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Pyrroloquinoline quinone (PQQ) is a factor influencing on the mitochondrial biogenesis. In this study the PQQ effect on viability, total cell number, antioxidant capacity, mitochondrial biogenesis and differentiation potential was investigated in human induced Pluripotent Stem Cells (iPSC) - derived: neural stem cells (NSC), early neural progenitors (eNP) and neural progenitors (NP). Here we demonstrated that sensitivity to PQQ is dependent upon its dose and neural stage of development. Induction of the mitochondrial biogenesis by PQQ at three stages of neural differentiation was evaluated at mtDNA, mRNA and protein level. Changes in NRF1, TFAM and PPARGC1A gene expression were observed at all developmental stages, but only at eNP were correlated with the statistically significant increase in the mtDNA copy numbers and enhancement of SDHA, COX-1 protein level. Thus, the "developmental window" of eNP for PQQ-evoked mitochondrial biogenesis is proposed. This effect was independent of high antioxidant capacity of PQQ, which was confirmed in all tested cell populations, regardless of the stage of hiPSC neural differentiation. Furthermore, a strong induction of GFAP, with down regulation of MAP2 gene expression upon PQQ treatment was observed. This indicates a possibility of shifting the balance of cell differentiation in the favor of astroglia, but more research is needed at this point.

All classes of phospholipids present in brush border membrane are exchanged in a 1:1 ratio for egg phosphatidylcholine when brush border membrane vesicles from rabbit small intestine are incubated with small unilamellar vesicles of egg phosphatidylcholine. The exchange reaction exhibits biphasic kinetics similar to those of the hydrolysis of brush border membrane phospholipids by phospholipase A2 and sphingomyelinase C. In both reactions there is an initial fast phase followed by a markedly slower one. The phospholipid exchange appears to be catalyzed by intrinsic brush border membrane protein(s), while the digestion by phospholipases is mediated by externally added enzymes. From a comparison of the kinetics of phospholipid exchange and phospholipid hydrolysis, the following conclusions can be drawn: Both sets of experiments indicate the presence of two phospholipid pools differing in the rate of phospholipid exchange and hydrolysis. Except for sphingomyelin, the size of the two phospholipid pools derived from phospholipid exchange is in good agreement with that derived from phospholipid hydrolysis. This is the main finding of this work, and on the basis of this result the two lipid pools are tentatively assigned to phospholipid molecules located on the outer and inner layer of the brush border membrane. The slow rate of phospholipid exchange reflects the rate of transverse or flip-flop movement of phospholipids. The half-time of this motion is approximately 8 h for isoelectric (neutral) phospholipids such as phosphatidylethanolamine and approximately 80 h for negatively charged phosphatidylserine and phosphatidylinositol. Isoelectric phospholipids (phosphatidylcholine, phosphatidylethanolamine) are preferentially located on the inner (cytoplasmic) side (to about 70%) while the negatively charged phospholipids are more evenly distributed: 55-60% are located on the inner side.

The phase behaviour of lipid X at high water concentrations (> 60%) is studied using mainly differential scanning calorimetry (DSC) and X-ray diffraction. The critical micellar concentration (CMC) decreases from 8 × 10−5 M at −2 °C to 4 × 10−5 M at 20 °C. The critical micellar temperature (CMT) is 0 °C and decreases slightly with increasing lipid X concentration. Above the CMC and below the CMT, lipid X forms a lamellar gel phase (Lβ). Above 0 °C and at concentrations ranging from the CMC up to about 0.2 M (20%), lipid X forms small micelles. At even higher concentrations there is a transition to a hexagonal phase, probably hexagonal I. Addition of excess NaCl to lipid X dispersions at concentrations 0.7 M NaCl, the bilayer repeat distance decreases to about 43 Å (1 Å = 0.1 nm).Key words: lipid X, phase behaviour, effects of NaCl, differential scanning calorimetry, X-ray diffraction.

