it is recognized that teeth with furcation involvement may undergo a more extensive and 
 rapid clinical probing attachment loss and are lost with greater frequency than 
 single - rooted teeth .
although 
 regeneration of periodontal tissue is the ultimate goal of periodontal therapy , 
 regeneration is not a predictable healing outcome after conventional treatment . with the advent of regenerative techniques ,
a new concept of periodontal treatment based 
 on the principle of cellular selection and guided tissue regeneration , permits the treatment of teeth with 
 furcation lesions by conservative surgical methods .
various studies reported gains in 
 horizontal and vertical clinical attachment , with complete closure of the furcation in 
 up to 67% of the cases using different types of mechanical barriers . despite positive results based on reports of substantial attachment 
 gain and bone fill of furcations , guided - tissue regeneration ( gtr ) is technique 
 sensitive and could be associated with increased postoperative problems . a major 
 drawback to expanded polytetrafluoroethylene
( eptfe ) and other non - resorbable membranes 
 is the need for their removal after 4 to 6 weeks , often with a surgical procedure that 
 adds to patient morbidity and may disturb the healing tissue .
bone replacement grafts have achieved similar results to gtr , demonstrating a 55% 
 overall improvement , with either complete or partial furcation fill compared to 52% for 
 gtr
. the use of bioactive glass 
 ( bg ) when compared to an eptfe barrier may produce similar clinical results .
when used in a variety of periodontal 
 and oral surgical applications , bg has shown positive clinical and histological 
 outcomes . in the past decade ,
several cytokines , or growth factors , have received attention 
 because they can regulate migration , attachment , proliferation and/or differentiation of 
 periodontal ligament cells into hard tissue cells and have been shown to enhance 
 regeneration with in vivo experiments .
a convenient 
 technique to obtain transforming growth factor  
 ( tgf ) and platelet - derived growth factor ( pdgf ) is by preparing 
 autologous platelet - rich plasma ( prp ) .
pdgf has shown to exert a favorable effect on periodontal 
 regeneration as measured by gain in clinical attachment and defect fill in 
 humans .
considering that the most favorable outcome of periodontal regenerative procedures in 
 furcations lesions has been achieved with a combination of graft material and 
 gtr , the goal of the present study was to investigate , histologically 
 and histometrically , the potential adjunctive benefits of prp when used with gtr and bg 
 in the treatment of class ii furcation lesions .
nine adult female mongrel dogs were included in the experiment ( mean weight=15 kg ) .
the 
 surgical procedures were performed under general anesthesia with intravenous injection 
 of a 3% sodium pentobarbital solution ( 0.5 ml / kg ) .
the maxillary and mandibular second 
 and fourth premolars had been bilaterally extracted previously and the extraction sites 
 had been allowed to heal for 2 months .
bilateral class ii furcation lesions were 
 surgically created at the buccal aspect of mandibular third premolars . in order to 
 create the defects
osteotomy was done in the furcation area with high - speed diamond burs with 
 constant irrigation and with nos . 1 and 2 oschenbein chisels
. the furcations lesions 
 were standardized with a millimeter probe and measured 5.0 mm in the apico - occlusal 
 direction and 2.0 mm in the buccolingual direction ( figure 1 ) , following a technique previously described .
clinical aspect of class ii furcation defect ( 5x 2 mm ) the defects were filled with gutta - percha to induce an inflammatory response due to 
 plaque accumulation for a period of one month ( figure 
 2 ) .
after this period , scaling and root planing was performed and a regimen of 
 daily brushing plus topical application of 0.1% chlorhexidine gluconate was instituted 
 for 7 days prior to the surgical procedures
. clinical aspect of class ii furcation defect filled with gutta - percha in the same day of the treatment of the mandibular defects , similar defects were created 
 at the buccal aspect of maxillary third premolars and were exposed to plaque 
 accumulation for 1 month .
forty - five days after this surgical intervention , the 
 maxillary defects were assigned to receive the same treatments .
three tubes 
 ( 5 ml ) , containing 0.5 ml of 3.2% sodium citrate solution as an anticoagulant , were 
 drawn from each dog .
