不耐热肠霉素和耐热肠毒素基因的融合

用重组基因技术将产肠毒素大肠杆菌（ＥＴＥＣ）的耐热肠毒素（ＳＴａ）基因与不耐热肠毒素Ｂ亚单位（ＬＴ－Ｂ）基因融合。表达的ＬＴ－Ｂ／ＳＴａ融合蛋白，既有ＬＴ－Ｂ的抗原性又有ＳＴａ的抗原性。由于基因融合的方式不同，对ＳＴａ的抗原性影响很大，但对ＬＴ－Ｂ的抗原性没有影响。ＬＴ－Ｂ／ＳＴａ融合多肽保留了ＥＴＥＣ肠毒素结合ＧＭ－１神经节甙酯的能力，却仍具有ＳＴａ的生物毒性。     

四种骨替代材料的人成骨细胞生物相容性研究

作者采用人胚成骨细胞体外培养法研究了活性玻璃陶瓷（ＢＧＣ）、羟基磷灰石（ＨＡ）、钛（Ｔｉ）、钴铬合金（ＣｏＣｒ）四种常用的骨替代材料对细胞附着、伸展、生长及代谢方面的影响，结果发现ＢＧＣ与ＨＡ可以促进细胞早期附着、增殖和细胞内钙合成，Ｔｉ无上述作用，而ＣｏＣｒ对细胞吸附、碱性磷酸酶活性及钙合成均有一定的抑制作用，因此认为ＣｏＣｒ不宜用作骨内种植材料。此外还发现材料对成骨细胞代谢的抑制作用要早于对细胞增殖的影响，这为进一步探讨实验细胞的敏感性问题积累了资料     

不耐热肠毒素和耐热肠毒素基因的融合

用重组基因技术将产肠毒素大肠杆菌（ＥＴＥＣ）的耐热肠毒素（ＳＴａ）基因与不耐热肠毒素Ｂ亚单位（ＬＴ－Ｂ）基因融合。表达的ＬＴＢ／ＳＴａ融合蛋白，既有ＬＴ－Ｂ的抗原性又有ＳＴａ的抗原性。由于基因融合的方式不同，对ＳＴａ的抗原性影响很大，但对ＬＴ－Ｂ的抗原性没有影响。ＬＴ－Ｂ／ＳＴａ融合多肽保留了ＥＴＥｃ肠毒素结合ＧＭ－１神经节甙酯的能力，却仍具有ＳＴａ的生物毒性。     

肝细胞肝癌和肝硬化组织中丙型肝炎病毒各区段抗 …

目的 研究丙型肝炎病毒（ＨＣＶ）在肝细胞肝癌（ＨＣＣ）和肝硬化（ＬＣ）中所起的作用。结合乙型肝炎病毒（ＨＢＶ）进行分析，并初步探讨了ＨＣＶ与ＨＢＶ感染是否有相互促进作用。方法 采用ＨＣＶ－Ｃ，Ｅ，ＮＳ３，ＮＳ４区单克隆抗体，ＨＢｓＡｇ多克隆抗体用免疫组化方法检测了５９例ＨＣＣ及３５例ＬＣ组织标本。结果 ＨＣＶ阳性反应主要分布在肝细胞及癌细胞的胞浆内，呈细颗粒状。ＨＣＣ中，ＨＣＶ感染率；北京（２９例     

