Effects of sciatic-conditioned medium on neonatal rat retinal cells in vitro

Schwann cells produce and release trophic factors that induce the regeneration and survival of neurons following lesions in the peripheral nerves. In the present study we examined the in vitro ability of developing rat retinal cells to respond to factors released from fragments of sciatic nerve. Treatment of neonatal rat retinal cells with sciatic-conditioned medium (SCM) for 48 h induced an increase of 92.5 +/- 8.8% (N = 7 for each group) in the amount of total protein. SCM increased cell adhesion, neuronal survival and glial cell proliferation as evaluated by morphological criteria. This effect was completely blocked by 2.5 microM chelerythrine chloride, an inhibitor of protein kinase C (PKC). These data indicate that PKC activation is involved in the effect of SCM on retinal cells and demonstrate that fragments of sciatic nerve release trophic factors having a remarkable effect on neonatal rat retinal cells in culture.     

Effects of systemic morphine on lamina I spinothalamic tract neurons in the cat

Lamina I spinothalamic tract (STT) neurons are an integral component of the central representation of pain and temperature and thus their sensitivity to various analgesics needs to be examined. In the present study, the effects of successive, cumulative doses (0.125-2.0 mg/kg) of intravenous morphine sulfate on the quantitative stimulus-response properties of nociceptive lamina I STT cells have been tested in the intact, barbiturate-anesthetized cat. Both nociceptive-specific (n = 7) and multireceptive (heat, pinch and cold sensitive; n = 7) lamina I STT cells were inhibited in a dose-dependent manner. Parallel dose-dependent effects on responses to noxious heat and pinch were generally observed that reduced ongoing discharge levels and the slopes of the stimulus-response functions. However, non-STT lamina I cells (n = 5) differed significantly; the responses of one multireceptive (heat, pinch and cold-sensitive) cell and the responses to pinch of 3 of 4 wide dynamic range cells were not inhibited. In addition, two-thirds of the nociceptive lamina I STT cells showed enhanced responses at the lowest dose of morphine (0.125 mg/kg). These results contrast with the varied effects of morphine reported for superficial dorsal horn cells with uncharacterized projections and they support the role of lamina I STT cells in pain. Furthermore, these observations are consistent with previous findings indicating that lamina I STT neurons are a distinct subpopulation of lamina I cells. These results support previous evidence that opiatergic modulation of sensory activity in lamina I is functionally organized.     

Modulation of retinal endothelial barrier in an in vitro model of the retinal microvasculature

The prediabetic/diabetic condition functionally alters the microvascular bed of the eye and the breakdown in the transvascular barrier may be produced by changes in the retinal endothelial barrier. To better understand how retinal microvessel barrier is maintained and is altered in vivo this study applies and extends our previously described in vitro permeability technique to study retinal endothelial monolayers. The model of the retinal microvasculature consists of retinal capillary endothelial cells cultured on porous microcarrier beads and perfused in chromatographic 'cell-columns'. This model design relies on indicator-dilution techniques to measure the permeability of the retinal endothelial monolayer and detects small changes in retinal endothelial permeability produced by treatments. Bovine retinal capillary endothelial cells (RCE) were obtained using an endothelial selective media. RCE were seeded at 3 x 10(4) cells cm-2 of fibronectin-coated gelatin microcarriers. After 7 days of microcarrier culture, microcarriers were poured to form columns 0.66 cm in diameter and 1.6 cm in length. The cell-column elution patterns of coinjected optically absorbing tracers (blue dextran 2 x 10(6) Da; cyanocobalamin 1355 Da; sodium fluorescein 376 Da) were analysed to estimate the permeability of the RCE monolayers covering the microcarriers. Scanning electron microscopic examination showed complete monolayer formation on the surface of the microcarriers. We found that baseline monolayer permeability averaged 7.57 +/- 0.57 x 10(-5) cm sec-1 for cyanocobalamin and 9.29 +/- 0.78 x 10(-5) cm sec-1 for sodium fluorescein (mean +/- S.E.M., n = 39). Permeability did not increase over 2 hr of cell-column perfusion. Permeability was decreased by 1 micron isoproterenol (n = 3) and increased by 1 microgram ml-1 cytochalasin D (n = 5). This is one of the first reports of in vitro permeability values for the transport barrier formed by retinal microvascular endothelial cells. Furthermore, the endothelial component of the retinal barrier is dynamic, and is enhanced by isoproterenol and diminished by cytochalasin D.     

