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Submitted: Aug 22, 2016 0RGL¿FDWLRQ'HF Accepted: Dec 28, 2016
Effects of hyaluronic acid on bleeding
following third m olar extraction
Obj ect ive: To explore t he effect s of hyaluronic acid ( HA) on bleeding and associat ed out com es aft er t hird m olar ext ract ion. Met hods: Forty pat ient s who had undergone m olar extraction were random ly divided into two groups; 0.8% ( w/ v) HA was applied t o t he HA group ( n= 20) whereas a cont rol group ( n= 20) was not t reat ed. Salivary and gingival t issue fact or ( TF) levels, bleeding t im e, m axim um int erincisal opening ( MI O) , pain scored on a visual analog scale ( VAS) , and t he swelling ext ent were com pared bet ween t he t wo JURXSV5HVXOWV+$GLGQRWVLJQL¿FDQWO\DIIHFWJLQJLYDO7)OHYHOV6DOLYDU\7) OHYHOVLQFUHDVHGVLJQL¿FDQWO\ZHHNDIWHU+$DSSOLFDWLRQEXWQRWLQWKHFRQWURO JURXS1HLWKHUWKH9$6SDLQOHYHOQRU0,2GLIIHUHGVLJQL¿FDQWO\EHWZHHQWKH t wo groups. The swelling ext ent on day 3 and t he bleeding t im e were great er in t he HA group t han in t he cont rol group. Conclusions: Local inj ect ion of HA at 0.8% prolonged the bleeding tim e, and increased hem orrhage and swelling in t he early post operat ive period aft er t hird m olar ext ract ions.
Ke yw or ds: Hyaluronic acid. Bleeding. Ext ract ion. Gokhan GOCMEN1
Sertac AKTOP1 Burcin TÜZÜNER2 Bahar GOKER3 Aysen YARAT2
http://dx.doi.org/10.1590/1678-77572015-0187
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Turkey.
2Marmara University, Faculty of Dentistry, Department of Biochemistry, Istanbul, Turkey. 3Marmara University, Faculty of Pharmacy, Department of Biochemistry, Istanbul, Turkey.
Corresponding address: Gokhan Gocmen Guzelbahce Buyukciftlik Sokak, No.6 - 34365
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I nt roduct ion
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effect during oral wound- healing, and is com m only applied aft er t oot h ext ract ion. Generally, previous st udies about t he subj ect have focused on t issue
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I t has been hy pot hesized t hat HA of appr opr iat e consist ency increases cell m ot ilit y1,13.
Tissue fact or ( TF) is best known as t he prim ary cellular init iat or of blood coagulat ion. TF levels vary as t he need for hem ost at ic prot ect ion changes4,10. The
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changes in t issue TF levels. The bleeding t im e m ay also be an obj ect ive m easure of HA act ivit y. Swelling, pain scor ed on a v isual analog scale ( VAS) , and
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t he hem orrhage ext ent .
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posit iv e ou t com es of HA t r eat m en t , m ay cau se r esear ch er s t o ov er look p ot en t ial sid e ef f ect s.
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pr ocess, hem ost at ic effect s of HA should also be con sid er ed . Ou r h y p ot h esis w as t h at HA m ig h t m odulat e hem ost asis and bleeding. Addit ionally it s t herapeut ic effect and relat ionship wit h side effect s can also com prom ise it s use in surgical procedures. The purpose of this study was to m easure the ability of high- m olecular weight HA t o induce hem ost asis aft er m andibular t hird m olar ( M3) ext ract ion. The prim ary m easures were t he TF level and bleeding ext ent ; t he VAS pain score, MI O, and swelling ext ent served as secondary clinical m easures.
