|Year : 2023 | Volume
| Issue : 3 | Page : 338-343
|Influence of the restoration after pulpotomy on the strength of electrical stimulus reaching the pulp space: An in vitro investigation
Aakanksha Chopra1, Sidhartha Sharma1, Vijay Kumar1, Amrita Chawla1, Suman Jain2, Ajay Logani1
1 Division of Conservative Dentistry and Endodontics, Centre for Dental Education and Research, All India Institute of Medical Sciences, New Delhi, India
2 Department of Physiology, All India Institute of Medical Sciences, New Delhi, India
Click here for correspondence address and email
|Date of Submission||25-Jan-2023|
|Date of Decision||10-Mar-2023|
|Date of Acceptance||22-Mar-2023|
|Date of Web Publication||16-May-2023|
| Abstract|| |
Introduction: The study evaluated the influence of coronal restoration after pulpotomy on the strength of electrical stimulus reaching the radicular pulp using an electric pulp test (EPT).
Materials and Methods: The pulp tissue from ten freshly extracted mandibular premolar teeth was removed and replaced with an electroconductive gel. The cathode probe of Powerlab was inserted into the pulp space and the anode probe was attached to the EPT handpiece. The EPT probe coated with electro-conducting material was positioned in the middle third of the buccal crown surface. The EPT stimulus reaching the pulp space of an intact tooth at 40 numerical readings was recorded. The tooth was removed from model and endodontic access was made. The 2-mm thick mineral trioxide aggregate was placed at the cementoenamel junction followed by composite resin restoration. The experimental setup was re-established and postpulpotomy EPT stimulus data were recorded. The data collected were compared using the Wilcoxon signed-rank test.
Results: There was a statistically significant difference (P = 0.038) between observed between the strength of EPT stimulus reaching the pulp space in prepulpotomy (mean 91.18 ± 101.02 V and median 25.79 V) and postpulpotomy (mean 58.49 ± 77.13 V and median 13.75 V) tooth samples.
Conclusion: The placement of the restoration and pulp capping agent after pulpotomy dampens the strength of EPT stimulus reaching the pulp canal space.
Keywords: Coronal restoration; electric pulp test; pulpotomy
|How to cite this article:|
Chopra A, Sharma S, Kumar V, Chawla A, Jain S, Logani A. Influence of the restoration after pulpotomy on the strength of electrical stimulus reaching the pulp space: An in vitro investigation. J Conserv Dent 2023;26:338-43
|How to cite this URL:|
Chopra A, Sharma S, Kumar V, Chawla A, Jain S, Logani A. Influence of the restoration after pulpotomy on the strength of electrical stimulus reaching the pulp space: An in vitro investigation. J Conserv Dent [serial online] 2023 [cited 2023 Oct 5];26:338-43. Available from: https://www.jcd.org.in/text.asp?2023/26/3/338/376917
| Introduction|| |
Minimally invasive endodontics aims to induce a biological response to preserve pulp health, prevent apical periodontitis and increase the success rate of vital pulp therapy (VPT)., VPT encompasses indirect pulp capping, direct pulp capping, and pulpotomy (partial or full) procedures. When the patient's symptoms are suggestive of irreversible pulpitis and the inflammation is believed to be confined to the coronal pulp (moderate/severe pulpitis), a pulpotomy procedure is prioritized over the root canal treatment (RCT). The pulpotomy involves partial or complete removal of the coronal pulp and application of a biomaterial directly onto the residual healthy pulp tissue before the placement of a permanent restoration.
Pulpotomy is considered a less technique-sensitive and cost-effective procedure compared to RCT.,,, Studies have reported high success rates (75% and 97%) for both partial and complete pulpotomy.,, The recent randomized control trial has reported no statistically significant difference between the outcome of partial (80.8%) and complete pulpotomy (89.8%) for the management of mature permanent teeth with irreversible pulpitis. However, the outcome of a partial and complete pulpotomy are difficult to compare due to the noninclusion of pulp sensibility tests in the outcome criteria of complete pulpotomy. It is the notion that pulp sensibility tests are unreliable in teeth after complete pulpotomy due to large coronal restoration and deposition of tertiary dentine, however, an experimental investigation is missing. On the contrary, literature reports that teeth demonstrate a positive response to the electric pulp test (EPT) after a successful pulpotomy., Therefore, the present in vitro study was designed to evaluate the influence of coronal restoration after pulpotomy on the strength of electrical stimulus reaching the radicular pulp using an EPT.
