Dextran synthesis by immobilized dextran sucrase

Dextran sucrase has been produced by fermentation of Leuconostoc mesenteroides NRRL B-512, with and without continuous sucrose addition to improve enzyme production. The enzyme preparation has been concentrated from the fermentation broth by ultrafiltration and purified by gel permeation chromatography on Ultrogel. The specific activity of the dextran sucrase was greatly enhanced by calcium chloride addition to the purified enzyme. This enzyme preparation has been immobilized by covalent coupling onto an amino porous silica support (Spherosil) activated with glutaraldehyde. Immobilized dextran sucrase derivatives with an activity up to 830 dextran sucrase units per g. support could thus be obtained. The effect of the support specific area on coupling efficiency and reaction kinetics has been investigated, and the effect of intraparticular diffusion underlined. The molecular weight distribution of the dextran has been determined when varying several parameters.     

Endocardial endothelium in the rat: junctional organization and permeability

Selective permeability of endocardial endothelium has been suggested as a mechanism underlying the modulation of the performance of subjacent myocardium. In this study, we characterized the organization and permeability of junctional complexes in ventricular endocardial endothelium in rat heart. The length of intercellular clefts viewed en face per unit endothelial cell surface area was lower, and intercellular clefts were deeper in endocardial endothelium than in myocardial vascular endothelium, whereas tight junctions had a similar structure in both endothelia. On this basis, endocardia endothelium. might be less permeable than capillary endothelium. However, confocal scanning laser microscopy showed that intravenously injected dextran 10000 coupled to Lucifer Yellow penetrated first the endocardial endothelium and later the myocardial capillary endothelium. Penetration of dextran 10000 in myocardium occurred earlier through subepicardial capillary endothelium than through subendocardial capillary endothelium. Penetration of tracer might thus be influenced by hydrostatic pressure. Dextran of MW 40000 did not diffuse through either endocardial endothelium or capilary endothelium. The ultrastructure of endocardial endothelium may constitute an adaptation to limit diffusion driven by high hydrostatic pressure in the heart. Differences in paracellular diffusion of dextran 10000 between endocardial endothelium and myocardial vessels, may result from differing permeability properties of the endocardium and underlying myocardium.     

A study of dextran production from maltodextrin by cell suspensions of Gluconobacter oxydans NCIB 4943

This study investigated dextran synthesis from a commercial maltodextrin substrate using cell suspensions of G. oxydans NCIB 4943 as catalysts. Experiments were arranged according to a central composite statistical design. The effects of substrate concentration (10-100 g l-1), cell concentration (0.32-32.0 g wet weight l-1), time of reaction (8-48 h) and pH (3.5-5.5), each at three levels, on dextran yield and dextran molecular weight (MW), were investigated. Response surface methodology was used to assess factor interactions, and empirical models describing the two responses were fitted. Most of the variance in dextran yield could be explained by the fitted model (R2 = 0.96). Dextran yield ranged from 1.21 to 41.69%. The presence of significant negative quadratic effects of cell concentration and time indicated that dextran yield reached a plateau and thus, optimum levels of cell concentration and time could be identified to maximize dextran yield. Dextran MW ranged from 6.6 to 38 kDa and was characterized by the significant interactions of reaction time with substrate concentration and cell concentration. The model, however, could account for only 60% of the variance in dextran MW. Possible reasons for this are discussed.     

Evaluation of crude dextran as phase-forming polymer for the extraction of enzymes in aqueous two-phase systems in large scale

