Periodontal Regeneration
Stuart Jay Froum, DDS,*† and Cynthia Gomez, DDS†"Veterans Administration Hospital, and +New York University Dental Center, New York, New York, USA

This paper reviews the current clinical and histologic literature in the field of periodontal regeneration. Various procedures and agents that have been used to attain regeneration and new attachment are reviewed, including open flap debridement, root conditioning, the use of bone grafts and bone substitutes, guided-tissue regeneration, coronally positioned flaps, and a combination of these. Future directions for clinical and research efforts are discussed, with emphasis on growth factors, their delivery, and enhancement of the healing response. Literature identifying factors that will aid in the regeneration of lost periodontium is reviewed and critiqued.

Current Opinion in Periodontology 1993:111-128

Periodontal regeneration

The ultimate goal of periodontal therapy is the reconstruction of supporting tissue lost as a result of disease. In the past, conflicting reports and terminology following various attempts at regeneration led many to question whether this goal was attainable. The standardization of precise definitions of terms involved in the healing phenomenon (ie, regeneration, repair, new attachment, and reattachment) has resulted in a more accurate assessment of the healing response [1|. This accuracy, together with improved measurement techniques, has allowed critical evaluation of new attachment procedures, with the recognition that periodontal regeneration and new attachment are possible following various surgical procedures [2-8], Regeneration is now defined as the reproduction or reconstruction of a lost or injured part' [1].  Periodontally, this definition implies the formation of new bone, new cementum, and a functionally oriented periodontal ligament. Regeneration therefore may only be assessed by histologic data. New attachment is defined as the reunion of connective tissue with a root surface that has been deprived of its periodontal ligament. This occurs with the formation of new cementum with inserting collagen fibers [1|. This again is a histologic phenomenon. The histologic determination and differentiation of regeneration, new attachment, or repair has been facilitated by the calculus notch marker system (3). This system assures that the pretreatment root surface being studied has been altered by disease and is free of periodontal ligament fibers. The submerged root model system has also been instrumental in determining whether the healing response is by regeneration or repair (9).

New clinical attachment (pocket closure) may occur as a result of regeneration, new attachment, or repair. However, from a clinical standpoint, this attachment implies a gain in clinical attachment level as measured by a decrease in probing pocket-depth, when measured from a fixed reference point. Current clinical studies use attachment-level gain, gingival shrinkage, and pocket-depth reduction and reentry or tissue probing bone measurements to describe the clinical response. Various procedures and agents have been used to attain periodontal regeneration. This article will review current literature relating to periodontal regeneration by organizing these efforts into the following topics: open flap debridement (OFD):

Abbreviations

CPF—coronally positioned flap; DFDBA—decalcified freeze-dried bone allograft: FDBA—freeze-dried bone allograft: CTR—guided-tissue regeneration: OFD—open flap debridement; PTFE—polytetratuoroethylene.

Periodontology root conditioning; bone grafting and bone substitutes; guided-tissue regeneration (GTR) (membranes); coronally positioned flaps (CPFs); combination procedures; and future directions.

Open flap debridement

The OFD procedure has been shown to clinically reduce probing pocket depth with a gain in probing attachment level. This effect was contrasted with resective procedures that also resulted in a decrease in probing pocket depth, but with a loss of attachment level [10].

Fig. I. Hematoxylin-eosin stain (xl0). Note soft-tissue closure within notch, and shape and position of alveolar crest. (From Stahl and Froum |18|; with permission.)

The OFD procedure is a viable clinical procedure, and often serves as a clinical and histologic control in assessing various regenerative models. Studies have shown that the OFD procedure results in a decrease in gingival inflammation, bleeding on probing, and pocket depth [6,11]. However, a recurrence of pocket depth and bleeding on probing was shown to occur with OFD [12.13]. Overall, various clinical studies using OFD procedures have shown an average gain of probing attachment level ranging from 1.1 to 1.4 mm [l4.15,l6]. Conflicting results were observed with reentry of sites treated by the OFD procedure. Variable amounts of bone fill were reported [14, 17, 18, 19]. Fill of osseous defects ranged from O.T to 3 mm [14.20-22]. Plaque control and professional maintenance seemed strongly correlated to the degree of osseous response [23] and new clinical attachment [14]. Human histology suggests that the retention of gingival fibers results in reattachment [24]. With injury to the root surface, only limited reattachment can be expected, and apical migration of the functional epithelium occurs [24l. Histologic research has demonstrated that the primary healing response to the OFD procedure is a soft tissue adhesion of the flap to the root, with the formation of a long junctional epithelium with little or no new cementum (25-27). Schroer et al. [28•] evaluated the clinical response of facial grade it furcations in molars comparing closed versus OFD procedures. The authors found a gain in probing attachment level of 0.6 mm in the midfurcal area in the scaling and root planing groups. This contrasted with a loss of 0.46 mm in the OFD group. The OFD procedure is often used in periodontal therapy in areas of shallow to moderate periodontal defects. Deep intrabony and furcation lesions may be more effectively treated by other regenerative techniques.

Citric acid conditioning

Citric acid conditioning of the root surface is based on the premise that demineralizing the root surface exposes the collagen fibers present in dentin or cementum, which form a barrier against epithelial migration. It has been demonstrated that a root surface affected by periodontal disease undergoes pathologic changes manifested by hyper-mineralization reported to a depth of 40 to 60 µm [29], and also by binding of endotoxin [30]. Establishment of a biologically acceptable root surface involves removal of plaque, calculus, and affected cementum. Studies have shown that instrumentation of the root surfaces with hand or ultrasonic devices results in the formation of a smear layer (31). This layer may act as a barrier to connective-tissue attachment to the root. Application of a demineralizing agent removes the smear layer, thereby exposing the collagen fibers present in cementum or dentin [32]. The exposed collagen fibers are chemotactic to fibroblasts and serve as a substrate for attachment, growth, and synthesis of new connective-tissue attachment on previously diseased roots (33].  These exposed fibrils were thought to allow interdigitation ot collagen fibers from repairing connective tissues [34] addition, adhesion of the wound coagulum or granulation tissue to the root surface is enhanced by demineralizing agents. The fibrin present in the blood clot of the early wound links with the collagen in the root surfaces, forming a stable bond, which prevents apical migration of the epithelium [35].

Fig. 2.  Higher magnification of crestal area from site shown in Figure 1 (x 25). Note resorption at the facial aspect of the labial plate, osteogenesis at the periodontal aspect of the plate, and cementogenesis at the root surface. Also note the tip of the crest tilting toward the root. (From Stahl and Froum [118|; with permission.)

Early animal model studies of previously denuded root surfaces treated with citric acid demonstrated new cementum, fiber linkage, and a minimal amount of alveolar bone growth [36—38]. In humans, the results of citric acid conditioning of the root surfaces have been varied [39-41]. Doubleblind clinical evaluations showed no benefits to the use of citric acid conditioning [42]. Eschler and Rapley [43] noted that citric acid alone was not effective in removing plaque and calculus. Hanes et al. [44••] noted that root planing was necessary before citric acid treatment. In another study, Hanes et al. [45••] noted that citric acid removed the smear layer on hand-instrumented roots, enlarged dentinal tubules, and increased tubule diameters.  LeBahn et al. [46•] agreed with Hanes et al., demonstrating that citric acid removed the smear layer and opened the dentin tubules. Sterret et al. [47] claimed that the peak solution for citric acid was pH l.42 for dentin demineralization. Chaves et al. [48••] found that citric acid did not alter the effects of scaling and root planing and caused no penetration of dentin tubules. These findings were contrary to those of Hanes [44••,45••] and LeBahn [46•].

The results of studies using citric acid and root conditioning are still inconclusive, and biologic mechanisms are still unclear.  Earlier studies documented the attachment of periodontal-ligament cells to root-planed roots, even with the smear layer [21]. At present, while adverse effects of citric acid root conditioning appear limited [49-51] further studies are necessary to ascertain true clinical or histologic advantage.

Bone grafts and bone substitutes

An excellent review by Bowers and Reddi [52••] of some current approaches and future directions for periodontal regeneration was recently published. (Although omitted from this paper, autogenous intraoral bone grafts have also been shown to facilitate successful regeneration [53-56].) An excellent review of the use of freeze-dried bone allografts (FDBAs) was recently provided [57]. Mellonig et al. [58●] used bone contaminated with HIV to demonstrate that demineralization and treatment with a virucidal agent inactivates HIV and renders the allograft safe for human use. The authors [58••] also provide a detailed explanation of the time-related cellular and molecular events in the bone-induction process. Future techniques and materials are mentioned, including the use of morphogenetic proteins produced by recombinant- DNA techniques. A recent paper by Bowers et al. [59••]. using the calculus notch marking system, histologically evaluated 36 submerged defects in eight patients and 50 nonsubmerged defects in six patients with 6-month biopsies. The findings indicated that osteogenin combined with decalcified freeze-dried bone allograft (DFDBA) enhanced regeneration in a submerged environment. In nonsubmerged defects, DFDBA and osteogenin and DFDBA alone formed significantly more new attachment than either tendon-derived matrix plus osteogenin or tendon-derived matrix alone. Also, osteogenin did not impair normal lymphocyte blastogenesis 6 months postsurgery.

Drury and Yukna [60•] used nylon mesh chambers placed in surgically created windows in the mandibles of baboons to determine if tetracycline rehydration of FDBA would enhance bone regeneration compared with rehydration of FDBA with sterile water. Three and 5-week histology demonstrated much greater (>5 times) new bone formation with tetracycline-rehydrated, compared with water-rehydrated FDBA. The osteoinduction capacity of demineralized bone matrix was confirmed in a recent paper by lsaksson and Alberius [(61•).