| Materials and Methods|| |
The in vitro investigation was performed after ethical approval from the institute's research ethics committee (ref. IEC/145/3/2021). A total of 10 single-rooted human mandibular premolar teeth extracted for orthodontic reasons from individuals aged 15–25 years were used after obtaining written informed consent. Teeth with preexisting restorations, cavities, fractures, and radiographic evidence of pulp canal obliteration were excluded. The extracted teeth were manually scaled and cleaned with a pumice slurry to eliminate any remaining soft tissue and stored at the room temperature in artificial saliva until further preparation.
Tooth sample preparation
Three millimeters of the apical root end were resected using a carborundum disc mounted on a straight handpiece (NSK E-Type Straight Handpiece, NSK, Japan). Tissue was removed from the pulp chamber and root canal space through retrograde instrumentation and copious irrigation with 3% NaOCl. A hole was drilled onto the lingual root surface 1 mm apical to the cementoenamel junction using a #2 round diamond bur (Mani, Inc, Japan) to gain access to the pulp canal space. An electroconductive gel (NP-1, Nissin Dental Products Inc., Kyoto, Japan) was used to replace the pulp tissue.
Study model preparation
The study model was prepared by modifying the model described for illustrating the use of electronic apex locators and the experimental setup for measuring the conductance of EPT.,, A dental model for electronic apex locator training (CON1002 Series, Nissin Dental Products Inc., Kyoto, Japan) was selected, and the right mandibular first premolar was removed. The root canal space and the socket were filled with electroconductive gel. The tooth was secured in the socket using sticky wax. The lip clip of an electric pulp tester (Vitality Scanner 2006, SybronEndo, California, USA) was connected to the dental model [Figure 1].
|Figure 1: Experimental setup: Powerlab electrode probes are attached to the tooth and EPT to record the electric stimulus reaching the pulp space. The radiograph demonstrates the level and thickness of the pulp capping agent, coronal restoration and location of the lingual hole for placement of the Powerlab electrode probe. EPT: Electric pulp test|
Click here to view
Electric stimulus measurement
A Powerlab device (DAQ, Data Acquisition Hardware Device, AD instrument Ltd, Sydney, Australia) was used. The cathode probe of Powerlab was covered with an insulating sleeve with only a 2 mm tip exposed and inserted into the pulp space through the opening on the lingual aspect. The anode was coupled to the EPT handpiece. The pulsating electric stimulation rate of the EPT was set at 5 to facilitate readings in a clinical scenario. The EPT probe was coated with conducting media (K-Y Jelly, Reckitt Benckiser) and placed at the middle third of the buccal tooth surface. The baseline readings of the electrical stimulus received at the pulp chamber were recorded using the PowerLab connected to a computer.
The tooth was then removed from the experimental model, and the root canal space was flushed with distilled water to remove the electro-conductive gel. The access opening was performed with the help of a round diamond bur under water spray, and the pulp space was again flushed with 3% NaOCl to remove any residual tissue or gel. A collagen sponge was placed inside the root canal up to the level of the cementoenamel junction. Mineral trioxide aggregate (MTA) (Pro Root MTA, Dentsply Sirona, NC, USA) was mixed according to the manufacturer's instructions. A 2-mm thick MTA was placed over the collagen sponge and gently condensed to simulate the placement of the pulpotomy agent in the clinical scenario. A moistened cotton pellet was placed over the MTA, and the teeth were stored at 100% humidity to allow the setting of the MTA. The moist cotton pellet and collagen sponge were removed after 24 h, and the setting of MTA was confirmed. A layer of resin-modified glass ionomer cement (Vitrebond, 3 m, ESPE, St. Paul, USA) was placed over MTA and light-cured for 20 s. The teeth were restored with restorative composite resin and further incubated for 24 h under 100% humidity and at the room temperature to allow the complete set of coronal restoration. Thereafter, the root canal space was filled with electroconductive gel and the experimental setup was re-established, as explained previously. The EPT probe was contacted at the middle third of the crown on the buccal surfaces, and measurements of the electrical stimulus from EPT transferred to the pulp chamber were recorded on the LabChart (AD instrument Ltd, Sydney, Australia) [Figure 2]. The pre- and postpulpotomy electrical stimulus measurements (in voltage) at EPT reading 40 were recorded three times per tooth, and the EPT was allowed to return to 0 before each reading. The mean value was recorded, and data were stored in a Microsoft excel sheet.