A detailed study of the influence of crude dextran on enzyme extractions in aqueous phase systems is presented in this article. The physical parameters of crude dextran, a purified T-500 fraction from Pharmacia, and a hydrolyzed crude dextran are compared and their influence on the phase system parameters investigated. Initially there is a drastic increase in the viscosity of the lower dextran-rich phase and a significant shift in the macroscopic structure of these phases, observed as the "gel-forming" properties of the dextran phases. The latter can be important for the partition of any enzyme by influencing the effect of phosphate concentration on the partition of proteins, although these experiments show that the partition coefficient of several enzymes is not much altered. The partition parameters allow the substitution of Dextran T-500 fractions by crude dextran or unfractionated, slightly hydrolyzed fractions. Using crude dextrans the performance and technical realization of enzyme extraction processes are demonstrated for pullulanase from Klebsiella pneumoniae and formate dehydrogenase from Candida boidinii.Both enzymes were recovered in comparable high yields. The equipment performance was quite good, as indicated by the high throughput values of the separators employed. Especially when using nozzle separators for phase separation there is a better performance in comparison to the Dextran T-500 fraction. No serious technical problems were encountered when replacing the expensive fractionated dextran with a crude dextran. In this way aqueous two-phase systems containing dextran become more feasible for enzyme purification from an economic point of view. The price of about 1.30 German marks (DM) per liter for a useful phase system already appears acceptable for the production of valuable intracellular enzymes.     

聚乙二醇、葡聚糖对杂交瘤细胞保护特性研究

利用鼠鼠杂交瘤２Ｆ７细胞（分泌抗小细胞肺癌单克隆抗体）研究聚乙二醇、葡聚糖与杂交瘤细胞的生物相容性及添加限制浓度,研究添加浓度对葡萄糖消耗速率及氨形成速率的影响。在高速搅拌、高剪切力下考察了添加剂的保护性能。实验结果表明,０．０４％－０．１０％的聚乙二醇能较好地保护杂交瘤细胞,添加聚乙二醇后葡萄糖的比消耗速率增加,而氨的比生成速率没有变化;添加葡聚糖对葡萄糖的比消耗速率没有影响,但降低了氨的比生成速率,并且葡聚糖抑制细胞生长,不适合作动物细胞保护剂;小牛血清能较好地保护细胞;细胞死亡动力学表征为一级反应。在２５０ｍｌ磁力搅拌瓶中培养,培养基中添加０．１０％聚乙二醇,培养温度为３７．０,搅拌转速为８０转／分,细胞能止常生长;而培养基不添加聚乙二醇时细胞不能生长。     

Decrease of saturation density of cells of hamster cell lines after treatment with dextran sulfate

Dextran sulfates of various molecular weights were added to cultures of 3 transformed cell lines of hamster, 3T6 cells and embryonic fibroblastic cells. Dextran sulfate of high molecular weight reduced the saturation densities of all the cell lines of hamster and 3T6 cells, but those of low molecular weight did not. The mitotic rate of the treated cells decreased at stationary cell density. Dextran sulfate had no effect on the growth of normal fibroblastic cells derived from mouse and hamster embryos. Viability of treated cells was indicated by the following results. Cells of cultures seeded at different cell densities grew at almost the same rate in the presence of dextran sulfate. Treated cells remaining in the monolayer stage began to grow after removal of dextran sulfate. The colony formation rate of treated cells was the same as that of untreated cells. With the exception of one cell line, the morphology of cells treated with dextran sulfate of high molecular weight was more flattened and there was less overlapping than in untreated cells. Treated cells were less agglutinable to concanavalin A than were untreated cells. These results suggest that dextran sulfate affects the cell surface, resulting in the decrease of saturation density of cell lines.     

