Evaluation of the Integrity of Horse Hoof Dermal and Epidermal Tissues Collected by Dorsal Transmural Access

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Effects of isocoumarins isolated from Paepalanthus bromelioides on mitochondria: Uncoupling, and induction/inhibition of mitochondrial permeability transition

Isolated mitochondria may undergo uncoupling, and in presence of Ca2+ at different conditions, a mitochondrial permeability transition (MPT) linked to protein,thiol oxidation, and demonstrated by CsA-sensitive mitochondrial swelling; these processes may cause cell death either by necrosis or by apoptosis. Isocoumarins isolated from the Brazilian plant Paepalanthus bromelioides (Eriocaulaceae) paepalantine (9,10-dihydroxy-5,7-dimethoxy-1H-naptho(2,3c)pyran-1-one), 8,8'-paepalantine dimer, and vioxanthin were assayed at 1-50 mu M on isolated rat liver mitochondria, for respiration, MPT, protein thiol oxidation, and interaction with the mitochondrial membrane using 1,6-diphenyl-1,3,5-hexatriene (DPH). The isocoumarins did not significantly affect state 3 respiration of succinate-energized mitochondria; they did however, stimulate 4 respiration, indicating mitochondrial uncoupling. Induction of MPT and protein thiol oxidation were assessed in succinate-energized mitochondria exposed to 10 mu M Ca2+; inhibition of these processes was assessed in non-energized organelles in the presence of 300 mu M t-butyl hydroperoxide plus 500 mu M Ca2+. Only paepalantine was an effective MPT/protein thiol oxidation inducer, also releasing cytochrome c from mitochondria; the protein thiol oxidation, unlike mitochondrial swelling, was neither inhibited by CsA nor dependent on the presence of Ca2+. Vioxanthin was an effective inhibitor of MPT/protein thiol oxidation. All isocoumarins inserted deeply into the mitochondrial membrane, but only paepalantine dimer and vioxantin decreased the membrane's fluidity. A direct reaction with mitochondrial membrane protein thiols, involving an oxidation of these groups, is proposed to account for MPT induction by paepalantine, while a restriction of oxidation of these same thiol groups imposed by the decrease of membrane fluidity, is proposed to account for MPT inhibition by vioxanthin. (c) 2006 Published by Elsevier B.V..

Correlation between light transmission and permeability of human dentin

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Assessment of apoptosis in epidermal lamellar cells in clinically normal horses and those with laminitis

Objective - To determine and compare the number, type, location, and distribution of apoptotic epidermal cells in the laminae of clinically normal horses and horses with laminitis.Sample Population - Formalin-fixed samples of digital lamellar tissue from 47 horses (including clinically normal horses [controls; n = 7], horses with acurte [4] and chronic [7] naturally acquired laminitis, and horses with black walnut extract-induced [11] or carbohydrate overload-induced [18] laminitis).Procedure - Blocks of paraffin-embedded lamellar tissues were stained for DNA fragmentation with the terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) technique. Differential immunohistochemical staining for caspases 3 and 14 were used to confirm apoptosis.Results - the number of TUNEL-positive epidermal cells per 0.1 mm of primary laminae was significantly greater in the acute laminitis group than in the other groups. In the acute laminitis group, there were 17 and 1,025 times as many TUN EL-positive basal layer cells and keratinocytes, respectively, compared with the control group. Apoptosis of TUNEL-positive basal layer cells was confirmed by results of caspase 3 immunohistochemical staining. The TUNEL-positive keratinocytes did not stain for caspases 3 or 14.Conclusions and Clinical Relevance - the large number of apoptotic basal layer cells detected in the lamellar tissue of horses with acute naturally acquired laminitis suggests that apoptosis may be important in the development of acute laminitis. The role of the large number of TUNEL-positive keratinocytes detected in the interface of primary and secondary epidermal laminae of horses with acute laminitis remains to be elucidated.