Effects of dental trauma on the pulp

Infection of the root canal system following dental trauma induces pulp and periapical disease and prevents healing of previously healthy pulp. A clinical goal in treating trauma is the maintenance of pulp vitality, and clinicians should be aware of factors that influence pulp healing. The learning objective of this article is to review the factors and techniques that influence pulp vitality and examine the influence pulp has on the healing of adjacent tissues. The potential routes for bacterial infection of the root canal system are discussed, with the clinical crown as the primary portal of entry. Uncomplicated and complicated crown fractures, as well as the crown-root and root fractures, are reviewed. Complications in pulp healing include canal obliteration, disturbed root development, apexogenesis, apexification, and the various forms of resorption.     

Effects of WR-2721 (amifostine) and its metabolite WR-1065 on the antiproliferative activity of chemotherapeutic agents on neuroblastoma cells in vitro

Approaches through the middle cranial fossa directed at reaching the internal auditory canal (IAC) invariably employ exposure of the geniculate ganglion, the superior semicircular canal (SSC) or the epitympanum. This involves risk to the facial nerve and hearing apparatus. To minimize this risk, we conducted a laboratory study on 9 cadaver temporal bones by using an image-interactive guidance system (StealthStation) to provide topographic orientation in the middle fossa approach. Surface anatomic fiducials such as the umbo of the tympanic membrane, Henle's spine, the root of the zygoma and various sutures were used as fiducials for registration of CT-images of the temporal bone. Accurate localization of the IAC was achieved in every specimen. Mean target localization error varied from 1.20 to 1.38 mm for critical structures in the temporal bone such as the apex of the cochlea, crus commune, ampula of the SSC and facial hiatus. Our results suggest that frameless stereotaxy may be used as an alternative to current methods in localizing the IAC in patients with small vestibular schwannomas or intractable vertigo undergoing middle fossa surgery.     

Engineering dental pulp-like tissue in vitro

Injury or infection of adult dental pulp often necessitates root canal therapy. This terminates dentin formation and subsequent tooth maturation. In addition, the synthetic materials currently utilized to replace lost tooth structure are not capable of completely replacing the function of the lost tissue, and often fail over time. This report describes a technique to engineer new pulp-like tissues utilizing cultured cells and synthetic extracellular matrices. Fibroblasts were obtained from human adult dental pulps and multiplied in culture. These cells were subsequently seeded onto synthetic matrices fabricated from fibers (approximately 15 microns in diameter) of polyglycolic acid (PGA). The pulp-derived fibroblasts adhered to the fibers, proliferated, and formed a new tissue over 60 days in culture with a cellularity similar to that of native pulp. These tissues may find application in the regeneration of oral tissues and may provide novel systems in which to study the biocompatibility of materials and chemicals used in dentistry.     

Effects of dental amalgam and heavy metal cations on cytokine production by peripheral blood mononuclear cells in vitro

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) is known to produce a differential toxicity in the nigrostriatal and mesolimbic dopaminergic pathways with the nigrostriatal pathway being more vulnerable. We, therefore, investigated whether oxidative stress and the antioxidant system play a role in this phenomenon. Balb/c mice were treated with either saline or MPTP (30 mg/kg/d) for 7 d, and were sacrificed on the next day. Results revealed that MPTP increased lipid peroxidation in the striatum (ST) and decreased glutathione concentration in the substantia nigra (SN) without markedly affecting these measures in the nucleus accumbens (NAc) and ventral tegmental area (VTA). Further, MPTP produced approximately twofold increases in both manganese superoxide dismutase (MnSOD) and copper-zinc superoxide dismutase (CuZnSOD) activities in the VTA while it only increased MnSOD activity in the SN. Both catalase and glutathione peroxidase (GPx) activities were not markedly altered by MPTP in both systems. However, the basal levels of catalase and GPx activities were higher in the VTA and NAc than in the SN and ST. These results together suggest that a lesser degree of oxidative damage and a more inducible CuZnSOD activity observed in the mesolimbic dopaminergic pathway may partially explain the differential toxicity MPTP produced in these two dopaminergic systems.     

