Characterization and expression of a spliced leader RNA in the parasitic nematode Ascaris lumbricoides var. suum.

The parasitic nematode Ascaris spp. contains a 22-nucleotide spliced-leader (SL) sequence identical to the trans-SL previously described in Caenorhabditis elegans and other nematodes. The SL comprises the first 22 nucleotides of a approximately 110-base RNA and is transcribed by RNA polymerase II. The SL RNA contains a trimethylguanosine cap and a consensus Sm binding site. Furthermore, the Ascaris SL RNA has the potential to adopt a secondary structure which is nearly identical to potential secondary structures of similar SL RNAs in C. elegans and Brugia malayi.

The ultrastructure of radiation-induced endosteal myelofibrosis in the dog.

A rapidly developing, progressive form of endosteal myelofibrosis (MF) (with myeloid metaplasia) has been shown to occur at low frequency (approximately 4%) in dogs exposed continuously to low daily doses (10 R/day) of whole-body gamma irradiation. We report in this study the morphological details of the endosteal surface during both preclinical and clinical phases of developing MF by combination light microscopy and scanning/transmission electron microscopy. Pronounced alterations of the endosteum were observed and included: (1) during the early preclinical phases, a progressive time-dependent transition of the endosteal surface from predominantly resting to actively formative and resorptive states; and (2) during the late preclinical phase, aberrant autonomous osteogenic process(es) characterized by a marked reduction in the resorptive, osteoclast-associated endosteal areas occurring concomitantly with further increases in formative areas of the endosteum. Localized patches of overlapping, morphologically transformed endosteal cells (i.e., round-osteoblastic to branched-reticular shaped) were observed within the morphologically reactive, formative endosteum. Osteogenic-endosteal changes coincided with major restructuring of the hematopoietic parenchyma and supporting stromal network. We discuss the possibility that the early occurring endosteal changes are causally linked to normal reparative functions that operate during regenerative hematopoiesis following local and systemic injury. Based on morphological data collected during the late preclinical phase, we speculate that the mechanism of myelofibrosis induction involves the failure to terminate early osteogenic-dependent repair sequences.

Sequential changes in bone marrow architecture during continuous low dose gamma irradiation.

Beagles continuously exposed to low daily doses (10 R) of whole-body 60Co gamma-radiation are prone to develop either early occurring aplastic anemia or late occurring myeloproliferative disorders (Seed et al., 1977). In this study, we have examined by a combination of light microscopy and scanning and transmission electron microscopy the sequential changes in the morphology of biopsied rib bone marrow of continuously irradiated dogs that developed either aplastic anemia, myelofibrosis, or myelogenous leukemia. Characteristic modification of key elements of marrow architecture have been observed during preclinical and clinical phases of these hemopathological conditions. The more prominent of these changes include the following. (i) In developing aplastic anemia: severe vascular sinus and parenchymal cord compression, and focally degenerate endosteal surfaces. (ii) In developing myelofibrosis: hyperplasia of endosteal and reticular stomal elements. (iii) In developing leukemia: hypertrophy of reticular and endothelial elements in the initial restructuring of the stromal matrix and the subsequent aberrant hemopoietic repopulation of the initially depleted stromal matrix. These architectural changes during preclinical phases appear to be related to the pathological progression to each of the radiation-induced hemopathological end points.

Preparation and observation by SEM of hemopoietic cells cloned in soft agar.

A method is described to prepare clones of hemopoietic cells grown in soft agar for scanning electron microscopy (SEM). A critical modification of the otherwise quite standard SEM processing procedure for biological samples involved the use of silver micropore disks as an adherent substrate to support the highly labile, deformable agar slabs. This support allows maintenance of the normal flat pancake shape of the specimen through the thiocarbohydrazide osmium ligand binding steps, dehydration, and critical point drying. With this support and careful dissection of the surface agar with a fine steel needle using a stereomicroscope, selected areas and depths within the colony can be exposed and examined by SEM. Surface topography of cloned cells can be correlated with intracellular cytological features by excising areas of interest and directly embedding them in plastic for thin-section preparation and viewing by transmission electron microscopy (TEM). The dried-specimen-teasing method appears useful, because of the ease of preparation of the specimens, its reproducibility, and the degree of visibility and preservation of cell surface structures and intraclonal relationships. Our initial observations, using combined EM techniques, indicate that clonal cell topography is highly variable and that this variability appears to be related both to the relative age and proliferative status of the colony. Based on work to date, we suggest that topographical and spatial analysis, in vitro of cloned, agar-embedded hemopoietic stem cells is possible with simple modifications of conventional SEM preparative techniques.