植入微型刺激器对大鼠神经损伤后自残及神经再生的影响

本实验利用大鼠坐骨神经钳夹损伤模型，观察了电针（Ａ组）、直流刺激器（Ｂ组）及双向平衡电脉冲刺激器（Ｃ组）对神经损伤后自残及神经再生的影响，Ｄ组为对照。结果：（１）术后１７ｄ，Ｃ组的右后足趾自残率为１３％，而Ａ、Ｂ及Ｄ组分别为４６％、５３％及８０％，Ｃ组的自残率显著低于其它各组（Ｐ＜０．００１）；（２）右侧屈肌反射阈：当刺激强度为３５±５Ｖ（Ｃ组）、４５±５Ｖ（Ａ和Ｂ组）时，可引出短潜伏期（４５～１５０ｍｓ）的Ａ类及长潜伏期（１２５～５２５ｍｓ）Ｃ类传入纤维反应，而当刺激强度增至６５±５Ｖ时，于Ｄ组仅见长潜伏期（１９０～６００ｍｓ）的Ｃ类传入纤维反应；（３）术后１７ｄ脊神经节的标记细胞百分享分别为１１．１％（Ｃ组）、５．７％（Ｂ组）、５．２％（Ａ组）及１．１％（Ｄ组）；脊髓前角的标记细胞百分率分别为１６．６％（Ｃ组）、８．１％（８组）、７．４％（Ａ组）及１．９％（Ｄ组）。上述结果表明：双向平衡电脉冲刺激器在抑制神经损伤后自残及促进周围神经再生等方面具有更显著的生理功效。     

肺淋巴上皮瘤样癌与EB病毒的相关性

目的为阐明肺淋巴上皮瘤样癌（ｌｙｍｐｈｏｅｐｉｔｈｅｌｉｏｍａ－ｌｉｋｅｃａｒｃｉｎｏｍａ，ＬＥＬＣ）与ＥＢ病毒（ＥＢＶ）的关系。方法对３０例肺ＬＥＬＣ和１９例肺非ＬＥＬＣ的石蜡包埋组织，采用原位杂交、免疫组化技术检测ＥＢＶ的存在情况及其基因表达特征。结果肺ＬＥＬＣ中原位杂交ＥＢＶ小ＲＮＡ（ＥＢＥＲｓ）阳性率为９３．３％（２８／３０）；而免疫组化潜在膜蛋白（ｌａｔｅｎｔｍｅｍｂｒａｎｅｐｒｏｔｅｉｎ，ＬＭＰ１）阳性率分别为５３．３％（１６／３０）和２３．３％（７／３０）；１９例病毒衣壳抗原（ｖｉｒｕｓｃａｐｓｉｄａｎｔｉｇｅｎ，ＶＣＡ）阳性率为２３．３％（７／３０）；１９例非ＬＥＬＣ肺癌中原位杂交检测ＥＢＥＲｓ、免疫组化检测ＬＭＰ１均为阴性。结论肺ＬＥＬＣ与ＥＢＶ密切相关，ＥＢＶ可能在肺ＬＥＬＣ发生发展中起重要的作用；ＥＢＶ在肺ＬＥＬＣ中主要以潜伏状态存在，只有少量病毒进入溶解周期。     

猪生物骨载体成份分析和对比研究

猪生物骨载体成份分析和对比研究胡晓波，吴多能（海南省人民医院，海南５７０３１１）１引言近年来，在关于骨诱导研究的文献中，经常可看到“载体”（Ｃａｒｒｉｅｒ）这个词。因为骨形态发生蛋白（ＢｏｘｅＭｏｒｐｈｏｇｅｎｅｔｌｃＰｒｏｔｅｉｎ，ＢＭＰ）含量甚微...     