Effects of beta-adrenergic blockers on retinal circulation

We evaluated the effects of 0.5% betaxolol hydrochloride and 0.5% timolol maleate on retinal blood flow. We measured the diameter of the retinal artery (Da) and vein (Dv), and retinal venous blood flow rate (V) in 8 healthy young volunteers by laser speckle velocimetry before and after the application of betaxolol or timolol topically to both eyes daily for one week. There were no any significant changes. Da before and after the application of betaxolol was 114.2 +/- 6.3 (mean +/- standard deviation) microns and 115.6 +/- 6.7 microns and before and after the application of timolol, 117.5 +/- 12.7 microns and 101.3 +/- 9.5 microns. Dv before and after the application of betaxolol was 149.5 +/- 11.1 microns and 148.4 +/- 13.3 microns, and before and after the application of timolol 148.5 +/- 9.2 microns and 146.2 +/- 10.3 microns. V before and after the application of timolol was 12.3 +/- 2.6 mm/s and 12.6 +/- 2.4 mm/s, and before and after the application of betaxolol, 11.6 +/- 1.5 mm/s and 11.4 +/- 2.1 mm/s. Thus one-week application of the beta-blockers, betaxolol and timolol did not change the retinal arterial or venous blood flow.     

Effects of neurotrophins on embryonic retinal outgrowth

In the literature it is generally assumed that venous reflux within the radicular veins is prevented by the presence of bicuspid valves and narrowing of the transdural part of these vessels. Recently, we performed a human cadaver study of the internal vertebral venous plexus. Surprisingly, a large number of radicular and perimedullary veins appeared to be filled with Araldite CY 221 mixture, after injection of this material into the vertebral venous system, implicating reflux via the radicular veins and suggesting insufficiency of the presumed anti-reflux mechanism. Therefore, it was decided to study the radicular veins in order to determine and to investigate the presence or absence of anti-reflux mechanisms within this system. The vertebral venous systems of ten fresh human cadavers, between 64 and 93 years of age, were injected with Araldite CY 221 mixture. After polymerization, all cadavers were dissected and the spinal nerve sheaths, including nerve roots, radicular veins and epidural veins, were excised as a whole. After macroscopical examination, serial sections (40 microm) were cut on a freezing microtome and stained in Von Gieson medium. Every third section was stained immunohistochemically with smooth muscle antigen (SMA), to visualize smooth muscle cells. In all cadavers, a number of intradural radicular veins was filled with Araldite. Employing microscopical examination, no bicuspid valves were found. However, four structures were encountered that might serve as ananti-reflux-mechanism: 1) intravenous dural folds, 2) meandrous configuration, and 3) narrowing of the radicular veins at the point of penetration of the dura mater, and 4) varying numbers of smooth muscle fibers in the walls of the intradural and extradural parts of the radicular veins. Reflux via the radicular veins seems to be a physiological phenomenon. Structural valves have not been encountered during this study. Intravenous dural folds, meandrous configuration and narrowing of the transdural part of the radicular veins, and the presence of large numbers of smooth muscle cells in the radicular venous walls suggest the existence of a dynamic reflux-regulating system that has the ability to increase the intravascular resistance under conditions of venous hyperpression, in order to protect the spinal cord from venous pressure waves. Possibly, venous reflux via the radicular veins has a role in selective cooling of the spinal cord.     

Fourth module of Xenopus interphotoreceptor retinoid-binding protein: activity in retinoid transfer between the retinal pigment epithelium and rod photoreceptors

