Qualitative and Quantitative Study of the Changes in the Ultrastructure of Mammalian Adrenal Cortex Caused by the Venezuelan Tigra Mariposa (Bothrops venezuelensis) Snake Venom.

The damage of the adrenal gland by snake venoms needs to be clarified. Lethality (LD50) of Bothrops venezuelensis (Bv) venom was established by intraperitoneally mice injections. Preparation of specimens for transmission electron microscopy samples from cortex adrenal gland biopsies at 3, 6, and 24 h was processed. The quantitative description by the principal component analysis (PCA) of the adrenal gland was as follows: thickening of the capillary endothelium, area of the capillary lumen, cell nucleus area, enlargement of the perinuclear space, number of mitochondria, area of the mitochondria, number of mitochondrial cristae, number of cristae per mitochondrial unit, and tubular diameter of the smooth endoplasmic reticulum (SER). Sections of the adrenal cortex, after 3 h postinjection with Bv venom showed in the cortical cells: mitochondria with tubular cristae and slightly swollen SER cisternae, nucleus with variable heterochromatin content, irregular edges, and swollen nuclear envelope. After 6 h, cells with swollen nucleus envelope, electron dense lipids and mitochondria with loss of their cristae were observed. Myelin figures, close to the microvilli of the cortical cell, multivesicular bodies, swollen profiles of the SER, and electron dense lipid drops were noticed. After 24 h, thickening of the endothelial wall, fenestrae and projections into the capillary lumen, loss of the mitochondrial cristae, destruction of the capillary and the plasma membrane of the cortical cell, multivesicular body, SER loss, and an enlargement of the perinuclear space were detected. In the quantitative PCA, there were significant changes after the venom treatments.

Biological activities of a new crotamine-like peptide from Crotalus oreganus helleri on C2C12 and CHO cell lines, and ultrastructural changes on motor endplate and striated muscle.

Crotamine and crotamine-like peptides are non-enzymatic polypeptides, belonging to the family of myotoxins, which are found in high concentration in the venom of the Crotalus genus. Helleramine was isolated and purified from the venom of the Southern Pacific rattlesnake, Crotalus oreganus helleri. This peptide had a similar, but unique, identity to crotamine and crotamine-like proteins isolated from other rattlesnakes species. The variability of crotamine-like protein amino acid sequences may allow different toxic effects on biological targets or optimize the action against the same target of different prey. Helleramine was capable of increasing intracellular Ca2+ in Chinese Hamster Ovary (CHO) cell line. It inhibited cell migration as well as cell viability (IC50 = 11.44 µM) of C2C12, immortalized skeletal myoblasts, in a concentration dependent manner, and promoted early apoptosis and cell death under our experimental conditions. Skeletal muscle harvested from mice 24 h after helleramine injection showed contracted myofibrils and profound vacuolization that enlarged the subsarcolemmal space, along with loss of plasmatic and basal membrane integrity. The effects of helleramine provide further insights and evidence of myotoxic activities of crotamine-like peptides and their possible role in crotalid envenomings.

Qualitative and quantitative ultrastructural analysis of the mitochondria from adrenal gland cortex under the action of viperidae family snake venoms / Análisis ultrastructural cualitativo y cuantitativo de la mitocondria de la glándula adrenal bajo la acción de los venenos de serpientes de la familia viperidae

SUMMARY: The Viperidae venoms are composed of a mixture of constituents with enzymatic and non-enzymatic actions, which act on ultrastructural components of cells and tissues. Here, the number of mitochondria, mitochondrial area and the number of mitochondrial cristae from adrenal glands cortex treated with snake venoms were tested after 3, 6 and 24 hours of venom injections. The mitochondria quantitative changes showed a statistically significant decrease, in the number of mitochondria past 3, 6 and 24 h. There was an increase in the mitochondrial area after 6 h, where Crotalus vegrandis venom did not present significant differences with Crotalus pifanorum or Bothrops venezuelensis venoms. After 24 h, there was an escalation of mitochondrial area in all tested venoms. The number of mitochondrial cristae after 3 h did not present important differences with the control treatment. After 6 h, the number of mitochondrial cristae initiated to decrease under the activities of the 3 venoms action, until 24 h of observation. In the qualitative observations it was possible to witness an intense damage of the mitochondria, with loss and swelling of membranes, disappearance of cristae and the appearance of myelin figures, which started at 3 h after the Crotalus and Bothrops venoms injections. These damages probably were due to cytotoxic effects of phospholipases, metalloproteases and/or other proteolytic activities present in Viperidae snake venoms, being more evident in Crotalus venoms. As far as we know, these results define a novel finding that suggest that Viperidae snake venoms are extremely toxic to mammalian mitochondria.

