Serosal ADH stimulation enhances water flow under an imposed osmotic gradient through insertion of water channels (aggrephores) into the mucosal plasma membrane of toad urinary bladder sacs. Following cessation of ADH actions, water channels are retrieved as endosomes that can be visualized by mucosal inclusion of horseradish peroxidase (HRP) into round vesicles, long tubules and multivesicular bodies within the cytosol (1,2,3). Endosomes also occur adjacent to golgi bodies or lysosomes (1,2,3). However, true nature of endosomes including their formation at the mucosal surface and their shuttling in granular cells is still unclear (4,5). Current studies were undertaken to understand the role of endosomes in water channel cycling in this renal membrane model.

Urinary bladder sacs removed surgically from doubly-pithed toads, were suspended at ends of glass tubes. Control (no hormone) and experimental bladder sacs were exposed to ADH for 10 min in the absence of osmotic gradient.

Several clinically important anti-cancer agents exert their effects on tumor cells through interference with the function of microtubules. In addition to the Vinca alkaloids, such as vinblastine and vincristine, the taxanes, such as paclitaxel (Trade Name: Taxol), kill tumor cells through a microtubular target. Treatment with taxol leads to the inability of microtubules to depolymerize, leading to the formation of large intracellular microtubular bundles. In tumor cells that progress through the cell cycle, this leads to the inability of these cells to disassembly interphase microtubule networks and a failure to form functional mitotic spindles. These cells arrest in M phase, from which they eventually progress, either by the induction of apoptotic cell death, or by micronucleation and the formation of tetraploid cells. There is also the possibility that taxol has other effects on the regulation of genes or other systems to enhance cell killing, perhaps through lowering the threshold of cells to the induction of apoptotic cell death.

The microtubule (MT) cytoskeleton plays a crucial role in a number of cellular functions including cytoplasmic organization, intracellular transport, cellular polarity, and cell division. Microtubules are linear polymeric structures composed of heterodimers of α- and β-tubulin with a class of specialized proteins, the microtubule-associated proteins, typically being present. Microtubules are dynamic structures that undergo repeated rounds of assembly/disassembly. The half-life of MTs, an index of their dynamics (i.e., assembly-disassembly), has been estimated from studies of single cells in culture and typically ranges from ≈ 5 - 20 minutes depending on the cell type. These dynamic properties of MTs have been explained in terms of the “dynamic instability” model. Five parameters have been used to describe dynamic instability of MTs: (1) elongation velocity; (2) shortening velocity; (3) nucleation frequency; (4) catastrophe frequency; and (5) rescue frequency. Catastrophe is the transition from elongation to shortening while rescue is the transition from shortening to elongation.

Following intracellular uptake, taxanes bind to β tubulin and suppress microtubular dynamics, resulting in stable microtubule bundles and mitotic arrest of cancer cells. This proximal taxane- β tubulin interaction depends on the amount of intracellular accumulation of taxanes. Since taxanes are substrates for the membrane transporter P-glycoprotein (PGP) but not the MRP, overexpression of PGP but not MRP reduces the intracellular accumulation and cytotoxicity of taxanes. Recent reports have also suggested that alterations in the levels of specific β tubulin isotypes can reduce the effects of taxanes on microtubule dynamics, thereby inhibiting the mitotic effects and resulting cytotoxicity of taxanes.6 For example, increased βIV and (βIII tubulin levels may confer resistance against microtubule and cytostatic effects of taxanes. Taxane-induced G2/M arrest is preceded by the activation of cell-cycle mitotic kinase CDK1, which promotes the G2/M transition. Recent studies have indicated that the loss of cell-cycle Gl checkpoint due to p53 mutations may enhance taxane-induced mitotic arrest and cytotoxicity.

Taxol, a plant-derived antimicrotubule agent, was originally isolated from the bark of the Pacific yew, Taxus brevifolia. This naturally occurring antineoplastic drug has been demonstrated to possess broad activity against a variety of human solid tumors, particularly in drug-refractory ovarian cancer and metastatic breast cancer (1). However, the exact mechanism of taxol's cytotoxicity against tumor cells is not entirely clear. Recent studies have demonstrated that taxol, besides causing microtubule bundling and mitotic arrest, is able to induce internucleosomal DNA fragmentation and typical morphological features of apoptosis in a number of solid tumor cells. These results clearly indicate that taxol, in addition to its classical activity against microtubules and cell cycle arrest, also possesses significant cell-killing activity by induction of apoptosis.

