Requirement for binding of catalytically active factor VIIa in tissue factor-dependent experimental metastasis.

Tissue factor (TF), the initiating cell surface receptor of the coagulation cascade, plays important roles in embryogenesis, angiogenesis, and tumor cell metastasis. It is controversial whether proteolytic function of TF complexed with its serine protease ligand VIIa is required for metastatic tumor dissemination. We show here in a model for TF-dependent experimental hematogenous metastasis, that TF supports metastasis by both proteolytic activity of the TF-VIIa complex and currently undefined functions of the cytoplasmic domain. We demonstrate that ligand binding of VIIa to TF is required for metastasis. Antimetastatic properties of covalently inactivated VIIa provide evidence that ligand binding is insufficient per se to support metastasis, emphasizing that proteolytic activity is necessary for the metastatic process. Ala or Asp mutations of cytoplasmic serine residues were introduced to preclude or mimic phosphorylation. In vivo analysis of these mutants suggests that local protease generation on the tumor cell surface does not serve simply to activate the cytoplasmic domain of TF by serine phosphorylation. Thus, extracellular functions of the catalytically active TF-VIIa complex cooperate with specific functions of the TF cytoplasmic domain to support the complex process of hematogenous tumor cell dissemination.     

Risk of recurrence of birth defects in Washington State

A population-based study was conducted using maternally-linked birth certificate records from Washington State for 1980–93 to evaluate the risk of birth defect occurrence among infants with pre-viously affected siblings, relative to infants whose siblings did not have birth defects. The risks of recurrence of similar and dissimilar defects were estimated, and the effects of change in paternity and/or city of residence were evaluated as proxies of genetic and environmental effects. At the first birth on record, 3322 women were identified in the linked certificates as giving birth to a child with a birth defect; 6620 women whose first birth did not result in an infant with a defect were randomly selected for comparison. Women with a malformed infant had an in-creased risk of having a malformed infant at the subsequent birth (Relative Risk = 1.9, [95% Confidence Interval (CI) = 1.5–2.4]), which did not vary by intervening changes in partner or residence. The risk of recurrence of the same general type of defect [RR = 11.7, 95% CI = 9.7–19.50] was much greater than that of occurrence of a dissimilar defect [RR = 1.5, 95% CI = 1.1–1.9]. This was consistent for all defect categories, and did not vary markedly by changes in partner or residence.     

Early postnatal hyperthyroidism alters hippocampal circuitry and improves radial-maze learning in adult mice

Inbred mice show strain-specific differences in the hippocampal mossy fiber projection. These differences are most pronounced in the portion of the projection that forms synaptic connections with the basal dendrites of the CA3 pyramidal neurons [intra- and infrapyramidal mossy fiber (IIP-MF) projection]. We have previously demonstrated that the extent of the IIP-MF subfield is positively correlated with the capacity to perform a spatial radial-maze task and that an experimentally induced enlargement of the IIP-MFs, by means of postnatal thyroxine treatment, predicted the ability of adult two-way avoidance learning. In the present study, we tested whether this treatment would also influence radial-maze performance. Forty-five male mouse pups from the inbred strain DBA/2 (chosen because of scanty IIP-MF projection and poor radial-maze learning) were divided into three groups that received daily injections of either 2 micrograms L-thyroxine, an alkaline vehicle solution, or physiological saline. Treatment lasted from postnatal days 0 to 11. At the age of 3 months, these animals were tested in an eight-arm radial maze. The extent of their IIP-MF projections was measured by means of planimetry on Timm-stained sections. Thyroxine-treated animals made significantly fewer errors and had larger IIP-MF projections as compared to both control groups. Within each group, the individual variability of the IIP-MF projection was significantly and positively correlated with performance. We conclude that experimentally modified IIP-MF projections mediate processes underlying spatial working memory. It would appear that the hippocampal circuitry alterations induced by postnatal hyperthyroidism can counteract a hereditary lack of talent, albeit only partially and in selected populations.     

Biological mechanisms and cardiovascular effects of omega-3 fatty acids

The mechanisms and cardiovascular effects of omega-3 fatty acids are reviewed. Omega-3 polyunsaturated fatty acids are the major ingredient found in commercially available fish oil products. The incidence of many diseases, including coronary heart disease, diabetes mellitus, and psoriasis, is lower in Eskimos, who ingest diets rich in omega-3 fatty acids, compared with European controls. Potential mechanisms by which these fatty acids cause their many physiologic effects include competing with omega-6 fatty acids for prostaglandin and leukotriene pathways and enhancing cell membrane fluidity by virtue of the high degree of unsaturation. Numerous studies have documented longer bleeding times and decreased platelet aggregation in subjects ingesting omega-3 fatty acids. Omega-3 fatty acids may reduce serum cholesterol concentrations by decreasing the synthesis of very low density lipoprotein and, therefore, low-density lipoprotein. Blood viscosity is significantly and uniformly lower in subjects receiving omega-3 fatty acids compared with controls. Potential risks of supplementation with fish oils include hypervitaminosis A and D, vitamin E deficiency, increased bleeding times, decreased platelets, and ingestion of contaminated fish. Supplementation with moderate amounts of omega-3 fatty acids appears to be relatively safe. Possible adverse effects include nausea, diarrhea, and a "fishy" taste. Properly controlled, long-term clinical trials are needed to determine whether supplementation with omega-3 fatty acids would be therapeutically beneficial in various patient populations and disease states.     