Fig. 3. Top left, Mandibular incisors with orthodontic brackets placed prior to surgery Top right Surgical flap exposing osseous defect Bottom left, Following defect ,and root debridement implications of citric acid (pH = 1) with cotton swabs for 3 minutes. Bottom right Coronally anchored flap with silk sutures.

A review of studies using alloplastic materials suggests that these materials result in improved probing-depth reduction, decreased defect depth, and added bulk of the osseous structures present. However, use of either beta tricalcium phosphate or hydroxyapatitt does not result in regeneration of the periodontium [2] From a clinical stand point. FDBA showed a greater improvement in clinical parameters than porous hydroxyapatite in intraosseous defects, ie osseous fill. 2 1 mm versus 1.3 mm: new clinical attachment gain. 2.2 mm versus 1.3 mm; decrease in probing depth, 3.0 mm versus 1.4 mm respectively [62] Recently, Galgut el at [63•1 reported on 4-year clinical results comparing 58 sites treated with non-resorbuble hydroxyapatite and 59 sites treated with OFD. At the 4th year, reduction in pocket depth was significantly greater for sites treated with the implant material From years 2 to 4. gains in probing attachment improved with the hydroxyapatite treated sites but deteriorated in the control sites.  Extraction of a hopeless tooth 1 year following treatment with a porous hydroxyapatite implant material was reported by Mellonig [64•|. Histologically a fibrous connective-tissue attachment to a planed root surface was “strongly suggestive of a new attachment.” However, as the author [65] points out, "the root surface was not notched in calculus prior to grafting" and new attachment "could not be confirmed." Previous histology on notched root surfaces "demonstrated increased bone mass but healing by long junctional epithelium without new attachment.' Frank et al [66••] reported on bone formation following implantation or three bioceramic powders into human periodontal lesions. They conclude that bone- formation around the three bioceramics occurred through similar mechanisms at the ultra-structural level. However, by 6 months, small microsized hydroxyapatite had a significantly greater amount of peripheral bone compared with the other ceramics. The authors conclude that crystal size may be important in determining the efficiency of bone formation. They recommend that the manufacturers indicate the crystal size or the various bioceramic powders. Particle size of the most commonly used alloplasts has been shown to be consistent and ranges from 100 to 400µm [67]. Two recent studies tested two different alloplastic graft materials. In the first study, Wilson and Low [68•] compared four bioactive ceramic materials. All four ceramics allowed bone repair but Bioglass (Bioglass Research Center. Gainesville, FL) appeared to limit the downgrowth of epithelium. Shamiri et al. [69•1 recently published a clinical study comparing HTR polymer versus OFD in 30 intrabony detects in 15 patients. Their findings demonstrated that whereas pocket reduction was significantly greater initially in the implanted sites versus control sites, there were no significant differences in pocket reduction responses at 12 months postsurgery. Histologically, HTR appears to perform similarly to other alloplastic materials, being well tolerated but of limited regenerative potential [70].

Guided-tissue regeneration

In 1976 Melcher [71] suggested that healing in the periodontal milieu was determined by the cell type that repopulated the wound surface. If epithelial cells were allowed to repopulate the wound, a long junctional epithelium resulted. Cells from the gingival connective tissue caused resorption of the root surfaces, while bone-derived cells brought about ankylosis. New attachment would result if cells from the periodontal ligament were allowed to repopulate the wound area. More recently, in two separate studies, Meicher et al. [72| and Meicher and McCulloch [73] demonstrated that cells derived from bone also have the potential to mediate a regenerative response. This concept of selective cell repopulation or GTR influenced Nyman et al. [74,75] to use occlusive barriers in periodontal healing studies. Recently, Duff [76•], Dowell et al. [77•]. Phillips and Palou [78]. Caffesse and Becker [79•]. Minabe [80•]. and Nyman [81••] wrote reviews on GTR and the use of barrier membranes.

Collagen’s effectiveness as a potential barrier received much attention in a number of studies. The rationale for the use of collagen is that type 1 collagen is the main constituent of connective tissues in the periodontium; collagen is a substrate for adhesion, migration, and proliferation, and aids in the orientation of periodontal ligament cells [82]: it is hemostatic; it is available in many forms; and it is weakly immunogenic in humans [83].

Fig. 4. Overview of root surface coronal to calculus notch at tooth #31, 18 weeks after surgical procedure (hematoxylineosin stain).  Note root surface resorption apical to junctional epithelium. Arrow points to apical position of functional epithelium. (From Stahl and Froum [119••]; with permission

Flanary et al. [84•] evaluated the use of Biobrane Temporary Wound Dressing (Withrop Pharmaceuticals, NY), a synthetic skin substitute in paired class II furcation defects in mandibular molars. They concluded that Biobrane had limited clinical application in GTR. Galgut et al. (85•) evaluated the histologic response of Millipore (Millipore Corp, Harrow. Middlesex. UK), Gore-Tex (W.L Gore. Flagstaff. AZ) and two biodegradable materials when placed transcutaneously in rats.

Quteish and Dolby [86•] developed and tested a biodegradable collagen membrane on paired defects in humans. The 6-month clinical evaluation showed a gain in attachment compared with OFD alone. Paul et al. [87•] used Collistat (The Kendal Co.) in paired class II furcation defects in seven patients. Sites were reentered in 6 months. Warrer and Kamng [88•] evaluated Tisseel Immuno AG. Vienna. Austria), a two-component adhesive system with fibronectin on four dogs. Tisseel failed to facilitate repopulation of the root with periodontal ligament cells on a predictable basis. Tal and Pitaru [89•] used collagen membranes in dogs. New cementum, new bone, and new periodontal ligament (at the apical half of the defect) were seen at 30 days. Lekovic et al. [90•] used autogenous periosteum in class II furcation defects and reentered the sites in 6 months. Yukna [91•] compared Gore-Tex with freeze-dried dura matter in class II furcation defects in human mandibular molars. Kon et al. [92••] compared the healing and regenerative capacity of the periodontal ligament using resorbable (Vicryl; Ethicon, East Brunswick, NJ) and nonresorbable (Gore-Tex) membranes in dogs. The nonresorbable membranes showed better regeneration potential than the resorbable ones. Metzler et al. [93••] evaluated paired class II furcation defects in maxillary molars treated with OFD or placement of Gore-Tex membranes. They concluded that results in class II furcation defects in maxillary molars were not predictable. Pontoriero et al. [94••] created three types of defects in dogs and treated these defects by OFD and placement of Gore-Tex membranes. The sites were evaluated after 4 months. The authors concluded that GTR may result in new attachment in furcations, but the size of the defect and the degree of bone loss determined the amount of tissue regeneration. Flap recession was cited as an obstacle to regeneration using the Teflon membranes. Caton et al. [95••] evaluated the use of GTR in one wall interproximal defects in monkeys. The authors concluded that the position of the barrier defined the magnitude of coronal regeneration. Moreover, regeneration is the result of participation of cells from the periodontal ligament and alveolar bone. This finding was similar to that of Melcher [72] and Melcher and McCulloch [73].

Lu [96•] studied the topography of root trunks and found that 94% of the molars studied had a concavity on the trunk that prevented the membrane from adapting to the root. Wenzel et al. [97•] noted that bone changes after GTR correlated best with clinical measurement of attachment gain when assessed with digital subtraction radiography. Bragger et al. [98•] used computer-assisted densitometer image analysis and found it to be a noninvasive, objective method of obtaining information on remodeling of alveolar bone at sites treated by GTR. Gottlow et al. [99•] noted that GTR can result in gains of attachment, which can be maintained for periods over 5 years. Various histologic studies performed used different defect types and morphologies. These studies seem to suggest that successful regeneration was hampered by flap recession. Clinical studies concurred with this observation. Exposure of the membrane led to incomplete soft or hard tissue fill of the defects. Three separate abstracts reported colonization or bacterial contamination of the Gore- Tex material that was exposed to the oral cavity [100-102]. The importance of defect morphology in the resultant healing cannot be discounted. Most of the studies stressed the impact of the size of the defect on the outcome of GTR procedures. This issue may relate to the adequacy of soft-tissue closure or complete coverage of the defect by the membrane barrier.

Coronally positioned flap

The rationale for employing CPFs in regeneration is based on the biologic principle of delaying migration of epithelial cells, thereby allowing the fibrin clot and connective-tissue cells to repopulate the root surface. By placing the flap in a more coronal position, preferably against the enamel of the tooth, the epithelium is displaced at a greater distance from the root surface [103]. This creates a space, allowing for cells from the periodontal ligament or alveolar bone to repopulate the wound and affect a new connective-tissue attachment (104). Coronally positioned flaps have been used mainly in conjunction with citric acid conditioning or demineralized bone allografts. When treated with the combination of CPF and allografts. furcal defects in dogs were decreased or completely closed [I05]. In humans, furcal defects were treated with citric acid (pH=l. 3 min) and CPF, and were compared with defects treated with CPF, citric acid, and allografts. There was a decrease in pocket depth in class II furcation defects from 5.5 mm to 2 mm, due to attachment gain rather than recession. Bone allografts did not enhance bone formation: only 48% of class II furcations were completely closed [106].

