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Comparison of Expanded and Nonexpanded Free Soft Tissue Autografts: A Clinical Comparison


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COMPARISON OF EXPANDED AND N ON-EXPANDED FREE SOFT TISSUE AUTOGRAFTS: A CLINICAL COMPARISON By MICHELE PAULA BEATY A THESIS PRESENTED TO THE GRADUATE SCHOOL OF THE UNIVERSITY OF FLOR IDA IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF MASTER OF SCIENCE UNIVERSITY OF FLORIDA 2006

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Copyright 2006 by Michele Paula Beaty

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iii ACKNOWLEDGMENTS I thank my committee members Dr.Hank Towle, Dr. Arthur Vernino, and Dr. Gregory Horning for their guidance through this study. I would like to also thank all of my friends and family for all of their love and support through the last 10 years of my academic career at the University of Florida.

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iv TABLE OF CONTENTS page ACKNOWLEDGMENTS.................................................................................................iii LIST OF FIGURES.............................................................................................................v ABSTRACT....................................................................................................................... vi CHAPTER 1 INTRODUCTION........................................................................................................1 Background...................................................................................................................1 Anatomy.......................................................................................................................2 Graft Harvesting and Wound Healing..........................................................................4 Purpose of the Study.....................................................................................................5 2 MATERIALS AND METHODS.................................................................................8 Inclusion Exclusion Criteria.........................................................................................8 Investigational Device..................................................................................................8 Study Protocol..............................................................................................................9 3 RESULTS...................................................................................................................13 Statistical Analysis......................................................................................................13 Histological Analysis..................................................................................................14 Aaesthetic Analysis....................................................................................................16 4 DISCUSSION.............................................................................................................17 Discussion of Statistical Results.................................................................................17 Discussion of Hist ological Results.............................................................................17 Modification of Device...............................................................................................18 Conclusions.................................................................................................................18 APPENDIX DATA COLLECTION SHEET...................................................................20 LIST OF REFERENCES...................................................................................................21 BIOGRAPHICAL SKETCH.............................................................................................22

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v LIST OF FIGURES Figure page 2-1 Patients with bilateral mucogingival de fects were accepted into the study if they met the inclusion criteria............................................................................................8 2-2 Investigational meshing device was used to perforate and mesh the grafts...............9 2-3 The grafts were affixed to the recipient bed with vicryl suture................................10 2-4 Side-by-side comparison of the non-mesh ed graft on the left versus the meshed graft on the right. Evaluators were aske d to make comparisons of the 6 month post operative results................................................................................................11 3-1 Equally spaced perforati ons are evident in this H&E stain with the perforation extending into the connective tissue.........................................................................15 3-2 Transverse section of a H&E slide of the perforation made by the device..............15

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vi Abstract of Thesis Presen ted to the Graduate School of the University of Florida in Partial Fulfillment of the Requirements for the Degree of Master of Science COMPARISON OF EXPANDED AND N ON-EXPANDED FREE SOFT TISSUE AUTOGRAFTS: A CLINICAL COMPARISON By Michele Paula Beaty May 2006 Chair: Arthur Vernino Major Department: Periodontics Background: Free gingival grafts (FGGs) ha ve been used for over 40 years to create a widened zone of attached gingiva. They were initially described by Bjorn in 1963 and have been extensively investigated and used since. The traditional FGG technique extends the incisions to approxima tely twice the desired width to allow for 50% contraction when healing is complete. Expanding the donor tissue will allow for a more conservative donor site preparation and less post-operative pain experienced by the patient. Prior attempts to expand FGGs have had limited success. Medical techniques and instruments although, have been deve loped to accomplish skin graft expansion consistently. These techniques currently use dermatomes to harvest the skin grafts and tissue meshers to expand the grafts. The tissue expanders allow for expansion of donor tissue in an attempt to gain more coverage fo r burn patients and for patients that require extensive grafting procedures.

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vii Methods: Fifteen patients with bilateral de fects will be used for this study. The study design will be a split mouth design. One side will represent the experimental unit (FGG will be meshed) and the other side of th e mouth will act as a control (a traditional FGG would be employed). The surgical technique would be identical for both sites, and would only differ by the mesh modification made in the donor tissue of the experimental unit. Purpose: The purpose of this study is to i nvestigate the healing effects of in vivo, free gingival grafts that have been modified by a tissue expander in patients with areas of inadequate attached gingiva. The FGG in order to see if less gi ngival donor tissue may be required to obtain satisfactory grafting re sults, and to observe the effects that expansion has on contraction after healing. Observations will also be made about aesthetics and color match. By perforating the FGGs we hope that the recipient tissue cells can profuse the donor tissue in order to help blend the borders of the graft.

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1 CHAPTER 1 INTRODUCTION Background Skin grafts were utilized first by the ancient Egyptians. Some of the first documented reports were by Reverdin in 1869, when he successfully transplanted small thin epidermal grafts, but th e foundation of modern skin gr afting began when Thiersch and Wolfe developed the technique of split-th ickness and full-thickness grafting in 1874. In 1894, the first intrao ral skin grafting pro cedure was completed by Schnitzer and Bjorn; in 1963, was the first to demonstrate the valu e of autogenous free gingival grafts (FGGs) in periodontal therapy. Gingival grafting was initia lly introduced into the United States in 1964 by King and Pennel. Subsequently, Nabers in 1966 re ported gingival grafti ng as beneficial for vestibular extension and repor ted that grafting could achieve a gain in attached tissue along with root coverage of previously denuded roots. With over 130 years of successful autogenous soft tissue grafting in the medical and dental fields, FGGs have become a rout inely and successfully used procedure to: 1) create an adequate zone of attached gingiva in areas wher e periodontal probing depths are close to or extend apical to the mucogingiva l junction, 2) to eliminate frena and muscle attachments that encroach upon the periodontal apparatus, 3) to co rrect recession causing denudation of root surfaces, 4) to deepen the ve stibule, and 5) to adequately deal with anatomic factors of thin alveolar hous ing, tooth position, prominent tooth roots, and other situations which predispose teeth to gingival recession.

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2 Controversy exists as to the minimum width of attached tissue that is necessary for gingival health. Prior studies have suggested that a total of 2 mm of keratinized tissue is required for periodontal health with 1 mm of attached tissue and 1 mm of unattached tissue. 1 Other studies have shown that patients with excellent oral hygiene may maintain healthy areas with no attached tissue, while Bo wers stated that for periodontal health a minimum amount of attached tissue is n ecessary but no exact dimension was given. 2, 3 Anatomy Gingival grafts are harvested from the pati ent’s maxillary palate typically, but other possible donor sites may include keratinized tissue from the tuberosity region or an edentulous ridge. Acceptable donor tissue is ch aracterized histologically by a keratinized or a parakeratinized epithelium and a dense lamina propria. Palatal tissue epithelium has been shown to range in thickness of 0.1 mm to 0.6 mm and is influenced by denture use, smoking or low grade inflammation .4 While palatal tissue is the most commonly used donor tissue for the harvesting of a FGG, it has anatomical limitations. The posteri or palate is the lo cation of the Greater Palatine artery, nerve, and accompanying vessels The foramen is typically located in the vicinity of the third molar 15 mm from th e midline and 4.8 mm from the most lateral point of the posterior border of the hard pa late. The Greater Palatine Nerve and vessels emerge through this foramen and run anteriorly in the submucosa of the palate between the palatal and alveolar proce sses. The location of these vital structures may limit the surgical access for harvesting donor tissue. The submucosa of the palate posterior to the first molars contains the majority of the minor salivary glands, which are more compact in the soft palate and extend anteriorly into the hard palate wher e they decrease in