RESUMEN: Los venenos de Viperidae tienen acciones enzimáticas y no enzimáticas, que actúan sobre la estructura celular. Aquí se probaron, a las 3, 6 y 24 horas de la inyección del veneno, el número de mitocondrias, el área mitocondrial y el número de crestas mitocondriales de la corteza de las glándulas adrenales. Los cambios cuantitativos de las mitocondrias mostraron una disminución en el número de mitocondrias a las 3, 6 y 24 h. Hubo un aumento en el área mitocondrial a las 6 h, donde el veneno de la serpiente Crotalus vegrandis no presentó diferencias significativas con los venenos de Crotalus pifanorum o Bothrops venezuelensis. Después de 24 h, hubo un aumento del área mitocondrial en todos los venenos. El número de crestas mitocondriales a las 3 h no presentó alteraciones o diferencias importantes con el tratamiento de control. Después de 6 h, el número de crestas mitocondriales comenzó a disminuir bajo la acción de los 3 venenos, hasta las 24 h de observación. En las observaciones cualitativas se observó un daño intenso de las mitocondrias, con pérdida y edema de las membranas, desaparición de las cristae y aparición de figuras mielínicas, que comenzó a las 3 h después de las inyecciones de veneno de Crotalus y Bothrops. Estos daños se debieron factiblemente a los efectos citotóxicos de componentes proteolíticos de los venenos. Creemos que estos resultados definen un nuevo y original hallazgo, que sugiere que los venenos de serpiente Viperidae son extremadamente tóxicos para las mitocondrias de mamíferos.

Crotamine-like from Southern Pacific rattlesnake (Crotalus oreganus helleri) Venom acts on human leukemia (K-562) cell lines and produces ultrastructural changes on mice adrenal gland.

Crotamine is a cationic, non-enzymatic, protein integrating a minor family of myotoxins, composed of 42 amino acid residues, described in Viperidae and Crotalidae snake's families that has been used in neuroscience research, drug progressing and molecular diversity reports. Crotamine-like protein (CLP) from C.o.helleri venom was isolated in fraction 5 from 7 peaks obtained by sulfopropyl waters protein pak cationic exchange column. In tricine-SDS-PAGE under non-reduced conditions this CLP showed a single band of ~8 kDa molecular weight. CLP induced toxicity of K-562 cells with a CC50 of 11.09 µM. In mice adrenal gland after 24 h of CLP injection, cortical cells exhibited swollen mitochondria with scarce tubular cristae, some elements of smooth and rough endoplasmic reticula, widened nuclear envelope, slightly osmiophilic lipid droplets, and autophagic vacuoles. In some areas cortical cells plasma membrane and endothelial walls disappeared, which indicated a necrosis process. In other areas, endothelial cell cytoplasm did not present the normal caveolae and pinocytotic vesicles. To our knowledge, this is the first report on mice adrenal gland damages, caused by the injection of CLP from rattlesnakes. Our results propose that adrenal cortex lesions may be significant in the envenoming etiopathogenesis by CLP.

Ultrastructure of colorectal adenocarcinoma and peritumoral tissue in untreated patients.

In this study, we describe, compare, and discuss several subcellular alterations found in Colorectal Adenocarcinoma and peritumoral tissue using transmission electron microscopy, morphometry, and statistical analysis. Tissue samples from anterior resections were collected from patients diagnosed with Colorectal Adenocarcinoma in the University Hospital of Caracas. Samples were processed according to the typical protocol for their observation through transmission electron microscopy. The resulting images were analyzed using specialized software for the collection of morphometric data. Several anomalies were common for both tissues, including but not limited to, rough endoplasmic reticulum and mitochondrial swelling, nuclear invagination, nuclear enlargement, and cellular swelling. In general, alterations within the tumor were more frequent and intense. Extensive organellar degradation and other evidences of cellular damage seemed to extend past the edge of the tumor into the peritumoral tissue. There seems to be a clear process of lateral cancerization present in the peritumoral area. The tissue layers composed of smooth muscle cells, probably due to their structural features, may allow greater diffusion of harmful substances produced by the tumor. A more in-depth analysis of peritumoral tissue considering organellar damage and morphometric data may provide relevant insight about the changing microenvironment promoted by the close proximity of a tumor.

Trypanosoma vivax Adhesion to Red Blood Cells in Experimentally Infected Sheep.

Trypanosomosis, a globally occurring parasitic disease, poses as a major obstacle to livestock production in tropical and subtropical regions resulting in tangible economic losses. In Latin America including Venezuela, trypanosomosis of ruminants is mainly caused by Trypanosoma vivax. Biologically active substances produced from trypanosomes, as well as host-trypanosome cellular interactions, contribute to the pathogenesis of anemia in an infection. The aim of this study was to examine with a scanning electron microscope the cellular interactions and alterations in ovine red blood cells (RBC) experimentally infected with T. vivax. Ovine infection resulted in changes of RBC shape as well as the formation of surface holes or vesicles. A frequent observation was the adhesion to the ovine RBC by the trypanosome's free flagellum, cell body, or attached flagellum in a process mediated by the filopodia emission from the trypanosome surface. The observed RBC alterations are caused by mechanical and biochemical damage from host-parasite interactions occurring in the bloodstream. The altered erythrocytes are prone to mononuclear phagocytic removal contributing to the hematocrit decrease during infection.

Myopathy in patients with Hashimoto's disease.