PURPOSE: Interphotoreceptor retinoid-binding protein (IRBP), an extracellular protein believed to support the exchange of retinoids between the neural retina and retinal pigment epithelium (RPE) in the vertebrate eye, exhibits a modular, i.e., repeat, structure. The present study was undertaken to determine whether an individual module of IRBP has activity in retinoid transfer between the RPE and rod photoreceptors. METHODS: The retinoid transfer activity of a recombinant protein corresponding to the fourth module of Xenopus laevis IRBP (X4IRBP) was examined in two ways. First, X4IRBP was tested for its ability to support the regeneration of porphyropsin in detached/reattached Xenopus retina/RPE-eyecups. Following illumination and removal of native IRBP, Xenopus eyecups supplemented with 42 microM X4IRBP or (as a control) Ringer's solution were incubated in darkness and then analyzed for regenerated porphyropsin. Second, toad (Bufo marinus) RPE-eyecup preparations were used to evaluate X4IRBP's ability to promote the release of 11-cis retinal from the RPE. RESULTS: The regeneration of porphyropsin in X4IRBP-supplemented Xenopus retina/RPE-eyecups (0.45 +/- 0.04 nmol; mean +/- SEM, n = 11) exceeded that in controls (0.13 +/- 0.02 nmol, n = 11). For promoting the release of 11-cis retinal from the toad RPE, 42 microM X4IRBP was more effective than equimolar bovine serum albumin although considerably less than that of 26 microM native bovine IRBP. CONCLUSIONS: The results indicate a low but significant activity of IRBP's fourth module in reactions relevant to retinoid exchange.     

Factors effecting dental composite wear in vitro

The dependence on pH of the absorption and circular dichroic spectra of iron(III), cobalt(III) and copper(II) transferrins has been (re)investigated. In the alkaline region, the CD profiles of iron(III) and cobalt(III) transferrin are essentially pH independent up to pH 11; only for very high pH values (pH > 11) is breakdown of the cobalt(III) and iron(III) transferrin derivatives observed, without evidence of conformational rearrangements. By contrast, the CD profiles of copper transferrin show drastic changes in shape around pH 10; these spectral changes, which are fitted to a pKa of approximately 10.4, are interpreted in terms of a substantial rearrangement of the local environment of the copper ions at high pH. Although the CD spectra of copper transferrin at alkaline pH strictly resemble those observed upon addition of modifier anions, the mechanism of site destabilization in the two cases is different; at variance with the case of modifier anions, our results suggest that the high pH form of copper transferrin still contains the synergistic anion. A 13C NMR experiment has confirmed this view. In the acidic region, iron(III) and cobalt(III) transferrins are stable down to pH approximately 6. For lower pH values progressive metal detachment is observed without evidence of conformational changes; around pH 4.5 most bound metals are released. In the case of the less stable copper-transferrin, metal removal from the specific binding sites is already complete around pH 6.0; in concomitance with release from the primary sites, binding of copper ions to secondary sites is observed. Additional information has been gained from CD experiments in the far UV. The pH dependent properties of iron(III), cobalt(III) and copper(II) transferrin are discussed in the frame of the present knowledge of transferrin chemistry, particular emphasis being attributed to the comparison between tripositive and bipositive metal derivatives.     

Effects of hindlimb contraction on pressor and muscle interstitial metabolite responses in the cat

We used the microdialysis technique to measure the interstitial concentration of several putative metabolic stimulants of the exercise pressor reflex during 3- and 5-Hz twitch contractions in the decerebrate cat. The peak increases in heart rate and mean arterial pressure during contraction were 20 +/- 5 beats/min and 21 +/- 8 mmHg and 27 +/- 9 beats/min and 37 +/- 12 mmHg for the 3- and 5-Hz stimulation protocols, respectively. All variables returned to baseline after 10 min of recovery. Interstitial lactate rose (P < 0. 05) by 0.41 +/- 0.15 and 0.56 +/- 0.16 mM for the 3- and 5-Hz stimulation protocols, respectively, and were not statistically different from one another. Interstitial lactate levels remained above (P < 0.05) baseline during recovery in the 5-Hz group. Dialysate phosphate concentrations (corrected for shifts in probe recovery) rose with stimulation (P < 0.05) by 0.19 +/- 0.08 and 0.11 +/- 0.03 mM for the 3- and 5-Hz protocols. There were no differences between groups. The resting dialysate K+ concentrations for the 3- and 5-Hz conditions were 4.0 +/- 0.1 and 3.9 +/- 0.1 meq/l, respectively. During stimulation the dialysate K+ concentrations rose steadily for both conditions, and the increase from rest to stimulation (P < 0.05) was 0.57 +/- 0.19 and 0.81 +/- 0.06 meq/l for the 3- and 5-Hz conditions, respectively, with no differences between groups. Resting dialysate pH was 6.915 +/- 0.055 and 6.981 +/- 0.032 and rose to 7.013 (P < 0.05) and 7.053 (P < 0.05) for the 3- and 5-Hz conditions, respectively, and then became acidotic (6. 905, P < 0.05) during recovery (5 Hz only). This study represents the first time simultaneous measurements of multiple skeletal muscle interstitial metabolites and pressor responses to twitch contractions have been made in the cat. These data suggest that interstitial K+ and phosphate, but not lactate and H+, may contribute to the stimulation of thin fiber muscle afferents during contraction.     