应用YAC探针进行荧光原位杂交检测慢性粒细胞白血病BCR基因重排

为检测慢性粒细胞白血病（ＣＭＬ）中ＢＣＲ基因重排，采用酵母人工染色体（ＹＡＣ）ＤＮＡ的Ｉｎ－ｔｅｒ－Ａｌｕ－ＰＣＲ产物为探针进行荧光原位杂交（ＦＩＳＨ）研究了１０例ＣＭＬ，其中包括初诊患者２例，ＣＭＬ急变并接受化疗２例，α干扰素治疗２例，自体骨髓移植术（ＡＢＭＴ）后３例和Ｐｈ染色体阴性ＣＭＬ１例。同时进行细胞遗传学和ＲＴ－ＰＣＲ检测。结果：９例ＣＭＬ的４６％～１００％的可分析核型显示ｔ（９；２２）易位，其中携带ｔ（９；２２）细胞最少者为１例自体骨髓移植术后８个月的患者，其６４％的分裂相存在ｔ（９；２２），３６％为正常核型；１例Ｐｈ（－）ＣＭＬ未见ＢＣＲ基因易位，而ＲＴ－ＰＣＲ（＋），提示ＡＢＬ基因片段插入２２ｑ１１，造成隐匿性Ｐｈ染色体。结果表明：应用ＹＡＣ探针进行原位杂交的定位明确。ＦＩＳＨ检测微小残留病（ＭＲＤ）比常规细胞遗传学方法更敏感，而且可以完成ＰＣＲ方法不易进行的定量分析。     

用热启动聚合酶链反应检测血清中乙型肝炎病毒DNA技术的建立和应用

建立了热启动聚合酶链反应（ＰＣＲ）检测乙型肝炎病毒ＤＮＡ（ＨＢＶＤＮＡ）的技术。ＰＣＲ所有反应成分被２次加样。先加Ａ液（含ｄＮＴＰｓ、一对引物和ＭｇＣｌ＿２）与一粒石蜡珠。７０℃加热后冷却至室温，使蜡珠先融化后凝固形成蜡盖封住Ａ液，然后在向其上加入Ｂ液（含耐热性ＤＮＡ聚合酶、ＨＢＶＤＮＡ模板和ＫＣｌ）并开始循环扩增。当反应管内第一次变性温度升至６０℃以上时，中隔蜡层融化，蜡上浮形成防蒸发屏障，Ａ、Ｂ两液则由于热分子运动而混匀，从而保证引物与靶基因在较高温度下严格退火以减少错导非特异性扩增和引物聚体形成。提高了ＰＣＲ的特异性和灵敏性，应用这种技术对７６例肝炎血清进行了检测，结果ＨＢＶＤＮＡ检出率为６８％，其中ＨＢｓＡｇ（＋）、ＨＢｅＡｇ（＋）、抗－ＨＢｃ（＋）血清检出率为１００％；ＨＢｓＡｇ（＋）、抗－ＨＢｅ（＋）、抗－ＨＢｃ（＋）血清检出率为７０％；ＨＢｓＡｇ（＋）、抗－ＨＢＣ（＋）血清检出率为８９％；抗ＨＢＣ（＋）血清检出率为对％；标记全阴性血清检出率为３３％。     

MTT比色法测定hM－CSF生物活性的研究

本文以小鼠骨髓细胞（ＢＭＣ）和经条件化筛选的分化程度较一致的小鼠骨髓巨噬细胞（ＢＭＭ）为研究对象，摸索了用ＭＴＴ法测定人巨噬细胞集落刺激因子（ｈＭ－ＣＳＦ）生物活性的最佳条件。结果显示ＢＭＣ细胞的线性检测范围为６００～２５００ＣＦＵ／ｍｌ；ＢＭＭ细胞的线性检测范围明显改善，为１５０～１２５０ＣＦＵ／ｍｌ。该法的建立与改善提供了简便、可靠、适于检测大批量Ｍ－ＣＳＦ样品生物活性的方法。     

