The ""single-section"" Golgi method adapted for formalin-fixed human brain and light microscopy

The Golgi method has been used for over a century to describe the general morphology of neurons in the nervous system of different species. The ""single-section"" Golgi method of Gabbott and Somogyi (1984) and the modifications made by Izzo et al. (1987) are able to produce consistent results. Here, we describe procedures to show cortical and subcortical neurons of human brains immersed in formalin for months or even years. The tissue was sliced with a vibratome, post-fixed in a combination of paraformaldehyde and picric acid in phosphate buffer, followed by osmium tetroxide and potassium dicromate, ""sandwiched"" between cover slips, and immersed in silver nitrate. The whole procedure takes between 5 and 11 days to achieve good results. The Golgi method has its characteristic pitfalls but, with this procedure, neurons and glia appear well-impregnated, allowing qualitative and quantitative studies under light microscopy. This contribution adds to the basic techniques for the study of human nervous tissue with the same advantages described for the ""single-section"" Golgi method in other species; it is easy and fast, requires minimal equipment, and provides consistent results. (C) 2010 Elsevier B.V. All rights reserved.

Characterization of juvenile and young adult mice following induction of hydrocephalus with kaolin

Hydrocephalus is a common neurological problem in humans, Usually caused by an impairment of cerebrospinal fluid (CSF) flow or absorption. A reliable induced model of chronic hydrocephalus in mice would be useful to test hypotheses using genetic mutants. Our goal was to characterize behavioral and histological changes in juvenile and Young adult mice with kaolin (aluminum silicate) -induced hydrocephalus. Seven-day old and 7-8 week old mice received injection of kaolin into the cisterna magna. Behavior was assessed repeatedly. Seven or 14 days following kaolin, magnetic resonance (MR) imaging was used to assess ventricle size. In hydrocephalic mice, body weight was significantly lower than in age-matched saline-injected sham controls and the gait and posture score were impaired. Juvenile mice developed severe ventriculomegaly and had reduced corpus callosum thickness with gross white matter destruction by 14 days. Reactive astroglial change in white matter and cortex and reduced cellular proliferation in the subependymal zone were also apparent. Young adult mice developed only moderate ventricular enlargement without overt white matter destruction, although there was corpus callosum atrophy and mild astroglial reaction in white matter. Glial fibrillary acidic protein content was significantly higher in juvenile and young adult hydrocephalic mice at 7 and 14 days, but myelin basic protein content was not significantly altered. In conclusion, hydrocephalus induced by percutaneous injection of kaolin in juvenile and young adult mice is feasible. The associated periventricular alterations are essentially the same as those reported in rats of comparable ages. (C) 2009 Elsevier Inc. All rights reserved.

Neuromuscular activity of BaTX, a presynaptic basic PLA(2) isolated from Bothrops alternatus snake venom

We have previously isolated a Lys49 phospholipase A(2) homolog (BaTX) from Bothrops alternatus snake venom using a combination of molecular exclusion chromatography and reverse phase HPLC and shown its ability to cause neuromuscular blockade. In this work, we describe a one-step procedure for the purification of this toxin and provide further details of its neuromuscular activity. The toxin was purified by reverse phase HPLC and its purity and molecular mass were confirmed by SIDS-PAGE, MALDI-TOF mass spectrometry, amino acid analysis and N-terminal sequencing. BaTX (0.007-1.4 mu M) produced time-dependent, irreversible neuromuscular blockade in isolated mouse phrenic nerve-diaphragm and chick biventer cervicis preparations (time to 50% blockade with 0.35 mu M toxin: 58 +/- 4 and 24 +/- 1 min, respectively; n = 3-8; mean +/- S.E.) without significantly affecting the response to direct muscle stimulation. In chick preparations, contractures to exogenous acetylcholine (55 and 110 mu M) or KCl (13.4 mM) were unaltered after complete blockade by all toxin concentrations. These results, which strongly suggested a presynaptic mechanism of action for this toxin, were reinforced by (1) the inability of BaTX to interfere with the carbachol-induced depolarization of the resting membrane, (2) a significant decrease in the frequency and amplitude of miniature end-plate potentials, and (3) a significant reduction (59 +/- 4%, n=12) in the quantal content of the end-plate potentials after a 60 min incubation with the toxin (1.4 mu M). In addition, a decrease in the organ bath temperature from 37 degrees C to 24 degrees C and/or the replacement of calcium with strontium prevented the neuromuscular blockade, indicating a temperature-dependent effect possibly mediated by enzymatic activity. (C) 2009 Elsevier Inc. All rights reserved.

Distinct patterns of somatic alterations in a lymphoblastoid and a tumor genome derived from the same individual

Although patterns of somatic alterations have been reported for tumor genomes, little is known on how they compare with alterations present in non-tumor genomes. A comparison of the two would be crucial to better characterize the genetic alterations driving tumorigenesis. We sequenced the genomes of a lymphoblastoid (HCC1954BL) and a breast tumor (HCC1954) cell line derived from the same patient and compared the somatic alterations present in both. The lymphoblastoid genome presents a comparable number and similar spectrum of nucleotide substitutions to that found in the tumor genome. However, a significant difference in the ratio of non-synonymous to synonymous substitutions was observed between both genomes (P = 0.031). Protein-protein interaction analysis revealed that mutations in the tumor genome preferentially affect hub-genes (P = 0.0017) and are co-selected to present synergistic functions (P < 0.0001). KEGG analysis showed that in the tumor genome most mutated genes were organized into signaling pathways related to tumorigenesis. No such organization or synergy was observed in the lymphoblastoid genome. Our results indicate that endogenous mutagens and replication errors can generate the overall number of mutations required to drive tumorigenesis and that it is the combination rather than the frequency of mutations that is crucial to complete tumorigenic transformation.

