DBP/PLGA 하이브리드 담체를 이용한 조직공학적 바이오 디스크 개발

고연경; 김순희; 정재수; 하현정; 윤선중; 이종문; 김문석; 이해방; 강길선; Youn Kyung Ko; Soon Hee Kim; Jae Soo Jeong; Hyun Jung Ha; Sun Jung Yoon; John M. Rhee; Moon Suk Kim; Hai Bang Lee; Gilson Khang

Polymer(Korea), Vol.31, No.1, 14-19, January, 2007

Export Citation

DBP/PLGA 하이브리드 담체를 이용한 조직공학적 바이오 디스크 개발

Biodisc Tissue-Engineered Using PLGA/DBP Hybrid Scaffold

고연경, 김순희 , 정재수, 하현정, 윤선중, 이종문 , 김문석 ¹, 이해방 ¹, 강길선 ^†

전북대학교 BK-21 고분자 BIN융합연구팀
¹한국화학연구원 나노생체재료연구팀

Youn Kyung Ko, Soon Hee Kim , Jae Soo Jeong, Hyun Jung Ha, Sun Jung Yoon, John M. Rhee , Moon Suk Kim¹, Hai Bang Lee¹, and Gilson Khang^†

BK-21 Polymer BIN Fusion Research Team, Chonbuk National University, 664-14, Dukjin Dong 1Ga, Dukjin Ku, Jeonju 561-756, Korea
¹Nanobiomaterials Laboratory, Korea Research Institute of Chemical Technology, P.O. Box 107, Yuseong, Daejeon 305-600, Korea

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초록

탈미네랄화된 골분(demineralized bone particle, DBP)은 골/연골 형성의 강력한 유도인자로 사용된다. 본 연구에서는 용매 캐스팅/염 추출법을 이용해 함량별 DBP와 PLGA가 하이브리드화된 다공성 지지체를 실제 디스크 형태와 유사하게 제조하였다. 제조된 지지체의 특성을 분석하기 위하여 다공도, 표면 젖음성 및 물 흡수성을 측정하였다. 디스크 세포인 섬유륜 및 수핵 세포는 토끼로부터 분리하여 제조된 지지체에 각각 파종한 후, 지지체를 재조합하여 배양하였다. 지지체에 파종된 디스크 세포의 생존율과 증식률은 MTT(3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyltetrazolium-bromide) 분석 방법을 이용하였고, 면역결핍 쥐의 피하에 삽입하여 이들의 디스크 조직 형성 정도를 확인하였다. 피하에 이식된 지지체를 적출하여 육안으로 관찰하고 모폴로지의 변화를 확인한 후, 조직을 파라핀으로 고정시켜 슬라이드를 제조하여 hematoxylin과 eosin 염색을 수행하였다. 천연/합성 하이브리드 담체로서의 DBP/PLGA 담체가 PLGA 단독으로 사용하였을 때와 비교하여 볼 때 디스크 조직의 형성이 우수하였으며, 특히 20, 40%의 DBP가 함유된 지지체가 세포의 성장과, 디스크 조직화에 유리함을 확인하였다.

Demineralized bone particle (DBP) has been used as one of the powerful inducers of bone and cartilage tissue specialization. In this study, we fabricated DBP/PLGA scaffold for tissue engineered disc regeneration. We manufactured dual-structured scaffold to compose inner cylinder and outer doughnut similar to nature disc tissue. The DBP/PLGA scaffold was characterized by porosity, wettability, and water uptake ability. We isolated and cultured nucleus pulposus (NP) and annulus fibrosus (AF) cells from rabbit intervertebral disc. We seeded NP cells into the inner core of the hybrid scaffold and AF cells into the outer portion of it. Cellular viability and proliferation were assayed by 3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyltetrazolium-bromide (MTT) test. PLGA and PLGA/DBP scaffolds were implanted in subcutaneous of athymic nude mouse to observe the formation of disc-like tissue in vivo. And then we observed change of morphology and hematoxylin and eosin (H&E). Formation of disc-like tissue was better DBP/PLGA hybrid scaffold than control. Specially, we confirmed that scaffold impregnated 20 and 40% DBP affected to proliferation of disc cell and formation of disc-like tissue.

