Biomedical subjects

Dirk K Hincha

Publications and source records attributed to Dirk K Hincha.

24 records · Page 2Linked to original sources

PubMed2002

Differential destabilization of membranes by tryptophan and phenylalanine during freezing: the roles of lipid composition and membrane fusion.

The stability of cellular membranes during dehydration can be strongly influenced by the partitioning of amphiphilic solutes from the aqueous phase into the membranes. The effects of partitioning on membrane stability depend in a complex manner on the structural properties of the amphiphiles and on membrane lipid composition. Here, we have investigated the effects of the amphiphilic aromatic amino acids Trp and Phe on membrane stability during freezing. Both amino acids were cryotoxic to isolated chloroplast thylakoid membranes and to large unilamellar liposomes, but Trp had a much stronger effect than Phe. In liposomes, both amino acids induced solute leakage and membrane fusion during freezing. The presence of the chloroplast galactolipids monogalactosyldiacylglycerol or digalactosyldiacylglycerol in egg phosphatidylcholine (EPC) membranes reduced leakage from liposomes during freezing in the presence of up to 5 mM Trp, as compared to membranes composed of pure EPC. The presence of the nonbilayer-forming lipid phosphatidylethanolamine increased leakage. Membrane fusion followed a similar trend, but was dramatically reduced when the anthracycline antibiotic daunomycin was incorporated into the membranes. Daunomycin has been shown to stabilize the bilayer phase of membranes in the presence of nonbilayer lipids and was therefore expected to reduce fusion. Surprisingly, this had only a small influence on leakage. Collectively, these data indicate that Trp and Phe induce solute leakage from liposomes during freezing by a mechanism that is largely independent of fusion events.

Daunorubicin↗

PubMed2002

A mechanism for stabilization of membranes at low temperatures by an antifreeze protein.

Polar fish, cold hardy plants, and overwintering insects produce antifreeze proteins (AFPs), which lower the freezing point of solutions noncolligatively and inhibit ice crystal growth. Fish AFPs have been shown to stabilize membranes and cells in vitro during hypothermic storage, probably by interacting with the plasma membrane, but the mechanism of this stabilization has not been clear. We show here that during chilling to nonfreezing temperatures the alpha-helical AFP type I from polar fish inhibits leakage across model membranes containing an unsaturated chloroplast galactolipid. The mechanism involves binding of the AFP to the bilayer, which increases the phase transition temperature of the membranes and alters the molecular packing of the acyl chains. We suggest that this change in acyl chain packing results in the reduced membrane permeability. The data suggest a hydrophobic interaction between the peptide and the bilayer. Further, we suggest that the expression of AFP type I in transgenic plants may be significant for thermal adaptation of chilling-sensitive plants.

Animals↗

PubMed2002

Vitrification of posterior corneal lamellae.

Cryopreservation of corneas has not yet been established as a routine method. Unsatisfactory experimental results with conventional techniques prompted us to explore the possibilities of vitrification. The aim of the present study was to optimize the heat exchange between the corneal tissue and cooling medium by reducing the corneal tissue volume and using a suitable sample container. A further objective was to promote vitrification by developing a new device for rapid cooling to -140 degrees C, just below the vitrification temperature of the cryopreservation medium. Experiments were done using posterior lamellar discs from pig corneas with a diameter of 7.5 mm and a thickness of 250-350 microm. The volume of tissue to be vitrified was 88% lower with posterior corneal lamellae than with the previously used corneoscleral discs. A very thin-walled (0.05 mm), teflon-coated bag served as the sample container. Immersed in only 0.1 ml of the vitrification solution VS41a, the lamellae were cooled to a final storage temperature of -196 degrees C. After warming and organ-culturing for 24h, the endothelium was stained with trypan blue and alizarin red, to determine cell viability. Vitrification of corneal lamellae without apparent ice formation or cracking of the specimen was achieved. Despite the successful vitrification, only a maximum of 10% of the endothelial cells was vital after warming. Thus, the toxicity of the cryoprotective agents and the devitrification that occurred during the heating process require further optimization of the method.

Animals↗

PubMed2002

Lipid unsaturation determines the interaction of AFP type I with model membranes during thermotropic phase transitions.