Lipid X forms small micelles of average diameter of 13 ± 2 nm in excess H2O pH 7 to 8, and under physiological conditions. The critical micellar concentration is 4 × 10–5 M, and the critical micellar temperature coincides with the ice melting temperature. Freezing of micellar dispersions of lipid X induces the conversion of the micellar phase to a lamellar gel phase with hexagonal hydrocarbon chain packing. Conversely, melting of ice in frozen lipid X dispersions induces the transition from the lamellar gel phase to a liquid crystalline phase. At room temperature different liquid crystalline phases are observed depending on the water content of lipid X dispersions. In the order of increasing water contents cubic, hexagonal and micellar phases are formed. At room temperature lipid X has therefore a preference for phases with highly curved lipid surfaces. The preference for non-lamellar phases in the fully hydrated state of lipid X at room temperature can be rationalized in terms of hydration. About 22 mol H2O/ mol lipid X are bound which is about twice as much water as is bound to phosphatidylcholine. The large amount of water which is associated with the α-D-glucosamine-l-phosphate group of lipid X gives rise to a wedge-shaped molecule. Molecules of this shape have been shown to have a preference for non-lamellar phases. The hydrocarbon chains of lipid X in the smectic (lamellar) phase pack in a typical, hexagonal lattice. Interdigitation of the hydrocarbon chains in the lamellar phase is not observed.

Resultats de l'etude du comportement de transition des phases de la dipalmitoyl-1,3 phosphatidylethanolamine dans H 2 O et D 2 O, par calorimetrie a balayage differentiel et par spectroscopie RMN- 31 P

The effect of increasing concentrations of lipid X (2,3-bis(3-hydroxymyristoyl)-alpha-D-glucosamine 1-phosphate) on the phase behaviour of EPC (egg phosphatidylcholine) and EPE (egg phosphatidylethanolamine) is studied at a pH greater than or equal to 7 where lipid X carries one to two negative charges. Small amounts of lipid X (molar ratio approximately 0.01) induce continuous swelling of EPC and EPE bilayers and consequently the formation of large unilamellar vesicles in excess water. In many respects, the effect of lipid X on EPC and EPE bilayers is similar to that of phosphatidic acid. However, lipid X/EPC mixtures form micelles in excess lipid X whereas mixtures of phosphatidic acid/EPC vesiculate at all ratios. The same is true for lipid X/EPE mixtures. Small unilamellar vesicles of an average diameter of 40 nm form spontaneously upon dispersion of a dry lipid X/EPE film (molar ratio = 10). Unsonicated dispersions of lipid X/EPC (molar ratio = 1) are subjected to pH-jump treatment which involves raising of the pH to 11-12 and subsequent lowering of the pH to between 7.5 and 8.5. Such a treatment has little effect on the vesicle size and size distribution as compared to a control dispersion at pH 8.2. The mean size is determined to be 92 +/- 60 nm. Electron micrographs of freeze-fractured samples of lipid X/EPC (molar ratio = 1) reveal the presence of mainly micelles at pH 12. Upon lowering the pH to neutrality these micelles become unstable and aggregate/fuse rapidly to unilamellar vesicles (average diameter 95 +/- 40 nm). Sonication of equimolar mixtures of lipid X and EPC at pH 7 yields small unilamellar vesicles of a diameter of 20-25 nm as well as mixed micelles of a size between 15 and 17 nm. This behaviour is again different from that of mixed EPC/phosphatidic acid dispersions which form small unilamellar vesicles. The presence of lipid X in such mixtures does not prevent the aggregation/fusion to larger vesicles during freezing of the dispersion. As with pure EPC bilayers, stabilization is, however, achieved in the presence of 10% sucrose. This indicates that the covalently bonded glucosamine group of lipid X cannot substitute water of hydration in neighbouring EPC molecules.