The characteristics of a hydroxyapatite-chitosan-PMMA bone cement

In this study, we propose a new bioactive bone cement (BBC), composed of natural bone powder (hydroxyapatite; HA), chitosan powder, and the currently available polymethylmethacrylate (PMMA) bone cement, for use in orthopedic surgeries such as vertebroplasty or as bone filler. Three types of BBCs (BBC I, BBC II, and BBC III) were prepared with different composition ratios. In vitro tests and animal studies were performed with the new BBCs, and with a currently available commercial PMMA bone cement. Surface morphology, chemical composition, changes in pH over time, exothermic temperatures, intrusion, and cellular responses were investigated in vitro. Scanning electron microscopy (SEM) and radiological and histological examinations were performed in animal studies. The results showed that the major components of the BBCs were C, O, Ca, P, Cl, Si, S, Ba, and Mg. The pH values of the BBCs decreased after 1 day, but eventually recovered to 7.2-7.4. The water absorbency, weight loss, and porosity of the BBCs were higher than those of pure PMMA, but the compressive Young's modulus and the ultimate compressive strength (UCS) of the BBCs were lower than those of pure PMMA. The exothermic temperatures of the BBCs were considerably lower than that of pure PMMA. BBC II and III required longer times to solidify than did pure PMMA. Intrusion tests showed that the BBCs were more intrusive than was pure PMMA. Cell proliferation tests demonstrated that BBC II was preferable to pure PMMA for cell attachment and proliferation. No cytotoxic characteristics were found associated with any of the BBCs. In animal tests, BBC II was more biocompatible and osteoconductible than was pure PMMA. The results of in vitro and animal studies indicated that the proposed BBCs have potential clinical application as replacements for the pure PMMA bone cements currently in use.     

载药硼酸盐骨水泥的制备及其体外生物活性和抗菌性能的研究

目的以硼酸盐生物活性玻璃和改性壳聚糖液相制备了新型的硼酸盐骨水泥,同时负载骨髓炎治疗药物硫酸庆大霉素,考察其体外抗菌性能,以探讨其治疗骨髓炎的可能性。方法以硼酸盐骨水泥为载体,制备了负载硫酸庆大霉素(GS)的骨水泥。探究负载GS对骨水泥的可注射性能、初凝时间的影响;将负载GS且预固化的硼酸盐骨水泥浸泡于磷酸盐缓冲溶液(PBS)中,考察其体外生物活性、生物降解性和药物释放;利用抑菌圈实验评估了负载GS的硼酸盐骨水泥的体外抗菌性能。结果制备的载药硼酸盐骨水泥能够被完全注射,初凝时间约6 min;在体外的磷酸盐缓冲溶液(PBS)中浸泡时,GS能够持续稳定的释放,药物释放长达26天;负载GS的硼酸盐骨水泥能够很好地抑制金黄色葡萄球菌(S.aureus)和大肠杆菌(E.coli)的生长。结论制备的负载GS的硼酸盐骨水泥具有优异的可注射性,合适的原位自固化时间,长期持续的药物释放和抗菌性能,可以用于骨髓炎治疗的进一步研究。     

Evaluation of the biphasic calcium composite (BCC), a novel bone cement,in a minipig model of pulmonary embolism

Polymethylmethacrylate (PMMA) bone cement, which is used as a filler material in vertebroplasty, is one of the major sources of pulmonary embolism in patients who have undergone vertebroplasty. In the present study, we established and evaluated two animal models of pulmonary embolism by injecting PMMA or biphasic calcium composite (BCC) bone cement with a negative surface charge. A total of 12 adults and healthy Wuzhishan minipigs were randomly divided into two groups, the PMMA and BBC groups, which received injection of PMMA bone cement and BBC bone cement with a negative surface charge in the circulation system through the pulmonary trunk, respectively, to construct animal models of pulmonary embolism. The hemodynamics, arterial blood gas, and plasma coagulation were compared between these two groups. In addition, morphological changes of the lung were examined using three-dimensional computed tomography. The results showed that both PMMA and BCC injections induced pulmonary embolisms in minipigs. Compared to the PMMA group, the BCC group exhibited significantly lower levels of arterial pressure, pulmonary artery pressure, blood oxygen pressure, blood carbon dioxide pressure, blood bicarbonate, base excess, antithrombin III and D-dimer. In conclusion, BCC bone cement with a negative surface charge is a promising filler material for vertebroplasty.     