Hashimoto thyroiditis (HT) is an autoimmune disease of the thyroid gland. Patients may present or not a hypothyroid state, and frequently have manifestations of myopathy. The present work was aimed to assess the clinical symptoms and signs of skeletal muscle alterations in HT, describe the muscular pathological changes and relate them to the functional thyroid status and to the autoimmune condition of the patient. Clinical and laboratory studies were performed in ten HT patients and three control subjects (hormonal levels and electromyography). Biopsies from their vastus lateralis of quadriceps femoris muscle were analyzed under light (histochemistry and immunofluorescense) and electron microscopy. All patients showed muscle focal alterations, ranging from moderate to severe atrophy, necrosis, activation of satellite cells, presence of autophagosomes, capillary alterations and macrophage and mast cell infiltration, common to autoimmune diseases. The intensity of clinical signs and symptoms was not related to the morphological muscle findings, the electromyography results, or to the state of the thyroid function. Reactions for immunoglobulin in muscle fibers were positive in 80% of the patients. Fiber type II proportion was increased in all patients, with the exception of those treated with L-thyroxine. In conclusion, autoimmune processes in several of the patients may be associated to the skeletal muscle alterations, independently of the functional state of the thyroid gland; however, fiber II type proportion could have been normalized by L-thyroxine treatment.

Myopathy in patients with Hashimoto´s Disease. / Miopatía en pacientes con enfermedad de Hashimoto

Hashimoto thyroiditis (HT) is an autoimmune disease of the thyroid gland. Patients may present or not a hypothyroid state, and frequently have manifestations of myopathy. The present work was aimed to assess the clinical symptoms and signs of skeletal muscle alterations in HT, describe the muscular pathological changes and relate them to the functional thyroid status and to the autoimmune condition of the patient. Clinical and laboratory studies were performed in ten HT patients and three control subjects (hormonal levels and electromyography). Biopsies from their vastus lateralis of quadriceps femoris muscle were analyzed under light (histochemistry and immunofluorescense) and electron microscopy. All patients showed muscle focal alterations, ranging from moderate to severe atrophy, necrosis, activation of satellite cells, presence of autophagosomes, capillary alterations and macrophage and mast cell infiltration, common to autoimmune diseases. The intensity of clinical signs and symptoms was not related to the morphological muscle findings, the electromyography results, or to the state of the thyroid function. Reactions for immunoglobulin in muscle fibers were positive in 80% of the patients. Fiber type II proportion was increased in all patients, with the exception of those treated with L-thyroxine. In conclusion, autoimmune processes in several of the patients may be associated to the skeletal muscle alterations, independently of the functional state of the thyroid gland; however, fiber II type proportion could have been normalized by L-thyroxine treatment.

La tiroiditis de Hashimoto (TH) es una enfermedad autoinmune de la glándula tiroides. Los pacientes pueden tener o no un estado hipotiroideo y suelen presentar manifestaciones de miopatía. Este trabajo estudia los síntomas y signos clínicos de alteración muscular esquelética que puedan estar presentes en pacientes con TH, describe los cambios patológicos musculares y los relaciona con el estado funcional de la glándula tiroides y la condición autoinmune del paciente. Diez pacientes y tres sujetos controles fueron examinados clínicamente, se midieron los niveles de hormonas tiroideas, se practicó electromiografía y se tomó biopsia del vasto lateral del músculo cuádriceps crural para microscopía de luz (histoquímica e inmunofluorescencia) y microscopía electrónica. Todos los pacientes mostraron alteraciones musculares focales, atrofia moderada a severa, presencia de autofagosomas (glucogenosomas), necrosis, activación de las células satélites, infiltración de macrófagos y mastocitos, así como alteraciones en los capilares, similares a las de las enfermedades autoinmunes. La intensidad de los signos y síntomas no estuvo relacionada con los hallazgos morfológicos en músculo, los resultados de la electromiografía ni con el estado funcional tiroideo. La reacción a las inmunoglobulinas fue positiva en el músculo de 80% de los pacientes. La proporción de fibras musculares tipo II estuvo incrementada en los pacientes excepto en aquellos que recibieron tratamiento con L-tiroxina. En conclusión, el proceso autoinmune hacia el músculo parece asociarse a las alteraciones en éste, independientemente del estado funcional tiroideo, sin embargo, la proporción de las fibras tipo II puede haber sido normalizada por el tratamiento con L-tiroxina.

Ultrastructural study on tissue alterations caused by trypanosomatids in experimental murine infections.

The ultrastructural study in different tissues of mice experimentally infected with isolates of Trypanosoma evansi, Trypanosoma cruzi, and Leishmania mexicana reveals changes in cardiac myocytes, skeletal muscle fibers, and hepatic, adrenal, kidney, and spleen cells. Some of these changes were cytoarchitectural and others consisted of necrosis. Alterations in the microvasculature were also found. The mononuclear cell infiltrate included neutrophils, eosinophils, and macrophages. This work shows that diverse mice tissues are important target for trypanosomatids.

