Studies on cryopreservation of luc gene transfected bluegill sunfish fibroblast cell line

The effect of cryopreservation on the survival of luc gene transfected bluegill sunfish fibroblast (BF-2) cells was investigated. Propane-1,2-diol was found to be the least toxic cryoprotectant when compared with DMSO and methanol. Both propane-1,2-diol and DMSO are effective in protecting cells from freezing damage. Whilst there were no significant differences in cell survival between cryoprotectant concentration (10 or 15%) and culture age used in this study, 7-day old culture appeared to be more resistant to freezing without cryoprotectant when compared with 3- and 14-day old culture. The highest cell survival values obtained with propane-1,2-diol (10%) and DMSO (10%) protection were 96.2 1.2% and 94.0 3.1% respectively. Initial subsequent cell growth after cryopreservation was slower than their non-frozen controls. The survival of transfected BF-2 cells (BF-2/luc1) after cryopreservation were very similar to those obtained with wild type cells being: 94.0 3.1% and 95.2 1.5% respectively with 10% DMSO protection. These results suggested that genetically modified fish cell lines may be equally amenable to cryopreservation as the wild type.     

Analysis of ploidy and the patterns of amplified fragment length polymorphism and methylation sensitive amplified polymorphism in strawberry plants recovered from cryopreservation

Shoot-tips of 10 strawberry genotypes were successfully cryopreserved using a modified encapsulation-dehydration method. All genotypes survived cryopreservation with high survival and regeneration rates. Eight Joho single-bud sibling lines were established as a model system for genetic analysis. Although cytological examination found chromosomal variation in both non-cryopreserved and cryopreserved samples, the ploidy constitution remained relatively stable after cryopreservation. DNA samples digested with MseI and PstI were used for amplified fragmentation length polymorphism (AFLP) assay. In 16 primer combinations, only one, namely, PCCA-MCAG, detected one site where band pattern changed after cryopreservation, which might be contributed to the change in DNA methylation status at PstI recognition site. Methylation sensitive amplified polymorphism (MSAP) assay was carried out for further investigation on the influence of cryopreservation on DNA methylation status. It was found that cryopreservation induced a significant change in DNA methylation status.     

动脉血管低温保存研究现状及展望

血管低温保存是满足临床移植的重要保证手段。从影响动脉血管低温保存的因素,低温保存后的活性恢复、细胞水平上的血管低温损伤机理、冻结/复温过程中的热物理性质和力学性质变化,以及低温断裂现象等方面综述了血管低温保存研究进展。     

Effects of cryopreservation on developmental competency, cytological and molecular stability of citrus callus

Cell suspensions of twelve citrus genotypes were successfully cryopreserved by vitrification. All genotypes survived cryopreservation with >90% viability and the surviving cells of some genotypes regenerated somatic embryos better than the controls. Single-cell sibling lines of cultivar Newhall were used for cytological and molecular examination. It was found that the ploidy constitution remained genetically stable and that no DNA sequence variation was detected by randomly amplified polymorphic DNA (RAPD) assay after cryopreservation. In addition, the methylation sensitive amplified polymorphism (MSAP) assay indicated that cryopreservation caused a significant change in DNA methylation status.     

Effects of aspiration on public cooperation in structured populations

We introduce a deterministic win-stay-lose-shift rule into the spatial public goods game, according to which a player will change its current strategy only if its payoff is below a predefined aspiration level. Simulation results on the square lattice and scale-free network indicate that the aspiration level greatly affects the evolution of cooperation. For small multiplication factors, the frequency of cooperation increases to 0.5 as the aspiration level increases. For large multiplication factors, intermediate levels of aspiration prove optimal for the successful evolution of public cooperation. Some qualitative analyses are provided to explain the above results. Besides, we have found that there exists a ping-pong vibration of cooperation at some specific values of multiplication factors and aspiration levels.     

