Ultrastructural, protein, and lipid changes in liver associated with chlordecone treatment of mice.

Pretreatment of mice with chlordecone (CD) reduced hepatic accumulation of a subsequent dose of [14C]CD without significantly changing [14C]CD biotransformation. To determine if CD-induced changes in hepatic [14C]CD accumulation were coincident with altered cell composition, we examined the effects of CD on hepatic protein and lipid content, on fatty acid profiles of liver and kidney, and on the ultrastructure of hepatocytes. SDS-polyacrylamide gel electrophoresis detected an apparent CD dose-related increase in a microsomal protein with a molecular weight of about 23 kDa. Total liver or kidney lipid contents were not altered by CD but relative amounts of several hepatic fatty acids were changed. CD caused marked hepatic mitochondrial swelling, increased amounts of endoplasmic reticulum, apparently increased numbers of peroxisome-like structures, and decreased numbers of lipid droplets in cytoplasm of hepatocytes. Numbers of lipid droplets were not decreased in perisinusoidal fat storage cells. In addition, the numbers of cytoplasmic lipoprotein vesicles were apparently increased in some hepatocytes. Overall these changes indicated an increased hepatocyte secretory activity and suggested that CD changed hepatocellular lipid transport, storage, and metabolism pathways.

Culture of cells from juvenile worms of Schistosoma mansoni.

Tissue disruption methods were developed and serum-free cell culture media formulated for the maintenance in vitro of cells from juvenile worms (day 18 after infection) of Schistosoma mansoni. Cultures maintained viability for up to 6 mo when plated on a feeder layer of irradiated rat liver cells and survived primarily as clusters of small (2.5-4 microns diameter) cells with a high nuclear-to-cytoplasmic ratio and relatively few organelles identified by electron microscopy. Cultures synthesized a protein profile similar to that of intact worms, and the cell clusters maintained a time- and concentration-dependent contractile response to serotonin. Cells synthesizing DNA were detected by precursor incorporation and flow cytometry in cultures initially and also after several weeks in vitro, although the percentage of cells synthesizing DNA decreased with time. Efforts to identify peptide growth factor-responsive tyrosine phosphorylation were negative, and the overall amount of S. mansoni phosphotyrosine-containing proteins identified by western blot with anti-phosphotyrosine monoclonal antibody was much less than that found in a peptide growth factor-responsive mouse cell line.

Light and scanning electron microscopic studies of the extrahepatic bile duct of sheep with experimentally induced Fasciola hepatica infection.

Changes in the common bile duct that contained adult Fasciola hepatica of sheep were evaluated by light (LM) and scanning electron microscopy (SEM). Nine ewes were inoculated with F hepatica metacercariae and necropsied 18 weeks after inoculation. The proximal portion of the common bile duct (CBD) that contained adult flukes was recovered and examined by LM and SEM. The CBD from 2 noninoculated ewes were used for control. On gross examination, CBD were markedly large because of the adult flukes, which were free in the lumen of the ducts. Extensive hemorrhage was not found either in intrahepatic or in extrahepatic bile ducts of any sheep. Histologic examination revealed changes, such as villous hyperplasia and hypertrophy of the epithelium; cell infiltration, predominantly with eosinophils or macrophages; and arterial intimal proliferation. By SEM, the epithelial surface of the CBD appeared intact. Villous hyperplasia and hypertrophy of the epithelium observed by LM was clearly seen by SEM. Epithelial damage, seen as small areas of denuded surface by LM and SEM, was confined to a few areas of the mucosa. Lack of extensive hemorrhage and confined epithelial damage were evaluated relative to the mode of feeding of adult flukes.

Enterovirulence of enterotoxigenic Bacteroides fragilis in gnotobiotic pigs.

A porcine isolate of enterotoxigenic Bacteroides fragilis colonized the intestinal tract and caused watery, nonhemorrhagic diarrhea when given orally to 12, 1- to 2-day-old gnotobiotic pigs. Diarrhea occurred 2 to 3 days post-inoculation and continued throughout the 4 to 6 day post-inoculation period. Diarrheic pigs became mildly anorexic and dehydrated. They developed intestinal lesions characterized by swelling, vacuolation, and exfoliation of enterocytes, and crypt hyperplasia throughout the large intestine and, to a lesser extent, in the distal small intestine. Bacterial adherence to, or invasion of, the intestinal mucosa was not detected. A porcine isolate of nonenterotoxigenic B. fragilis was administered orally to six control pigs. The isolate colonized the intestinal tract, but the pigs did not develop clinical disease or intestinal lesions. The pathogenetic mechanism of the disease may involve mediation by a soluble enterotoxin (or toxins) elaborated by B. fragilis.

Toxicity to isolated hepatocytes caused by the intracellular calcium indicator, Quin 2.

