In vitro biocompatibility of a novel membrane of the composite poly(vinylidene-trifluoroethylene)/barium titanate

This study was aimed at investigating the in vitro biocompatibility of a novel membrane of the composite poly(vinylidene-trifluoroethylene)/barium titanate (P(VDF-TrFE)/BT). Osteoblastic cells were obtained from human alveolar bone fragments and cultured under standard osteogenic condition until subconfluence. First passaged cells were cultured on P(VDF-TrFE)/BT and expanded polytetrafluoroethylene (e-PTFE - control) membranes in 24-well plates. Cell adhesion and spreading were evaluated at 30 min, and 4 and 24 h. For proliferation assay, cells were cultured for 1, 7, and 10 days. Cell viability was detected by trypan blue at 7 and 10 days. Total protein content and alkaline phosphatase (ALP) activity were measured at 7, 14, and 21 days. Cultures were stained with Alizarin red at 21 days, for detection of mineralized matrix. Data were compared by ANOVA and Student t test. Cell attachment (p = 0.001), cell number (p = 0.001), and ALP activity (p = 0.0001) were greater on P(VDF-TrFE)/BT. Additionally, doubling time was greater on P(VDF-TrFE)/BT (p = 0.03), indicating a decreased proliferation rate. Bone-like nodule formation took place only on P(VDF-TrFE)/BT. The present results showed that both membranes are biocompatible. However, P(VDF-TrFE)/BT presented a better in vitro biocompatibility and allowed bone-like nodule formation. Therefore, P(VDF-TrFE)/BT could be an alternative membrane to be used in guided tissue regeneration. (c) 2006 Wiley Periodicals, Inc.

On non-linear dynamics and a periodic control design applied to the potential of membrane action

The Fitzhugh-Nagumo (fn) mathematical model characterizes the action potential of the membrane. The dynamics of the Fitzhugh-Nagumo model have been extensively studied both with a view to their biological implications and as a test bed for numerical methods, which can be applied to more complex models. This paper deals with the dynamics in the (FH) model. Here, the dynamics are analyzed, qualitatively, through the stability diagrams to the action potential of the membrane. Furthermore, we also analyze quantitatively the problem through the evaluation of Floquet multipliers. Finally, the nonlinear periodic problem is controlled, based on the Chebyshev polynomial expansion, the Picard iterative method and on Lyapunov-Floquet transformation (L-F transformation).

CHOLESTEROL MODULATION OF LIPID PROTEIN INTERACTIONS IN LIVER MICROSOMAL MEMBRANE - A SPIN LABEL STUDY

ESR spectra of spin probes were used to monitor lipid-protein interactions in native and cholesterol-enriched microsomal membranes. In both systems composite spectra were obtained, one characteristic of bulk bilayer organization and another due to a motionally restricted population, which was ascribed to lipids in a protein microenvironment. Computer spectral subtractions revealed that cholesterol modulates the order/mobility of both populations in opposite ways, i.e., while the lipid bilayer region gives rise to more anisotropic spectra upon cholesterol enrichment, the spectra of the motionally restricted population become indicative of increased mobility and/or decreased order. These events were evidenced by measurement of both effective order parameters and correlation times. The percentages of the motionally restricted component were invariant in native and cholesterol-enriched microsomes. Variable temperature studies also indicated a lack of variation of the percentages of both spectral components, suggesting that the motionally restricted one was not due to protein aggregation. The results correlate well with the effect of cholesterol enrichment on membrane-bound enzyme kinetics and on the behavior of fluorescent probes [Castuma & Brenner (1986) Biochemistry 25, 4733-4738]. Several hypothesis are put forward to explain the molecular mechanism of the cholesterol-induced spectral changes.

New attachment achieved by guided tissue regeneration using a bioresorbable polylactic acid membrane in dogs

Created periodontal defects in dogs were randomly assigned for experimental (Guidor bioresorbable membranes) or control (conventional therapy) treatment the results showed that the new connective issue attachment was significantly greater in test sites than in controls. This new attachment averaged 2.79 +/- 0.74 mm and 1.47 +/- 0.20 mm at test and control sites, respectively (P < 0.05). Epithelial downgrowth was also reduced in the test sites (P < 0.05). No differences in bone response were found. The bioresorbable barrier was effective in blocking gingival epithelial downgrowth and connective tissue proliferation, promoting new attachment according to the principles of guided tissue regeneration.