Diabetes and Vascular Disease: Basic Concepts of Nitric Oxide Physiology, Endothelial Dysfunction, Oxidative Stress and Therapeutic Possibilities

The vascular manifestations associated with diabetes mellitus (DM) result from the dysfunction of several vascular physiology components mainly involving the endothelium, vascular smooth muscle and platelets. It is also known that hyperglycemia-induced oxidative stress plays a role in the development of this dysfunction. This review considers the basic physiology of the endothelium, especially related to the synthesis and function of nitric oxide. We also discuss the pathophysiology of vascular disease associated with DM. This includes the role of hyperglycemia in the induction of oxidative stress and the role of advanced glycation end-products. We also consider therapeutic strategies.

Kaposi sarcoma and paracoccidioidomycosis in the same fragment of oral mucosa biopsy: a rare association in human immunodeficiency virus-positive patient. A case report

The immunossuppression caused by HIV infection makes the affected individuals more susceptible to some diseases including infections, neoplasms, or even the association between them. Kaposi sarcoma (KS) is the most common AIDS-related neoplasm, featured as an angioproliferative disorder. Its cause seems to be related to the human herpesvirus type 8 and it is usually associated with lower CD4+ T cell count. Oral involvement is frequent, presenting red to blue-purplish plaques, maculaes, and nodules. On the other hand, paracoccidioidomycosis (PCM) is a systemic mycosis, endemic in Latin America, caused by Paracoccidioides brasiliensis. This mycosis is not commonly related to human immunodeficiency virus (HIV) infection, although PCM can be present in immunosuppression cases. Oral lesions, as granulomatous ulcers, are often identified in seropositive patients with PCM. A rare case, in which a male HIV-positive patient presented simultaneously Kaposi sarcoma and PCM in the same fragment of oral mucosa biopsy, is described. To the best of our knowledge, this concomitant association had not been previously described. (C) 2011 Elsevier Inc. All rights reserved.

Influence of pH on slow crack growth of dental porcelains

Objectives. To evaluate the effect of pH of storage medium on slow crack growth (SCG) parameters of dental porcelains. Methods. Two porcelains were selected: with (UD) and without (VM7) leucite particles, in order to assess if the microstructure would affect the response of the material to the pH variation. Disc specimens were produced following manufacturers` instructions. Specimens were stored in artificial saliva in pHs 3.5, 7.0 or 10.0 for 10 days and after that the fatigue parameters (n: SCG susceptibility coefficient and sigma(0): scaling parameter) were obtained by the dynamic fatigue test using the same pH of storage. Microstructural analysis of the materials was also performed. Results. For VM7, the values of n obtained in the different pHs were similar and varied from 29.9 to 31.2. The sigma(0) value obtained in pH 7.0 for VM7 was higher than that obtained in the other pHs, which were similar. For porcelain UD, n values obtained in pHs 7.0 and 10.0 were similar (40.8 and 39.6, respectively), and higher than that obtained in pH 3.5 (26.5). With respect to sigma(0), the value obtained for porcelain UD in pH 10.0 was lower than those obtained in pHs 3.5 and 7.0, which were similar. Significance. The effect of pH on the stress corrosion susceptibility (n) depended on the porcelain studied. While the n value of VM7 was not affected by the pH, UD presented lower n value in acid pH. For both porcelains, storage in acid or basic pH resulted in strength degradation. (C) 2007 Academy of Dental Materials. Published by Elsevier Ltd. All rights reserved.

Effect of sodium, amine and stannous fluoride at the same concentration and different pH on in vitro erosion

Objective: This study aimed to compare the effects 0.5% and 1% sodium, amine and stannous fluoride at different pH on enamel erosion in vitro. Methods: Bovine enamel samples were submitted to a cyclic de- and remineralisation for 3 days. Each day, the samples were exposed for 120 min to pooled human saliva and subsequently treated with one of the fluoride solutions for 3 min: amine fluoride (AmF, 0.5% and 1% F(-)), sodium fluoride (NaF, 0.5% and 1% F(-)), each at pH 3.9 and 7.0, and stannous fluoride (SnF(2), 0.5% and 1% F-), at pH: 3.9. Additionally, two groups were treated with fluoride-free placebo solutions (pH: 3.9 and 7.0) and one group served as control (no fluoridation). Ten specimens each group were inserted in a so-called artificial mouth and eroded six times daily with hydrochloric acid (pH 2.6) for 90 s each intermitted by exposure to artificial saliva (1 h). After 3 days, enamel loss was analyzed profilometrically and evaluated statistically by ANOVA. Results: Only the acidic 0.5% and 1% SnF(2) and 1% AmF solutions were able to reduce erosive enamel loss significantly, while all other solutions and placebos did not differ significantly from the control. Between the acidic SnF(2) and the 1% AmF solutions no significant differences could be detected. Conclusion: At the same concentrations, acidic SnF(2) and AmF may be more effective than NaF to protect enamel against erosion. (C) 2009 Elsevier Ltd. All rights reserved.