Corneal tolerance of vitrifiable concentrations of propane-1,2-diol

The merit of corneal cryopreservation by vitrification as opposed to conventional freezing is the avoidance of ice damage which is believed to disrupt the integrity of the corneal endothelium resulting in loss of corneal transparency. The cornea must be equilibrated with high concentrations of cryoprotectant in order to achieve vitrification at practicable cooling rates. In an earlier study, corneas were exposed to 3.4 mol/liter propane-1,2-diol (Rich and Armitage (1990) Cryobiology 27, 42-54). The present study exposed rabbit corneas to concentrations of propane-1,2-diol between 3.4 and 5.4 mol/liter in a Hepes-buffered Ringer's solution containing glutathione, adenosine, 5 mmol/liter sodium bicarbonate, 6% (w/v) bovine serum albumin, and 2.5% (w/v) dextran sulfate. Dextran sulfate was as effective as chondroitin sulfate at improving endothelial tolerance of 3.4 mol/liter propane-1,2-diol. This beneficial effect may be linked to the polyanionic nature of these molecules. Corneas exposed to 5.4 mol/liter propane-1,2-diol were cooled in liquid nitrogen vapor at a temperature of -140 degrees C for 2 h. Warming was achieved by direct transfer to a dilution solution at -10 degrees C. Endothelial function was assessed by monitoring corneal thickness during perfusion of the endothelial surface at 34 degrees C for 6 h. Endothelial structure was observed by specular microscopy during the perfusion and by scanning electron microscopy after perfusion. Corneas tolerated exposure to 3.4 mol/liter propane-1,2-diol for 20 min at 0 degrees C and to 4.1 mol/liter for 10 min at -10 degrees C. Exposure to 4.8 and 5.4 mol/liter for 10 min at -10 degrees C caused endothelial damage, although a degree of endothelial function was retained. Function following exposure to 5.4 mol/liter was improved by reducing the temperature of exposure to -15 degrees C. Corneas cooled after exposure to 5.4 mol/liter propane-1,2-diol for 10 min at -15 degrees C apparently vitrified, but devitrified on warming. The corneas swelled to such an extent during perfusion that the endothelium could not be viewed by specular microscopy, subsequent scanning electron microscopy showed a severely disrupted endothelium.     

Selective effects of three herbicides on the fungus flora of egyptian soil

2,4-D (2,4-dichlorophenoxyacetio acid) when applied to the soil at three doses (1.9, 7.6 and 15.2 mg per kg dry soil) had a stimulating effect on the total count of soil fungi and on several fungal species especially between 5 and 20 d after treatment. When the herbicide was incorporated in the agar medium it had a stimulating effect on the counts of total fungi, Aspergillus sp., A. niger, A.fumigatus and Fusarium app. at the low dose (6.3 ppm), but wag toxic at this dose toward Humicola grisea and Myrothecium verrucaria at the medium and high doses (25.2 and 50.4 ppm), it was toxic to the total count of fungi and to the majority of fungal species. VCS-438 [2-(3,4-dichlorophenyl)-4-methyl-l,2,4-oxadiazolidine-3,5-dione] was beneficial to the total count of fungi 2 and 5 d after soil treatment with the medium dose (8.0 mg per kg dry soil). Some fungal species could benefit from the low and the high doses (2.0 and 16.0 mg per kg dry soil) after these experimental periods. In the agar medium the counts of total fungi, Aspergillus sp., A. niger and A.fumigatus were almost significantly reduced by the three doses (6.8, 27.2 and 54.8 ppm). Planavin (4-methylsulphonyl-2,6-dinitro-N,N-dipropylaniline) was stimulating for the total count of fungi, Aspergillus, A.niger and A.ochraceus 2 and 5 d after treatment with the medium dose (8.0 mg per kg dry soil), and was also stimulating to Fusarium population at the medium dose after 2 d and at the high dose (16 mg per kg dry soil) after 20 d. In the agar medium Planavin at the low dose (6.8 ppm) was stimulating to A.terreus and inhibitory to A.nidulans and A.fumigatus. The medium and high doses (27.2 and 54.8 ppm) were generally toxic to the total count of fungi.     