Características histológicas del epidídimo de Baba (Caiman crocodilus crocodilus) sexualmente maduro / Histological characteristics of the epididymis of the sexually mature spectacled Caiman (Caiman crocodilus crocodilus)

El epidídimo es un órgano que forma parte del aparato reproductivo masculino en todas las especies vertebradas, y que posee características histológicas que permiten lograr su identificación. Se realizó el estudio histológico del órgano reproductor de las babas Caiman crocodilus crocodilus en diferentes meses del año, incluyendo las épocas de gran actividad sexual o apareamiento y la época de no apareamiento, para conocer si las características histológicas de este órgano son similares durante todo el año. Se extrajeron 11 ejemplares sexualmente maduros de su medio ambiente natural, obteniéndose por necropsia, las muestras del epidídimo. Las muestras se fijaron con formol buferado al 10%, y se incluyeron en parafina. Posteriormente, se realizaron cortes de 5µm de grosor y fueron teñidos con Hematoxilina - Eosina. Como característica histológica común durante todo el año, se observó la presencia de unos conductillos epididimarios que confluyen en un conducto principal o conducto del epidídimo con un epitelio que varió de cúbico simple en la porción anterior a cilíndrico simple en los segmentos medio y posterior. De igual manera, se apreció un aumento de la capa muscular a medida que se avanzaba hacia las porciones media y posterior. En época de apareamiento resalta la característica del epitelio visiblemente festoneado, el aumento en la altura del mismo, la presencia de un gran número de células claras sin estereocilios y el aumento del diámetro de los conductos, así como gránulos de secreción eosinofílicos y un gran número de espermatozoides en el lumen, detalle que no es observado en la época de no apareamiento, durante la cual no se observa el epitelio de forma festoneado y su diámetro es mucho menor.

The epididymis is an organ which forms part of the male reproductive tract in all vertebrate species, and it possesses histological characteristics which allow its identification. A study was conducted to verify if the histological characteristics of the epididymis of the spectacled caiman (Caiman crocodilus crocodilus) are similar during different seasons of the year, including seasons of high and low mating activity. A total of 11 mature animals were selected from their natural environment. Samples of the epididymis were obtained from necropsy, fixed with 10% buffered formaldehyde and embedded in paraffin. Subsequently, they were stained with hematoxilin-eosin. The results show a common histological characteristic: the presence of some epididymis tiny ducts that converge into a main duct or the epididymis duct, with an epithelium which varies from a simple cubic in the anterior portion to a cylindrical one in the medial and posterior segments. In a similar way, as one moves forward to the medial and posterior portions, an increase in the muscular layer is observed. During mating time, the visibly festooning of the epithelium, the increase in height, the presence of a large amount of clear cells without stereocilium, an increase in the diameter of the ducts, as well as eosinophillic secretion granules and a great quantity of sperms in the lumen, stand out, a detail which is not seen during non-mating season, where the presence of a festooned epithelium is not observed, and its diameter is much smaller.

Activities against hemostatic proteins and adrenal gland ultrastructural changes caused by the brown widow spider Latrodectus geometricus (Araneae: Theridiidae) venom.

Brown widow spider (BrWS) (Latrodectus geometricus) venom produces intense systemic reactions such as cramps, harsh muscle nociceptive, nauseas, vomiting and hypertension. The proposed pathogenic mechanisms resulting in these accidents have principally been damages occurring at the nervous system. However, it is suspected that there is also damage of the adrenal glands, as a result of the experimental animal's clinical manifestations, which developed symptoms compatible with acute adrenal insufficiency. We have currently found that the adrenal gland is damaged by this venom gland homogenates (VGH) producing severe alterations on cortex cells resulting in death by acute adrenal insufficiency. In general, the ultrastructural study on the glands of mice under transmission electronic microscopy observations showed alterations in the majority of the intracellular membranes within 3 to 24h. BrWSVGH also showed specific actions on extracellular matrix proteins such as fibronectin, laminin and fibrinogen. In addition, zymogram experiments using gelatin as substrates detected gelatinolytic activity. The molecular exclusion fractionation of crude BrWSVGH resulted in 15 fractions, of which F1 and F2 presented alpha/beta-fibrinogenase and fibronectinolytic activities. Fractions F6, F14 and F15 showed only alpha-fibrinogenase activity; in contrast, the gelatinolytic action was only observed in fraction F11. Only metalloproteinase inhibitors abolished all these proteolytic activities. Our results suggest that adrenal cortex lesions may be relevant in the etiopathogenesis of severe brown widow spider envenoming. To our knowledge, this is the first report on adrenal gland damages, fibrinogenolytic activity and interrelations with cell-matrix adhesion proteins caused by L.geometricus VGH. The venom of this spider could be inducing hemostatic system damages on envenomed patients.