EPIDERMAL GROWTH-FACTOR AND TRANSFORMING GROWTH-FACTOR-ALPHA CHARACTERISTICS OF HUMAN ORAL-CARCINOMA CELL-LINES

This study examined the expression of epidermal growth factor (EGF) cell-surface receptors, the response to exogenous ligand and the autocrine production of transforming growth factor a (TGF-a) in normal and carcinoma-derived human oral keratinocytes. One of eight malignant cell lines overexpressed EGF receptors, while the remainder expressed receptor numbers similar to normal cells. Exogenous EGF stimulated incorporation of tritiated thymidine in a dose-dependent manner. In keratinocytes expressing normal numbers of EGF receptors, the cellular response to exogenous EGF correlated positively with total EGF receptor number. SCC-derived keratinocytes produced more TGF-a than normal cells. There was no statistical correlation between the autocrine production of TGF-a, EGF cell-surface receptor expression and cellular response to exogenous EGF. While the growth-stimulatory effects of exogenous TGF-cl were inhibited by the addition of a neutralising antibody, the presence of this antibody in conditioned medium failed to produce a similar decrease in growth. The results indicate that overexpression of EGF receptors is not an invariable characteristic of human oral squamous carcinoma-derived cell lines. Further, the contribution of TGF-a to the growth of normal and carcinoma-derived human oral keratinocytes in vitro may be less significant than previously documented.

Morphological characterization of the venom secretory epidermal cells in the stinger of marine and freshwater stingrays

Marine and freshwater stingrays are characterized by the presence of one to three mineralized serrated stingers on the tail, which are covered by epidermal cells secreting venom. When these animals are dorsally touched, the stinger can be introduced into the aggressor by a whip reflex mechanism of the tail, causing severe mechanical injuries and inoculating the venom. Accidents in humans are frequent causing intense local pain, oedema and erythema. Bacterial secondary infection is also common. In addition, injuries involving freshwater stingrays frequently cause a persistent cutaneous necrosis. The exact localization of the venom secretory epidermal cells in the stinger is controversial, but it is known that it is preferentially located in the ventrolateral grooves. A comparative morphological analysis of the stinger epidermal tissue of different marine and freshwater Brazilian stingray species was carried out. The results indicate that in freshwater species there is a larger number of protein secretory cells, of two different types, spread over the whole stinger epidermis, while in marine species the protein secretory cells are located only around or inside the stinger ventrolateral grooves. These differences between the stingers of the two groups can justify the more severe envenomation accidents with the freshwater species when compared with the marine species. (c) 2007 Elsevier Ltd. All rights reserved.

AMINO-ACID-SEQUENCE OF TSTX-V, AN ALPHA-TOXIN FROM TITYUS-SERRULATUS SCORPION-VENOM, AND ITS EFFECT ON K+ PERMEABILITY OF BETA-CELLS FROM ISOLATED RAT ISLETS OF LANGERHANS

Highly purified Tityustoxin V (TsTX-V), an alpha-toxin isolated from the venom of the Brazilian scorpion Tityus serrulatus, was obtained by ion exchange chromatography on carboxymethylcellulose-52. It was shown to be homogeneous by reverse phase high performance liquid chromatography, N-terminal sequencing (first 39 residues) of the reduced and alkylated protein and by polyacrylamide gel electrophoresis in the presence of sodium dodecylsulfate and tricine. Following enzymatic digestion, the complete amino acid sequence (64 residues) was determined. The sequence showed higher homology with the toxins from the venoms of the North African than with those of the North and South American scorpions. Using the rate of Rb-86(+) release from depolarized rat pancreatic beta-cells as a measure of K+ permeability changes, TsTX-V (5.6 mu g/ml) was found to increase by 2.0-2.4-fold the rate of marker outflow in the presence of 8.3 mM glucose. This effect was persistent and slowly reversible, showing similarity to that induced by 100 mu-M veratridine, an agent that increases the open period of Na+ channels, delaying their inactivation. It is suggested that, by extending the depolarized period, TsTX-V indirectly affects beta-cell voltage-dependent K+ channels, thus increasing K+ permeability.