the blood was thus separated into 3 basic parts : red blood cells ( at the 
 bottom of the tube ) , platelet enriched plasma ( a discrete grey line in the middle of 
 the tube ) and platelet poor plasma ( at the top of the tube ) .
the portion 
 corresponding to the platelet poor plasma was discarded from each tube and the 
 remaining content was centrifuged again at 1,200 rpm for 15 min .
four hundred 
 microliter of the middle portion , corresponding to the platelet enriched plasma , were 
 pipetted from each tube . in order to obtain the gel ,
30 l of 10% calcium chloride 
 was added to prp and heated in a water bath at 37c .
mucoperiosteal flaps were carefully reflected on the buccal aspect of the 
 experimental sites , the granulation tissue was removed by curettage , and the exposed 
 root surface was instrumented . on the root surface ,
the base of the defect was marked 
 with curettes to establish a landmark for the histomorphometric analysis .
each 
 contralateral defect in each animal was randomly assigned to one of the following 
 treatments : gtr+bg ( perioglas , us biomaterials , alachua , fl , usa ) - control group : bg particles 
 were applied filling the defect and absorbable membrane ( resolut xt 
 gore - tex , gore associates , flagstaff , az , usa ) was adapted to the defect ; gtr+bg+prp - test group : bg was incorporated to the prp ( 1:1 ) and this assemblage was 
 immediately taken to a water bath at 37c what enabled the formation of a mixture of 
 the gel and the bg graft .
this mixture was applied filling the defect and an 
 absorbable membrane ( resolut xt gore - tex , gore associates ) was adapted to 
 the defect .
primary , tension - free wound closure was accomplished following defect treatment , with 
 the gingival flaps positioned and sutured with interrupted sutures ( vicryl , ethicon 
 inc , so jos dos campos , sp , brazil ) in order to cover the membranes . immediately 
 after the surgeries , an intramuscular injection of penicillin ( 1.5 ml - 150,000 iu ) 
 was given to the animal .
forty five days after the mandibular treatment , the maxillary defects ( 2 defects/ 
 animal ) were randomly assigned to receive the same treatments ( group a and group 
 b ) .
postoperative plaque control was performed by irrigation with a solution of 1% 
 chlorhexidine gluconate ( every other day ) .
dogs were sacrificed 90 days after the 
 first treatment , under general anesthesia , with a perfusion of 10% neutral formalin 
 solution .
they were decalcified in a solution of equal parts of 50% 
 formic acid and 20% sodium citrate for 5 months .
the decalcified specimens were 
 washed in running water , dehydrated and embedded in paraffin .
mesiodistal 7-m - thick 
 sections were obtained from the tissue blocks longitudinally , in a buccolingual 
 direction , in a way to obtain a panoramic view of the furcation area .
the sections 
 were stained with hematoxylin and eosin . for histometric and descriptive histological analyses , 5 sections 
 per site representing the middle portion of the defect ( 30 m 
 apart )
were used to obtain the mean for each parameter in each dog , measured with an 
 image analysis system ( image pro plus version 3.0 , media cybernetics , silver spring , 
 md , usa ) .
the following linear measurements ( mm ) were obtained in the mesial and distal roots 
 under the furcation : 
 defect extension ( de ) : linear extension of the total root surface 
 between the two notches ; 
 new cementum ( nc ) : linear extension of the root surface covered by 
 new cementum ; 
 connective tissue adaptation ( ca ) : extension of the root surface 
 covered by connective tissue without cementum . 

new bone ( nb ) : measured from a horizontal line connecting both 
 notches ( apical limit of the defect ) to the most coronal extent of new bone .
the following 
 parameters ( mm ) were obtained by the point counting technique : 
 mineralized bone area ( mba ) : portion of the defect area filled with 
 mineralized bone tissue ; 
 residual bg particle area ( rbg ) : portion of the defect area filled 
 with bg remaining particles ; 
 non - mineralized bone area ( nmba ) : portion of the defect area filled 
 with non - mineralized tissue ; the measurements were performed using a microscope ( zeiss axioskop 2 , carl zeiss 
 instruments , gottingen , lower saxony , germany ) with a 1.25/.035 objective associated 
 with a video camera / computer / software ( image pro plus version 3.0 , media 
 cybernetics ) .
the mean 
 values for all groups were determined using the individual means from the 9 dogs . 