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3 quantity. These glands occur between the mu cosal connective tissue and the periosteum protecting the underlying vessels and nerves. Palatal tissue harvested from the tuberosity region yields significantly thicker grafts averaging over 5.4 mm. The thickness of tissue in the first molar vicinity is usually 1.8 mm and increases as the first premolar is approached. The tissue thickness in the first premolar area averages 3.9 mm, and the tissue wi ll also increase in thickness as grafts are harvested, further away from the margins of th e crowns of the teeth, thus thicker grafts can be harvested if grafts are obtained several millimeters away from the gingival margin of teeth. The thinnest tissue occurs 1–8 mm from the gingival margin of the first molar, hence the palatal root of the first molar represents an anatomical barrier. 5 The palate has other limitations, including palatal rugae that begins around the mesial of the first pre-molar. Studies have shown that the underlying lamina propria contains genetic pre-determinants for the specific character of the overlying epithelium and its structure. 6 Hence, incorporating donor tissue th at includes these rugae will result in the transfer of the tissue topography to the recipient site Attempts to surgically remove rugae by blades and burs are usua lly not successful, a nd aesthetics can be compromised when the rugae return. Anothe r limitation of palata l donor tissue is the underlying submucosa which can have a variab le amount of adipose tissue. Adipose tissue can act as a diffusion and va scularization barrier if it is included in the palatal graft, and care must be taken to remove it with a su rgical blade before the graft is stabilized onto the recipient site. Adipose tissue in the ha rd palate increases in quantity anterior to the first premolar, limiting the extent of available donor tissue, in this area.

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4 Graft Harvesting and Wound Healing Harvested grafts are classified into full or split-thickness grafts. A full-thickness graft refers to the inclusion of all of the lamina propria, while the split-thickness graft includes only a partial amount. FGGs that ar e harvested can be either thickness because of the challenges of tissue contour, poor access due to curvature of the palate, proximity of the teeth, and instrument design. The pr oper thickness of the graft can be important for graft survival. The graft should be thin enough to permit diffusion of nutritive fluid from the recipient site; but if it is too thi n, it may necrose and e xpose the recipient site. 7 The ideal thickness of a graft has been shown to be 1.0–1.5 mm. 8 Epithelial tissue lacks the presence of blood vessels and it must derive its vascularization and exchange of metabolites and waste solely by diffusion. When grafts are transplanted, its epithelium and lamina propria will maintain its viability through diffusion from the recipient bed in the initial st ages of healing. The lamina propria of the graft hinders diffusion; hence, the thinner the graft the more easily it can be maintained by diffusion and can be revascularized. Thi nner grafts (0.75 mm) have been shown to heal more rapidly than thicker grafts (>1.75 mm ). Microscopically, th in grafts complete their healing at 10.5 weeks and thicker grafts require 16 weeks or longer. 9 The process of graft healing occurs in th ree stages: plasmatic circulation stage, capillary circulation stage, and the organization stage. Fluorescein angiography studies have shown blood recirculation in grafts to be first established through capillary “budding” and are maintained for the firs t 2–3 days by plasmatic circulation which provides graft sustenance. A formation of a regular pattern of capillary loops will begin on the seventh day and is followed by complete revascularization on the ninth to fourteenth day. 10

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5 The thickness of the graft can influence it s behavior during the healing process and the final results. Thicker grafts, because th ey contain more elastin, will undergo more immediate contracture when the graft is rem oved from the donor site. This shrinkage is termed primary shrinkage since it occurs befo re the healing process. This higher amount of elastin in thick grafts has shown that contracture of as much as 43% can occur. Thinner grafts with less connective tissue wi ll have less primary shrinkage but will experience more secondary contra cture termed “cicatrization”. 11 Secondary contracture is caused by the scarring or fibrosis of the tissue that unites the graft with its recipient base. The effect of this cicatrization on the graft is dependant upon the rigidity of the recipient bed and the thickness of th e lamina propria of the graft. The traditional FGG technique extends incisions for the donor harvest site to approximately twice the desired width allo wing for initial shrinka ge after harvesting allowing for a 50% graft contraction with fina l healing. Grafts can either be placed on denuded bone or on periosteum. Grafts when placed on denuded bone have shown to have less post-operative mobility, le ss swelling, and better hemostasis. 12 Several biometric studies analyzed that grafts pl aced on denuded bone shrink 25% versus grafts placed on a periosteal bed shrink 50% after a 24-week healing period. The greatest amount of shrinkage typically o ccurs during the firs t 6 weeks, however a healing lag of 2 weeks is observed when graf ts are placed on denuded bone. 13, 14 Purpose of the Study Disadvantages of FGGs not only include shrinkage but also present a potential challenge. FGGs have been known to heal in a “postage stamp” fashion since the donor tissue color may not match the r ecipient tissue. This color disparity leads to an obvious

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6 demarcation of where the FGG was placed and a potentially unaesthetic result. No prior attempts to date have been made to improve the aes thetics of FGGs. Pervious attempts have been made to expand FGGs, but none have been truly successful. Rateitschak create d a technique that was termed the “accordion technique”. 15 This technique involves trying to gain expans ion by making incisions by hand in alternate sides of the graft using a scal pel blade. These alternating incisions attempted to gain expansion, but limited expansion is actually achieved. The accuracy and reproducibility of these incisions is suspected to be the limitation to expansion. This study hopes to determine a true and predictable expansion ratio that a tissue expander may achieve with donor tissue. The ability to expand FGGs would allow for smaller donor tissue to be harvested, less post-operative pain for the patient, and increased coverage from the donor tissue. We al so expect that the perforations made in our donor tissue will allow for recipient tissu e to perfuse the FGG, allowing for better aesthetics and eliminating the “postage” like appearance. This study applies the techniques already used in modern medical grafting to dental procedures in order to improve surgical tr eatment for dental patients. The medical profession has been using devices to mesh sk in grafts since the 1960s in order to expand donor grafts for large surgical skin defects as those associat ed with burn patients. Their investigations have proven that meshed gr afts are both safe and effective. Medical research has shown that the minimal thickness fo r a skin graft to remain viable and grow is 1.02 mm. 6 Our research utilized grafts be tween 1.0– 2.0 mm in thickness. Zimmer, Padgett, and Brennen are medical corporations marketing skin graft meshers that have the ability of producing expansion ratios of up to 4:1. The grafts used in this study were

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7 thicker in nature and required less expansion than what is required in medical applications; hence, the meshing of our grafts were done in a more conservative nature. The medical specialties have demonstrated th at perforating a graf t improves the graft’s viability. Regardless, if a sk in graft is intended to be e xpanded the medical protocol still requires perforation of the graf t by this meshing process. Th is perforation is completed on every graft in order to prevent hematomas or seromas, and ultimately graft acceptance. The purpose of our study was to employ the standard surgical technique used for dental grafting procedures, but to incorporate some of the principles of perforation and expansion as utilized with skin grafting in or der to determine an expansion ratio, aesthetic improvements, and to determine the amount of gain in keratinized ti ssue. Expansion of the graft was attempted with a custom device that is not yet available commercially. The device was designed to bring the benefits of skin grafting to intraoral tissue grafting.