Ultraestructura renal en infecciones murinas experimentales con un aislado venezolano de Trypanosoma evansi / Renal ultrastructure in experimental murine infections with a Venezuelan isolate of Trypanosoma evansi

Infecciones murinas experimentales con un aislado venezolano de Trypanosoma evansi ocasionan cambios ma - cros copicos y ultraestructurales en el rinon de los ratones. La modificacion macroscopica se restringe a palidez progresiva del organo. Por su parte, las alteraciones ultraestructurales incluyen incremento del grosor de la membrana basal glomerular, asi como del espesor de la membrana basal de los tubulos contorneados proximales, cambios en la colocacion y el agrupamiento de las mitocondrias asociadas a la lamina basal, asi como extenso deterioro epitelial. Los tripanosomas intravasculares fueron detectados en la circulacion renal a partir de dia 9 post-infeccion. Se discute al respecto del dano glomerular, sugiriendo que las alteraciones submicroscopicas debilitarian la funcion tubular, contribuyendo mediante un proceso nefritico al desarrollo de una falla renal aguda que culminaria con la muerte del hospedador experimental. La significancia de los cambios ultraestructurales fue corroborada estadisticamente.

Murine experimental inoculations with a Venezuelan isolate of Trypanosoma evansi cause macroscopic and ultrastructural changes in the kidney of the infected animals. The macroscopic modifications are restricted to the progressive paleness of the organ. By its part, the ultrastructural alterations include thickness enlargement of the glomerular basal and the proximal tubule basal membranes, changes in the mitochondrial grouping and layout close to the basal membrane, as well as extensive epithelial damage. Intra vascu - lar trypanosomes were seen in the renal circulation from day 9 post-infection and on. The glomerular injury is discussed, suggesting that submicroscopic alterations could weaken the tubular function contributing through a nephritic process with the development of an acute renal failure culminating in the experimental host¡¦s death. The significance of the sequential increasing ultrastructural changes was statistically corroborated.

Neurotoxic activity and ultrastructural changes in muscles caused by the brown widow spider Latrodectus geometricus venom.

Brown widow spider (Latrodectus geometricus) venom (BrWSV) produces few local lesions and intense systemic reactions such as cramps, harsh muscle pains, nausea, vomiting and hypertension. Approximately 16 protein bands under reducing conditions and approximately 14 bands under non-reducing conditions on a 12.5% sodium dodecyl sulfate-polyacrylamide gel electrophoresis were observed. Neurotoxic clinical manifestations were confirmed in vivo, while proteolytic activity was demonstrated on gelatine film. Severe ultrastructural damages in mice skeletal muscles were observed at 3, 6, 12 and 24 h postinjection with at total of 45 microg of venom protein. Infiltration of eosinophils and ruptures of the cellular membranes were observed in the muscles along with swelling of the nuclear cover and interruption of the collagen periodicity. Altered mitochondrias and autophage vacuoles, nuclear indentation and mitochondria without cristae, slight increment of intermyofibrillar and subsarcolemic spaces and myelinic figures formation were also observed. In the capillary, endothelial membrane unfolding into the lumen was noticed; along with myelinic figures compatible with a toxic myopathy. Swollen sarcotubular systems with lysis of membrane, intense mitochondria autophagia and areas without pinocytic vesicles were observed. Swollen mitochondria surrounded by necrotic areas, myofibrillar disorganization and big vacuolas of the sarcotubular system, degenerated mitochondrium with formation of myelinic figure was seen. Glycogenosomes with small particulate, muscle type glycogen was noticed. Autophagic vacuole (autophagolysosomes) and necrotic areas were also noticed. These damages may be due to interactive effects of the multifactorial action of venom components. However, Latrodectus geometricus venom molecules may also be utilized as neuro therapeutic tools, as they affect neuronal activities with high affinity and selectivity. To our knowledge, the present study is the first ultrastructural report in the literature of muscle injuries and neurological and proteolytic activities caused by BrWSV.

Neurotoxic activity and ultrastructural changes in muscles caused by the brown widow spider Latrodectus geometricus venom / Actividad neurotóxica y cambios ultraestructurales en musculos causados por el veneno de la araña viuda marrón Latrodectus geometricus

Brown widow spider (Latrodectus geometricus) venom (BrWSV) produces few local lesions and intense systemic reactions such as cramps, harsh muscle pains, nausea, vomiting and hypertension. Approximately 16 protein bands under reducing conditions and ~ 14 bands under non-reducing conditions on a 12.5% sodium dodecyl sulfate-polyacrylamide gel electrophoresis were observed. Neurotoxic clinical manifestations were confirmed in vivo, while proteolytic activity was demonstrated on gelatine film. Severe ultrastructural damages in mice skeletal muscles were observed at 3, 6, 12 and 24 h postinjection with at total of 45 µg of venom protein. Infiltration of eosinophils and ruptures of the cellular membranes were observed in the muscles along with swelling of the nuclear cover and interruption of the collagen periodicity. Altered mitochondrias and autophage vacuoles, nuclear indentation and mitochondria without cristae, slight increment of intermyofibrillar and subsarcolemic spaces and myelinic figures formation were also observed. In the capillary, endothelial membrane unfolding into the lumen was noticed; along with myelinic figures compatible with a toxic myopathy. Swollen sarcotubular systems with lysis of membrane, intense mitochondria autophagia and areas without pinocytic vesicles were observed. Swollen mitochondria surrounded by necrotic areas, myofibrillar disorganization and big vacuolas of the sarcotubular system, degenerated mitochondrium with formation of myelinic figure was seen. Glycogenosomes with small particulate, muscle type glycogen was noticed. Autophagic vacuole (autophagolysosomes) and necrotic areas were also noticed. These damages may be due to interactive effects of the multifactorial action of venom components. However, Latrodectus geometricus venom molecules may also be utilized as neuro therapeutic tools, as they affect neuronal activities with high affinity and selectivity. To our knowledge, the present study...