after testing the assumptions confirming normal distribution , two - way anova was 
 carried out to check any significance among the different treatment groups , any 
 significance among the different time periods and the interaction between them .
three tubes 
 ( 5 ml ) , containing 0.5 ml of 3.2% sodium citrate solution as an anticoagulant , were 
 drawn from each dog .
the blood was thus separated into 3 basic parts : red blood cells ( at the 
 bottom of the tube ) , platelet enriched plasma ( a discrete grey line in the middle of 
 the tube ) and platelet poor plasma ( at the top of the tube ) .
the portion 
 corresponding to the platelet poor plasma was discarded from each tube and the 
 remaining content was centrifuged again at 1,200 rpm for 15 min .
four hundred 
 microliter of the middle portion , corresponding to the platelet enriched plasma , were 
 pipetted from each tube . in order to obtain the gel ,
30 l of 10% calcium chloride 
 was added to prp and heated in a water bath at 37c .
mucoperiosteal flaps were carefully reflected on the buccal aspect of the 
 experimental sites , the granulation tissue was removed by curettage , and the exposed 
 root surface was instrumented . on the root surface , the base of the defect was marked 
 with curettes to establish a landmark for the histomorphometric analysis . each 
 contralateral defect in each animal was randomly assigned to one of the following 
 treatments : gtr+bg ( perioglas , us biomaterials , alachua , fl , usa ) - control group : bg particles 
 were applied filling the defect and absorbable membrane ( resolut xt 
 gore - tex , gore associates , flagstaff , az , usa ) was adapted to the defect ; gtr+bg+prp - test group : bg was incorporated to the prp ( 1:1 ) and this assemblage was 
 immediately taken to a water bath at 37c what enabled the formation of a mixture of 
 the gel and the bg graft .
this mixture was applied filling the defect and an 
 absorbable membrane ( resolut xt gore - tex , gore associates ) was adapted to 
 the defect .
primary , tension - free wound closure was accomplished following defect treatment , with 
 the gingival flaps positioned and sutured with interrupted sutures ( vicryl , ethicon 
 inc , so jos dos campos , sp , brazil ) in order to cover the membranes . immediately 
 after the surgeries ,
an intramuscular injection of penicillin ( 1.5 ml - 150,000 iu ) 
 was given to the animal .
forty five days after the mandibular treatment , the maxillary defects ( 2 defects/ 
 animal ) were randomly assigned to receive the same treatments ( group a and group 
 b ) .
postoperative plaque control was performed by irrigation with a solution of 1% 
 chlorhexidine gluconate ( every other day ) .
dogs were sacrificed 90 days after the 
 first treatment , under general anesthesia , with a perfusion of 10% neutral formalin 
 solution .
they were decalcified in a solution of equal parts of 50% 
 formic acid and 20% sodium citrate for 5 months .
the decalcified specimens were 
 washed in running water , dehydrated and embedded in paraffin .
mesiodistal 7-m - thick 
 sections were obtained from the tissue blocks longitudinally , in a buccolingual 
 direction , in a way to obtain a panoramic view of the furcation area .
the sections 
 were stained with hematoxylin and eosin . for histometric and descriptive histological analyses , 5 sections 
 per site representing the middle portion of the defect ( 30 m 
 apart )
were used to obtain the mean for each parameter in each dog , measured with an 
 image analysis system ( image pro plus version 3.0 , media cybernetics , silver spring , 
 md , usa ) .
the following linear measurements ( mm ) were obtained in the mesial and distal roots 
 under the furcation : 
 defect extension ( de ) : linear extension of the total root surface 
 between the two notches ; 
 new cementum ( nc ) : linear extension of the root surface covered by 
 new cementum ; 
 connective tissue adaptation ( ca ) : extension of the root surface 
 covered by connective tissue without cementum . 

