The septin cytoskeleton in myelinating glia.

Myelin is organized in subdomains with distinct protein and lipid composition. How these domains are established and maintained is currently unknown. Cytoskeletal elements interacting with membrane components could generate and sustain such structural domains. Here, we demonstrate that the transmembrane myelin protein MAL interacts with the cytoskeleton protein septin 6. Septins represent a fourth filamentous system involved in membrane compartmentalization, vesicle transport and scaffold formation. We report that multiple septin complexes are associated with myelin, and that they display an overlapping but non-identical composition in the central and peripheral nervous system. The expression of distinct subsets of septins was upregulated during myelin formation in peripheral nerves and oligodendrocytes. In the PNS, septins were highly enriched in non-compact myelin compartments, particularly in the paranodal loops and the microvilli at the node of Ranvier. Importantly in myelin lacking Septin 6, the abundance of its closest homolog Sept11 was increased, suggesting a functional compensatory role. Our data demonstrate that the septin cytoskeleton is an integral component of the myelin sheath and interacts with distinct myelin constituents such as MAL. We suggest that septins are intriguing candidates for membrane compartmentalization in myelin internodes.

The myelin protein MAL affects peripheral nerve myelination: a new player influencing p75 neurotrophin receptor expression.

The myelin and lymphocyte protein (MAL) is a raft-associated membrane protein predominantly expressed by oligodendrocytes and Schwann cells. Here we show that MAL regulates myelination in the peripheral nervous system. In mice overexpressing MAL, myelination was retarded and fibers were hypomyelinated, whereas myelination in MAL knockout mice was accelerated. This was not due to impaired Schwann cell proliferation, differentiation or axonal sorting. We found that the expression level of p75 neurotrophin receptor mRNA and protein was strongly reduced in developing sciatic nerves in MAL-overexpressing mice. This reduction is well correlated with the observed alterations in myelination initiation, speed of myelination and alterations in Remak bundle development. Our results suggest a functional role for MAL in peripheral myelination by influencing the expression of membrane components that mediate axon-glia interaction during ensheathment and myelin wrapping.

Magnetization behavior of ferrofluids with cryogenically imaged dipolar chains.

Theories and simulations have demonstrated that field-induced dipolar chains affect the static magnetic properties of ferrofluids. Experimental verification, however, has been complicated by the high polydispersity of the available ferrofluids, and the morphology of the dipolar chains was left to the imagination. We now present the concentration- and field-dependent magnetization of particularly well-defined ferrofluids, with a low polydispersity, three different average particle sizes, and with dipolar chains that were imaged with and without magnetic field using cryogenic transmission electron microscopy. At low concentrations, the magnetization curves obey the Langevin equation for noninteracting dipoles. Magnetization curves for the largest particles strongly deviate from the Langevin equation but quantitatively agree with a recently developed mean-field model that incorporates the field-dependent formation and alignment of flexible dipolar chains. The combination of magnetic results and in situ electron microscopy images provides original new evidence for the effect of dipolar chains on the field-dependent magnetization of ferrofluids.

Complex magnetic susceptibility setup for spectroscopy in the extremely low-frequency range.

A sensitive balanced differential transformer was built to measure complex initial parallel magnetic susceptibility spectra in the 0.01-1000 Hz range. The alternating magnetic field can be chosen sufficiently weak that the magnetic structure of the samples is only slightly perturbed and the low frequencies make it possible to study the rotational dynamics of large magnetic colloidal particles or aggregates dispersed in a liquid. The distinguishing features of the setup are the novel multilayered cylindrical coils with a large sample volume and a large number of secondary turns (55 000) to measure induced voltages with a good signal-to-noise ratio, the use of a dual channel function generator to provide an ac current to the primary coils and an amplitude- and phase-adjusted compensation voltage to the dual phase differential lock-in amplifier, and the measurement of several vector quantities at each frequency. We present the electrical impedance characteristics of the coils, and we demonstrate the performance of the setup by measurement on magnetic colloidal dispersions covering a wide range of characteristic relaxation frequencies and magnetic susceptibilities, from chi approximately -10(-5) for pure water to chi>1 for concentrated ferrofluids.

Motor neuropathy in porphobilinogen deaminase-deficient mice imitates the peripheral neuropathy of human acute porphyria.