Macrophage response to peripheral nerve injury: the quantitative contribution of resident and hematogenous macrophages

Whereas local microglial cells of the CNS rapidly respond to injury, little is known about the functional role of resident macrophages of the peripheral nervous system in nerve pathology. Using bone marrow chimeric rats, we recently identified individual resident endoneurial macrophages that rapidly became activated after nerve injury. However, the extent of local macrophage activation and its quantitative contribution to the total macrophage response is unknown. We now have created chimeric mice by transplanting bone marrow from green fluorescent protein (GFP)-transgenic mice into irradiated wild-type mice, allowing easy differentiation and quantification of hematogenous and resident endoneurial macrophages. After sciatic nerve crush injury, both GFP(-) and GFP(+) resident macrophages, the latter having undergone physiological turnover from the blood before injury, rapidly underwent morphological alterations and increased in number. Proliferating GFP(-) and GFP(+) resident macrophages were abundant and peaked 3 days after injury. A major lesion-induced influx of hematogenous macrophages with a disproportionate increase of GFP(+) macrophages was not observed until Day 4. Throughout all time points examined, GFP(-) resident macrophages were strikingly frequent, reaching maximum numbers 9.5-fold above baseline. There was also a notable proportion of GFP(-) resident endoneurial macrophages phagocytosing myelin and expressing major histocompatibility complex class II. Our results demonstrate for the first time that the rapid response of resident endoneurial macrophages to nerve injury is quantitatively important and that local macrophages contribute significantly to the total endoneurial macrophage pool during Wallerian degeneration.     

Physical Factors affecting Maximum Precipitation of the BSA-AntiBSA Fowl System

Studies were made ascertaining the effects of temperature and length of incubation period on the amount of precipitate formed in the BSA-antiBSA fowl serum system. The specific factors considered were centrifugal temperature, temperature of incubation and length of incubation. Reactions were analysed for the entire precipitin curve, using doubling dilutions of antigen, and also for the region of equivalence using intervals of 1 μg. nitrogen.

Reactions mixtures were incubated at 37° for 3 hours and then centrifuged at 22° or at 4° and the precipitate was analysed for total N precipitated. In addition, secondary incubation periods, following this initial incubation treatment of 3 hours at 37°, were made. The secondary periods were either 18, 66 or 118 hours at 4° or 18 hours at 37° and were followed by centrifugation at either 22° or 4°.

Evaluation of the data showed centrifugation at 22° gave higher amounts of precipitate than at 4° in all cases and the differences were statistically significant in most of the instances. In eighteen of the twenty sera tested, precipitation was at a maximum after 3 hours' incubation at 37° and warm centrifugation in contrast to additional incubation periods and/or cold centrifugation.

     

Immunohistochemical, in situ hybridization, and ultrastructural localization of SARS-associated coronavirus in lung of a fatal case of severe acute respiratory syndrome in Taiwan

This article describes the pathological studies of fatal severe acute respiratory syndrome (SARS) in a 73-year-old man during an outbreak of SARS in Taiwan, 2003. Eight days before onset of symptoms, he visited a municipal hospital that was later identified as the epicenter of a large outbreak of SARS. On admission to National Taiwan University Hospital in Taipei, the patient experienced chest tightness, progressive dyspnea, and low-grade fever. His condition rapidly deteriorated with increasing respiratory difficulty, and he died 7 days after admission. The most prominent histopathologic finding was diffuse alveolar damage of the lung. Immunohistochemical and in situ hybridization assays demonstrated evidence of SARS-associated coronavirus (SARS-CoV) infection in various respiratory epithelial cells, predominantly type II pneumocytes, and in alveolar macrophages in the lung. Electron microscopic examination also revealed coronavirus particles in the pneumocytes, and their identity was confirmed as SARS-CoV by immunogold labeling electron microscopy. This report is the first to describe the cellular localization of SARS-CoV in human lung tissue by using a combination of immunohistochemistry, double-stain immunohistochemistry, in situ hybridization, electron microscopy, and immunogold labeling electron microscopy. These techniques represent valuable laboratory diagnostic modalities and provide insights into the pathogenesis of this emerging infection.     

Expression of interleukin-8 gene in inflammatory bowel disease is related to the histological grade of active inflammation.

Interleukin-8 (IL-8) is a potent cytokine for recruitment and activation of neutrophils. To visualize its distribution in the intestinal mucosa and to understand better its possible role in the induction and promotion of inflammatory bowel disease, expression of the IL-8 gene was analyzed in resected bowel segments of 14 patients with active Crohn's disease or ulcerative colitis. In situ hybridization with IL-8 anti-sense RNA probes revealed strong and specific signals in the histologically affected mucosa. The number of cells expressing IL-8 gene correlated with the histological grade of active inflammation. In accordance with the characteristic histological signs of active disease, IL-8-expressing cells were diffusely distributed over the entire affected mucosa in patients with ulcerative colitis, whereas in patients with Crohn's disease, IL-8-expressing cells showed a focal distribution pattern. Cells expressing IL-8 were mainly located at the base of ulcers, in inflammatory exudates on mucosal surfaces, in crypt abscesses, and at the border of fistulae. Analysis of semi-serial sections pointed to macrophages, neutrophils, and epithelial cells as possible sources of this cytokine in active inflammatory bowel disease. We consistently failed to detect IL-8 messenger RNA in the mucosa of uninvolved bowel segments and in normal-appearing control mucosa of patients with colon cancer. In contrast, tissue specimens from two patients with acute appendicitis displayed IL-8-expressing cells in the mucosa. These results support the notion that IL-8 plays and important but nonspecific role in the pathogenesis of inflammatory bowel disease and that the production of IL-8 messenger RNA is restricted to areas with histological signs of inflammatory activity and mucosal destruction.