电离辐射致股骨头坏死早期病理特点和机制的研究

Objective To observe the earlier pathological character and mechanism of radiation osteonecrosis in femoral head, in order to provide evidences for the earlier diagnosis and prevention of radiation osteonecrosis of femoral head. Methods Single femoral head of rats were irradiated singly with 30 Gy of 137 Cs γ-ray. The rats were executed after 2, 6 and 12 weeks, then the femurs were stained with HE and histopatholngical changes were observed by light microscope. The bone marrow mesenchymal stem cells (BMSCs) were cultured after 2 weeks and its proliferation and the colony formation were observed. The rats were endo-perfused with microfili contrast medium 12 weeks later, and the 3-dimensional structure of capillaries by Micro-CT was re-estabhshed to detect the pathological changes of capillaries after irradiation. Result The irradiated femur showed deranged cbondrocyte, decreased osteocyte, shrinking nucleus, increased empty bone lacuna and reduced bone trabocnla (P < 0.05). Micro-CT showed the discontinued small vessels and absence(6.65 %) capillaries in irradiated femur were obviously less than those of the unirradiated (12.3 %)(P < 0.001). The proliferation of BMSCs was slowed, the number of colony in irradiated group (10 %) was less than that of control (21 %) (P < 0.001). Conclusions The preliminary histopathological changes of osteoradionecrosis on femoral head could be increased the empty bone lacuna, and the bone lacuna above 30 % was the sign of the earlier period of osteoradionecrosis. The osteonecrosis of femoral head induced by radiation is not only correlated to the damages to the bone, but also to the damages to BMSCs and capillaries.     

卵巢辐射损伤对大鼠腰椎骨密度和生物力学的影响

目的探讨大鼠卵巢辐射损伤后腰椎骨密度、微结构和生物力学的改变。方法手术暴露大鼠双侧卵巢并应用50Gy的γ射线局部照射,术后90d取大鼠腰椎,DEXA测定骨密度,扫描电镜显示微结构,并行压缩实验检测腰椎最大载荷。结果与假手术组相比,卵巢辐射组大鼠的腰椎骨密度显著减少(P<0.05),骨微结构破坏,生物力学性能下降(P<0.05)。结论卵巢辐射损可导致大鼠腰椎的骨质疏松样改变。     

p38MAPK在17β-雌二醇和雷洛昔芬促成骨细胞増殖和分化中的作用

目的:观察p38MAPK信号转导通路在17β-雌二醇和雷洛昔芬促成骨细胞增殖和分化过程中的作用。方法:取第一继代BALB/c小鼠头盖骨成骨细胞,药物刺激组分别加入不同浓度17β-雌二醇或雷洛昔芬;含阻断剂组预先添加不同浓度的SB202190(p38MAPK的阻断剂)阻断信号转导通路,再加17β-雌二醇或雷洛昔芬,作用72h后用MTT法与PNPP法测定细胞的增殖能力和碱性磷酸酶活性。结果:加入雌激素或雷洛昔芬后,成骨细胞的增殖和分化明显增强,与空白对照组比较差异具有显著性意义(P<0·05);阻断p38MAPK信号转导通路后,细胞增殖分化受到明显抑制,与未阻断组比较差异具有非常显著性意义(P<0.01)。结论:p38MAPK在17β-雌二醇和雷洛昔芬诱导的小鼠成骨细胞的増殖和分化过程中发挥重要作用。     