Establishment of a porcine corneal endothelial organ culture model for research purposes

Human corneas usually are not available for research, as they are used for transplantation only. At the same time, scientific studies on cultured human endothelial cells can produce misleading results due to inevitable dedifferentiation. Therefore, an organ-culture model of porcine corneas—displaying endothelial cell death rates comparable to those of cultured human corneas—would be very desirable. Fresh pig eyes were prepared under sterile conditions to obtain corneoscleral buttons, corneal buttons and so called “split corneal buttons” (new preparation method) and cultivated for 15 days. Morphology of the endothelial cell layer was observed by light microscopy on day 1, 8 and 15. On day 15 staining with trypan blue and alizarin red S was performed. Photographs were evaluated in a randomized, blinded manner. Here, the morphology of the corneal endothelium and the number of endothelial cells per mm² were analyzed. After 15 days of cultivation the endothelial cell layer was maintained only in corneal buttons and split corneal buttons. Alizarin red S stained areas and the existence of polymorphisms like rosette figures and reformation figures were significantly less frequent in split corneal buttons than in corneal buttons. Loss of endothelial cells was significantly greater in corneal buttons [575 ± 25/250 cells/mm² (median ± 25%/75%-quantile); 14.8%] than in split corneal buttons [417 ± 138/179 cells/mm² (median ± 25%/75%-quantile); 10.2%]. The new preparation method of split corneal buttons allows the cultivation of porcine corneas for 2 weeks with cell death rates comparable to those of the corresponding human tissue in cornea banks without the need to add de-swelling additives to the media. This is therefore a simple and highly reliable method model to be applied in intervention studies on corneal endothelial cells in their natural compound.     

Clinical cryobiology of tissues: preservation of corneas

It is well recognized that the clarity of the cornea is a function of its hydration, and that this hydration is controlled by a "pump-and-leak" mechanism operating across the posterior monolayer of cells called the endothelium. A breakdown of the endothelium through disease or injury causes a marked increase in corneal thickness as the stroma imbibes fluid from the aqueous humor in the anterior chamber of the eye. This thickened, edematous condition of the stroma results in a cloudy cornea with an associated marked decrease in visual acuity. Treatment for this condition is usually by full-thickness corneal transplantation (penetrating keratoplasty), the success of which is dependent upon the donor cornea having an intact and healthy endothelium. It is essential, therefore, that any method of corneal storage for penetrating keratoplasty should protect and preserve the endothelium in a viable state. Current clinical practice relies upon short-term methods of preservation by two principal methods. Moist Chamber Storage is the time-honored corneal preservation method; it consists of keeping enucleated eyes at 0-4 degrees C in a sealed jar containing a pad of cotton gauze soaked in saline to provide a humid environment. The time limit placed upon this method of storage is 24-48 hr after which the viability of the endothelium deteriorates rapidly. Storage in M-K (McCarey-Kaufman) Medium involves excision of the corneoscleral segment from the donor eye and immersing it, endothelial side uppermost, in a medium consisting of tissue culture medium, 5% Dextran 40, and antibiotics. Laboratory and clinical studies indicate that storage in M-K medium at 4 degrees C preserves human endothelial cells for up to 4 days when the eye has been removed from the cadaver in less than 10 hr postmortem. Long-term preservation of corneas by freezing has long been a major goal in eye banking because indefinite storage by cryopreservation offers significant advantages for the quality and the quantity of material for use in keratoplasty, as well as for its distribution. However, procedures that have been developed for the cryopreservation of corneas have not been widely used, and a number of studies have shown that these procedures are inadequate for maintaining the integrity of the corneal endothelium. Not surprisingly, clinicians are now reluctant to accept corneas that have been frozen by these methods, though the clinical need is now greater than ever.(ABSTRACT TRUNCATED AT 400 WORDS)     

不同缝合方式对大鼠角膜穿通伤后内皮细胞影响的研究

目的:通过手术缝合治疗大鼠角膜穿通伤,探索全层缝合、深板层缝合、及不缝合对角膜内皮细胞的影响.方法:建立大鼠角膜穿通伤模型,同一手术者对角膜切口进行全层、深板层、及不缝合操作.在裂隙灯下动态观察角膜创伤愈合情况;对不同时间点愈合角膜行内皮细胞台盼蓝-茜素红联合活细胞染色及HE染色,观察内皮细胞损伤、修复及白细胞浸润情况.结果:无论是全层缝合还是板层缝合以及不缝合组角膜内皮细胞均损伤明显.但从第1天观察至1月,三组损伤区面积大小无明显差别.结论:全层和深板层缝合及未缝合组可直接造成角膜内皮细胞受损,继发性炎症反应损伤后角膜内皮细胞的损害;伤口周围1.5 mm处角膜内皮几乎损失殆尽,内皮细胞受损可使角膜损伤区水肿迁延不愈,最终形成瘢痕愈合,所以角膜内皮损伤及最终愈合程度三组间无明显差异.     

