Original Article http://dx.doi.org/10.1590/1678-7757-2016-0599 Apical Negat ive Pressure irrigat ion present s t issue com pat ibilit y in im m at ure t eet h Abst ract Carolina Maschietto PUCINELLI1 Aim : To com par e t he apical negat ive pr essur e ir r igat ion ( ANP) w it h Léa Assed Bezerra da SILVA1 convent ional irrigat ion in t he t eet h of im m at ure dogs wit h apical periodont it is. Nestor COHENCA2 Priscilla Coutinho ROMUALDO1 Met hods: Fift y- t wo im m at ure pre- m olar root canals were random ly assigned int o 4 groups: ANP ( n= 15) ; convent ional irrigat ion ( n= 17) ; healt hy t eet h ( cont rol) ( n= 10) ; and t eet h wit h unt reat ed apical periodont it is ( cont rol) Raquel Assed Bezerra da SILVA1 ( n= 10) . Aft er induct ion of apical periodont it is, t eet h were inst rum ent ed using Alberto CONSOLARO3 EndoVac® ( apical negat ive pressure irrigat ion) or convent ional irrigat ion. Alexandra Mussolino de QUEIROZ1 The anim als were eut hanized aft er 90 days. The sect ions were st ained by Paulo NELSON-FILHO1 +( DQG DQDO\]HG XQGHU FRQYHQWLRQDO DQG ÀXRUHVFHQFH PLFURVFRS\ 75$3 hist oenzym ology was also perform ed. St at ist ical analyses were perform ed ZLWK WKH VLJQL¿FDQFH OHYHO VHW DW  5HVXOWV 7KHUH ZDV GLIIHUHQFH LQ t he hist opat hological param et ers bet w een ANP and convent ional groups ( p< 0 . 0 5 ) . Th e ANP gr ou p sh ow ed a pr edom in an ce of low m agn it u de LQÀDPPDWRU\LQ¿OWUDWHDVPDOOHUSHULRGRQWDOOLJDPHQWDQGORZHUPLQHUDOL]HG t issue resorpt ion. There were no differences in t he periapical lesion ext ensions bet w een t he ANP and convent ional gr oups ( p> 0.05) . How ever, a low er num ber of ost eoclast s was observed in t he ANP group ( p< 0.05) . Conclusion: The EndoVac® irrigat ion syst em present ed bet t er biological result s and m ore advanced repair process in im m at ure t eet h wit h apical periodont it is t han t he FRQYHQWLRQDOLUULJDWLRQV\VWHPFRQ¿UPLQJWKHK\SRWKHVLV Keyw ords: Apical negat ive pressure irrigat ion. Apical periodont it is. Apical posit ive pressure irrigat ion. I m m at ure t eet h. Submitted: November 22, 2016 0RGL¿FDWLRQ$SULO Accepted: June 02, 2017 Corresponding address: Paulo Nelson Filho Universidade de São Paulo Faculdade de Odontologia de Ribeirão Preto. Avenida do Café S/N, 14040-904. Ribeirão Preto - SP - Brazil. Phone: +55-16-3315-3995 - Fax: +55-16-3315-4102. e-mail: nelson@forp.usp.br 1 Universidade de São Paulo, Faculdade de Odontologia de Ribeirão Preto, Departamento de Clínica Infantil, Ribeirão Preto, SP, Brasil. 2 University of Washington & Seattle Children’s, Department of Endodontics, Seattle, USA. 3 Universidade de São Paulo, Faculdade de Odontologia de Bauru, Departamento de Cirurgia, Estomatologia, Patologia e Radiologia, Bauru, SP, Brasil. J Appl Oral Sci. 612 2017;25(6):612-9 Apical Negative Pressure irrigation presents tissue compatibility in immature teeth Three beagle dogs, 4 m ont hs old, were used. Upper I nt roduct ion ( second and t hir d) and low er ( second, t hir d, and I nfect ion cont rol in endodont ic t herapy is ext rem ely fourt h) im m at ure prem olars were included. Fift y- t wo im port ant t o achieve a successful out com e in t eet h root s were random ly divided int o 4 groups, as follows: wit h apical periodont it is7,16,25,27 . This can be obt ained t h r o u g h t h e st e p s i n v o l v e d i n t h e e n d o d o n t i c t r eat m en t 1 5 , in clu din g ir r igat ion 8 . Th e associat ion bet w een ir r igat ion and biom echanical pr eparat ion opt im izes