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Endovenous laser treatment to promote venous occlusion.

BACKGROUND AND OBJECTIVES: Current treatment methods of superficial venous insufficiency (SVI) can be painful or result in incomplete occlusion. The objective of this study was to evaluate a technique for laser endovenous ablation with a newly developed diffuser fiber. STUDY DESIGN/MATERIALS AND METHODS: Six lateral saphenous veins in three goats were used. A specifically designed diffuser laser fiber tip was employed in all trials to deliver a wavelength of 1,064 nm. Each segment was treated with a different energy fluence by changing the power setting of the laser or the withdrawal rate of the fiber. Energy fluence was calculated by dividing the Joules employed for each segment over the internal surface area of the vessel. Segments were evaluated with ultrasound and histologically. RESULTS: Seventy-five percent of the segments were occluded when an energy fluence of greater than 85 J/cm2 was employed. No perforations were observed, but perivascular changes were more common at higher energy fluence. CONCLUSIONS: Endovenous laser occlusion of veins with minimal perivascular effects can be achieved with laser wavelengths of 1,064 nm if an energy fluence of 84.9-224 J/cm2 is employed and circumferential effect is achieved.

Animals↗

Classification and quantification of neuronal fiber pathways using diffusion tensor MRI.

Quantitative characterization of neuronal fiber pathways in vivo is of significant neurological and clinical interest. Using the capability of MR diffusion tensor imaging to determine the local orientations of neuronal fibers, novel algorithms were developed to bundle neuronal fiber pathways reconstructed in vivo with diffusion tensor images and to quantify various physical and geometric properties of fiber bundles. The reliability of the algorithms was examined with reproducibility tests. Illustrative results show that consistent physical and geometric measurements of novel properties of neuronal tissue can be obtained, which offer considerable potential for the quantitative study of fiber pathways in vivo.

Algorithms↗

Omental graft improves functional recovery of transected peripheral nerve.

The omentum has several properties that are advantageous for neuronal sprouting and direction. We have therefore analyzed functional recovery following transection of rat sciatic nerve using omental graft to bridge the nerve defect. In group 1, a 25-30-mm nerve defect was produced and bridged with omental graft, whereas in group 2, an end-to-end repair was performed. The sciatic function index (SFI) was assessed at 2-week intervals until 8 weeks after surgery. Functional recovery was faster in group 1 than in group 2. After 8 weeks, SFI was improved significantly from -100% to -45% (+/- -4%) in group 1 (P < 0.001) compared to -72% +/- -2% in group 2 (n = 10). Histologically, the omental graft contained more newly developed nerve fibers and less scar tissue than the end-to-end repair. Thus, omental graft appears to improve directional growth of regenerating axon sprouts and may be a means of treating peripheral nerve injury.

Animals↗

Short- or long-term effects of adult myoblast transfer on properties of reinnervated skeletal muscles.

Skeletal muscle demonstrates a force deficit after repair of injured peripheral nerves. Data from the literature indicate that myoblast transfer enhances recovery of muscle function. Thus, we tested the hypothesis that transfer of adult myoblasts improves the properties of reinnervated rabbit tibialis anterior (TA) muscles in both the short term (4 months) and long term (14 months). Two months after transection and immediate suture of the common peroneal nerve, TA muscles were made to degenerate by cardiotoxin injection and then transplanted with adult myoblasts cultured for 13 days. Under these conditions, muscles studied at 4 months were heavier, contained larger fibers, and developed a significantly higher maximal force than muscles that had only been denervated-reinnervated. In the long term, although muscles made to degenerate were heavier and developed a significantly higher maximal force than denervated-reinnervated muscles, myoblast transfer failed to improve these parameters. However, the overall characteristics of long-term operated muscles tended clearly to approach those of the controls. Taken together, these results may have significant implications in certain orthopedic contexts, particularly after immediate or delayed muscle reinnervation.

Animals↗

Induction of spheroid cytoplasmic bodies in a rat muscle by local tetanus.

