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1 Biomaterials Research (2006) 10(2) : Biomaterials Research 7 The Korean Society for Biomaterials e œ Novel Fabrication and Characterization of Asymmetrically Porous Membrane for Effective Guided Bone Regeneration ½ yá wá y* Jun Ho Kim, Se Heang Oh, and Jin Ho Lee* Š Š g Š Department of Advanced Materials, Hannam University, Daejeon , Korea (Received March 15, 2006/Accepted May 12, 2006) Porous guided bone regeneration (GBR) membranes with selective permeability, hydrophilicity, and adhesiveness with bone were prepared using polydioxanone (PDO) and Pluronic F127 by an immersion precipitation method. The PDO/ Pluronic F127 membranes were fabricated by immersing PDO/Pluronic F127 mixture solution [in N-Methyl-2-pyrrolidone (NMP)] in a mold into water. The PDO/Pluronic F127 mixture was precipitated in water by the diffusion of water into PDO/Pluronic F127 mixture solution. It was observed that the membrane has an asymmetric column-shape porous structure. The top surface of the membrane (water contact side) had nano-size pores (~100 nm) which can effectively prevent from fibrous connective tissue invasion but permeate nutrients, while the bottom surface (mold contact side) had micro-size pores (~70 µm) which can improve adhesiveness with bone. From the investigations of mechanical property, water absorbability, and model nutrient permeability of the membranes, the hydrophilized PDO/ F127 (3 wt%) membrane seems to be a good candidate as a GBR membrane for the effective permeation of nutrients as well as the good mechanical strength to maintain a secluded space for the bone regeneration. Key words: Guided bone regeneration, Bone defect, Selective permeability, Polydioxanone (PDO), Pluronic F127 ilf g f hœd, Œd, x ( d ) Š f l hš l d jdš ex rlš f. w Œ fš h, ilf h, Œf hf Š xjlœ f ilf f Š d, e Œ il, Œ il, e Šil fš h seh ilf g f ŠŠ. 1-3) f Š il g hhf Šf Š il e Šilf x hf r Š, il g edš l f h z eš i lf g f e Š x f g e [guided bone regeneration (GBR)]f Š, f d f g e (GBR membrane)f Š. x f ƒ x i h f f f xš d f ƒ f ilf h l Š d. ƒ x w f ƒ f v l hhf e Š. u v f ƒ *sf hf: jhlee@hannam.ac.kr f tg g e f fdš Š f hf d f. 4) g e f f 1980 Nyman 5) Gottlow 6) f h Š ilg e Š Dahlin 7) f ut g e f dš g f Šf Š thf h fšf, g e f td (in vitro) ilf g Š f Š il Šh h Š d d x f f hšš g e f f f f f df f. g e f d g } s f [z (collagen), 8-11) (alginate) 3,12) ] Š f [ (expanded-polytetrafluoroethylene (e- PTFE)), 13-15) (poly(lactic acid) (PLLA)), 16,17) z (poly(glycolic acid) (PGA)), - z j Št (poly(dl-lactic-co-glycolic acid) (PLGA)) 18-20) ] f. s ff d, thš f ŒŠ f Š t g e f f f vi ~ l l Š Š Š hf f f h f f, Š ff d, thš f d Š e-ptfe fd Š g hhf f ff, t Š l g fr f Š h Š hf f. 21) f Š fe u 55

2 56 jœá ŠÁflŒ Š Š f fdš j f f. f hf g e f f d f viš f lš h f. h thš f h Š, ilf f f lš e Šilf x l Š eš hhš f l Š, g f f f el ~ ff h f h f h Š. Š f gf e Šilf x lš eš j il g hrš f Š, g Š hf Š, j h ilf l df eš e Š ƒ f l Š, g g f Š h Š d Š f l Š. 18,22-25) u Š f Š l hf f Š f t Š Guidor, Vicryl Periodontal Mesh, Biomesh f h ˆf v fl, Š ff Š (brittle) l f Š f d il f f hr ( f e Šilf x ) g e f f lš d Šf vi ~l Š f hf, f fš Š ff g g f Šd gh Š df d hšhf. df g Š hf f Šl e Šilf x hš f lf l ~h f f h Š hf. f Š f d Š h e f l polydioxanone (PDO) x ff Pluronic F127f fh e ŒŠŠ, f f ŒŠd f ŠxÁxh (immersion precipitation method) fš x, x