The phase transition properties of aqueous suspensions of a series of nonhydrated (not heated above room temperature) and hydrated 1,2 diacylphosphatidylethanolamines (PE's) have been examined by high sensitivity differential scanning calorimetry at scan rates of 0.02–1.0 K min-1. At all scan rates nonhydrated PE's show a single asymmetric transition curve of excess heat capacity as a function of temperature. Multilamellar dispersions of hydrated PE's, however, exhibit transitions with fine structure, which can be fitted as the sum of three two-state component transitions, at scan rates of 0.02–0.1 K min-1, but give only a single asymmetric transition at 1.0 K min-1. At all scan rates the transition(s) of hydrated samples occur at lower temperatures than those of nonhydrated samples. One of the component transitions of hydrated PE's may be analogous to the pretransition that occurs in 1,2 diacylphosphatidylcholines.

The influence of the linear, positively charged detergent cetyltrimethylammoniumchloride (CTAC1) on the well known phase behaviour of fully hydrated dipalmitoylphosphatidylcholine (DPPC) has been studied along fourteen isopleths of the phase diagram using high sensitivity scanning microcalorimetry. Particular interest was focused towards the effects of very small amounts of detergent down to the region of 1 CTAC1 per 10 4 DPPC-molecules. Even starting with such low concentrations distinct changes of the thermograms are detectable by high sensitivity DSC. Essentially, the phase diagram is cut into four domains between the two limiting cases of pure phospholipid multilamellar dispersion and isotropic solution of detergent micelles, respectively. At the present stage these four regions in the phase diagram are identified as follows : 1. 0–1 Mol % CTAC1: a modified lamellar phase 2. 1–50 Mol % CTAC1: a two-phase region with this modified lamellar phase coexisting with a newly formed detergent-rich phase. 3. 50–90 Mol % CTAC1: large mixed micelles. 4. > 90 Mol % CTAC1: an isotropic solution of small mixed phospholipid/detergent micelles.

The behavior of mixed bile salt micelles consisting of sodium taurocholate, egg phosphatidylcholine, and cholesterol has been studied by ESR spin labeling and synchrotron x-ray scattering. Consistent with published phase diagrams, pure and mixed bile salt micelles have a limited capacity to incorporate and, hence, solubilize cholesterol. Excess cholesterol crystallizes out, a process that is readily detected both by ESR spin labeling using 3-doxyl-5 alpha-cholestane as a probe for cholesterol and synchrotron x-ray scattering. Both methods yield entirely consistent results. The crystallization of cholesterol from mixed bile salt micelles is indicated by the appearance of a magnetically dilute powder spectrum that is readily detected by visual inspection of the ESR spectra. Both the absence of Heissenberg spin exchange and the observation of a magnetically dilute powder spectrum provide evidence for the spin label co-crystallizing with cholesterol. In mixed bile salt micelles containing egg phosphatidylcholine, the solubility of cholesterol is increased as detected by both methods. With increasing content of phosphatidylcholine and increasing mole ratio cholesterol/phosphatidylcholine, the anisotropy of motion of the spin probe increases. The spin label 3-doxyl-5 alpha-cholestane is a useful substitute for cholesterol provided that it is used in dilute mixtures with excess cholesterol: the cholesterol/spin label mole ratio in these mixtures should be greater than 100. Despite the structural similarity between the two compounds, there are still significant differences in their physico-chemical properties. These differences come to the fore when cholesterol is totally replaced by the spin-label: 3-doxyl-5a-cholestane is significantly less soluble in bile salt and mixed bile salt micelles than cholesterol and, in contrast with cholesterol, it interacts only very weakly, if at all,with phosphatidylcholine. The potential of the ESR method for detecting cholesterol crystal growth in human bile is discussed.

Interaction of lanthanum ions (La3+) with 1,2 dipalmitoyl-sn-glycero-3-phosphorylcholine (DPPC) causes an increase in Tc, the temperature of maximal excess heat capacity, and the width of the gel-to-liquid crystalline transition. At a mole ratio of La3+ to DPPC sufficient to remove the hydrocarbon chain tilt angle of DPPC, the changes in the thermodynamic parameters of the pretransition are minor, Tc and the width were unaltered and the enthalpy was reduced by only 10%. This suggests that the change in tilt angle is not a necessary concomitant of the pretransition.