Keywords:biodisc;demineralized bone particle;scaffold;annulus fibrosus;nucleus pulposus

[References]

Kraemer J, Spine, 20, 635 (1995)
Buckwalter JA, Spine, 20, 1307 (1995)
Nishida K, Kang JD, Gilbertson LG, Moon SH, Suh JK, Vogt MT, Robbins PD, Evans CH, Spine, 24, 2419 (1999)
Mochida J, Nishimura K, Nomura T, Toh E, Chiba M, Spine, 21, 1556 (1996)
Lee CK, Spine, 12, 357 (1988)
Schlegel JD, Smith JA, Schleusener RL, Spine, 21, 970 (1996)
Mizuno H, Roy AK, Vacanti CA, Kojima K, Ueda M, Bonassar LJ, Spine, 29, 1299 (2004)
Seguin CA, Grynpas MD, Pillar RM, Walden SD, Kanda RA, Spine, 29, 1299 (2004)
Bibby SR, Jones DA, Lee RB, Jing Y, Urban JP, Joint Bone Spine, 68, 537 (2001)
Bogduk N, "The inter-body joints and the intervertebral disc", in Clinical Anatomy of the Lumbar Spine and Sacrum, N. Bogduk, Editor, New York, Churchill Livingstone, p. 13 (1997)
Rudert M, Tillmann B, Spine, 20, 1307 (1995)
Khang G, Lee SJ, Lee HB, "Polymer-cell interaction"; in Tissue Engineering: Concepts and Application, 2nd Eds., J. J. Yoo, and I. Lee, Editors, Korea Med. Pub. Co., Seoul, p. 297 (2002)
Khang G, Lee SJ, Han CW, Rhee JM, Lee HB, Adv. Exper. Med. Biology, 657, 235 (2003)
Khang G, Kim MS, Cho SH, Lee I, Rhee JM, Lee HB, Tissue Eng. Reg. Med, 1, 9 (2004)
Khang G, Kim SK, Hong KD, Jang WY, Han CW, Lee I, Lee HB, Tissue Eng. Reg. Med, 1, 136 (2004)
Khang G, Kim EJ, Kim SH, Park KS, Han CW, Yang YS, Lee HB, Tissue Eng. Reg. Med., 2, 20 (2005)
Perrin DE, English PE, "Polyglycolide and Polylactide". in Handbook of Biodegradable Polymer, A. J. Domb, J. Kost, and D. M. Wiseman, Editors, Harwood Academic Publishers, Netherland, Chap. 1, p. 3 (1997)
Hellier WG, Hedman TP, Kostuik JP, Spine, 17, S86 (1992)
Dove J, Hardcastle PH, Davis JK, US Patent 4,904,261 (1990)
Kim SH, Park KS, Choi BC, Ha HJ, Rhee JM, Kim MS, Yang YS, Lee HB, Khang G, Adv. Exper. Med. Biology, 585, 209 (2006)
Mikos AG, Sarakinos G, Leite SM, Vacanti JP, Langer R, Biomaterials, 14, 323 (1993)
Smith DM, Hua DW, Earl WL, MRS Bull., 19, 44 (1994)
Lee MS, Yang JC, Kim SH, Song IB, Kim MS, Khang G, Lee HB, Polym.(Korea), 30(3), 259 (2006)
Kim SK, Kim SH, Lee HR, Cho MH, Kim MS, Khang G, Lee HB, Tissue Eng. Reg. Med., 2, 388 (2005)
Jang JW, Lee B, Han CW, Kim MS, Cho SH, Lee HB, Khang G, Polym.(Korea), 28(5), 382 (2004)
Khang G, Lee SJ, Kim MS, Lee HB, "Scaffolds; Tissue Engineering", in Webster's Biomedical Engineering Handbook, S. Webster, Editor, John & Wiley Press, NY, pp 366-383 (2006)