i ~h f f f hiš, f f f morphology, h wh, x f f f Š g e f f d f ~lš f Š. x g e hi eš Š ff PDO (Mw, ~200,000) h dš f, g e x f Š eš Pluronic F127 (EG99PG65EG99, Mw 12,500; BASF, USA)f t h d. f f d (co-solvent) ft fh d f l N-Methyl-2-pyrrolidone (NMP, Junsei, Japan) 26) d. Š ff PDO x ff Pluronic F127 f fh e(0, 3, 5 wt%; PDO base) ŒŠŠ, f NMP 100 o C Š 10 wt%f f ŒŠd f hiš. f fhš } f ( mm) x Š, f ŒŠd f ŠŠ d (nonsolvent)f t 1 Šx z. t htš, d - d xœ (solvent-nonsolvent exchange) fš f d f (top surface) xh f Œ f, f d f (sublayer)f f Œ (diffusion) fš xhf Œ. Œ f xh f f t 6 sš g NMP h Š j f, l iš x Œ g e f hiš (Figure 1). f hi f 0.4 mmf f, Š ff PDOf Š lš eš dš h l l Š. Morphology hi g e f }, Œ~ Figure 1. Schematic diagram showing porous PDO or PDO/F127 membrane fabrication process. Biomaterials Research 2006

3 d x g e f hi 57 f scanning electron microscopy (SEM; S-3000N, Hitachi, Japan) dš. hi g e f 5 5 mmf } g platinumf l lrš, (top & bottom surface) f } Œ~ rš.» fg (Tensile strength) wh hi g e f h f Š eš fg f Š f, f eš f hi (ASTM D 638 type V) fdš hfš, UTM fg (AG- 5000G, Shimadzu, Japan) ex z wh. f, crosshead speed 10 mm/min Š f, data acquisition systemf fdš stress-strain f, f f fg e h Š. Š (Suturing strength) wh g e f ft hd, d Šf Š f il j h ~ Šd ff, hiš g e f f hšš f l l ŒfŠ eš Š whš. i f g xjlœ f ƒ h hd f Bio-Gide (collagen membrane; Geistlich Pharm AG, Switzerland) h. f 1 cm 2 cm } hiš f, f f Šo f h j ( gf 2 mm ex)f Nylon Š (5-0, Ailee, Korea) ŠŠ, f UTM ex z 10 mm/minf cross-head speed fg f ŠŠ. Š data acquisition systemf fdš l e fracture force hfš f, f i ~ hf ~ Š Á Š. g e f x Pluronic F127f h f f hf Š hf Šf Š g e f x h (absorption time) whf Š rš. j f fdš g e f e (top surface) t f l whš f x h Š. Š, hi g e f t (37 o C) fh (1f 7f) j f (50 rpm), f t i s, l iš e fš Š g e Š f Pluronic F127f f h (leaching stability)f Š. g e f g e f Š f h Š eš, fluorescein isothocyanate-bovine serum albumin (FITC-BSA; Sigma, USA)f f f d f, f eš 3 mlf l side-by-side diffusion cell (diffusion area, 0.64 cm 2 )f fd. FITC-BSA phosphate buffered saline (PBS, ph ~7.4) 1 mg/mlf dšš f, f PDO, PDO/Pluronic F127 Bio-Gide f gr diffusion cellf Šo cell (donor cell) 3 ml sej, o cell (receptor cell) 3 mlf PBS sej. f 37 o Cf f} f 12 z j f, fh receptor cellf ht PBSf swš d PBSf sej f fdš swš f, Fluorescence spectrophotometer (RF PC, Shimadzu) fdš FITC-BSAf f whš (excitation wavelength, 494 nm; emission wavelength, 520 nm). š PDO f df d Š ff PGA, PLLA PLGA Š Ž e (flexibility) l l (toughness)f l ff, 27) PDO dš ~ f d hšhf h fdf l Š f. Šf d e g e f hd Š lš (e l l)f l f, PDO g e hi eš ƒ g hš, f x f Š eš t h Pluronic F127f hš. f ˆf t (FDA)f t df f f f f. f f NMP dš z d ~ hiš ŠxÁxh f fdš x, ~h x i l g e f hiš. Figure 2 PDO/Pluronic F127 (3 wt%) hi g e f SEM lf ~ f. f, f f y Œ~f x i l f d e Š 0.4 mm ff ŒfŠ f. Š g e f e (t ht )f e Šilf x f fl f f ~ ff h f 100 nmf }, ( ht )f j f il g hr Š f h } } ( 70 µm) h, g hf Œ~f i fff rš f. 28,29) f Š ƒš Œ~f Œ f fd d (nonsolvent)f f f f. 30) ~ l l, Pluronic F127 t f PDO f hi f d fš Œ~ lf r Š. hi g e f Pluronic F127 t Œ fg Š whf Š Š f, Figures 3 4 ~. f, Pluronic F127f ŒŠ ef l Š Pluronic F127f plasticizer effect f, fg f e f l Vol. 10, No. 2

58 김준호 오세행 이진호 SEM photographs showing the morphologies of the top, cross-sectional, and bottom surfaces of PDO/F127 (3 wt%) membrane (*, pore size). Figure 2. 가 현상을 관찰할 수 있었으나, 그 차이는 그리 크지 않았다.