Acute porphyrias are inherited disorders caused by partial deficiency of specific heme biosynthesis enzymes. Clinically, porphyrias are manifested by a neuropsychiatric syndrome that includes peripheral neuropathy. Although much is known about the porphyrias' enzyme defects and their biochemical consequences, the cause of the neurological manifestations remains unresolved. We have studied porphyric neuropathy in mice with a partial deficiency of porphobilinogen deaminase (PBGD). PBGD-deficient mice (PBGD-/-) imitate acute porphyria through massive induction of hepatic delta-aminolevulinic acid synthase by drugs such as phenobarbital. Here we show that PBGD-/- mice develop impairment of motor coordination and muscle weakness. Histologically femoral nerves of PBGD-/- mice exhibit a marked decrease in large-caliber (>8 microm) axons and ultrastructural changes consistent with primary motor axon degeneration, secondary Schwann cell reactions, and axonal regeneration. These findings resemble those found in studies of affected nerves of patients with acute porphyria and thus provide strong evidence that PBGD deficiency causes degeneration of motor axons without signs of primary demyelination, thereby resolving a long-standing controversy. Interestingly, the neuropathy in PBGD-/- mice developed chronically and progressively and in the presence of normal or only slightly (twofold) increased plasma and urinary levels of the putative neurotoxic heme precursor delta-aminolevulinic acid. These data suggest that heme deficiency and consequent dysfunction of hemeproteins can cause porphyric neuropathy.

Dipolar structures in magnetite ferrofluids studied with small-angle neutron scattering with and without applied magnetic field.

Field-induced structure formation in a ferrofluid with well-defined magnetite nanoparticles with a permanent magnetic dipole moment was studied with small-angle neutron scattering (SANS) as a function of the magnetic interactions. The interactions were tuned by adjusting the size of the well-defined, single-magnetic-domain magnetite (Fe3O4) particles and by applying an external magnetic field. For decreasing particle dipole moments, the data show a progressive distortion of the hexagonal symmetry, resulting from the formation of magnetic sheets. The SANS data show qualitative agreement with recent cryogenic transmission electron microscopy results obtained in 2D [Klokkenburg, Phys. Rev. Lett. 97, 185702 (2006)] on the same ferrofluids.

Laparoscopic gastric re-banding versus laparoscopic gastric bypass as a rescue operation for patients with pouch dilatation.

BACKGROUND: The authors assessed whether laparoscopic rebanding or laparoscopic Roux-en-Y gastric bypass (LRYGBP) is the best approach for failed gastric banding after pouch dilatation. METHODS: Between January 2000 and June 2005, 489 patients underwent laparoscopic gastric banding, and of these, 33 (6.7%) required rescue procedures for pouch dilatation. Each reoperated patient was contacted to obtain information about their postoperative course. Additionally, preoperative weight and BMI, weight loss at 1 year postoperatively, weight at time of pouch dilatation and the time-period between the primary operation and pouch dilatation were analyzed. RESULTS: The most common operation for pouch dilatation was band repositioning or rebanding (16 patients). Band removal without replacement was performed in 7 patients. 8 patients underwent conversion to a LRYGBP. 1 patient underwent laparoscopic gastric sleeve resection and 1 patient received an intragastric balloon. Patients who underwent conversion to LRYGBP are very content and, although weight loss has been nearly the same as after gastric banding, they would prefer the gastric bypass operation to the gastric banding. CONCLUSION: Conversion to LRYGBP appears to offer significant advantages, and appears to be the rescue therapy of choice after failed laparoscopic gastric banding.

Paraproteinaemic neuropathies.

In this article, we review the main clinical and pathological features of paraproteinaemic neuropathies and discuss recent experimental findings. Further knowledge of the disease process at the molecular level has allowed a better characterization of clinical syndromes and has given new insights into their pathogenesis. The most convincing evidence for a causal relationship can be drawn from IgM monoclonal gammopathies with specificities directed against carbohydrate determinants of the myelin associated glycoprotein (MAG). There remain however, many unresolved questions, such as how monoclonal anti-MAG IgM antibodies cross the blood-nerve barrier and trigger a chronic demyelinating polyneuropathy while the central nervous system is essentially spared. IgM paraproteins with specificity for other molecules, such as neurofilaments, sulphatide, gangliosides, chondroitin sulphate and tubulin, have also been identified, but their pathogenetic importance remains to be elucidated. Other paraproteinaemic neuropathies such as IgG and IgA neuropathies have to be considered separately. The paraneoplastic endocrine and cytokine manifestations of rare osteosclerotic myelomas provide valuable insights into the interaction between the immune and the nervous system. The antigen-specificity of IgG and IgA monoclonal antibodies are only poorly characterized but some have been found to be directed against endoneurial determinants and a few against axonal proteins such as neurofilaments.