Although it is well recognized that taxol can cause both mitotic arrest and apoptotic cell death, it remains unclear whether taxol-induced cell death is a secondary event resulting from mitotic arrest or represents a novel mechanism of taxol against tumor cells.

Correlative microscopy is employed in a great variety of settings by both diagnostic and investigative pathologists. Combinations of conventional light microscopy (LM), immunohistology, and electron microscopy (EM) are used in a wide range of diagnostic settings, including the analysis of tumors, autoimmune disorders, and infectious diseases. Valuable diagnostic information is also frequently obtained by simultaneous or sequential examination of exfoliated or aspirated cell suspensions (cytopathology) and tissue sections (histopathology) from the same lesion. An even wider range of correlative microscopic methods is employed by pathologists in basic and clinical research. The rationales for using correlative techniques are many and varied, but in most cases fall within a limited number of categories.

Pathologists frequently use a second microscopic or preparative technique to improve on the resolution afforded by an initial technique. Electron microscopy is often used to refine the analysis of features initially detected by routine LM or immunohistology.

Molecular pathology represents a distinctive new discipline in medicine with many potential applications that have been heretofore untapped. This discipline is introducing important changes in the concepts of disease causation and in the classification of diseases. Transfer of molecular genetic technology and knowledge to the pathology laboratory is giving the pathologist new and powerful tools to diagnose, classify, and monitor disease. Methodologies such as in situ hybridization (ISH), immunohistochemistry (IHC), and the polymerase chain reaction (PCR) will continue to allow more systematic and accurate classification of human diseases. IHC and ISH allow the identification and localization of cells containing specific antigens or nucleic acid sequences. The main advantage of these methods is their ability to localize targets and define their morphologic features in various tissues, allowing correlations with histopathologic changes.

Cisplatin (cw-diamminedichloroplatinum II; CDDP) is a broad spectrum anti-cancer agent with severe toxic side effects. New platinum based chemotherapeutic agents with similar properties are in constant development. Poly-plat (Poly-[(teans-l,2-diaminocyclohexane) platinum]-carboxyamylose), a novel second generation platinum chemotherapeutic agent, has been shown to be less toxic compared to CDDP. The purpose of this paper was to examine the morphological changes in the Kupffer cells in the liver after poly-plat treatment in rats and compare the results obtained after CDDP. Peripheral blood was also examined for levels of circulating leukocytes.

Wistar rats (120-160 g) were given intraperitoneal injections of either CDDP (9 mg/kg in 0.9% sodium chloride) or poly-plat (10 mg/kg in 0.9% sodium chloride) over a five day period. Controls were treated with equal amounts of the vehicle of injection. Peripheral blood was collected via tail vein beginning 1 day prior to treatment and continuing 10 days after treatment.

Rotaviruses are complex, relatively large (1000 Å) nonenveloped icosahedral viruses. They are the major causative agents of severe diarrhea in children and animals. Annually an estimated 1 million children die from rotavirus-induced gastroenteritis worldwide. Medical relevance, intriguing structural complexity, and several unique strategies in the morphogenesis and replication have provoked extensive molecular biological and structural studies on rotaviruses in recent years.

Rotaviruses are double-stranded RNA (dsRNA) viruses belonging to the family Reoviridae. Each of the 11 segments codes for one protein. Of the eleven proteins encoded by the genomic RNA, six are structural and five are non-structural. Structural studies have shown that mature rotaviruses are triple-layered particles (TLPs). During the process of cell entry the outer-most layer consisting of VP7 and the spike protein VP4 is lost and the resulting double layered particles (DLPs) become transcriptionally active (Fig. 1 A). During the process of transcription, the DLPs maintain their structural Integrity.

Endemic and epidemic viral gastroenteritis are major causes of morbidity worldwide and are frequently a major cause of death in the developing world. Rotavirus, adenovirus, calicivirus, astrovirus, and the Norwalk virus family are the viruses most often associated with gastroenteritis. Of these, the major cause of epidemic viral gastroenteritis in the US is Norwalk-like viruses. Gastroenteritis caused by this virus group is responsible for such outbreaks in settings such as cruise ships, schools, camps, hospitals, day-care and extended care facilities, football games and festivals.

As early as the 1950s and 1960s electron microscopy (EM), including negative stain EM, was established as a diagnostic tool for identifying viruses. Negative stain EM is especially useful for identifying viruses that are difficult or impossible to culture (eg. viruses in the Norwalk family). The first Norwalk virus was identified in the 1969 by immune-negative stain EM. Many antigenic types were identified soon afterward that did not cross-react with the prototype Norwalk virus.