|Figure 2: (a) Pre pulpotomy, (b) post pulpotomy The output signal from the EPT reaching the radicular pulp space recorded in the LabChart in pre and postpulpotomy tooth samples. The magnified view represents a single output burst with ten pulses. EPT: Electric pulp test|
Click here to view
The data collected were analyzed using IBM SPSS statistics version 25 software (IBM Corporation, NY, USA). The data of electrical stimulus reaching pulp space in pre-and post-pulpotomy tooth samples were not normally distributed (Shapiro–Wilk test P = 0.034). The nonparametric test for two related samples (Wilcoxon signed-rank test) was used to compare the cumulative strength of electrical stimulus reaching pulp space in the pre-and postpulpotomy tooth samples at an EPT reading of 40.
| Results|| |
The output signal was in the form of a waveform, with each number corresponding to the burst of 10 pulses followed by a resting period. The output voltage increased gradually from 0 to 40 readings, but the pulse duration and distance between the two output bursts remained constant [Figure 2]. The output voltage (V) (mean ± standard deviation and median, p25–p75) reaching the radicular space varied considerably in prepulpotomy (91.18 ± 101.02 V and 25.79 V, 7.10–165.15 V) and postpulpotomy (58.49 ± 77.13 V and 13.75 V, 7.19–113.05 V) tooth samples. There was a statistically significant difference (P = 0. 038) between the pre-and post pulpotomy tooth samples for the strength of electrical stimulus reaching pulp space, and the values were diminished after the placement of pulpotomy agent and coronal restoration when compared with an intact tooth [Figure 3] and [Table 1].
|Figure 3: Strength of electrical stimulus reaching the pulp space in pre-and post-pulpotomy tooth samples|
Click here to view
|Table 1: Descriptive statistics of power lab readings (V) from pre-and post-tooth samples|
Click here to view
| Discussion|| |
The outcome of pulpotomy depends upon the reparative potential of residual radicular pulp and the asepsis followed during the clinical procedure. The radicular pulp can develop irreversible inflammation as the part of an acute or chronic process. The occurrence of spontaneous, intense, or continuous pain in a pulpotomized tooth during a period of 2 months postoperatively constitutes an acute adverse event. The appearance of condensing osteitis, pulp canal obliteration, internal resorption, and periapical pathology on the radiograph indicates chronic inflammation of the radicular pulp., While an acute adverse event and early failure are easily detectable, the chronic pulp inflammation may often go unnoticed as the patient remains asymptomatic and it may take several months before changes are evident on radiographs. Therefore, periodic monitoring of pulp health status following a pulpotomy procedure is of paramount importance. Pulse oximetry and laser doppler flowmetry are the ideal tools to assess pulp vitality, however, removal of coronal pulp could result in a false-negative response to these pulp-vitality tests. Clinical and radiographic examinations are currently used as surrogate measures to determine the pulp health in pulpotomized teeth.
The dental pulp is richly innervated with the sensory nerves which pass through the radicular pulp to reach the pulp chamber where they diverge and branch outward and form a mesh at the pulpo-dentine junction. The thermal and electric pulp sensibility tests provide an indirect assessment of dental pulp status. The hydrodynamic theory of pulp sensitivity suggests that electrical stimulation and cold stimulation have two different mechanisms of action. According to this theory, cold may cause neurons to act as mechanoreceptors that react to movement from the thermal contraction of dentinal fluid. The presence of vital nerves in the dentine or pulp directly beneath the cold stimulus is presumably necessary to obtain a response. Based on the clinical evidence, where teeth adjudged clinically and radiographically successful after partial or complete pulpotomies but did not respond to the cold test, it cannot be considered a reliable technique for pulp-sensibility assessment., The EPT is believed to cause ionic shifts within the dentin fluid, resulting in the localized depolarization of nerve membranes and the production of action potentials. Logically the use of bipolar EPT which requires the placement of two probes in the buccal and lingual surfaces and is believed to elicit a response by the stimulation of coronal pulp seems ineffective in full pulpotomy cases. With monopolar EPT, stimulus passes through the dentinal tubules beneath the tooth electrode to the coronal and radicular pulp. Apart from the pulp chamber's occlusal wall, the nerve plexus's density is well-developed in the peripheral pulp along the lateral wall of the coronal and cervical dentine. The histologic study demonstrated a close contact between fibroblast-like cells/osteoblast-like cells and nerve terminals at the calcification front in the early healing process after pulpotomy. Thus, it can be hypothesized that if an electrical stimulus reaches the pulp space, an EPT can elicit a positive response from a radicular pulp after pulpotomy.