Evaluation of the CPF and citric acid conditioning in dogs showed an enhanced flap adaptation to root surfaces and new cementum in 21 of 23 furcations in one study [107]. New connective-tissue attachment with CPF with or without citric acid occurred in 13 of 14 sites evaluated in another study [104]. Connective-tissue adhesion was observed if the flaps were placed in apposition to the roots [108]. Resorption of the roots by the cells of the gingival corium was a variable finding, as was ankylosis [104].

Gantes et al. [109•] evaluated the response of class III furcations treated with citric acid and CPF or with citric acid, CPF, and allografts. They noted that complete soft-tissue coverage of the furcation was critical for success, with only one of 14 nongrafted and three of 13 grafted defects showing complete soft-tissue clinical closure. In another review, Gantes and Garrett [110••] presented the healing responses of beagle dogs to CPF. Successful regeneration once more depended on adequate soft-tissue coverage of the defects. The most dramatic results were observed when the clinical crown was completely covered by the soft-tissue flaps. Even when complete soft-tissue coverage was not obtained, positive results were reported.
Combination techniques

In an attempt to produce more predictable results, various studies have attempted to combine the biologic principles and elements of different regenerative procedures. Combining the techniques of GTR, root conditioning, and composite grafting showed a clinical advantage of the membrane-graft group over the membrane alone [111]. Handelsman et al. [112••] compared the use of polytetrafluoroethylene (PTFE) membrane (W.L. Gore. Flagstaff. AZ) in the treatment of intraosseous defects with and without prior root conditioning with citric acid (pH = 1. for 3 min). Although in both groups clinical parameters improved, there were no statistically significant differences between the two groups. Citric acid root conditioning did not enhance clinical findings when used with PTFE membranes. Anderegg et al. [113••] compared the use of a membrane barrier alone (PTFE) or in combination with DFDBA in the treatment of class II and III furcation invasions. Six months posttreatment, surgical reentry on each site (27 class II and three class III furcations) showed no difference in attachment level changes but greater reduction of probing depths in the sites treated with DFDBA and membrane versus membrane alone. Although hardtissue changes were comparable for alveolar crest resorption, there was a statistically greater horizontal and vertical bone repair in defects treated with the DFDBA and membrane combination. However, the authors noted that only four sites of 30 closed completely. This finding differed from six of 14 noted in the previous study [111].

A recently published abstract [114] compared treatment of two and three-wall bony defects after GTR with and without hydroxyapatite implants. Conventional surgery served as a control group. There were no statistically significant differences among the groups and among the 6 to 12 months' results. The addition of hydroxyapatite implants when using membranes showed no advantage. Another abstract presented by Yamada et al. [115] compared the effects of the use of a PTFE barrier membrane with and without DFDB in created class III furcation defects in four mongrel dogs. Results suggested the use of DFDB in conjunction with the membrane can support regeneration of new bone and periodontal tissue. A recent study by Warren and Karring [116•], using surgically created horizontal defects in three dogs compared a Teflon membrane (Zitex: Norton Chemplast. New York) plus a fibrin sealant (Tisseel) with the membrane, sealant, and Kielbone (Braun; Melsungen. Germany). Histologic analysis after 3 to 4 months of healing failed to demonstrate consistent periodontal regeneration in either of the two groups. Combined Teflon membrane and porous hydroxyapatite was used by Stahl and Froum [11••] in the treatment of seven vertical lesions at seven teeth in three adults. Notches were placed in calculus and blocks were taken 10 to 28 weeks postsurgery. Histologically, two sites exhibited closure by long junctional epithelium. The remaining five sites showed gingival recession to be apical to the calculus notch (or epithelium in the notch). However, cellular cementum was seen just apical to the notch. Within the osseous crater, increased bone mass and functionally oriented periodontal ligament were seen. Two recent histologic studies explored healing of human suprabony defects, which may be the most difficult type of defect to regenerate. In one study, Stahl and Froum [118] applied a Teflon membrane over four suprabony lesions and sutured flaps as coronally as possible using orthodontic brackets as anchors. Using a calculus notch marker, 2- to 3-month block sections showed new cementum and functionally oriented fibers in three of four membrane-treated sites and immediately apical to the notch in one site (Fig. 1) Cementogenesis adjacent to supracrestal osteogenesis was seen with the new bone filling the space created by the membrane (Fig. 2). In a second study [119••], seven suprabony pockets were treated with debridement, citric acid root demineralization, and coronal positioning to the marginal flaps with orthodontic brackets as anchors. (Fig. 3) Block sections at 7 and 18 weeks demonstrated histologic evidence of new cementum with functionally inserted fibers within the calculus notch in all coronally anchored sites (Figs. 4 and 5).

Fig. 5. Site of calculus notch in specimen shown in Figure 4 (x 25). This was located apically to root surface resorption. Note osteogenesis, cementogenesis. and oriented periodontal ligament fibers (new attachment) in the calculus notch [From Stahl and Froum [119••] with permission).  

Future directions

Several years ago recommendations were made for future directions in clinical practice and research into regenerative procedures [2]. Many of the "combination therapy" regenerative procedures discussed in that review now appear to offer the opportunity for enhancing clinical results while improving the predictability of the regenerative response. A combination of CPFs in conjunction with membranes, grafts, or root conditioning has demonstrated promising clinical results (Figs. 6 and 7).

When using combination-type procedures, one must avoid the "throwing everything in" philosophy, which may be no more effective (or even less effective) than any of the component techniques used individually. This certainly was the case in a study by Wikesjo et al. [120••]. Test protocol on created supra-alveolar defects included root surface treatment with citric acid and tetracycline, followed by placement of a composite graft including hydroxyapatite, freeze-dried decalcified bone, tetracycline, and fibronectin with flaps placed and sutured to cover most of the crown. Controls included citric acid root conditioning and similarly placed flaps. Six-week histologic results showed that the composite graft protocol did not offer any advantages over citric acid alone for periodontal reconstruction. Regeneration is a complex phenomenon that depends on a coordinated response from several cell types, extra-cellular matrix, and proteins. The more that is understood about the biology of events and mechanisms regulating periodontal tissue healing, the easier it will be to transfer this knowledge to clinical techniques and to achieve more predictable regenerative results. To this end, Wang and Somerman [121••] presented a review of the proteins and factors considered essential for the maintenance and regeneration of periodontal tissues. A review of extra-cellular matrix, growth, and morphogenetic factors was presented by Ripamonti and Reddi [122••]. Further examination of osteogenin and recombinant human bone morphogenetic protein-2B was presented by Luyten et al. [123].

Several recent studies examined other growth factors related to their possible use in periodontal regeneration. A combination of platelet-derived and insulin-like growth factors was shown to enhance regeneration of the periodontal structures in periodontitis-affected teeth in beagle dogs [124••]. The use of recombinant human platelet-derived growth factor and insulin-like growth factor in debrided, experimentally produced lesions in monkeys (Rutherford et al. [125••]) suggests that further study of these growth factors is needed to treat adult periodontitis. Matsuda et al. [126••] demonstrated that recombinant human platelet-derived growth factor and insulin-like growth factor stimulate mitogenesis. proliferation, and chemotaxis of periodontal ligament fibroblastic cells. Moreover,  pH recombinant human platelet-derived growth factor also stimulates collagen synthesis by periodontal ligament cells. Takeshita et al. [127] demonstrated that interleukin 1β is a regulatory cytokine involved in the regeneration of the human periodontal ligament. Using growth factors in periodontal regenerative procedures requires a biocompatible and effective delivery system. To that end, various bone graft materials and gels have been used. Decalcified freeze-dried bone allograft combined with osteogenin, a bone-inductive protein, was used in submerged and nonsubmerged defects in humans

Fig. 6. Top left, Class II furcation on tooth #19. Top center, Furcation detect exposed. Top right, Probe penetrates 5.0 mm horizonttally. Middle left. Gore-Tex (W.L. Gore, Flagstaff, AZ) periodontal material positioned and sutured. Middle center, Coronal flap positioning utilizing Gore-Tex sutures. Middle right.  Healing open probing new attachment following membrane removal. Bottom left,  Coronally sutured flap. Bottom center, Reentry 16 months postsurgery showing bone filling the furcation. Bottom right Tooth #19, 2 years postsurgery

[59••]. Ripamonte et al [128•] used resorbable and non-resorbable hydroxyapatite rods with and without osteogenin in intramuscular sites in baboons.  Only non-resorbable hydroxyapatite with and without osteogenin showed differentiation of bone.  Osteogenin did not increase the amount of bone.  An abstract by Yewey et al. [129•] used a biodegradable polymer (Atrigel, Atrax Lab Inc. Fort Collins, CO) to release in vitro fibronectin and fibroblast growth factor. The results may hold promise for this method as a delivery system. Another future direction of regenerative research was discussed by Wikesjo et al [130••] in their review of early healing events in periodontal repair. The authors note the importance of establishing and maintaining a root surface-adhering fibrin clot to prevent apical migration of gingival epithelium and thus favor connective tissue attachment. Another attempt to prevent apical migration of epithelium during initial stages of wound healing was studied in dogs by Pitaru et al [131••]. Using bi-layered collagen harriers enriched with fibronectin and heparan sulfate, the authors repopulated 95% of the occlusal-apical length of created defects in dogs by connective-tissue cells.

Fig. 7. Top left, Eight millimeter probing depth on the mesial of tooth #5. Top center Mucoperiosteal flap reflected and mesial defect and furcation probed. Top right, Defect depth 8 mm Middle left, Following debridement, fill of the defect with decalcified freeze dried bone allograft under Gore-Tex (W.I Gore. Flagstaff, AZ), periodontal material. Middle center, Membrane positioned and sutured over the defect. Middle right, One year reentry following membrane removal. Note defect fill. Bottom left Two year probing with apparent gain in clinical attachment (pocket closure).

