the study were used in other studies in Turkish validated format [14,19].
The same parameters were tested 1 week later while performing the left/right upper tooth extraction using the dental anaesthesia technique that had not been applied in the rst session based on randomisation in each patient (Fig. 1).

IOA anaesthesia procedure
The installation of the SleeperOne TM (DHT, Cholet, France) anaesthesia device (needle diameter and length, 30 Gauge/9 mm, a double row needle that provides better penetration into the bone) [20] was made in accordance with the manufacturer's instructions. Before local anaesthesia, 1-2 drops of articaine solution was applied to the buccal region after super cially passing the epithelium for topical purposes, and then the IOA method was used. IOA was performed directly on the interproximal bone between the two teeth. By approaching from the buccal part, the needle was gently pushed into the cortical bone under the interdental papilla at an angle of 45° with clockwise and counter clockwise movements, [4] and a total of half carpule (1 cc) of the anaesthetic was administered (Fig. 2).

NFA anaesthesia procedure
Likewise, the installation of the Comfort-In TM (Mika Medical; Busan, Korea) anaesthesia device was performed in accordance with the manufacturer's instructions. Before anaesthesia, 1 shot of 0.1 ml was applied to the buccal and palatal regions for topical purposes, and after 10-15 seconds of anaesthesia, 2 shots of 0.3 ml each to the buccal region and 2 shots of 0.1 ml each to the palatal region were administered (total 1 cc) [7] (Fig. 3).
Obtaining saliva samples Saliva samples were taken from the individuals in the patient group in a biochemistry tube with separator gel. Consent forms were collected along with the saliva samples. The tubes were centrifuged at 1800 ×g for 10 min. After centrifugation, the supernatants were carefully transferred to capped microfuge tubes and stored at −80ºC until analysis. The saliva samples were brought to room temperature and thawed before analysis. The thawed samples were homogenised by vortexing and the biochemical parameters were measured.
Determining opiorphin levels SOL was determined using the enzyme-linked immunosorbent assay (ELISA) kit (Cusabio, Cat No: CSB-EQ027423HU, Wuhan, China) [21] in accordance with the manufacturer's instructions. The opiorphin standards (40-0.625 ng/mL) were prepared according to the kit-speci ed procedure. Absorbances of the samples and standarts were measured at 450 nm using a microplate reader spectrophotometer (Versamax, Molecular Devices, California, USA). Each sample was analysed in duplicate, and their results were averaged and presented in ng/mL units. The intra-assay repeatability (%CV) of the ELISA kit used was calculated as 3.2% (n=20).

Statistical analysis
A statistical data programme was used for statistical analyses. Descriptive values were assigned to sex and age. Descriptive values were expressed as mean±standard deviation, median (minimum-maximum) or % values. Cohen's Kappa value was calculated for inter-observer agreement. Normal distribution of the data was evaluated using the Shapiro-Wilk test. The Friedman and Wilcoxon signed-rank tests and Bonferroni correction were applied for both continuous (SOL, PR and SSAS-SAAS scores) and categorical variables (WB, Frankl and FLACC scores). In all analyses, a p-value of <0.05 was considered statistically signi cant.

Blinding
The Frankl Behavioural Scale and the Face, Legs, Activity, Cry, Consolability (FLACC) Scale datas were scored from the video recording images of the procedure by two other pediatric dentist researchers who were blinded to the applied techniques. The researchers who determined the levels of opiorphin in the collected saliva and performed the statistical analyses blinded to the grouping of the patients. Although the patients had partial awareness of the technique applied, since all evaluations were performed on video recordings, the risk of subjective results during the pain scoring of the patients was avoided and objective blinding could be established.

