摘要目的 观察唑来膦酸(ZA)对破骨细胞(OC)分化、增殖和骨髓微环境的影响,探讨ZA抗辐射(IR)后小鼠骨量丢失的机制.方法 50只雄性BALB/c小鼠经6.0Gy 60Co γ射线一次性全身辐射后,随机分成辐射组10只,高、中、低剂量治疗组30只和骨髓输注组10只.治疗组分别按0.00025、0.00050、0.00100mg/g进行尾静脉注射ZA,骨髓输注组4h内尾静脉回输异体骨髓细胞2×106个.4周后应用小动物成像仪测定小鼠骨密度,流式细胞术检测骨髓中破骨原始细胞(CD34+CD115+细胞)、破骨前体细胞[CD34+CD115+核因子-κB受体活化因子配体(RANKL)+细胞]及其活化细胞[CD34+CD115+RNAKL+CXC趋化因子受体4(CXCR4)+细胞]的百分比,骨及骨髓病理分析骨小梁及破骨前体细胞的变化,小动物五分类血细胞分析仪检测外周血破骨前体细胞(单核细胞)的百分比及绝对值.流式微球阵列(CBA)和酶联免疫吸附试验(ELISA)法检测血清中OC活化相关因子的含量.结果 辐射组,低、中、高剂量治疗组,骨髓输注组小鼠骨密度分别为(0.51±0.06)、(1.26±0.17)、(2.24±0.29)、(1.27±0.20)、(2.52±0.23)g/cm3.辐射组显著低于各治疗组和骨髓输注组(P均为0.000),中剂量治疗组高于低剂量治疗组和高剂量治疗组(P=0.000、0.942),与骨髓输注组接近(P=0.004).中剂量治疗组CD34+CD115+细胞、CD34+CD115+RANKL+细胞、CD34+CD115+RANKL+CXCR4+细胞的百分比分别为(4.09±0.97)%、(4.11±1.64)%、(18.71±6.23)%,显著低于辐射组[(7.19±1.11)%、(9.01±4.06)%、(40.16±10.31)%],差异有统计学意义(P=0.000、0.000、0.000).与骨髓输注组[(10.14±2.13)%、(2.86±0.82)%、(11.86±3.39)%]接近,差异有统计学意义(P=0.000、0.241、0.056).辐射组小鼠骨病理组织表现为骨小梁变细、中断、疏松,分布不均匀,含大量脂肪空泡充填,局部出现囊性区,原始细胞比例显著下降,骨髓间质血窦充血,局部可见多核细胞.经ZA治疗后,各治疗组小鼠骨小梁增多增粗,髓腔内原始细胞数量升高,变性组织明显减少,其中以中剂量治疗组最为明显.骨髓输注组小鼠骨小梁致密丰满,排列整齐,原始细胞比例明显增多,造血组织结构尚完整.结论 ZA可改善骨髓微环境,刺激骨髓的自我修复,原始细胞数量增加,并抑制其向OC增殖分化,改善IR后小鼠骨量丢失情况.

abstractsObjective To study the mechanism of how zoledronic acid (ZA) inhibit the bone loss in mice which were suffered total ionizing radiation (IR), we would discuss through three aspects including osteoclast (OC) proliferation, OC differentiation and the bone marrow mirco-environment.Methods 50 male BALB/c mice suffered one-time 6.0 dose radiation were randomly divided into 5 groups, 10 in radiation group, 30 in low, middle, high treatment group, 10 in bone marrow group.Low, middle, high treatment groups were each injected with 0.000 25, 0.000 50, 0.001 00 mg/g ZA through caudal vein.Bone marrow group were injected with 2×106 allograft bone marrow cells in 4 hours.4 weeks later, detect bone mineral density by small animal imaging measurement.Count the percentage of osteoclast primitive cells (CD34+CD115+), osteoclast precursor cells [CD34+CD115+ receptor activator of nuclear factor κB ligand (RANKL)+] and its activate cells [CD34+CD115+RANKL+CXC chemokine receptor 4 (CXCR4)+] in bone marrow by flow cytometry.Analyzed bone trabecular and osteoclast precursor cells by bone marrow pathology.Detected the percentage and absolute value of blood cells and osteoclast precursor cells by mall animals 5 classification blood cell analyzer.Assess the level of cytokines in serum related with OC activation by cytometric beads array (CBA) and enzyme-linked immuno sorbent assay (ELISA) kits.Results The bone mineral density of mice in radiation group, low treatment group, middle treatment group, high treatment group and bone marrow group were correspondingly (0.51±0.06), (1.26±0.17), (2.24±0.29), (1.27±0.20), (2.52±0.23) g/cm3.The bone mineral density of mice in radiation group were lower than others in four groups (P=0.000).The bone mineral density of mice in middle treatment group were higher than low treatment group and high treatment group (P=0.000, P=0.942) and got close to bone marrow group (P=0.004).The present of cells with CD34+CD115+, CD34+CD115+RANKL+, CD34+CD115+RANKL+CXCR4+ were (4.09±0.97)%, (4.11±1.64)%, (18.71±6.23)% in middle treatment group, which are absolutely lower than radiation group (7.19±1.11)%, (9.01±4.06)%, (40.16±10.31)% (P=0.000, P=0.000, P=0.000) and moved towards to bone marrow group (10.14±2.13)%, (2.86±0.82)%, (11.86±3.39)% (P=0.000, P=0.241, P=0.056).Bone trabecular in the pathology of mice in radiation group became tenuous, discontinuous and messy.Limited bone marrow primitive cell were observed, which were instead of much necrotic tissue and adipose tissue.We could observe hyperaemia in bone marrow stromal mostly and polykaryocyte locally.After treatment of ZA, bone trabecular became stronger and more continuous and primitive cells were increased in the marrow cavity and degenerative tissues were decreased, especially in middle treatment group.Bone trabecular of mice in bone marrow group become compact, fullness and well-ordered.The percentage of primitive cell was obviously increased.Hemopoietic tissues were integrity.Conclusion ZA can restore bone marrow mirco-environment, accelerate self-rehabilitation.The number of marrow blast was increased and inhibit differentiation of OC.So we could conclude that ZA could prevent the bone mass loss after radiation.