肾小管上皮细胞对成骨细胞骨代谢相关基因表达的体外调节

目的观察近端肾小管上皮细胞对成骨细胞骨代谢相关基因表达的直接调节作用。方法体外分离培养大鼠近端肾小管上皮细胞与头盖骨成骨细胞;用甲状旁腺激素（PTH）、137^Cs-γ线及25（OH）D3预作用前者,再与后者分为4组共培养：①单培养组（成骨细胞单培养）、②共培养组（肾小管上皮细胞与成骨细胞共培养）、③刺激共培养组（肾小管上皮细胞经PTH＋25（OH）D3处理,再与成骨细胞共培养）、④抑制共培养组（肾小管上皮细胞经137^Cs-γ线照射＋25（OH）D3处理,再与成骨细胞共培养）;并用RT-PCR方法检测共培养体系中成骨细胞的BGP、ALP、ColI、RANKL与OPG mRNA表达。结果共培养组成骨细胞的骨形成相关基因BGP、ALP、Col I mRNA表达较单培养组显著降低（P〈0．001）;刺激共培养组各基因表达较共培养组显著上升（P〈0．05-P〈0．001）;抑制共培养组ALP表达水平较共培养组显著降低（P〈0．001）,BGP与Col I mRNA表达无显著变化。共培养组与单培养组比较,RANKL与OPC;mRNA表达显著降低（P〈0．001）,但RANKL／OPG比值显著高于后者（P〈0．01）;刺激共培养组与共培养组比较,OPG mRNA表达明显上调（P〈0．01）, RANKL mRNA表达变化无显著意义。RANKL／OPG比值显著降低（P〈0．05）;抑制共培养组与共培养组比较, OPG表达显著下降（P〈0．01）,RANKL/OPG比值则显著增高（P〈0．01）。结论培养的近端肾小管上皮细胞对成骨细胞骨代谢相关基因表达具有直接的调节作用。建立的共培养体系可用于骨代谢相关药物筛选及作用机制研究。     

补肾富钙中药对大鼠卵巢辐射损伤所致骨质疏松形态

目的 探讨补肾富钙中药对大鼠卵巢辐射损伤所致骨质疏松形态计量学的影响.方法 40只SD大鼠随机分为四组(n=10),分别为假手术组(灌服1%纤维素90 d)、模型组(以50 Gy γ射线局部照射大鼠卵巢建立卵巢辐射损伤模型,1%纤维素灌服90 d)、尼尔雌醇组(造模后给予尼尔雌醇灌胃90 d)和补肾富钙中药组(造模后给予补肾富钙中药灌胃90 d).各组大鼠于处死前第14天和第4天分别腹腔注射盐酸四环素(30 mg/kg)进行标记,处死后取右侧股骨组织和L4腰椎组织分别进行骨组织形态计量和扫描电镜观察.结果 与模型组比较,补肾富钙中药组和尼尔雌醇组的骨小梁体积显著增加,骨吸收表面减少;尼尔雌醇组的骨形成表面和矿化沉积率下降,但在补肾富钙中药组则无显著变化.扫描电镜观察显示,模型组骨小梁变细、中断、排列稀疏,而补肾富钙中药组和尼尔雌醇组上述变化较轻微.结论 补肾富钙中药可有效预防卵巢辐射损伤诱发的骨质疏松.     

体外培养人成骨细胞Ⅰ型胶原合成与表达的增龄性改变

目的探讨体外培养不同年龄组人成骨细胞Ⅰ型胶原（COL-1）表达的增龄性改变. 方法选择＜5岁、30～40岁、50～60岁及70～79岁4个年龄组骨折患者的松质骨,用1：1 DMEM-Ham F-12培养液进行培养.第2代细胞培养至7 d和13 d,分别定量留取培养48 h的条件培养液,第2代细胞一部分经诱导培养至14 d进行电镜观察,另一部分培养至14 d抽提细胞内总RNA,用逆转录聚合酶链反应方法半定量检测COL-1基因表达,并进行各年龄组比较. 结果成骨细胞经体外诱导培养后透射电镜下观察,细胞外基质中可见大量原胶原;细胞培养至7 d和13 d Ⅰ型前胶原羧基端前肽（PICP）检测值均随年龄增长逐渐降低,与年龄之间呈显著负相关（男性7 d： r=-0.503, P＜0.05, 13 d： r=-0.662, P＜0.01; 女性7 d： r=-0.658, P＜0.01, 13 d： r=-0.770, P＜0.01）;各年龄组成骨细胞均检测到COL-1基因mRNA表达,表达量＜5岁组较低,30～40岁组最高,之后随年龄增长逐渐降低,其表达量和年龄呈显著正相关（男性r=0.331,女性r= 0.327,均为P＜0.05）,性别间差异无统计学意义. 结论体外培养人成骨细胞可合成原胶原,PICP分泌随增龄而降低,不同年龄组人成骨细胞均表达COL-1 mRNA,30～40岁后表达量随年龄增长而降低.     

