A micro-anatomical model of the distribution of myocardial endomysial collagen.

Myocardial connective tissue probably provides passive support for regulating heart tensile strength and stiffness and ultimately for controlling heart mechanics through its endomysial part. However, endomysial collagen micro-arrangement is still a matter of debate. In order to define the fine distribution of left ventricle endomysial collagen, we applied the NaOH-scanning electron microscopy (SEM) maceration method (one of the techniques of choice for studying collagen micro-arrangement) to rabbit heart. Gomori-reticulum staining was used for correlated light microscopy (LM) observations. The SEM-NaOH method allowed isolation of collagen by removing other extracellular matrix components and cells and preserved collagen structure and position. Endomysial collagen appeared arranged in laminae that delimited the lacunae that were left empty by macerated myocytes and small vessels (mostly capillaries). These laminae were formed by reticular fibers, as confirmed by LM observations of Gomorireticulum-stained samples, and were organized in irregularly meshed networks made of thin (single) and thick (composed) filaments. In longitudinal views, collagen laminae extended the entire length of lacunae. In transversal views, the cut surface of the laminae appeared to be made of collagen bundles. These observations provide an updated microanatomical view of endomysial collagen distribution, which integrates previous studies. This model is based on the evidence that collagen laminae enveloped the surface of small vessels and myocytes. Thus, a type of myocyte-myocyte or capillary-myocyte "laminar connection" anchored to the entire cell length here is emphasized, rather than a type of "strut connection" anchored to defined loci, as usually described. This structure explains better how endomysium may provide the necessary support for heart compliance and protection against overstretch.

The "human"-like morphology of the swine papilla ilealis.

The morphology of the ileocecocolic transition, between the small and large intestine was studied in 33 male, adult pigs of unknown breed. The results were analyzed by the Wilcoxon Test (alpha < or = 0.05). The ileum opened into the large intestine, in all of the cases, at the level of the cecocolic junction in the form of a conical papilla (ileal papilla) turned towards the large intestine, with an oblique caudal and lateral direction, pointing to the cecal lumen. The cranial contour is higher than the caudal one, with average values of 3.0 cm, and 1.5 cm in the fresh specimens and 2.8 cm and 1.4 cm after fixation of the specimens. The average diameter of the papilla apex was 1.5 cm, both in the fresh specimens and in the fixed ones. In 100% of the cases the ileal papilla appeared with a star-shaped orifice in its apex in both the fresh and in the fixed specimens. Our studies showed that the transition between the small and large intestine of the swine is ileocecocolic, thus similar to humans and different from the ileocecal (Perissodactyla, Rodentia, Lagomorpha) and, according to most descriptions, ileocolic (dog) transitions found in other animals.

Arrangement of the tunica muscularis in the ileal papilla of swine.

The arrangement of the bundles of muscular fibers in the ileal papilla was studied in 32 adult male crossbred swine by dissection of the muscular bundles in this region, after immersion in 50% nitric acid, and by observation under the stereomicroscope of material preserved in a 10% formaldehyde solution. In the ileal papilla, the cranial (ileocolic) contour of the tunica muscularis was formed by the circular strata of the ascending colon and the ileum, and the caudal (ileocecal) contour by the circular strata of the cecum and the ileum. The longitudinal stratum was located between the circular strata with predominance of muscular bundles originating in the terminal ileum. The tunica muscularis showed an increased thickness at the base of the eminence and at the free border, not reaching the apex. In the ileal papilla, the circular and longitudinal components characterize a "pylorus", the ileocecocolic or ileal pylorus as a transit regulation device for the flow of contents between the small and large intestine of swine.

Arrangement of the tunica muscularis of the ileum, cecum and colon at the transition between the small and the large intestines of swine.

The arrangement of the bundles of muscular fibers in the transition between the small and the large intestines was studied in 12 male adult crossbred swine by dissection, after immersion in 50% nitric acid solution. The connection between the small and the large intestines was formed by the association of the muscular tissue, the connective tissue and the fat tissue. The tunica muscularis of these organs was organized in thin thread-like bundles and wide ribbon-like bundles of variable width and thickness. The superficial bundles of the longitudinal stratum of the tunica muscularis of the ileum established the continuity with the large intestine; the deep bundles penetrated into the ileal papilla. The limit between the cecum and the ascending colon was externally marked by the sulcus cecocolicus dorsalis and ventralis. The connection between the cecum and the ascending colon was formed by bundles of muscular fibers coming from the ileum, and the taeniae ventralis, lateralis and medialis of the cecum. Some bundles of muscular fibers from the ascending colon and the cecum headed toward the sulcus cecocolicus. The median bundles of muscular fibers of the taenia ventralis of the cecum, near the termination of the ileum, were arranged to form a loop around the termination of the ileum, mixing with the musculature of the ascending colon at the level of this junction.