4 58 김준호 오세행 이진호 SEM photographs showing the morphologies of the top, cross-sectional, and bottom surfaces of PDO/F127 (3 wt%) membrane (*, pore size). Figure 2. 가 현상을 관찰할 수 있었으나, 그 차이는 그리 크지 않았다. 봉합강도 측정으로부터, 제조된 뼈재생유도막의 경우 상품화되 어 사용되고 있는 Bio-Gide 막의 경우와는 달리 건조 및 젖 은 상태의 물성 차이가 없음을 관찰할 수 있었으며 (Figure 4), 제조된 뼈재생유도막 (Pluronic F127의 첨가량에 상관없이) 의 물성이 젖은 상태의 Bio-Gide 막 (인체 적용시 생리식염 수에 10여분 정도 적셔서 사용)과 유사한 봉합강도, 즉 실제 인체 적용시 충분한 봉합강도를 가질 수 있음을 확인하였다. 첨가된 Pluronic F127에 의한 봉합강도 차이는 그다지 크게 나타나지 않았다. 제조된 PDO/F127 막의 친수성 정도와 PDO/F127 막 내에 존재하는 Pluronic F127의 수용액상에서의 안정성 (leaching stability)을 물 흡수성 측정을 통해 각각 관찰하였으며, 그 결과 를 Table 1에 나타내었다. 표에서 보듯이 Pluronic F127이 첨가되지 않은 PDO 막의 경우, 물에 서서히 젖어드는 현상 (absorption time, ~140 sec)을 관찰하였는데, 이는 기존의 생분해성 합성고분자들 (PGA, PLLA, PLGA 등)이 물에 젖음 현상이 전혀 나타나지 않는 것과는 다른 현상이며, 이는 기 존 생분해성 고분자들에 비해 PDO의 소수성이 다소 약하기 때문인 것으로 판단된다. Pluronic F127이 첨가된 PDO/ F127 막의 경우, Pluronic F127 첨가량이 증가할수록 물 흡 수시간이 빨라짐을 관찰할 수 있었으며, Pluronic F127의 첨 가량이 3 wt% 이후로는 물 흡수시간의 차이가 크지 않아, 3 wt%가 친수성 부여를 위한 최적농도로 판단되었다. 친수성이 부여된 막은 산소 및 자양분을 포함하는 체액의 침투가 쉽게 이루어져 손상부위로의 효과적인 공급에 의해 뼈 재생에 매우 긍정적이리라 판단된다. PDO/F127 막에 포함되어 있는 Pluronic F127의 막 내 안정성 (leaching stability)을 평가한 결과, PDO/F127 막을 1주일간 물 속에 담갔다 꺼낸 후에도 막의 물 흡수시간이 변하지 않는 것으로 보아 Pluronic F127 이 막 내부에 안정하게 존재함을 확인할 수 있었다. 이는 Pluronic F127에 존재하는 PPG 사슬과 PDO matrix 간의 31) 27) Stress-strain curves of PDO and PDO/F127 membranes (membrane thickness, ~0.40 mm). Figure 3. Absorption time of PDO and PDO/F127 membranes with different Pluronic F127 compositions Absorption time (sec) Pluronic F127 compositions (PDO base, wt%) 0 day* 1 day 7 day 0 ~ 140 ~ 140 ~ ~ 45 ~ 45 ~ 45 5 ~ 40 ~ 40 ~ 40 o *Incubation time of membrane in water (37 C, 50 rpm shaking) Table 1. Comparison of suturing forces of PDO and PDO/F127 membranes with commercial Bio-Gide membrane (n=3). Figure 4. Biomaterials Research 2006

5 d x g e f hi 59 Figure 5. Cumulative FITC-BSA permeation profiles through PDO, PDO/F127 (3 wt%), and Bio-Gide membranes (n=3). hydrophobic interaction PDO matrix Pluronic F127 f entanglement fš f f. f f h, Pluronic F127f 3 wt% t d PDO/F127 hif uhi f, f f d. f f, g e f Š f f g d jdš d f ff, f Š eš f f FITC-BSA d. Figure 5 PDO, PDO/F127 (3 wt%) Bio-Gide f Š FITC-BSAf f ~. f, f Š FITC-BSAf f i f f Œhf l Š ff, PDO/F127 (3 wt%), PDO, Bio-Gide f f h d Š f ~. Type I type III collagenf Bio-Gide f d, x f Š i Š i (compact structure) fš FITC- BSAf ~ f, hiš PDO PDO/F127 (3 wt%) f y Œ~f ƒ Š i fš Bio-Gide FITC-BSAf f d Š ~ f, PDO/F127 (3 wt%) f d, Pluronic F127 fš x fš hf f PDO Š f d Š, g hf f f ~ f. x, ~h j il f d Š hr f l f g e f PDO Pluronic F127 ŒŠd f ŠxÁxh fš hiš. hi g e f g Š hf f f dfš g f Š f e Šilf x h ~ f ~h Š xh Œ~ ( 100 nmf } l j il f d Š hr f Š f 70 µmf } l f ) h, g hf Œ h f Š f wš f. h PDO/F127 (3 wt%) f x Œ g e f hšš i f Šf, f f FITC-BSAf f Š PDO/F127 (3 wt%) f d Š f ƒ f lf ŒfŠ. f f hi PDO/F127 (3 wt%) f g e f f fd f ŒfŠ f f, g PDO/F127 (3 wt%) i f PDO Bio-Gide f fdš in vivo g eš (SD ratf fdš skull defect model)f lš jf Š Š Š le f Š f l f f. š x 1. J. N. Kent and M. F. Zide, Wound healing: bone and biomaterials, Otolaryngol. Clin. North Am., 17, (1984). 