Effects of ovariectomy on meal pattern in the albino rat.

In a study with 24 female albino Sherman rats, it was found that after ovariectomy most Ss increased food intake while continuing to eat discrete meals. Meal size increased in ovariectomized Ss, whereas meal frequency decreased. It is suggested that ovariectomy impairs the onset of satiety during a meal but not the ability to regulate total intake through modification of intermeal interval. (21 ref) (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Rat liver ADP-ribose pyrophosphatase-I as an in vitro target of the acetaminophen metabolite N-acetyl-p-benzoquinoneimine

N-acetyl-p-benzoquinoneimine (NAPQI) is the metabolite responsible for acetaminophen hepatotoxicity. ADP-ribose pyrophosphatase-I (ADPRibase-I; EC 3.6.1.13) hydrolyzes protein-glycating ADP-ribose. The results show NAPQI-dependent alterations of ADPRibase-I leading to strong inhibition: a fast Km increase produced by low concentrations, and a time-dependent Vmax decrease by higher NAPQI concentrations. Both effects were prevented by thiols, but not reverted by them, nor by gel filtration of NAPQI-treated enzyme. Liver ADPRibase-I can be a target of NAPQI-dependent arylation. The inhibition or inactivation of the enzyme would contribute to increasing the free ADP-ribose concentration and nonenzymatic ADP-ribosylation, which is coherent with results linking free ADP-ribose-producing pathways to acetaminophen toxicity.     

Artificial salivas for in vitro studies of dental materials

In a preliminary study, levels of activity of enzymes involved in anaerobic glycerol catabolism by Enterobacter agglomerans grown in batch cultures regulated in a pH range of 6.5-8.0 were monitored. That study showed that activities of key enzymes of the downstream metabolism of glycerol--glyceradehyde-3-phosphate dehydrogenase (GAP-DH), lactate dehydrogenase and pyruvate formate lyase--were strongly dependent on the culture pH. To investigate the influence of pH on the physiology of the strain, E. agglomerans was grown anaerobically in a continuous culture supplied with glycerol as the sole carbon source and regulated at pH 8. A complete biochemical analysis was performed and was compared with that previously described for the continuous culture regulated at pH 7. A limitation of the glycolytic flux at the level of GAP-DH was demonstrated at high dilution rate, resulting in an overflow metabolism through the 1,3-propanediol formation pathway. Increasing the specific rate of glycerol consumption also resulted in enhanced lactate production due to limitation by the pyruvate decarboxylation step. Finally, changing the culture pH significantly modified the enzymatic profile of E. agglomerans, and it enabled the stability of the culture to be increased by preventing the accumulation in the fermentation broth of 3-hydroxypropionaldehyde, an inhibitory metabolite, when the glycerol supply was suddenly increased.     

In vitro corrosion measurements of dental alloys

Unguided protrusive and retrusive mandibular movement was investigated in 76 asymptomatic volunteers and temporomandibular joint (TMJ) patients using computerized axiography. In 38 of these subjects, guided pro-/retrusion was also recorded. Measurements were performed in a three-dimensional Cartesian coordinate system (x = anteroposterior; y = transversal, toward the hinge axis; z = craniocaudal). Transversal shift (y) was measured and evaluated as well as the distance "s" and horizontal condylar inclination (HCl) at the point of maximum transversal deviation. The patterns of the tracings were analyzed with a view to diagnostic improvements. The objective of this study was to evaluate transversal shift and to examine the postulate of symmetrical behavior of the TMJs in pro-/retrusion. The mean mandibular shift was 0.15 mm to the left. This maximum deviation was seen in a mean distance "s" of 7.42 mm in relation to the Cartesian coordinate system in reference position. These results suggest a functional dominance of the right part of the mandible in protrusive and retrusive movement.