El veneno de la araña viuda marrón (Latrodectus geometricus) produce pocas lesiones locales pero intensas reacciones sistémicas, tales como calambres, dolores musculares severos, nauseas, vómitos e hipertensión arterial. Se observaron ~ 16 bandas de proteina bajo condiciones reducidas y ~14 bandas bajo condiciones no reducidas en electroforesis en geles de poliacrilamida al 12.5%. Las manifestaciones neurotóxicas clínicas fueron confirmadas in vivo, mientras que la actividad proteolítica fue demostrada en una placa de gelatina. Los músculos de ratón se estudiaron durante las 3, 6, 12 y 24 horas después de ser inyectados con 45 µg de proteina de veneno. Los músculos fueron seriamente dañados por este veneno. Se demostró una infiltracción de células eosinofílicas y rupturas de membranas celulares en tejido muscular, al mismo tiempo un fuerte incremento de la membrana nuclear y una interrupción de la periodicidad del colágeno. Se observaron daños en la mitocondria y sin cristaes, vacuolas autofágicas e indentación nuclear. Se notó un aumento de la luz de los espacios intermiofibrilares y subsarcolemicos. En los capilares fue visible un desdoblamiento de la membrana endotelial hacia el lúmen vascular. Del mismo modo, fue visto un hinchamiento del sistema sarcotubular con lisis de las membranas; intensa autofagia de mitocondrias y áreas sin vesículas pinocíticas. Fue además observado, glucogenosomas con glucogeno particulado. Se observaron vacuolas autofágicas (autofagolisosomas) y áreas de necrosis. Estos daños podrían ser atribuídos a los efectos interactivos de una acción multifactorial de los componentes del veneno.

Morphological alterations in skeletal muscle of spontaneously hypertensive rats.

The Extensor digitorum longus (EDL) and the soleus muscles of spontaneously hypertensive rats (SHR) were studied in comparison with those of their normal counterparts, the Wistar Kyoto (WKY) rats. Quantitative assessment of capillaries and muscle fibre typing was done with optical microscopy, while the study of capillary abnormalities was performed by ultrastructural observation. There were no differences in fibre type proportion or in capillarity indexes between the SHR and the control rats. A reduction in the area of IIB fibres was found in the EDL muscle of the hypertensive animals. The ultrastructural study showed abnormalities in the capillaries of both muscles in SHR, the cross section of the endothelial cells was enlarged; there was irregular distribution of caveolae and pinocytic vesicles, the capillary basement membrane showed irregular width, with parts engrossed and reduplicated. Some pericytes were prominent. There were macrophages present in the interstitial space. In some muscle fibres there was disorganization of the sarcomere structure, swelling of the sarcotubular system, abundant autophagic vacuoles, and proliferative satellite cells. There were abundant collagen fibrils. The presence of cellular rests, autophagic vacuoles and loss of sarcolemma indicated necrosis. It can be concluded, that in SHR, muscle capillaries showed alterations that may be the substrate of functional rarefaction, although anatomical rarefaction (number reduction) could not be demonstrated. In EDL and soleus muscles of SHR, signs of a mild myopathy with focal fibrosis were present.

Alteraciones ultraestructurales en eritema gingival lineal en pacientes vih / Ultrastructural alterations in linear gingival erythema in HIV patients

La enfermedad periodontal asociada mas frecuentemente a infección por VIH es el eritema gingival lineal, gingivitis ulcerativa necrozante y periodontitis ulcerativa necrozante. Los cambios que sufre el periodonto en presencia de VIH se sugiere que pueden ser indicativos de alteraciones sistémicas como infección por VIH. Los virus en la enfermedad periodontal, juegan un papel muy importante estando relacionados con el sistema inmune del huésped sobre todo en relación con patologías periodontales como eritema gingival lineal y periodontitis ulcerativa necrozante. Se encontró disminución de linfocitos T, macrófagos y alto porcentaje de neutrófilos e IgG en el plasma sanguíneo en eritema gingival lineal, existiendo cambios importantes en las células inflamatorias. Alto número de neutrófilos en el epitelio gingival se asocia con la severa necrosis gingival reportada en pacientes con SIDA El objetivo de este trabajo es establecer algunas alteraciones ultraestructurales en una patología periodontal como es el eritema gingival lineal en pacientes VIH. Material y Método Se tomaron biopsias de tejido periodontal con diagnostico de eritema gingival lineal en pacientes que acudieron al Centro de Atención a Pacientes con Enfermedades Infectocontagiosas "Dra. Elsa La Corte", adultos, masculinos. Pacientes VIH+, bajo terapia HAART. Biopsias procesadas por técnicas rutinarias para M.E.T Resultados: alteración de melanocitos presencia de queratinocitos y nucleoedema, mitocondrias electrondensas, escasos tonofilamentos y poco retículo endoplásmico rugoso y Golgi. Núcleo de contorno ligeramente irregular (espongiosis), pérdida de desmosomas, específicamente mácula adherens, membrana basal continua, con pequeñas áreas de discontinuidad (fenestras), pocas vesículas pinocíticas y algunas caveolas. Abundante colágeno, fibroblastos atípicos en forma de masa celular. Conclusión: las alteraciones ultraestructurales encontradas en eritema gingival lineal sugieren