Conclusions

Periodontal regeneration has been demonstrated to be possible. However, to date, no single technique has been shown to be predictable for all defects. Identification of the healing events and factors that control or enhance these events has led to development of clinical techniques and new agents that we hope will improve predictability. Testing combinations of the various procedures has shown that some do not enhance (and may even inhibit) healing whereas others hold promise for improved responses. With more research data, regenerative treatment may become site-specific, with certain growth factors, root conditioning agents, and membrane or flap designs used with specific defect types. At present, non-resorbable Teflon membranes have shown the best results in
furcation defects.  Research using membranes or citric acid root conditioning with CPFs has shown the potential for supracrestal attachment.  Certainly, resorbable membranes and delivery systems for growth-regulating proteins lie in the near future. We hope the studies cited in this paper will bring us closer to the reality of predictable regeneration.

References and recommended reading

Papers of particular interest, published within the annual period of review have been highlighted as:

• of special interest
• • of outstanding interest

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14. FROUM S., CORAM, I., THULLER, B.  Periodontal healing following flap debridement procedures 1. Clinical assessment of soft tissue and osseous repair.
J Periodontal 1982, 53:8-14.

15. DURWIN, A., CHAMBERLAIN H., GARRETT S., RENVERT, S., EGELBERG, J Healing after treatment of periodontal intraosseous defects IV. Effect of a non-resective vs. a partially resective approach. J Clin Periodontol 1985. 12:525-539

16. RENVERT, S., BADERSTEN, A., NlLVEUS, R., EGELBERG, J. Healing after treatment of periodontal intraosseous defects. I. Comparative study of clinical methods.  J Clin Periodontol 1981. 8:387-399

17. RENVERT, S., EGELBERG, J  Healing after treatment of periodontal intraosseous defects II Effects of citric acid conditioning of the root surface. J Clin Periodontal 1981, 8:-459-473.

18. ELLEGAARD, B., LOE, H.  New attachment of periodontal tissue after treatment of infrabony lesions. J Periodontal 1971. 42:648-652

19. ELLEGAARD, B., KARRING, T., DAVIS R, LOE, H: New attachment after treatment of infrabony lesions.  J Periodontal 1974. 45: 308-

HlATT, W., LARATO, D., HIATT, W.,  LINDFORS, F. The induction of new bone and cementum formation. V: A comparison of graft and control sites in deep infrabony lesions. Int J Periodont Rest Dent 1986. 6:8-21.

21 .  POLSON, A., HEIL, L: Osseous repair in infrabony periodontal defects. J Clin Periodontal 1978. 5: 13-23

22.  EGELBERG, J: Regeneration and repair of periodontal tissues. J Periodontal Res 1987. 22:223-242

23. ROSLING, B., NYMAN S., LINDHE, J: The effect of systematic plaque control on bone regeneration in intrabony pockets. J Clin Periodontol 1976, 3:38-53.

24. STAHL, S., LEVINE, H.  Repair of peroodontal flap surgery with retention of gingival fibers. J. Periodontol 1972, 43:99—103

25. LlSTGARTEN, M., ROSENBERG, M., Histologic repair of new attachment procedures in human periodontal lesions. J Periodontal 1979, 50:333-344.

26. STAHL, S., FROUM, S.J.,  KUSHNER, L.  Periodontal healing of open flap debridement flap procedures. II. Histologic observations. J Periodontal 1982. 53-15.

27. CATON, J., ZANDER, H: The attachment between tooth and gingival tissue after periodic root planing and soft tissue curettage. J Periodontol 1979), 50:462.

28. SCHROER, M., WAHL, T., HUTCHENS, L. MORIARTY, J., BERGENHOLTZ, B: Closed versus open debridement of facial grade II molar furcations. J Clin Periodontol 1991. 18323-329. Twenty-five teeth with facial class II furcation problems were treated with scaling and root planing and were evaluated at 4 months. Twelve of 25 teeth were treated with OFD. 13 were treated with scaling and root planing. There was a decrease in probing depth, gingival inflammation, and Plaque Index at 16 months. Scaling and root planing groups gained probing attachment level by 0.6 mm; OFD groups lost 0.46 mm

29.  SELVIG, K., HALS, D: Periodontally diseased cementum studied by correlated micro-radiography, electron-probe analysis and electro-microscope. J Periodont Res 1977. 12:419-429

30. ALEO, J., DERENZIS, T., FARBER, P.  In vitro attachment of human gingival fibroblasts to root surfaces.  J Periodontol 1975 46:639-645.

31. JONES, S., LOZDAN, J., BOYDE, A. Tooth surfaces treated in situ with peroidontal instruments SEM studies.  Br Dent J 1972. 132:57-64

32. POLSON, AM., FREDERICK, GT., LADENHEIM, S., HANES, PI., The production of a root surface smear layer by instrumentation and its removal by citric acid.
J Periodontol 1984.  55:443-446

33. FERNYHOUGH, W., PAGE, R: Attachment, growth and synthesis by human gingival fibroblasts on demineralizcd or fibronectin treated normal and diseased tooth roots, j Penodontol 1983. 54:133-140

34. LOPEZ, NJ: Connective tissue regeneration to periodontally diseased roots, planed and conditioned with citric acid and implanted into the oral mucosa. J Periodontol 1984. 55: 381-390

35. POLSON, AM., PROYE, MF. Fibrin linkage: A precursor for new attachment.
J Periodontol 1983. 54:l4l-147

36. REGISTER, A., BURD1CK, F.  Accelerated reattachment with cementogenesis to dentin, demineralized in situ. II Defect repair. J Periodontol 47:497-505

37. RlRIE, CM., CRIGGER, M., BOGLE, G. EGELBERG, J, SELVIG, K. Healing of periodontal connective tissue following surgical wounding and application of citric acid in dogs. J. Periodontal Res 1980 15:314-327.

38. NILVEUS, R., EGELBERG, J:  The effect of topical citric acid application on the healing of experimental furcation defects in dogs. III. The relative importance of coagulum support, flap design and systemic antibiotics J Periodont Res 1980. 15:551

39. COLE, RT., CRIGGER. M., BOGLE, G., EGELBERG, J., SELV1G, K: Connective regeneration to periodontally diseased teeth. J Periodont Res 1980, 15:1-9.

40. FROUM, SJ., KUSHNER, L., STAHL, SS. Healing responses in human intraosseous lesions following the use of debridement, grafting and citric acid root treatment. II. Clinical and histologic observations, one year postsurgery. J Periodontol 1983, 54:325-338.

41. REVERT, S., EGELBERG, J  Healing after treatment of periodontal intraosseous defects. II. Effect of citric acid conditioning of the root surface. J Clin Periodontol 1981. 8:459-473.

42. MOORE, J., ASHLEY, KP., WATERMAN CA The effect on healing of application of citric acid during replaced flap surgery J Clin Periodontol 1987. 14:130-135

43. ESCHLER, B., RAPLEY, J Mechanical and chemical root preparation in vitro Efficiency of plaque and calculus removal J Periodontol 1991, 62:755-760

44. HANES, P., POLSON, F., FREDERICK, T. Citric acid treatment of •• periodontitis affected cementum: a SEM study. J Clin Periodontol 1991. 18:567.
Cementum surfaces from roots of extracted human teeth from areas beneath periodontal ligaments (normal) and calculus deposits (periodontitis-affected) were evaluated. Only scaling was done on affected roots. 

Both groups were treated with citric acid (pH=l, 3 min). Citric acid treatment of normal roots exposed a fibrillar collagen substrate, while periodontitis-affected roots were not altered in appearance by citric acid treatment, implying that these roots had undergone changes that reduced the effects of a demineralizing agent. 

45. HANES, P., O'BRIEN, N., GARNICK, J: •• A morphologic comparison of radicular dentin following root planing and treatment with Citric acid or tetracycline HCL. J Clin Periodontol 1991, 18:660-668. An evaluation using scanning electron microscopy is presented of the morphology of dentin after the use of hand instruments, citric acid (pH-l. 3 min), and tetracycline (pH-3 2, 5 min). Hand instruments produced a smear layer, with no dentinal orifices exposed Citric acid produced round-to-oval dentinal orifices that were irregular in shape, flared, or funneled in appearance. Tetracycline produced numerous dentinal orifices that were variable in size and shape with some specimens exhibiting a layer of organic debris.

46. LEBAHN, R. FAHRENBACH, W., CLARK, S., LIE, T., ADAMS, D. •Root dentin morphology after different modes of citric acid and tetracycline hydrochloride conditioning. J Periodontal 1992. 63:303-309
Twenty dentin blocks from freshly extractted non-diseased human molars were root-planed and treated with citric acid or tetracycline for 30, 60, 120, and 240 seconds.  There was a time dependent increase in dentinal tubule diameter with citric acid. Citric acid caused more extensive changes than tetracycline.

47. STERRET, J., DELANEY, B., RIZKALLA, A., HAWKINS, C. Optimal citric acid concentration for dentinal demineralization. Quintessence Int 1991. 22.371-375

48. CHAVES, E., COX, C., MORRISON, E., CAFFESSE, R. The effect of •• citric acid application on periodontally involved root surfaces. I. An in vitro light microscopic study, Int J Periodont Rest Dent 1992. 12:219-229. Forty periodontally involved teeth were divided into four groups: no treatment, citric acid treatment, root planing, and citric acid with root planing. Citric application alone had no effect on the root surface and did not alter the appearance or penetrate the dentinal tubules of the planed root surface. The authors noted an incomplete removal of the cementum without careful root planing in an in vitro model. The effects obtained after scaling and root planing were not altered by citric acid application on a light microscopic level.