2. A. Linde, P. Alberius, C. Dahlin, K. Bjurstam, and Y. Sundin, ÁOsteopromotion: a soft-tissue exclusion. principle using a membrane for bone healing and bone neogenesis, J. Periodontol., 64, (1993). 3. Y. Ueyama, K. Ishikawa, T. Mano, T. Koyama, H. Nagatsuka, K. Suzuki, and K., Ryoke, Usefulness as guided bone regeneration membrane of the alginate membrane, Biomaterials, 23, (2002) S. Nyman, J. Gottlow, T. Karring, and J. Lindhe, The regenerativ potential of the periodontal ligament. An experimental study in the monkey, J. Clin. Periodontol., 9, (1982). 6. J. Gottlow, S. Nyman, J. Lindhe, T. Karring, and J. Wennstrom, New attachment formation in the human periodontium by guided tissue regeneration. Case reports, J. Clin. Periodontol., 13, (1986). 7. C. Dahlin, L. Sennerby, U. Lekholm, A. Lindhe, and S. Nyman, Int. Generation of new bone around titanium implants using a membrane technique: an experimental study in rabbits, J. Oral. Maxillofac. Impl., 4, (1989). 8. N. Ozmeric, B. Bal, T. Oygur, and K. Balos, The effect of a collagen membrane in regenerative therapy of two-wall intrabony defects in dogs, Periodontal Clin. Investin., 22, (2000). 9. R. D. Mundell, M. P. Mooney, M. I. Siegel, and A. Losken, Osseous guided tissue regeneration using a collagen barrier membrane, Int. J. Oral. Maxillofac. Surg., 51, (1993). 10. H. L. Wang, R. B. O'Neal, C. L. Thomas, Y. Shyr, and R. L. MacNeil, Evaluation of an absorbable collagen membrane in Vol. 10, No. 2

6 60 jœá ŠÁflŒ treating Class Ä furcation defects, J. Periodontol., 65, (1994). 11. A. K. Schlegel, H. Mohler, F. Busch, and A. Mehl, Preclinical and clinical studies of a collagen membrane (Bio-Gide), Biomaterials, 18, (1997). 12. H. Jianqi, H. Hong, S. Lieping, and G. Gengbua, Comparison of calcium alginate film with collagen membrane for guided bone regeneration in mandibular defects in rabbits, J. Oral Maxillofac. Surg., 60, (2002). 13. R. K. Schenck, D. Buser, W. R. Hardwick, and C. Dahlin, Healing pattern of bone regeneration in membrane-protected defects: a histologic study in the canine mandible, Int. J. Oral. Maxillofac. Impl., 9, (1994). 14. C. H. F. Hammerle, U. Bragger, B. Schmid, and N. P. Lang, Successful bone formation at immediate transmucosal implants: a clinical report, Int. J. Oral. Maxillofac. Impl., 13, (1998). 15. D. Buser, K. Dula, H. P. Hirt, and R. K. Schenk, Lateral ridge augmentation using autografts and barrier membranes: a clinical study with 40 partially edentulous patients, J. Oral Maxillofac. Surg., 54, (1996). 16. K. Warrer, T. Karring, S. Nyman, and S. Gogolewski, Guided tissue regeneration using biodegradable membranes of. polylactic acid or polyurethane, J. Clin. Periodontol., 19, (1992). 17. A. Piattelli, A. Scarano, P. Russo, and S. Matarasso, Evaluation of guided bone regeneration in rabbit tibia using bioresorbable and non-resorbable membranes, Biomaterials, 17, (1996). 