The European society of endodontology's position statement on management of deep caries and the exposed pulp states that “the teeth which have undergone full pulpotomy are expected to be unresponsive to pulp sensibility testing on follow-up evaluation.” However this is contradictory to the findings of the previous clinical studies that evaluated the patient response to EPT after pulpotomy., It can be asserted that the positive response in teeth after full pulpotomy could be due to stimulation of nerver fibers in supporting periodontal tissues as it is belived that large coronal restoration could block the EPT stimulus from reaching the radicular pulp. This is the first study evaluating the effect of restoration after pulpotomy on the electrical conductance of teeth during electronic pulp testing. Single rooted pre molar teeth were used for the study because of ease of availability of intact premolar teeth extracted due to orthodontic reasons. However, the results can be replicated in multi rooted teeth as well. For the electric stimulus to reach the pulp space, it must overcome the electrical resistance of enamel and dentine which varies with their thickness. Therefore, the extracted teeth used in the study were obtained from individuals aged between 15 and 25 years to circumvent the chances of encountering receding pulp chambers, calcifications, etc., which may alter the conductance of electric current thereby the EPT values.
The experimental setup was designed as a modification of the model described for illustrating the use of electronic apex locators using the electrical root length measurement training model. The conductance media used during EPT also influences the strength of the electrical signal reaching the pulp and the use of media with maximal conduction allows for the most reliable EPT results. Water-based electroconductive media are reported to have higher conductance compared to wax or petroleum-based media. Therefore water-based electroconductive gel was used in the present study.
The comparative study of two EPTs established that the analytic technology pulp tester has the most suitable output characteristics, efficient, and easy to use. The modified version of this pulp tester was used in the present study. The voltage output is generated electronically and each reading on EPT corresponds to a burst of ten pulses. The rate of increase in the strength of the stimulus can be controlled by a panel wheel. It has been recommended that the output from any pulp tester should start at a low level and have an initial slow rate of increase which then accelerates towards the middle and upper range to minimize patient discomfort. Similarly, for a particular dial setting of vitality scanner, the output level rises slowly at the beginning of the test and more quickly at the end. The dial 5 setting for a rate of increase of stimulus up to 40 numerical readings on EPT display was used in the in-vitro experiment to simulate the actual clinical scenario.
VPT has been buttressed with the introduction of calcium silicate-based cements such as MTA, Neo MTA plus, Biodentine, Totalfill and calcium-enriched mixture cement which seal the pulp wound interface, enhancing pulp healing.,,, MTA, the most widely used pulp capping agent after pulpotomy, was utilized in the present study. MTA and other hydraulic calcium silicate cement are considered bioactive, which implies that they release calcium ions, produced calcium hydroxide, and possess electroconductive properties. The MTA has an electric conductivity of 1388.7 263.5 S/cm. In the present study, A significant difference was observed in the cumulative voltage reaching pulp space from 0 to 40, the values were significantly reduced (P = 0.038) after placement of pulpotomy agent and coronal restoration when compared with an intact tooth. This can be attributed to the weak electrical conductance of composite resin used as coronal restorative material which can attenuate the magnitude of current reaching the radicular pulp space. These results are in accordance with clinical observations in previous studies where a significant difference in the pulpal response was observed before and after pulpotomy., Additionally, the presence of fewer myelinated axons with smaller diameters having higher thresholds to electrical stimulation could be attributed to the increased response threshold to EPT observed in pulpotomized teeth.
The electrical stimulus can reach the pulp through the pulp-capping agent. Also, as the dentinal tubules have waveform and there is a possibility that the tubules in the middle and cervical third of the crown transverse up to the radicular pulp. Electric pulp testers based on the production of impulses of negative polarity are claimed to reduce the voltages required to stimulate a pulpal response and so avoid stimulation of the nerves in the periodontal membrane. The histologic examination of the tooth showing a positive response to EPT 10 months after the pulpotomy revealed the presence of vital radicular pulp. Thus, albeit at higher numerical readings, the positive response to EPT after pulpotomy could be used as an indirect indicator of pulp vitality and can be used to monitor the health status of the remaining pulp tissue.