TNF-alpha expression in painful and nonpainful neuropathies.

OBJECTIVE: To determine whether the cytokine tumor necrosis factor alpha (TNF-alpha) acts as a pain mediator in neuropathic pain in humans. BACKGROUND: In animal models, inflammatory cytokines such as TNF-alpha have been shown to facilitate neuropathic pain. METHODS: The expression of TNF-alpha was analyzed immunohistochemically in 20 human nerve biopsy specimens of patients with painful (n = 10) and nonpainful (n = 10) neuropathies. Additionally, serum soluble TNF-alpha receptor I (sTNF-RI) levels were determined in 24 patients with neuropathies, 16 of which were painful and 8 that were painless. RESULTS: Colocalization studies by confocal fluorescence microscopy for S-100 and TNF-alpha showed expression of TNF-alpha in human Schwann cells. Patients with painful neuropathies showed a stronger TNF-alpha immunoreactivity in myelinating Schwann cells relative to the epineurial background staining compared with patients with nonpainful neuropathy (0.949 +/- 0.047 vs 1.010 +/- 0.053, p < 0.05). Although there was no difference in sTNF-RI levels between painful (n = 16) and nonpainful (n = 8) neuropathies (sTNF-RI: 1412 +/- 545 pg/mL vs 1,318 +/- 175 pg/mL), patients with a mechanical allodynia (n = 9) had elevated serum sTNF-RI (1627 +/- 645 pg/mL vs 1233 +/- 192 pg/mL, p < 0.05) compared with patients without allodynia (n = 15). CONCLUSIONS: TNF-alpha expression of human Schwann cells may be up-regulated in painful neuropathies. The elevation of sTNF-RI in patients with centrally mediated mechanical allodynia suggests that systemic sTNF-RI levels may influence central pain processing mechanisms.

Gene dosage effects in hereditary peripheral neuropathy. Expression of peripheral myelin protein 22 in Charcot-Marie-Tooth disease type 1A and hereditary neuropathy with liability to pressure palsies nerve biopsies.

A duplication of a 1.5-Megabase genomic region encompassing the gene for the peripheral myelin protein 22 (PMP22) is found on chromosome 17p11.2-12 in Charcot-Marie-Tooth disease type 1A (CMT1A), whereas the reciprocal deletion is associated with hereditary neuropathy with liability to pressure palsies (HNPP). Since most CMT1A patients harbor three copies of the PMP22 gene, and most HNPP patients carry only a single copy, a gene dosage effect has been proposed as a mechanism for both diseases. We have analyzed the steady-state expression of PMP22 protein in sural nerve biopsies from three CMT1A and four HNPP patients. Quantitative immunohistochemical determination showed that PMP22 protein expression relative to that of myelin protein zero and myelin basic protein was increased in all CMT1A patients and reduced in all HNPP patients, as compared with biopsy samples of patients with normal PMP22 gene expression. These data demonstrate that both neuropathies result from an imbalance of PMP22 protein expression.

Matrix metalloproteinase upregulation in chronic inflammatory demyelinating polyneuropathy and nonsystemic vasculitic neuropathy.

OBJECTIVE: To determine the expression pattern and cellular source of matrix metalloproteinases (MMPs) in chronic inflammatory demyelinating polyneuropathy (CIDP) and nonsystemic vasculitic neuropathy (NSVN). BACKGROUND: MMPs are endopeptidases involved in tissue destruction and infiltration by immune cells in multiple sclerosis and Guillain-Barré syndrome. Enzyme inhibitors of MMPs attenuate clinical symptoms in corresponding animal models of these diseases. MMP inhibition may therefore be a novel approach for the treatment of CIDP and NSVN. However, the spectrum of MMPs expressed in chronic inflammatory neuropathies has not been established. METHODS: The expression of MMP-2, MMP-3, MMP-7, and MMP-9 in T cells, macrophages, and stromal cells in CIDP, NSVN, and noninflammatory neuropathies (NIN) was quantitated by immunohistochemistry. Results were correlated with clinical and electrophysiologic findings. RESULTS: The production of MMP-2 and MMP-9 is increased in nerve tissue in CIDP and NSVN compared with NIN. T cells are the predominant source of MMP-2 and MMP-9 in CIDP and NSVN, whereas macrophages contribute only to a minor extent. Stromal cells of the perineurium/epineurium are an additional source of MMP-2 in NSVN, but not in CIDP. Expression of MMP-3 and MMP-7 was not detectable in CIDP or NSVN. Expression of MMP-2 and MMP-9 did not correlate with clinical disease activity and electrophysiologic measurements. CONCLUSIONS: The upregulation of MMP-2 and MMP-9 is a specific feature of CIDP and NSVN, and selective inhibitors of these enzymes could be used to prevent inflammatory tissue damage. The similar increase of MMP-2 and MMP-9 in both demyelinating (CIDP) and nondemyelinating (NSVN) neuropathies raises doubts about whether MMPs play a primary role in demyelination.