18. L. Dupoirieux, D. Pourquier, M. C. Picot, and M. Neves, Comparative study of three different membranes for guided bone regeneration of rat cranial defects, Int. J. Oral. Maxillofac. Surg., 30, (2001). 19. L. Mayfield, N. Nobreus, R. Attstrom, and A. Linde, Guided bone regeneration in dental implant treatment using a bioabsorbable membrane, Clin. Oral. Implants Res., 8, (1997). 20. M. Simion, A. Scarano, L. Gionso, and A. Piatelli, Guided bone regeneration using resorbable and nonresorbable membranes: a comparative histologic study in humans, Int. J. Oral. Maxillofac. Impl., 11, (1996). 21. B. Boyce, Physical characteristics of expanded polytetrafluoroethylene grafts, In J. C. Stanley (ed). Biologic & Synthetic Vascular Prothesis, Grune and Stratton, Philadephia, 1982, pp C. Bosch, B. Melsen, and K. Vargervik, Importance of the critical-size bone defect in testing bone-regenerating materials, J. Craniofac. Surg., 9, (1998). 23. D. Lundgren, S. Nyman, T. Mathisen, S. Isaksson, and B. Klinge, Guided bone regeneration of cranial defects, using biodegradable barriers: an experimental pilot study in the rabbit, J. Craniomaxillofac. Surg., 20, (1992). 24. J. T. Melloning and R. G. Triplett, Guided tissue regeneration and endosseous dental implants, Int. J. Periodontal. Res. Dent., 13, (1993). 25. R. Hardwick, T. D. Scantlebury, R. Sanchez, N. Whitley, and J. Ambruster, in Guided Bone Regeneration in Implant Dentistry, D. Buser, C. Dahlin, and R K. Schenk (Ed.), Quintessence Publishing Co., Chicago, 1994, pp R. L. Dunn, J. P. English, D. R. Cowsar, D. P. Vanderbilt, Biodegradable in situ forming implants and methods of producing the same, US patent No. 5,278,201, K. J. Cho, D. K. Song, S. H. Oh, Y. J. Koh, S. H. Lee, M. C. Lee, and J. H. Lee, Fabrication and characterization of hydrophilized polydioxanone scaffold for meniscus regeneration, Tissue Eng. Regen. Med., 2, (2005). 28. S. Nyman, Bone regeneration using the principle of guided tissue regeneration, J. Clin. Periodontol., 18, (1991). 29. L. A. Salata, G. T. Craig, and I. M. Brook, Bone healing following the use of hydroxyapatite or ionomeric bone substitutes alone or combined with a guided bone regeneration technique: an animal study, Int. J. Oral. Maxillfac. Impl., 13, (1998). 30. L. Broens, F. W. Altena, C. A. Smolders, and D. M. Koenhen, Asymmetric membrane structures as a result of phase separation phenomena, Desalination, 32, (1980). 31. S. H. Oh, S. G. Kang, E. S. Kim, S. H. Cho, and J. H. Lee, Fabrication and characterization of hydrophilic poly(lactic-coglycolic acid)/poly(vinyl alcohol) blend cell scaffolds by meltmolding particulate-leaching method, Biomaterials, 24, (2003). Biomaterials Research 2006

12-3전체

12-3전체 The histologic study of the effect of enamel matrix protein and -tricalcium phosphate in gap defect model in dogs : pilot study Lim Hyun-Chang 1, Yang Jin-Hyuk 1, Chae Gyoung-Joon 1, Kim Sung-Tae 1, Jung