The in vitro study model and inability to simulate periodontal tissue was the limitation of the study. Future studies can be conducted to compare the electrical conductivity of different pulp capping agents and restorative materials. Histologic and animal studies are necessary to establish the reliability of EPTs after full pulpotomy.
| Conclusion|| |
The electrical stimulus from the electric pulp tester reaches the radicular pulp space despite the presence of restoration and pulp capping agent but the stimulus strength is significantly reduced.
Statement of clinical relevance
The EPT can be used for the assessment of pulp sensibility after full pulpotomy. When compared to the contralateral control teeth, it is anticipated that the pulpotomized teeth will respond at greater EPT readings.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
| References|| |
Wolters WJ, Duncan HF, Tomson PL, Karim IE, McKenna G, Dorri M, et al
. Minimally invasive endodontics: A new diagnostic system for assessing pulpitis and subsequent treatment needs. Int Endod J 2017;50:825-9.
Gutmann JL. Minimally invasive dentistry (Endodontics). J Conserv Dent 2013;16:282-3.
] [Full text]
Cohenca N, Paranjpe A, Berg J. Vital pulp therapy. Dent Clin North Am 2013;57:59-73.
European Society of Endodontology (ESE) developed by:, Duncan HF, Galler KM, Tomson PL, Simon S, El-Karim I, et al
. European Society of Endodontology position statement: Management of deep caries and the exposed pulp. Int Endod J 2019;52:923-34.
Simon DS. Endodontic case difficulty assessment: The team approach. Gen Dent 1999;47:340-4.
Zafar K, Nazeer MR, Ghafoor R, Khan FR. Success of pulpotomy in mature permanent teeth with irreversible pulpitis: A systematic review. J Conserv Dent 2020;23:121-5. [Full text]
Asgary S, Ehsani S. Permanent molar pulpotomy with a new endodontic cement: A case series. J Conserv Dent 2009;12:31-6.
] [Full text]
Cushley S, Duncan HF, Lappin MJ, Tomson PL, Lundy FT, Cooper P, et al
. Pulpotomy for mature carious teeth with symptoms of irreversible pulpitis: A systematic review. J Dent 2019;88:103158.
Elmsmari F, Ruiz XF, Miró Q, Feijoo-Pato N, Durán-Sindreu F, Olivieri JG. Outcome of partial pulpotomy in cariously exposed posterior permanent teeth: A systematic review and meta-analysis. J Endod 2019;45:1296-306.e3.
Baranwal HC, Mittal N, Yadav J, Rani P, Naveen Kumar PG. Outcome of partial pulpotomy verses full pulpotomy using biodentine in vital mature permanent molar with clinical symptoms indicative of irreversible pulpitis: A randomized clinical trial. J Conserv Dent 2022;25:317-23. [Full text]
Ramani A, Sangwan P, Tewari S, Duhan J, Mittal S, Kumar V. Comparative evaluation of complete and partial pulpotomy in mature permanent teeth with symptomatic irreversible pulpitis: A randomized clinical trial. Int Endod J 2022;55:430-40.
Aravind A, Rechithra R, Sharma R, Rana A, Sharma S, Kumar V, et al
. Response to pulp sensibility tests after full pulpotomy in permanent mandibular teeth with symptomatic irreversible pulpitis: A retrospective data analysis. J Endod 2022;48:80-6.
Simon S, Perard M, Zanini M, Smith AJ, Charpentier E, Djole SX, et al
. Should pulp chamber pulpotomy be seen as a permanent treatment? Some preliminary thoughts. Int Endod J 2013;46:79-87.
Jacobson JJ. Probe placement during electric pulp-testing procedures. Oral Surg Oral Med Oral Pathol 1984;58:242-7.
Mickel AK, Lindquist KA, Chogle S, Jones JJ, Curd F. Electric pulp tester conductance through various interface media. J Endod 2006;32:1178-80.
Tchorz JP, Hellwig E, Altenburger MJ. An improved model for teaching use of electronic apex locators. Int Endod J 2012;45:307-10.
