CAJ | 학술논문

背景:课题组以往研究显示:体外培养条件下,冻存骨髓基质细胞复苏后仍保持较高的细胞存活率、细胞增殖及成骨分化能力.上述结果仍然需要进一步在体内环境下证实.目的:观察经超低温冻存后的骨髓基质细胞和支架材料胶原膜BME-10X复合体植入裸鼠体内后Ⅰ型胶原的合成情况.方法:体外分离培养Beagle犬骨髓基质细胞,冻存12个月后复苏,体外构建骨髓基质细胞和胶原膜材料复合体.分别经矿化诱导培养液、基础培养液培养5 d后,植入裸鼠体内,于术后第4周取出标本,进行大体观察、组织病理学和免疫组化分析,并应用图像分析系统对各组标本中的Ⅰ型胶原进行定量分析.以矿化诱导培养液培养的单纯胶原膜材料为对照组.结果与结论:对照组在植入胶原膜后,胶原膜边界清晰,膜边缘及内部基本没有细胞生长,Ⅰ型胶原分布很少;在未诱导矿化组,术后第4周可见,胶原膜内有细胞长入,并有细小的条索状新生胶原形成,Ⅰ型胶原分布明显增多;在诱导矿化组,植入后也可见支架材料的分解降解和更多的细胞生长,大量新生的胶原形成类骨质样组织,与前两组对比,Ⅰ型胶原分布增多有显著性意义.结果表明冻存骨髓基质细胞复苏后进行体外培养扩增与诱导分化,并在体内环境下复合胶原支架材料,仍然具有较强成骨能力.
배경:과제조이왕연구현시:체외배양조건하,동존골수기질세포복소후잉보지교고적세포존활솔、세포증식급성골분화능력.상술결과잉연수요진일보재체내배경하증실.목적:관찰경초저온동존후적골수기질세포화지가재료효원막BME-10X복합체식입라서체내후Ⅰ형효원적합성정황.방법:체외분리배양Beagle견골수기질세포,동존12개월후복소,체외구건골수기질세포화효원막재료복합체.분별경광화유도배양액、기출배양액배양5 d후,식입라서체내,우술후제4주취출표본,진행대체관찰、조직병이학화면역조화분석,병응용도상분석계통대각조표본중적Ⅰ형효원진행정량분석.이광화유도배양액배양적단순효원막재료위대조조.결과여결론:대조조재식입효원막후,효원막변계청석,막변연급내부기본몰유세포생장,Ⅰ형효원분포흔소;재미유도광화조,술후제4주가견,효원막내유세포장입,병유세소적조색상신생효원형성,Ⅰ형효원분포명현증다;재유도광화조,식입후야가견지가재료적분해강해화경다적세포생장,대량신생적효원형성류골질양조직,여전량조대비,Ⅰ형효원분포증다유현저성의의.결과표명동존골수기질세포복소후진행체외배양확증여유도분화,병재체내배경하복합효원지가재료,잉연구유교강성골능력.
BACKGROUND: Our previous studies have demonstrated that cryopreserved bone marrow stromal cells (BMSCs) still maintain high survival rate, cell proliferation and osteogenic differentiation potentials after thawing. However, this result needs confirmed in vivo environment. OBJECTIVE: To explore the effects of cryopreserved BMSCs and collagenic membrane BME-10X complex on type Ⅰ collagen synthesis in vivo. METHODS: Beagle dog BMSCs were cultured in vitro and cryopreserved for 12 months, which were thawed and prepared complexes with collagenic membrane. The complexes were cultured with mineralization induction medium or normal medium for 5 days, followed by implanting into nude mice. The specimens were harvested and analyzed by gross observation, histopathological and immunohistochemistry at 4 weeks after implantation. The collagenic membrane cultured with mineralization induction medium served as controls. RESULTS AND CONCLUSION: In the control group, the boundary of collagenic membrane was distinctly, without cell growth around boundary or intra collagenic membrane, additionally, there was little type Ⅰ collagen. In the non-induction group, cells grew into collagenic membrane, trabes-like collagen formed, and type Ⅰ collagen distribution increased at 4 weeks. In the induction group, scaffold degraded, more cells grew, and plenty of collagen formed osteoid-like tissues. The distribution of typeⅠcollagen was obviously increased than that of other groups. The findings demonstrated that cryopreserved BMSCs possess strong osteogenic differentiation potentials after proliferation and induction combined with collagenic membranes in vitro.