The root surface walls of periodontal pockets harbour bacteria and endotoxins, which hinders the reattachment of gingival and periodontal cells1.Though scaling and root planing are followed conventionally as a part of periodontal therapy, it is not completely effective in removing the diseased cementum.Therefore, chemical root conditioning is advocated to achieve a biologically suitable root surface for epithelial and connective tissue attachment2.
Dentinal collagen exposed by demineralization enhances binding of the extracellular matrix glycoprotein to dentin.3 This increases the attachment and growth of fibroblasts on dentin surfaces.4
` Tetracyclines are one of the effective agents used to enhance periodontal regeneration.5 Root dentin treatment by tetracycline removes the smear layer and exposes the dentin with open tubules.6 It promotes the binding of fibronectin to dentin.The adsorbed fibronectin stimulates fibroblast attachment and growth.Tetracyclines reduce excessive collagenase activity.Also, tetracycline is adsorbed to and subsequently released from dentin, maintaining its antimicrobial activity.7
Tetracycline hydrochloride concentration ranges from 0.5% to 200% and application periods between 0.5 to 10 minutes.8,9 Higher concentrations of tetracycline hydrochloride are required to achieve complete demineralization .8,10 The efficacy of application times and optimal concentration of tetracycline hydrochloride to achieve root surface characteristics to support periodontal reconstructive therapy have yet to be established.11
The purpose of this study is to evaluate and compare surface changes of human root dentin at different Tetracycline hydrochloride concentrations of 0,10,25,50,75,100,125 and 150mg/ml solutions, and also different application times i.e, 1,3 and 5 minutes, using Scanning Electron Microscope.The surface characteristics are compared and the width of the dentinal tubule orifices will be measured.
Periodontally affected single rooted teeth with poor prognosis indicated for extraction.
1)Teeth with root caries.
2)Teeth with root surface demonstrating abrasive lesions.
3)Teeth with root surface showing demineralized areas.
4)Teeth with root surface showing hypercementosis.
5)Teeth with fractured root surface
6)Patients who give history of scaling and root planning in the previous 6
7)Teeth with root restorations.
8)Patients who give history of using desensitizers and mouth washes.
3.Sampling technique: 48 specimen samples will be prepared from the proximal surface of 24
periodontally affected single rooted teeth indicated for extraction.
Prior to specimen preparation,root surface apical to cementoenamel junction
is subjected to thorough scaling and root planning using Gracey curettes
Sectioning procedure: The crown portion of the extracted teeth will be sectioned following the cementoenamel junction as the guideline using high speed diamond bur.Following the separation of the crown from the roots,a longitudinal sectioning will be done at the line marked on the facial and lingual or palatal surface of the roots extending 6mm apically from the CEJ and two proximal halves will be separated with cross section at the termination of 6mm vertical cut.The root blocks will measure individually in 3 dimensions as follows
8 groups for conditioning at tetracycline hydrochloride concentration(weight
per volume) of 0,10,25,50,75,100,125 and 150mg/ml and
- 3 application periods of 1,3, and 5 minutes(i.e, 2 samples per group).
Preparation of Tetracycline hydrochloridesolutions: Fresh tetracycline hydrochloride solutions will be prepared at room temperature by weighing pure tetracycline hydrochloride powder and dissolving it in 20 to 35 ml of distilled water, using a magnetic stirrer to mix the solution to a final volume of 50ml.The concentrations(weight per volume) of tetracycline hydrochloride test solutions are 0,10,25,50,75,100,125,and 150mg/ml.The pH of each solution will be tested with a hand-held, battery operated pH meter and recorded.
The specimens in each group will be conditioned with one of the 8 concentrations and the dentin surface is burnished for 1,3 or 5 minutes with a solution-soaked cotton pellet changed every 30 seconds;the area will then be flushed for 30 seconds with distilled water.
Specimen preparation for SEM: Immediately after applying Tetracycline hydrochloride and rinsing,the dentin specimens are fixed in 2.5% Glutaraldehyde in phosphate buffer (pH 7.3) for 24 hours at 40C, washed three times in same phosphate buffer, post-fixed in phosphate-buffered 1.5% osmium tetroxide for 2 hours,and washed three more times in phosphate buffer.The specimens will then be dehydrated in a graded series of aqueous ethanol solutions(50%,70%,85%,95%,and 100% ethanol) for 10 minutes each.After two more 10- minute washings in absolute ethanol,the samples will then be dried overnight in a desiccator jar.
The specimens thus prepared will be subjected to Scanning Electron Microscopic analysis to evaluate-
Smear layer removal
Morphological changes on the root surface
Opening of dentinal tubules;the width of dentinal tubules for each concentration of Tetracycline hydrochloride solution and for each application time.
4. Statistical analysis The means and standard deviation of diameter of the tubule orifices for each solution is determined for 1,3, and 5 minute application periods.One-Way analysis of variance is used to examine effect of application times and tetracycline hydrochloride concentrations on the width of dentinal tubule orifices.All groups are tested by multiple range test (Tukey test).Paired t-test is performed for each combination of application time and concentration.
List of References
Andriaens PA,De Boever JA,Loesche WJ-Bacterial invasion in root cementum
and radicular dentin of periodontally diseased teeth in humans A reservoir of
J Periodon Res 1986;21:322-329
7) Delazari FMC,Gerlach RF,Joly JC,Lima AFM-Scanning Electron Microscopy Study of the effect of Tetracycline HCl on smear layer removal and fibrin network formation .Braz Dent J (1999)10(2):81-87
8) Hanes PJ,O’Brien NJ,Garnick JJ-A morphological comparison of radicular