Investigating the effects of bone cement, cyanoacrylate glue and marine mussel adhesive protein from Mytilus edulis on human osteoblasts and fibroblasts in vitro

Bone cement is a widely used standard fixation substance in Orthopaedic Surgery. Cyanoacrylate glue is available for wound closure to supplement suturing. The mussel adhesive protein extracted from Mytilus edulis (Cell-Tak©, BD Biosciences, Heidelberg, Germany) is an experimental fixation device used for in vitro purposes of cell adhesion.

The aim of this study is to introduce a cell culture model investigating the effects of commonly applied and experimental glues on human fibroblasts and osteoblasts in vitro. Cells cultured without additives served as a control group. Microscopic examination was performed to evaluate the morphologic changes. An apoptosis test (Apo-Tag©, Chemicon International, Temecula, CA, U.S.A.) was applied to determine the rate of natural cell death at the end of the study.

It could be demonstrated that morphological changes in bone cement are different in fibroblasts and osteoblasts. Osteoblasts seem to grow on bone cement and develop an orderly formation. Fibroblasts grow in a confluent monolayer around bone cement but do not adhere to the cement itself. This is a desirable effect since most Orthopaedic applications aim at osteointegration as opposed to fibrous tissue overgrowth. Apoptosis attributed to bone cement is comparable to the respective natural rate of apoptosis. Cyanoacrylate glue and the mussel adhesive protein lead to an almost complete apoptosis in the investigated cells. Their routine application should be avoided. The developed cell culture model seems appropriate for performing further investigations.     

贻贝多糖的工业化制备工艺研究

目的 建立适于工业放大的贻贝多糖提取工艺。方法 以厚壳贻贝为对象,对贻贝多糖制备过程中原料筛选、浸提条件、除蛋白质、醇沉等技术环节进行改进和优化。最后利用改进的工艺进行了贻贝多糖的中试制备。结果 厚壳贻贝多糖为白色粉末,多糖平均得率为贻贝干重的14.1%。MP-A为厚壳贻贝多糖的主要成分,占比约为89.42%,相对分子质量为1,268.15 kD。结论 建立了流程简单、绿色环保、易于放大的贻贝多糖工业化制备工艺,能获得纯度90%左右的贻贝多糖样品。     

东海染毒紫贻贝中贝类毒素的分离和鉴定

目的掌握紫贻贝的染毒情况,对其含有的贝类毒素成分进行分离和鉴定。方法样品经丙酮提取,提取物经乙醚-水液分配,乙醚可溶部分再经硅胶柱层析粗分离、反相高效液相色谱半制备细分离,UPLC-MS/MS监测。结果分离得到4个毒素组分,根据质谱数据以及与pectenotoxin-2(PTX-2)的酶水解产物比较,鉴定组分Ⅰ和Ⅱ分别为pectenotoxin-2 seco acid(PTX-2sa)和7-epi-PTX-2 seco acid(7-epi-PTX-2sa),经LC-MS/MS分析与标准品对照鉴定组分Ⅲ和IV分别为大田软海绵酸(OA)和鳍藻毒素-1(DTX-1)。结论从苍南县市售贻贝中同时分离出腹泻性贝类毒素(DSP)和扇贝毒素类(PTXs),其中游离态的OA、DTX-1及总OA含量超过欧盟限量值,为这次腹泻性贝类中毒的主要原因。     

Influence of rearing water temperature on induced gonadal development and spawning behaviour of tropical green mussel,Perna viridis

ObjectiveTo standardize the technique of induced breeding and spawning of green mussel Perna viridis (P. viridis), in captivity.MethodsIn Experiment-A, the temperature was increased at a rate of 2 °C/5 days interval. In Experiment-B, a rise of 3 °C/5 days was practiced, whereas in Experiment C and D, respectively 4 and 5 °C was increased in 5 days interval. The temperature was maintained constant at 20 °C in the Control.ResultsThe increase in temperature showed a progressive effect on the gonadal development of mussels. The gonads ripped at 30 to 32 °C in all the experimental tanks, irrespective of the difference in temperature hike. Complete spawning in P. viridis was achieved by gradually raising the temperature from 20 to 35 °C at a rate of 3 or 4 °C/5 days.ConclusionAccording to the present study temperature induced spawning method is very simple and cost effective and can accelerate the production of mussel seeds in hatchery units and further stock improvement through genetic manipulation.     

Profiles and levels of fatty acid esters of okadaic acid group toxins and pectenotoxins during toxin depuration, Part I: Brown crab (Cancer pagurus)

In 2002, two outbreaks of diarrhetic shellfish poisoning (DSP) occurred in Norway, which was later confirmed to be caused by the consumption of brown crab (Cancer pagurus) contaminated predominantly by esters of okadaic acid (OA) after feeding on toxic blue mussels (Mytilus edulis). In addition to OA-group toxins, pectenotoxins (PTXs) are commonly detected in the toxin-producing algae (i.e. Dinophysis). In this paper, an experiment was set up to study the fatty acid ester profiles and depuration rates of OA-group toxins and PTXs from C. pagurus after feeding on M. edulis containing these toxin groups. OA, DTX1, DTX2 and PTX2 SA were all detected primarily in the form of fatty acid esters in the crab hepatopancreas (HP). Crabs preferentially assimilated toxins of the OA group after feeding on the mussels for 1 week. Detailed analysis of the fatty acid ester profile in crabs and mussels showed that the ester profiles in the crabs differed slightly from profiles of the fatty acid esters in M. edulis, but neither ester profile nor ester to free toxin ratio appeared to change in the crabs during the first 2 weeks of depuration. Calculations of depuration rates of the free forms of toxins resulted in similar reduction rates for OA and DTX2, whereas the depuration rate of DTX1, PTX2 and PTX2 SA was considerably faster. From the industrial perspective, the PTX-compounds are of minor importance compared to the OA group toxins in crabs, considering (1) the uncertainty regarding the oral toxicity of the PTXs, (2) the preferential ingestion of OA-group toxins compared to PTXs and (3) the faster depuration of PTXs.     