The term "cytoplasmic body" or "spheroid body" myopathy refers to a heterogeneous group of familial or sporadic diseases characterized primarily by the presence of abundant spheroid or cytoplasmic bodies in the muscles. The morphogenesis of these inclusions remains unclear. This article describes the induction and evolution of spheroid cytoplasmic bodies (SCBs) in the rat plantaris muscle (PL) with local tetanus, which was induced in rats by the injection of a minute amount of tetanus toxin. In contrast to the tetanized soleus muscle (SOL), which developed core fibers (central cores, minicore, target fiber, targetoid fiber, and rods), the tetanized PL produced numerous SCBs with a predictable time course. They were induced in both type 1 and 2 fibers of PL, which is composed predominantly (95%) of type 2 fibers, in contrast to SOL (85% type 1 fibers). Factors inducing SCBs may include immobilization, shortening, intact innervation, and disuse atrophy.

Actins↗

Nicotine decreases agrin signaling and acetylcholine receptor clustering in C2C12 myotube culture.

The clustering of acetylcholine receptors (AChRs) in skeletal muscle fibers is a critical event in neuromuscular synaptogenesis. AChRs in concert with other molecules form postsynaptic scaffolds in response to agrin released from motor neurons as motor neurons near skeletal muscle fibers in development. Agrin drives an intracellular signaling pathway that precedes AChR clustering and includes the tyrosine phosphorylation of AChRs. In C2C12 myotube culture, agrin application stimulates the agrin signaling pathway and AChR clustering. Previous studies have determined that the frequency of spontaneous AChR clustering is decreased and AChRs are partially inactivated when bound by the acetylcholine agonist nicotine. We hypothesized that nicotine interferes with AChR clustering and consequent postsynaptic scaffold formation. In the present study, C2C12 myoblasts were cultured with growth medium to stimulate proliferation and then differentiation medium to stimulate fusion into myotubes. They were bathed in a physiologically relevant concentration of nicotine and then subject to agrin treatment after myotube formation. Our results demonstrate that nicotine decreases agrin-induced tyrosine phosphorylation of AChRs and decreases the frequency of spontaneous as well as agrin-induced AChR clustering. We conclude that nicotine interferes with postsynaptic scaffold formation by preventing the tyrosine phosphorylation of AChRs, an agrin signaling event that precedes AChR clustering.

Agrin↗

The activated insulin-like growth factor I receptor induces depolarization in breast epithelial cells characterized by actin filament disassembly and tyrosine dephosphorylation of FAK, Cas, and paxillin.

Insulin-like growth factor I (IGF-I) promotes the motility of different cell types. We investigated the role of IGF-I receptor (IGF-IR) signaling in locomotion of MCF-7 breast cancer epithelial cells overexpressing the wild-type IGF-IR (MCF-7/IGF-IR). Stimulation of MCF-7/IGF-IR cells with 50 ng/ml IGF-I induced disruption of the polarized cell monolayer followed by morphological transition toward a mesenchymal phenotype. Immunofluorescence staining of the cells with rhodamine-phalloidin revealed rapid disassembly of actin fibers and development of a cortical actin meshwork. Activation of phosphatidylinositol (PI)3-kinase downstream of the IGF-IR was necessary for this process, as blocking PI 3-kinase activity with the specific inhibitor LY 294002 at 10 microM prevented disruption of the filamentous actin. In parallel, IGF-IR activation induced rapid and transient tyrosine dephosphorylation of focal adhesion proteins p125 focal adhesion kinase (FAK), p130 Crk-associated substrate (Cas), and paxillin. This process required phosphotyrosine phosphatase (PTP) activity, since pretreatment of the cells with 5 microM phenylarsine oxide (PAO), an inhibitor of PTPs, rescued FAK and its associated proteins Cas and paxillin from IGF-I-induced dephosphorylation. In addition, PAO-pretreated cells were refractory to IGF-I-induced morphological transition. Thus, our findings reveal a new function of the IGF-IR, the ability to depolarize epithelial cells. In MCF-7 cells, mechanisms of IGF-IR-mediated cell depolarization involve PI 3-kinase signaling and putative PTP activities.

Actins↗

Measurement of rat brain spectra.

A method of optically monitoring oxygen saturation of blood haemoglobin in the rat brain stem was developed. Optical fiber bundles were attached to the orifices of both ears of the rat to irradiate incident light from one ear and receive transmitted light from the other ear. Absorption spectra were measured using a white-light source and a photodiode array spectrophotometer. Stable measurements of optical time courses and absorption spectra were made using this "ear-to-ear" path. Oxygen saturation levels were calculated from spectra by using a suitable reference and an improved method of determining spectral peak heights.

