体外状态下新西兰白兔骨髓中CFU-F集落及成骨性CFU-F集落数量观察

Observations in vitro on the CFU-F Number and Osteogenic CFU-F Number in Bone Marrow of New Zealand White Rabbits

目的 观察体外状态下新西兰白兔骨髓中成纤维细胞集落形成单位 (CFU F)及成骨性的CFU F集落数量 ,为探讨骨髓的成骨能力提供实验依据。方法 将实验动物分为幼年组 (6个月～ 1岁 )、壮年组 (2 5～ 3 2岁 )两组 ,在无菌状态下 ,在每只动物的股骨上穿刺点抽取骨髓 0 8ml,每个培养瓶 (2 5ml)接种含 8× 10 5个有核细胞的骨髓量 ,在体外培养 12d时计数CFU F集落数量。计数后将培养瓶中CFU F集落进行ALP染色 ,在肉眼下计数着色的CFU F集落 (即成骨性CFU F集落 )数量。结果 按照集落着色的深浅 ,本研究把CFU F集落分为 3种 :①强阳性集落 ;②弱阳性集落 ;③阴性集落。两组成骨性CFU F集落数量约占CFU F集落总数的 1 4 ,主要为弱阳性集落 ,占成骨性CFU F集落的 70 %～ 80 % ;幼年组与壮年组强阳性集落分别占CFU F集落总数的 1 13和 1 17,幼年组明显高于壮年组 ,两组差异有显著性 (P <0 0 5 )。由于每个CFU F集落均来源于一个骨髓基质干细胞 ,通过集落形成率可推算出新西兰白兔骨髓中每 110 0 0～ 130 0 0个有核细胞中有一个骨髓基质干细胞 ,后者仅占骨髓细胞的百万分之一。结论 骨髓中骨髓基质干细胞数量极少 ,成骨性骨髓基质干细胞就更少了 ,但就骨髓细胞总量来说骨髓中骨髓基质干细胞也是相当可观的。

Objective To investigate the number of colony forming unit-fibroblastic (CFU-F) and osteogenic CFU-F colonies in bone marrow in New Zealand white rabbits for evaluating osteogenic potential of marrow.Methods The animals were divided into two groups:juvenile group (6?months-1?year);adult group (2.5?years-3.2?years).0.8ml of marrow were obtained in each animal by aspiration aseptically from superior femoral puncture points (0.3-0.5 cm below the third trochanter apex of femur).A aliquot of marrow containing 8×105 nucleated cells was planted per flask.The CFU-F number in each flask was counted at 12th day and then CFU-F in all flasks were stained for alkaline phosphatase (ALP) activity.The positive CFU-F colonies (namely osteogenic CFU-F colonies) were counted by eye.Results Three kinds of CFU-F colonies were classified according to the degree of ALP staining:strongly positive CFU-F colony (SPC),weakly positive CFU-F colony(WPC)and negative CFU-F colony (NC).The number of osteogenic CFU-F colonies of the two groups accounted for 1/4 of total colonies approximately; osteogenic CFU-F colonies consisted mainly of WPC(70%-80%).The SPC number of juvenile group and adult group accounted for 1/13 and 1/17 respectively,the former being statistically higher than the latter (P<0.05).Because each CFU-F colony derived from one marrow stromal stem cell (MSSC),it could be calculated that using colony forming efficiency there are 1 MSSC per 11000-13000 nucleated cells in the narrow.Conclusion The number of MSSC in the marrow is few and the number of osteogenic MSSC is much fewer than MSSC.Newertheless from the viewpoint of vast amount of marrow cells,the number of MSSC is magnificent.