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8 CHAPTER 2 MATERIALS AND METHODS Inclusion Exclusion Criteria Patients from the University Of Florida College Of Dentistry in Gainesville, Florida will be recruited for this study a nd will be examined for their levels of periodontitis, recession and amounts of keratinized tissue. The inclusion criteria are medically healthy patients between the ages of 16–65 with a dia gnosis of insufficient width of keratinized gingiva or other mucogingival defects that would benefit from an increase in keratinized or attached tissue. Pa tients were excluded from the study if they had spontaneous bleeding upon probi ng, suppuration, active infection were pregnant, or had any other severe concurrent systemic diseases that may im pact on the periodontal condition and wound healing. Figure 2-1. Patients with bilateral mucogingi val defects were accepte d into the study if they met the inclusion criteria. Investigational Device The prototype instrument was manufactur ed by Champagne designs in West Palm Beach, Florida. The instrument was modeled after the medical meshers that are currently

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9 available for skin graft meshing. The instru ment has a rolling drum with cylindrical cones as the perforation mechanism. The cones are symmetrically spaced 1.0 mm apart with the base of the cones measuring 2 mms in diameter. The instrument was designed to be rolled over the free gingival graft (FGG) graft in a length wise direction on the outer epithelial side. The graft is held on a poly carboxylte cutting board with the edge of a periosteal elevator. The rolli ng action of the mesher is meant to perforate and mesh the graft symmetrically. Figure 2-2. Investigational meshing device wa s used to perforate and mesh the grafts. Study Protocol Thirteen healthy patients with bilateral mucogingival defects were used for this study. The study design was a split mouth design. One side chosen at random represented the experimental unit and the FGG on that side was meshed. The contralateral side acted as the control unit with a traditional FGG technique employed and the graft was not meshed. The surgical technique would be identical fo r both sites and would only differ by the mesh modification made in the donor tissue on the experimental side. The instrument would be rolled across the outer epithelial surface of the experimental graft in a mesial-distal direction and a ll other procedures would be identical. Patients included in this study were required to have five visits. The initial visit included an explanation of th e purpose of the study and the signing of informed consent forms. Initial measurements of the mucogingiva l defects were recorded to include: length

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10 and width in millimeters, and the amount keratinized gingival tissue. A Miller Classification was determined for each site, probing depths bleeding on probing, mobility, and furcation involvement was also not ed. The second surgical visit consisted of pre-operative photographs to document the patient’s mucogingival defect before the surgery. Patients rinsed with a 60-second pre-operative chlo rhexidine 0.12% rinse. The recipient bed was created in the standard split-thickness method, a nd the involved root surface was prepared with hand instrumentati on and Pref Gel was applied to the root surface for a minimum of 3 minutes. Pref Gel is a neutral acid that is safe on the tissues. The purpose of this acid is to expose collagen fi bers to allow for attachment of the soft tissue grafts to the surfaces of the roots. The donor tissue was then harvested from the patient’s palate and the thickness ranged 0.75–2.00 mm. Grafts were considered “thin” if they were less than 1.0mm, “average” if they were 1.0–1.5 mm, and “thick” if they exceeded 1.5mm. The sizes of the harvested grafts were documented in length and width in millimeters and the expanded grafts were also measured post-expansion for length and width in millimeters. The harvested graft fo r both the control and experimental unit was affixed to recipient site with 5.0 vicryl suture using a p-3 needle. Surgical photos were taken documenting each step of the procedure. Figure 2-3. The grafts were affixed to the recipient bed w ith vicryl suture.

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11 One graft that was harvested and expanded had a 3 x 4 mm sample removed for histological analysis. Statistical analysis was completed by using both a t-test and a signed ranked t-test, since our population sample was small (n=13). P values < 0.05 were considered significant. The subsequent post-operative visits occurred 10 days and 4 weeks postoperatively to document the healing stages a nd to enforce oral hygi ene. Patients were placed on a chlorhexidine rinse 0.12% to aid in soft tissue healing and inflammation reduction for duration of 4 weeks from the day of surgery. Motrin 800mg was prescribed and patients were instructed to take 3 200 mgs per day for control of discomfort. Figure 2-4. Side-by-side comparison of th e non-meshed graft on the left versus the meshed graft on the right. Evaluators we re asked to make comparisons of the 6-month post-operative results. The final visit occurred at 6 months post surgery and final photographs were taken in order to determine the final aesthetic result. The aesthetic comparisons were completed by three board certified periodontists, 3 periodontal resident s, and 3 dental students. A computer presentation was created with before and after photographs of each case. The evaluators were given a questionnaire and were instructed on how to complete the comparison (Appendix A). The presentation of the cases included side by side comparisons of the experimental graft next to the control. The two treatments were randomly assigned a letter A or B and the evalua tors were asked to judge each graft for 1)

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12 graft detectibility 2) graft color match to su rrounding tissue and 3) aes thetics of the graft being superior for A or B or if they are similar.

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13 CHAPTER 3 RESULTS Statistical Analysis Of the 13 patients recruited, all complete d the study. Healing was uneventful for all patients. The study populati on consisted of nine female s and three males with the mean age of 52. A total number of 17 mucogingival defects were located in the mandibular premolar region, 3 in the mandibul ar canine region, and 6 in the mandibular anterior region. The data collection sheets we re reviewed and statistical analysis was completed by the University Of Florida Biostatistics Department. Both a t-test and a signed t-test were conducted (n=13) for length, width, gender, and thickness of the grafts to determine statistical significance (p<0.05) related to expansion. No difference in gender was found in the study. Pre-meshing and postmeshing measurements were rounded to the ne arest 0.5mm to determine if there was any statically significant difference in length and width after the meshing was completed. The mean change in length was calculate d at 0.65mm and a standard deviation of 0.66mm. The t-test for length expansion wa s statistically signifi cant (p=0.0038) and the signed-rank test was also stat istically significant (p=0.0078). The mean change in width was calculated at 0.04 mm and a standard deviation of 0.14mm. No statistical significance was found with either the t-test or the signed-rank ed t-test for a change in expansion of width pre-meshing and post-mesh ing. When considering the thickness of the grafts harvested, no statistical difference was calculated between thin, average, or thick grafts.

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14 Table 3-1. Statistical analysis parameters. Subject Length Before Length After Width Before Width After Graft Thickness M/F 1 13.5 13.5 8.0 8.0 1.5 thick F 2 10.5 11.5 7.0 7.0 1.5 thick M 3 11.5 11.5 7.5 7.5 1.5 thick F 4 25.5 27.0 8.0 8.0 1.5 thick M 5 12.0 12.0 9.5 9.5 1.5 thick F 6 15.0 15.0 7.0 7.0 1.5 thick M 7 15.0 15.5 5.0 5.0 0.75 thin F 8 18.0 20.0 4.0 4.0 0.75 thin F 9 21.0 22.0 7.5 7.5 1.0 average F 10 30.0 30.0 5.0 5.0 0.75 thin M 11 17.0 17.5 10.0 10.0 1.5 thick F 12 17.0 18.0 10.0 10.5 1.5 thick F 13 13.0 14.0 8.5 8.5 1.0 average F Histological Analysis A histology sample was sent to the Un iversity Of Florida Department Of Pathology. The tissue sample was first fixe d in 10% neutral buffered formalin. The tissue was processed on Leica tissue pro cessor followed by dehydration with serial alcohol intervals at 80%, 90%, 100% concen trated alcohol and followed by xylene to clear the sample. The samples were placed in paraffin and the sample was embedded top down in order to visualize the surface penetr ations from the mesher. The paraffin block was cut into 5 m sections with a Micron m achine. Leica Auto Stainer XL was used to complete the Hemolysin & Eosin stain. Histologic analysis was completed by light microscopy using a Leica microscope at 40 magnification. Photomicrographs were captured using the Q capture Pro program. Histological analysis revealed intact pa rakeratinized epithelium with alternating perforations equally distanced from each othe r. The perforations were seen in cross-

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15 section and they completely incised thr ough the epithelium and partially through the connective tissue. Figure 3-1. Equally spaced perforations are evident in this H&E stain with the perforation extending into the connective tissue. Determination of whether the incisions tr ansversed completely through the entire thickness of the graft could not be complete d through histology sin ce the sample was in cross-section, the entire pe rforation from epithelium through connective tissue may not have been captured in the 5m slice. Alte rnate transverse sections were completed and histology revealed consistently spaced perf orations through the connective tissue. No signs of incisions or tearing could be noted from the rolling of the drum over the graft, only perforations were detected. Figure 3-2. Transverse secti on of a H&E slide of the perf oration made by the device.