49. WIKESJO, U., CLAFFEY, N., NILVEUS, R., EGELBERG, J.  Periodontal repair in dogs: effect of root surface treatment with stannous fluoride or citric acid" on root resorption. J Periodontol 1991. 62 180-184.

50. AUKHIL, I. PETTERSSON, E: Effect of citric acid conditioning on fibroblast cell density in periodontal wounds. J Clin Periodontol 1987. I4:80-84.

51. PETTERSSON, E., AUKHIL, I., Citric acid conditioning of roots affects guided tissue regeneration in experimental periodontal wounds. J Periodont Res 1986 21
543-552.

52. BOWERS, GM., REDDI, AH: Regenerating the periodontium •• in advanced periodontal disease. J Am Dent Assoc 1992. 122:45-48.
An excellent review of some current approaches and future directions for periodontal regeneration. The authors reference histologic research, documenting regeneration using autologous cancellous bone and marrow. References are cued for use of DFDBA materials with the limiting factor being limited performance in supracrestal (and furcation) defects due to the small particle size allowing displacement by overlying flap. The authors discuss the fear of disease transmission and quote from the literature estimating the risk of transmitting HIV as being one in 8 million after the graft material is frozen and with proper tissue banking. The authors also provided a detailed explanation of the name-related cellular and molecular events in the bone-induction process. Future techniques and materials are mentioned and include the use of morphogenic proteins produced by recombinant DNA techniques. One such bone-inductive protein, osteogenin. in association with a bone-derived collagenous matrix, appears to be safe in humans and is currently being evaluated.

53. FROUM, SJ., THALER, R., SCOPP, IW., STAHL, SS. Osseous autografts. II Histological responses to osseous coagulum-bone blend grafts. J Periodontol 1976.
46:656-661

54. SCHALLHORN, RG., Present status of osseous grafting procedures. J Periodontol 1977 48:570-576

55. HIATT, WH., SCHALLHORN, RG., AARONIAN, AI: The induction of new bone and cementum formation IV. Microscopic examination of the periodontium following human bone and marrow allograft, autograft, and nongraft periodontal regenerative procedures J Periodontol 1978. 49-495-512

56 BOWERS, GM., SCHALLHORN, RG., MELLONIG, JT. Histologic evaluation of new attachment in human intrabony defects: A literature review J Periodontol 1982. 53:509-514.

 greater for sues treaied with the implant material From vears 2 to •+. Rains in probing attachment improved with the hvdroxvapautetreated sites but deteriorated in the control sites.

57. MELLONIG, J T. Freeze-dried bone allografts in periodontal reconstructive surgery Dental Clinics of North America 1991. 35:505-520

58.  MELLONIG, J T., PREWETT AB., MOYER, MP.  HIV inactivation in •• a bone allograft J Periodontol 1992. 63:979-983.
Direct evidence is presented demonstrating that processing of a DFDBA renders the allograft safe for human use. Using disease free cortical bone that was then contaminated with viral particles, and cortical bone from a donor who died of AIDS. the authors show that all treated samples were negative when assayed for HIV.

59. BOWERS, G., FELTON, F., MIDDLETON, C., GYLNN, D., SHARP, S., • • MELLONIG, J., CORIO, R., EMERSON, J., PARK, S., SUZUKI, J. MA S, ROMBERG, E., REDD, AH: Histological comparison of regeneration in human intrabony defects when osteogenin is combined with demineralized freeze-dried bone allograft and with purified bone collagen. J Periodontol 1991. 62:690-702.
Using the calculus notch marking system, the authors histologically evaluated 36 submerged defects in eight patients and 50 non-submerged defects in six patients with 6-month biopsies. The findings indicate that osteogenin combined with DFDBA enhanced regeneration in a submerged environment In non-submerged defects DFDBA and osteogenin and DFDBA alone formed significantly more new attachment than either tendon-derived matrix plus osteogenin or tendon-derived matrix alone. Also, osteogenin did not impair normal lymphocyte blastogenesis 6 months postsurgery.

60. DRURY, GI. YUKNA, RA: Histologic evaluation of combining • tetracycline and allogenic freeze-dried bone on bone regeneration in experimented defects in baboons
J Periodontol 1991. 62,052-658
The authors used nylon mesh chambers placed in surgically created windows in the mandible of baboons to determine it tetracycline rehydration of FDBA would enhance bone regeneration compared with rehydration of FDBA with sterile water. Three and five-week histology demonstrated much greater (>5 x 1) new bone formation with tetracycline rehydrated compared with the water-rehydrated FDBA.

61. ISAKSSON, S. ALBERIUS, P: Comparison of regenerative capacity elicited by demineralized bone matrix of different embryonic origins. J. Craniomaxillofac Surg 1992, 20:73-80.
The osteoinduction capacity of demineralized bone matrix was confirmed Fifteen-week results with demineralized bone matrix (of both membranous and enchondral origin) implanted in trephined calvarial defects in adult rabbits displayed extensive osteoinductive capacity and early bone production that significantly exceeded the two controls, autogenous bone chips and an unfilled defect.

62. BARNETT, JD.,  MELLONIG, IT., GRAY, JL., TOWLE HJ: Companson of freeze-dried bone allograft and porous hydroxylapatite in human periodontol defects. J. Periodontol 1989. 60:231-237

63. GALGUT, PN., WAITE, IM., BROOKSHAW, JD, KINGSTON, CP.  A • 4-year controlled clinical study into the use of a ceramic hydroxylapatite implant material for the treatment of peridontal bone defects. J Clin Periodontal 1992, 19: 57O-577
The authors reported on 4-year clinical results comparing 58 sites treated with nonresorbable hydroxyapatite and 59 sites treated by OFD. Pockets were categorized as shallow (less than 3 mm), moderate ( 3 mm to 6 mm) and deep (greater than 6 mm). Both test and control groups lost attachment (0.9 mm and 1.09 mm) in shallow sites. For moderate and deep pockets, both groups gained attachment. At the 4th year, reduction in pocket depth was significantly greater for sites treated with the implant material.  From years 2 to 4, gains in probing attachment improved with the hydroxyapatite treated sites but deteriorated in the control sites. 

64. MELLONIG, JT: Porous particulate hydroxyapatite in a human • periodontal osseous defect: a case report. Int J Periodont Res Dent 1991, 11:217-223.
A tooth diagnosed as periodontally hopeless was treated with a synthetic bone graft (porous hydroxyapatite) and was extracted 1 year postsurgery. Clinical findings indicated a fractured root. Histologic examination revealed previously planed fibrous connective tissue attached to the root surface. This connective tissue contained particles of hydroxyapatite. Because the author did not delineate the extent of root planing, a determination of new attachment or reattachment cannot be made.

65. STAHL, S., FROUM, SJ: Histologic and clinical responses to porous hydroxyapatite implants in human periodontal defects three to twelve months post-implantation. J Periodontol 1987. 58:689-695.

66. FRANK, RM., KLEWANSKY, P., HEMMERLE, J , TENENBAUM, H: ••Ultra-  structural demonstration of the importance of crystal size of bioceramic powders implanted into human periodontal lesions. J Clin Periodontol 1991. 18: 669-680. The authors reported on bone formation following implantation of three bioceramic powders into human periodontal lesions. Twelve-month biopsies of sites implanted with B tricalcium phosphate or hydroxyapatite were compared with 6-month biopsies of sites implanted with micro-sized hydroxyapatite. The authors concluded that bone formation around the three bioceramics occurred through similar mechanisms at the ultrastructural level. However, by 6 months, small micro-sized hydroxyapatite had a significantly greater amount of peripheral bone compared with the other ceramics at 12 months. The authors concluded that crystal size may be important in determining the efficiency of bone formation.  They recommend that the manufacturer indicate the crystal size of the various bioceramic powders.  

67. ZANER, DJ., YUKNA, RA: Particle size of periodontal bone grafting materials. J Periodontal 1984. 55 406-409.

68. WILSON, J., Low, SB. Bioactive ceramics for periodontal treatment: comparative studies in patus monkey J Appl Biomat 1992. 3:123-129. The authors compared four bioactive ceramic materials. Two forms of Bioglass: synthagraft and augment, and alveolograf and periograf were implanted in surgically created defects in 6 adult monkeys (Erythrocebus patus).  All four ceramics allowed bone repair but Bioglass appeared to limit the downgrowth of epithelium. The authors claim that bone growth from the surface of the alveolus is augmented by bone growth on the Bioglass surface, thus causing a more rapid fill of the defect with Bioglass.

69. SHAMIRI, S., SINGH, IJ., STAHL, SS: • Clinical response to the use of HTR polymer implant in human intrabony lesions. Int J Periodont Rest Dent 1992. 23:295-299 This study compared hard-tissue replacement polymer versus OFD in 30 intrabony defects in 15 patients. Their findings demonstrated that whereas pocket reduction was significantly greater initially in the control sites, there was no significant difference in pocket reduction responses at 12 months postsurgery.