Mat erials and m et hods
Pat ient s and Met hods:
An a pr ior i pow er calculat ion indicat ed t hat a sam ple size of 18 pat ient s was required for each gr oup. This com parat ive, pr ospect ive random ized st udy included 40 pat ient s t reat ed in t he Depart m ent of Oral and Maxillofacial Surgery, at t he Facult y of Den t ist r y, Mar m ar a Un iv er sit y, I st an bu l, Tu r k ey. Approval was obt ained from t he appropriat e Et hics Com m it t ee ( approval no. 2011- 1) . Writ t en inform ed consent was obt ained from all pat ient s. Eligibilit y crit eria t o part icipat e in t he st udy required pat ient s t o h av e v er t ical h alf- im pact ed M3 w it h ou t bony
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according t o t he guidelines of t he Am erican Societ y of Anest hesiology. Exclusion crit eria included t he use of
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t he 2 weeks prior t o surgery, and/ or any pat hological or inflam m at or y condit ion in t he im pact ed t oot h area ( n: 65) . Addit ionally, aft er subm ission t o t he sur ger y, if t he ext ract ion was t raum at ic, pat ient s were excluded from t he st udy – i.e., t he t oot h m ay r equir e or under w ent separat ion, fract ur e dur ing ext ract ion, rem oval surrounding bony support or full
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ext ract ion t im e ( t im e passed during local anest het ic adm inist rat ion and anest hesia onset were excluded, only ex t ract ion per iod w as account ed fr om st ar t t o end) t ook m ore t han 30 m inut es. Bot h groups con t ain ed elect iv e pat ien t s w h o h ad u n der gon e unilat eral m andibular M3 ext ract ion ( Figure 1) .
The prim ary predictor, HA application, was coded as a binary variable. Half of t he pat ient s were random ly assigned t o r eceive a local HA gel ( 0.8% [ w / v ] ; Gengigel, Ricerfarm a, I t aly) following t oot h rem oval ( n= 20)5,6. A 0.2 m l HA gel was applied im m ediat ely aft er M3 rem oval t o t he edge of t he ext ract ion socket . The cont rol group was not t reat ed.
Before each surgical procedure, MI O was recorded ( in m m ) and the m outh was rinsed with distilled water. Saliva sam ples ( 2 m L) were collect ed by spit t ing int o a funnel while in t he rest ing posit ion. Teet h were rem oved under local anest hesia ( 2% [ w/ v] art icaine HCl wit h 1: 100,000 [ v/ v] epinephrine HCl; Ult racaine D- S Fort e; Avent is, Bridgewat er, NJ, USA) . Tissue sam ples approxim ately 1 m m3 in volum e were collected im m ediately after extraction ( T0) from the edge of the buccal wound. Wound closures were m ade wit h 3.0 silk sut ures. Bleeding t im e( s) aft er wound closure was not ed. Saliva sam ples ( 2 m L) were collect ed again exactly 1 h (T1) following extraction. Bleeding tim e and TF levels ( salivary and gingival) served as t he prim ary outcom e variables. Postoperatively, am oxicillin ( 1,000 m g every 12 h for 1 week) and ibuprofen ( 400 m g every 6 h for 48 h) were prescribed for all pat ient s. No st eroid was prescribed for any pat ient . All pat ient s were inst ruct ed t o rinse t heir m out hs t wice daily wit h 0.2% ( w/ v) chlorhexidine.
( t hus from t he m ost inferior region of t he t ragus t o t he oral com m issure, and t o t he soft t issue m argin, respect ively) on t he operat ed side were not ed11. VAS pain scores were recorded 1 h, and 3 and 7 days, aft er surgery. At 1 week ( T2) , t he m out h was rinsed wit h dist illed wat er and 2- m L saliva sam ples collect ed once m ore as described above. The sut ured regions were locally anesthetized and tissue sam ples were obtained from sit es very close t o t he init ial sam pling sit es.