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16 Aesthetic Analysis The aesthetic evaluation was completed by a panel of evaluators. As previously described for graft detectibilit y, the meshed graft was chosen 23.1% of the time as being “non-detectible”, 53.8% for “above averag e” blend, 23.1% of the time as having “average” blend, and 0% as having “poor” blend. The control was chosen 15.4% as “non-detectible”, 30.8% “above average” bl end, 53.8% “average” blend, and 0% chosen as having “poor” blend. Evalua tors were also asked to examine the photos for color march to the surrounding tissues. The meshed graft was chosen 46.1% as an “excellent” match, 38.5% “above average”, 15.4% “average”, and 0% as “poor” color match to the surrounding tissues. The control graft was chosen 15.4% as “excellent”, 53.8% as “above average”, 30.8% as “average”, and 0% as a “poor”. Evaluators were also asked to choose which graft appeared more aesthetic when the experimental unit was compared side-by-side to the control. The meshed gr aft was chosen as bei ng superior in 46.1% of the cases, and no difference in aesthetics wa s determined in 38.5% of the photos when the meshed side was compared to the non-me shed side. The control, non-meshed side was chosen as being superior than th e meshed side in 15.4% of the cases.

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17 CHAPTER 4 DISCUSSION Discussion of Statistical Results Evaluation of the statistical results yielde d a mean expansion in length of 0.63mm. The mean expansion of width was calculated only at 0.14mm When considering the statistical significance versus the clinical si gnificance, 0.63mm of expansion is clinically insignificant. One must also take into c onsideration the error in measurements. The harvested grafts were not pe rfectly rectangular and someti mes had irregular proportions due to the difficulty in harvesting. The measurements were made as accurate as possible with a ruler and a probe to th e nearest .05mm, but inherent error in measurements could have occurred. Discussion of Histological Results A review of the histologic sample did re veal successful perf oration of the device though the epithelium and into the underl ying connective tissue (Fig. 3-1). The perforations were equally spaced, but the samp le was taken in cross-section, and it could not be determined if the perforation extended th rough the entire width of the graft. It is possible that the cross-secti onal 5m cut did not include th e entire incision made by the instrument and only the surface perforation wa s included in the slice. A transverse sample was then analyzed and equally spaced perforations were detected in the deeper layers of the graft (Fig. 3-1). Visual exam ination during the meshing of the grafts did reveal that complete perforation was achie ved by the instrument, since the cones were able to be seen entering and exiting the graft.

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18 Modification of Device Since histological evaluation only rev ealed a perforating effect from our instrument, it is believed that modifications to the instrument could improve the possible expansion achieved in free gingi val grafts (FGGs). Dr. Neel Bhatavadekar was consulted regarding making modifications to the prototype in order to improve expansion ratios. Dr. Bhatavadekar has recently just complete d some research rega rding the development of a newly designed dermatome and improved skin meshing device. His work was completed at the University of Florida in th e Department of Biomedical Engineering. Consultation with him suggested that a rede sign of the device to incorporate a motorized or hand-crank mechanism in order to provide positive pressure to the graft during the meshing process. This positive pressure would allow for an even and controlled meshing to occur. A suggestion was also made to modi fy the perforating cones. The layout of the cones was determined to be appropriate but a suggestion to modify th e cones into surgical cutting blades. These alternating blades w ould provide a cutting effect instead of a perforating effect. The cutting e ffect of the blades would allo w for better expansion to be achieved. Conclusions Aesthetic results of the meshed versus the traditional non-meshed graft were compared by a panel of evaluators. Overall, the meshed graft was chosen 46.1% of the time to be superior in overall aesthetics, but 38.5% of the time no di fference in aesthetics could be determined between the two sites. The control was actua lly chosen to more aesthetic than the meshed site in 15.4% of the cases. Alternative uses for this device were consid ered, since it is succe ssful at perforation of the grafts consistently. Th e perforations could be used as a carrier for either proteins

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19 or amelogins. Combing these healing enhanci ng mediators within the grafts perforations could act as a carrier for these substances. Since root coverage with FGGs is not as predictable as connec tive tissue grafts, the addition of these substances may allow for better root coverage by increasing attachment to the root surface. It would then be possible to gain both keratinized tissue and r oot coverage by a single surgical procedure. In conclusion, the device seemed to have improved the aesthetic appearance of the graft in 46.1% of the cases, but clinically the expansion wa s non-significant. Future research could be conducted by modifying the mesher in order to improve the expansion ratio, or keeping the current design and using th e perforations as a ca rrier for proteins to improve soft-tissue procedures.

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20 APPENDIX DATA COLLECTION SHEET Please Circle or Highlight one of the Choi ces Below for Comparison of Photo A to B Please Fill in the Case Number #1-12 Case# ________ Graft Detectibility: Graft A: Graft is not Detectible (excellent blend) Above Average Blend of Graft Average Blend of Graft Poor Blend of Graft Graft B: Graft is not Detectible (excellent blend) Above Average Blend of Graft Average Blend of Graft Poor Blend of Graft Graft Color Match to Surrounding Tissue: Graft A: Excellent Above Average Average Poor Graft B: Excellent Above Average Average Poor Aesthetics of The Graft is Superior For: Graft A Graft B Neither, Grafts ha ve similar Aesthetic Appearance

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21 LIST OF REFERENCES 1. Lang NP, Loe H. The relationship between the width of keratinized gingiva and gingival health. J Pe riodontol 1972; 43:623-7. 2. Wennstrom J, Lindhe J. Role of attached gingiva for maintenance of periodontal health. Healing following excisional and grafting procedures in dogs. J Clin Periodontol 1983; 10:206-21. 3. Wennstrom JL. Role of attached gingiv a for maintenance of periodontal health. Healing following excisional and grafting procedures in dogs. J Clin Periodontol 1983 Mar; 10(2):206-21. 4. Soehren S, Allen A, Cutright D. Clini cal and histologic stud ies of donor tissues utilized for free grafts of masticat ory mucosa. J Periodontol 1973; 12:727-41. 5. Studer S, Allen E, Rees T. The thickness of masticatory mucosa in the human hard palate and tuberosity as potential donor sites for ridge augmentation procedures. J Periodontol 1997; 68: 145-151. 6. Karring T, Loe H. Conservation of tis sue specificity after hetertropic transplantation of gingiva and alve olar mucosa. J Perio Res 1971; 6:282. 7. Pennel BM, Tarbor JC, King KO. Free ma sticatory mucosa grafts. J Periodontol 1969; 40:162. 8. Mormann W, Schaer F, Firestone AC. The relationship between success of free gingival grafts and transplant th ickness. J Periodontol 1981; 52:74. 9. Gordon HP, Sullivan HC. Free autogenous gingival grafts. Part II, supplemental findingshistology of the graf t site. Periodontics 1968; 6:130. 10. Mormann W, Bernimoulin JP, Schmid MO. J Clin Periodontol 1975; 2:177-189. 11. James WC, McFall WR Jr. Placement of fr ee gingival grafts on denuded alveolar bone. Part I: Clinical Evalua tions. J Periodont ol 1978; 49:283. 12. Donnenfeld OW, Marks R, Glickman M. Th e apically positioned flap: a clinical study. J Periodontol 1964; 35:381. 13. Matter J, Cimasoni G. Creeping reatt achment after free gingival grafts. J Periodontol 1976; 47:574.

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22 BIOGRAPHICAL SKETCH Michele P. Beaty was born and raised in Boca Raton, FL. She graduated with a Bachelor in Science in 1999 from the Univer sity of Florida. She continued her postgraduate education at the Univ ersity Of Florida College Of Dentistry. In 2003 she earned her Doctorate of Dental Medi cine and began a residency in the Graduate Periodontics Department at the University Of Florida. Sh e currently is in private practice specializing in Periodontics and Implant Dentistry.