Priming of the fluid pump by osmotic gradients across rabbit corneal endothelium

The present study shows that the inclusion of 5% Dextran (average mol. wt. 40 000) in solutions to preserve in vitro rabbit corneal endothelium induces a sizable osmotic flow across the preparation which is superimposed on the existing fluid transport. Furthermore, even after fluid transport ceases due to in vitro deterioration, the Dextran-induced flow remains for some addition time. The osmotic permeability was 162 +/- 17 micrometer/s in the presence of glucose and 451 +/- 84 micrometer/s in its absence. The latter, comparatively high value suggests that such osmotic flow traverses the intracellular junctions. In addition, temporary (10--15 min) imposition of an osmotic gradient has a separate stimulatory 'priming' effect on the rate of fluid transport. Thus, the rate of fluid pumping increased by about 40% after challenge with Dextran. It was further noted that, after addition of Dextran, preparations in the absence of glucose escape gross deterioration for a time longer than those in the presence of glucose. On the other hand, mere addition of Dextran to a glucose-containing solution does not appear to prolong the estimated 'survival time' of the pumping mechanism. The sizable osmotic flows and the priming effect described here may provide a physiological context with which previously described Dextran effects on cornea preservation can now be compared.     

洛伐他汀在巴马香猪体内的长期毒性试验

为研究洛伐他汀在巴马香猪体内为期42 d的长期毒性特点,以抗动脉粥样硬化药物洛伐他汀为模型药,选择健康6月龄雄性巴马香猪为实验对象,经灌胃途径给药(12 mg.kg-1和135 mg.kg-1),观察给药后的临床表现、血液学、血生化、脏器系数和组织病理学等指标进行药物毒性的评价。结果主要毒性反应在135 mg.kg-1组,给药后巴马香猪出现了竖毛、腹泻等症状,总采食量和体重减轻;血液血检查显示白细胞总数有所增加,红细胞总数、血红蛋白含量和血小板计数明显下降;血液生化检查显示ALT、AST、ALP和CK升高2~10倍,肌酐和尿素有所升高。同时,组织病理学分析结果显示,巴马香猪出现了与人相似的肝肾病理改变。可见洛伐他汀给药后,巴马香猪在高剂量组可大部分呈现洛伐他汀在人临床出现的毒理反应,表明巴马香猪能敏感的反映洛伐他汀的潜在毒性。     

The Hydrolysis of Dextran by Gram Negative Non-sporing Anaerobic Bacilli

The ability of Gram negative anaerobic bacilli to hydrolyse dextran was determined in liquid and solid media containing Blue Dextran 2000. Released blue chromophore in the liquid medium was detected spectrophotometrically. Results obtained with 334 strains of Bacteroidaceae grown on the solid medium indicated that most strains did not hydrolyse the substrate. Hydrolysis of Blue Dextran 2000 occurred with certain strains of Bacteroides thetaiotaomicron , B. melaninogenicus ss. melaninogenicus, B. oralis, B. ovatus and B. ochraceus.     

The effect of dextran sulphate on CFU-S and nucleic acids in blood and haemopoietic tissues in irradiated mice.