new bone ( nb ) : measured from a horizontal line connecting both 
 notches ( apical limit of the defect ) to the most coronal extent of new bone .
the following 
 parameters ( mm ) were obtained by the point counting technique : 
 mineralized bone area ( mba ) : portion of the defect area filled with 
 mineralized bone tissue ; 
 residual bg particle area ( rbg ) : portion of the defect area filled 
 with bg remaining particles ; 
 non - mineralized bone area ( nmba ) : portion of the defect area filled 
 with non - mineralized tissue ; the measurements were performed using a microscope ( zeiss axioskop 2 , carl zeiss 
 instruments , gottingen , lower saxony , germany ) with a 1.25/.035 objective associated 
 with a video camera / computer / software ( image pro plus version 3.0 , media 
 cybernetics ) .
the mean 
 values for all groups were determined using the individual means from the 9 dogs . 
 after testing the assumptions confirming normal distribution ,
two - way anova was 
 carried out to check any significance among the different treatment groups , any 
 significance among the different time periods and the interaction between them .
no suppuration or 
 abscess formation was observed during the 90 days and no exposure of membranes was 
 observed in both groups during the healing period .
a clinical examination performed 
 immediately before sacrifice , disclosed that the gingival margin of all sites was 
 clinically normal and positioned coronal to the fornix of the buccal furcation .
there were no cases of epithelial downgrowth , ankylosis or extensive root resorption 
 observed in all sites .
a similar healing pattern was observed in the two groups , in both 
 periods , regarding the filling of the defect ( figure 
 3 ) .
mesiodistal histologic section of class ii furcation lesion stained with 
 hematoxilyn and eosin ( h&e ) , ( original magnification x20 ) ; a ) site of the 
 control group guided - tissue regeneration + bioactive glass ( gtr+bg ) after 45 
 days ; b ) site of the test group guided - tissue regeneration + bioactive glass + 
 platelet - rich plasma ( gtr+bg+prp ) after 45 days ; c ) site of the control group 
 after 90 days ; d ) site of the test group after 90 days .
both treatments promote 
 defect filling in both periods however a greater mineralized bone area was 
 observed for test group in both periods when compare with the control group a continuous layer of new cementum had formed in the specimens of both groups 
 extending coronally from the apical notches to a varying degree .
however , in the 
 group b , a more evident layer of new cementum was observed in the fornix region 
 ( coronal portion of the defect ) . in both groups ,
some small areas of the defects 
 presented a very dense connective tissue surrounding the remaining bg particles . 

however , connective tissue adaptation with collagen fibers running parallel to the 
 root surface seemed to be more frequent in group a. various defects in all groups presented bone surrounding bg particles and bone in 
 direct contact with these graft particles ( figure 
 4 ) .
however , in group b the defects were filled by a higher density bone 
 with a more evident mineralized bone area with islands of highly cellular bone 
 marrow . in group a , the bone presented larger marrow spaces what could indicate a 
 more immature bone when compared to group b. a ) photomicrograph of a mesiodistal histologic section of class ii furcation 
 lesion platelet - rich plasma + bioactive glass + guided - tissue regeneration 
 ( prp+bg+gtr ) 90 days after treatment defect ( masson trichrome staining , 
 original magnification x40 ) .
remaining bg particles were observed surrounded by new bone ( nb ) , 
 connective tissue ( ct ) and areas suggesting the presence of immature bone ( ib ) 
 in contact with bg ; b ) histologic section of the prp+bg+gtr defect , 90 days 
 after staining with hematoxilyn and eosin ( h&e ) , ( original magnification 
 x100 ) .
some 
 areas indicated the new bone formation occurring around bg particles 
 ( arrows ) the histometric results are shown in tables 1 
 and 2 .
data analysis showed a superior length 
 of new cementum for group b in both periods ( 45 days and 90 days ) ( p<0.05 ) .
no 
 statistically significant differences were observed between periods for the linear 
 parameters for the two groups .
the addition of prp provided a greater mineralized 
 bone area in both periods ( p<0.05 ) .
the non - mineralized bone area was greater in 
 the control group ( p<0.05 ) in both periods while no statistically significant 
 difference was observed in the area occupied by the remaining bg particles between 
 groups .