Animals↗

Changes in extracellular potassium concentration in cat spinal cord in response to innocuous and noxious stimulation of legs with healthy and inflamed knee joints.

In 20 cats anaesthetized with alpha-chloralose and spinalized at the thoracolumbar junction we investigated the role of stimulation induced accumulation of extracellular potassium in the spinal cord in the processing of nociceptive discharges from the knee joint. For that we electrically stimulated the posterior articular nerve of the knee. We further performed innocuous and noxious stimulation of the knee and of other parts of the leg and studied the effect of an acute inflammation of the knee on [K+]0 in the spinal cord. Innocuous stimulation of the skin (brushing or touching) and innocuous movements in the knee joint all induced rises in [K+]0 which were maximal at recording depths of 1500 to 2200 microns below the surface of the cord dorsum. Peak increases were 0.4 mM for touching the leg and 1.7 mM during rhythmic flexion/extension of the knee joint. Noxious stimulation of the skin, the paw, the tendon and noxious movements of the knee joint also produced rises in [K+]0, which were somewhat larger for the individual types of stimuli than those produced by innocuous intensities. Electrical stimulation of the posterior articular nerve induced rises in [K+]0 by up to 0.6 mM. Stimulus intensities sufficient to activate unmyelinated group IV fibers were only slightly effective in raising [K+]0 above the levels reached during stimulation of myelinated group II and III fibers. During development of an acute inflammation of the knee joint (induced by kaolin and carrageenan), increases in [K+]0 and associated field potentials became larger by about 25%. We assume that this reflects an increase in neuronal responses. In conclusion, changes in [K+]0 in the spinal cord are somewhat larger during noxious stimulation than during innocuous stimulation. The absolute level reached depended more on the site and type of stimulation than on the actual stimulus intensity itself. Hence a critical role of spinal K+ accumulation for nociception is unlikely.

Animals↗

Role of fibroblast migration in collagen fiber formation during fetal and adult dermal wound healing.

Adult dermal wounds, in contrast to fetal wounds, heal with the formation of scar tissue. A crucial factor in determining the degree of scarring is the ratio of types I and III collagen, which regulates the diameter of the combined fibers. We developed a reaction-diffusion model which focuses on the control of collagen synthesis by different isoforms of the polypeptide transforming growth factor-beta (TGF beta). We used the model to investigate the current controversy as to whether the fibroblasts migrate into the wound from the surrounding unwounded dermis or from the underlying subcutaneous tissue. Numerical simulations of a spatially independent, temporal model led to a value of the collagen ratio consistent with that of healthy tissue for the fetus, but corresponding to scarring in the adult. We investigated the effect of topical application of TGF beta and show that addition of isoform 3 reduces scar tissue formation, in agreement with the experiment. However, numerical solutions of the reaction-diffusion system do not exhibit this sensitivity to growth factor application. Mathematically, this corresponds to the observation that behind healing wave-front solutions, a particular healed state is always selected independent of transients, even though there is a continuum of possible positive steady states. We explain this phenomenon using a caricature system of equations, which reflects the key qualitative features of the full model but has a much simpler mathematical form. Biologically, our results suggest that the migration into a wound of fibroblasts and TGF beta from the surrounding dermis alone cannot account for the essential features of the healing process, and that fibroblasts entering from the underlying subcutaneous tissue are crucial to the healing process.

Animals↗

Biosynthesis and structure of glycosyl diglycerides, steryl glucosides, and acylated steryl glucosides.