Characterisation of autoantibodies to peripheral myelin protein 22 in patients with hereditary and acquired neuropathies.

To investigate the possibility that an autoimmune mechanism may play a role in the hereditary neuropathy Charcot-Marie-Tooth type 1A (CMT1A), sera were analysed by Western blot for anti-peripheral myelin protein 22 (PMP22) autoantibodies. These sera were compared with sera from patients with CMT type 2 (CMT2), acquired peripheral neuropathies such as chronic inflammatory demyelinating neuropathy (CIDP), anti-MAG IgM neuropathy, Miller-Fisher syndrome (MFS), diabetic neuropathy and with control blood donors. Anti-PMP22 positive sera were detected in 70% of patients with CMT1 and unexpectedly in 60% of patients with CMT2. Interestingly, 44% of the patients with other peripheral neuropathies and 23% of the apparently healthy controls showed also anti-PMP22 antibody reactivity. Immunohistochemical analysis of the human anti-PMP22 antisera on healthy sural nerve sections and on PMP22-expressing COS cells revealed that these sera did not recognise endogenous PMP22. Our results indicate that anti-PMP22 autoantibodies are found in sera of patients with different types of peripheral neuropathies, but their role in the pathogenesis of these diseases remains to be determined.

Reciprocal expression of myelin-associated glycoprotein splice variants in the adult human peripheral and central nervous systems.

The L- and S-MAG isoforms differ only at their C-terminus and are believed to be functionally distinct. To obtain information on the relative expression of these alternatively spliced isoforms in humans, we cloned an S-MAG cDNA fragment. The deduced amino-acid sequence of the human S-MAG C-terminus shows fairly conservative substitutions of 4 out of the 10 residues compared to the rodent peptide. Using reverse transcription and a competitive polymerase chain reaction, we show that, in contrast to rodents, the L-MAG splice variant predominates in adult human brain while, like in rodents, S-MAG transcripts are most abundant in peripheral nerve. The results obtained by Western blot analysis and immunohistochemistry are in good agreement with the findings at the mRNA level. Animal experiments may thus be more representative for the role of MAG in human nerve than in brain.

Ultrastructural analysis of rat articular cartilage following treatment with dexamethasone and glycosaminoglycan-peptide complex.

This ultrastructural study describes a stereological analysis of rat articular cartilage, with and without exposure to dexamethasone and a chondroprotective drug used in the treatment of osteoarthritis. Normal rat cartilage was found to contain 27.6 x 10(4) chondrocytes/mm3 which occupied approximately 10% of the cartilage tissue, and the organelle content of each chondrocyte was calculated to be about 20% of the cytoplasmic volume. After 3 weeks of treatment with dexamethasone the chondrocytic volume was increased by 23% with a proportionate increase in the cellular volume of the whole cartilage, and in addition the organelle content was significantly reduced to about half that of the control chondrocytes. By contrast the administration of GP-C (RUMALON) to dexamethasone-treated animals reduced these steroid effects. No significant change was seen in the total chondrocyte numbers with either of the treatments. Whereas dexamethasone inhibits chondrocyte metabolism and produces concomitant ultrastructural changes, GP-C was found to counteract these effects, a result which supports the contention that GP-C helps to preserve chondrocyte function.

The mode of action of a glycosaminoglycan-peptide-complex (Rumalon) on articular cartilage of the rat in vivo.