significant differences between periods were observed in the mineralized bone 
 area and non - mineralized bone area ( p<0.05 ) .
mean and standard deviation ( sd ) for the linear parameters evaluated after all 
 the treatments ( bioactive glass+guided - tissue regeneration and bioactive 
 glass+platelet - rich plasma+guided - tissue regeneration ) in both periods .
capital letters compare treatments within each time and non - capital letter 
 compare times within each treatment .
means followed by different letters are 
 statistically different mean and standard deviation ( sd ) for the area parameters evaluated after all 
 the treatments ( bioactive glass+guidedtissue regeneration and bioactive 
 glass+platelet - rich plasma+guided - tissue regeneration ) in both periods capital letters compare treatments within each time and non - capital letter 
 compare times within each treatment .
no suppuration or 
 abscess formation was observed during the 90 days and no exposure of membranes was 
 observed in both groups during the healing period .
a clinical examination performed 
 immediately before sacrifice , disclosed that the gingival margin of all sites was 
 clinically normal and positioned coronal to the fornix of the buccal furcation .
there were no cases of epithelial downgrowth , ankylosis or extensive root resorption 
 observed in all sites . no inflammation and foreign body reactions indicating toxicity 
 were observed .
a similar healing pattern was observed in the two groups , in both 
 periods , regarding the filling of the defect ( figure 
 3 ) .
mesiodistal histologic section of class ii furcation lesion stained with 
 hematoxilyn and eosin ( h&e ) , ( original magnification x20 ) ; a ) site of the 
 control group guided - tissue regeneration + bioactive glass ( gtr+bg ) after 45 
 days ; b ) site of the test group guided - tissue regeneration + bioactive glass + 
 platelet - rich plasma ( gtr+bg+prp ) after 45 days ; c ) site of the control group 
 after 90 days ; d ) site of the test group after 90 days .
both treatments promote 
 defect filling in both periods however a greater mineralized bone area was 
 observed for test group in both periods when compare with the control group a continuous layer of new cementum had formed in the specimens of both groups 
 extending coronally from the apical notches to a varying degree .
however , in the 
 group b , a more evident layer of new cementum was observed in the fornix region 
 ( coronal portion of the defect ) . in both groups ,
some small areas of the defects 
 presented a very dense connective tissue surrounding the remaining bg particles . 

however , connective tissue adaptation with collagen fibers running parallel to the 
 root surface seemed to be more frequent in group a. various defects in all groups presented bone surrounding bg particles and bone in 
 direct contact with these graft particles ( figure 
 4 ) .
however , in group b the defects were filled by a higher density bone 
 with a more evident mineralized bone area with islands of highly cellular bone 
 marrow . in group a , the bone presented larger marrow spaces what could indicate a 
 more immature bone when compared to group b. a ) photomicrograph of a mesiodistal histologic section of class ii furcation 
 lesion platelet - rich plasma + bioactive glass + guided - tissue regeneration 
 ( prp+bg+gtr ) 90 days after treatment defect ( masson trichrome staining , 
 original magnification x40 ) .
remaining bg particles were observed surrounded by new bone ( nb ) , 
 connective tissue ( ct ) and areas suggesting the presence of immature bone ( ib ) 
 in contact with bg ; b ) histologic section of the prp+bg+gtr defect , 90 days 
 after staining with hematoxilyn and eosin ( h&e ) , ( original magnification 
 x100 ) .
some 
 areas indicated the new bone formation occurring around bg particles 
 ( arrows )
data analysis showed a superior length 
 of new cementum for group b in both periods ( 45 days and 90 days ) ( p<0.05 ) .
no 
 statistically significant differences were observed between periods for the linear 
 parameters for the two groups .
the addition of prp provided a greater mineralized 
 bone area in both periods ( p<0.05 ) .
the non - mineralized bone area was greater in 
 the control group ( p<0.05 ) in both periods while no statistically significant 
 difference was observed in the area occupied by the remaining bg particles between 
 groups .
significant differences between periods were observed in the mineralized bone 
 area and non - mineralized bone area ( p<0.05 ) .