A particulate enzyme fraction from Mycobacterim smegmatis catalyzed the transfer of -14C-glucose from the UDP--14C-glucose into neutral glycolipids. The two major radioactive components were purified by column chromatography on O-diethylamino ethyl cellulose (acetate) and thin layer chromatography on silica gel in several solvents. The first product yielded a water-soluble component upon saponification, which had a hexoseglycerol ratio of 1:1 with all of the hexose being identified as glucose. The second product yielded a water-soluble component upon saponification which contained hexose and glycerol in a 2:1 ratio and, in addition to glucose, contained lesser amounts of mannose and galactose. Palmitate and oleate were the predominant fatty acids and were present in equimolar amounts. The products thus have been identified as monoglycosyldiglyceride and diglycosyldiglyceride. The diglycosyldiglyceride could also be labeled with -14C-galactose when UDP--14C-galactose served as the donor, but the monoglycosyldiglyceride was only slightly labeled with -14C-galactose. Membrane fractions from mung bean seedlings catalyzed the transfer of -14C-glucose from UDP--14C-glucose into a neutral glycolipid which has been purified by thin layer chromatography and analyzed by combined gas liquid chromatography-mass spectrometry. It was determined to be a steryl glucoside with the two major sterol components being betasitosterol and stigmasterol linked to beta-D-glucose. Particulate fractions from developing cotton fibers also catalyzed the formation of steryl glucosides and, in addition, they catalyzed the esterification of steryl glucosides at the 6 position of glucose with fatty acids (primarily palmitate and oleate) from an andogenous acyl donor. Both the glucosyl transferase and the acyltransferase have been solubilized with Triton X-100 and partially purified by chromatography on Sephadex G-200. The acyltransferase activity was reconstituted by the addition of the steryl glucoside and a phospholipid acyl donor.

Acyltransferases↗

Detection of heavy metals in water by fluorescence spectroscopy: on the way to a suitable sensor system.

In order to develop a fiber optical heavy metal ion detection system, the applicability of selected complexing agents with fluorescent properties has been studied. Beginning with the application of known chelators, like BTC-5N, Newport Green, neocuproine, and chromotropic acid, a sensor configuration has been found, which allows the detection of Cd2+, Ni2+, and Cu2+ well below the chemical parameter threshold values of the new Water Quality Directive 98/83/EU. The sensor itself uses a membrane separation of the chelator flow from the sample volume. The diffusion across the membrane limits the response time to about 15 to 20 min. Applications are seen in monitoring networks.

Journal Article↗

Vulnerability to aging in the rat serotonergic system.

Distributional changes of serotonergic fibers associated with aging were demonstrated immunohistochemically. Old rat brains were morphologically characterized by the presence of peculiar features of serotonergic fibers not found in the young adult brain. In 24-month-old rats, these aberrant serotonergic fibers were subdivided into two groups according to morphological alterations: type 1 fibers consisting of thin fibers with moderately enlarged varicosities, and type 2 fibers consisting of much thicker fibers that have even larger varicosities and a tortuous course. These two types of fibers were distributed differentially in the forebrain. Type 1 fibers were found mainly in the striatum and frontoparietal cortex, whereas type 2 fibers were found in the posterior cingulate cortex and dentate gyrus of the hippocampal formation. Both types of aberrant fibers were seen in amygdala, frontoparietal cortex, hypothalamus, and thalamus. In 36-month-old rats, more highly degenerating arborizations were detected, and these aberrant ramifications were classified as follows based on shape as: type 3 fibers consisting of highly arborized thin fibers with a larger number of larger varicosities, and type 4 fibers consisting of thick fibers with abundant larger varicosities. Distributional difference indicated that type 1 fibers progress into type 3 fibers, whereas type 2 develop into type 4 fibers. These findings suggest the possibility that one set of pathological fibers emanate from the dorsal raphe nucleus and the other from the median raphe. Moreover, both two sets of serotonergic fibers show age-related aberrations in their morphology over same time course.

Aging↗

Immunohistochemical localization of aromatic L-amino acid decarboxylase in mouse taste buds and developing taste papillae.