Analysis of Hepatitis C virus (HCV) assembly and packaging, and the study of virus-cell interactions, have been impeded by the lack of a robust cell culture system and a viable small animal model. Recently, an alternative approach utilizing recombinant gene technology has been implemented. We describe a system in which specific HCV genes are cloned into a baculovirus to form a recombinant baculovirus expression vector that produces HCV proteins that self-assemble into enveloped virus-like particles (VLP). Four HCV structural genes were cloned into the baculovirus expression vector under the control of the polyhedrin promoter, and were then recombined with baculovirus DNA. A baculovirus with no HCV insert served as a negative control (wild-type). Ovary cells (Spodoptera frugiperda) were infected with either the recombinant or wild type baculovirus. Five-days post-infection, a group of cells were harvested, fixed in McDowell-Trumps solution, and routinely prepared for transmission electron microscopy (TEM).

The application of both classical and advanced microscopy to the exploration of abnormal, human, fetal development (birth defects) and to other forms of human, fetal disease has generated a wealth of information, often documented by dramatic images. We have studied, at varying levels of inquiry (simple inspection to molecular analysis), some 10,000 normal and abnormal specimens. For presentation purposes, we first demonstrate the use of imaging in explorations of the fetal habitat (the gestational sac). Our studies reveal a variety of fetal surface encounter lesions (skin-to-amnion). Many of these lesions in the past have been erroneously interpreted. These include fetal amputations, strangulations, and also the reimplantation of amputated parts to ectopic sites. We have also explored, by various imaging techniques, developmental abnormalities within a variety of fetal organs.

The comparative morphology of the tongue microvasculature has been studied in different animals, such as in man (Okada et al, 1993, Matsukawa and Okada, 1994), rats (Nagato et al., 1989, Ohshima, et al, 1990, Selliseth and Slvig, 1993), dogs (Kishi et al, 1986, Kishi et al 1988), goats(Scala et al, 1992), monkeys (Patterson, et al, 1992, Okada et al, 1993, Matsukawa and Okada, 1994), marmosets (Okada et al, 1993, Matsukawa and Okada, 1994) and shrews (Okada et al, 1993, Matsukawa and Okada, 1994), but apparently none in rabbits. The disposition of the blood vessels of the rabbit's lingual papillae were studied by injecting them with Mercox resin and posterior corrosion, thus obtained microcorrosion casts were observed in the scanning electron microscope.

Male and adult rabbits were used to obtain our models. After perfusion with heparin 0,1M, pH 7.4 and glutaraldehyde, both in the same buffer, the vessels were filled as reported above.

Hepatomegaly, characterized by increased liver weight due to hyperplasia or hypertrophy, can be induced in many species by various drugs or environmental toxins. In rats, hepatomegaly is more usually associated with increases of smooth endoplasmic reticulum in hepatocytes and the associated induction of cytochrome P450 enzymes or with hepatocellular peroxisomal proliferation. In this study, a substituted pyrimidine derivative given to rats resulted in hepatomegaly with an unusual selective increase in hepatocellular rough endoplasmic reticulum (RER).

Female Sprague-Dawley rats were given repeated oral doses of a substituted pyrimidine derivative for 28 consecutive days. Livers were collected at necropsy, weighed, fixed in 10% neutral buffered formalin and processed to paraffin sections stained with hematoxylin and eosin (H&E) for light microscopic evaluation. Portions of formalin-fixed liver were transferred to buffered 2.5% glutaraldehyde for ultrastructural evaluation. Specimens were processed into epoxy resin and thin sections were examined with a JEOL 1200EX transmission electron microscope.

Some pathologic alterations in animal tissue sections, because of low resolution, are not identified using light microscopy. Availability of higher resolution in electron microscopy (EM) confers a decisive role on EM for diagnostic pathology that is well accepted. Much has already been contributed by EM in understanding of liver diseases. However, EM has been further refined by adding a quantitative facet to the results. Morphometric technique allows transfer of facts acquired from 2-dimensional samples, e.g., electron micrographs, to a 3-dimensional perspective. In addition, the technique is valuable in determining the significance of the alterations especially in instances where the changes numerically are either scanty or indistinguishable from a normal appearance of the cells or tissues (Fig. 1).

In our work on the effects of several different PCB congeners, one of the compounds, namely, congener number 153 was studied to obtain morphometric data on the lesions in the liver of rats given the congener in the diets for 13 weeks.