The effect of synthetic polyanion dextran sulphate on the development and recovery of radiation-induced haemopoietic damage in mice was investigated. Dextran sulphate (mol. wt. 500,000 D) in the dose of 40 mg.kg-1 of body weight was injected i.p. 3 days before single total body irradiation with a dose of 7.8 Gy gamma-rays. The animals were examined from hour 6 to day 26 after irradiation, i.e. from hour 78 to day 29 after DS-treatment. In irradiated mice DS-pretreatment showed some positive effect on the CFU-S number in bone marrow (less in spleen and blood), bone marrow cellularity, attenuated the radiation-induced changes of erythrocytes (number, MCV) and of RNA concentration in blood. The changes of other parameters (spleen cellularity, liver CFU-S, leukocyte count and DNA concentration in blood) were the same as in unprotected animals. In conclusion, we can say that DS-pretreatment had a beneficial effect on the recovery of radiation-induced damage of erythropoiesis but not on granulopoiesis or lymphopoiesis.     

Axenic mass cultivation of the free-living soil amoeba,Acanthamoeba castellanii in a laboratory fermentor

Axenic mass cultivation of Acanthamoeba castellanii in laboratory fermentors (14 l) yielded after 20 days approximately 3 g cells (wet weight). After a short lag phase amoebal cell numbers increased exponentially to a maximum of 3.5×10⁵ cells per ml until cell death occurred after 20 days. Optical density and protein concentrations revealed identical patterns. During amoebal growth only 12–19% of the initially added glucose (100 mM) as sole carbon source was used. Large amounts of ammonia (1 g in 10.5 l culture volume) were excreted into the medium which subsequently raised the pH from 6.6 to 7.7, and from 6.6 to 6.8 in 2 and 20 mM buffered media, respectively. Growth inhibition and cell death could not be explained by a depletion of glucose or oxygen limitations during growth. The production of ammonia had a growth inhibitory effect, however, the sudden termination of the exponential growth phase and cell death could not be explained by the toxic influence of ammonia only.     

Corneal endothelial structure and function under normal and toxic conditions.

Our understanding of the function of the corneal endothelium in corneal thickness regulation, and the role of ion transport mechanisms in endothelial physiology, has expanded greatly over the past 25 years. The basic events occurring across the apical and basolateral membranes of the cells are far better understood today, although gaps still exist in the area of the relationship of the cellular and paracellular pathways and their relative contribution to the overall behavior of the endothelium. Little is known about the movement of ions or fluid between the cells or in what proportion this may occur compared to the cellular events. Furthermore, although our knowledge of the ionic movement processes has been enhanced, the link between fluid transfer across the endothelium and ion movements remains an enigma. Important questions also remain concerning the link between electrical characteristics and either ion movement or fluid transport. Improved storage solutions are needed that will preserve endothelial function after transplantation through the provision of a significant improvement in long-term cell survival. The limit to preservation time at present is about 14 days, and the use of other variables in the storage solution may extend this time. In reality, however, extension of preservation time is now of secondary importance relative to the need to enhance cell survival and reduce cell loss following surgery. Whether such improvement can be made with manipulation of the solution alone, or whether refinements are needed in the surgical technique awaits further study. Our comprehension of the biochemical linkage between energy supply and ion movement also remains uncertain in view of the particular intracellular localization of the anionic ATPases to mitochondrial loci. Despite numerous attempts there have been only a few chemicals identified that stimulate the fluid pump, but the level of stimulation has been relatively small and short-lived. No sustained effects have been found that would be of clinical benefit in reducing corneal thickness. A considerable variety of chemicals has been tested on the endothelium and it is unlikely that any new compounds will be identified that will cause enhancement of the fluid pump that would be of clinical benefit in dystrophic, or otherwise swollen, corneas. Of all the toxic responses of the endothelium the majority have been identified because of a malfunction of corneal thickness regulation, with the resultant corneal swelling, or by morphological examination. Only in a few instances has the permeability to non-electrolytes (carboxyfluorescein, inulin/dextran) been measured, and even more rarely have ion fluxes, or pump activity (3H-ouabain binding), been measured.(ABSTRACT TRUNCATED AT 400 WORDS)     

Corneal cryopreservation with dextran.