益骨胶囊含药血清对大鼠成骨细胞IL-6 mRNA及其蛋白表达的影响

目的：从mRNA及蛋白水平探讨益骨胶囊含药血清对成骨细胞表达IL-6的影响。方法：分离、培养新生SD大鼠成骨细胞，传代后分为3组即含药血清组；空白血清组；DMEM组。采用RT-PCR法检测IL-6 mRNA相对表达量，用放射免疫法检测成骨细胞培养上清中的IL-6含量。结果：益骨胶囊含药血清组IL-6 mRNA相对表达量显著低于对照组（P<0.05）；益骨胶囊含药血清组IL-6蛋白表达量也低于对照组（P<0.05）。结论：益骨胶囊含药血清能下调成骨细胞IL-6 mRNA 表达；亦能在蛋白水平降低成骨细胞分泌骨吸收因子IL-6，这可能是益骨胶囊防治骨质疏松症的机制之一。     

Are bilateral breast cancers and breast cancers coexisting with ovarian cancer different from solitary tumors? A pair‐matched immunohistochemical analysis aimed at intrinsic tumor phenotype

Patients with bilateral breast cancer (BBC) and breast‐ovarian cancer syndrome (BOCS) constitute populations potentially enriched for molecular defects involved in the pathomechanisms of these malignancies. The aim of our study was to compare tumor morphology and expression of estrogen and progesterone receptor, HER2, Ki67, cytokeratin 5/6, E‐cadherin, vimentin and epidermal growth factor receptor in tissue microarrays from 199 tumors from BBC or BOCS patients and 199 age‐matched solitary tumors. Compared to controls, BBC and BOCS considered jointly had lower incidence of DCIS, lower expression of PgR and HER2, and higher expression of Ki67 and vimentin. BOCS tumors were of higher grade, had lower expression of ER and PgR and higher expression of Ki67, CK5/6, vimentin and EGFR. BBC had less DCIS component, lower HER2 expression and higher Ki67 expression. Metachronous BBC (mBBC) had lower expression of ER, PgR and HER2, and higher expression of Ki67 and vimentin. Synchronous BBC (sBBC) had less DCIS component, higher expression of ER, and lower expression of CK5/6, EGFR and E‐cadherin. BBC and breast cancers in BOCS differ in many aspects from solitary tumors. BBC are a heterogeneous group of tumors, differing between sBBC and mBBC. mBBC phenotype shares many features with BOCS tumors.     

A delivery system of linezolid to enhance the MRSA osteomyelitis prognosis: in vivo experimental assessment

Staphylococcus aureus, a major responsible microorganism of osteomyelitis, represents a challenge to treat because of the poor penetration of antibiotics in bone and increasing minimum inhibitory concentrations (MICs) to glycopeptides. The calcium-deficient apatites (CDA), closer to the biological components found in bone and other calcified tissues, have osteoconductive properties. So, to process severe osseous infections, CDA can be used to deliver in the infectious site antibiotics like linezolid. The acute experimental osteomyelitis due to methicillin-resistant Staphylococcus aureus (MRSA) was induced in rabbit’s femurs and surgery mimicking human procedures was performed at day three after inoculation. Animals were randomly assigned to treatment groups: L_(IV) [4-day linezolid IV infusion, human-equivalent dose of 10 mg/kg/12 h], L_(CDA50%) (100 mg CDA with linezolid 500 μg/mg) and L_(CDA50%)?+?L_(IV). Surviving bacteria were counted in bone marrow (BM) and bone (Bo) at day 3 (before treatment), day 7 (4-day treatment) or day 17 (14-day treatment). L(iv) was effective after a 4-day treatment with a log₁₀CFU/g decrease of ?2.63?±?1.92 and ?2.17?±?1.58 in bone marrow and bone, respectively. CDA loaded with linezolid enhance the efficacy of the IV linezolid regimen by more than one log₁₀CFU/g.     