70. STAHL, SS., FROUM, SJ.,  TARNOW, D: Human clinical and histological responses to the placement of HTR polymer particles in intrabony lesions. J Periodontol

71. MELCHER, AH: On repair potential of periodontal tissues. J Periodontol 47:256-260

72. MELCHER, AHJ., CHEONG, T., COX, J., NEMETH, E., SHIGA, A: Synthesis of cementum-like and bone-like tissue in vitro may migrate into the periodontal ligament in vivo. J Periodont Res 1986. 21: 592-612.

73. MELCHER, AH., MCCU LLOCH, C: Cells from bone synthesize cementum-like and bone-like tissue in vitro and may migrate into periodontal ligament in vivo. J Periodont Res 1987 22:246-247

74. NYMAN, S., GOTTLOW, J. KARRING, T.  The regenerative potential of periodontal ligament: an experimental study in the monkey. J Clin Periodontol 1982. 9:257-265

75. NYMAN, S., LINDHE, J., KARRING, T: New attachment following surgical treatment of periodontal disease. J Clin Periodontol 1982. 9: 290-296

76. DUFF, B: Guided tissue regeneration: a new frontier in periodontics. J Mich Dent Assoc 1991. Apr/May: 15 -19
In this literature review, the author notes that the use of membranes in GTR is best in three wall intrabony lesions, and class II furcation with vertical bone loss. He points out that careful case selection is crucial, and that the surgical technique is demanding. He states that the exact nature of attachment with OTR in humans is not completely elucidated

77. DOWELL, P., MORAN, .J, QUTEISH, D: Guided tissue regeneration. Br Dent J
 1991. 171:125-127.
The authors discussed the materials used in GTR, and their limitations Gore-Tex has a tendency to adhere to root surfaces during healing, preventing cell regeneration in the area of contact. It is expensive, and recession is a common sequel. Millipore fillers are brittle and difficult to manipulate. Collagen membranes do not consistently provide new attachment in humans. There is the risk of developing Creutzfeldt-Jakob disease if dura mater is used. Polylactic acid has not been sufficiently studied.

78. PHILLIPS, J., PALOU, M: A review of the guided tissue regeneration concept Gen Dent 1992. Mar/Apr: 118-123
The authors reviewed the various materials used in GTR and pointed out the problems in using non-resorbable membranes. These problems include abscess formation, membrane exfoliation, and recession They also pointed out the variety of responses and the lack of long-term predictability and Mobility of GTR-treated defects.

79. CAFFESSE, R., BECKER, W: •• Principles and techniques of guided tissue regeneration. Dent Clin North Am 1991. 35:479-494
A review of the literature is presented. The authors point out that the ideal defects for GTR procedures are three-wall defects, two- t three-wall defects funnel-shaped defects with definite osseous stops greater than 5 mm deep, and class II furca with or without a vertical component. Full thickness flaps are used in the surgical procedure.  The interdental papillae are retained, and vertical releasing incisions are recommended.

80. MlNABE, M: • • A critical review of the biologic rationale for guided tissue regeneration. J Periodontol 1991. 62:171-179.
This is a good literature review with a tabulation of results from animal and human studies. The author pointed out that defects beyond a certain size may not be completely restored by GTR. and the morphology of the defect as well as postsurgical recession are important in the success of GTR. The author also pointed out the advantage nt CPFs to decrease recession. When discussing resorbable membranes, the author noted that the time degradation of the membrane is crucial, but the critical period for degradation of the resorbable membrane is still not known.

81. NYMAN S: •• Bone regeneration using the principle of guided tissue regeneration. J Clin Periodontol 1991. I8:494-498 
Reviewed the history of GTR and discussed the use of membranes in endodontic surgery, ridge augmentation, and implants.

82. POSTLEWAITE, A., SEYER, J.,  KANG, A Chemotactic attraction of human fibroblasts to type 1. II. III collagens and collagen derived peptides. Proc Natl Acad Sci USA 1978. 75:871-875.

83. COOPERMAN, L., MICHAELI, D:  The immunogenicity of injectable collagen I: A one year prospective study J Amer Acad Dermatol 1984. 10:638-646
84. FLANARY, D., TWOHEY, S., GRAY, J., MELLONIG, J., GHER, M: •  The use of synthetic skin substitute as a physical barrier to enhance healing in human periodontal furcation defects: a follow up report. J Periodontol 1991. 62:684-689.
The authors evaluated the use of Biobrane, a synthetic skin substitute of polydimethylsiloxane bonded to nylon fabric with peptides from porcine dermal collagens. In paired class II furcation defects in mandibular molars in humans, none of the furca treated with the material were completely closed in a horizontal direction at the 6-month reentry point. The authors concluded that Biobrane has limited clinical application in GTR.

85. GALGUT, R., PITROLA, K., WAITE, I., DOYLE, I., SMITH, R: • Histologic  evaluation of biodegradable membranes placed transcutaneously in rats.
J Clin Periodontol 1991. 18 581-586.
The authors evaluated the histologic responses to Millipore, Gore-Tex, and two biodegradable materials of different molecular weights in transcutaneous wounds in rats. Healing was governed by a variety of factors, including host responses, the chemical composition of the materials, the physical and surface character of the material, its porosity, and the depth of insertion into the tissue.

86. QUTEISH, D., DOLBY, AE: • The use of irradiated cross-linked human collagen membrane in guided tissue regeneration. J Clin Periodontol 1992. 19:476-484
Type I collagen was extracted, prepared, irradiated, and cross-linked. The collagen was placed in 26 periodontal defects in 19 patients. No untoward reactions were observed in the test period.  There was no reentry. Clinical examination revealed a gain of attachment with the use of this collagen preparation.

87. PAUL, B., MELLONIG, J., TOWLE, H., GRAY, J:  • Use of collagen barrier to enhance healing in human periodontal furcation defects. Int J Periodont Rest Dent 1992. 12:123-131. The authors used Collistat in paired class II furcation defects in seven patients. The study reported an improvement in probing depths and horizontal osseous support with Collistat, but the changes adjacent to the material may have been due to difference in defect morphology rather than to a function of the barrier.

88. WARRER, K., KARRIM. T: • The effect of Tisseel on healing after periodontal flap surgery. J Clin Periodontol  1992.
Tisseel, a two-component adhesive system with fibronectin, was used in 5-mm defects in four dogs. The animals were sacrificed in 4 months. Tisseel failed to facilitate repopulation of the root surface with periodontal ligament cells on a predictable basis. Tisseel may not favor new attachment unless the root is pretreated with citric acid.

89. TAL, H., PITARU, S: • Formation of new periodontal attachment apparatus after experimental root isolation with collagen membranes in the dog. Int J Periodont Rest Dent 1992. 12:231-242.
Four-mm buccal defects were induced in maxillary canines, and U-shaped defects were induced in premolars of nine dogs. Membranes made of collagen from rat tail were placed. Animals were sacrificed at 10 and 30 days. Histology showed functional epithelium of the coronal quarter of the defect, with new bone, periodontal ligament, and cementum at the apical half. Connective tissue adhesion was seen between the functional epithelium and new cementum.  No bone formation was seen in the canines.

90 LEKOVIC, V., KENNY, B., CARRANZA, F., MARTIGNONI, M: • The use of autogenous periosteal grafts as barriers for the treatment of class II furcations involvements in lower molars. J Periodontal 1991. 61:775-780.
Fifteen patients with class II furcation defects were treated using periosteum as a barrier to utilize progenitor cells in the periosteum The sites were reentered in 6 months. The use of periosteum yielded similar results to those reported in the Gore-Tex studies. The periosteum may modify the cellular dynamics of wound healing.

91 YUKNA, R: Clinical human comparison of expanded polytetrafluorethylene barrier membrane and freeze dried dura mater allografts for guided tissue regeneration of lost periodontal support. I. Mandibular molar class II furcations.
J Periodontol 1992. 63:431-442
Yukna compared Gore-Tex and freeze-dried dura mater in the treatment of class II furcation defects in mandibular molars in humans, with a 12-month reentry evaluation. He saw no "complete” fill in furcations with soft or reformed tissue, leading him to question the use of GTR in the management of class II furcation defects.

92. KON, S., RUBEN, ML., BLOOM, A., DEY, WM., BOFFA, J: ••Regeneration of periodontal ligament using resorbable and non-resorbable membranes. Clinical, histological and histometric study in dogs. Int J Periodont Rest 1991. 11:59-71.
Defects measuring 5 mm by 6 mm were created in three dogs and were treated with Vicryl, Gore-Tex and control sites. Gore-Tex was removed in 5 weeks. New attachment divided in to three zones, apical zone had periodontal ligament, bone, and cementum. Middle zone had fibers into cementum and cementoid. The coronal zone had loose connective tissue in an undefined structure. Vicryl had 1.48 mm of regeneration. Sixty percent of the lesion (0-9 mm) had new attachment apically. 40% (0.58 mm) had connective tissue attachment inserted in cementoid (or undefined structure) coronal to the bone.

93. METZLER, D., SEAMONS, B., MELLONIG, J., GHER, M., GRAY, J: •• Clinical evaluation of guided tissue regeneration in the treatment of maxillary class II molar furcation invasions. J Periodontol 1991. 62:355-560.
Seventeen patients with paired class II furcation problems on maxillary molars were treated with OFD or placement of Gore-Tex membrane.  Membranes were removed in 6 weeks, and sites were reentered at 6 months.  Recession was seen in 50% of the cases. No defect “filled" with bone in 6 months. There was an increase in attachment levels and a decrease in probing depths, but regeneration was not predictable.  The authors suggested the use of citric acid, tetracycline or allografts to enhance results.