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of t est s. On t he day of TF m easurem ent s, t hawed t issue sam ples were washed wit h saline, all veins and
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paper, and wet weight s recorded. The t issue sam ples were t hen hom ogenized in saline and TF act ivit ies in t he hom ogenat es, and saliva sam ples m easured using Quick’s one- st age m et hod em ploying pooled plasm a from healt hy part icipant s. Each TF act ivit y m easurem ent was perform ed by m ixing 0.1 m L of tissue hom ogenate or saliva with 0.1 m L 0.02 M CaCl2, and t he clot t ing react ion st art ed by t he addit ion of 0.1 m L of plasm a9. All reagent s were brought t o t he reaction tem perature (37° C) prior to adm ixture. As the clotting tim e is inversely proportional to the TF activity
level, lengt hening of clot t ing t im e is a m anifest at ion of reduced TF act ivit y.
All st at ist ical analysis was perform ed using t he SPSS for Window s soft w ar e pack age ( v er. 1 2 . 0 ; SPSS I nc., Chicago, I L, USA) . Descript ive st at ist ics w er e com put ed for all var iables. The t - t est , and Mann-Whit ney U t est were used t o assess differences bet ween t he HA and cont rol groups. A p value < 0.05
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Result s
The m ean pat ient age was 24.8 years ( range: 18- 35 years) . Com parison of t he gingival TF bet ween
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differ ence w it h TF w hen com par ed t o t he cont r ol group’s m ean values.
I n t he cont rol group, t here was no st at ist ically
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week in HA group in One-Way ANOVA for Repeat ed Measurem ent t est ( p< 0,05) . I n t his group, binary
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Difference) t est . I n which, m ean of T2 was less t hen
bot h T0 and T1. LSD t est show ed no significant difference bet ween T0 and T1 ( Table 2) .
The m ean ± SD of bleeding t im e in HA group
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group. Bleeding t im e was longer in HA group t han
in t he cont rol group. ( 7.17± 1.36 versus 5.64± 1.46 m inut es, p< 0.05 ) . Com parison of pre- op, 1 hour, 3rd day and 7t h day MI O and VAS were not st at ist ically si g n i f i ca n t b et w een g r o u p s. Sw el l i n g w a s n o t
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TF Control
Group
H.A. Group
Mean ± SD Mean ± SD
Pre-op 102.15±31 89.80±24.88
Post-op (1 week) 63.10±16.2 54.15±14.03
+p
Median í í *p
(Min ; Max) í í
0.758 +: t test
*: Mann-Whitney U test
Table 1- Comparison of gingival Tissue Factor (TF) between control and hyaluronic acid groups
TF Control
Group
H.A. Group
Mean ± SD Mean ± SD
Pre-op 87.1±26.5 122.3±48 Post-op (1 hour) 97.55±37.48 114.20±31.48 Post-op (1 week) 84.8±29.2 68.6±22.98
+p 0.081
(Pre-op vs Post-op 1 hour) *p
Median í í 0.001
(Min ; Max) (-40.96 ; 59.72) í
(Pre-op vs Post-op 1 week)
Median í í 0.002
(Min ; Max) í í
+: Repeated measures ANOVA *: Mann-Whitney U test
Table 2- Comparison of salivary Tissue Factor (TF) between control and hyaluronic acid groups
MIO Swelling VAS
Mean ± (SD) Mean ± (SD) Mean± (SD)
HA Control p HA Control
HA Group
(mm)
Control Group
(mm)
p Group (mm) Group (mm) Group Group p
O.T M.T. O.T. M.T. O.T. M.T.
Pre-op 39.4
±(2.3) ±(2.2)40.1 ! ±(3.32)104.36 ±(4.6)140.4 105.2±(3.4) 140.18±(4.3) ! ! 0 0 1 hour 39.2
±(2.5) ±(2.3)39.4 ! ±(3.76)105.8 ±(4.1)142.2 106.23±(2.8) ±(3.8)140.7 ! ! ±(0.4)1.2 ±(0.5)1.4 ! 3th day 35.6
±(1.6) ±(1.8)34.82 ! ±(2.66)113 ±(2.48)148.3 ±(2.1)110 ±(2.6)144.6 ±(1.6)4.3 ±(1.2)4.5 ! 7th day 38.6
±(2.2) ±(2.5)37.8 ! ±(2.8)106.5 141.48±(4.4) ±(3.8)107.8 ±(3.2)141.2 ! ! ±(1.4)3.1 ±(1.83)3.4 !
m easurem ent s( p< 0.05) . However 3rd day out com es of orot ragus and m ent ot ragus m easurem ent s showed m ore swelling in t he HA group ( p< 0.05) ( Table 3) .