Pathological changes of hepatocytes from rats fed a 30% ethanol-derived calories diet for three weeks include noticeable ultrastructural changes including steatosis and hypertrophy of the smooth endoplasmic reticulum. We sought to examine hepatocytes of deer mice administered ethanol in an inhalation chamber for two weeks to determine if subtle changes occur in hepatocyte organelles prior to steatosis.

Two strains of Peromyscus maniculatus, ADH-positive possessing hepatic cytosolic alcohol dehydrogenase and ADH-negative deer mice lacking this enzyme were purchased from the Peromyscus Genetic Stock Center (Univ. of South Carolina). They tested negatively for Hanta viruses. A morphometric analysis of the ultrastructure of ADH+(n=14) and ADH- (n=14) controls as well as experimentals exposed to chronic, intoxicating levels of ethanol was conducted. Blood ethanol levels were maintained between 1.25-1.75 mg/ml for two weeks in the experimentals.

Cancer is a disease characterized by uncontrolled cell divisions in which the molecular controls for cytoskeletal regulation are bypassed. Cell division is governed by centrosomes, microtubule-organizing cell organelles which are crucial for the organization of the mitotic apparatus during mitosis and cell division. Because centrosome abnormalities are observed in the most common human cancers, we used immunofluorescence and transmission electron microscopy to determine centrosome organization in the human androgenresponsive prostate cancer cell line LNCaP and the androgen-independent prostate cancer cell line DU145. During interphase, centrosomes are located in close vicinity to the outer nuclear membrane, duplicate during S-phase, and become separated to the mitotic poles during the transition from interphase to mitosis. Centrosome regulation is based on a number of different factors which are only partly understood. Hormones play a role during developmental regulation of prostates which might trigger the activation of centrosome proteins and consequent cell divisions in order to ensure tissue growth.

Cell adhesion and cell adhesion molecules (CAMs) play a crucial role in testicular development and function. The seminiferous epithelium, the functional unit of the testis, represents a three dimensional architecture of supporting Sertoli cells (SC), and developing germ cells (GC). The seminiferous epithelium, therefore, must be receptive not only to individual cell growth and differentiation, but also to cell-cell interactions. Morphologically distinct cell-cell interactions occur between SC and GC and also between SC.[1] In general, these junctions can be categorized into three types: adhesive, occluding, and gap junctions. The orientation and function of these junctions are interaction dependent. For example, desmosome-like junctions (spot desmosomes) are found between SC and GC. These junctions are present in the basal and intermediate compartments of the testis and serve to translocate developing GC. SC-SC interactions, like the zonula occludens (tight junction), function as vectorial mediators, maintaining the blood-testis barrier and SC polarity.

The vascular endothelium is the interface with hyperglycemia in uncontrolled diabetes mellitus and its complications, including nephropathy, myocardial infarction, blindness, and gangrene. To mimic the vascular endothelium in a controlled experimental situation, we use the endothelial cell line (CRL 1998) obtained from The American Type Culture Collection. Cells are grown to confluency in Media 199 containing 10% fetal calf serum and gentamicin (50mg/ml). The growth media is replaced with experimental media containing high glucose (30mM), insulin (20 Units/ml), or heparin (500 Units/ml) either separately or in combinations. Cells are briefly trypsinized, pelleted, embedded, and sectioned with a Diatome diamond knife for TEM or for SEM, grown on 0.45um filters, followed by fixation and air-drying in situ: this approach to culture allows cells to obtain a polar, more natural phenotype than monolayers on glass or plastic.

Incubation in high glucose results in loss of confluency, production of intercellular gaps, and fewer mitotic figures.

Monocrotaline (M) induces pulmonary hypertension (PH) characterized by increased pulmonary arterial pressure, thickened pulmonary arterial walls, and right ventricular hypertrophy (RVH). In addition, numerous mast cells (MC) have been localized in M treated rats and these cells may have a role in causing PH and other lung injury because of their content of serotonin, which is a vasoconstrictor. Nitric oxide (NO) is a vasodilator that has been used in treatments of human patients and experimental animals with PH. The purpose of this study is to determine if NO inhalation can prevent or reverse RVH or MC accumulation induced by M.

Adult male Sprague-Dawley rats were divided into 5 groups (C, M3, MN3, M5, MN5). Group C served as controls without treatments. Groups M3 and M5 received one single subcutaneous injection of M (80mg/kg BW) and were sacrificed at 3 weeks (M3) or 5 weeks (M5) after injection.