Aromatic L-amino acid decarboxylase (AADC) catalyses the decarboxylation of all aromatic L-amino acids. In mammals, AADC is expressed in many tissues besides the nervous system, and is associated with additional regulatory roles of dopamine and serotonin in a wide range of tissues. We examined the expression of AADC by using reverse transcription-polymerase chain reaction (RT-PCR) and immunohistochemistry. RT-PCR analysis showed that mRNA of AADC was detected in the taste bud-containing epithelium of the circumvallate papilla of mice. By immunohistochemical analyses, AADC was detected in a subset of taste bud cells of fungiform, foliate, and circumvallate papillae. Double-label studies showed that AADC colocalized with serotonin, NCAM, PLCbeta2, and PGP9.5. On the other hand, AADC never colocalized with alpha-gustducin. Our results of double staining with AADC and taste cell markers indicate that only the type III cells could convert 5-hydroxytryptophan (5-HTP) to serotonin within taste buds. Taken together with previous studies, the properties of the type III cell of taste buds exactly fit into the APUD (amine and amine precursor uptake and decarboxylation) cell scheme. Furthermore, in the developing circumvallate papilla, AADC are first detected in a small number of papillary epithelial cells at E14.5. By E18.5, AADC-positive epithelial cells also express PGP9.5, which is one of marker of taste cells, and these cells have been contacted by developing nerve fibers. These results suggest that AADC expression begins at early stages of taste bud cell differentiation, and biogenic amines may act on taste bud differentiation of tongue epithelial cells, and further may regulate innervation of taste bud progenitor cells.

Animals↗

Hepatic stellate cells (vitamin A-storing cells) change their cytoskeleton structure by extracellular matrix components through a signal transduction system.

When cultured on a polystyrene surface or aminoalkylsilane-coated cover glasses, rat and human hepatic stellate cells exhibit a flattened, fibroblast-like shape with well-developed stress fibers. However, culturing the cells on type I collagen gel results in the elongation of long, multipolar cellular processes, whereas cells cultured on Matrigel maintain their round shapes. Dual fluorescence staining of microtubules and fibrillar actin indicated that the processes extend together with collagen fibers and contained microtubules as the core, whereas the periphery contained fibrillar actin. Immunofluorescence staining of vinculin showed that the focal adhesions were distributed mainly in lamellipodia when cultured on aminoalkylsilane-coated cover glasses, whereas in the cells cultured on type I collagen gel they were localized to the tips of the processes and along their bottom surface contacting collagen fibers. Wortmannin, as well as staurosporin and herbimycin A, inhibited the elongation process and induced the retraction of elongated processes. The wortmannin treatment also resulted in an alteration in focal adhesion distribution from the processes to cell bodies. These results indicate that the cell surface integrin binding to interstitial collagen fibers induces the elongation of processes through signaling events and the subsequent cytoskeleton assembly in hepatic stellate cells.

Androstadienes↗

Hemicellulose bioconversion.

Various agricultural residues, such as corn fiber, corn stover, wheat straw, rice straw, and sugarcane bagasse, contain about 20-40% hemicellulose, the second most abundant polysaccharide in nature. The conversion of hemicellulose to fuels and chemicals is problematic. In this paper, various pretreatment options as well as enzymatic saccharification of lignocellulosic biomass to fermentable sugars is reviewed. Our research dealing with the pretreatment and enzymatic saccharification of corn fiber and development of novel and improved enzymes such as endo-xylanase, beta-xylosidase, and alpha- l-arabinofuranosidase for hemicellulose bioconversion is described. The barriers, progress, and prospects of developing an environmentally benign bioprocess for large-scale conversion of hemicellulose to fuel ethanol, xylitol, 2,3-butanediol, and other value-added fermentation products are highlighted.

Biotechnology↗

Oxidized low density lipoproteins exert arrhythmogenic effects in rabbit Purkinje fibers.

The electrophysiological effects of oxidized low density lipoproteins (ox-LDLs) have been studied in rabbit Purkinje fibers using standard microelectrode techniques, in comparison with native LDLs (n-LDLs) and lysophosphatidylcholine (LPC). At the concentration of 100 micrograms protein/ml, ox-LDL but never n-LDL induced the abrupt occurrence of abnormal electrical activities during the basic stimulation of 1 Hz (6/13 fibers) and the development of either early afterdepolarizations (6/13 fibers) or abnormal automaticity (4/13 fibers) at low frequencies (0.1 and 0.03 Hz). Short trains of rapid stimulation (2, 3, 4 and 5 Hz) did not trigger delayed afterdepolarizations. However, early afterhyperpolarizations were commonly seen after each action potential. 30 microM LPC caused quite similar electrophysiological derangements. The results suggest that ox-LDLs may exert arrhythmogenic effects partly explained by their LPC content.

Action Potentials↗