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Permanent Link: http://ufdc.ufl.edu/UFE0014311/00001

Material Information

Title: Comparison of Expanded and Nonexpanded Free Soft Tissue Autografts: A Clinical Comparison
Physical Description: Mixed Material
Copyright Date: 2008

Record Information

Source Institution: University of Florida
Holding Location: University of Florida
Rights Management: All rights reserved by the source institution and holding location.
System ID: UFE0014311:00001

Permanent Link: http://ufdc.ufl.edu/UFE0014311/00001

Material Information

Title: Comparison of Expanded and Nonexpanded Free Soft Tissue Autografts: A Clinical Comparison
Physical Description: Mixed Material
Copyright Date: 2008

Record Information

Source Institution: University of Florida
Holding Location: University of Florida
Rights Management: All rights reserved by the source institution and holding location.
System ID: UFE0014311:00001


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Full Text












COMPARISON OF EXPANDED AND NON-EXPANDED FREE SOFT TISSUE
AUTOGRAFTS: A CLINICAL COMPARISON
















By

MICHELE PAULA BEATY


A THESIS PRESENTED TO THE GRADUATE SCHOOL
OF THE UNIVERSITY OF FLORIDA IN PARTIAL FULFILLMENT
OF THE REQUIREMENTS FOR THE DEGREE OF
MASTER OF SCIENCE

UNIVERSITY OF FLORIDA


2006

































Copyright 2006

by

Michele Paula Beaty















ACKNOWLEDGMENTS

I thank my committee members Dr.Hank Towle, Dr. Arthur Vernino, and Dr.

Gregory Horning for their guidance through this study. I would like to also thank all of

my friends and family for all of their love and support through the last 10 years of my

academic career at the University of Florida.















TABLE OF CONTENTS

page

A C K N O W L E D G M E N T S ......... .................................................................................... iii

LIST OF FIGURES ................................. ...... ... ................. .v

ABSTRACT .............. ......................................... vi

CHAPTER

1 IN TR OD U CTION ............................................... .. ......................... ..

B background ............................................................... .. ........ ...............
A n atom y ................................................................ ................ .. 2
Graft Harvesting and W ound Healing ........................................ ...................... 4
Purpose of the Study ............... ............... .................................. .5

2 M ATERIALS AND M ETHOD S ........................................... ........... ............... 8

Inclusion E exclusion C riteria ............................................................................. 8
Investigational D vice ....................................................... 8
Study Protocol ...................... ........ .. ... .... ............... ....

3 R E S U L T S ........................................................................................................1 3

Statistical A nalysis................................................... 13
H istological A nalysis................................................... 14
A aesthetic A naly sis .............................................................. 16

4 D ISC U SSIO N ...................................................... 17

D discussion of Statistical R results .......................................................... .. .....17
D discussion of H istological R results ................................................................... 17
M odification of D ev ice ......................................................................................... 18
C on clu sion s..................................................... 18

APPENDIX DATA COLLECTION SHEET .......................... ........ ...............20

LIST OF REFERENCES ..................................... .............. ....................21

BIOGRAPHICAL SKETCH .................................................................22
















LIST OF FIGURES


Figure page

2-1 Patients with bilateral mucogingival defects were accepted into the study if they
m et the inclusion criteria .......................................................... .............8

2-2 Investigational meshing device was used to perforate and mesh the grafts ...........9

2-3 The grafts were affixed to the recipient bed with vicryl suture..............................10

2-4 Side-by-side comparison of the non-meshed graft on the left versus the meshed
graft on the right. Evaluators were asked to make comparisons of the 6 month
post operative results .................. ............................. .. ...... .. ........ .... 11

3-1 Equally spaced perforations are evident in this H&E stain with the perforation
extending into the connective tissue................ .......................... ............... 15

3-2 Transverse section of a H&E slide of the perforation made by the device .............15















Abstract of Thesis Presented to the Graduate School
of the University of Florida in Partial Fulfillment of the
Requirements for the Degree of Master of Science

COMPARISON OF EXPANDED AND NON-EXPANDED FREE SOFT TISSUE
AUTOGRAFTS: A CLINICAL COMPARISON

By

Michele Paula Beaty

May 2006

Chair: Arthur Vemino
Major Department: Periodontics

Background: Free gingival grafts (FGGs) have been used for over 40 years to

create a widened zone of attached gingiva. They were initially described by Bjorn in

1963 and have been extensively investigated and used since. The traditional FGG

technique extends the incisions to approximately twice the desired width to allow for

50% contraction when healing is complete. Expanding the donor tissue will allow for a

more conservative donor site preparation and less post-operative pain experienced by the

patient. Prior attempts to expand FGGs have had limited success. Medical techniques

and instruments although, have been developed to accomplish skin graft expansion

consistently. These techniques currently use dermatomes to harvest the skin grafts and

tissue meshers to expand the grafts. The tissue expanders allow for expansion of donor

tissue in an attempt to gain more coverage for burn patients and for patients that require

extensive grafting procedures.









Methods: Fifteen patients with bilateral defects will be used for this study. The

study design will be a split mouth design. One side will represent the experimental unit

(FGG will be meshed) and the other side of the mouth will act as a control (a traditional

FGG would be employed). The surgical technique would be identical for both sites, and

would only differ by the mesh modification made in the donor tissue of the experimental

unit.

Purpose: The purpose of this study is to investigate the healing effects of in vivo,

free gingival grafts that have been modified by a tissue expander in patients with areas of

inadequate attached gingiva. The FGG in order to see if less gingival donor tissue may

be required to obtain satisfactory grafting results, and to observe the effects that

expansion has on contraction after healing. Observations will also be made about

aesthetics and color match. By perforating the FGGs we hope that the recipient tissue

cells can profuse the donor tissue in order to help blend the borders of the graft.














CHAPTER 1
INTRODUCTION

Background

Skin grafts were utilized first by the ancient Egyptians. Some of the first

documented reports were by Reverdin in 1869, when he successfully transplanted small

thin epidermal grafts, but the foundation of modern skin grafting began when Thiersch

and Wolfe developed the technique of split-thickness and full-thickness grafting in 1874.

In 1894, the first intraoral skin grafting procedure was completed by Schnitzer and Bjorn;

in 1963, was the first to demonstrate the value of autogenous free gingival grafts (FGGs)

in periodontal therapy.

Gingival grafting was initially introduced into the United States in 1964 by King

and Pennel. Subsequently, Nabers in 1966 reported gingival grafting as beneficial for

vestibular extension and reported that grafting could achieve a gain in attached tissue

along with root coverage of previously denuded roots.

With over 130 years of successful autogenous soft tissue grafting in the medical

and dental fields, FGGs have become a routinely and successfully used procedure to: 1)

create an adequate zone of attached gingiva in areas where periodontal probing depths are

close to or extend apical to the mucogingival junction, 2) to eliminate frena and muscle

attachments that encroach upon the periodontal apparatus, 3) to correct recession causing

denudation of root surfaces, 4) to deepen the vestibule, and 5) to adequately deal with

anatomic factors of thin alveolar housing, tooth position, prominent tooth roots, and

other situations which predispose teeth to gingival recession.









Controversy exists as to the minimum width of attached tissue that is necessary for

gingival health. Prior studies have suggested that a total of 2 mm of keratinized tissue is

required for periodontal health with 1 mm of attached tissue and 1 mm of unattached

tissue. 1 Other studies have shown that patients with excellent oral hygiene may maintain

healthy areas with no attached tissue, while Bowers stated that for periodontal health a

minimum amount of attached tissue is necessary but no exact dimension was given. 2, 3

Anatomy

Gingival grafts are harvested from the patient's maxillary palate typically, but other

possible donor sites may include keratinized tissue from the tuberosity region or an

edentulous ridge. Acceptable donor tissue is characterized histologically by a keratinized

or a parakeratinized epithelium and a dense lamina propria. Palatal tissue epithelium has

been shown to range in thickness of 0.1 mm to 0.6 mm and is influenced by denture use,

smoking or low grade inflammation .4

While palatal tissue is the most commonly used donor tissue for the harvesting of a

FGG, it has anatomical limitations. The posterior palate is the location of the Greater

Palatine artery, nerve, and accompanying vessels. The foramen is typically located in the

vicinity of the third molar 15 mm from the midline and 4.8 mm from the most lateral

point of the posterior border of the hard palate. The Greater Palatine Nerve and vessels

emerge through this foramen and run anteriorly in the submucosa of the palate between

the palatal and alveolar processes. The location of these vital structures may limit the

surgical access for harvesting donor tissue. The submucosa of the palate posterior to the

first molars contains the majority of the minor salivary glands, which are more compact

in the soft palate and extend anteriorly into the hard palate where they decrease in









quantity. These glands occur between the mucosal connective tissue and the periosteum

protecting the underlying vessels and nerves.