[Referenced By]

[Related Articles]

[1] Kraemer J, Spine, 20, 635 (1995)

[2] Buckwalter JA, Spine, 20, 1307 (1995)

[3] Nishida K, Kang JD, Gilbertson LG, Moon SH, Suh JK, Vogt MT, Robbins PD, Evans CH, Spine, 24, 2419 (1999)

[4] Mochida J, Nishimura K, Nomura T, Toh E, Chiba M, Spine, 21, 1556 (1996)

[5] Lee CK, Spine, 12, 357 (1988)

[6] Schlegel JD, Smith JA, Schleusener RL, Spine, 21, 970 (1996)

[7] Mizuno H, Roy AK, Vacanti CA, Kojima K, Ueda M, Bonassar LJ, Spine, 29, 1299 (2004)

[8] Seguin CA, Grynpas MD, Pillar RM, Walden SD, Kanda RA, Spine, 29, 1299 (2004)

[9] Bibby SR, Jones DA, Lee RB, Jing Y, Urban JP, Joint Bone Spine, 68, 537 (2001)

[10] Bogduk N, "The inter-body joints and the intervertebral disc", in Clinical Anatomy of the Lumbar Spine and Sacrum, N. Bogduk, Editor, New York, Churchill Livingstone, p. 13 (1997)

[11] Rudert M, Tillmann B, Spine, 20, 1307 (1995)

[12] Khang G, Lee SJ, Lee HB, "Polymer-cell interaction"; in Tissue Engineering: Concepts and Application, 2nd Eds., J. J. Yoo, and I. Lee, Editors, Korea Med. Pub. Co., Seoul, p. 297 (2002)

[13] Khang G, Lee SJ, Han CW, Rhee JM, Lee HB, Adv. Exper. Med. Biology, 657, 235 (2003)

[14] Khang G, Kim MS, Cho SH, Lee I, Rhee JM, Lee HB, Tissue Eng. Reg. Med, 1, 9 (2004)

[15] Khang G, Kim SK, Hong KD, Jang WY, Han CW, Lee I, Lee HB, Tissue Eng. Reg. Med, 1, 136 (2004)

[16] Khang G, Kim EJ, Kim SH, Park KS, Han CW, Yang YS, Lee HB, Tissue Eng. Reg. Med., 2, 20 (2005)

[17] Perrin DE, English PE, "Polyglycolide and Polylactide". in Handbook of Biodegradable Polymer, A. J. Domb, J. Kost, and D. M. Wiseman, Editors, Harwood Academic Publishers, Netherland, Chap. 1, p. 3 (1997)

[18] Hellier WG, Hedman TP, Kostuik JP, Spine, 17, S86 (1992)

[19] Dove J, Hardcastle PH, Davis JK, US Patent 4,904,261 (1990)

[20] Kim SH, Park KS, Choi BC, Ha HJ, Rhee JM, Kim MS, Yang YS, Lee HB, Khang G, Adv. Exper. Med. Biology, 585, 209 (2006)

[21] Mikos AG, Sarakinos G, Leite SM, Vacanti JP, Langer R, Biomaterials, 14, 323 (1993)

[22] Smith DM, Hua DW, Earl WL, MRS Bull., 19, 44 (1994)

[23] Lee MS, Yang JC, Kim SH, Song IB, Kim MS, Khang G, Lee HB, Polym.(Korea), 30(3), 259 (2006)

[24] Kim SK, Kim SH, Lee HR, Cho MH, Kim MS, Khang G, Lee HB, Tissue Eng. Reg. Med., 2, 388 (2005)

[25] Jang JW, Lee B, Han CW, Kim MS, Cho SH, Lee HB, Khang G, Polym.(Korea), 28(5), 382 (2004)

[26] Khang G, Lee SJ, Kim MS, Lee HB, "Scaffolds; Tissue Engineering", in Webster's Biomedical Engineering Handbook, S. Webster, Editor, John & Wiley Press, NY, pp 366-383 (2006)