1. 김홍성 1,3, §, 박상민 1, 윤상준 1, 김지현 1, 류수착 2,3, 신상훈 4 Hong Sung Kim 1,3, §, Sang Min Park 1, Sang Jun Yoon 1, Ji Hyun Kim 1, Su Chak Ryu 2,3, Sang Hun Shin 4 1 부산대학교 생명자원과학대학 바이오소재공학과, 2 나노과학기술대학 나노정보소재공학과, 3 PNU-IGB 국제공동연구소, 4 치과대학 구강악안면외과 1 Dept. of Biomaterial Engineering, College of Natural Resources & Life Science, 2 Dept. of Nanomaterial Engineering, College of Nano Science & Technology 3 Joint Research Center of PNU-Fraunhofer IGB, Pusan National University, 3 Joint Research Center of PNU-Fraunhofer IGB, Pusan National University, 4 Dept. of Oral and Maxillofacial Surgery, College of Dentistry, Pusan National University. § khs@pusan.ac.kr 개요 소재 제법 & 특성 In vitro 물성 In vivo 조직 특성 특허 특허 등록 ( 제 10-0845002) 특허 등록 ( 제 10-0845002) 국제 특허 (PCT) (KR2008/001085) 국제 특허 (PCT) (KR2008/001085) Porosity Figure 7. Variation of cumulative filter flow of the scaffold with average diameter: about 3.4 ㎛. Figure 6. The SBF absorptivity behavior accor ding to the weight ratios of hydroxyapatite in the hybrid scaffolds. Absorptivity of SBF Figure 9. Relative viability of osteoblast cells o n the matrices. The cell activity was determine d by the MTT assay and normalized the respe ctive living cell numbers. Activity of osteoblast Protein release Figure 8. FITC-BSA release profiles from the hybrid scaffolds with HAP weight rati o of 0.2 and 0.5. Figure 10. Photographs taken from cranial bone tissue implanted porous hybrid scaffolds after 3 days from implantation. The newly born spicules are briskly produced in the boundary of hybrid matrix by bone medullar cells (left image). The osteoconduction is noticed inside of the scaffold(red color) by permeation of osteoblasts (right image). Publication: 1. Macromolecular Research, 15, 1, 65 (2007) Preparation of a porous chitosan/fibroin-hydroxyapatite composite matrix for tissue engineering 2. Biomaterials Research, 11, 3, 96 (2007) Porous structure and characteristic of protein release on biodegradable chitosan/fibroin-hydroxyapatite hybrid scaffold Figure 1. SEM images taken from the inside of the hybrid with biopolymers/HAP weight ratios of (A) 100/0, (B) 80/20, (C) 50/50 at a magnification of ×30, (D) 70/30 at ×200, and (E) comparison images of the surface and the back in 40/60 hybrid toward the sublimated direction of the solvent; the upper image is the surface at ×70 and the lower image is the back at ×30, and (F) lateral section at ×200. 1. 생체적합성 2. 세포의 부착, 분화, 증식할 수 있는 표면 조성 3. 혈관신생을 위한 적절한 크기의 서로 연결된 공극 구조 4. 분해 흡수성 : 부산물 대사 5. 물성유지와 시술을 위한 역학적 물성 6. 다양한 크기와 형태의 제작성  결손 되거나 손상된 조직을 자가세포와 세포지지체 (scaffolds) 를 이용하여 재생 복구.  Implantation of porous 3D matrix that impregnated cells and/or bioactive factors or blank. - Cells: 성체세포, 전구세포, 중배엽 줄기세포 - Factors: 분화유도인자, 성장촉진인자 ; Insulin, GH, IGF, PDGF, TGF, FGF, etc.  경조직 지지체 : 조골, 치주, 연골  연조직 지지체 : 지방, 피부, 신경  차폐재 : 치공 PLUG, 조직융착 성형 구조 Specifications 1) 공극크기 : 평균 3.4  m, 최대 11.3 3.4  m 2) 공극율 : 95% 이상 3) 분해시간 : 8 주 ~ 16 주 ( 조절가능 ) 4)SBF 흡수율 : x6 ~ 20 5) 단백질 투과율 : 1mg/cm 2 day 6) 파단강도 : 25~45 gf/mm 2 생체모방형 hybrids  Polysaccharide: Chitosan - (acetyl) glucosamine  Protein: Fibroin (silk) - amino acids o 세포외기질 (ECM) 의 구성 성분  Bioceramic: Hydroxyapatite Ca 10 (PO 4 ) 6 (OH) 2 calcium phosphate o 뼈 ( 연골 ) 의 주성분  다층구조의 투과성 다공체 : o 적절한 크기의 상호연결형 공극구조 o 표면의 등방성 공극과 내면의 비등방성 공극  Chitosan (Mw 400kD): Acetic acid solution  Fibroin ( Bombyx mori ): dialysis of CaCl 2 /H 2 O/ EtOH solution  Hydroxyapatite (synthesis): 0.5~2  m, Ca/P=1.55  Thermally inducing phase separation (TIPS) of mechanically foamy solution in PET mold; -98  C, 0.5mmHg, 24hr  7 kinds of hybrids were prepared as shown in Table 1.  FT-IR spectroscopy: 4000~400 cm-1  X-ray diffraction analysis: Cuka, 50kV, 30mA  SEM morpholgy: 15kV  Simulated body solution(SBF) absorptivity  Protein permeability: FITC-bovine serum albumin  MTT viability: Osteoblasts  Tenacity: UTM 5kgf  Surgical procedure - 3 cm midline scalp incision of rat - 5 mm bilateral round bony defect in parietal bone - Defect site was packed with scaffold, opposite site was left as control - Rats were euthanized 3 days, 2 weeks, 4 weeks, 12 weeks post-operatively.  Histomorphometric analysis - 5  m section, H&E stain, light microscopy § Corresponding to Hong Sung Kim, PH.D., Professor Dept. of Biomaterial Science, School of Applied Resources & Life Science, Pusan National University khs@pusan.ac.kr HP. 011-840-5385 LAB 055-350-5385, 5912 FAX 055-350-5389 Figure 1. SEM images from the surfaces of chitosan/fibroin- hydroxyapatite hybrid with biopolymers/HAP weight ratios of (A) 100/0, (B) 70/30, (C) 50/50 at a magnification of ×200, (D) 50/50 at ×1000, (E) 50/50 at ×5000, and (F) 30/70 at ×2000. Crystallography Figure 3. XRD patterns of chitosan/fibroin - hydroxyapatite hybrids with biopolymers /HAP weight ratios of (a) 100/0, (b) 90/10, (c) 80/20, (d) 70/30, (e) 60/40, and (f) 50/50, and (g) the 70/30 hybrid immersed in SBF for 2 weeks. Schematic diagram Figure 4. The formation of hydroxyapatite nuclei on the acetylated chitosan surface in 1.5X SBF. Hap crystal formation Figure 5. The surf ace morphologies of acetylated chit osans after imme rsion in 1.5X SBF solutions for 3 we ek at 37 ℃ ; each magnification of X20,000 and X80, 000. Figure 12. Photograph of cranial bone section implanted the scaffold on 4 weeks. The neobone was formed from the trabecular bone to a border of the scaffold by osteointegration. Figure 11. Photograph of cranial bone section implanted the scaffold on 2 weeks. The degradation of the scaffold occured inside of it. The neobone was formed on the boundary of scaffold contacting fibrous layer-like osteo cytes way. 3 Day 2 Week 4 Week Polymeric Biomaterials Lab., Biomaterial Engineering Dept., Pusan Nat. Univ.