建立自体骨碎末移植材料恢复骨缺损的实验模型

目的:建立恢复种植体周围骨缺损的自体骨碎末骨移植材料的实验模型。方法:实验于2005-08/2006-04在大连医科大学动物实验基地(辽宁省重点实验室)完成。①实验材料:健康杂交家犬5只,体质量15～20kg。Bio-Oss骨移植材料为引导骨/组织再生多孔骨无机材料,白色颗粒状,颗粒大小1.0～2.0mm。②实验方法:拔除家犬下颌第1,2,3前臼齿,3个月后行种植术。预备种植体窝,每只犬左右两侧各预备4个,共40个。在每个种植窝内,各植入种植体钛钉1枚,共40枚。用种植转孔时收集的自体骨碎末、Bio-Oss骨移植材料及两者1∶1混合骨碎末恢复种植体颊侧单壁人为骨缺损,以未植骨作空白对照。③实验评估:第9周时观察各组骨量的恢复情况、X线片观察牙槽骨高度、骨小梁致密度及骨整合情况,亚甲基蓝-碱性品红法观察组织学变化。结果:5只家犬钛钉均无脱落,均纳入结果分析。①一般情况:9周时,创口愈合均良好,钛钉稳定,总存留率为100%。骨缺损处已有不同程度恢复,与正常骨组织无明显差别。②9周时各组骨缺损量的测量结果:植入自体骨碎末、Bio-Oss骨移植材料、混合骨碎末及空白对照组的平均骨缺损量分别为1.8125、1.6975、1.5025、2.6375mm。植入混合骨碎末的平均骨缺损量最小,说明恢复最佳。③X线观察骨量的恢复情况:40颗钛钉外周均与骨组织紧密接触,愈合良好。不同组间未见明显骨质密度区别。④组织学观察骨量的恢复情况:低倍镜下见所有钛钉均被周围淡红色的致密骨组织紧密包绕,种植体与骨组织间无蓝色的软组织,产生了直接骨结合界面。结论:应用家犬建立自体骨碎末移植材料恢复种植体周围骨缺损的实验模型效果理想。     

Western Australian cigarette smokers have fewer small lung nodules than North Americans on CT screening for lung cancer

To determine the prevalence of small lung nodules on low‐dose helical computed tomography (CT) in a Western Australian cohort of asymptomatic long‐term cigarette smokers and to compare this with a large, similarly derived cohort of North Americans from the Mayo Clinic Lung Cancer Screening Trial. Forty‐nine asymptomatic long‐term cigarette smokers of minimum age 50 years underwent a low‐dose 64‐slice helical CT of the lungs. Images were viewed on a soft copy reporting station with thin section axial and coronal images, maximum intensity projection images, and advanced image manipulation tools. The prevalence of all nodules was 39%, significantly lower than the Mayo Clinic cohort prevalence of 51% (P < 0.01, Fisher's exact test), despite the use of more advanced imaging technology and image manipulation designed to increase the sensitivity for nodules. The prevalence of small nodules in asymptomatic long‐term cigarette smokers in Western Australia is high, though significantly less than that found in a large study in North America. The authors postulate this is due to the relatively low rates of mycobacterium tuberculosis and soil‐derived fungal pulmonary infections in Western Australia, as well as a lower degree of urban air pollution.     

Synthesis of fluorescent muramyl dipeptide congeners. 2

Muramyl dipeptide (MDP) analogues were prepared and utilized in the synthesis of new fluorescently labeled MDP derivatives for use as biologic probes. Thus, N alpha-(N-acetylmuramyl)-L-lysyl-D-isoglutamine (Lys-MDP, 4) and N alpha-(N-acetylmuramyl)-L-alanyl-D-isoglutaminyl)-L-lysine [MTP, 5] were synthesized and then reacted with 2-(fluoresceinylamino)-4,6-dichloro-s-triazine (DTAF, 2) to yield the fluorescent adducts, DTAF-Lys-MDP (6) and DTAF-MTP (7). The adjuvant activity of the fluorescent MDP derivatives was determined by the ability of the compounds to promote delayed skin test responses in guinea pigs immunized with ovalbumin (OA) and by evaluating the anti-OA activity of these guinea pigs.     

Determination of absolute amounts of heparin and of dextran sulfate in plasma in microgram quantities.

Heparin and dextran sulfates 8000 are separated from citrated plasma by absorption on epichlorohydrin triethanolamine cellulose columns followed by elution with 1.1 and 1.4 mol/L NaCl in 0.05 mol/L glycine-HCl buffer. The eluate is desalted with Sephadex G25-40, dried, and dissolved in water. A 1 microliters sample is applied to an agarose gel slide. After electrophoresis, the slide is fixed and stained with toluidine blue. The sulfated polysaccharide band(s) is identified by relative electrophoretic migration. The total amount of drug is estimated by matching its optical density with that of a band on one of a set of slides with graded amounts of heparin or dextran sulfate. The reaction with toluidine blue measures the total polyelectrolyte, not just the small proportion of the drug with anticoagulant activity. Pooled normal plasma showed a trace of chondroitin and no heparin. Recovery of heparin and hydrogenated dextran sulfate that was added to pooled normal plasma was complete (lowest concentration tested was 10 micrograms/ml); however, recovery for unhydrogenated dextran sulfate declined consistently by 9 micrograms/ml for concentrations below 50 micrograms/ml, setting a limit for its recovery. Plasma samples taken from patients for coagulation tests were examined by this procedure, and in so doing, steps were ascertained to improve the procedure for routine use. Results were compared with values for prothrombin time and activated partial thromboplastin times obtained on the same samples by the clinical laboratory. Because the procedure provides an independent parameter for measurement in patients who have received heparin therapy, insight into different patient responses to the drug is therefore possible. With minor modifications, the procedure can be used for heparans, dermatans, and chondroitins, because it allows identification and microscale quantitation on the basis of charge, molecular weight, and carbohydrate structure.     