Quantitative ultrastructural morphometry and autoradiography of articular cartilage were used to assess in 3 months old rats the effects of in vivo administration of dexamethasone alone or in combination with a glycosaminglycan-peptide-complex (GAGPC). Dexamethasone treatment (3 mg/kg week for three weeks) induced a decrease of 35S-sulphate incorporation in cartilage and ultrastructural changes of articular chondrocytes, mainly characterized by an increase in cell mortality rate, a decrease in length of endoplasmic reticulum, in the number of Golgi bodies and in mitochondrial pool and size. These autoradiographic and ultrastructural changes were reversed or prevented when GAGPC was administered concomitantly with dexamethasone. These results show that the modifications measured by quantitative ultrastructural morphometry of chondrocytes are consistent with changes in biosynthetic functions and that the GAGPC protects cartilage from the inhibitory effects of corticoids.

[Effect of the learning phase on safety and efficiency of laparoscopic fundoplication]. / Einfluss der Lernphase auf Sicherheit und Effizienz der laparoskopischen Fundoplicatio.

The introduction of laparoscopic techniques into surgical practice has required a learning process on the part of the surgeons involved. The duration, morbidity, and functional outcome of laparoscopic fundoplication were evaluated in our institution's first 146 cases. During a 34-month period the patients underwent laparoscopic Nissen (n = 102) or Toupet (n = 44) fundoplication. Conversion to open access was necessary in 7 cases, re-operation for complications in 2, all among the first 40 cases of the series. The median operating time was 165 min (range 75-375) in the first 40 cases, and 105 min (range 50-235) thereafter (P < 0.001). Body mass index, grade of esophagitis, and the surgeon's experience were independent predictors of the operating time. One hundred and thirty-four patients (92%) could be evaluated for recurrence of reflux, which was encountered in 2 (5%) of the first 40 cases and 8 (8%) of 94 patients in the later group.

Composition, concentration and charge profiles of water-water interfaces.

The properties of interfaces are discussed between coexisting phases in phase separated aqueous solutions of polymers. Such interfaces are found in food, where protein-rich and polysaccharide-rich phases coexist. Three aspects of such interfaces are highlighted: the interfacial profiles in terms of polymer composition and polymer concentration, the curvature dependence of the interfacial tension, and the interfacial potential, arising when one of the separated polymers is charged. In all three cases a theoretical approach and methods for experimental verification are presented.

Diverging electrophoretic and dynamic mobility of model silica colloids at low ionic strength in ethanol.

Electroacoustics and laser Doppler electrophoresis were employed to measure the mobility of surface-modified silica colloids in ethanol as a function of the ionic strength. Sufficiently low volume fractions were chosen to exclude effects of interparticle interactions. At high ionic strength, the electrophoretic mobility µ(e) is equal to the (electroacoustic) dynamic mobility µ(d) at 3.3 MHz. However, the ratio µ(d)/µ(e) increases significantly to ∼5 at low ionic strength. This increase may be related to the porous outer layer of the surface-modified silica spheres.

A differential dielectric spectroscopy setup to measure the electric dipole moment and net charge of colloidal quantum dots.

A sensitive dielectric spectroscopy setup is built to measure the response of nanoparticles dispersed in a liquid to an alternating electric field over a frequency range from 10(-2) to 10(7) Hz. The measured complex permittivity spectrum records both the rotational dynamics due to a permanent electric dipole moment and the translational dynamics due to net charges. The setup consists of a half-transparent capacitor connected in a bridge circuit, which is balanced on pure solvent only, using a software-controlled compensating voltage. In this way, the measured signal is dominated by the contributions of the nanoparticles rather than by the solvent. We demonstrate the performance of the setup with measurements on a dispersion of colloidal CdSe quantum dots in the apolar liquid decalin.

Size-dependent second virial coefficients of quantum dots from quantitative cryogenic electron microscopy.

Cryogenic transmission electron microscopy (cryo-TEM) is utilized to determine the second virial coefficient of osmotic pressure of PbSe quantum dots (QDs) dispersed in apolar liquid. Cryo-TEM images from vitrified samples provide snapshots of the equilibrium distribution of the particles. These snapshots yield radial distribution functions from which second virial coefficients are calculated, which agree with second virial coefficients determined with analytical centrifugation and small-angle X-ray scattering. The size dependence of the second virial coefficient points to an interparticle interaction that is proportional to the QD surface area. A plausible cause for this attraction is the interaction between the surface ions on adjacent QDs.