A new method for the selective quantitation of cyanogenic glycosides by membrane introduction mass spectrometry

A new method is described for the rapid, sensitive, virtually interference-free, and selective quantitation of cyanogenic glycosides in aqueous extracts using membrane introduction mass spectrometry (MIMS). Selective monitoring, by either conventional MIMS or cryotrap-MIMS, not of HCN but of the co-released ketones (acetone and butan-2-one), when performed for both the crude cassava extracts and the linamarase-NaOH-hydrolyzed extracts, is found to offer an advantageous alternative to classic spectrophotometric methods based on HCN analysis for the selective quantitation of the two cyanogenic glycosides linamarin and lotaustralin expressed as both the free HCN content and the total cyanogenic potential (total HCN).

IDENTIFICATION OF A PUTATIVE MEMBRANE-INSERTED SEGMENT IN THE ALPHA-TOXIN OF STAPHYLOCOCCUS-AUREUS

To gain a fuller understanding of the regions of the Staphylococcus aureus alpha-toxin important in pore formation, we have used Forster dipole-dipole energy transfer to demonstrate that a central glycine-rich region of alpha-toxin (the so-called ''hinge'' region) inserts deeply into the bilayer on association of toxin with liposomes. Mutant alpha-toxins with unique cysteine (C) residues at positions 69 and 130 [Palmer, M., et al. (1993) J. Biol. Chem. 268, 11959) were reacted with the C-specific fluorophore acrylodan, which acted as an energy donor. The chosen acceptor was N-(7-nitrobenz-2-oxa-13-diazol-4-yl)-1,2-bis(hexadecanoyl) -sn-glycero-3-phosphoethanolamine (NBD-PE). Measurement of the degree of donor quenching with increasing NBD-PE in the inner bilayer leaflet enables the distance of closest approach between donor and acceptor to be estimated. For toxin labeled with acrylodan at position 130 (in the hinge region), this distance is approximately 5 +/- 2 Angstrom, showing that the probe is close to the inner surface of the liposomes. A second probe labeled at position 69 (in the N-terminal domain) shows negligible energy transfer, indicating a distance of closest approach >40 Angstrom. This implies that this N-terminal region remains ''outside'' the liposome. We propose a model in which the central region of the alpha-toxin inserts into the membrane and possibly participates in forming the wall of the pore.

Secretion of metalloendopeptidase 24.15 (EC 3.4.24.15)

The metalloendopeptidase EP24.15 (EC3.4.24.15) is a neuropeptide-metabolizing enzyme present in neural and endocrine tissues, presumably functioning extracellularly, Because the majority of the EP24.15 activity is identified in the soluble fraction of cellular homogenates, suggesting that the enzyme is primarily an intracellular protein, we addressed the issue of how EP24.15 arrives in the extracellular environment, We utilized a model system of neuroendocrine secretion, the AtT20 cell, According to both enzymatic activity and immunologic assays, EP24.15 was synthesized in and released from AtT20 cells. Under basal conditions and after stimulation by corticotropin-releasing hormone or the calcium ionophore A23187, EP24.15 activity accumulated in the culture medium. This secretion was not attributable to cell damage, as judged by the absence of release of cytosolic enzyme markers and the ability to exclude trypan blue dye. Pulse-chase analysis and subcellular fractionation of AtT20 cell extracts suggested that the mechanism of EP24.15 secretion is not solely via classical secretory pathways, Additionally, drugs which disrupt the classical secretory pathway, such as Brefeldin A and nocodazole, blocked A23187-stimulated EP24.15 release yet had no effect on basal EP24.15 release, suggesting differences in the basal and stimulated pathways of secretion for EP24.15. In summary, EP24.15 appears to be secreted from AtT20 pituitary cells into the extracellular milieu, where the enzyme can participate in the physiologic metabolism of neuropeptides.