94. PONTORIERO, R., NYMAN, S, ERICSSON, I., LlNDHE, J: ••  Guided tissue regeneration in surgically produced furcation defects, an experimental study in the beagle dog. J Clin Periodontol 1992. 19: 159-163
Part I of the study involved creating "keyhole" defects (2 x 2 mm) in the furcations of dogs. Gore-Tex was placed and removed in 30 days, and animals were sacrificed at 4 months. Histology revealed new cementum and collagen in the roots lacing the furcation detect. Bone regeneration of 0.99 to 2 3 mm was seen (50% to 100% of original bone height restored). Functionally oriented periodontal ligament was seen where new bone and cementum formed. Part II involved through-and-through furcation defects (3 mm high. 4 mm wide) treated with Gore-Tex. Animals were sacrificed at 45 days. Histology showed new collagen in one of five test sites. The remaining sites had partial healing with new attachment. Lack of new attachment was correlated to flap recession and membrane exposure 2 weeks after the surgery. Part III involved creating wide, through-and-through defects (5 mm high, -4 mm wide) Gore-Tex was placed, and animals were sacrificed in 45 days.  There was recession in all the teeth, and no new cementum in eight of the 10 furcations studied. Guided tissue regeneration may result in new attachment in furcation defects, but the size of the defect and the degree of bone loss adjacent to the defect and the soft-tissue recession determines the outcome of regenerative procedures.

95. CATON, J., WAGENER, C., POLSON, A., NYMAN, S., FRANTZ, B., BOUWSMA, O., BLIEDEN, T: •• Guided tissue regeneration in interproximal defects in monkeys.  Int J Periodont Rest Dent 1992. 12:267-277
One-wall interproximal defects were created in monkeys and Gore-Tex was placed. The membranes were left for 1 and 3 months, at which times the animals were sacrificed. At 1 month, there was junctional epithelium to the membrane, new cementum. cementoblast-like cells in the apical area, immature periodontal ligament, new bone, and osteoid. At 3 months, there was mature cementum with Sharpey s fibers in the coronal one third of the root, and new bone. Some of the angular defects filled completely with bone, maintaining a periodontal ligament space. The new periodontal ligament had normal morphology. The position of the barrier defined the magnitude of coronal regeneration. The authors suggested leaving barriers in place for longer periods of time - anywhere from 1 to 3 months.

96. LU, J:  • Topographical characteristics of root trunk lengths related to guided tissue regeneration. J Periodontol 1992. 63:215-216
Lu studied the topography of root trunks and found that 94% of the molars studied had concavity depths on the trunks that prevented the membranes from adapting to the root. The author suggested a supragingival placement of the membrane to avoid this concavity.

97. WENZEL, A., WARRER, K., KARRING, T:  • Digital subtraction radiography in assessing bone changes in periodontal defects following guided tissue regeneration.
J Clin Periodontol 1992. 19:208-213
Clinical gain of attachment is accompanied by formation of new alveolar bone. The amount of bone formation alter GTR vanes considerably. Changes in the marginal bone level determined by conventional radiography are terminated in 6 months Bone changes after GTR correlated better with clinical measurements of attachment gain when assessed with digital subtraction radiography.

98. BRAGGER, U., HAMMERLE, CHF., MOMBELLI, A., BURGlN, W., LANG, NP: Remodelling of periodontal tissues adjacent to sites treated according to the principles of guided tissue regeneration. J Clin Periodontol 1992. 19:615-624.
The authors noted that because hard-tissue changes are complete in 1 year, reentry should not he done until that time.

99. GOTTLOW, J., NYMAN, S., KARRING, T: • Maintenance of new attachment gained through guided tissue regeneration. J Clin Periodontol 1992. 19:315-317
Gain of probing attachment was observed in 80 new attachment sites: 80 were monitored for 1 year. 67 for 2 years. 40 for 3 years. 17 for 4 years, and nine for 5 years. Nine of nine sites at 5 years were found to have retained the improved probing attachment level value. The gain of probing attachment observed may reflect a new connective tissue attachment that can be maintained on a long-term basis.

100. GREVSTAD, HJ., LEKNES, KN: Ultrastructure of PTFE membrane associated plaque. J Dent Res 1992. 72:580.

101. KARRING, T: Clinical results of guided tissue regeneration: what are its limitations? J Dent Res 1992. 72:519

102. GOMEZ, C., LlNKE, H:  Microbiota around the Gore-Tex periodontal material.
J Dent Res 1992. 71:216.

103. MARTIN, M., GANTES, B., GARRETT, S., EGELBERG, J: Treatment of periodontal furcation defects. I. Review of literature and description of regenerative surgical technique. J Clin Periodontol 1988. 15:227-231.

104. GOTTLOW, J., NYMAN, S., KARRING, T., LINDHE, J: Treatment of localized gingival recessions with coronally displaced flaps and citric acid: an experimental study in the dog. J Clin Periodontol 1986. 13:57-63.

105. NILVEUS, R., JOHANNESSON, D., EGELBERG, J: The effects of autogenous cancellous bone grafts on healing of experimental defects in dogs. J Periodont Res 1978, 13:532-537

106. GANTES, B., MARTIN, M., GARRETT, S., EGELBERG, J: Treatment of perodontal furcation defects. II. Bone regeneration of mandibular class II defects. J Clin Periodontol 1988, 15:232-239

107. CRIGGER, M., BOGLE, G., NILVEUS, R., EGELBERG, J., SELVIG, K: The effect of topical citric acid application to the healing response of experimental furcation defects in dogs. J Periodont Res 1978, l3:538-542.

108. BOGLE, G., CLAFFEY, N., EGELBERG, J: Healing of horizontal circumferential periodontal defects following regenerative surgery in beagle dogs. J Clin Periodontol 1985, 12:837-849

109. GANTES, B., SYNOWSKY, B., GARRETT, S., EGELBERG, J: Treatment of periodontal furcation defects: Mandibular class III defects. J Periodontol 1991. 62:361-365.
Twenty-seven class III furcation defects in mandibular molars were treated by citric acid root conditioning and CPF (in CPF 14 cases, citric acid, and DFDBA in 13 cases). Clinical measurements were taken and reevaluated 6 months after treatment. Vertical probing depth-reduction and mean probing attachment-level gain in the furca were 2.6 mm and 2.2 mm for the nongrafted defects and 1.9 mm and 1.5 mm for the grafted defects. No differences were observed between the defects treated with or without bone grafts.

110. GANTES, B., GARRETT, S: •• Coronally displaced flaps in reconstructive periodontal therapy. Dent Clin North Am 1991. 35:495-504.
This paper was a presentation of studies performed on dogs and humans. The first part was a histologic documentation of healing responses in dogs to CPF and citric acid (pH-1. 3 min) Small furcation defects were successfully regenerated, but large defects had no primary closure and were not treated with success. Citric acid with extreme CPFs (clinical crowns completely covered) showed complete regeneration, with flaps gradually receding and stabilizing at the cementoenamel junction. Citric acid with moderate coronal position (flaps anchored 1 mm above the cementoenamel junction. sutures attached to enamel with composite) showed that if the flaps stayed attached to the crowns, regeneration occurred. Clinical techniques and flap design for use in clinical situations were presented. The authors also reported on the treatment of class II furcation problems using citric acid and CPF. Only 50% of the lesions were partially closed, while 48% completely closed. Probing depths were reduced from 5.5 mm to 2 mm. Decreases in probing depth were due to gain in attachment, not recession.

111. SCHALLHORN, KG.,  MCCLAIN, PK: Combined osseous composite grafting, root conditioning and guided tissue regeneration. Int J Periodont Rest Dent 1988. 8: 9-31

112. HANDELSMAN, M., DAVARPANAH, M., CELEETI, R: •• Guided tissue  regeneration with and without citric acid treatment in vertical osseous defects. Int J Periodont Rest Dent 1991. 11:351-363.
The authors compared the use of a PTFE membrane in the treatment of intraosseous defects with and without prior root conditioning with citric acid (pH=l for 3 minutes). Although in both groups clinical parameters were improved, there were no statistically significant differences between the two groups Citric acid root conditioning did not enhance clinical findings when compared to the membrane-only group.

113.  ANDEREGG, CR., MARTIN, SJ., GRAY, JL., MELLONIG, JT., GHER, ME:
•• Clinical evaluation of the use of decalcified freeze-dried bone allograft with guided tissue regeneration in the treatment of molar furcation invasions. J Periodontol 1991. 62:264-268. The authors compared the use of membrane barrier alone (PTFE) or in combination with DFDBA in the treatment of class II and III furcation invasions. Six months post-treatment, surgical reentry on each site (27 class II and three class III furcations) showed no difference in attachment level changes but greater reduction of probing depths in the sites treated with DFDBA and membrane versus membrane alone. Although, hard-tissue changes were comparable for alveolar crest resorption, there was a statistically greater horizontal and vertical bone repair in defects treated with the DFDBA and membrane combination. However the authors note that only four sites of 30 completely closed.

114. FATH, S. WACHTEL, HC., BERNIMOULIN, JP: Treatment of periodontal intrabony lesions with the GTR technique and hydroxylapatite implants. J Dent Res 1992. 72(special issue):624.