t he cleaning of root canals6,22 . Alt hough t he conv ent ional ir r igat ion sy st em is widely used 12 , in 2007 a novel syst em called EndoVac® ANP ( Apical Negat ive Pressure) : ( n= 15) . Co n v e n t i o n a l I r r i g a t i o n ( Po si t i v e Pr e ssu r e ) : ( n= 17) . Healt hy Teet h ( Negat ive Cont rol) : ( n= 10) Teet h wit h unt reat ed apical periodont it is ( Posit ive Cont rol) : ( n= 10) . $OOWHHWKKDGDQ;UD\WDNHQWRFRQ¿UPDQRSHQ ( Discus Dent al, Culver Cit y, CA, USA) was launched t o ap ex . Th e cor on al access w as p er f or m ed in t h e t he dent al m arket . I nst ead of posit ive pressure, t he En d oVac ® , con v en t ion al, an d ap ical p er iod on t it is EndoVac® syst em uses an apical negat ive pressure groups. Aft er pulp t issue rem oval, root canals were irrigat ion ( ANP) and has been considered a prom ising left exposed in t he oral cavit y for 7 days for m icrobial disinfect ion prot ocol in t he endodont ic lit erat ure 8,23 . cont am inat ion, as recom m ended by Leonardo, et al. 17 Previous st udies dem onst rat ed t hat t he ANP decreases ( 1993) . I n order t o prom ot e t he induct ion of apical t he risk of irrigant solut ion ext rusion t hrough t he apical periodont it is, t he pulp cham ber was sealed wit h zinc foram en 9, 14, 26 . This sy st em enables t he cir culat ion oxide eugenol cem ent ( SS Whit e, Rio de Janeiro, RJ, of irrigant solut ion t o all working lengt hs ( WL) 2 and Brazil) . Bone t hinning occurs bet ween 15 and 25 days facilit at es m icrobiological cont rol 13,23 . The ANP is also in im m at ure dog t eet h 17 . HI¿FLHQWLQUHPRYLQJWKH³VPHDUOD\HU´10,18 and debris, m ost ly at t he apical t hird of a root canal 1,4,10,11 . Af t er t h is p er iod , a r u b b er d am w as u sed t o isolat e t he t eet h and t he t em porary rest orat ion was The anat om y of t he apical t hird in t he root canals r em oved. Root canal disinfect ion was per for m ed 3 of im m at ur e t eet h incr eases t he r isk of accident al m m short er t han t he radiographic apex, followed by in j ect ion of ir r ig an t solu t ion in t o t h e p er iap ical WL det er m inat ion, est ablished 1 m m shor t er t han t issues3,26 . However, an in vit ro st udy dem onst rat ed t he radiographic apex. The com plem ent at ion of root t hat open apex t eet h had sim ilar ext rusion t o closed canal disinfect ion was perform ed at t he WL. ANP and apex t eet h when t he ANP was used 20 . convent ional groups were inst rum ent ed wit h K- t ype To dat e, no in vivo st udies com paring t he EndoVac® ¿OHV WR WKH :/ $W HDFK ¿OH H[FKDQJH URRW FDQDOV sy st em w i t h t h e con v en t i on al i r r i g at i on sy st em were irrigat ed wit h 10 m L of NaOCl 2.5% for bot h t he separat ely in im m at ure t eet h have been published. EndoVac® syst em and convent ional syst em . Th er ef or e, t h e aim of t h is in v iv o st u d y w as t o Th e r ecom m en d ed p r ot ocol f or t h e En d oVac ® perform t he hist opat hological and hist oenzym ological syst em includes 2 m ain st eps: m acro- irrigat ion and evaluat ion t o com par e t he EndoVac ® sy st em w it h m icro- irrigat ion. I n t his part icular st udy wit h im m at ure t he convent ional ir r igat ion in im m at ur e dog t eet h t eet h, canals were irrigat ed using t he m acro- cannula wit h experim ent ally induced apical periodont it is. The only aft er det erm ining t he apical size of t he canal hy pot hesis is t hat ANP pr esent ed bet t er biological due t o t he large apical size, as