改性羟基磷灰石骨修复纳米复合材料的制备及生物学评价

目的:制备羟基磷灰石/聚乳酸聚乙醇酸骨修复材料,并对其进行生物学评价。方法:实验于2006-06/2007-02在中科院长春应用化学研究所完成材料制备,在吉林大学基础医学院实验动物中心完成动物实验。将低聚乳酸的羧基与羟基磷灰石表面的钙原子用化学键连接,得到表面接枝聚左旋乳酸的羟基磷灰石,将其与聚乳酸聚乙醇酸共混,得到复合材料PLLA-g-HA/PLGA。溶于氯仿后铺膜(厚0.2mm),用DMEM培养液浸泡材料膜制备浸提液。首先,进行材料生物安全性实验:①细胞毒性实验:将浸提液与培养液混合,接种兔成骨细胞,培养24h,MTT法检测细胞增殖,计算细胞增殖率和细胞毒性级(细胞毒性级0或1级为合格)。②全身毒性实验:小鼠以50mL/kg的剂量静脉注射浸提液,观察72h内小鼠中毒症状。③皮肤刺激实验:兔脊柱两侧皮内注射材料浸提液,观察72h内皮肤有无异常反应。④热原实验:自兔耳缘静脉注入浸提液(10mL/kg)。注射后每0.5h测肛温1次,共6次,以6次中最高的1次减去正常体温,计为升高度数。其次,对复合材料进行细胞黏附性检测:将复合材料制成1%氯仿溶液,涂于硅化的盖玻片上,置于6孔板,每孔接种1×105个成骨细胞,培养3d,在2,24,72h行FITC荧光染色,数码摄像系统拍摄细胞荧光照片。结果:制备了新型PLLA-g-HA/PLGA复合材料。①生物安全性实验结果:MTT实验检测复合材料细胞增殖率为94.8%,细胞毒性级为1级;全身毒性实验中动物无死亡、惊厥、瘫痪、呼吸抑制、腹泻和体质量下降等不良反应;热原实验中兔体温最大的变化值是0.25℃(国家标准为<0.6℃);皮肤刺激实验中未见任何刺激反应,无红斑、焦痂、水肿表现。②细胞黏附性实验结果:细胞接种后2h可见少量细胞开始贴壁;24h时可见贴壁细胞明显增多,并呈聚集生长;培养3d后可见细胞逐渐融合,细胞状态良好。结论:新型PLLA-g-HA/PLGA复合材料符合生物材料细胞毒性要求,按毒性剂量分级属无毒级,无致热原性、对皮肤无刺激作用,具有良好的生物相容性和细胞黏附性。     

Fused pyrazine mono-n-oxides as bioreductive drugs. II Cytotoxicity in human cells and oncogenicity in a rodent transformation assay

PURPOSE: To determine what structural moieties of the fused pyrazine mono-N-oxides are determining factors in their in vitro cytotoxicity and oncogenicity. METHODS AND MATERIALS: A new series of experimental bioreductive drugs, fused pyrazine mono-N-oxides, was evaluated in vitro for aerobic and hypoxic cytotoxicity in the HT29 human colon adenocarcinoma cell line by using clonogenic assays. The relative oncogenicities of these compounds were also determined in aerobic cultures of C3H 10T1/2 mouse embryo fibroblasts by using a standard transformation assay. RESULTS: Removal of the 4-methyl piperazine side chain from the parent compound, RB 90740, reduced the potency of the hypoxic cytotoxin. Reduction of the N-oxide function increased the aerobic cytotoxicity and eliminated most of the hypoxic/aerobic cytotoxic differential. The reduced N-oxide also had significant oncogenicity, consistent with a mechanism of genotoxicity following bioreduction of RB 90740. CONCLUSION: This new series of bioreductive compounds may be effective in cancer therapy, particularly the lead compound RB 90740. The oncogenic potential of these compounds is similar to that for other cancer therapies. Further studies should include evaluation of these compounds in vivo and the development of analogs with reduced oncogenic potential and retention of the hypoxic/aerobic cytotoxicity differential.     

Influence of late Pleistocene sea-level variations on midocean ridge spacing in faulting simulations and a global analysis of bathymetry

It is established that changes in sea level influence melt production at midocean ridges, but whether changes in melt production influence the pattern of bathymetry flanking midocean ridges has been debated on both theoretical and empirical grounds. To explore the dynamics that may give rise to a sea-level influence on bathymetry, we simulate abyssal hills using a faulting model with periodic variations in melt supply. For 100-ky melt-supply cycles, model results show that faults initiate during periods of amagmatic spreading at half-rates >2.3 cm/y and for 41-ky melt-supply cycles at half-rates >3.8 cm/y. Analysis of bathymetry across 17 midocean ridge regions shows characteristic wavelengths that closely align with the predictions from the faulting model. At intermediate-spreading ridges (half-rates >2.3 cm/y and ≤3.8 cm/y) abyssal hill spacing increases with spreading rate at 0.99 km/(cm/y) or 99 ky (n = 12; 95% CI, 87 to 110 ky), and at fast-spreading ridges (half-rates >3.8 cm/y) spacing increases at 38 ky (n = 5; 95% CI, 29 to 47 ky). Including previously published analyses of abyssal-hill spacing gives a more precise alignment with the primary periods of Pleistocene sea-level variability. Furthermore, analysis of bathymetry from fast-spreading ridges shows a highly statistically significant spectral peak (P < 0.01) at the 1/(41-ky) period of Earth’s variations in axial tilt. Faulting models and observations both support a linkage between glacially induced sea-level change and the fabric of the sea floor over the late Pleistocene.