Seasonality of diarrhetic shellfish poisoning at a coastal lagoon in Portugal: rainfall patterns and folk wisdom.

Of the three types of toxicity known so far in Portuguese shellfish, only diarrhetic shellfish poisoning (DSP) and amnesic shellfish poisoning (ASP) are produced by microalgae that seem to have been present in the last decades or centuries. The most important paralytic shellfish poisoning (PSP) producer, Gymnodinium catenatum, is hypothesised to have been introduced quite recently as only in 1976 PSP toxicity was detected for the first time in shellfish from Galicia, NW Iberian Peninsula. While ASP presents very short episodes of contamination, the concentration of DSP toxins in some years surpasses human safety values for much longer periods. It is traditionally stated that shellfish should be consumed in 'months with R' (September-April). A retrospective study of the maximum monthly DSP levels attained in mussels from a coastal lagoon-Ria de Aveiro-between 1994 and 2001, showed that the highest frequency of months with concentrations surpassing the safety level of 2 microg/g digestive glands were found in June-September, followed by May and October. These months correspond with the months of lowest historical average rainfall in the period 1941-1998.Oscillations in the rainfall pattern coincided with earliest (or latest) detection by HPLC of DSP toxins in mussel in the years studied. In a semi-closed lagunar environment prone to in situ growth of DSP-producer microalgae, like Dinophysis acuminata, rainfall affects river output, lowering salinity and disrupting water column stability that favours Dinophysis growth. The seasonality of DSP recurrence may be connected to the folk adage on safety of shellfish consumption, after many years of empirical observations by coastal populations of diarrhoea episodes in summertime.     

Biomonitoring of heavy metals using Mytilus galloprovincialis in Safi coastal waters, Morocco

Heavy metal concentrations of mercury, cadmium, lead, zinc, cooper, nickel, manganese, and chromium in Mytilus galloprovincialis were investigated to provide information on pollution of Safi coastal area, since these metals have the highest toxic potential. The concentration of Hg and Pb was determined by AFS and ICP-MS methods, respectively, whilst the remaining metals (Cd, Cr, Cu, Mn, Zn, and Ni) were quantified by AAS. High lead, cadmium, chromium, and mercury levels were registered in tissue samples collected from two stations near the Jorf Lihoudi and Safi city, while elevated concentration of manganese and zinc (14.70-25.30 mg kg(-1) and 570-650 mg kg(-1) dry wt, respectively) were found in mussel specimens from Cap Cantin. The high levels of nickel found respectively in the areas near the industrial area being of concern in terms of environmental health need frequent monitoring. The metal concentrations recorded at the clean stations may be considered as useful background levels to which to refer for comparison within the Atlantic coast. M. galloprovincialis are suitable biomonitors to investigate the contamination levels of heavy metals pollution face a different human activity in this coastal area of the Atlantic coast.     

A Biomonitoring Study Assessing the Residual Biological Effects of Pollution Caused by the HAVEN Wreck on Marine Organisms in the Ligurian Sea (Italy)

Residual biological effects of the 1991 HAVEN oil spill off the Ligurian (Arenzano) coast were assessed in this study. Samples of the fish species Boops boops, Mullus barbatus, and Uranoscupus scaber were collected from two polluted sites near the HAVEN wreck and from an uncontaminated area. In addition to this, mussels were caged along the coast affected by the HAVEN disaster. The physiological status of fish and mussels was assessed using a battery of stress and exposure biomarkers. The PAH content of mussel and fish tissues was also analyzed. Significant biological responses were observed in lysosomal membrane stability, neutral lipid and lipofuscin accumulation and micronucleus frequency for mussels caged at two sites close to the HAVEN wreck. Chemical analyses indicated, however, that these effects are not caused by aromatic hydrocarbons. For this reason, we suggest that the aftermath of the HAVEN disaster contributes very little to coastal ecosystem pollution. This was also confirmed by the few biological effects observed in fish specimens (Boops boops) collected from surface waters. Nevertheless, it is important to point out that benthic fish displayed a stress syndrome potentially caused by aromatic hydrocarbons released from the oil tanker, as witnessed by an enhanced EROD activity and increased lipofuscin and neutral lipid lysosomal contents.     

双组分快速固化生物软组织黏合剂的研制

贻贝足丝蛋白(MFp)是一类能够在水下迅速固化,并能黏附在不同基材表面的特殊蛋白质,作为蛋白类生物黏合剂具有广泛的应用前景。本文在大肠杆菌中高效可溶性地表达了贻贝足丝融合蛋白Sumo-Fp3(SFp3),并通过蘑菇酪氨酸酶在柱催化,将其中约5%的酪氨酸残基转化为DOPA。在含DOPA的SFp3(DSFp3)中加入分子量为1 500 kD的透明质酸后,形成的双组分生物胶水在牛皮表面的黏附力超过氰基丙烯酸盐组织粘合剂Dermabond?的两倍,并在5 min内达到最大黏附强度的52%。通过生物膜层干涉技术和扫描电子显微镜,观测到透明质酸与DSFp3存在静电层层组装行为,并形成紧密片层结构。本研究为蛋白质类生物胶水黏附强度低、固化慢提供了一种解决方法和理论基础。