Discussion
We explored whet her HA affect ed hem ost asis aft er M3 ext ract ion. We used t he TF level and bleeding t im e as prim ary out com es and VAS- m easured pain, MI O, and swelling as secondary out com es. To t he best of our knowledge, the effect of high- m olecular weight HA on hem ost asis aft er M3 ext ract ion has not previously been exam ined.
HA inhibit s plat elet aggr egat ion and adhesion and, at high concent rat ions, prolongs bleeding t im es. As HA exert s ant it hrom bot ic effect s, t he m at erial is used t o coat endovascular devices15. HA plays t wo very im port ant roles during wound- healing. First , it creat es a t em porary st ruct ure during t he early st ages of healing. Second, and m ost im port ant ly, it t riggers cell proliferat ion and m igrat ion14. Therefore, HA is oft en used t o aid oral wound- healing and increases
OHXNRF\WH GLDSHGHVLV DQG ¿EUREODVW SUROLIHUDWLRQ12. How ev er, alt h ou gh HA aids w ou n d- h ealin g, t h is m ay be associat ed w it h in cr eased bleedin g ( an ant it hrom bot ic effect )2,15. We m easured t he TF levels, t he blood coagulat ion cascade init iat or, t o explore t he relat ionship bet ween HA applicat ion and bleeding.
TF is oft en overexpressed aft er wounding, t raum a, or surgery. TF- induced hypercoagulabilit y encourages wound- healing3. However, we found t hat HA did not increase gingival TF act ivit y. The salivary TF level
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cont rols at 1 week aft er operat ion, when we expect ed a reduct ion. This m ay be associat ed wit h prolongat ion of bleeding in t he HA group. HA increased salivary but not gingival TF levels, perhaps because physiological changes in t he salivary glands and gingiva differ.
HA is a nat ural com ponent of t he ext racellular m at r ix , enabling a st r uct ural fram ew or k , helping hydrat ion, and t hus cr eat ing a non- im m unogenic environm ent t hat assist s regenerat ion6. HA m ight
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wound- healing and t hus having clinical applicat ions. Koray, et al.8 ( 2014) report ed less pain and a reduced MI O aft er M3 ext ract ion in pat ient s t reat ed wit h HA. Hanci and Alt un7 ( 2015) report ed post- t onsillect om y pain relief and increased w ound- healing upon HA ap p licat ion . HA also con t r olled g ast r oin t est in al
bleeding aft er failed endoscopic t herapy9. I n our st udy, we m easured early clinical out com es and t he ext ent of bleeding im m ediat ely aft er ext ract ion. HA usage was associat ed wit h m ore swelling and m ore prolonged bleeding. However, t he bleeding effect was short- t erm ( 2 or 3 days) ; out com es becam e sim ilar lat er in bot h groups.
Conclusion
Local injection of HA at 0.8% prolonged the bleeding t im e, increased hem orrhage and swelling in t he early post operat ive period aft er M3 ext ract ions. However, hem ost asis and hem orrhage t o induce wound healing ar e com plex m ech an ism s an d inv olv e n u m er ou s param eters. Further work on other coagulation factors, m easuring other clotting param eters in m ore patients, is required.
References
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J Case Rep. 2014; 15: 199- 202.
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ext ract ion. J Craniom axillofac Surg. 2015; 43( 7) : 1033- 7.
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12- Mesa FL, Aneiros J, Cabrera A, Bravo M, Caballero T, Revelles F, et al. Ant iproliferat ive effect of t opic hyaluronic acid gel. St udy in gingival biopsies of patients with periodontal disease. Histol Histopathol. 2002; 17( 3) : 747- 53.
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