Application of phosphoenolpyruvate into canine red blood cell cryopreservation with hydroxyethyl starch

Phosphoenolpyruvate (PEP) is a phosphorylated glycolytic intermediate that can penetrate the RBC membrane and be metabolized to 2,3-DPG and ATP. In this study, we evaluated the effects of PEP treatment on canine red blood cells (RBCs) cryopreserved with 12.5% (w/v) HES. RBCs were incubated for 30, 60, and 90 min at 37 degrees C with PEP solution containing 60 mM mannitol, 30 mM sodium chloride, 25 mM glucose, 1 mM adenine and 50 mM PEP (340 m osm/kg), pH 6.0 and then cryopreserved in liquid nitrogen with 12.5% (w/v) HES for 2 weeks. 2,3-DPG and saline stabilities of the PEP treated groups were increased and osmotic fragility indices were significantly decreased compared to the untreated control group. There were no differences in 2,3-DPG levels within the PEP treated groups with different PEP incubation times. These results suggest that PEP treatment may be beneficial for the cryopreservation of canine RBCs with HES.     

医用生物材料的低温保存研究

生物材料的低温保存一般都要经历降温过程、低温储存过程及复温过程,其中降温过程中对生物细胞的影响最大.每一种生物细胞都有自己合适的降温速率,如能满足其这种降温速率,细胞所受到的低温损伤最小,则生物细胞的复活率最高.文中介绍程序控制变速降温装置的主要结构及几种典型生物体的降温过程.最后,对器官的低温保存进行分析讨论.     

莴苣种子的生长与低温保存研究

文中主要介绍高原莴苣种子生长和低温保存时的相关性,为保护保存该种子提供理论和实验依据。种子在低温和适宜温度两种不同情况下培养,进行实验比较,得出低温预培养的种子对于低温保存无作用;快速降温下,DSC曲线出现两个波峰HTE和LTE,当温度低于LTE时,无种子发芽存活;慢速降温下,DSC曲线只有一个波峰HTE;随着温度降低,种子发芽率降低,当温度低于-40℃时,无种子存活。     

Nonequilibrium statistical mechanics of dividing cell populations

We present and study a model for the nonequilibrium statistical mechanics of protein distributions in a proliferating cell population. Our model describes how the total protein variation is shaped by two processes: variation in protein production internal to the cells and variation in division and inheritance at the population level. It enables us to assess the contribution of each of these components separately. We find that, even if production is deterministic, cell division can generate a large variation in protein distribution. In this limit we solve exactly a special case and draw an analogy between protein distribution along cell generations and stress distribution in layers of granular material. At the other limit of extremely noisy protein production, we find that the population structure restrains variation and that the details of division do not affect the tail of the distribution.     

Development of a shoot-tip vitrification protocol and comparison with encapsulation-based procedures for plum (Prunus domestica L.) cryopreservation

Cryopreservation of plum (Prunus domestica L.), cv Regina Claudia, was obtained by a vitrification/one-step cooling procedure of shoot tips from cold-hardened in vitro-grown plants. Best survival (57%) was obtained when the shoot tips were precultured at 4 degree C for 2 days on 0.09 M sucrose-containing Quoirin and Le Poivre medium, loaded for 30 min with a cryoprotectant (2 M glycerol and 0.4 M sucrose), incubated with the PVS2 solution at 0 C for 90 min, and directly plunged into liquid nitrogen. After re-warming in a waterbath at 40 degree C, the shoot tips were washed in a 1.2 M-sucrose MS solution for 20 min and finally plated on a regrowth medium. In comparison with the one-step cooling procedure, both the slow cooling (-0.5 degree C/min up to -45 degree C), and the two-step cooling (-160 degree C for 25 min, then -196 degree C) gave lower percentages of shoot-tip survival. Among the other cryogenic procedures tested, the performance of the encapsulation-vitrification method was similar to the vitrification protocol in terms of shoot-tip regrowth (47.5%), while encapsulation-dehydration was unsatisfactory.     