To determine whether incubation for several hours with intracellular Ca++ indicators caused toxicity to freshly isolated hepatocytes from rats, cells were incubated under 95% O2-5% CO2 in medium containing 2 mM Ca++ and the acetoxymethyl (AM) esters of Quin 2, Indo 1, Fluo 3, 5,5'-Dimethyl BAPTA, 4,4'-Difluoro BAPTA or Fura 2 for up to 5 hr. Quin 2-AM and Indo 1-AM (2.5 microM) induced lipid peroxidation in the cells after 1 or 3 hr of treatment, respectively. Additional experiments with Quin 2-AM (25 microM) revealed that it also caused lactate dehydrogenase leakage, cell blebbing and vitamin E loss in cells, but did not affect reduced glutathione or intracellular Ca++ content. The ability of Quin 2-AM to cause toxicity was dependent on the amount of Quin 2 which was present in the cell. Ca++ appeared to be involved in the mechanism of Quin 2-AM toxicity, for modulation of the extracellular Ca++ concentration partially inhibited lipid peroxidation, vitamin E loss, cell blebbing and lactate dehydrogenase leakage.

Death of serum-free mouse embryo cells caused by epidermal growth factor deprivation.

Serum-free mouse embryo (SFME) cells, derived in medium in which serum is replaced with growth factors and other supplements, are proastroblasts that are acutely dependent on epidermal growth factor (EGF) for survival. Ultrastructurally, an early change found in SFME cells deprived of EGF was a loss of polysomes which sedimentation analysis confirmed to be a shift from polysomes to monosomes. The ribosomal shift was not accompanied by decreased steady-state level of cytoplasmic actin mRNA examined as an indicator of cellular mRNA level. With time the cells became small and severely degenerate and exhibited nuclear morphology characteristic of apoptosis. Genomic DNA isolated from cultures undergoing EGF deprivation-dependent cell death exhibited a pattern of fragmentation resulting from endonuclease activation characteristic of cells undergoing apoptosis or programmed cell death. Flow cytometric analysis indicated that cultures in the absence of EGF contained almost exclusively G1-phase cells. Some of the phenomena associated with EGF deprivation of SFME cells are similar to those observed upon NGF deprivation of nerve cells in culture, suggesting that these neuroectodermal-derived cell types share common mechanisms of proliferative control involving peptide growth factor-dependent survival.

Ultrastructure of hepatocellular neoplasms in aflatoxin B1 (AFB1)-initiated rainbow trout (Oncorhynchus mykiss).

The fine structure of hepatocellular neoplasms from aflatoxin B1 (AFB1)-initiated rainbow trout was studied by transmission electron microscopy. Large, usually uniform hepatic nuclei, large nucleoli, abundant, dilated rough-surfaced endoplasmic reticulum, and reduced glycogen storage were common findings in both hepatocellular adenomas and hepatocellular carcinomas. In addition, the presence of poorly developed microvilli in the space of Disse and in bile canaliculi, the occurrence of few or no bile preductule cells and a striking increase in the size and number of intercellular spaces characterized hepatocellular carcinomas. The three latter characteristics of hepatocellular carcinomas suggest loss of inter-relationships between hepatocytes and the microvascular system (sinusoids), between hepatocytes and the biliary system, and between individual hepatocytes, respectively. With respect to these parameters, adenomas were more similar to normal liver than to carcinomas.

Myelitis in a cat infected with Toxoplasma gondii and feline immunodeficiency virus.

Severe necrotizing myelitis secondary to localization and reactivation of Toxoplasma gondii within the spinal cord of a domestic shorthair cat was diagnosed by use of light and electron microscopy and immunohistochemistry. The cat also was infected with feline immunodeficiency virus. This case may have useful comparative features to T gondii infections in human patients with acquired immunodeficiency syndrome.

Comparative virulence of two porcine group-A rotavirus isolates in gnotobiotic pigs.

The virulence of 2 porcine group-A rotavirus isolates was compared. Forty hysterotomy-derived 3-day-old gnotobiotic pigs were inoculated orally with 2 ml of intestinal homogenate containing either the Ohio State University (OSU) or the South Dakota State University (SDSU) strain of porcine rotavirus or were inoculated with medium only. Clinical signs of disease, body weight, distribution of viral antigen, fecal excretion of virus, and histologic lesions (observed by light and scanning electron microscopy) were determined. Morphometric measurements of villi and crypts were made. In pigs inoculated with OSU or SDSU strains, diarrhea began at postinoculation hours (PIH) 19 to 48 and PIH 24 to 54, respectively. None of the virus-infected pigs died as a consequence of infection and all had similar clinical signs of disease, body weight changes, and virus-shedding patterns, regardless of the strain of rotavirus with which they were infected. Microscopic findings in the small intestine of virus-infected pigs were similar, except that the SDSU strain caused more severe villus atrophy and villus fusion in the duodenum at PIH 72 and 168 than was associated with the OSU strain. Viral antigen in the small intestine of pigs infected with either virus was observed by use of immunofluorescence at PIH 24 and 72, but was seldom seen at PIH 168.

Infection of gnotobiotic pigs with an Escherichia coli O157:H7 strain associated with an outbreak of hemorrhagic colitis.

Gnotobiotic pigs inoculated with an Escherichia coli O157:H7 strain isolated from a human with hemorrhagic colitis developed anorexia, lethargy, and watery diarrhea. Bacteria diffusely colonized the cecum and colon surfaces and the crypt epithelium. At bacterial attachment sites, microvilli were effaced, and epithelial cells were irregularly shaped, rounded, or detached. Submucosa, lamina propria, and mesentery were markedly edematous and contained many inflammatory cells.