We have previously shown that antifreeze protein (AFP) type I from winter flounder interacts with the acyl chains of lipids in model membranes containing a mixture of dimyristoylphosphatidylcholine (DMPC) and the plant thylakoid lipid digalactosyldiacylglycerol (DGDG), most likely through hydrophobic interactions. By contrast, in studies with pure phospholipid membranes, no such interaction was seen. DGDG is a highly unsaturated lipid, which renders these studies quite different from the previous studies of AFP-membrane interaction where the lipids were saturated or trans-unsaturated. Therefore, it seemed possible that either the digalactose headgroups or the unsaturated DGDG acyl chains, or both, may be important for interactions of membranes with AFP type I. To distinguish between these possibilities, we catalytically hydrogenated the DGDG to obtain a galactolipid with completely saturated fatty acyl chains. The results with the hydrogenated DGDG were strikingly different from those obtained previously with the unsaturated DGDG; the clear binding of AFPs to the bilayer appeared to be lost. Nevertheless, the temperature-dependent folding of AFP type I was inhibited in the presence of liposomes containing either the unsaturated or the hydrogenated DGDG. The results indicate that the liposomes and protein still interact, even following hydrogenation of the acyl chains, perhaps at the membrane-solution interface.

Animals↗

PubMed2002

Looking beyond sugars: the role of amphiphilic solutes in preventing adventitious reactions in anhydrobiotes at low water contents.

Plants and animals that can survive dehydration accumulate high concentrations of disaccharides in their cells and tissues during desiccation. These sugars are necessary both for the depression of the membrane phase transition temperature of the dry lipid and for the formation of a carbohydrate glass. In the past decade, however, it has become clear that certain types of adventitious enzymatic reactions are possible at low water contents, which along with free-radical mediated damage, can cause hydrolysis of lipids and loss of membrane barrier function. Disaccharides do not necessarily prevent these types of reactions, which suggests that other compounds might also be necessary for protecting organisms from this type of degradation during anhydrobiosis. Arbutin, one possible example, accumulates in large quantities in certain resurrection plants and has been shown to inhibit phospholipase A(2) activity at low water contents. The direct effect of arbutin on membranes under stress conditions depends on the membrane lipid composition. It can serve a protective function during desiccation- or freeze/thaw-induced stress in the presence of nonbilayer-forming lipids or a disruptive function in their absence. Other possible amphiphiles, including certain naturally occurring flavonols, may serve as anti-oxidants and some might have similar lipid composition-dependent effects. Such compounds, therefore, are likely to be localized near specific membranes, where they might provide the greatest benefit at the least liability to the organism.

Adaptation, Physiological↗

PubMed2002

Specific effects of fructo- and gluco-oligosaccharides in the preservation of liposomes during drying.

The fructan family of oligo- and polysaccharides is a group of molecules that have long been implicated as protective agents in the drought and freezing tolerance of many plant species. However, it has been unclear whether fructans have properties that make them better protectants for cellular structures than other sugars. We compared the effects of fructans and glucans on membrane stability during air-drying. Although glucans of increasing chain length were progressively less able to stabilize liposomes against leakage of aqueous content after rehydration, fructans showed increased protection. On the other hand, glucans became more effective in protecting liposomes against membrane fusion with increasing chain length, whereas fructans became less effective. Fourier transform infrared spectroscopy showed a reduction of the gel to liquid-crystalline phase transition temperature (T(m)) of air-dried liposomes by approximately 25 degrees C in the presence of sucrose and maltose. For the respective pentasaccharides, the reduction of T(m) of the lipids was 9 degrees C lower for samples containing fructan than for those containing glucan, indicating increased sugar--membrane interactions for the fructan compared to the glucan. A reduced interaction of the longer-chain glucans and an increased interaction of the respective fructans with the phospholipid head groups in the dry state was also indicated by dramatic differences in the phosphate asymmetric stretch region of the infrared spectrum. Collectively, our data indicate that the fructo-oligosaccharides accumulated in many plant species under stress conditions could indeed play an important role in cellular dehydration tolerance.

Cichorium intybus↗