Effect of cryopreservation on the structural and functional integrity of cell membranes of sugarcane (Saccharum sp.) embryogenic calluses

In this paper, we investigated if the differences consistently noted in survival and plantlet production between cryopreserved and non-cryopreserved, control sugarcane embryogenic calluses were related to modifications induced during cryopreservation in the structural and functional integrity of cell membranes. For this, the evolution of electrolyte leakage, lipid peroxidation products and cell membrane protein contents was measured during 5 d after cryopreservation. Differences between control and frozen calluses were observed only during the first 2 (electrolyte leakage) or 3 d (lipid peroxidation products and membrane protein content) after freezing. It was not possible to link these differences with the differences noted in survival and plant production between control and cryopreserved calluses. Additional studies are thus needed to elucidate which biochemical factors, linked to survival and plantlet regeneration, are affected by cryopreservation.     

Effect of cryopreservation protocols on the phenotypic stability of yeast

Eight cryopreservation protocols were assessed for their effects on the viability and phenotypic stability of the yeast Saccharomyces cerevisiae during a five-year study. It is found that viability and phenotypic features have remained largely unchanged when the yeast was preserved in glycerol, dimethyl sulphoxide, or sucrose at -80 degrees C or in liquid nitrogen. When sorbitol was used as a cryoprotectant, yeast cells frozen and stored at -80 degrees C manifested great decreases in viability after six months in storage and concomitantly large fluctuations in the rate of the trpl auxotrophic reversion. This phenotypic reversion was stable passage after passage. Such a degree of phenotypic fluctuations, however, was not observed for yeast cells preserved in the same sorbitol solution that went through a controlled freezing program and were subsequently stored in liquid nitrogen. These results indicate that some combinations of cryoprotective agent, freezing program, and storage temperature disturb biomaterials more profoundly during cryopreservation and imply a genetic basis of this phenotypic change.     

Effects of cryopreservation with polyethylene glycol on the expression of CD11b and CD62L on the surface of polymorphonuclear leukocytes

In experimental and clinical studies, expression of surface adhesion molecules such as ?2-integrine (CD11b) and L-selectin (CD62L) on polymorphonuclear leukocyte (PMNL) are investigated to assess certain crucial innate immune functions. Because the expression of CD11b and CD62L on PMNL can alter they cannot be quantified reliably when the time between blood draw and measurements is prolonged. Goals of this study were to test effects of cryopreservation on the expression of CD11b and CD62L on human PMNLs either under native conditions as well as after stimulation-dependant adhesion molecules′ expression pattern. CD11b and CD62L expression on PMNL can be cryopreserved with 10% of PEG-solution for at least one month at -60 degree C. This was observed in native, unstimulated as well as in stimulated cell-preparations. CD11b is very stable in contrast to CD62L expression which appears to be more susceptible to alteration due to freezing-thawing. However, the relative stimulus-dependant changes of activation can still be reflected.     

生物材料玻璃化保存方法研究进展

生物材料的玻璃化保存技术一直是低温生物学领域的研究热点,提高冷却速率是实现玻璃化保存的一条途径。介绍了常用玻璃化保存方法的操作方法,分析比较了其冷却速率和保存效果,并提出了一种新颖的玻璃化保存法。     

Effects of traffic noise on paserine populations in Mediterranean wooded pastures

The effect of traffic noise on the breeding density of 20 passerine species was examined over a 2-year period in three different road types passing through pasture-woodlands in western-central Spain. No statistically significant differences were observed during the 2 years studied. An average of 19.6 birds/10 ha was recorded for the low-traffic road (LT), 21.7 birds/10 ha for the medium-traffic road (MT) and 19.1 birds/10 ha for the high-traffic road (HT). A total of 11 species (55%) did not show any statistically significant differences in breeding density among the different types of roads. By contrast, other species, such as the Blackbird, the Iberian Shrike and the Linnet, did point to differences between the MT and HT roads. House and Rock Sparrows, as well as the Corn Bunting, showed higher breeding densities near the HT road. The opposite effect was observed for the Wheatear, the Iberian chiffchaff and the Woodlark, for which high breeding densities were recorded in the vicinity of the LT road. Our results suggest that traffic noise constitutes a serious problem for at least 15% of the breeding bird community.     