补肾中药与阿法骨化醇联合应用对卵巢摘除大鼠原发性骨质疏松的防治作用

 目的 研究补肾中药与阿法骨化醇联合应用对卵巢去势（ovariectomy,OVX）大鼠原发性骨质疏松的防治作用。方法 手术摘除6月龄雌性大鼠双侧卵巢,随机分成4组：OVX组（ovariectomy,OVX）、OVX给予补肾中药组（OVX administrated with Bushen Chinese drugs,OVX+TCM）、OVX给予补肾中药和阿法骨化醇组（OVX administrated with Bushen Chinese drugs and alfacalciferol,OVX+TCM+VD）和OVX给予尼尔雌醇组（OVX administrated with nilestriol,OVX+CEE₃）,另设假手术对照组（SHAM）。术后35 d起给予药物至13周后处死取血、子宫、腰椎、股骨等标本,并测定比较各组以下变化：取子宫称重,计算子宫重/体重、子宫内膜厚度;放免法血测血清雌二醇（E₂）应用固体物理密度仪自动测量左股骨和腰椎L₂的骨密度;应用图像分析仪IPP计量软件分析椎体骨的组织病理形态计量;以万能材料测试仪检测右股骨与腰椎L₃的骨生物力学性能的指标;自动生化仪检测血清碱性磷酸酶（ALP）;酶免法检测尿吡啶酚（Pyd）,测尿肌酐（Cr）,计算Pyd/Cr作为骨代谢生化标志物,综合判断骨质疏松情况。结果 OVX大鼠双侧卵巢摘除完全,雌激素缺乏诱发骨质疏松指标明显;联合给药组骨量明显增加,骨结构明显改善,抗骨折性能明显提高,骨形成和骨吸收生化标志物降低;其防治骨质疏松的作用优于单用补肾中药组,并与尼尔雌醇组相仿。结论 补肾中药与阿法骨化醇联合应用对卵巢去势大鼠原发性骨质疏松有明显防治作用,避免了雌激素引起的子宫内膜增生等毒副作用。     

猪大脑灰质组织提取液促进体外培养大鼠成骨细胞的增殖

背景:有研究表明大鼠脑组织提取液能够促进大鼠成骨细胞和骨髓基质干细胞的增殖。目的:观察跨种属脑组织固有成分对体外培养大鼠成骨细胞增殖的影响。方法:提取猪和大鼠大脑灰质、白质、垂体、肢体肌肉组织,分别制成匀浆提取液,并设空白对照组,各组提取液分别用于培养大鼠成骨细胞。比较各种组织提取液对成骨细胞增殖的影响。结果与结论:与空白对照相比,猪和大鼠的大脑灰质、白质和垂体组织提取液均显著促进了成骨细胞的增殖(P<0.05),各组间比较显示猪和大鼠的大脑灰质较白质、垂体和肌肉组织提取液促进成骨细胞增殖的能力更加显著(P<0.05)。结果证实,猪脑组织提取液能跨种属促进大鼠成骨细胞的增殖,其中以大脑灰质的促进作用最为显著。     

电离辐射致股骨头坏死早期病理特点和机制的研究

Objective To observe the earlier pathological character and mechanism of radiation osteonecrosis in femoral head, in order to provide evidences for the earlier diagnosis and prevention of radiation osteonecrosis of femoral head. Methods Single femoral head of rats were irradiated singly with 30 Gy of 137 Cs γ-ray. The rats were executed after 2, 6 and 12 weeks, then the femurs were stained with HE and histopatholngical changes were observed by light microscope. The bone marrow mesenchymal stem cells (BMSCs) were cultured after 2 weeks and its proliferation and the colony formation were observed. The rats were endo-perfused with microfili contrast medium 12 weeks later, and the 3-dimensional structure of capillaries by Micro-CT was re-estabhshed to detect the pathological changes of capillaries after irradiation. Result The irradiated femur showed deranged cbondrocyte, decreased osteocyte, shrinking nucleus, increased empty bone lacuna and reduced bone trabocnla (P < 0.05). Micro-CT showed the discontinued small vessels and absence(6.65 %) capillaries in irradiated femur were obviously less than those of the unirradiated (12.3 %)(P < 0.001). The proliferation of BMSCs was slowed, the number of colony in irradiated group (10 %) was less than that of control (21 %) (P < 0.001). Conclusions The preliminary histopathological changes of osteoradionecrosis on femoral head could be increased the empty bone lacuna, and the bone lacuna above 30 % was the sign of the earlier period of osteoradionecrosis. The osteonecrosis of femoral head induced by radiation is not only correlated to the damages to the bone, but also to the damages to BMSCs and capillaries.