Palatal tissue harvested from the tuberosity region yields significantly thicker grafts

averaging over 5.4 mm. The thickness of tissue in the first molar vicinity is usually 1.8

mm and increases as the first premolar is approached. The tissue thickness in the first

premolar area averages 3.9 mm, and the tissue will also increase in thickness as grafts are

harvested, further away from the margins of the crowns of the teeth, thus thicker grafts

can be harvested if grafts are obtained several millimeters away from the gingival margin

of teeth. The thinnest tissue occurs 1-8 mm from the gingival margin of the first molar,

hence the palatal root of the first molar represents an anatomical barrier.

The palate has other limitations, including palatal rugae that begins around the

mesial of the first pre-molar. Studies have shown that the underlying lamina propria

contains genetic pre-determinants for the specific character of the overlying epithelium

and its structure. 6 Hence, incorporating donor tissue that includes these rugae will result

in the transfer of the tissue topography to the recipient site. Attempts to surgically

remove rugae by blades and burs are usually not successful, and aesthetics can be

compromised when the rugae return. Another limitation of palatal donor tissue is the

underlying submucosa which can have a variable amount of adipose tissue. Adipose

tissue can act as a diffusion and vascularization barrier if it is included in the palatal graft,

and care must be taken to remove it with a surgical blade before the graft is stabilized

onto the recipient site. Adipose tissue in the hard palate increases in quantity anterior to

the first premolar, limiting the extent of available donor tissue, in this area.









Graft Harvesting and Wound Healing

Harvested grafts are classified into full or split-thickness grafts. A full-thickness

graft refers to the inclusion of all of the lamina propria, while the split-thickness graft

includes only a partial amount. FGGs that are harvested can be either thickness because

of the challenges of tissue contour, poor access due to curvature of the palate, proximity

of the teeth, and instrument design. The proper thickness of the graft can be important

for graft survival. The graft should be thin enough to permit diffusion of nutritive fluid

from the recipient site; but if it is too thin, it may necrose and expose the recipient site. 7

The ideal thickness of a graft has been shown to be 1.0-1.5 mm.8

Epithelial tissue lacks the presence of blood vessels and it must derive its

vascularization and exchange of metabolites and waste solely by diffusion. When grafts

are transplanted, its epithelium and lamina propria will maintain its viability through

diffusion from the recipient bed in the initial stages of healing. The lamina propria of the

graft hinders diffusion; hence, the thinner the graft the more easily it can be maintained

by diffusion and can be revascularized. Thinner grafts (0.75 mm) have been shown to

heal more rapidly than thicker grafts (>1.75 mm). Microscopically, thin grafts complete

their healing at 10.5 weeks and thicker grafts require 16 weeks or longer. 9

The process of graft healing occurs in three stages: plasmatic circulation stage,

capillary circulation stage, and the organization stage. Fluorescein angiography studies

have shown blood recirculation in grafts to be first established through capillary

"budding" and are maintained for the first 2-3 days by plasmatic circulation which

provides graft sustenance. A formation of a regular pattern of capillary loops will begin

on the seventh day and is followed by complete revascularization on the ninth to

fourteenth day. 10









The thickness of the graft can influence its behavior during the healing process and

the final results. Thicker grafts, because they contain more elastin, will undergo more

immediate contracture when the graft is removed from the donor site. This shrinkage is

termed primary shrinkage since it occurs before the healing process. This higher amount

of elastin in thick grafts has shown that contracture of as much as 43% can occur.

Thinner grafts with less connective tissue will have less primary shrinkage but will

experience more secondary contracture termed "cicatrization". 1 Secondary contracture

is caused by the scarring or fibrosis of the tissue that unites the graft with its recipient

base. The effect of this cicatrization on the graft is dependant upon the rigidity of the

recipient bed and the thickness of the lamina propria of the graft.

The traditional FGG technique extends incisions for the donor harvest site to

approximately twice the desired width allowing for initial shrinkage after harvesting

allowing for a 50% graft contraction with final healing. Grafts can either be placed on

denuded bone or on periosteum. Grafts when placed on denuded bone have shown to

have less post-operative mobility, less swelling, and better hemostasis. 12 Several

biometric studies analyzed that grafts placed on denuded bone shrink 25% versus grafts

placed on a periosteal bed shrink 50% after a 24-week healing period. The greatest

amount of shrinkage typically occurs during the first 6 weeks, however a healing lag of 2

weeks is observed when grafts are placed on denuded bone. 13, 14

Purpose of the Study

Disadvantages of FGGs not only include shrinkage but also present a potential

challenge. FGGs have been known to heal in a "postage stamp" fashion since the donor

tissue color may not match the recipient tissue. This color disparity leads to an obvious









demarcation of where the FGG was placed and a potentially unaesthetic result. No prior

attempts to date have been made to improve the aesthetics of FGGs.

Pervious attempts have been made to expand FGGs, but none have been truly

successful. Rateitschak created a technique that was termed the "accordion technique". 15

This technique involves trying to gain expansion by making incisions by hand in alternate

sides of the graft using a scalpel blade. These alternating incisions attempted to gain

expansion, but limited expansion is actually achieved. The accuracy and reproducibility

of these incisions is suspected to be the limitation to expansion.

This study hopes to determine a true and predictable expansion ratio that a tissue

expander may achieve with donor tissue. The ability to expand FGGs would allow for

smaller donor tissue to be harvested, less post-operative pain for the patient, and

increased coverage from the donor tissue. We also expect that the perforations made in

our donor tissue will allow for recipient tissue to perfuse the FGG, allowing for better

aesthetics and eliminating the "postage" like appearance.

This study applies the techniques already used in modern medical grafting to dental

procedures in order to improve surgical treatment for dental patients. The medical

profession has been using devices to mesh skin grafts since the 1960s in order to expand

donor grafts for large surgical skin defects as those associated with burn patients. Their

investigations have proven that meshed grafts are both safe and effective. Medical

research has shown that the minimal thickness for a skin graft to remain viable and grow

is 1.02 mm. 6 Our research utilized grafts between 1.0- 2.0 mm in thickness. Zimmer,

Padgett, and Brennen are medical corporations marketing skin graft meshers that have the

ability of producing expansion ratios of up to 4:1. The grafts used in this study were









thicker in nature and required less expansion than what is required in medical

applications; hence, the meshing of our grafts were done in a more conservative nature.

The medical specialties have demonstrated that perforating a graft improves the graft's

viability. Regardless, if a skin graft is intended to be expanded the medical protocol still

requires perforation of the graft by this meshing process. This perforation is completed

on every graft in order to prevent hematomas or seromas, and ultimately graft acceptance.

The purpose of our study was to employ the standard surgical technique used for

dental grafting procedures, but to incorporate some of the principles of perforation and

expansion as utilized with skin grafting in order to determine an expansion ratio, aesthetic

improvements, and to determine the amount of gain in keratinized tissue. Expansion of

the graft was attempted with a custom device that is not yet available commercially. The

device was designed to bring the benefits of skin grafting to intraoral tissue grafting.














CHAPTER 2
MATERIALS AND METHODS

Inclusion Exclusion Criteria

Patients from the University Of Florida College Of Dentistry in Gainesville,

Florida will be recruited for this study and will be examined for their levels of

periodontitis, recession and amounts of keratinized tissue. The inclusion criteria are

medically healthy patients between the ages of 16-65 with a diagnosis of insufficient

width of keratinized gingiva or other mucogingival defects that would benefit from an

increase in keratinized or attached tissue. Patients were excluded from the study if they

had spontaneous bleeding upon probing, suppuration, active infection, were pregnant, or

had any other severe concurrent systemic diseases that may impact on the periodontal

condition and wound healing.