Effects of migration on the evolutionary game dynamics in finite populations with community structures

We investigate the impacts of migration on the evolutionary game dynamics in finite populations with community structures in the framework of evolutionary game theory. In contrast to deterministic dynamics, our model incorporates stochastic factors induced by the finite population size. Based on the analysis of the stationary distribution of the evolutionary process in the limit of rare mutations, we prove that it is most likely to find the population in the community where all individuals have the lower migration rate. Furthermore, we show that reducing the difference between the migration rates of distinct communities can increase the first hitting time to the homogeneous absorbing state and can prolong the coexistence time of different species, promoting the conservation of biodiversity.     

Classifying the expansion kinetics and critical surface dynamics of growing cell populations

We systematically study the growth kinetics and the critical surface dynamics of cell monolayers by a class of computationally efficient cellular automaton models avoiding lattice artifacts. Our numerically derived front velocity relationship indicates the limitations of the Fisher-Kolmogorov-Petrovskii-Piskounov equation for tumor growth simulations. The critical surface dynamics corresponds to the Kardar-Parisi-Zhang universality class, which disagrees with the interpretation by Bru et al. of their experimental observations as generic molecular-beam-epitaxy-like growth, questioning their conjecture that a successful therapy should lead away from molecular beam epitaxy.     

量化对干涉成像仪复原光谱幅值不确定度的影响

推导并分析了干涉图量化对干涉成像光谱仪复原光谱幅值的影响。结果表明,复原光谱幅值的不确定度受量化条件的影响。在确定了干涉图采样点数后,干涉成像光谱仪复原光谱幅值的不确定度与A/D转换器的量化步长成正比;在固定了量化步长后,不确定度随干涉图采样点数的增加而增大;由干涉图量化引起的复原光谱幅值不确定度的大小还与在实际算法中选取的切趾函数形式有关。     

Influence of cryopreservation on the cytosine methylation state of potato genomic NDA

Shoot tips of Solanum tuberosum (Désirée) were successfully cryopreserved by the DMSO droplet method and stored for almost 7 years, while control material was maintained in vitro for the same period of time. To analyse potential epigenetic changes, the DNA methylation status was assayed by methylation-sensitive amplified polymorphism (MSAP) analysis using restriction endonucleases MspI and HpaII. An amount of 93.6% of the analysed MSAP signals were stable among all cryopreserved and in vitro maintained samples tested, indicating extensive stability of DNA methylation. Only 0.9 % of MSAP signals showed results that differed between the two treatments and at the same time matched for all three biological replications within each treatment. These can be seen as indicating directed effects of the two treatments on the DNA methylation. Cryopreserved samples displayed in comparison to in vitro stored samples consistent hypomethylation for 0.6 % (3 of 469) of MSAP signals (Table 4, pattern 4) and consistent hypermethylation for 0.2% (1 of 469), respectively. For 5.6% of all MSAP signals, inconsistent results were observed among the three biological replications at least for one of the two treatments. These were interpreted as resulting from stochastic DNA methylation changes in individual samples. As results for two biological replications were identical and different from the result for the third biological replication, the direction of methylation change could be determined in those cases. Cases of stochastic loss of CG methylation in cryopreserved samples were most frequent among them, adding up to 3.4% of MSAP signals. Stochastic loss of CG methylation was also found in material maintained in vitro, only for 0.6% of all MSAP signals. In conclusion, methylation changes occurred in long-term cryopreservation of potato, in a random rather than directed fashion. Hence, cryopreservation and long-term in vitro maintenance both induce limited changes of DNA methylation status. The order of magnitude of methylation changes observed was consistent with other studies, where similar rates of DNA methylation changes have been found.     

Mössbauer spectroscopic study of meteorites recovered on Antarctica

The chemical states of iron in sixteen Antarctic meteorites belonging to H-group chondrites were studied by means of Mössbauer spectroscopy. An Fe-Ni alloy, troilite, paramagnetic Fe(III), and two kinds of paramagnetic Fe(II) were observed in each meteorite. The Mössbauer parameters indicated that the Fe(II) components can be assigned to olivine and some pyroxenes. The relative area intensities of Fe(III) in the chondrites correlated positively with iodine content, which was determined by radiochemical neutron activation analysis, and those of two Fe(II)-species correlated negatively with the content. On the basis of the data on the halogen and the Mössbauer spectrocopy, the terrestrial contamination on Antarctic meteorites is discussed.