The effect of 0-1.0 M sucrose on the phase-transition properties of 1,2-dipalmitoyl-3-sn-phosphatidylcholine (1,2-DPPC) was examined by high-sensitivity differential scanning calorimetry at a scan rate of 0.1 K min-1. Increasing the concentration of sucrose caused a small, but experimentally significant, increase in the temperature (Tm) of maximal excess apparent specific heat (Cmax) and in delta T 1/2 (the transition width at 1/2 Cmax), a reduction in Cmax, and a small decrease (approximately 8-10% at 1.0 M sucrose compared with 0 M sucrose) in the calorimetric enthalpy (delta Hcal) of the gel-to-liquid crystalline transition. The calorimetric parameters of the pretransition of 1,2-DPPC were not significantly affected by sucrose in the concentration range examined, except there was a 1.0 degree C increase in the temperature (Tp) of maximal excess apparent specific heat in the presence of 1.0 M sucrose. The results are discussed in terms of the possible molecular mechanisms that could have caused the observed changes and are contrasted with the results obtained by C. -H. Chen et al. (1981, Biophys. J., 36:359-367).

Two independent parameters, two characteristic temperatures, one indicating the change in the molecular organization of the core, T e and the other in the surface layer, T p were measured for a number of natural and triglyceride-enriched porcine low-density lipoprotein (LDL 1 (buoyant density 1.020–1.063 g/ml) and LDL 2 (buoyant density 1.063–1.080 g/ml) samples T c was determined by differential scanning calorimetry (DSC), whereas T 3 was measured by Mn(II) binding to the lipoprotein surface followed by electron spin resonance (ESR) spectroscopy. A significant causal relationship between T c and T s is both LDL subfractions demonstrates the surface-core interaction in LDL. The significance of that interaction is emphasized as a possible link in the chain diet → lipoprotein changes → atherosclerosis.

The B, or binding, subunit of cholera enterotoxin forms a pentameric ring structure in the intact toxin, and also when the subunit is isolated from the A subunit. The thermal denaturation of the B subunit ring was examined by differential scanning calorimetry in the presence and absence of ganglioside GM1, its natural 'receptor'. In the absence of ganglioside an irreversible endotherm was observed with maximal excess apparent heat capacity, Cmax, at 74.6 degrees C. When the ganglioside was added in increasing amounts, multiple transitions were observed at higher temperatures, the most prominent having a Cmax at 90.8 degrees C. At high ganglioside concentrations, the 74.6 degrees C transition was not observed. In addition to the thermodynamic results a model is proposed for the interaction of GM1 and B subunit pentamer. This model is derived independently of the calorimetric results (but is consistent with such data) and is based upon considerations of the geometry of the GM1 micelle B subunit pentamer.

The phase behaviour of aqueous dispersions of lipid X, a precursor of bacterial lipopolysaccharides has been investigated by a variety of physico-chemical techniques. The results are consistent with the presence of disk-shaped micelles with an average diameter of 13 +/- 1.8 nm. The critical micellar concentration in water and physiological saline is 4 x 10(-5) M. Consistent with the formation of micelles in water and physiological saline is the finding that lipid X is in the liquid-crystalline state at temperatures higher than 0 degrees C. The packing and the dynamics of lipid X are characteristic of micelles. Close to the polar group the hydrocarbon chains are significantly more mobile and disordered than in the corresponding region of lipid bilayers. From monolayer studies an estimate of the molecular area of lipid X is derived; under physiological conditions the area/molecule is about 0.50 nm2 at 30 mN/m indicating that lipid X has a wedge-like shape. The two pK values of the primary phosphate group of lipid X are pK1 approximately 1.3 and pK2 = 8.2. At pH values less than 7, the area/molecule decreases, i.e. the packing of the lipid X molecules becomes tighter, and there is also a decrease in the solubility of lipid X. As is characteristic of charged lipids, the state of aggregation (phase behaviour) of lipid X depends on pH, the ionic strength and the nature of the counterion.

Resultats de l'etude du comportement de transition des phases de la dipalmitoyl-1,3 phosphatidylethanolamine dans H 2 O et D 2 O, par calorimetrie a balayage differentiel et par spectroscopie RMN- 31 P