Linear Gingival Erythema, Ulcerative Necrotizing Gingivitis and Periodontitis are the most frequent pathology associated to HIV. The alterations in periodontal tissue could indicate the presence of a systemic disease like HIV infection. The virus in periodontal disease have a closer relations with host human immune system, specifically in Linear Gingival Erythema and Ulcerative Necrotizing Periodontitis and Gingivitis. In the blood plasma of Linear Gingival Erythema it was found decrease of T lymphocyte, histiocytes and high percent of neutrophils and Ig G, and important changes in the inflammatory cell. High number of neutrophils in the gingival epithelial are associated with mostly severe gingival necrosis that reported in AIDS patient. Propose: evaluation of ultrastructural alterations in periodontal disease like lineal gingival erythema. Material and Methods: biopsies were taken from periodontal lesions with diagnostic of lineal gingival erythema, patients attending the Centre for the Care of Patients with Infections and Contagious diseases "Dra Elsa La Corte". The patients are males using HAART therapy. Biopsies were processed according to rutinary techniques for Transmission Electron Microscopy. Results: melanocytes alteration, and nucleoedema, different electron densities mitochondria, scarce tonofilaments, rough endoplasmic reticulum and Golgi apparatus. Irregular shape nucleus, loosed desmosome specifically macula adherens, spongyosis continuous basement membrane, with little areas of discontinue membrane, scarce pynocitc vesicles and caveolas, plentiful collagen, atipic fibroblast like cell mass. Conclusion: it suggested that ultrastructural alterations founded in lineal gingival erythema are irreversible changes similar to those described in autoimmune disease and cancer

Patología ultraestructural del infiltrado inflamatorio en la displasia epitelial y carcinoma espinocelular bucal / Ultrastructural pathology of inflammatory infiltrate in epithelial dysplasia and oral spinocellular carcinoma

Se ha observado que muchas neoplasias se desarrollan en asociación con la inflamación, infección e irritación crónica, siendo éste el caso del Carcinoma Espinocelular bucal y los estados previos de Displasia Epitelial. El propósito del presente trabajo fue estudiar ultraestructuralmente el infiltrado inflamatorio en lesiones epiteliales con Displasias y Carcinomas Espinocelulares bucales, mediante técnicas convencionales de la microscopia electrónica de transmisión. La presencia de un infiltrado inflamatorio y las frecuentes interacciones observadas entre sus componentes con las células epiteliales neoplásicas, sugieren su participación activa en el desarrollo y progresión del Carcinoma Espinocelular bucal dada la etiopatogenia de estas lesiones y la estrecha relación entre el microambiente generado por la inflamación crónica y las células epiteliales tumorales

It has been observed that many neoplasms are developed in association with inflammation, infection and chronic irritation, which is the case of oral Spinocellular Carcinoma and previous states of Epithelial Dysplasias. The purpose of the present work was to study ultrastructurally : inflammatory infiltrate in epithelial lesions related to Dysplasia and oral Spinocellular Carcinomas by routine techniques for transmission electron microscopy. Observed presence of inflammatory infiltrate and the frequent interactions between cells of infiltrate and with neoplastic cells suggests its active participation in the development and progression of oral Spinocellular Carcinoma given the etiopathogenia of these lesions and the intimate relation among microenvironment generated by chronic inflammation and tumor epithelial cells

Alterations in the ultrastructure of cardiac autonomic nervous system triggered by crotoxin from rattlesnake (Crotalus durissus cumanensis) venom.

This study explored the toxic effects of crotoxin isolated from Crotalus durissus cumanensis venom on the ultrastructure of mice cardiac autonomic nervous system. Mice were intravenously injected with saline (control group) and crotoxin diluted in saline venom (study group) at a dose of 0.107 mg/kg mouse body weight. Samples from the inter-ventricular septum were prepared for electron microscopy after 6 h (G1), 12 h (G2), 24 h (G3) and 48 h (G4). The G1 group showed some cardiomyocyte with pleomorphic mitochondria. Capillary swollen walls, nerve cholinergic endings with depleted acetylcholine vesicles in their interior and other depletions were observed. A space completely lacking in contractile elements was noticed. The G2 group demonstrated a myelinic figure, a subsarcolemic region with few myofibrils and nervous cholinergic terminal with scarce vacuoles in their interior. The G3 group demonstrated a structure with a depleted axonic terminal, mitochondrias varying in size and enhanced electron density. In addition, muscular fibers with myofibrillar structure disorganization, a depleted nervous structure surrounded by a Schwann cell along with an abundance of natriuretic peptides, were seen. An amyelinic terminal with depleted Schwann cell and with scarce vesicles was also observed. Finally, axonic lysis with autophagic vacuoles in their interior and condensed mitochondria was observed in the G4 group. This work describes the first report of ultrastructural damage caused by crotoxin on mice cardiac autonomic nervous system.