previously published 23 . result s in im m at ure t eet h wit h apical periodont it is t han Finally, root canals were irrigat ed wit h saline solut ion, t he convent ional irrigat ion syst em . dried wit h absorbent paper point s and sealed wit h ProRoot MTA ( Dent sply Tulsa Dent al, Johnson Cit y, TN, USA) and silver am algam ( Sybraloy, Kerr Corporat ion, Orange, CA, USA) . Mat erial and m et hods HE- st aining Th i s r e se a r ch p r o j e ct w a s a p p r o v e d b y t h e Aft er 90 days, t he anim als were eut hanized. The I nst it ut ional Anim al Et hics Com m it t ee ( # 006/ 2012) . m axillas and m andibles wit h t eet h were dissect ed and All t he experim ent al procedures were perform ed as sect ioned t o obt ain individual root s. The hist ot echnical in our previous st udies8,24 . procedures were perform ed as previously published 5,8 . J Appl Oral Sci. 613 2017;25(6):612-9 PUCINELLI CM, SILVA LAB, COHENCA N, ROMUALDO PC, SILVA RAB, CONSOLARO A, QUEIROZ AM, NELSON-FILHO P The HE- st ained sect ions were analyzed in an Axio St at ist ical Analysis I m ager.M1 m icroscope ( Zeiss, Göt t ingen, Germ any) , The GraphPad Prism 5.a ( GraphPad Soft ware I nc., using scores based on t he following hist opat hological San Diego, CA, USA) was used for st at ist ical analyses. SDUDPHWHUV D  LQÀDPPDWRU\ LQ¿OWUDWH DEVHQW RU Chi- square or Fisher ’s exact t est were used t o evaluat e m ild ( scor e 1) , m oderat e or sever e ( scor e 2) ; ( b) t he scores. One- way ANOVA wit h Tukey’s post hoc t hickness of periodont al ligam ent : norm al ( score 1) , t est was used t o evaluat e m ean difference. The level slight ly incr eased ( scor e 2) , m oderat ely incr eased RIVLJQL¿FDQFHZDVVHWDW ( score 3) , or severely increased ( score 4) ; and ( c) process of resorpt ion of m ineralized t issues: absent ( score 1) or present ( score 2) . I n addit ion, descript ions of t he apical and periapical regions were conduct ed for each group. Fluorescence m icroscope m orphom et ry I n t he ANP, convent ional, and apical periodont it is groups, t he area of t he periapical lesion was m easured in square m illim et ers in t he HE- st ained sect ions using DQ$[LR,PDJHU0PLFURVFRSHDWîPDJQL¿FDWLRQ DQGRSHUDWLQJLQWKHÀXRUHVFHQFHPRGHDVSUHYLRXVO\ described 5,19,24 . I n t he healt hy t eet h group, t he t hickness of t he healt hy periodont al ligam ent area was m easured by drawing a line perpendicular t o t he root apex, locat ed 0.5 m m above t he opening of each specim en, t o delim it t he m axim um height of t he m easured area. Ta r t r a t e - r e s i s t a n t a c i d p h o s p h a t a s e hist oenzym ology ( TRAP) The TRAP act iv it y w as per for m ed t o m ar k t he ost eoclast s 8 , 1 9 . Th e sect ion s w er e depar af f in ized, hydrat ed, and placed in a solut ion of 50% et hanol/ acet one for 1 m inut e and dried at room t em perat ure. Ne x t , a b u f f e r so l u t i o n co n t a i n i n g a ce t i c a ci d , dim et hylform am ide, Fast Red, and phosphoric acid napht hol AS- BI ( Sigm a-Aldrich Corporat ion, St . Louis, MO, USA) w as pipet t ed ov er t h e sect ion s, w h ich were m aint ained at 37° C for 40 m inut es prot ect ed fr om light . The count er- st ain w it h Fast Gr een was Result s Microscopic analysis of apical and periapical regions Th e ANP gr ou p pr esen t ed m ix ed an d dif f u sed LQÀDPPDWRU\ LQ¿OWUDWH UDQJLQJ IURP PLOG   t o m oderat e ( 26.7% ) . The periapical region showed rich neovascularizat ion and bet t er repair process wit h ¿EUREODVWSUROLIHUDWLRQ7KHUHZDVSDUWLDOO\PLQHUDOL]HG connect ive t issue in t he r egion of t he lar ge apical foram en t o t he m iddle t hird of t he root canal. This t issue or iginat ed fr om t he per iodont al t issue t hat invaginat ed int o t he root canal in 53.6% of t he cases, ZLWKDGHQVHSUHVHQFHRI¿EUREODVWVDQGEORRGYHVVHOV I n 80% of t he specim ens t here was no dent in, bone, or cem ent um resorpt ion ( Figure 1) . I n t he convent ional group, t here were m ixed and GLIIXVHGLQÀDPPDWRU\LQ¿OWUDWHLQDOOFDVHV   The m aj orit y of t he cases ( 58.8% ) were m oderat ely in f lam ed. Th e per iodon t al ligam en t w as sev er ely increased in 82.4% of t he cases, wit h areas of edem a DQG¿EULOODUGLVVRFLDWLRQ,QRIWKHFDVHVWKHUH was severe bone, cem ent um , and dent in resorpt ion. I n som e cases, t here were no cem ent oblast s on t he cem ent surface and unrepaired root resorpt ion was also frequent ( Figure 2A and B) . All param et er s ( pulp t issue, odont oblast lay er, periodont al ligam ent , and alveolar bone) in t he healt hy t eet h group were norm al ( Figure 2C) . perform ed. The sam ples were exam ined under t he Axio I m ager. M1 m icroscope under convent ional light t o count t he num ber of m ult inucleat e TRAP–posit ive cells present in t he resorpt ion lacunae t hat were in direct cont act wit h t he alveolar bone around t he periapical lesion. There was an invaginat ion of part ially m ineralized con n ect iv e t issu e in t o t h e r oot can al in t h e ANP specim ens. I n t he healt hy specim ens, t he ost eoclast count r egion was est ablished as descr ibed for t he ÀXRUHVFHQFH PLFURVFRSH PRUSKRPHWU\ $OO UHVXOWV were expressed in cell num bers. J Appl Oral Sci. I n t he apical periodont it is group, t he apical and per iapical r egion s pr esen t ed sev er ely m ix ed an d d if f u sed in f lam m at or y in f ilt r at e. Th e p er iod on t al ligam ent was severely increased wit h int ense edem a DQG¿EULOODUGLVVRFLDWLRQ,QWKHFHPHQWXPVXUIDFHWKH resorpt ion areas were not repaired. The alveolar bone was dist ant from t he root apex, indicat ing advanced bone resorpt ion. There were no ost eoblast s on t he su r f ace an d ost eoclast s w er e f r eq u en t ly p r esen t ( Figure 2D) . I n f l am m at o r y i n f i l t r at e sco r es d em o n st r at ed 614 2017;25(6):612-9 Apical Negative Pressure irrigation presents tissue compatibility in immature teeth VLJQL¿FDQWGLIIHUHQFHEHWZHHQWKH$13DQGFRQYHQWLRQDO an d con v en t ion al gr ou ps. St at ist ically sign if ican t groups ( p= 0.03) . Regarding t he periodont al ligam ent , difference was also observed bet ween t hese groups difference was observed ( p= 0.02) bet ween t he ANP ( p= 0.003) , regarding m ineralized t issue resorpt ion Figure 1- Representative photomicrographs of the ANP group, 90 days after the endodontic treatment, in conventional light microscopy: (A) Panoramic view of the periapical and apical regions showing intense invagination of the connective tissue into the root canal (HE, Zeiss, 5X). (B) Detail of panel A: part of the invaginated tissue after the mineralization (HE, Zeiss, 20X). (C) Panoramic view of the periapical and apical regions showing that the periodontal ligament was slightly increased (HE, Zeiss, 5X). (D) Photomicrography of the SHULDSLFDODQGDSLFDOUHJLRQVZLWK¿EHUVYHVVHOVDQGPLOGLQÀDPPDWRU\FHOOV +(=HLVV; +( KHPDWR[\OLQ HRVLQ Figure 2- Representative photomicrographs of the conventional, healthy, and apical periodontitis groups, 90 days after the endodontic treatment, in conventional light microscopy: (A) Panoramic view of the apical and periapical regions of the conventional group showing an increased periodontal ligament (HE, Zeiss, 1.25X). (B) Detail of the alveolar bone of the conventional group with no osteoblast in its surface and presence of osteoclast (HE, Zeiss, 20X). (C) Panoramic view of the healthy teeth group with incomplete root formation, normal pulp, and an odontoblastic layer. Periodontal ligament and alveolar bone were healthy (HE, Zeiss, 10X). (D) Panoramic view of the apical SHULRGRQWLWLVJURXSVKRZLQJDIRFDOVHYHUHLQÀDPPDWRU\LQ¿OWUDWH +(=HLVV; +( KHPDWR[\OLQ HRVLQ J Appl Oral Sci. 615 2017;25(6):612-9 PUCINELLI CM, SILVA LAB, COHENCA N, ROMUALDO PC, SILVA RAB, CONSOLARO A, QUEIROZ AM, NELSON-FILHO P Table 1-5HVXOWVIRU,QÀDPPDWRU\LQ¿OWUDWHWKLFNQHVVRISHULRGRQWDOOLJDPHQWDQGUHVRUSWLRQRIPLQHUDOL]HGWLVVXHEHWZHHQWKHJURXSV Groups Scores EndoVac® Conventional p-Value Healthy Apical EndoVac® EndoVac® EndoVac® Periodontitis x x x Conventional Conventional x Conventional Healthy Apical Healthy Periodontitis x Apical Periodontitis ,QÀDPPDWRU\LQ¿OWUDWH Absent or Mild Moderate or Severe                 Normal Slightly increased Moderately increased Severely increased                                 Present Absent             0.03 0.06 0.001 0.0002 0.263 ” 0.005 ” 0.273 ” ” 0.263 Thickness of periodontal ligament 0.02 Process of resorption of mineralized tissues     0.003 0.250 Figure 3- Photomicrographs of microscopic sections representing the 4 groups evaluated, 90 days after the endodontic treatment: (A) 5HSUHVHQWDWLYH SKRWRPLFURJUDSKV RI WKH FRQYHQWLRQDO JURXS +(6WDLQHG DQG REVHUYHG XQGHU ÀXRUHVFHQFH PLFURVFRS\ +( =HLVV ;  % 5HSUHVHQWDWLYHSKRWRPLFURJUDSKVRIWKH$13JURXS+(6WDLQHGDQGREVHUYHGXQGHUÀXRUHVFHQFHPLFURVFRS\ +(=HLVV 1.25X). HE = hematoxylin & eosin ( Table 1) . Fluorescence m icroscopy m orphom et ry The m ean lesion size was 12.94 ( ± 7.73) m m 2 in t he ANP group, 17.91 ( ± 8.85) m m 2 in t he convent ional g r o u p , a n d 2 1 . 4 7 ( ± 1 . 4 8 ) m m 2 i n t h e a p i ca l periodont it is group. I n t he healt hy t eet h group, t he periodont al ligam ent area in t he apical region was 0.67 m m 2 ( ± 0.38) m m 2 7KHUHZDVQRVLJQL¿FDQWGLIIHUHQFH bet ween t he ANP and convent ional groups ( p> 0.05) . Figure 3 shows represent at ive phot om icrographs of t he GLIIHUHQWJURXSVDIWHUÀXRUHVFHQFHPLFURVFRS\ $%  TRAP hist oenzym ology The m eans for t he ost eoclast count s were 26.25 ( ± 18.78) for t he ANP group, 50.94 ( ± 26.74) for t he J Appl Oral Sci. Figure 4- Distribution of the number of osteoclasts. The different OHWWHUVUHSUHVHQWJURXSVZLWKVLJQL¿FDQWGLIIHUHQFH S!  616 2017;25(6):612-9 Apical Negative Pressure irrigation presents tissue compatibility in immature teeth Figure 5- Photomicrographs of microscopic sections representing the 4 groups evaluated, 90 days after the endodontic treatment, stained E\75$3WHFKQLTXHIRULGHQWL¿FDWLRQDQGFRXQWRIRVWHRFODVWLFFHOOV $ 5HSUHVHQWDWLYH SKRWRPLFURJUDSKV RI WKH $SLFDO 3HULRGRQWLWLV group, where it was observed that the apex and alveolar bone had intense presence of osteoclasts (Zeiss, 5X). (B) Image A detail, highlighting intense presence of osteoclasts (Zeiss, 20X). (C) Representative photomicrographs of healthy teeth group, where there is healthy tissues with reduced presence of osteoclasts (Zeiss, 5X). (D) Representative photomicrograph of the conventional irrigation group. The surface of the alveolar bone showing moderate presence of osteoclasts, albeit in larger quantity than in the ANP group (Zeiss, 40X). (E) Representative