Bcl-2 is an apoptotic target suppressed by both c-Myc and E2F-1.

Malignant transformation occurs in cells that overexpress c-Myc or that inappropriately activate E2F-1. Transformation occurs after the selection of cells that have acquired resistance to apoptosis that is triggered by these oncogenes, and a key mediator of this cell death process is the p53 tumor suppressor. In IL-3-dependent immortal 32D.3 myeloid cells the ARF/p53 apoptotic pathway is inactivated, as these cells fail to express ARF. Nonetheless, both c-Myc and E2F-1 overexpression accelerated apoptosis when these cells were deprived of IL-3. Here we report that c-Myc or E2F-1 overexpression suppresses Bcl-2 protein and RNA levels, and that restoration of Bcl-2 protein effectively blocks the accelerated apoptosis that occurs when c-Myc- or E2F-1-overexpressing cells are deprived of IL-3. Blocking p53 activity with mutant p53 did not abrogate E2F-1-induced suppression of Bcl-2. Analysis of immortal myeloid cells engineered to overexpress c-Myc and E2F-1 DNA binding mutants revealed that DNA binding activity of these oncoproteins is required to suppress Bcl-2 expression. These results suggest that the targeting of Bcl-2 family members is an important mechanism of oncogene-induced apoptosis, and that this occurs independent of the ARF/p53 pathway.     

The DNA damage mark pH2AX differentiates the cytotoxic effects of small molecule HDAC inhibitors in ovarian cancer cells

High grade epithelial ovarian cancers are relatively sensitive to DNA damaging platinum-based chemotherapy, suggesting that the dependencies of ovarian tumors on DNA damage response pathways can be harnessed for therapeutic purposes. Our goal was to determine if the DNA damage mark gamma-H2AX phosphorylation (pH2AX) could be used to identify suitable cytotoxic histone deacetylase inhibitors (HDACi) for ovarian cancer treatment. Nineteen chemically diverse HDACi compounds were tested in 7 ovarian cancer cell lines. Fluorescent, biochemical and cell-based assays were performed to assess DNA damage by induction of pH2AX and to measure cell viability and apoptosis. The relationships between pH2AX and the cellular effects of cell viability and apoptosis were calculated. Selected HDACi were tested in combination with cisplatin and other DNA damaging agents to determine if the HDACi improved upon the effects of the DNA damaging agents. The HDACi compounds induced differing levels of pH2AX expression. High levels of pH2AX in HDACi-treated ovarian cancer cells were tightly associated with decreased cell viability and increased apoptosis. Consequently, a ketone-based HDACi was chosen and found to enhance the effects of cisplatin, even in ovarian cancer cells with extreme resistance to DNA damaging drugs. In conclusion, a fluorescent-based assay for pH2AX can be used to determine cellular responses to HDACi in vitro and may be a useful tool to identify potentially more effective HDACi for the treatment of ovarian cancer. In addition, these results lend support to the inclusion of ketone-derived HDACi compounds for future development.     

Correlation between patterns of horizontal connectivity and the extend of short-term representational plasticity in rat motor cortex

Plasticity of representational maps in adult cerebral cortex has been documented in both sensory and motor cortex, but the anatomical basis for cortical plasticity remains poorly understood. To investigate horizontal connectivity in primary motor cortex (M1) as a putative anatomical substrate for short-term, functional plasticity of adult motor cortical representations, a combination of electrical stimulation and biocytin labeling was used to examine pre-existing patterns of intrinsic connections in adult rat M1 in relationship to the pattern of reorganization of the motor movement may induced by transection of the contralateral facial nerve. Two hours after nerve cut, small, circumscribed regions of the forelimb representation expanded medially into territory previously devoted to the vibrissae representation. Outside of this novel, expanded forelimb region, no forelimb movement could be evoked from the former vibrissae representation at any time over the period of hours tested, thus representing silent cortex. Injections placed into vibrissae cortex representing the newly expanded forelimb representation gave rise to labeled axons and dense terminal fiber labeling which crossed the forelimb/vibrissae border and extended up to 1.2 mm within the low-threshold forelimb representation. In contrast, injections placed into silent vibrissae cortex gave rise to labeled axons and terminal boutons which remained mostly restricted to the original vibrissae representation, with only sparse projections that crossed into the low-threshold forelimb representation. Thus, these results suggest that the extent of short-term, functional reorganization of M1 induced within the first several hours following peripheral nerve cut is mediated, and constrained, by an anatomical framework of pre-existing, horizontal projections which traverse representation borders.      

南蛇藤素抑制豚鼠体外精子的受精能力

南蛇藤素(Cel)对豚鼠精子前向运动(FM)、获能(Cap)、顶体反应(AR)和穿透去透明带仓鼠卵(SPA)均有明显的抑制作用,其作用强度随剂量而增加;对Cel的敏感性依次为精子Cap>FM>SPA>AR。Cel对豚鼠精子AR,FM和Cap的抑制作用比乙酸棉酚(GA)明显为强。     

Implementation of an image-guided radiation therapy program: Lessons learnt and future challenges

The aim of this paper is to detail the experience obtained in implementing an image-guided radiation therapy program at the Northern Sydney Cancer Centre. This required retrofitting a Varian Clinac 21EX with an on-board imager. The commissioning and quality assurance procedures, organisation of a multidisciplinary image guided radiation therapy group, and the development of clinical protocols for orthogonal kV and cone beam computed tomography implementation are described. Reassessment of the image-guided radiation therapy program has continued as new equipment and software versions were made available in the department.