115. YAMADA, S., TAKAHASHI, V., YAMANOICHI, K: Periodontal healing following guided tissue regeneration with bone matrix application. J Dent Res 1992, 72(special issue):581

116. WARREN, K., KARRING, T: Guided tissue regeneration combined with osseous grafting in suprabony periodontal lesions. J Clin Periodontal 1992. 19:373-380 Surgically created horizontal defects in three dogs compared the use of a Teflon membrane (Zitex) with a fibrin sealant (Tisseel) with the membrane, sealant, and Kielbone. Various complications and exposure of the membrane occurred during the study phase Histologic analysis after 3 to 4 months of healing failed to demonstrate consistent periodontal regeneration in either of the two groups. The authors caution against the use of large membranes that may result in poor wound closure and disturbed healing, leading to a loss of the grafted materials.

117. STAHL, SS., FROUM, SJ: •• Human intrabony lesion responses to debridement. porous hydroxylapatite implants and teflon barrier membranes. J Clin Periodontol 1992. 18:605-610
A Teflon membrane and porous hydroxyapatite were used in the treatment of seven vertical lesions at seven teeth in three adults. Notches were placed in calculus and blocks were taken 16 to 28 weeks postsurgery. Histologically, two sites exhibited closure by long junctional epithelium. The remaining five sites showed gingival recession to be apical to the calculus notch for epithelium in the notch). However, cellular cememtum was seen just apical to the notch. Within the osseous crater, increased bone mass, and functionally oriented periodontal ligament was seen.

118. STAHL SS. FROUM SJ: Healing of human suprabony lesions treated with guided tissue regeneration and coronally anchored flaps: case reports. J Clin Periodontol 1991. 18: 69-74.

119.  STAHL, SS., FROUM, SJ: • • Human suprabony healing responses following root demineralization and coronal flap anchorage: histologic responses in 7 sites. J Clin Periodontol 1991. Seven suprabony lesions on anterior teeth of two patients were treated with root debridement and citric acid conditioning.  Flaps were coronally positioned or coronally anchored to orthodontic brackets bonded to enamel Block sections were taken at 7 and 18 weeks after surgery.  The average gain of attachment was 1.8 mm in CPF, and 4.5 mm with coronal anchorage.  Coronally anchored sites had new attachment in all instances. One site gained 6.7 mm in 18 weeks.  New cellular cementum was present, with attached fibers on dentin. There were varying degrees of crestal osteogenesis in all specimens.

120. WIKESJO, UME., BOOLE, GC., NILVEUS, RE: • • Periodontal repair in dogs; effect of a composite graft protocol on healing in supra alveolar periodontal defects. J Periodontol 1992, 63: 107-113. The study evaluated a composite graft protocol for treating surgically created supra-alveolar periodontal defects in beagle dogs. A comparison was made between a test group treated with citric acid and tetracycline root conditioning followed by placement of a composite graft (including hydroxyapatite. FDBA, tetracycline, and fibronectin) and flaps sutured to cover most of the crowns of the teeth and a control group of citric acid root conditioning and similarly positioned flaps. Connective-tissue repair was greater in the control group (98% vs 60% of defect height) compared with the composite graft group.  Root resorption was observed on almost all teeth with ankylosis present on two citric-acid-treated teeth. The composite graft protocol did not offer any advantages over citric acid conditioning alone as an adjunct to CPFs in this model system.

121. WANG, HL., SOMERMAN, Ml: • • Periodontal connective tissue. Curr Opin Dent 1991. 1:816-825. The review summarizes current information on proteins and factors associated with the periodontium. It also references literature on growth factors associated with periodontal tissue regeneration.

122. RIPAMONTI, U., REDDI, AH:  • Growth and morphogenetic factors in bone induction: role of osteogenin and related bone morphogenetic proteins in craniofacial and periodontal bone repair. Crit Rev Oral Biol Med 1992. 3:1-14
The developmental cascade for collagenous matrix-bone formation, including migration of progenitor cells by chemotaxis, attachment of cells through fibronectin, proliferation of mesenchymal cells, and differentiation of bone is discussed.  The bone-inductive protein osteogenin, its derivation as recombinant osteogenin and applications to oral and orthopedic surgery are presented.

123. LUYTEN, HP., YU, MY., YANAGISHITA, M., VUKICEVIC. S., HAMMONDS, RG., REDDI, AH: Natural bovine osteogenin and recombinant human bone morphogenetic Protein-2B are equipotent in the maintenance of proteoglycans in bovine articular cartilage explant cultures. J Biol Chem 1992. 267:3691-3695

124.  LYNCH, SE., DECASTILLA, GR., WILLIAMS, RC., KlRITSY, CP., HOWELL, TH. REDDY, MS., ANTONIADES, HN:  • • The effects of short-term application of a combination of platelet-derived and insulin like growth factors on periodontal wound healing. J Peridontol 1991. 62:458-467
This study on beagle dogs contrasted a test group utilizing a combination of recombinant platelet derived growth factor-B and insulin-like growth factor 1, in a methylcellulose gel, with a control group receiving the gel alone.  Two and 5 weeks postsurgery revealed a significant (five to 10-fold) increase in new bone and cementum in the test sites compared with the control gel.  A physiologic ligament space was formed between the new bone and cementum and no increase in ankylosis was seen at the test sites.

125. RUTHERFORD, RB, NIEKRASH, CE., KENNEDY, JE., CHARETTE, MF: • • Platelet-derived and insulin-like growth factors stimulate regeneration of periodontal attachment in monkeys. J Periodont Res 1992. 27:285-290
The regimen of platelet-derived and insulin-like growth factors induced regeneration of nearly 50% of lost attachment in 4 weeks in ligature-induced defects in adult monkeys. New attachment in some cases included regeneration of horizontally resorbed interdental septa.

126. MATSUDA, N., KUMAR, W-L NM., CHO, MI., GENCO, RJ: • Mitogenic, chemotactic, and synthetic responses of rat periodontal ligament fibroblastic cells to polypeptide growth factors in vitro. J Periodontol 1992, 63:515-525. Using periodontal ligament cells obtained from the coagulum of healing tooth sockets in rats, epidermal growth factor transforming growth factor β. recombinant human platelet-derived growth factor rh platelet-derived growth factor ββ, natural platelet-derived growth factor aβ, and insulin-like growth factor-1 were examined in vitro. Transforming growth factor β revealed no and epidermal growth factor slightly increased chemotactic effects. Both rh platelet-derived growth factor-a β and transforming growth factor β stimulated collagen synthesis.  Insulin-like growth factor-1 had no effect on collagen synthesis. Findings suggest that rh platelet-derived growth factor ββ and insulin-like growth factor-1 stimulate mitogenesis proliferation and chemotaxis of periodontal ligament fibroblastic cells and may thus be useful for clinical application in periodontal regenerative procedures.

127. TAKESHITA, A., ZHON GYING, NJU., HANAZAWA, S., TAKARA, I., HIGUCHI, H., KATAYAMA, I., KITANO, S: Effect of interleukin- 1β on gene expressions and functions of fibroblastic cells derived from human periodontal ligament. J Periodont Res 1992. 27:250-255

128. RIPAMONTI, U., MA, SS., REDDI, AH: • Induction of bone in composites of osteogenin and porous hydroxyapatite in baboons. Plast Reconstr Surg 1992. 89:731-739 This study examined the osteogenic potential of osteogenin in combination with porous nonresorbable and resorbable hydroxyapatite. Rods impregnated with osteogenin were implanted intramuscularly in eight adult baboons. Specimens at 30 and 90 days showed no evidence of bone in the resorbable hydroxyapatite with and without osteogenin. The nonresorbable hydroxyapatite showed evidence of bone independent of the osteogenin. Osteogenin did not increase significantly the amount of bone formation.

129. YEWEY, GL., TlPTON, AJ., SOMERMAN, MJ., DUNN, RL: • Delivery  of periodontal tissue-regeneration factors.  J Dent Res 1992. 7l (special issue):298 Fibronectin and fibroblast growth factor were incorporated in a biodegradable polymer (Atrigel) in vitro to test the efficacy of this as a delivery system. The bioactivity of fibronectin released was measured by in Vitro periodontal ligament cell attachment and recorded 45% to 94% of native fibronectin bioactivity.  Released fibroblast growth factor gave a bioactivity of 43% to 100%

130 WIKESJO, UMF., NILVEUS, RE., SELVIG, KA: •• Significance of early healing events on periodontal repair: a review. J Periodontol 1992. 63: 158-165
The authors review healing studies and conclude that connective tissue repair to the root surface following reconstructive periodontal surgery is a function of the establishment and maintenance of a stable root surface adhering fibrin clot. This clot will prevent apical migration of gingival epithelium and allow for new connective-tissue attachment without ankylosis or long junctional epithelium.

131. PITARU, S,, NOFF, M., GROSSKOPF, A., ROSES, O., TAL, H., SARION, N: •• Hcparan sulfate and fibronectin improve the capacity of  collagen barriers to prevent apical migration of the junctional epithelium.  J Periodontal 1991. 62: 598-601
Bilayered collagen barriers enriched with fibronectm and heparan sulfate were compared with non enriched bilayered collagen barriers using the GTR technique in experimentally produced osseous defects in 12 maxillary canines in eight dogs. Histologic and histomorphometric examinations 20 days postsurgery revealed a short junctional epithelium with 95% of the occlusal-apical length of the defects repopulated by connective-tissue cells in the enriched collagen group.  In the iron enriched bilayered collagen barrier and mono-layered non-enriched collagen barrier group, a long junctional epithelium developed with only 65% of defect being repopulated by connective-tissue cells.

defects repopulated by connective-tissue cells in the enriched collagen group. In the iron enriched bilavered collagen barrier und

mono-layered nonennched collagen barrier group, a lung |unc-