mean and standard deviation ( sd ) for the linear parameters evaluated after all 
 the treatments ( bioactive glass+guided - tissue regeneration and bioactive 
 glass+platelet - rich plasma+guided - tissue regeneration ) in both periods .
capital letters compare treatments within each time and non - capital letter 
 compare times within each treatment .
means followed by different letters are 
 statistically different mean and standard deviation ( sd ) for the area parameters evaluated after all 
 the treatments ( bioactive glass+guidedtissue regeneration and bioactive 
 glass+platelet - rich plasma+guided - tissue regeneration ) in both periods capital letters compare treatments within each time and non - capital letter 
 compare times within each treatment .
the present study evaluated if the application of prp on class ii furcation lesions 
 would enhance periodontal regeneration when associated with gtr and bioactive glass 
 ( bg ) .
clinical reports have demonstrated that the association of gtr , bone substitutes 
 and prp could provide good results in the treatment of intrabony defects and class ii 
 furcation lesions , as revealed by gain in clinical attachment , defect fill at reentry , 
 reducing probing depth , vertical defect fill and horizontal defect fill .
however , to the best of our knowledge , this is the first study that 
 evaluated histologically the effect of the association of prp with gtr and bg on 
 periodontal regeneration of class ii furcation lesions in dogs .
it has been demonstrated that when a bone graft or substitute is combined with the gtr 
 procedure in furcation lesions , a greater amount of defect fill is observed .
this advantage can be explained by the 
 osteoconductive and/or osteoinductive properties of the graft materials .
one of the bone 
 substitutes that have been investigated is the bg , a material that has been suggested as 
 an osteoconductive graft .
the use of bg in the present study was not associated with any 
 foreign body reaction .
it was observed that some remaining bg granules showed 
 dissolution of their core with the formation of new bone , as described by vogel , et 
 al .
the sites treated with grt+bg ( group a ) showed new cementum formation and new bone 
 regeneration in both periods .
these results are in accordance with previous reports , 
 which demonstrated a beneficial effect of the association of gtr and bone 
 substitutes .
however , clinical studies do not provide information 
 about the nature of the healing process after these procedures .
in contrast with previous studies , in the present investigation , there were no instances of epithelial 
 downgrowth in all sites .
( 2007 ) observed a correlation between the exposure of the 
 membrane and the epithelial migration .
the presence of epithelial tissue was observed in 
 all histological sections of the sites that shown a membrane exposure and was ascribed 
 to the degradation of the exposed portions of this material .
the use of prp for periodontal regeneration has been based on the idea that this 
 preparation contains polypeptide growth factors ( pgfs ) .
some specific pgfs , like pdgf 
 and tgf , could promote the growth and differentiation of the periodontal ligament and 
 alveolar bone cells and could be responsible for the clinical improvement observed in 
 experimental sites
. other interesting feature of prp is its sticky 
 consistency due to its high fibrin content .
the fibrin component of prp may work as a 
 hemostatic agent aiding in stabilizing the graft material and the blood clot .
blood clot immobilization has been 
 suggested as an important event in the early phases of wound healing of periodontal 
 regenerative procedures .
it has been previously shown that prp may stimulate pdl cells and fibroblastic cell 
 proliferation but may suppress epithelial cell division in 
 vitro .
consequently , by 
 ordering these cellular responses into a series of related events , prp may facilitate 
 wound - healing and tissue regeneration . in the present study ,
the sites in the group b 
 showed a superior cementum formation , in both observation periods , when compared with 
 the sites of group a ( p<0.05 ) . to the best of our knowledge ,
no study provided direct 
 evidence that the prp could stimulate the differentiation of cells from the periodontal 
 ligament to cementoblasts , inducing cementogenesis .
however , previous investigations 
 have suggested that the treatment with recombinant growth factors , such as pdgf and igf 
 may enhance the formation of cementum on denuded dentin surfaces .
in addition to it , a similar result was 
 observed when the prp was associated with a subepithelial connective tissue graft for 
 the treatment of gingival recessions created in dogs .
the positive effect of prp in the bone response was demonstrated by previous 
 studiesand could be evidenced in the present study by the 
 largest area of mineralized bone observed in group b. sites treated with prp showed a 
 superior mineralized bone area with a concomitant inferior non - mineralized bone area , in 
 both periods , when compared with group a ( p<0.05 ) . however , the difference between 
 treatments is more evident in the first period ( 45 days ) when the area of mineralized 
 bone in group b was almost twice the one observed in group a. this observation is in 
 agreement with an in vitro study that suggested that pdgf acts upon osteoblastic cell 
 proliferation , exerting most of its effects during the early phase of wound healing . 