Porous titanium and silicon-substituted hydroxyapatite biomodification prepared by a biomimetic process: Characterization and in vivo evaluation

Porous titanium with a pore size of 150–600 μm and a porosity of 67% was prepared by fiber sintering. The porous titanium had a complete three-dimensional (3D) interconnected structure and a high yield strength of 100 MPa. Si-substituted hydroxyapatite (Si-HA) was coated on the surface by a biomimetic process to improve the surface bioactivity. X-ray diffraction results showed that Si-HA coating was not well crystallized. New bone tissue was found in the uncoated porous titanium after 2 weeks of implantation and a significant increase (p < 0.05) in the bone ingrowth rate (BIR) was found after 4 weeks of implantation, indicating the good osteoconductivity of the porous structure. The HA-coated and Si-HA-coated porous titanium exhibited a significantly higher BIR than the uncoated titanium at all intervals, highlighting the better surface bioactivity and osteoconductivity of the HA- and Si-HA coatings. Also, the Si-HA-coated porous titanium demonstrated a significantly higher BIR than the HA-coated porous titanium, showing that silicon plays an active role in the surface bioactivity. For Si-HA-coated porous titanium, up to 90% pore area was covered by new bone tissue after 4 weeks of implantation in cortical bone. In the bone marrow cavity, the pore spaces were filled with bone marrow, displaying that the interconnected pore structure could provide a channel for body fluid. It was concluded that both the 3D interconnected pore structure and the Si-HA coating contributed to the high BIR.     

聚甲基丙烯酸甲酯骨水泥与Cortoss骨水泥生物力学性能及聚合温度比较

背景：聚甲基丙烯酸甲酯骨水泥是其最常用的一种填充材料,但由于其存在聚合时放热、单体毒性等缺点,所以目前出现了以Cortoss为代表的生物新型骨水泥。 目的：对比两种骨水泥在体外的力学性能。 方法：将聚甲基丙烯酸甲酯与新型骨水泥Cortoss按照ISO5833:2002标准分别制作成抗压及抗弯模型,将制作的模型使用生物力学机测试两种骨水泥的抗弯及抗压模量,同时在制模过程中测量骨水泥聚合温度。 结果与结论：与聚甲基丙烯酸甲酯骨水泥比较,Cortoss骨水泥在聚合过程中最高温度较低、抗压强度较强、抗弯模量降低,差异均有显著性意义(P < 0.05),而抗弯强度间差异无显著性意义(P > 0.05)。说明Cortoss生物力学强度优于聚甲基丙烯酸甲酯骨水泥,由于Cortoss在聚合时放热较少,所以其用于手术时有较好的安全性。      

生物可吸收高强度左旋聚丙交酯材料在体内的力学特征

目的观察生物可吸收固态压缩法增强的、高强度左旋聚丙交酯poly-L-lactide,PLLA材料在体内的力学变化特征,评价其作为骨固定装置材料的价值. 方法用特殊的加热压缩方法-固态压缩法(solid-state compressionSC)加工左旋聚丙交酯,得SC-PLLA试棒(3.2mm×30mm),植入兔的皮下和股骨内,在长达48周的降解时间里,观察材料的力学性能(弯曲强度和剪切强度)和扫描电镜(SEM)下的微观形态. 结果植入24周后,各组SC-PLLA在体内的力学强度均可维持在弯曲强度180Mpa以上,剪切强度75Mpa以上,它们都大于皮质骨强度.剪切强度的下降比弯曲强度快.SEM见SC-PLLA降解前内部有大量排列整齐的纵向纤维,并随降解而破坏并出现孔隙. 结论固态压缩法可获得高初始强度和维持强度的PLLA,满足一般的骨内固定要求.SC-PLLA是一种有前途的生物可吸收接骨材料.     

骨的应力适应性

加载骨细胞或成骨细胞膜上的力传导器,通过对第二信使、生长因子、骨基质、原癌基因等的作用,促进骨代谢。骨的应力反应与骨所受应力大小成正比,应力的大小、性质、作用时间与骨的应力适应性均有关系。