Zanini M, Hennequin M, Cousson PY. A review of criteria for the evaluation of pulpotomy outcomes in mature permanent teeth. J Endod 2016;42:1167-74.
DeRosa TA. A retrospective evaluation of pulpotomy as an alternative to extraction. Gen Dent 2006;54:37-40.
Taha NA, Ahmad MB, Ghanim A. Assessment of mineral trioxide aggregate pulpotomy in mature permanent teeth with carious exposures. Int Endod J 2017;50:117-25.
Asgary S, Eghbal MJ, Shahravan A, Saberi E, Baghban AA, Parhizkar A. Outcomes of root canal therapy or full pulpotomy using two endodontic biomaterials in mature permanent teeth: A randomized controlled trial. Clin Oral Investig 2022;26:3287-97.
Alghaithy RA, Qualtrough AJ. Pulp sensibility and vitality tests for diagnosing pulpal health in permanent teeth: A critical review. Int Endod J 2017;50:135-42.
Jafarzadeh H, Abbott PV. Review of pulp sensibility tests. Part II: Electric pulp tests and test cavities. Int Endod J 2010;43:945-58.
Moody AB, Browne RM, Robinson PP. A comparison of monopolar and bipolar electrical stimuli and thermal stimuli in determining the vitality of human teeth. Arch Oral Biol 1989;34:701-5.
Pantera EA Jr., Anderson RW, Pantera CT. Reliability of electric pulp testing after pulpal testing with dichlorodifluoromethane. J Endod 1993;19:312-4.
Careddu R, Duncan HF. A prospective clinical study investigating the effectiveness of partial pulpotomy after relating preoperative symptoms to a new and established classification of pulpitis. Int Endod J 2021;54:2156-72.
Lin J, Chandler NP. Electric pulp testing: A review. Int Endod J 2008;41:365-74.
Inoue H, Muneyuki H, Izumi T, Taguchi K, Nishigawa Y, Watanabe K, et al
. Electron microscopic study on nerve terminals during dentin bridge formation after pulpotomy in dog teeth. J Endod 1997;23:569-71.
Dummer PM, Tanner M. The response of caries-free, unfilled teeth to electrical excitation: A comparison of two new pulp testers. Int Endod J 1986;19:172-7.
Taha NA, Al-Rawash MH, Imran ZA. Outcome of full pulpotomy in mature permanent molars using 3 calcium silicate-based materials: A parallel, double blind, randomized controlled trial. Int Endod J 2022;55:416-29.
Taha NA, Al-Khatib H. 4-Year follow-up of full pulpotomy in symptomatic mature permanent teeth with carious pulp exposure using a stainproof calcium silicate-based material. J Endod 2022;48:87-95.
Borkar SA, Ataide I. Biodentine pulpotomy several days after pulp exposure: Four case reports. J Conserv Dent 2015;18:73-8.
] [Full text]
Parirokh M, Torabinejad M, Dummer PM. Mineral trioxide aggregate and other bioactive endodontic cements: An updated overview – Part I: Vital pulp therapy. Int Endod J 2018;51:177-205.
Gonçalves JL, Viapiana R, Miranda CE, Borges AH, Cruz Filho AM. Evaluation of physico-chemical properties of Portland cements and MTA. Braz Oral Res 2010;24:277-83.
ElKestawy MA, Saafan SA, Shehata MM, Saafan AM. Investigation of the electrical properties of some dental composite restorative materials before and after laser exposure. Dent Mater 2006;22:885-95.
Ikeda M, Oka Y. The relationship between nerve conduction velocity and fiber morphology during peripheral nerve regeneration. Brain Behav 2012;2:382-90.
Chueh LH, Chiang CP. Histology of Irreversible pulpitis premolars treated with mineral trioxide aggregate pulpotomy. Oper Dent 2010;35:370-4.
Dr. Vijay Kumar
Room No. 308, Third Floor, Division of Conservative Dentistry and Endodontics, Centre for Dental Education and Research, All India Institute of Medical Sciences, New Delhi
Source of Support: None, Conflict of Interest: None
[Figure 1], [Figure 2], [Figure 3]
| Article Access Statistics|
| Viewed||974 |
| Printed||102 |
| Emailed||0 |
| PDF Downloaded||69 |
| Comments ||[Add] |