although the mineralized bone formation was superior in group b of the present study , it 
 should be noticed that recent animals studies could not demonstrate that prp increases 
 bone regeneration when used in peri - implant bone defects . to date
, there is insufficient information about growth factor interactions and how they 
 influence the activations of gene expression and protein production .
also , the ideal 
 platelet and growth factor concentrations to promote periodontal regeneration has not 
 been established .
( 2003 ) reported that growth factors may act at specific times and at appropriate 
 concentrations and this may be other explanation for the different results in studies 
 with prp . in the present study ,
the achieved percentage mean of platelet concentration 
 was 227.02 % ( mean of 463,015 platelets/l ) in relation to blood platelet count .
the technique to produce prp used in this study was previously described by suaid , et 
 al.(2008 ) .
this technique 
 enables to produce a platelet concentration generally four times greater than the whole 
 blood .
( 2003 ) 
 performed a study that measured the concentrations of bfgf , vegf , pdgf - bb and tgf - b1 
 released from platelet concentrate and whole blood , before and after the addition of 
 calcium alone , thrombin alone and various concentrations of calcium and thrombin .
they 
 observed that calcium chloride , regardless of the use of thrombin , released platelet 
 growth factors . in this study ,
the concentration of growth factors that was released was 
 sufficient to promote endothelial cell proliferation in vitro .
since the use of thrombin in this type 
 of investigation is not allowed in our country , instead of thrombin , 10% calcium 
 chloride was added to prp and heated in a water bath at 37c .
within the limits of this animal study , it was concluded that prp in association with 
 gtr and bg may enhance the amount of new cementum and provide a more mineralized bone 
 ( mature ) in a shorter period of time when applied to the treatment of class ii furcation 
 lesions in dogs .
future studies are required to clarify the mechanism of action of prp 
 in the periodontal regeneration process and the ideal platelet and growth factor 
 concentrations to adequately promote periodontal regeneration .
the authors thank the financial support of fapesp ( the state of so paulo research 
 foundation , # 04/12428 - 6 ) and cnpq ( national counsel of technological and scientific 
 development # 300817/2007 - 0 ) .
 objective this study was designed to evaluate the potential adjunctive benefits of 
 platelet - rich plasma ( prp ) when used with guided - tissue regeneration ( gtr ) and 
 bioactive glass ( bg ) in the treatment of class ii furcation lesions . 
 material and methods bilateral class ii furcation lesions were surgically created and allowed to 
 become chronic in the mandibular third premolars of 9 dogs . 
 the defects were 
 randomly assigned to : a ) gtr+bg and b ) gtr+bg+prp . 
 similar defects were created in 
 the maxillary third premolars and received the same treatments after 45 days . 
 dogs 
 were sacrificed 90 days after the first treatment . 
 the histometric parameters 
 evaluated were : connective tissue adaptation , new cementum , new bone , mineralized 
 bone area , non - mineralized bone area , and residual bg particle area . 
 results data analysis showed a superior length of new cementum and a greater mineralized 
 bone area for group b in both periods ( p<0.05 ) . 
 the non - mineralized bone area 
 was greater in the control group ( p<0.05 ) in both periods . 
 conclusion within the limits of this study , it can be concluded that the use of prp in the 
 treatment of class ii furcation defects may enhance the amount of new cementum and 
 provide a more mineralized bone in a shorter period of time . 
INTRODUCTION
MATERIAL AND METHODS
PRP Preparation
Defect treatment
Linear measurements
Area measurements
Statistical analysis
RESULTS
Clinical observations
Histological observations
Histometric measurements
DISCUSSION
CONCLUSIONS
ACKNOWLEDGEMENTS