Different methods of corneal cryopreservation have been introduced, those employing intracellular cryoprotectants such as Me2SO or glycerol being the most widely favored. We investigated the influence of several freeze-thaw trauma variables on the survival of porcine endothelial monolayers when employing the extracellular cryoprotective agent dextran. We first examined the effects of various dextran concentrations and then, having ascertained the optimal concentration, further investigated the influence of fetal calf serum (FCS) concentration in the cryopreservation medium, the cooling rate, the thawing temperature, and the length of the preincubation in the freezing medium prior to cryopreservation. The numerical densities of endothelial cells were determined at dissection in hypoosmotic balanced salt solution and after organ culture by staining with alizarin red S and trypan blue. Morphological evaluation was not performed directly after thawing but after a subsequent organ culture at 37 degrees C to detect latent cell damage after freeze-thaw trauma. Our data revealed that corneas cryopreserved in minimal essential medium containing 10% dextran but lacking FCS, preincubated for 3 h, frozen at a cooling rate of 1 degrees C/min, and thawed at 37 degrees C incurred the lowest cell losses (22.4%, SD +/- 3.8). We conclude that dextran is an effective cryoprotectant for freezing of porcine corneas. However, variations between species in the results of cryopreservation require further investigation of an in vivo animal model and studies with human corneas before its clinical use can be recommended.     

An antibiotic-free medium for the xenic cultivation of Entamoeba gingivalis.

Diamond's TYI-S-33 (Trypticase-Yeast Extract-Iron-Serum) medium was used as the basis for a new antibiotic-free medium for xenic growth of Entamoeba gingivalis. Nutritional requirements of the oral protozoan were determined in an effort to optimize growth. TYI-S-33 medium did not support E. gingivalis growth prior to modification. The changes included: (a) deletion of L-cysteine.HCl and thioctic acid, (b) substitution of glucose for dextran I (mol. wt 185,000) or rice starch, (c) reduction of concentrations of tryptone (2.5 g l-1), yeast extract (1.25 g l-1) and dextran I (1 g l-1), (d) increased concentration of ferric ammonium citrate (0.2 g l-1), and (e) addition of gastric mucin (2.4 g l-1). Dextran I was chosen as the major carbon source; its use in the medium limited growth of accompanying bacteria. This new antibiotic-free medium significantly increased E. gingivalis growth (16-20 E. gingivalis trophozoites observed per field) as compared to growth in Diamond's TYSGM-9 (Trypticase-Yeast Extract-Serum-Gastric Mucin) medium (six to 10 E. gingivalis trophozoites observed per field).     

Dextran vitrification media prevents mucin coat and zona pellucida damage in rabbit embryo

Vitrification of embryos is being increasingly important for cryopreservation in mammals. However, damage and toxicity has to be reduced even more. The composition of cryoprotective medium used to immerse the embryos affects viability and developmental potential. The aim of this work was to assess the effect of the Polyvinylalcohol-PVA- and Dextran addition to vitrification media on the in vitro development of rabbit embryos from superovulated and non-superovulated females. Superovulation group were treated intramuscularly with 25 IU rhFSH. The vitrification media contained the same permeable cryoprotectans (Ethylene Glycol-ET- and Dimethyl Sulfoxide-Me₂SO-) and different macromolecules (PVA and Dextran) in different combinations. There was a significantly higher proportion of embryos without damages in mucin coat or zona pellucida after warming (undamaged embryos) in the control than in the superovulation group (95.8% vs. 83.2%, respectively). The proportion of undamaged embryos was significantly affected by the vitrification solution composition. The rate of undamaged embryos after warming in media containing 20% Me₂SO was significantly lower in media supplemented with PVA than in media with dextran (67.3 vs. 93.8, respectively). However, the proportion of undamaged embryos for the medium supplemented with dextran was similar for media with 15 or 20% Me₂SO. In conclusion, the addition of dextran to the vitrification media improve the preservation of rabbit embryos and permits to reduce the amount of Me₂SO for vitrification. Additionally, in vitro developmental ability of undamaged embryos were not affected by superovulation treatment nor vitrification media.