Figure 2-1. Patients with bilateral mucogingival defects were accepted into the study if
they met the inclusion criteria.

Investigational Device

The prototype instrument was manufactured by Champagne designs in West Palm

Beach, Florida. The instrument was modeled after the medical meshers that are currently









available for skin graft meshing. The instrument has a rolling drum with cylindrical

cones as the perforation mechanism. The cones are symmetrically spaced 1.0 mm apart

with the base of the cones measuring 2 mms in diameter. The instrument was designed to

be rolled over the free gingival graft (FGG) graft in a length wise direction on the outer

epithelial side. The graft is held on a polycarboxylte cutting board with the edge of a

periosteal elevator. The rolling action of the mesher is meant to perforate and mesh the

graft symmetrically.









Figure 2-2. Investigational meshing device was used to perforate and mesh the grafts.

Study Protocol

Thirteen healthy patients with bilateral mucogingival defects were used for this

study. The study design was a split mouth design. One side chosen at random

represented the experimental unit and the FGG on that side was meshed. The contra-

lateral side acted as the control unit with a traditional FGG technique employed and the

graft was not meshed. The surgical technique would be identical for both sites and would

only differ by the mesh modification made in the donor tissue on the experimental side.

The instrument would be rolled across the outer epithelial surface of the experimental

graft in a mesial-distal direction and all other procedures would be identical.

Patients included in this study were required to have five visits. The initial visit

included an explanation of the purpose of the study and the signing of informed consent

forms. Initial measurements of the mucogingival defects were recorded to include: length








and width in millimeters, and the amount keratinized gingival tissue. A Miller

Classification was determined for each site, probing depths, bleeding on probing,

mobility, and furcation involvement was also noted. The second surgical visit consisted

of pre-operative photographs to document the patient's mucogingival defect before the

surgery. Patients rinsed with a 60-second pre-operative chlorhexidine 0.12% rinse. The

recipient bed was created in the standard split-thickness method, and the involved root

surface was prepared with hand instrumentation and Pref Gel was applied to the root

surface for a minimum of 3 minutes. Pref Gel is a neutral acid that is safe on the tissues.

The purpose of this acid is to expose collagen fibers to allow for attachment of the soft

tissue grafts to the surfaces of the roots. The donor tissue was then harvested from the

patient's palate and the thickness ranged 0.75-2.00 mm. Grafts were considered "thin" if

they were less than 1.0mm, "average" if they were 1.0-1.5 mm, and "thick" if they

exceeded 1.5mm.

The sizes of the harvested grafts were documented in length and width in

millimeters and the expanded grafts were also measured post-expansion for length and

width in millimeters. The harvested graft for both the control and experimental unit was

affixed to recipient site with 5.0 vicryl suture using a p-3 needle. Surgical photos were

taken documenting each step of the procedure.



TtllT!


Figure 2-3. The grafts were affixed to the recipient bed with vicryl suture.









One graft that was harvested and expanded had a 3 x 4 mm sample removed for

histological analysis. Statistical analysis was completed by using both a t-test and a

signed ranked t-test, since our population sample was small (n=13). P values < 0.05 were

considered significant.

The subsequent post-operative visits occurred 10 days and 4 weeks post-

operatively to document the healing stages and to enforce oral hygiene. Patients were

placed on a chlorhexidine rinse 0.12% to aid in soft tissue healing and inflammation

reduction for duration of 4 weeks from the day of surgery. Motrin 800mg was prescribed

and patients were instructed to take 3200 mgs per day for control of discomfort.











Figure 2-4. Side-by-side comparison of the non-meshed graft on the left versus the
meshed graft on the right. Evaluators were asked to make comparisons of the
6-month post-operative results.

The final visit occurred at 6 months post surgery and final photographs were taken

in order to determine the final aesthetic result. The aesthetic comparisons were completed

by three board certified periodontists, 3 periodontal residents, and 3 dental students. A

computer presentation was created with before and after photographs of each case. The

evaluators were given a questionnaire and were instructed on how to complete the

comparison (Appendix A). The presentation of the cases included side by side

comparisons of the experimental graft next to the control. The two treatments were

randomly assigned a letter A or B and the evaluators were asked to judge each graft for 1)






12


graft detectibility 2) graft color match to surrounding tissue and 3) aesthetics of the graft

being superior for A or B or if they are similar.














CHAPTER 3
RESULTS

Statistical Analysis

Of the 13 patients recruited, all completed the study. Healing was uneventful for

all patients. The study population consisted of nine females and three males with the

mean age of 52. A total number of 17 mucogingival defects were located in the

mandibular premolar region, 3 in the mandibular canine region, and 6 in the mandibular

anterior region. The data collection sheets were reviewed and statistical analysis was

completed by the University Of Florida Biostatistics Department.

Both a t-test and a signed t-test were conducted (n=13) for length, width, gender,

and thickness of the grafts to determine statistical significance (p<0.05) related to

expansion. No difference in gender was found in the study. Pre-meshing and post-

meshing measurements were rounded to the nearest 0.5mm to determine if there was any

statically significant difference in length and width after the meshing was completed.

The mean change in length was calculated at 0.65mm and a standard deviation of

0.66mm. The t-test for length expansion was statistically significant (p=0.0038) and the

signed-rank test was also statistically significant (p=0.0078). The mean change in width

was calculated at 0.04 mm and a standard deviation of 0.14mm. No statistical

significance was found with either the t-test or the signed-ranked t-test for a change in

expansion of width pre-meshing and post-meshing. When considering the thickness of

the grafts harvested, no statistical difference was calculated between thin, average, or

thick grafts.









Table 3-1. Statistical analysis parameters.
Subject Length Length Width Width Graft M/F
Before After Before After Thickness
1 13.5 13.5 8.0 8.0 1.5 thick F
2 10.5 11.5 7.0 7.0 1.5 thick M
3 11.5 11.5 7.5 7.5 1.5 thick F
4 25.5 27.0 8.0 8.0 1.5 thick M
5 12.0 12.0 9.5 9.5 1.5 thick F
6 15.0 15.0 7.0 7.0 1.5 thick M
7 15.0 15.5 5.0 5.0 0.75 thin F
8 18.0 20.0 4.0 4.0 0.75 thin F
9 21.0 22.0 7.5 7.5 1.0 average F
10 30.0 30.0 5.0 5.0 0.75 thin M
11 17.0 17.5 10.0 10.0 1.5 thick F
12 17.0 18.0 10.0 10.5 1.5 thick F
13 13.0 14.0 8.5 8.5 1.0 average F

Histological Analysis

A histology sample was sent to the University Of Florida Department Of

Pathology. The tissue sample was first fixed in 10% neutral buffered formalin. The

tissue was processed on Leica tissue processor followed by dehydration with serial

alcohol intervals at 80%, 90%, 100% concentrated alcohol and followed by xylene to

clear the sample. The samples were placed in paraffin and the sample was embedded top

down in order to visualize the surface penetrations from the mesher. The paraffin block

was cut into 5 sm sections with a Micron machine. Leica Auto Stainer XL was used

to complete the Hemolysin & Eosin stain.

Histologic analysis was completed by light microscopy using a Leica microscope at

40 magnification. Photomicrographs were captured using the Q capture Pro program.

Histological analysis revealed intact parakeratinized epithelium with alternating

perforations equally distanced from each other. The perforations were seen in cross-









section and they completely incised through the epithelium and partially through the

connective tissue.















Figure 3-1. Equally spaced perforations are evident in this H&E stain with the
perforation extending into the connective tissue.

Determination of whether the incisions transversed completely through the entire

thickness of the graft could not be completed through histology since the sample was in

cross-section, the entire perforation from epithelium through connective tissue may not

have been captured in the 5m slice. Alternate transverse sections were completed and

histology revealed consistently spaced perforations through the connective tissue. No

signs of incisions or tearing could be noted from the rolling of the drum over the graft,

only perforations were detected.