photomicrograph of the ANP group with slightly increased presence of osteoclasts (Zeiss, 5X). (F) Detail of image E, highlighting the osteoclasts in the bone surface (Zeiss, 40X). convent ional group, 7.9 ( ± 4.99) for t he healt hy t eet h for t eet h wit h closed apex. However, t here are no in group, and 103 ( ± 23.27) for t he apical periodont it is vivo st udies t hat evaluat ed t he EndoVac® syst em in JURXS )LJXUH 6LJQL¿FDQWGLIIHUHQFHZDVREVHUYHG t eet h wit h im m at ure root form at ion, com pared wit h bet ween t he ANP and convent ional groups ( p< 0.0001) . convent ional irrigat ion separat ely. Thus, t his in vivo Figure 5 shows represent at ive phot om icrographs st udy aim ed t o add im port ant inform at ion t o provide of all groups aft er TRAP hist oenzym ology. VFLHQWL¿FEDFNJURXQGIRUWKHFOLQLFDODSSOLFDWLRQRI$13 in t eet h wit h open apex. This is ext rem ely im port ant t o ach iev e a su ccessfu l ou t com e post - en dodon t ic t reat m ent in im m at ure t eet h wit h apical periodont it is. Discussion Th i s h i st o p a t h o l o g i ca l st u d y co m p a r e d t w o Our research group’s previous st udy 8 dem onst rat ed t h a t t h e AN P p r e s e n t e d m a n y a d v a n t a g e s i n com parison wit h t he convent ional irrigat ion syst em J Appl Oral Sci. different t ypes of root irrigat ion syst em . The init ial experim ent al st ages evaluat ion enables t o observe DQDFXWHLQÀDPPDWRU\UHVSRQVHI n vivo m et hodology 617 2017;25(6):612-9 PUCINELLI CM, SILVA LAB, COHENCA N, ROMUALDO PC, SILVA RAB, CONSOLARO A, QUEIROZ AM, NELSON-FILHO P LQYROYHVGLI¿FXOWLHVLQREWDLQLQJWKHDQLPDOVKLJKFRVW m ight increase t his success rat e. t echnique, as well as et hical im plicat ions. For t hese There was no st at ist ical difference bet ween t he reasons, we decided t o evaluat e a longer period t o En doVac ® an d conv en t ion al gr ou ps r egar din g t h e observe t he occurrence of persist ent inj ury t o t issues r esult s of t he post - ir r igat ion per iapical lesion ar ea DQGFRQVHTXHQWO\WKHLQÀDPPDWRU\UHVSRQVHLQWKH PHDVXUHPHQWLQWKHÀXRUHVFHQFHDVVHVVPHQW2QWKH lat e st age aft er use of different root irrigat ion syst em s. ot her hand, t hrough t he hist opat hological analysis of This m et hodology represent s t he “ gold st andard” of sect ions st ained wit h HE, advanced st age t issue repair t issue response against t he use of different m at erials in t he ANP specim ens was ev ident . The per iapical or t echniques. I n t he endodont ic lit erat ure, only 2 lesions t hat were present in som e specim ens of t he papers8,23 evaluat ed t he m icroscopic response of t he ANP group did not com plet ely repair and present ed ® EndoVac syst em in vivo using dog t eet h. Silva, et al. 23 ( 2010) com pared t he revascularizat ion and t he apical and periapical repair in im m at ure dog t eet h wit h periapical lesions aft er irrigat ion wit h t he EndoVac ® syst em and convent ional irrigat ion plus t riant ibiot ic an ext ensive lesion area. This m ay explain t he lack RIVLJQL¿FDQWGLIIHUHQFHEHWZHHQWKHOHVLRQDUHDVLQ t he ANP and convent ional groups in t he assessm ent XQGHUÀXRUHVFHQFH Bone is a dynam ic tissue with continuous rem odeling LQWUDFDQDOGUHVVLQJ$OWKRXJKRQO\WKHLQÀDPPDWRU\ process. I n pat hological condit ions, such as chronic in f ilt r at e w as sig n if ican t ly d if f er en t b et w een t h e apical periodont it is, bone resorpt ion is great er t han groups, t he cases t reat ed wit h t he ANP had a higher bone for m at ion. One of t he