Discussion
One of the most important factors affecting success in pediatric dentistry is pain control [22,23]. In the literature, methods such as the use of computercontrolled anaesthesia delivery systems, modern devices and new NFA systems have been investigated to reduce anaesthesia-induced pain [23,24]. However, according to our knowledge, no study has evaluated the effects of IOA and NFA methods on patients' pain levels. Therefore, this study was designed and conducted to eliminate this research gap and to compare two different advanced dental anaesthesia devices.
In this study, the cross-over design was preferred as it eliminates subject variability and allows greater biological homogeneity [13]. The reason for choosing children aged 8-10 years was that there are studies [18, 23,24] reporting that the scales used herein are reasonable and reliable for children in this age group. [17,25,26] In addition, the tooth extraction treatment was chosen in this study because of it was most painful procedure for children [27,28].
WB, which was developed for the subjective assessment of pain, is an easy-to-use scale with adequate psychometric properties [18]. FLACC, one of the scales used in pain measurement, has been shown to have excellent validity and reliability for the objective assessment of pain in young or cognitively sound children [29]. In this study, WB and FLACC were used to evaluate children's pain behaviours both subjectively and objectively, taking advantage of both these scales. The general anxiety level of the individuals may affect the speci c anxiety level related to the procedure [14]. Baygin et al. [14] evaluated the relationship between pre-procedural anxiety level and perceived pain level during the procedure in children undergoing tooth extraction and reported that the measured anxiety levels were correlated with each other [14]. In this study, SSAS-SAAS was used to determine state and trait anxiety levels of the patients when they came to the clinic, and the Frankl Behavioural Scale was used to evaluate the anxiety status throughout the procedure. While the trait anxiety subscale scores of the patients included in the study did not change due to the cross-over study design, the absence of a statistically signi cant difference between the state anxiety subscale scores between sessions (p=0.636; p>0.05) suggested that the initial anxiety levels of the patients were similar. This outcome may have simpli ed to compare the anaesthesia methods during the study period.
In addition to the pain and anxiety scales used in the study, PR [1,30] and SOL, which are physiological and biological markers [28] that indirectly measure pain and anxiety, [15,31] were used to obtain measurements independent of observer bias or subjective reporting of the patient [1]. Similarly, there are studies in the literature that have used PR [1,28] and SOL [15,16,31] to analyse pain and anxiety.
The hypothesis of the study that there would be no difference in felt pain during local anaesthesia between IOA and NFA systems is partially accepted.
According to increases FLACC scores during local and topical anaesthesia, IOA and NFA systems produced pain during local and topical anaesthesia, respectively. However, the increase in FLACC during local and topical anaesthesia with IOA and NFA systems may also have occurred owing to the fear of injection [27]. Although it was not statistically signi cant, two methods showed similar patterns of change according to the Frankl, WB and PRs.
According to our knowledge as there is no study in the literature that compares IOA and NFA systems, it may not be appropriate to compare the results of this study entirely. In general, studies in the literature have compared traditional injection and CCLAD, they report that CCLAD to be more advantageous [2,32,33]. However, there are also studies that did not nd a statistical difference between the two injection methods [17,25,34].
IOA has high e cacy rates, but it is rarely used in young patients [36]. In previous studies conducted on adults that compared IOA with conventional syringes and computerised devices, pain ratings were found to be higher in the early stages of the traditional procedure than in the computerised procedure [36]. Pain due to osseous perforation was mild to moderate in 8-78% and severe in 0-15% of patients [35,36]. In a study by Sixou et al., [35] in which they compared IOA and conventional anaesthesia, they noted symptoms of pain/discomfort during mucosal anaesthesia and bone perforation, but these ndings were not associated with high pain scores. The ndings of pain/discomfort during IOA in patients can be considered as a result similar to the patient's feeling of pain during local anaesthesia in this study. Because of the device used in the study by Sixou et al. [35] was an advanced model of SleeperOne TM , it can be assumed that their results are not entirely comparable with the results of this study.
A study by Versloot et al. [37] found that using CCLAD (Wand) did not provide any bene t compared to traditional injection as it creates additional discomfort for the child owing to the long injection time in extremely anxious children. Wand is one of the CCLAD similar to SleeperOne TM [4]. Nieuwenhuizen et al. [4] compared Wand and SleeperOneTM in their study. They found no signi cant difference in pain between two devices but the duration of anaesthesia injection for SleeperOneTM was shorter than that for Wand. Although no measurements were made in this study, it can be thought that the longer duration of anaesthesia injection in the SleeperOne TM injection method [38] may have been affected the patient's feeling of pain during local anaesthesia.
In this study, the statistically signi cant decrease in the SOL for the NFA system, both 5 min and 10 min after anaesthesia, was accepted as important biological data showing the effectiveness of anaesthesia. Similar results were obtained in a study by Parida et al. [15] evaluating the relationship between SOL and the use of different local anaesthetic techniques. They found that SOL increased, albeit not statistically signi cantly, during anaesthesia techniques that cause more pain compared to other techniques. In a study by Ozdogan et al. [31] evaluating toothache due to endodontic dental diseases and SOL concentrations, it was reported that SOL increased signi cantly in toothache due to in ammation and a strong correlation was observed between the reported pain level and SOL. The signi cant decrease in the SOL in this study can be explained that local anaesthetics control pain or the element of psychological relief after the onset of lethargy is involved [15].
In the literature, in studies where NFA method was evaluated, high patient and operator acceptance has been reported [39,40]. There is one study in the literature evaluating the Comfort-In™ system also the study in question used injection for topical anaesthesia. In the study, Yıldırım et al. [23] reported that the NFA system signi cantly decreased the pain scores compared to traditional topical anaesthesia methods, but did not signi cantly affect patient preferences.
In our study, for the NFA system, statistically signi cant increases in scores of the FLACC suggested that the patients were more incompliant and felt more pain during topical anaesthesia. This situation can be supported that NFA requires the use of pressure to rapidly apply the anaesthetic solution to the tissue, indicated by Yıldırım et al. [23]. Hence, a sharp popping sound and a feeling of pressure are experienced during the administration of the anaesthetic solution.
Yıldırım et al. [23] also stated that the negative relationship they reported in their study may be related to the discomfort that younger children felt from the pressure sensation from the Comfort-In™ injection system, and they may have interpreted this as a higher level of pain.
Further, for NFA system, according to the WB scores showed that the patients felt pain during tooth extraction. However, in contrast to WB scores, PRs have decreased during tooth extraction statistically signi cant for NFA method. In addition, the decreases in SOL, which shows the effectiveness of anaesthesia, was statistically signi cant in NFA method. Considering these ndings, it can be considered valid because of that physiological and biological marker such as SOL results as their measurements are not subject to observer bias or not based on the subjective reporting of the patient [1].
The limitations of this study are that the injection systems used in the study were not compared in terms of post-operative discomfort and patient preferences, and that a conventional dental injection group was not established as a control group. In future studies, these limitations should be addressed with larger patient populations, mostly on the basis of biological and physiological markers.

Conclusion
1. The results showed that IOA and NFA systems are similar during local anaesthesia.
2. Further, decreases the SOL, which is a robust biological marker, suggested that the NFA system may have provided higher anaesthetic e ciency.  Intraosseous anesthesia device and its intraoral application Needle free anesthesia device and its intraoral application

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