Figure 3-2. Transverse section of a H&E slide of the perforation made by the device.









Aesthetic Analysis

The aesthetic evaluation was completed by a panel of evaluators. As previously

described for graft detectibility, the meshed graft was chosen 23.1% of the time as being

"non-detectible", 53.8% for "above average" blend, 23.1% of the time as having

"average" blend, and 0% as having "poor" blend. The control was chosen 15.4% as

"non-detectible", 30.8% "above average" blend, 53.8% "average" blend, and 0% chosen

as having "poor" blend. Evaluators were also asked to examine the photos for color

march to the surrounding tissues. The meshed graft was chosen 46.1% as an "excellent"

match, 38.5% "above average", 15.4% "average", and 0% as "poor" color match to the

surrounding tissues. The control graft was chosen 15.4% as "excellent", 53.8% as

"above average", 30.8% as "average", and 0% as a "poor". Evaluators were also asked to

choose which graft appeared more aesthetic when the experimental unit was compared

side-by-side to the control. The meshed graft was chosen as being superior in 46.1% of

the cases, and no difference in aesthetics was determined in 38.5% of the photos when

the meshed side was compared to the non-meshed side. The control, non-meshed side

was chosen as being superior than the meshed side in 15.4% of the cases.














CHAPTER 4
DISCUSSION

Discussion of Statistical Results

Evaluation of the statistical results yielded a mean expansion in length of 0.63mm.

The mean expansion of width was calculated only at 0.14mm. When considering the

statistical significance versus the clinical significance, 0.63mm of expansion is clinically

insignificant. One must also take into consideration the error in measurements. The

harvested grafts were not perfectly rectangular and sometimes had irregular proportions

due to the difficulty in harvesting. The measurements were made as accurate as possible

with a ruler and a probe to the nearest .05mm, but inherent error in measurements could

have occurred.

Discussion of Histological Results

A review of the histologic sample did reveal successful perforation of the device

though the epithelium and into the underlying connective tissue (Fig. 3-1). The

perforations were equally spaced, but the sample was taken in cross-section, and it could

not be determined if the perforation extended through the entire width of the graft. It is

possible that the cross-sectional 5m cut did not include the entire incision made by the

instrument and only the surface perforation was included in the slice. A transverse

sample was then analyzed and equally spaced perforations were detected in the deeper

layers of the graft (Fig. 3-1). Visual examination during the meshing of the grafts did

reveal that complete perforation was achieved by the instrument, since the cones were

able to be seen entering and exiting the graft.









Modification of Device

Since histological evaluation only revealed a perforating effect from our

instrument, it is believed that modifications to the instrument could improve the possible

expansion achieved in free gingival grafts (FGGs). Dr. Neel Bhatavadekar was consulted

regarding making modifications to the prototype in order to improve expansion ratios.

Dr. Bhatavadekar has recently just completed some research regarding the development

of a newly designed dermatome and improved skin meshing device. His work was

completed at the University of Florida in the Department of Biomedical Engineering.

Consultation with him suggested that a redesign of the device to incorporate a motorized

or hand-crank mechanism in order to provide positive pressure to the graft during the

meshing process. This positive pressure would allow for an even and controlled meshing

to occur. A suggestion was also made to modify the perforating cones. The layout of the

cones was determined to be appropriate but a suggestion to modify the cones into surgical

cutting blades. These alternating blades would provide a cutting effect instead of a

perforating effect. The cutting effect of the blades would allow for better expansion to be

achieved.

Conclusions

Aesthetic results of the meshed versus the traditional non-meshed graft were

compared by a panel of evaluators. Overall, the meshed graft was chosen 46.1% of the

time to be superior in overall aesthetics, but 38.5% of the time no difference in aesthetics

could be determined between the two sites. The control was actually chosen to more

aesthetic than the meshed site in 15.4% of the cases.

Alternative uses for this device were considered, since it is successful at perforation

of the grafts consistently. The perforations could be used as a carrier for either proteins









or amelogins. Combing these healing enhancing mediators within the grafts perforations

could act as a carrier for these substances. Since root coverage with FGGs is not as

predictable as connective tissue grafts, the addition of these substances may allow for

better root coverage by increasing attachment to the root surface. It would then be

possible to gain both keratinized tissue and root coverage by a single surgical procedure.

In conclusion, the device seemed to have improved the aesthetic appearance of the

graft in 46.1% of the cases, but clinically the expansion was non-significant. Future

research could be conducted by modifying the mesher in order to improve the expansion

ratio, or keeping the current design and using the perforations as a carrier for proteins to

improve soft-tissue procedures.















APPENDIX
DATA COLLECTION SHEET

Please Circle or Highlight one of the Choices Below for Comparison of Photo A to B
Please Fill in the Case Number #1-12


Case#


Graft Detectibility:

Graft A:
0 Graft is not Detectible (excellent blend)
I Above Average Blend of Graft
I Average Blend of Graft
0 Poor Blend of Graft

Graft B:
0 Graft is not Detectible (excellent blend)
I Above Average Blend of Graft
I Average Blend of Graft
0 Poor Blend of Graft


Graft Color Match to Surrounding Tissue:


Graft A:

Graft B:


Excellent Above Average

Excellent Above Average


Average

Average


Poor

Poor


Aesthetics of The Graft is Superior For:


Neither, Grafts have similar Aesthetic Appearance


Graft A


Graft B















LIST OF REFERENCES


1. Lang NP, Loe H. The relationship between the width of keratinized gingiva and
gingival health. J Periodontol 1972; 43:623-7.

2. Wennstrom J, Lindhe J. Role of attached gingiva for maintenance of periodontal
health. Healing following excisional and grafting procedures in dogs. J Clin
Periodontol 1983; 10:206-21.

3. Wennstrom JL. Role of attached gingiva for maintenance of periodontal health.
Healing following excisional and grafting procedures in dogs. J Clin Periodontol
1983 Mar; 10(2):206-21.

4. Soehren S, Allen A, Cutright D. Clinical and histologic studies of donor tissues
utilized for free grafts of masticatory mucosa. J Periodontol 1973; 12:727-41.

5. Studer S, Allen E, Rees T. The thickness of masticatory mucosa in the human hard
palate and tuberosity as potential donor sites for ridge augmentation procedures. J
Periodontol 1997; 68: 145-151.

6. Karring T, Loe H. Conservation of tissue specificity after hetertropic
transplantation of gingiva and alveolar mucosa. J Perio Res 1971; 6:282.

7. Pennel BM, Tarbor JC, King KO. Free masticatory mucosa grafts. J Periodontol
1969; 40:162.

8. Mormann W, Schaer F, Firestone AC. The relationship between success of free
gingival grafts and transplant thickness. J Periodontol 1981; 52:74.

9. Gordon HP, Sullivan HC. Free autogenous gingival grafts. Part II, supplemental
findings- histology of the graft site. Periodontics 1968; 6:130.

10. Mormann W, Bernimoulin JP, Schmid MO. J Clin Periodontol 1975; 2:177-189.

11. James WC, McFall WR Jr. Placement of free gingival grafts on denuded alveolar
bone. Part I: Clinical Evaluations. J Periodontol 1978; 49:283.

12. Donnenfeld OW, Marks R, Glickman M. The apically positioned flap: a clinical
study. JPeriodontol 1964; 35:381.

13. Matter J, Cimasoni G. Creeping reattachment after free gingival grafts. J
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BIOGRAPHICAL SKETCH

Michele P. Beaty was born and raised in Boca Raton, FL. She graduated with a

Bachelor in Science in 1999 from the University of Florida. She continued her post-

graduate education at the University Of Florida College Of Dentistry. In 2003 she earned

her Doctorate of Dental Medicine and began a residency in the Graduate Periodontics

Department at the University Of Florida. She currently is in private practice specializing

in Periodontics and Implant Dentistry.