charact er ist ics of t his m ineralized t issue form at ion in t he apical region. The condit ion is t he accum ulat ion of ost eoclast s in t he ® aut hors concluded t hat t he EndoVac syst em m ight be bone resorpt ion areas28 . I n t his st udy we observed considered a valuable disinfect ion prot ocol in im m at ure VLJQL¿FDQW GLIIHUHQFH LQ WKH QXPEHU RI RVWHRFODVWV perm anent t eet h wit h apical periodont it is, and t hus bet w een t he t w o t y pes of ir r igat ion. The av erage int racanal ant ibiot ics w ould not be necessar y. The num ber of TRAP- posit ive cells was lower in EndoVac® - result s of our st udy are in agreem ent wit h Silva, et t r eat ed t eet h t han in convent ional ir r igat ion. This 23 ® ( 2010) , in which EndoVac specim ens present ed result does not agree wit h t hat previously published st ruct ured connect ive t issue, rich vascularizat ion, and by Cohenca, et al. 8 ( 2015) , who observed no st at ist ical repair process in advanced st age. difference in t he num bers of ost eoclast s. This was al. 8 Cohenca, et al. ( 2015) com pared t he ANP irrigat ion probably due t o t he fact t hat t he aut hors used t eet h wit h convent ional irrigat ion and ult rasonic irrigat ions w it h com plet e r oot for m at ion and closed apex as in d og t eet h w it h com p let e r oot f or m at ion an d experim ent al m odel. apical periodont it is. The result s of t heir m icroscopic DQDO\VLV VKRZHG VLJQL¿FDQW GLIIHUHQFH RQO\ LQ WKH LQÀDPPDWRU\ LQ¿OWUDWH ZKLFK ZDV ORZHU LQ WKH $13 specim ens in com parison wit h convent ional irrigat ion. The per iodont al ligam ent t hick ness, r esor pt ion of m ineralized t issues, size of per iapical lesions, and t he num ber of ost eoclast s did not present st at ist ically VLJQL¿FDQW GLIIHUHQFH EHWZHHQ WKH JURXSV 7KHVH result s do not agree wit h ours, probably due t o t he par t icular anat om ical charact er ist ics of t eet h w it h in com p let e r oot f or m at ion , w h ich p r esen t r ich er vascular supply 21,29 enabling short er repair process. Conclusion The r esult s fr om t his in v iv o st udy allow ed us t o con clu de t h at t h e n egat iv e pr essu r e ir r igat ion ( EndoVac ® ) dem onst rat ed bet t er biological r esult s t han t he convent ional irrigat ion in im m at ure t eet h wit h apical periodont it is and present ed a m ore advanced UHSDLUSURFHVVWKXVFRQ¿UPLQJWKHK\SRWKHVLV&OLQLFDO st u dies sh ou ld be per f or m ed in or der t o pr ov ide addit ional inform at ion for dent al pract ice. I n our st udy, t her e was a par t ially m ineralized t issue invaginat ion in 53.6% of t he root canals 90 days aft er use of t he ANP. The invaginat ion of healt hy periodontal tissue into the root canal in the apical region indicat ed t hat t he int ense m icrobial cont am inat ion was cont rolled in 53.6% of t he cases. We hypot hesize t hat Acknowledgm ent s 7KHDXWKRUVGHFODUHQRFRQÀLFWVRILQWHUHVWUHODWHG t o t his st udy. This invest igat ion was support ed in part by a scholarship from CNPq – Nat ional Council for 6FLHQWL¿FDQG7HFKQRORJLFDO'HYHORSPHQW t he ant im icrobial dressing bet ween t he appoint m ent s J Appl Oral Sci. 618 2017;25(6):612-9 Apical Negative Pressure irrigation presents tissue compatibility in immature teeth 16 - Kawashim a N, Wadachi R, Suda H, Yeng T, Parashos P. Root canal References m edicam ent s. I nt Dent J. 2009; 59( 1) : 5- 11. 1 - Abaraj it han M, Dham S, Velm urugan N, Valerian- Albuquerque D, Ballal S, Sent hilkum ar H. 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