Parallel to the global system of midocean ridges is an orderly progression of fault-bounded topographic highs called abyssal hills. Hypothesized models for abyssal-hill formation generally involve magma delivery rates, either directly through variable magma emplacement at the surface (1) or indirectly by modulating the amount of plate extension that must be accommodated by faulting (2–5). Faults would be especially likely to occur during times of reduced magma supply, when stress accumulates as young lithosphere is pulled away from a spreading axis (6, 7).Abyssal hills exhibit regularity in spacing and symmetry across the ridge axis (1) that has motivated hypotheses of periodic or quasiperiodic variability in magma supply (4, 6–10), but the causes of such variability have been obscure. One proposed mechanism involves sea-level change driven by glacial cycles (11–13). Melting beneath ridges is driven by depressurization of upwelling mantle and hence the melting rate can be modulated by pressure variations associated with fluctuating sea level. During the last deglaciation, for example, sea level rose by ∼1 cm/y for 10,000 y, suggesting an ∼10% reduction in rates of melt generation for mantle upwelling at 3 cm/y, where the factor of 3 difference in density between mantle and seawater is taken into account. More sophisticated models that account for melt migration also predict sea-level–induced variations in melt supply beneath ridges (14) and that such variations will depend upon factors including spreading rate and mantle permeability (12).Throughout the late Pleistocene, variations in sea level occur most prominently at 18- to 23-, 41-, and 100-ky periods (15). The 18- to 23- and 41-ky periods are, respectively, associated with variations in the orientation and tilt of Earth’s spin axis, whereas the origin of the 100-ky variability appears to involve a combination of orbital variations, glacial dynamics, and variations in atmospheric CO₂ (16). Although the 100-ky cycle dominates the magnitude of sea-level variability during the late Pleistocene, melt-supply variations respond to the rate of change of sea level, accentuating the importance of higher-frequency variations (SI Appendix, Fig. S1) (12, 13).The hypothesis that Pleistocene variations in sea level influence melt supply is supported by several lines of evidence. Proxies of ridge hydrothermal circulation indicate a response to sea-level change (17–19), as do variations in the abundance of sediment-hosted volcanoclastics (20). Axis-parallel variations in crustal thickness along the East Pacific Rise of 200 to 800 m vary with wavelengths that are consistent with the sea-level hypotheses (21). Bathymetry along portions of the Australian–Antarctic ridge (12) and Chile Rise (22) was found to contain significant concentrations of variability at orbital and 100-ky periods. Finally, an analysis of characteristic spacing involving five regions whose half-spreading rate exceeds 3 cm/y found that the spacing across the fastest-spreading regions appeared to increase with spreading rate, consistent with the possibility that orbital cycles influence spacing (23).It has been argued, however, that sea-level–induced variations in melt supply would have negligible implications for surface bathymetry on two counts. First, the presence of a magma chamber at the ridge axis could damp the influence of melt supply, and, second, flexural rigidity limits surface bathymetry variations associated with emplacement of magma at the base of the crust (24). Furthermore, there is a long-standing interpretation that the regularity in midocean ridge bathymetry reflects preferred length scales inherent to lithospheric structural controls (3, 25, 26), such that the presence of quasiperiodicity in bathymetry is not itself indicative of a response to orbital or 100-ky variations in climate (24).We seek to distinguish between abyssal-hill length scales that are inherent to lithospheric structural controls and those that reflect temporal periodicity in sea level and magma supply, such as at orbital periods. Our primary approach is to examine differing predictions for the relationship between spreading rate and abyssal-hill spacing at intermediate and faster spreading ridges. An assumption that melt supply accommodates a constant fraction of spreading (3) leads to a conclusion that abyssal-hill spacing becomes constant at half-spreading rates above ∼3 cm/y (e.g., ref. 24, their figure 3a and supplemental material). Conversely, if abyssal-hill spacing is controlled by quasiperiodic variability in melt, characteristic wavelengths should increase with spreading rate in proportion to the period of melt variability (23). To explore these distinct predictions, we first revisit a faulting model that was used to illustrate how quasiperiodic bathymetry can be independent of sea-level variations (24) to explore conditions under which sea-level–induced changes in melt supply influence simulated bathymetry. We then compare our simulations against analyses of bathymetric variability across 17 different regions (Fig. 1 and Open in a separate windowFig. 1.Global bathymetry with details of the 17 abyssal-hill regions used in this study. Regional maps show ridge flank profiles along which center-beam bathymetry is extracted (black lines) that extend from the ridge axis (circles) to the estimated location of the Brunhes–Matuyama magnetic reversal (crosses). A total of 182 ridge flank profiles adjoin to give 91 complete transects of the ridge, whereas 23 ridge flank profiles are unpaired because the other side is disturbed, the Brunhes–Matuyama reversal is not identifiable, or bathymetry data are missing. Mapped bathymetry is from the Marine Geoscience Data System. White shading indicates that data are unavailable. Regional maps show a 10-km horizontal scale (horizontal black bar).Table 1.Data sources for each region from which bathymetry is analyzed in this study