Alteraciones musculares periféricas en la enfermedad de Chagas

La enfermedad de Chagas sigue siendo un problema en Venezuela y en varios países de Latinoamérica. Este trabajo presenta datos sobre las alteraciones que esta enfermedad produce en los músculos esqueléticos periféricos, lo cual puede contribuir a la incapacidad funcional que presentan muchos de los pacientes. Fueron estudiados 19 pacientes con enfermedad avanzada, a quienes se les hizo evaluación cardiorrespiratoria, ecocardiograma, prueba de ejercicio y biopsia del músculo cuádriceps. El 42% presentó incapacidad funcional moderada a severa, medida por el consumo máximo de oxígeno, que estuvo relacionada con cambios morfológicos y metabólicos en las fibras musculares. Se hallaron alteraciones en la microvasculatura muscular compatibles con daño autoinmune. En conclusión, los pacientes con enfermedad de Chagas sufren alteraciones en la musculatura periférica que puede contribuir a la incapacidad funcional y al grado de afectación por esta enfermedad. La determinación de la capacidad funcional debería agregarse a la evaluación de la enfermedad

Chagas disease is still a problem in Venezuela and other Latin American countries, in spite of efforts to control the disease. Peripheral muscle alterations that contribute to decreased functional capacity in moderate to advanced Chagas’ disease patients are analyzed in the present work. Nineteen Chagas’ disease patients were studied, including cardio-respiratory evaluation, echocardiogram, exercise test and quadriceps muscle biopsy. The results showed 42% patients with moderate to severe decrease of maximal oxygen consumption. Muscle biopsies presented morphologic and metabolic alterations, some related to functional capacity. Capillary vessel damage in muscle was similar to that found in autoimmune diseases. In conclusion, Chagas’ disease patients showed peripheral muscle alterations related to decreased functional capacity. The functional capacity changes may contribute to the severity of the disease and should be added to the evaluation of the patients

Alteraciones ultraestructurales en la microvasculatura de la región de la cara en pacientes VIH / Ultrastructural alterations in face microvasculature in HIV patients

Se han descrito las alteraciones ultraestructurales en fibras musculares esqueléticas en pacientes VIH+. En fibras del músculo orbicular de los labios se efectuaron las siguientes observaciones: atrofia, desorganización del sistema sarcotubular y núcleos hipercrómaticos. Los cambios encontrados en la microvasculatura, fueron compatibles con los encontrados en la fibra en enfermedades autoinmunes: Lupus Eritematoso Sistémico, Diabetes Autoinmune, Síndrome de Sjõrgen. Propósito: Evaluar las alteraciones ultraestructurales en la microvasculatura asociadas a desórdenes en el músculo orbicular de los labios de pacientes VIH+. Material y Método: se tomaron biopsias del músculo orbicular de los labios de pacientes del Centro de Atención a Pacientes con Enfermedades Infectocontagiosas Dra. Elsa La Corte entre 2001- 2002, masculinos , femeninos, edades entre 38 y 53 años. Pacientes VIH+, bajo terapia HAART, presentando miopatía. Biopsias procesadas por técnicas rutinarias para M.E.T. Resultados: citoplasma capilar proliferativo, ruptura de la membrana plasmática de la célula endotelial, membrana basal engrosada, endotelio con áreas electrón densas y electrón transparentes, prolongaciones del citoplasma hacia la luz. Conclusión: se sugiere que los efectos del VIH sobre la microvasculatura del músculo esquelético son similares a los descritos en otras enfermedades autoinmunes.

Ultrastructural alterations of squeletic muscle fibers in HIV+ patients have been described. In orbicular muscle fibers of lips were realized the following observations: atrophy, sarcotubular system desorganization and hyperchromatic nuclei. Changes found in the microvasculature were similar to there seen in autoimmune disorders as Systemic Lupus Erythematosus, Autoimmune Diabetes and Sjörgen Syndrome. Propose: evaluation of microvascular ultrastructural alterations in orbicular muscles in HIV patients. Material and Methods: biopsies were taken from lips orbicular muscles in patients attending the Centre for the Care of Patients with Infections and Contagions diseases "Dra Elsa La Corte" between years 2001 and 2002, females and males, with ages between 38 and 53 years. Patients used HAART therapy, present myopathy. .Biopsies were processed according to rutinary techniques for Transmission Electron Microscopy. Results: proliferative endothelial cell cytoplasm in some case rupture of plasma membrane with necrosis, widening of basement membrane, endothelial cell cytoplasm with different electron densities and infolding of cytoplasm into the lumen. Conclusion: it is suggested that the effects of HIV on the skeletal muscle microvasculature are similar to those described in other autoimmune diseases.