Morphology of the swine ileum terminale.

The morphology of the terminal ileum was studied in 33 male adult pigs of unknown breed. The ileum ended in the cecocolic junction at an acute angle to the cecum. The terminal ileum displayed a cylindrical form (63.6% +/- 8.4 of the cases) more frequently than an ampullary (21.2% +/- 7.1) and/or an infundibular form (15.2% +/- 6.2). The ampullary form is not related to the quadrupedal position of the animal, but is related to the functional phase of the intestine at the time of death. In the mucous tunica of the terminal ileum, longitudinal folds and aggregated lymphoid nodules appeared concentrated along the antimesenteric border.

Three-dimensional distribution of the collagen fibers in the submucosa of the swine terminal ileum.

Collagen has an important role in controlling mechanical function and physiopathology of intestinal wall. Swine small intestine may be used as biomaterial source for tissue repairing. Changes of collagen arrangement and three-dimensional (3D) distribution may be related to the dissimilar biomechanical proprieties showed by different intestine tracts. 3D spatial distribution of collagen bundles of swine submucosal terminal ileum (SSTI) was studied by a correlated analysis of light (LM) and scanning electron microscopy (SEM) of NaOH macerated samples. Bundles of collagen fibers were greatly represented in the submucosa at the mesenteric border and also extended along the longitudinal folds beneath mucosa layer. Polarized LM of picrosirius stained samples evidenced yellow and red fibers (type I collagen), and green fibers (type III collagen). Silver-impregnated sections showed predominant brown-stained fibers and, in a smaller amount, black-stained ones. By SEM submucosal collagen, isolated by NaOH maceration, appeared arranged in wide bundles forming a complicated 3-D network. The bundles presented a sinuous course, opened and closed repeatedly forming meshes fashioned in a regular net. These observations originally demonstrated that 3-D distribution of SSTI collagen is different from that observed in other gut segments and species. The arrangement of SSTI collagen fibers that we observed seems to be morphofunctionally adjusted to provide appropriate resistance to mechanical forces and to assure compliance to deformations induced by intestinal wall motion. The studies for selection of optimal intestinal patches for surgical replacement should take into consideration the basic morphological evaluation of parietal collagen 3D distribution.

Flufenamic acid as an inducer of mitochondrial permeability transition.

To assess the mechanism by which mitochondrial permeability transition (MPT) is induced by the nonpolar carboxylic acids, we investigated the effects of flufenamic acid (3'-trifluoromethyl diphenylamine-2-carboxylic acid, FA) on mitochondrial respiration, electrical transmembrane potential difference (delta psi), osmotic swelling, Ca2+ efflux, NAD(P)H oxidation and reactive oxygen species (ROS) generation. Succinate-energized isolated rat liver mitochondria incubated in the absence or presence of 10 microM Ca2+, 5 microM ruthenium red (RR) or 1 microM cyclosporin A (CsA) were used. The dose response-curves for both respiration release and delta psi dissipation were nearly linear, presenting an IC50 of approximately 10 microM and reaching saturation within 25-50 microM, indicating that FA causes mitochondrial uncoupling by a protonophoric mechanism. Within this same concentration range FA showed the ability to induce MPT in energized mitochondria incubated with 10 microM Ca2+, followed by delta psi dissipation and Ca2+ efflux, and even in deenergized mitochondria incubated with 0.5 mM Ca2+. ADP, Mg2+, trifluoperazine (TFP) and N-ethylmaleimide (NEM) reduced the extent of FA-promoted swelling in energized mitochondria by approximately one half, whereas dithiothreitol (DTT) slightly enhanced it. NAD(P)H oxidation and ROS generation (H2O2 production) by mitochondria were markedly stimulated by FA; these responses were partly prevented by CsA, suggesting that they may be implicated as both a cause and effect of FA-induced MPT. FA incubated with mitochondria under swelling assay conditions caused a decrease of approximately 40% in the content of protein thiol groups reacting with 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB). The present results are consistent with a ROS-intermediated sensitization of MPT by a direct or indirect FA interaction with inner mitochondrial membrane at a site which is in equilibrium with the NAD(P)H pool, namely thiol groups of integral membrane proteins.